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ALEX. AGASSIZ.

Pibrary of the Museum

OF
COMPARATIVE ZOOLOGY,
AT HARVARD COLLBGE, CAMBRIDGE, MASS,

Founded by private subscription, in 1861.

Deposited by ALEX. AGASSIZ.

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ape:

QUARTERLY JOURNAL

OF

MICROSCOPICAL SCIENCE:

EDITED BY

E. RAY LANKESTER, M.A., LL.D., F.R.S.,
Honorary Fellow of Exeter College, Oxford, and Deputy Linacre Professor of Human
and Comparative Anatomy in the University of Oxford.

WITH THE CO-OPERATION OF

Be KLELN, MD: P.R.S:,

Lecturer on General Anatomy and Physiology in the Medical School of
St. Bartholomew's Hospital, London ;

ADAM SEDGWICK, M.A., F.R.S.,
Fellow and Assistant-Lecturer of Trinity College, Cambridge ;
AND

A. MILNES MARSHALL, M.A., D.Sc., M.D., F.R.S.,
Late Fellow of St. John’s College, Cambridge; Professor in the Victoria University ;
Beyer Professor of Zoology in the Owens College, Manchester.

VOLUME XXXII.—New Sertzs.
With Aithographic Plates and Engrabings on Wood.

LONDON:
J & A. CHURCHILL, 11, NEW BURLINGTON STREET.
1891,

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CONTENTS.

CONTENTS OF No. CXXV, N.S., JANUARY, 1891.

MEMOIRS :

Studies on the Comparative Anatomy of Sponges. III.—On the
Anatomy of Grantia labyrinthica, Carter, and the so-called
Family Teichonide. By Artuur Denny, M.Sc., F.L.S., De-
monstrator and Assistant Lecturer in Biology and Fellow of
Queen’s College in the University of Melbourne. (With Plates
I—IV) : : 5 5 ;

Studies on the Comparative Anatomy of Sponges. IV.—On the
Flagellated Chambers and Ova of Halichondria panicea.
By Artuur Drwpy, M.Sc., F.L.S., Demonstrator and Assistant
Lecturer in Biology and Fellow of Queen’s College in the
University of Melbourne. (With Plate V)

On Megascolex ceruleus, Templeton, from Ceylon; together
with a Theory of the Course of the Blood in Harthworms. By
ALFRED GipBs Bourne, D.Sc.Lond., C.M.Z.S., F.L.8., Fellow
of University College, London; Fellow of the Madras Uni-
versity; Professor of Biology in the Presidency College, Madras.
(With Plates VI—IX)

On a Little-known Sense-organ in Salpa. By Artuur BoLies
Lex, Assistant in the Russian Laboratory of Zoology at Ville-
franche-sur-Mer (Nice). (With Plate X) : ;

Immunity against Microbes. By M. Armayp Rurrer, M.A.,
M.D.(Oxon.)

PAGE

41

49

89

98

1V CONTENTS.

CONTENTS OF No. CXXVI, N.S., MARCH, 1891.

MEMOIRS :

On a New Species of Phymosoma, with a Synopsis of the Genus
and some Account of its Geographical Distribution. By ARTHUR
EK. Surptey, M.A., F.L.S., Fellow and Lecturer of Christ’s
College, Cambridge, and Demonstrator of Comparative amuse
in the University. (With Plate XI)

On the British Species of Crisia. By Sipyey F. ae M.A,,
B.Se., Fellow and Lecturer of es s oe Garntonidee!
(With Plate XII) :

The Later Larval Development of ree ee By Asmite
Wituey, B.Sc. (of Pe age ante His Plates
XITI—XV)

On the Structure of Two New Gohan of Masilowaviss sane
to the Eudrilide, and some Remarks on Nemertodril us.
By Franx E. Brepparp, M.A., Prosector to the Zoological So-
ciety of London. (With Plates XVI—XX) ;

CONTENTS OF No. CXXVII, N.S., JUNE, 1891.

MEMOIRS:

The Renal Organs of Certain Decapod Crustacea. By W. F. R.
Wexpovn, M.A., F.R.S., Jodrell Professor of Zoology and Com-
parative Anatomy in University College, London ; Fellow of St.
John’s College, Cambridge. (With Plates XXI & XXII)

The Nephridium of Lumbricus and its Blood Supply; with Re-
marks on the Nephridia in other Chetopoda. By W. BuaxLanp
Brnuam, D.Se.(Lond.), Anatomical Rian Oxford. (With
Plates XXITI—XXV) :

Notes on the Naidiform Oligocheta; containing a Desoeiptien of
New Species of the Genera Pristina and Pterostylarides, and
Remarks upon Cephalization and Gemmation as Generic and
Specific Characters in the Group. By Atrrep Gisps Bourne,
D.Sc.(Lond.), C.M.Z.8., F.L.S., Fellow of University College,
London; Fellow of the Madras University; Professor of Bio-
logy in the Presidency cil Madras. an Plates XXVI &
XXVIII).

PAGE

lll

127

183

235

279

293

335

CONTENTS.

On Pelomyxa viridis, sp. n., and on the Vesicular Nature of
Protoplasm. By ALFRED GiBBs Bourne, D.Sc.(Lond.), C.M.Z.S.,
F.L.S., Fellow of University College, London; Fellow of Madras
University; Professor of Biology in the Presidency College,
Madras. (With Plate XXVIII)

The Meduse of Millepora murrayi and the Gonophores of
Allopora and Distichopora. By Sypney J. Hickson, M.A.,
D.Sc., &., Fellow of Downing ck fates (With
Plates XXIX & XXX) : :

On a Red Pigment-forming Organism, B. corallinus. By
CuHarLEs StatTer, M.B.(Cantab.) (With Plate XX XI)

357

375

409

CONTENTS OF No. CXXVIII, N.S., OCTOBER, 1891.

MEMOIRS :

Immunity against Microbes. By M. Anmanp Rurrzr, M.A.,,
M.D.(Oxon.). (With Plates XXXII & XXXIII)

The Formation and Fate of the Primitive Streak, with Observations
on the Archenteron and Germinal Layers of Rana temporaria,
By ArtHur Rosinson, M.D., Senior Demonstrator of Anatomy
in the Owens College; and Ricnarp Assueton, M.A., Demon-
strator of Zoology in the Owens College. (With Plates XXXIV
& XXXV) :

On some Points in the FEatelty and Development of Myrio-
thela phrygia. By W.B. Harpy, B.A., Shuttleworth Scholar
of Gonville and Caius College, and Junior Demonstrator of
Physiology to the University of aie (With Plates
XXXVI&XXXVII). - ‘

On the Structure of an Earthworm allied to =e had [alate
Mich., with Observations on the Post-embryonic Development of
Certain Organs. By Franx EH. Bepparp, M.A., Prosector of
the Zoological Society of London, Lecturer on Biology at Guy’s
Hospital. (With Plates XXXVIII & XXXIX) :

Some Points in the Development of Scorpio fulvipes, By
Matcorm Lauriz, B.Sc., F.L.S. (With Plate XL) .

417

451

505

539

587

vi CONTENTS.

a

Abstract of Maupas’s Researches on Multiplication and Fertili-
sation in Ciliate Infusorians. By Marcus M. Hartoe, D.Sc.,

M.A., F.L.S., Professor of Natural History at Queen’s College,
Cork : é ; : -

On the Occurrence of Pseudopodia in the Diatomaceous Genera,
Melosira and Cyclotella. By J. G. Grenrett, B.A., F.GS.,
F.R.M.S. (With Plate XLI)

REVIEW:

A Monograph of the Victorian Sponges. By ArtHur Denpy,
D.Sc., F.L.S., Fellow of Queen’s College, and Demonstrator and
Assistant Lecturer in Biology in the University of Melbourne.
Part I.—The Organisation and Classification of the Calcarea
Homocela, with Descriptions of the Victorian Species (with
plates i—xi)

TitLE, CoNTENTS, AND INDEX.

PAGE

599

615

625

Studies on the Comparative Anatomy of
Sponges.
III—On the Anatomy of Grantia labyrin-
thica, Carter, and the so-called Family
Teichonide.

By
Arthur Dendy, M.Sc., F.L.S.,

Demonstrator and Assistant Lecturer in Biology and Fellow of Queen’s
College in the University of Melbourne.

With Plates I—IV.

INTRODUCTION.

Durine the past few years Mr. J. Bracebridge Wilson has,
by perseveringly dredging in the neighbourhood of Port Phillip,
accumulated a collection of sponges which already numbers
something like 2000 specimens, and is probably the most com-
plete collection ever brought together from the shores of any
one country. The entire collection has been generously placed
in my hands for investigation, and Professor McCoy has like-
wise kindly placed at my disposal the collection contained in
the National Museum at Melbourne. The task of dealing
with so large a mass of material is, I need hardly say, one of
great magnitude, and the systematic investigation must neces-
sarily extend over several years. The difficulty of the work,
so far as identification of species is concerned, has been greatly
lessened by the courtesy of Mr. H. J. Carter, who has gene-
rously sent me his own copy of his work on the Port Phillip
Calcispongiz (1), containing a large number of unpublished
sketches ; and of Dr. Giinther, to whom I am deeply indebted

VOL. XXXII, PART I.—NEW SER. A

2 ARTHUR DENDY.

for a large series of duplicate pieces of named sponges from
the British Museum collection, most kindly sent to me since
I left England.

It is hoped that the final account of the collection will be
embodied in one of a series of reports on the marine zoology
of Port Phillip, which it is intended to publish under the
auspices of the Port Phillip Exploration Committee of the
Royal Society of Victoria. Meanwhile, even during the pre-
liminary arrangement of the collection, forms are constantly
being met with which deserve special anatomical investigation.
One such I have already described in the present series of
studies (2); and I hope, as time permits, to be able to deal
similarly with a number of others.

The more one studies the group, the more is one convinced
of the necessity of thorough and minute anatomical investiga-
tion as a basis for classification. Especially in the present
transitional state of our knowledge of the sponges anatomical
investigation must precede systematic work; and the greater
the number of types selected for such investigation, the greater
will be the value of the scheme of classification ultimately
arrived at. Polymorphism and homoplasy occur so generally
and to such an extraordinary degree amongst the Porifera, that
true genetic relationships can be determined only by most
careful examination of the internal anatomy, and especially of
the skeleton and canal system, although even these systems
are by no means exempt from the general rule.

The sponge which forms the principal subject of the present
contribution is a large and singularly beautiful calcisponge,
originally described (3) by Mr. Carter under the name Tei-
chonella labyrinthica. As Mr. Carter subsequently dis-
covered, the sponge is very far removed from. the genus
Teichonella, and must be placed amongst the Sycons, where
for the present, at any rate, it may be classed in the genus
Grantia,!

The material at my disposal for investigating this sponge
consisted principally of a number of fine adult specimens col-

1 Vide Vosmaer’s diagnosis of this (20).

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 3

lected by Mr. Wilson, and preserved in ordinary methylated
spirit. This was supplemented by some fragments taken from
afresh specimen by myself when out dredging with Mr. Wilson,
and preserved in absolute alcohol, and by three young examples
of great interest. Sections were cut in different directions by
hand, by the freezing microtome, and by the paraffin method.
For studying the skeleton, hand-cut sections of unstained ma-
terial and preparations of the spicules boiled out with caustic
potash are most serviceable. For minute anatomical and his-
tological work, thinner sections of material stained with borax
carmine, and cut by the ordinary paraffin method, were found
to yield good results.

The Anatomy of Grantia labyrinthica.

(a) Historical.

The sponge under consideration was, as I have stated above,
originally described by Mr. Carter in the ‘ Annals and Maga-
zine of Natural History ’ in 1878 (8), apparently from a dry
and imperfect specimen in the British Museum collection.
The brief description is confined almost entirely to the ske-
leton and the external characters, but the author observes that
“in spiculation and in the structure of the lamina it is
closely allied to Grantia compressa, Fleming.” He refers
the sponge, however, as already noted, to his genus
Teichonella.

In 1886 Mr. Carter was able (1) to supplement his original
description from the examination of specimens dredged and
sent to England by Mr. Bracebridge Wilson. He observes
that “the sponge is goblet-shaped in general form, and not
simply ‘vallate,’ like T. prolifera; also thata quadriradiate
forms part of its spiculation; hence these additional facts
render it necessary that it should be relegated to the vicinity
of Grantia compressa, where its generic name might be
changed from ‘Teichonella’ to ‘Grantia,’”

- Previously to this date, however, Mr. Carter had published
in the ‘ Annals and Magazine of Natural History’ a remark-

4 ARTHUR DENDY.

able paper (4) on the ‘“‘ Mode of Circulation in the Spongida,”
in which he advocates the theory that “the particles that are
taken in with the water through the pores of the dermis fall
directly into the subdermal cavities, and pass thence into
the large excretory canals, from which they are afterwards
deflected to their destination through smaller branches,
whose apertures may be seen in the walls of the former.” He
adds, “‘ This perhaps is best seen in Teichonella labyrin-
thica, wherein the chambers, which are arranged in juxtapo-
sition perpendicularly to the lamina of which the sponge is
composed, thus pass directly through it from one side to the
other, having therefore on one side the pores or pore-dermis,
and on the other the vent; in short, exactly like those of
Grantia compressa, only there is no cloaca. We must,
however, regard this chamber as at once ampullaceous sac and
excretory canal; for the pore-dermis being at one end or side
of the lamina and the vent at the other, the circulation passes
into the former and out at the latter, through the chamber,
where the nutritive particles are instantly taken up by the
spongozoa lining its cavity. Hence the holes in the walls of
the chamber, which are very numerous, may serve for the
purpose of intercommunication, where the walls of the neigh-
bouring chambers are in direct contact with each other, or for
the purpose of allowing the ova developed in the intercameral
tissue to pass into the chamber and thus be expelled. There-
fore these holes would seem to have more functions than those
ascribed to them in the wall of the ampullaceous sac of the so-
called ‘ siliceous sponges,’ ex. gr. Spongelia avara.” For
a further elucidation of Mr. Carter’s views on the canal
system of sponges in general, the student is referred to his
paper (5) ‘On the Position of the Ampullaceous Sac and the
Function of the Water Canal System in the Spongida;” at
present I wish to consider only the particular ease of Grantia
labyrinthica. Mr, Carter’s account of the arrangement of
the canal system in this sponge is supplemented by a figure
(Pl. LV, fig. 7), wherein the flagellated chamber (= ampulla-
ceous sac or radial tube) is represented as being perfectly

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 5

straight and cylindrical, with a pore-sieve opening into it at
one end and a vent at the other.

Such, then, was the state of our knowledge of Grantia
labyrinthica, one of the most remarkable of all the calca-
reous sponges, when I commenced my investigations. How
far my own observations agree or disagree with Mr. Carter’s
descriptions will appear subsequently.

(6) External Characters.

Grantia labyrinthica is very large for a calcareous
sponge, well-grown specimens being about three inches in
height and a little more in breadth, so that it is probably the
largest of all the Sycons. Hence it is peculiarly well adapted
for anatomical investigation.

A better idea of the external appearance will perhaps be
gathered from an examination of fig. 4 than from any descrip-
tion which I can give. It will be seen that the adult sponge
consists essentially of a thin-walled cup or basin, with a widely
open mouth showing no signs of constriction, and thus differ-
ing markedly from the oscula of all other known Sycons.
The wall of the cup, however, which, as I shall show later on,
is in the young sponge simple and not folded, in the adult
becomes greatly convoluted and folded upon itself, without,
however, ever losing its character of a single continuous
lamella. Thus, while in most Sycon sponges the circumfer-
ential growth of the tube or cup after a time diminishes as the
sponge grows older, giving rise to a more or less constricted
osculum, in Grantia lahyrinthica precisely the reverse
takes place, and the circumferential growth so far outstrips
the vertical growth that the sponge wall becomes thrown into
numerous deep folds, while the osculum becomes enormously
wide and bounded by a deeply sinuous margin. The cup
thus formed is attached by the middle of its lower surface to
a stout cylindrical stalk, which is also a subsequent develop-
ment not present in the very young sponge. At its lower end
the stalk is fixed to the rock or other body upon which the
free swimming embryo may have chanced to come to rest.

6 ARTHUR DENDY.

This peculiar external form is to some extent paralleled by
certain species of the genus Phyllospongia amongst the
horny sponges, as will be evident on referring to Lendenfeld’s
figures (6). By far the most remarkable parallelism, however,
is exhibited by a Renieriz species, not yet determined, which
so closely resembles a young Grantia labyrinthica in
external appearance that I at first placed it along with other
specimens of that sponge to be figured, and only found out
my mistake on microscopical examination. This interesting
example of homoplasy serves well to show the necessity of
microscopical examination before even the approximate posi-
tion of any particular sponge can be safely determined.

Both surfaces of the cup are smooth, and the inner surface
is at once seen to be perforated by innumerable minute and
closely placed apertures, the exhalant openings of the flagel-
lated chambers. These give to the surface a minutely punc-
tate appearance, which is absent immediately below the free
margin, where the sponge wall becomes very thin and translu-
cent. On the outer surface of the cup the pore-sieves form
less obvious markings.

(c) The skeleton.

The Spicules.—The calcareous spicules composing the
skeleton of the sponge are of three main types—triradiate,
quadriradiate, and uniaxial (oxeote). Each of these types
occurs in the sponge under more than one modification,
according to its position.

Triradiate Spicules.

These form the main mass of the skeleton. The different
modifications which they present in different parts of the
sponge depend chiefly upon the relative length of the rays;
and, to some extent, upon the proportions of the angles
between them. Thus we find a more or less gradual series
between approximately equiradiate and equiangular spicules
on the one ,hand, in which all three rays are of about the
same length, and the angles between them nearly equal (fig.

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 7

16), and extremely inequiradiate and inequiangular spicules,
of the form termed by Haeckel (7) ‘‘ sagittal,’’ on the other.
In the sagittal spicules two rays are about equal in length,
while the third has either grown out into a very long and
slender shaft, which may attain to three or four times the
length of either of the others (figs. 13, 14), or (much more
rarely) remained short while the other two have grown long
(fig. 18). The angle between the two paired arms, or the oral
angle, as Haeckel terms it, is greater than either of the other
_two angles, which are equal.

Figs. 13 to 19 represent seven triradiate spicules, and
illustrate the variation in the proportions of the rays and
angles. The commonest form is the sagittal, with one ray
(the shaft) much longer than the other two. It is important
to notice that, as a general rule at any rate, the three rays of
the triradiate do not all lie in exactly the same plane, so that
if the spicule were laid down upon an even surface it would
rest upon the ends of the three rays, with the centre elevated.
This fact is not shown in the figures, which are merely out-
lines drawn with the camera. In the sagittal spicules the
shaft is usually perfectly straight, but often of the beautiful
spear-like form shown in figs. 13 and 14. The two paired
rays, on the other hand, are often slightly curved.

The triradiates vary considerably in size. The following
measurements are taken from a well-grown spicule, of the
form shown in fig. 13, and all the other spicules (from figs.
7 to 20 inclusive) are drawn to the same scale.

Length of the shaft 0°38 mm.
Length of the paired rays 0°12 mm.

Quadriradiate Spicules.

These are probably to be regarded only as further modifi-
cations of the fundamental triradiate type, for although they
are quadriradiate they are still only triaxial. Their shape is
usually that represented in fig. 20. It will be seen that the
third ray of a normal triradiate spicule has become much

8 ARTHUR DENDY.

shortened, while an additional fourth ray has appeared as a
direct continuation of the third ray in the angle between the
two paired rays. The cessation of growth of the third ray in
its usual direction is perhaps to be accounted for by the out-
growth of the new fourth ray in the exactly opposite direction.
The additional fourth ray is usually slightly hastate in form,
the surface of the spear-head being at the same time slightly
roughened. Occasionally, as shown in fig. 21, this peculiarity
in form is very much more strongly marked. The size of the
quadriradiates may be estimated from fig. 20.

Uniaxial Spicules.

The shape of these spicules is a modification of the oxeote
type, in which one end is markedly broader than the other,
and often decidedly hastate (figs. 10,11). Fig. 11 represents
a spicule of the more usual size and form; figs. 7, 8, 9 repre-
sent giant modifications of the same from the margin of the
osculum, and fig. 12 represents the large-sized form found in
the stem. All the figures are drawn to the same scale as the
triradiates.

The Arrangement of the Skeleton.—In dealing with
this part of my subject I propose to follow the plan laid down
by Haeckel in his ‘Monograph of the Calcareous Sponges,’
which seems to be in all respects the most satisfactory.

In the Sycons Haeckel distinguishes the following skeletal
systems, which, for the sake of convenience, I arrange in an
order slightly different from his :

1. The dermal skeleton, protecting the outer or dermal
surface of the tube or cup, of which the Sycon individual
consists.

2. The gastral skeleton, protecting the inner or gastral
surface.

3. Theskeleton of the peristome, protecting the margin
of the osculum.

4, The tubar skeleton, lying between the dermal and
gastral systems, and affording support to the flagellated
chambers,

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 9

5. The skeleton of the base or stalk (where one is
present).

It will be evident from the following account that Grantia
labyrinthica conforms very closely in the arrangement of
the skeleton to the normal Sycon plan.

The Dermal Skeleton.

This forms a distinct cortex of somewhat varying thickness
over the outer surface of the sponge (figs. 22, 25). The main
mass of this cortex consists of large sagittal triradiates, arranged
so that the long unpaired ray points towards the base of the
sponge. Even within the limits of the dermal cortex the tri-
radiates exhibit considerable individual variation in shape and
size. In the inner portion of the cortex numbers of the
singularly beautiful sagittal triradiates represented in figs. 13
and 14 occur.

The outermost portion of the dermal skeleton consists of
great numbers of small oxeote spicules of the form represented
in figs. 10 and 11, placed at right angles to the surface, with
their narrow ends embedded amongst the large triradiates, and
their broad ends projecting freely (fig. 22). In the pore-areas
the dermal skeleton is practically reduced to this outer layer
of small oxea, many of which appear to be quite flat in the
thin dermal membrane.

The Gastral Skeleton.

The gastral skeleton (figs. 22, 25) forms a protective cortex
over the inner surface of the cup, exactly as the dermal
skeleton does over the outer. The gastral and dermal cortex
do not differ very greatly in thickness, although in this respect
there seems to be a good deal of variation. In the specimen
before me as I write the gastral cortex is decidedly thinner
than the dermal. Like the latter it is made up princi-
pally of triradiate spicules interwoven to form a feltwork ;
as before, the spicules are usually sagittal, with the long arm
pointing towards the base of the sponge. The beautiful

10 ARTHUR DENDY.

sagittal triradiates represented in figs. 138 and 14 occur
also in the gastral cortex; and here, too, we meet with
the quadriradiate spicules. The latter surround the short
exhalant canals of the flagellated chambers, being arranged as
shown in figs. 21 and 24, with the additional fourth ray pro-
jecting towards or into the lumen of the canal. Occasionally
an unusually small exhalant canal is met with, cut transversely,
in tangential sections of the cup-wall, a little below the level
of the ordinary exhalant canals. Such small canals (fig. 21)
are surrounded by quadriradiate spicules of the slightly different
form already described.

On the extreme outside of the gastral cortex (i. e. the extreme
inside of the cup-wall) there occur abundant oxeote spicules
like those of the dermal skeleton, and arranged, in close-set
tufts, perpendicularly to the surface with their broad ends
outwards (fig. 22). Mr. Carter (8) states that the linear
(oxeote) spicules on the ‘‘ vent-side” of the cup-wall are twice
the length of those on the “ pore-side.”” This does not hold
good as a general rule, for in the specimen before me the
reverse is the case (fig. 22).

The Skeleton of the Peristome.

This consists of a fringe of the giant oxeote spicules already
described (figs. 7—9), arranged with their broad ends pro-
jecting freely around the margin of the osculum, and their
narrow ends embedded amongst a mass of approximately equi-
radiate triradiates, into which the dermal and gastral skeletons
merge. The oscular fringe (fig. 25, 0. sp.) thus formed is so
very slightly developed in proportion to the size of the whole
sponge that it is scarcely noticeable with the naked eye.

The Tubar Skeleton.

According to the manner in which the spicules are arranged
around the flagellated chambers (radial tubes) Haeckel (7)
distinguishes two types of tubar skeleton, (1) articulate (geglie-
dertes) and (2) inarticulate (ungegliedertes). The articulate

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 11

tubar skeleton is always composed of triradiates, and is dis-
tinguished by the fact that along the length of the chamber
there are always two or more transverse zones or “joints ” of
triradiates, one behind the other. The triradiates are usually
sagittal, and the shaft is directed towards the dermal surface.
The longer the flagellated chamber the greater is the number
of joints in its skeleton.

Haeckel adds that in most Sycons with an articulate tubar
skeleton the separate joints of the latter become specially
differentiated. Thus the first or innermost joint is longer
than the following; the outermost, on the other hand, is the
shortest. The triradiates of the first joint, again, are most
markedly sagittal, while their basal ray is unusually elongated,
and their paired lateral rays are placed with convex, curved
margin beneath the gastral surface. The triradiates of the
following joints are usually less markedly sagittal, their basal
ray less hypertrophied, and their oral angle generally smaller.
Finally, at the distal end of the tube, towards the dermal
surface, the sagittal triradiates generally pass over into the
regular or subregular, and often into the irregular form.

The tubar skeleton in Grantia labyrinthica (fig. 22) is
articulate, and agrees very exactly with the general description
of such a skeleton given by Haeckel. The first joint is much
longer than any of the others, and the sagittal triradiates
composing it (=subgastral triradiates of Haeckel) are modified
precisely as he describes, their short, curved, lateral arms
lying beneath, or, indeed, forming a portion of, the gastral
skeleton. The number of joints depends upon the length of
the flagellated chamber, and this again varies with its position
in the sponge, the older and longer chambers being situate
nearer the base, and the younger and shorter ones nearer the
margin of the cup.

It is hoped that fig. 22, which is drawn on the same plan as
that adopted by Haeckel, will make all these points clear
without further description,

It remains to be added that the two shorter arms of each
triradiate curve slightly towards one another, so as partly to

12 ARTHUR DENDY.

embrace the chamber which they help to support. This is
seen in figs. 28 and 29, where, owing to the direction of the
section at right angles to the long axes of the flagellated
chambers, the shafts of the triradiates are cut off, while the
paired lateral arms are seen partially surrounding the chambers.

The Skeleton of the Stalk.

This consists essentially of a confused mass of closely inter-
woven sagittal triradiates with very long and slender arms.
One of these spicules is represented in fig. 19; often they are
more or less irregular in form. On the extreme outside there
is a layer of oxeote spicules disposed at right angles to the
surface. Most of these spicules are small, and very like those
found over the surfaces of the cup, but a large number are
modified into giant forms (fig. 12), differing somewhat in shape
from those which form the oscular fringe. These giant
spicules give to the surface of the stalk a more or less hispid
character. They are remarkable from the fact that they
project to an unusual extent, so that commonly less than a
“quarter of the length of the spicule is embedded in the tissues
of the sponge. The outer ends of the spicules are consequently
generally worn or broken.

(d) The canal system.

Grantia labyrinthica appears to agree more closely with
respect to the arrangement of the canal system with Haeckel’s
Sycortis levigata (7) than with any other described form.
The canal system of all the Sycons is, of course, fundamentally
the same, but numerous, by no means insignificant, variations
occur, especially with regard to the inhalant pores and canals.

The canal system of calcareous sponges may be described in
precisely the same terms as that of the siliceous and horny
sponges, and since it is advisable to preserve uniformity of
nomenclature wherever possible, I shall follow Poléjaeft’s
example (8) in making use of such general terms as “ flagel-
lated chamber” and “inhalant canal” in preference to such
special terms as “radial tube” and ‘‘intercanal” used by
Haeckel for the Calcarea. The term “ gastral cavity” I pro-

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. IBS

pose to retain in the present paper, because, although the
gastral cavity corresponds functionally to the oscular tube of
siliceous and horny sponges, it is very improbable that the
two structures are homologous.

The Inhalant Pores.

According to Haeckel (7), in certain of the Sycons a portion
of the inflowing water obtains direct access to the tubular
flagellated chambers by means of ‘ dermal ostien ” situate at
the distal extremity of the latter. This never takes place in
Grantia labyrinthica, although Carter maintains, as I
have already mentioned, that the pore-sieves are placed over,
and lead into the distal ends of the chambers. As a matter of
fact, the pore-sieves, or pore-areas, lie between the ends of
the flagellated chambers, and over the ends of the inhalant
canals. The blind ends of the flagellated chambers, on the other
hand, are covered over by the well-developed dermal cortex.

Each group of pores—for which we may conveniently use
the term pore-area as in other sponges—is usually more or
less oval in outline, and contains a dozen or more small round
pores (fig. 30). Thelonger diameter of the pore-areas averages
about 0°33 mm. in length, and the diameter of the pores
themselves about 0:033 mm. The pore-areas and pores are
best studied in tangential sections of the dermal surface, a
method the importance of which cannot be too strongly insisted
upon. In the pore-areas the cortex is reduced to a mere thin
membrane, corresponding to the dermal membrane of other
sponges (e.g. Monaxonider), containing large numbers of
small oxeote spicules, and perforated by the pores. If the sec-
tion be very thin—only of about the thickness of the dermal
membrane—it is not easy to determine the boundaries of the
pore-areas, which lie very close together. If, however, the
section be fairly thick, then a portion of the mesodermal
trabecule separating the subdermal cavities will be included,
and the appearance shown in fig. 30 will be presented, where
each pore-area is seen overlying the end of an inhalant canal.
Fig. 31 shows a single pore more highly magnified.

14 ARTHUR DENDY.

The Inhalant Canals.

These commence as widely expanded cavities immediately
underlying the pore-areas. These cavities correspond in posi-
tion to the subdermal cavities of other sponges, but they
merge so gradually into the deeper parts of the inhalant
canals, with which they are directly continuous, that it is
impossible to distinguish the boundaries between the two.
The inhalant canals (intercanals of Haeckel) are by no means
regular; they may anastomose and they may branch. The
anastomosis. takes place—most frequently, at any rate—just
below the surface, so that the pores of two contiguous areas
may lead almost directly into one and the same inhalant
canal. The branching takes place chiefly at the far end of
the canals, towards the gastral surface.

At first very wide, the inhalant canals, as they penetrate
below the dermal cortex and between the flagellated chambers,
rapidly diminish in diameter, and finally come to an end just
below the gastral cortex where the flagellated chambers are
just commencing (figs. 25, 26, im. c.). In figs. 28 and 29,
which represent sections taken at right angles to the loug
axes of the flagellated chambers, the inhalant canals are seen
cut transversely between the chambers. In fig. 28, which
represents a section taken not very far from the middle of the
sponge-wall, the inhalant canals are still wide; but in fig. 29,
which represents a section from near the gastral surface, the
inhalant canals have become very much reduced in size.

The Prosopyles.

The term “ prosopyle ” is used by Sollas (9) to designate the
openings of the inhalant canals into the flagellated chambers,
and to distinguish them from the inhalant pores on the surface
of the sponge. As it is a decided advantage to employ two
separate terms for these two very distinct structures, I shall
adopt Sollas’s nomenclature.

The prosopyles in Grantia labyrinthica are numerous
small circular apertures, each about 0°018 mm. in diameter,

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 15

which place the inhalant canals in direct communication with
the flagellated chambers. On looking down upon the wall of
a chamber the prosopyles are seen fairly regularly scattered
over it (figs. 23, 25). In this way they are most readily
recognised, but in order to see the actual communication
between the flagellated chambers and the inhalant canals it is
necessary to examine very thin sections. We then see that
the amount of mesodermal tissue intervening between the
inhalant canals and the chambers which they supply is by no
means great. On approaching a prosopyle (fig. 26, pr.) it
thins away altogether, and the flattened epithelial lining of the
inhalant canal meets the lining of collared cells of the chamber
around the margin of the circular aperture. The prosopyles
are scattered over the whole of the chamber, so that some are
found quite close to the exhalant aperture (fig. 26); in such
cases the water can only just enter the chamber and leave it
again almost immediately.

Mr. Carter gives a very different account of the prosopyles
in Grantia labyrinthica, which I have already quoted on
a previous page. They do not, as he suggests, ‘‘ serve for the
purpose of intercommunication where the walls of the neigh-
bouring chambers are in direct contact with each other, or for
the purpose of allowing the ova developed in the intercameral
tissue to pass into the chamber and thus be expelled.” In
the first place it appears, as I shall show presently, that the
walls of neighbouring chambers never are in direct contact
with each other, nor have I found their cavities ever in direct
communication ; and, secondly, I have been able to prove (10)
that the embryos escape in a very different manner from that
suggested.! In short, the prosopyles of Grantia labyrin-
thica agree in function and position with those of other Sycons
as described and figured by Schulze (11), Poléjaeff (8), and
others.

The Flagellated Chambers.
These have the usual Sycon character of more or less
1 Cf. next page.

16 ARTHUR DENDY.

cylindrical tubes penetrating the sponge-wall at right angles
to its two surfaces—not extending, however, completely from
surface to surface, but terminating at either end just beneath
the cortex (figs. 23, 25—27). In transverse section (figs.
28, 29) the chambers appear approximately circular, or at all
events more or less rounded in outline, and not, as in many
Sycons, polygonal from mutual pressure. The retention of
the primitive cylindrical character is doubtless due to the fact
that the chambers are not very closely packed, but separated
by a fair amount of intervening mesoderm. At their peri- |
pheral ends the chambers terminate blindly beneath the dermal
cortex, there being, as already stated, no dermal ostia to place
them in direct communication with the exterior. At their
peripheral ends also the chambers exhibit a marked inclination
towards branching. I have endeavoured to represent the most
striking instance of this which has come under my notice in fig.
23. This tendency towards branching of the chambers appears
to be not very uncommon amongst the Sycons, and is mentioned
by Schulze in the case of Syeandraraphanus (11). I hope
to be able to discuss its possible significance at a later date.

In specimens, or in those parts of specimens which contain
pretty far advanced embryos, the walls of the flagellated
chambers are frequently seen to exhibit little shallow pits on
their inner surface (fig. 23, em. c.). These little pits or
pockets, instead of being lined by the usual collared cells, are
lined by flattened pavement-cells. They are the remains of
cavities in the mesoderm from which embryos have escaped
by bursting through the wall of the chamber and tearing away
part of it with them. The collared cells of the part torn away
first become stretched out and flattened, as shown in fig. 38,
by the pressure of the growing embryo beneath them; finally
they appear to degenerate altogether, so as to form a structure-
less membrane, which is carried away bodily by the escaping
embryo. For further particulars as to the mode of escape of
the embryos the student is referred to my paper “On the
Pseudogastrula Stage in the Development of Calcareous
Sponges ” already cited.

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 17

The stream of water leaves the flagellated chamber through
the gastric ostium or exhalant aperture, a wide opening guarded
by a delicate, membranous, sphincter diaphragm, as already
described (4) by Carter (vide figs. 28—26).

In sections such as that represented in fig. 29 I have
not infrequently met with peculiar structures having the
appearance shown at w in the figure. These structures
evidently represent some probably normal phase in the
life-history of the flagellated chambers, and it appears to
me uot improbable that they may be chambers in process of
dying. )

That the Sycon chamber, as an individual, can die, is perhaps
a somewhat novel idea; but if, as everyone will admit, an
Ascon individual dies, there is no difficulty in supposing that
a single chamber of a Sycon, which in many respects corre-
sponds to an entire Ascon, should also die. Indeed, it
is evident from the ontogeny of Grantia labyrinthica
(vide infra) that as the stalk develops the first formed
chambers must perish (fig. 27); and if so, why not indi-
vidual chambers later on? MHaeckel, in his work on the
‘ Challenger Deep-sea Keratosa’ (12), suggests that the flagel-
lated chamber is to be regarded as the individual, comparable
to an individual person of a hydroid colony; and in accord-
ance with this view we may certainly expect to find individual
chambers perishing, while the sponge as a whole continues to
exist healthily.

In the cases alluded to (fig. 29, x), the skeleton of what I
believe to have been an originally normal and healthy chamber
is still present in exactly its normal position and enclosing
the normal space. This space, however, is no longer com-
pletely filled by the chamber, but the latter has shrunk away
from the surrounding tubar skeleton into the centre, where it
occupies little more than a third of its original diameter. The
remains of the collared cells are distinctly visible, closely,
if not exactly, resembling the collared cells which line the
surrounding chambers, only less regularly arranged and in
more than one layer. In the process of shrinking the meso-

VOL. XXXII, PART I.—-NEW SER. B

18 ARTHUR DENDY.

derm around the chamber has been pulled out into delicate
strands forming a kind of network, but mostly radially dis-
posed, and thus serving to suspend the dying or dead chamber
as shown in the figure.

Fig. 28 (#@) shows what is perhaps a later stage in events.
The chamber occupies a still smaller space, and the surrounding
mesoderm has become solid and homogeneous again. The
arrangement of the spicuies and of the surrounding chambers
still indicates the space originally occupied by the dying
chamber.

If the individual chambers die it is probable that they are
replaced by new chambers ; and, indeed, I shall give reasons
later on, in describing the exhalant canals, for supposing that
new chambers are actually interpolated between the old ones.
Thus the older parts of the sponge may be kept alive and
vigorous by the gradual replacement of the old flagellated
chambers, as they reach their limits of existence and die off,
by new ones. I have unfortunately found no evidence to show
how the new chambers originate, but since the older flagellated
chambers frequently branch it is not unlikely that they may
also bud, or the new chambers may be developed from ameeboid
cells as in the embryo of Stelospongos (2). The problem
is on much the same footing as the question, how are new
chambers constantly added around the margin of the growing
sponge-cup? and, so far as I am aware, no one knows. All I
can say is that they commence life very small, and gradually
increase in size as they grow older (figs. 25, 27) ; they make
their first appearance in about the middle of the thickness of
the sponge-wall, and apparently do not originate as out-
growths of the gastral cavity.

Another explanation of the unusual condition of the flagel-
lated chambers described above is suggested by some observa-
tions of Sollas, in his report on the “ Challenger” Tetracti-
nellida (18), to the effect that the walls of the flagellated
chambers in this group sometimes appear contracted, under
which condition ‘fine filaments may be frequently observed
produced from the base of the choanocytes and extending

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 19

radially from the chambers into the surrounding matrix.”
“The whole appearance is suggestive of a contraction of the
choanocytal wall under the influence of some strong stimulus,
possibly of the alcohol into which the sponge was plunged on
removal from the dredge.”

Doubtless the unusual appearances of the chambers observed
by Sollas and myself are in both cases due to contraction ; and
had I seen only such cases as that represented in fig. 29, v, I
should have been strongly inclined to regard the contraction
either as a merely temporary condition or as a post-mortem
condition, produced, as suggested by Sollas, by the action of
the alcohol. The appearances presented in fig. 28, however,
in which the gelatinous matrix around the chamber has regained
its normal condition, while the chamber is more contracted
than ever, seem to me to demand a different explanation,
which I have endeavoured to give above.

The Exhalant Canals.

Since the flagellated chambers do not extend to the actual
gastral surface, but are separated therefrom by the entire
thickness of the gastral cortex (fig. 26), the existence of special
exhalant canals becomes necessary in order to place the
chambers in communication with the gastral cavity. These
exhalant canals are short, wide, cylindrical tubes, sharply
marked off by the sphincter diaphragms already described
from the chambers at the one end, and opening directly, with-
out any narrowing or diaphragm, into the gastral cavity at the
other (fig. 26).

I have already had occasion to mention that sometimes an
unusually small exhalant canal is met with, cut transversely,
in tangential sections of the cup wall a little below the level of
the ordinary exhalant canals. Such small canals are surrounded
by quadriradiate spicules of a slightly unusual form (fig. 21).
I am at a loss to explain the existence of these smaller canals,
with their slightly peculiar spicules, unless they be simply
the exhalant canals of young interpolated flagellated chambers,
surrounded by young spicules. This view is supported by the

20 ARTHUR DENDY.

fact that the spicules in question are somewhat smaller than
the more ordinary quadriradiates.

The Gastral Cavity and Osculum.

As already pointed 6ut in my description of the external
form of the sponge, the gastral cavity and osculum are greatly _
modified by the peculiar mode of growth of the sponge. The
gastral cavity, instead of being narrow and tubular, has become
wide and basin-like, and at the same time, owing to the con-
volutions of its wall, extremely irregular. The osculum has
thus become wider than any other part of the gastral cavity—
a condition the opposite of that which obtains in other Sycon
sponges.

Sometimes traces of the gastral cavity may be found in the
stalk even of adult sponges, causing the latter to be more or
less hollow. This indicates that the gastral cavity originally
extended all through the sponge—a fact which is proved, as I
shall show later on, by the ontogeny.

(ec) The histology of the soft tissues.

The terms ectosome and choanosome, proposed by Sollas
(15) and adopted by myself (2) in describing siliceous and
horny sponges, are not convenient for at any rate the great
majority of the Calcarea, and it is better to classify the tissues
simply under the heads ectoderm, mesoderm, and endoderm.
I must follow the example of Schulze (14) in considering that
the ectoderm of the larval sponge (in the case of the Sycons, at
any rate) furnishes not only the epithelium of the dermal
surface, but also the epithelial lining of the inhalant canal
system ; while the endoderm lines the remainder of the canal
system from the prosopyles to the margin of the osculum,
and the mesoderm furnishes all the remainder of the sponge
body.

The Ectoderm.

The ectoderm resembles exactly what Schulze has described
(11) in Sycandra raphanus, consisting of a single layer of
flat, polygonal epithelial cells lining the dermal surface of the
sponge and the inhalant canal system. These cells are most

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 21

readily distinguished around the inhalant canals, where they
are less obscured by spicules and other mesodermal structures
than on the dermal surface. The nucleus is surrounded by
the very characteristic granules described by Schulze in
Sycandra. In my preparations I have only after some
trouble succeeded in making out the boundary lines between
the individual cells, and Schulze himself observes that it is
remarkable that the boundaries of these cells—sometimes so
distinct—are not always clearly visible. Nevertheless I have
been able to determine the shape of the cells pretty accurately,
and found them to agree precisely with Schulze’s drawings.

The Endoderm.

This consists, as in other Heteroczla, of two parts: (1) a
layer of flattened epithelial cells lining the gastral cavity and
the short exhalant canals of the chambers; (2) a layer of
collared cells lining the flagellated chambers themselves. The
epithelial portion of the endoderm exhibits no features of
special interest and needs no further description, so we may
pass on at once to the collared cells.

Dr. R. von Lendenfeld has recently (16) called in question
the accuracy of my description of the collared cells, with their
connecting membrane, in Stelospongos (2), observing, “ Es
ist jedoch seine schematische Darstellung dieser Membran
(Taf. xxxii, fig. 9) keineswegs Vertrauen-einfléssend, sondern
eher ein Beweis der theoretischen Unwahrscheinlichkeit der
Existenz derselben.’? Notwithstanding this criticism, I still
maintain the correctness of my original description and figures,
and have already published a note (17) in the ‘ Zoologischer
Anzeiger’ in reply to Dr. von Lendenfeld’s observations. The
latter are apparently based partly on imperfect observation,
and partly on the convenient, albeit somewhat unphilosophical,
assumption that all sponges must be exactly alike in this
respect. Dr. von Lendenfeld finds that in certain sponges
examined by him “der Raum zwischen den Kragenzellen
von einer durchsichtigen, der gewohnlichen Grundsubstanz
der Zwischenschicht der Spongien sehr ihnlichen Substanz

ae, ARTHUR DENDY.

ausgefallt sei; and the only conclusion at which he is able
to arrive with regard to the question of Sollas’s membrane
is that it does not exist, and that Professor Sollas and I
have only misinterpreted what it has been reserved for him
to correctly describe. But in spite even of a preconceived
“ theoretische Unwahrscheinlichkeit ” I adhere to my original
opinion. For my own part, I am unable to see where the
theoretical improbability comes in. Quite recently Mr. Carter
has, in a private letter, afforded me valuable corroborative
evidence of the existence of Sollas’s membrane. He says,
“T have seen in the brim of the collar of the Calcisponge
spongozoon plastic amalgamation like that produced by two
semi-fluid bits of gum—under which circumstances, if all
became amalgamated, then you would have Sollas’s membrane.
Might it not so happen that at one time they may be so
amalgamated and at another not, and thus produce the dif-
ference?’’ That Sollas’s membrane originated in the almost
accidental manner here indicated there can be no doubt, but I
am inclined to think that in many sponges it has become a
more or less fixed and constant character—a view supported
by the fact that, as I shall show later on, it is still recognisable
when both collars and flagella are withdrawn. It is exceed-
ingly likely from the nature of the case that it may have
originated independently in several groups, so that in each
group forms with and forms without it may exist. If so it is
only another instance of that homoplasy so characteristic of
the Porifera.

I am not aware that there is anything particularly new in
Dr. von Lendenfeld’s observation that ‘‘die Kragenzellen
stehen nicht frei auf der Oberfliche der Zwischenschicht,
sondern sie sind in dieselbe Eingesenkt” (16). Indeed, in
my paper on Stelospongos (2) I have said, ‘“‘I have not
been able to trace any definite outline to the body of the cell
which is embedded in the highly granular ground-substance.”
It is amusing to see Dr. von Lendenfeld so vigorously opposing
one of my observations which does not happen to fit in with
his idea of the fitness of things, and at the same time taking

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 23

another from my very next page and putting it forward as
though it were original.

Sollas’s membrane, however, and the intercellular substance
which exists between the bases (and, so far as I have seen,
between the bases only) of the collared cells have nothing
whatever to do with one another. The former is endodermal
in origin, and is separated by a wide empty space from the
probably mesodermal ground-substance between the collared
cells. In the case of Stelospongos, if the mesodermal
ground-substance really filled up the whole of the intervals
between the collared cells it would be at once recognisable
by its highly granular appearance ; but it does not, as a glance
at my figures will show ; it stops at the bottom of the neck.

I must now describe the condition of things in Grantia
labyrinthica. All the collars and flagella of the collared
cells are retracted in my preparations. This is not to be re-
garded as a purely artificial and post-mortem condition, but
probably rather as a periodically recurring phase in the life-
history of the cells. Carter has shown long since (18) that the
individual collared cells may become ameeboid, and probably
in the living sponge they often spontaneously retract their
collars and flagella and enjoy a period of rest.

In this retracted condition the collared cells (figs. 382, 33) of
Grantia labyrinthica are somewhat pyramidal bodies, poly-
gonal in transverse section, and with the narrow end pointing
towards the lumen of the chamber, They measure about
0:0048 mm. in height and about the same in breadth at the
base. The nucleus is situate in the apex of the pyramid (fig. 33).
This position of the nucleus appears at first sight a little curious,
but it is interesting to observe that it agrees with the position
of the nucleus in the long prismatic cells of the embryo (fig. 38)
from which the collared cells of the adult are admittedly derived.
Thus the collared cells in a state of rest revert more or less to
their embryonic condition, the chief distinction being that they
are now very much shorter.

Even at their bases the collared cells appear to be separated
from one another by distinct intervals (fig. 32), but these may

24 ARTHUR DENDY.

possibly be due to shrinkage. The apices of the cells are still
further apart, and in longitudinal sections (fig. 33) are seen to
be connected by a fine, sharp line running from one to the
other. This line is Sollas’s membrane seen in section, no
longer supported on the tops of the collars, which have been
retracted, drawing the membrane after them close down into
the apices of the cells. So Sollas’s membrane remains visible
even when the collars of the cells are retracted, which indicates
that it is probably a more or less permanent structure, and no
mere temporary fusion of the margins of adjacent collars.

Owing to the great transparency of the gelatinous meso-
dermal ground-substance—which is a very characteristic feature
of calcareous sponges—it is not possible to determine, as in the
case of Stelospongos, exactly how far it extends between the
collared cells. Probably there is considerable individual varia-
tion in this respect, and it is a matter of little importance to
ascertain exactly how far each cell happens to be sunk in the
matrix. I cannot believe, however, that the actual collars are
ever embedded.

T understand that Mr. Bidder has already pointed out that
Sollas’s membrane occurs in a calcareous sponge, Leuconia
aspera, but I have neither been able to see his paper nor to
find out where it was published.

The Mesoderm.

The constituents of the mesoderm may be divided into two
main classes, the cells and the cell-products. We will con-
sider the cell-products first ; they consist of the intercellular
ground-substance and the spicules, and as the spicules have
been already fully described we have only to deal with the
ground-substance. The ground-substance' consists of the
usual transparent jelly, exhibiting no differentiation excepting
a slight concentration around the spicules, forming the so-called
spicule sheaths. These are very distinct, and are always seen
to be continuous with the surrounding ground-substance
when the spicules themselves have been dissolved out by the

1 Maltha of Haeckel (12),

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 25

action of weak hydrochloric acid. This view regarding the
nature of the spicule sheaths has been already expressed by
Haeckel and Schulze (11), and is doubtless the correct one.

The mesodermal cells, which lie embedded in the ground-
substance, may be classified as follows :

(1) Ameeboid.

(2) Stellate.

(3) Glandular:

(a) Spicule-secreting.
(6) Slime- or cuticle-secreting.

(4) Endothelial.

(5) Muscular.

(6) Nervous.

(7) Reproductive.

Ameboid Cells.—Concerning the ameeboid cells proper I
have no information to add to that which we already possess.
They are distinguished from the stellate cells by their more
rounded and massive form, and their more abundant and more
granular protoplasm. Certain of them, as is well known,
develop into ova, and these I shall describe later on.

Stellate Cells.—These, which may be regarded as the con-
nective-tissue cells of the sponge body, have also the usual
form, characterised by the long, slender, and often branched
processes given off from an inconspicuous central mass of
protoplasm surrounding the nucleus (fig. 26, sé. c.). It is very
probable that, as in the case of Stelospongos (2) and other
sponges, adjacent stellate cells may be united by their slender
processes, but I have not succeeded in clearly demonstrating
the connection here.

Glandular Cells.—These are of two kinds—spicule-secret-
ing, and slime- or cuticle-secreting cells. We will consider the
spicule-secreting cells, or calcoblasts, as they have been termed
by Poléjaeff (8), first.

It is generally admitted that both calcareous and siliceous
spicules originate within special mother-cells, but probably in
both cases they subsequently receive additional layers of the
calcareous or siliceous material from other cells. Poléjaeff (8)

26 ARTHUR DENDY.

figures a “‘ conjectural calcoblast ”’ attached to the outside of a
spicule of Leuconia multiformis. This cell has the form
of an ordinary stellate mesodermal cell, and such, so far as
structure is concerned, I believe the calcoblast to be. In the
horny sponges the corresponding spongoblasts, although some-
what specialized in form, are clearly only slight modifications
of stellate cells, as I have elsewhere shown (2). In the cal-
careous sponges the calcoblasts have acquired the function of
secreting carbonate of lime without undergoing any correspond-
ing modification in form. It should, however, be borne in
mind that there are probably two kinds of calcoblasts, primary
and secondary. The primary calcoblasts are the mother-cells
in which the spicules originate, and the secondary calcoblasts
are the cells which secrete additionai layers of calcareous matter
around the spicule after it has been formed. In the same way I
have no doubt that there are primary and secondary silicoblasts.+

The slime- or cuticle-secreting cells have, so far as I am
aware, not hitherto been observed in calcareous sponges,
although well known in the Keratosa through the researches of
von Lendenfeld. In Grantia labyrinthica the gland-cells
in question occur in a single layer just beneath the epithelium
of the surface of the sponge. They are very distinct and
plentiful on the gastral surface (fig. 26, gl. c.), but less so on
the dermal. Each gland-cell consists of an irregular, granular,
nucleated body, which closely resembles an ordinary ameeboid
cell, but which may be seen under favorable conditions to be
connected with the overlying epithelium by one or more pro-
cesses. Fig. 26 shows the layer of gland-cells beneath the
gastral epithelium, and fig. 34 is a more highly magnified
drawing of an individual gland-cell and its surroundings, as it
appeared under a Zeiss F objective and ocular 4, At the par-
ticular spot figured an irregularity in the surface caused the
epithelium to be cut somewhat tangentially, while the gland-
cell itself was cut vertically ; the connection between the two
is, however, very well shown. Four processes are shown in this

1 For an illustration of a primary silicoblast the student is referred to the
‘Report on the “Challenger” Monaxonida,’ pl. xxi, fig. 13.

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 27

instance connecting the gland-cell with the epithelium, but it is
very unusual to see so many. Just on the right of the gland-
cell the epithelium is raised up by a projecting spicule which
does not itself appear in the section. On the dermal surface
of the sponge the gland-cells are more like ordinary amceboid
cells immediately underlying the epidermis.

The essential agreement of these cells with those described
by von Lendenfeld in, for example, Dendrilla cavernosa
(19) is obvious, both as regards structure and arrangement.
According to this author the cells secrete a cuticle which is not
to be distinguished from the spongin of the horny fibres. I
agree that they probably secrete a cuticle, but, as Vosmaer
points ont (20), no grounds are given for supposing that this is
identical with the spongin, and the fact that similar gland-cells
occur in the Calcisponges argues against the assumption.
Very possibly the cuticle, after a time, more or less entirely
replaces the original epithelium, and not only, as suggested by
von Lendenfeld,! so much of it as may have been accidentally
damaged. Hence, perhaps, arises the difficulty of making
out the structure of the surface epithelium.

Endothelial Cells.—The embryos, which I have described
elsewhere (10), lie each in a separate cavity in the mesoderm
around the flagellated chambers. These embryo-containing
cavities (fig. 38, em. c.) are lined each by a single layer of
pavement-cells, which are not to be distinguished in my
sections from the pavement-cells already described. These
cells I regard with Schulze (11) as being of mesodermal origin,
and hence endothelial.

In my memoir on Stelospongos flabelliformis (2) I
regarded the embryo-containing cavities in that sponge as pro-
bably, though by no means certainly, specialized parts of the
exhalant canal system. As the result of further study, how-
ever, I must relinquish this view, and regard the cavities as
special excavations in the mesoderm, and the remarkable giant
pavement-cells which line them as of mesodermal origin.

1 For an abstract of von Lendenfeld’s observations on the subject
vide 20.

28 ARTHUR DENDY.

Muscular Cells.—Certain spindle-shaped cells lying in the
membranous diaphragms of the exhalant apertures of the
flagellated chambers are probably muscular in function. The
cells in question are shown in fig. 24, which renders further
description unnecessary.

Nerve-cells.— The only cells which I have found in
Grantia labyrinthica to which a nervous function can
possibly be assigned are certain structures which occur around
the margins of the inhalant pores, as shown in fig. 31.
These cells are elongated radially in relation to the circular
pores which they surround. The nucleus is distinct and is
placed at the distal end of the cell, and the main mass of
protoplasm stretches from the nucleus to the edge of the pore,
where it ends in an expansion along the free margin. There
may also be indications of smaller processes given off near the
base of the cell. It is probable that the thickening of the
main protoplasmic process of each cell as it touches the margin
of the pore may simply indicate a retracted, sensitive, hair-like
process such as Stewart (21) and Lendenfeld (22) describe ; but
it seems also just possible that each nerve-cell naturally ends
in a sensitive plate or expansion at the free margin of the pore.
If we adopt the former of these two views the sensitive cells
will be seen to agree pretty closely in structure with those de-
scribed and figured by Stewart in Grantia compressa; but
I have met with no evidence of the grouping of the cells into
“ synocils,” as described and figured by von Lendenfeld. Von
Lendenfeld, however, has also described single sensitive cells
(“‘Sinnes-Ganglienzelle”’) around the inhalant pores of his
Chalinissa communis, var. flabellum (23), which in struc-
ture and arrangement almost exactly agree with those found by
mein Grantia labyrinthica except that he figures the end of
the main protoplasmic process projecting for a short distance
beyond the margin of the pore. This slight difference may, as
will be gathered from what I have already said, be due to dif-
ferences in state of contraction in the two cases. It is interest-
ing to note that the sense-cells of Grantia labyrinthica
agree more closely with those of so different a sponge as

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 29

Chalinissa communis than they do with those of the closely
related Grantia compressa. We must associate with this
fact the fact that the arrangement of the pores in Grantia
labyrinthica also agrees much more closely with that which
usually obtains in the Chalinine than with that found in
Grantia compressa, thus affording another instance of
homoplasy.

The cells which Sollas (18) describes and figures as possible
sense-cells (“‘ zesthocytes ”) in the Tetractinellida appear to be
of a more problematical character.

Reproductive Cells.—The only reproductive cells which
I have met with are the ova. ‘These, as in other sponges, are
obviously derived from ameeboid cells, and in the earliest
stages of their development it is impossible to distinguish
between the two. later, however, the ova become more
rounded off, and the nucleus becomes large and distinct.

It is now generally admitted that the ova of sponges are
fertilised by spermatozoa probably of other sponges, which
gain admission to the sponge with the inflowing current of
water. Noone, however, so far as I can discover, has attempted
to find out whereabouts in the sponge the union of ovum and
spermatozoon takes place. Having followed the spermatozoon
into the canal system they leave it there to take care of itself,
forgetting that, unless some special arrangement exists to
prevent such a catastrophe, it will speedily be washed out
again through the osculum without ever having had a chance
of fulfilling its errand. The general assumption would seem
to be that the spermatozoon loses its way through the walls of
the canals, and wanders about in the gelatinous mesoderm
until it happens to come across an ovum. It seems highly
improbable that this should be the case, for it would be a
strange thing if the spermatozoa bored their way through the
epithelium, as they would have to do in order to get into the
gelatinous ground-substance, without any obvious inducement
todo so. Mr. Carter seems to have come nearer to the truth
than anyone, but without realising the true significance of
what he saw. He says (24) that the ovum in Grantia com-

30 ARTHUR DENDY.

pressa ‘“‘ may be seen to be hanging, pear-shaped, upon the
surface of the excretory canals, where it remains for a certain
time locomotive, until, after further development, it becomes
permanently fixed, and the locomotive envelope seems to pass
into a capsule.” It is obvious that by ‘‘ excretory canals ”
Mr. Carter means the inhalant canals, and he is probably
wrong in considering that the locomotive envelope of the ovum
passes into a capsule; but the main fact, that the ovum ata
certain stage of its existence hangs freely from the surface of
a portion of the canal system, is clearly brought out.

In Grantia labyrinthica I have distinctly observed the
ova hanging from the epithelial lining of the inhalant canals
by means of short peduncles, and projecting freely into the
lumen of the canal, where they must be washed by the incoming
stream of water. Fig. 35 represents an amceboid ovum
approaching the surface of an inhalant canal; and figs. 36
and 37 represent it, after having passed through the epithe-
lium, hanging freely by its short peduncle, awaiting ferti-
lisation. In fig. 37 the nucleus of the ovum appears very
near the surface, in a position suggestive of the formation of
a polar body. Probably the ova pass through the epithelium
of the inhalant canals in the same way that the white blood-
corpuscles pierce the walls of the capillaries in higher animals.

After fertilisation the ovum probably migrates back into the
gelatinous ground-substance, and takes up its position near the
wall of aflagellated chamber, there to undergo the earlier stages
of its development. This seems more probable, from what we
know (10) of the position of the embryos, than the supposition
that it remains and develops in the spot where it is fertilised.

It is probably a general rule in sponges that the ova are
fertilised while hanging from the walls of the canal system,
and that they migrate first of all through the canal-wall to be
fertilised, and then back again into the gelatinous ground-
substance to undergo development ; hence the necessity for
the amceeboid movements so characteristic of sponge ova. Thus
we see that the sponge ovum plays an unusually active part in
the process of fertilisation, as it were meeting the spermatozoon

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 81

halfway. The migrations of the ova in sponges remind one
forcibly of Weissmann’s descriptions of the migrations of the
ova in various hydroids (25).

The Post-embryonic Development of Grantia
labyrinthica.

The embryology of Grantia labyrinthica, so far as the
material at my disposal has allowed me to work it out, forms
the subject of a special memoir (10), so that it is unnecessary
for me to deal with the question in this place. The post-
embryonic development, however, has not as yet been dealt
with, and although there is but little to be said about it, yet
that little is of interest as clearly indicating the manner in
which the very peculiar external form has been evolved.

Fig. 1 shows the youngest stage found, in which it will be
seen that, as regards both size and shape, the sponge differs
but little from an ordinary Grantia. The wall of the tube
is even and not convoluted, although already the osculum is
the widest part of the gastral cavity. Fig. 27 represents
somewhat diagrammatically a longitudinal section of this
specimen, and is important chiefly because it shows how the
stalk arises by the filling up of the lower portion of the gastral
cavity with a copious growth of mesodermal tissue in which
numerous spicules are developed. As I have already pointed
out, remnants of the gastral cavity may sometimes be recog-
nised in the stem even of adult specimens. Figs. 2 and 3
represent two stages intermediate between that just described
and the full-grown sponge (fig. 4). The specimen drawn in
fig. 8 is very much compressed, so as to be bilaterally sym-
metrical. This may either be accidental, or it may indicate
the commencement of folding in the wall of the cup. All
figures are drawn of the natural size, and they render further
description needless.

I may conveniently here describe the only case of budding
with which I have met in the species. On the outer surface
of the wall of the cup in a well-grown specimen, and not
far below the margin, a small individual had budded off

oe ARTHUR DENDY.

from the parent sponge. This is represented, twice the natural
size, in fig. 5. The gastral cavity of the young sponge is still
connected with that of the mother by a very evident aperture,
and it is interesting to notice that in this case the osculum is
distinctly constricted, so that the bud has the typical Sycon
form.

The So-called Family Teichonide.

The very artificial group of sponges now generally known
under the name Teichonide has been unfortunate from first
to last. It was not even christened properly, for the title of
Mr. Carter’s original paper (8) is printed “ On Teichonia, a
New Family of Calcareous Sponges, with Descriptions of Two
Species ;” while subsequently, on the same page, the name
Teichonellide is given to the new family, which is concisely
diagnosed as “vallate.” Then the genus Teichonella is
diagnosed as follows: ‘“ Vallate or foliate, without cloaca.
Vents numerous, confined to the margin or general on one side
of the lamina only; naked.”

Teichonella prolifera is described as the first species of
the genus, and in the same paper Grantia labyrinthica is
described as a second species. Subsequently, as I have already
said, Mr. Carter withdrew Grantia labyrinthica from the
genus Teichonella, and recognised its true position amongst
the Sycons.

Now we see upon what slight foundations spongologists
build their edifices. Poléjaeff (8) adopts the new family,
altering the name, however, to Teichonide. He states that
the main character of the family consists in the differentiation
of the outer surface into two planes, one bearing oscula, the
other pores exclusively ; and he enters into a curious specula-
tion as to whether a Teichonid is “a colony with dislocated
oscula and pores,” whatever that may be. But what authority
had Poléjaeff for stating that the outer surface was differen-
tiated into two planes, one bearing oscula and the other pores ?
Mr. Carter never said so. His diagnosis of the family is
“ vallate,” while even his generic diagnosis says nothing about

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 35

the pores. In the description of the type species (Teichonella
prolifera), however, we find the words “ pores invisible to
the naked eye, scattered over the surface thickly, and generally
vents slightly marginated, . . . arranged more or less in a
single line along the margin only.” I have myself most care-
fully examined Teichonella prolifera by means of stained
sections cut by the paraffin method, and I find that it is
nothing but an ordinary Leucon, with pores on both surfaces
of the low, thick walls of which the sponge consists, and oscula
along the raised margin (fig. 6). The surface is certainly not
differentiated into anything like a pore-bearing and an osculum-
bearing plane.

The fact appears to be that Poléjaeff wanted a place for his
new genus Hilhardia, and so he seized upon the Teichonel-
lide and altered both the name and the characters of the
family to suit his own ideas, apparently without having so
much as ever seen a specimen of Teichonella. Of course,
both Teichonella and Hilhardia duly appear in an ela-
borate genealogical tree.

Eilhardia, if Poléjaeff’s figures be correct, is a Leucon.
True, the pores appear to be on one surface and the oscula on
the other, but can anyone possessed of the slightest know-
ledge of the subject regard this as a family character? I
think certainly not, and in support of my opinion venture to
call attention to the following extract from Ridley and Dendy’s
report on the “Challenger” Monaxonida (26) :—“ Before
leaving the question of the pores we must consider briefly the
condition of flabellate sponges in this respect. It is an almost
invariable rule that in flabellate sponges the pores are to be
found on one surface and the oscula on the other. Thus in
Phakellia ventilabrum, var. connexiva (pl. xxxv, figs.
3, 3a; pl. xlix, fig. 3), and Phakellia flabellata, nobis
(pl. xxxiv, figs. 2, 3, 3a), this arrangement is very well illus-
trated ; and the same condition occurs in Myxilla frondosa,
nobis (pl. xxvi, figs. 1, la), and Gellius flabelliformis,
nobis (pl. xxvi, figs. 5,5a). Again, in that very remarkable
sponge, Esperiopsis Challengeri (pl. xviti), the pores

VOL. XXXIJ, PART IL.—NEW SER. ©

34. ARTHUR DENDY.

occur only on the concave surfaces of the lamelle (pl. xviii,
fig. 4), while the oscula are all on the convex surfaces.

“* By far the most remarkable instance of this kind is, how-
ever, afforded by a boring Suberite which we have described
under the name Cliona dissimilis (p. 227, pl. xxv, fig. 5,
&c.). Here the sponge has bored its way into a flattened
coral which it completely surrounds ; hence it has itself acquired
a flattened, lamellar form, and we find the pores collected in
areas (woodcut, fig. 11, pa.) on one side of the sponge, and the
oscula (woodcut, fig. 11, 0.) on the other.’

«There is no other known example, so far as we are aware,
of a lamellar Suberitid sponge ; and even the species in question
is lamellar only because it has bored into a lamellar coral, and
yet the pores and oscula are arranged just as they would be in
a free-living frondose sponge, suchas Phakellia. There must
be some strong reason why as soon as a sponge, for any cause,
acquires a lamellar form, the oscula become confined to one
surface and the pores to the other, and to account for the occur-
rence of this condition in genera so widely separated as
Gellius, Myxilla, Phakellia, and Cliona, What this
reason may be we cannot at present say.”

I am still no better able to give an explanation of this
curious phenomenon than I was when the above passage was
written ; but the facts appear to me to be conclusive evidence
against the value of the peculiar arrangement of the pores and
oscula as a family character.

But even if it were allowed that the arrangement of the
pores and oscula were a character of family importance we
could not put Hilhardia and Teichonella in the same
family, for, as I have shown, they differ widely from one
another in this respect. Then, according to Poléjaeff’s dia-
gnosis, not Hilhardia, but Teichonella, would have to
come out of the family Teichonide. As a matter of fact the
family ought to be abandoned altogether, and the three species
which have been at various times placed in it distributed as
follows ;

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 395

Teichonella prolifera . ; : . Leuconide.
Hilhardia Schulzei : : : . Leuconide.
Grantia labyrinthica ~< : 5 - Syconide.

It would not have been necessary to deal with this question
so carefully had not Poléjaeff’s emended family Teichonidez
met with such general and unquestioning acceptance. Thus
Vosmaer adopts it in his most important work (20), and Len-
denfeld (6) gives it a place in his system and in the inevitable
genealogical tree. Haeckel also accepts the family in his
latest work on sponges (12).

In conclusion I wish again to record my great indebtedness
to Professor Howes for his kindness in correcting the proofs
of this paper in my absence from England.

List oF MEMOIRS REFERRED TO.

1. Carter, H. J.—‘ Descriptions of Sponges from the Neighbourhood of
Port Phillip Heads, South Australia,” ‘Ann. and Mag. Nat. Hist.,’
1886, et seq.

2. Denpy, AnTHUR.—*“ Studies on the Comparative Anatomy of Sponges.
II.—On the Anatomy and Histology of Stelospongos flabelli-
formis, Carter; with Notes on the Development,” ‘Quart. Journ,
Mier. Sci.,’ December, 1888.

8. Carter, H. J.—‘ On Teichonia, a New Family of Calcareous Sponges,
with Descriptions of Two Species,” ‘Ann. and Mag. Nat. Hist.,’
1878.

4. Canter, H. J.—“ Mode of Circulation in the Spongida,’” ‘Ann, and
Mag. Nat. Hist.,’ 1885.

5. Carter, H. J.—“‘On the Position of the Ampullaceous Sac and the
Function of the Water Canal System in the Spongida,” ‘ Ann. and
Mag. Nat. Hist.,’ 1887.

6. LENDENFELD, R. von.—‘ A Monograph of the Horny Sponges,’ Royal
Society of London, 1889.

. Harcxet, Ernst.—‘ Die Kalkschwamme.’

8. Pottsarrr, N.—‘ Report on the Calcarea collected by H.M.S. ‘ Chal-
lenger” during the Years 1873—1876.’

9. Sontas, W. J.—‘‘ Sponges,” ‘ Encyclopedia Britannica,’ ed, ix,

~J

36

10.

11.

12.

13.

14.

15.

16.

aT:

18.

19.

20.

21.

22.

23.

24.

25.
26.

ARTHUR DENDY.

Denpy, ArtHuR.—“ On the Pseudogastrula Stage in the Development
of Calcareous Sponges,” ‘ Proceedings of the Royal Society of
Victoria’ for 1889.

Scuuuze, F. E— Ueber den Bau und die Entwicklung von Sycandra
raphanus, Haeckel,” ‘ Zeitschrift fiir wissensch. Zoologie,’ xxv
Band, Suppl.

HakckeEL, Ernst.—‘ Report on the Deep-sea Keratosa collected by
H.M.S. “Challenger” during the Years 1873—1876.’

Sortas, W. J.—‘ Report on the Tetractinellida collected by H.M.S.
* Challenger” during the Years 1878—1876.’

Scuvuze, F. E.—‘‘ Untersuchungen ueber den Bau und die Entwicklung
der Spongien. IX.—Die Plakiniden,” ‘ Zeitschrift fiir wissensch.
Zoologie,’ xxxiv Band.

Sottas, W. J.—“ Preliminary Account of the Tetractinellid Sponges
dredged by H.M:S. “ Challenger,” 1872-6,” ‘ Sci. Proc. Royal Dublin
Soc.,’ vol. v, pt. vi. :

LENDENFELD, R. von.—“ Notiz tiber den Bau der Geisselkammer der
Spongien,” ‘ Zoologischer Anzeiger,’ No. 311, 1889.

Denpy, ArTHUR.—‘“ Some Old and New Questions concerning Sponges,”
‘Zoologischer Anzeiger,’ No. 325, 1890.

Carter, H. J.—“ Notes Introductory to the Study and Classification of
the Spongida. Part I.—Anatomy and Physiology,” ‘Ann. and Mag.
Nat. Hist.,’ 1875.

LENDENFELD, R. von.— Studies on Sponges. I.—The Vestibule of
Dendrilla cavernosa, Nova Species,” ‘Proc. Linn. Soc. New
South Wales,’ vol. x, part 4.

VosmaER, G. C. J.—‘‘Spongien (Porifera),’ ‘Bronn’s Klassen und
Ordnungen des Thier-reichs,’ vol. ii.

Srewart, C.—“ Sense-cells in Sponges,” ‘ Bell’s Comparative Anatomy
and Physiology,’ p. 431.

LENnDENFELD, R. von.— Synocils, Sinnesorgane der Spongien,” ‘ Zoolo-
gischer Anzeiger,’ No. 16, 1887.

LENDENFELD, R. von.—‘ Die Chalineen des australischen Gebietes,”
‘ Zoologische Jahrbiicher,’ Band ii.

Carter, H. J.—“ Development of the Marine Sponges from the Earliest
Recognisable Appearance of the Ovum to the Perfected Individual,”
‘Ann. and Mag. Nat. Hist.,’ 1874.

Werissmann.—‘ Die Entstehung der Sexualzellen bei den Hydromedusen,’

Riptey, 8. O., and Denpy, A.—‘ Report on the Monaxonida collected
by H.M.S. “Challenger” during the Years 1873—1876

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 37

DESCRIPTION OF PLATES I—IV,

Illustrating Mr. Arthur Dendy’s paper “ Studies on the Com-
parative Anatomy of Sponges. III.—“On the Anatomy
of Grantia labyrinthica, Carter, and the So-called
Family Teichonide.”’

PEATE

Figs. 1, 2, 3.—Three early stages in the post-embryonic development of
Grantia labyrinthica. Nat. size.

Fic. 4.—Adult specimen of Grantia labyrinthica, growing upon a mass
of foreign matter. Nat. size.
- Fig. 5.—Portion of an adult specimen of Grantia labyrinthica, witha
bud attached to the outer surface. xX 2.

Fic. 6.—Specimen of Teichonella prolifera, showing the oscula dis-
tributed along the raised margin. Nat. size. . :

PLATE II.
SKELETON OF GRANTIA LABYRINTHICA.

Fics. 7, 8, 9.—Large linear spicules (oxea) from the margin of the osculum.
Zeiss, D, ocular 2, camera.

Fies. 10, 11.—Oxea from the surface of the cup. Zeiss, D, ocular 2,
camera.

Fig. 12.—Large oxeote spicule from the surface of the stem. Zeiss, D,
ocular 2, camera.

Fics. 13, 14, 15, 16, 17, 18.—Different forms of triradiate spicules from
the wall of the cup. Zeiss, D, ocular 2, camera.

Fic. 19.—Sagittal triradiate spicule from the stem. Zeiss, D, ocular 2,
camera. .

Fie. 20.—Quadriradiate spicule from the gastral cortex. Zeiss, D, ocular
2, camera.

Fic. 21.—Arrangement of quadriradiate spicules around the exhalant canal
of a young flagellated chamber.

Fic. 22.—Arrangement of the skeleton as seen in vertical longitudinal
section of a chamber from gastral to dermal surface.

PLATE III.

ANATOMY OF GRANTIA LABYRINTHICA,

Fic. 23.—A single flagellated chamber and its exhalant canal, in part laid
open by longitudinal section. ea. c. Exhalaut canal. di. Diaphragm. pr.

38 ARTHUR DENDY.

Prosopyle. ft. c. Cavity of flagellated chamber. em. Embryo. em. c. Embryo-
containing cavity (the embryo has already escaped in this case). ‘The collared
cells are coloured red, and the embryos brown. Zeiss, D), ocular 2.

Fic. 24.—Exhalant opening of a flagellated chamber, with its membranous
diaphragm (di.). mus. Muscle-cells. The spicules are coloured blue, and the
collared cells (which lie somewhat below the level of the diaphragm) red.
Zeiss, F, ocular 2.

Fic. 25.—Part of a section vertical to the margin and to the two surfaces
of the wall of the cup. em. Embryo. o. sp. Spicules of the oscular fringe.
p- Inhalant pore. y.a. Pore-area. iz.c. Inhalant canal. g.s. Gastral
skeleton. d.s. Dermal skeleton. 7.5. Tubarskeleton. Other lettering and
colouring as before. Zeiss, A, ocular 3.

Fie. 26.—Gastral portion of a thin section similar to the last, but more
highly magnified. g/. c. Gland-cells lying beneath the gastral epithelium.
st. c. Stellate cell. Other lettering and colouring as before. Zeiss, F,-
ocular 2.

Fic. 27.—Vertical section through the young specimen represented in Fig.
1. gas.c. Gastral cavity. mes. Growth of mesodermal tissue filled with
spicules, to form the stalk. Flagellated chambers red, spicules blue.

PLATE IV.
Awatomy oF GRANTIA LABYRINTHICA.

Fic. 28.—Portion of a section taken parallel to the surface of the sponge-
wall, and somewhat nearer to the dermal than to the gastral surface. It will
be noticed that the inhalant canals are still of large size. s. sp. Sections of
the shafts of the triradiate spicules of the tubar skeleton. z. Peculiarly
modified flagellated chamber. Other lettering and colouring as before. Zeiss,
D, ocular 2.

Fic. 29.—Portion of a section similar to the last, but taken near the gastral
surface, showing the diminution in diameter of the inhalant canals, &c.
Lettering and colouring as before. Zeiss, D, ocular 2.

Fic. 30.—Portion of the dermal surface sliced off, showing the pores
arranged in pore-areas. Some of the spicules are omitted. Lettering and
colouring as before. Zeiss, A, ocular 2.

Fig. 31.—A single pore more highly magnified, showing the nerve-cells
(m. c.) around its margin. Zeiss, F’, ocular 2.

Ftc. 32.—A group of collared cells (with retracted collars and flagella),
seen from above or below. #. Nucleus. Zeiss, F, ocular 3.

¥ie. 83.—A row of four collared cells (with retracted collars and flagella),
seen from the side. . Nucleus. s, m. Sollas’s membrane shrunk down
upon the apices of the cells. Zeiss, I’, ocular 3.

Fic. 34.—A single gland-cell (g/. c.) connected by four processes with the

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 389

overlying epithelium (ep.), which latter is cut almost tangentially owing to the
unevenness of the surface. =p. ep. Projection of the surface epithelium
around a protruding spicule, which latter is not itself seen in the section.
Spicules blue. Zeiss, F, ocular 4.

Fig. 35.—An ameboid ovum (ov.) lying in the gelatinous ground-substance
of the mesoderm, near the epithelium (ep.) of an inhalant canal. Zeiss, F,
ocular 3.

Fic. 36.—An ovum (ov.) hanging by a stalk from the epithelium (ep.) of an
inhalant canal. Zeiss, F, ocular 3.

Fie. 37.—Another example of an ovum (ov.) hanging by a stalk from the
epithelium (ep.) of an inhalant canal. Zeiss, F, ocular 3.

Fic. 38.—An embryo (approaching the pseudogastrula stage) lying within
the embryo-containing capsule in the mesoderm, between two flagellated
chambers of the mother sponge. col. Collared cells of mother sponge. sp.
Spicule of mother sponge. ed. Prismatic cells (future endoderm) of the
embryo. mes. Incipient mesoderm of the embryo. gr. Layer of large
granular cells of the embryo. Other lettering as before.

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STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 41

Studies on the Comparative Anatomy of
Sponges.

IV.—On the Flagellated Chambers and Ova of
Halichondria panicea.

By

Arthur Dendy, M.Se., F.L.S.,
Demonstrator and Assistant Lecturer in Biology and Fellow of Queen’s
College, in the University of Melbourne.

With Plate V.

THE purpose of the present short communication is, as indi-
cated in the title, to call attention to the minute structure of
the flagellated chambers and ova in the common British sponge
—Halichondria panicea.

The material upon which my observations are based I owe
chiefly to Mr. Martin Woodward, who kindly gave me some
fine pieces of the sponge in question, collected by himself in
the English Channel. Some of the material thus obtained
was treated while fresh with osmic acid, to which fact is due
the excellent state of preservation. Material not so preserved
I find to be practically valueless for the study of the histology
of the chambers, which, indeed, can only be made out where
the osmic acid has penetrated before the death of the sponge.

Part of the material was dredged by myself in the Solent,
and it was in sections of this that I first recognised the exist-
ence of Sollas’s membrane in the species.!_ The sections were

1 “ Studies on the Comparative Anatomy of Sponges,” II, ‘Quart. Journ,
Micr. Sci.,’? December, 1888.

42 ARTHUR DENDY.

stained with borax carmine, and cut by the ordinary paraffin
method.

I at one time hoped to be able to publish a complete account
of the minute anatomy of Halichondria panicea; but my
removal from England and the pressure of other work render
this impracticable, and I must content myself with describing
only the most important points. This is particularly de-
sirable, because I find in this sponge the best example with
which I have yet met of Sollas’s membrane, the very existence
of which has lately been called in question by Dr. R. von
Lendenfeld.!' I need not here reply to the arguments of this
observer, because I have already done so in my article on
“Some Old and New Questions concerning Sponges,” in the
‘Zoologischer Anzeiger,’* and more fully in the third of
my “Studies on the Comparative Anatomy of Sponges,” re-
cently sent to England for publication in the ‘ Quarterly
Journal of Microscopical Science.’ To these papers and to
the second of my “studies,” already cited, I must refer the
reader for further information, and for the literature of the
subject.

Halichondria panicea may be so readily obtained in
England, that any student of sponges acquainted with the
modern method of research can easily satisfy himself as to the
correctness of the observations here recorded.

Before describing the flagellated chambers of Hali-
chondria panicea it will be advisable to say a few words as
to the canal system. Thisisofthe lacunartype. The inhalant
pores, scattered over the surface of the sponge, lead into a
system of irregular lacune, scarcely definite enough to deserve
the name of canals. The ectosome is thin, and the subdermal
cavities are not recognisable as distinct structures. As they
penetrate below the surface, branching again and again, the in-
halant lacunz become smaller and smaller, being recognisable
in sections only as minute cavities surrounded by the flagel-
lated chambers. Interdigitating with the inhalant lacune in

1 «Zoologischer Anzeiger,’ No, 311, 1889, p. 362.
2 No. 325, 1890.

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 43

the most complicated and irregular manner we find the pre-
cisely similar exhalant lacune. The ultimate inhalant and
exhalant lacune are separated only by narrow strands of gela-
tinous mesodermal tissue, in which lie the spicules and the fla-
gellated chambers (fig. 1). Thus every chamber lies wedged
in between an inhalant lacuna on the one side, and an exha-
lant lacuna on the other; and it is only by noticing which
way the exhalant openings of the chambers are turned that it
is possible to tell whether a given lacuna is inhalant or exha-
lant. The exhalant lacune gradually unite together into
larger and larger channels, and open finally on to the surface
of the sponge by means of the wide oscula, generally, if not
always, passing first into well-defined oscular tubes.

The chambers themselves (fig. 1) may be roughly described
as subspherical. When, however, they are cut along the
imaginary axis running through the inhalant and exhalant
poles they frequently appear somewhat compressed, and we
see also that the exhalant opening (fig. 1, ¢, e. 0. c.) is very
wide. When cut in a direction at right angles to this axis
they appear circular in outline, and the wall of the chamber is
uninterrupted (fig. 1,a@). The diameter of the chamber in this
case is about 0:047 millimetre in length. (In the report of
the ‘Challenger’? Monaxonida the average diameter of the
chambers is given as 00336 millimetre. This figure appears
from my subsequent researches to be rather too low; but it
must be remembered that a certain amount of variation occurs
in this respect, in accordance with the state of preservation
of the specimens examined.)

We now come to the most important consideration, viz. the
form and arrangement of the collared cells; and we will first
describe them as they appear when the chamber is seen in
section (fig. 1, a,c). The collared cells stand some little dis-
tance apart from one another on the gelatinous ground-sub-
stance surrounding the chamber. They have each a short
nucleated body, indistinguishable from the collum or neck, and
surmounted by the delicate funnel-shaped collar. The outlines
of the collars are extremely fine, but all the collars are con-

44 ARTHUR DENDY.

nected at their margins by a very distinct membrane, which
appears in section as a thicker line running from one to the
other, but interrupted by the mouths of the collars, as shown
in the figure. The structure and relations of this membrane
(Sollas’s membrane) appear exactly as I have described them
in Stelospongos. The collared cells nearest to the exhalant
opening of the chamber are shorter than those farther away,
so that the membrane gradually approaches more and more
closely to the gelatinous ground-substance around the chamber,
and finally seems to run into it at the opening itself (fig.
1, c). The fact that Sollas’s membrane appears here as a
thicker line than the outlines of the collars is, I believe,
simply due to the thickness of the sections causing us to see a
little more than the mere cut edge of the membrane.

Professor Sollas, in his valuable ‘ Report on the Tetracti-
nellida of the Challenger Expedition,’ observes, “ I have never
yet seen the flagella of the concrescent choanocytes, though I
have never failed to find them in the case of choanocytes which
are not concrescent. It might be explained on the supposition
that the flagella are retracted in the former case; but that
naturally leads to the inquiry as to why they are not retracted
in the latter.” In Stelospongos I was also unable to detect
the flagella, but I expressed my belief in their existence in the
living sponge, and gave adiagram showing them. My researches
on Halichondria panicea fully justify this view. Fig. 1,
which is a careful drawing of an actual preparation, shows the
flagella plainly, projecting from the bodies of the collared cells
through the collars and into the cavity of the chamber (a).
Thus the question as to the coexistence or otherwise of Sollas’s
membrane and the flagella of the collared cells is settled.

If we now look down upon the wall of a chamber from the
outside, instead of examining it in section, another important
point is brought to light. ‘The collared cells, which, it will be
remembered, stand well apart from one another, are connected
at their bases by broad protoplasmic processes, so that the wall
of the chamber appears to be made up of a network of broad
protoplasmic strands with nuclei at the nodes of the net (fig.

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 45

1, 6). This fact has also been observed by Sollas in the case
of the Tetractinellida (op. cit.), and is well shown in his
diagram of the protoplasmic continuity of a sponge. I have
also seen and figured at z. in fig. 1 what appears to be a con-
nection of a collared cell with a stellate cell in the surrounding
mesoderm, as described by Sollas.

We have now to face the difficult question, ‘‘ How does the
water from the inhalant lacune enter the flagellated chambers?”
Sollas (op. cit.) observes, ‘‘ Since the fenestrated membrane
stretches across the flagellated chamber transversely there
must be an aperture or apertures in it for the passage of water
from the prosodus to the aphodus, though I have never suc-
ceeded in finding any; it is possible that the apertures have
the form of pores no larger than the lumen of the choanocytal
collars, and in this case they would be very difficult to dis-
tinguish.”

In spite of the excellent preservation of my sections I can
find in Halichondria panicea no apertures in Sollas’s mem-
brane to allow of the passage of the water, and I am inclined
to believe that either there are none, or that they exist merely
as temporary and not definite openings, which is quite possible,
The idea that there are no openings may seem at first sight
rather strange, but I believe it to be quite reasonable, if not
probable.

After having arrived at this point in the present paper I at
length obtained access to Bidder’s “ Note on the Physiology of
Sponges,’”! which I had hitherto been unable to see. I find
that this author has forestalled me in a suggestion which I was
about to make concerning the function of Sollas’s membrane,
namely, that it serves to filter food particles from the current
of water flowing through the sponge. As the note in question
is very brief I may perhaps be allowed to quote it in full.

«¢ After feeding with suspended carmine a calcareous sponge
(Leucandra aspera, Vosmaer) the author found that in it
the carmine was at no time in any but the collared cells. The

1 «Proceedings of the Cambridge Philosophical Society,’ vol. vi, part 4
p. 183.

46 ARTHUR DENDY.

water is filtered of the particles. suspended in it by a membrane,
formed by the coalescence of the collars, which stretches com-
pletely across the current. This coalescence has been figured
by Sollas in certain siliceous sponges. The whole evolution
of the canal system in sponges consists in increasing the energy
of the oscular flow and diminishing the velocity in the flagellate
chambers. In these are alike specialised the functions of
absorption and propulsion, since to each a low velocity is
advantageous. The author believes that the collared cells
primitively both ingest and digest the food, the collars having
as their function its retention; digestion is only secondarily
passed to the mesoderm.”

It appears almost certain that in some sponges Sollas’s
membrane completely stretches across the prosopyle, so that
the water has actually to pass through it on its way into the
chamber. We may compare the passage of the water through
Sollas’s membrane to the passage of liquids through organic
membranes by osmosis, only in sponges it is the flagella of the
collared cells which supply the necessary motive power.

Nevertheless, although Sollas’s membrane probably acts in
all cases as a trap to catch food particles—a conclusion arrived
at independently by Bidder and myself—yet it is not likely
that it always stretches completely across the prosopyles or
inhalant apertures of the chambers. Whether it does so or
not probably depends upon the diameter of the prosopyles.
Thus in the Sycons, such as Grantia labyrinthica,' with
large prosopyles, we may safely assume that the membrane
ceases around the margin of the prosopyle. In many cases it
must, from the nature of the case, be impossible to tell with
absolute certainty whether the membrane is continuous across
the prosopyles or otherwise ; for even if, as figured by myself
in Stelospongos, a gap appears in the membrane, we can
never be certain, in dealing with such a delicate structure,
that the membrane has not been accidentally ruptured by the
mode of preparation adopted. Again, it is quite possible and
even probable that temporary gaps frequently make their

1 Vide Dendy, “ Studies on the Comparative Anatomy of Sponges,” III.

7

STUDIES ON THE COMPARATIVE ANATOMY OF SPONGES. 47

appearance in the membrane, which is, after all, only an
extension of the delicate protoplasmic collars of the collared
cells, and it is well known how plastic and changeable such
structures are.

The main point, however, which I wish to establish concern-
ing Sollas’s membrane in the present article is its coexistence
with the flagella of the collared cells, which has not hitherto
been proved. I wish also to support Bidder’s view that its
function is the retention of food particles, which function
would be most effectually fulfilled by its stretching completely
across the prosopyles, and less so in cases where the prosopyles
are too wide to admit of this.

I have now to describe the structure of the ova, which are
remarkable for their great complexity. In my third study on
the comparative anatomy of sponges I have described how the
mature ova of Grantia labyrinthica migrate through the
walls of the inhalant lacune and remain suspended therefrom,
each by a distinct peduncle, awaiting fertilisation by the sper-
matozoa which enter the sponge with the incoming stream of
water. This is probably the mode in which fertilisation is
effected in most if not in all sponges. Fig. 2 represents a
mature ovum of Halichondria panicea suspended by a
peduncle from the wall of a lacuna. I have been unable to
prove that the lacune in which I have found ova suspended in
the case of Halichondria are inhalant, but from the analogy
of Grantia labyrinthica we may assume that this is the
case. The adult ovum is about 0:067 mm. in total diameter.
Its outermost portion forms a distinct envelope around the
ovum proper. The envelope (fig. 2, env.) is fairly thick, and
projects at one side to form the peduncle (ped.), by means of
which the ovum is attached to the wall of the lacuna. The
substance of the envelope is but faintly granular, and stains
lightly ; the peduncle in the specimen figured appears to be
hollow, but I should doubt if this is a constant character.
Within the envelope the ovum proper is suspended as in a
bag. It is spherical, about 0053 mm. in diameter, uniformly
and rather coarsely granular, and stains deeply with borax

48 ARTHUR DENDY.

carmine. In its centre is the nucleus, a spherical body
invested by a very thick and distinct membrane, and measur-
ing, together with its membrane, about 0:02 mm. in diameter.
The substance of the nucleus itself is very finely granular, and
stains but lightly. The nuclear membrane, however, stains
deeply. There is alsoa single spherical nucleolus which stains
very deeply, and is placed excentrically, touching the nuclear
membrane.

I have found several ova exhibiting the structure here
described ; they appeared to be worth mentioning as show-
ing to what a degree of complexity the ovum even of a sponge
may attain, and also as affording some confirmation of my
views as to the mechanism of fertilisation of the ovum in
sponges generally.

The above remarks deal only with certain points in the
structure of Halichondria panicea; for the benefit of the
student I may add that a fairly complete literature of the
species is to be found on p. 2 of the Report on the ‘ Challenger ’
Monaxonida.

EXPLANATION OF PLATE V,

Illustrating Mr. Arthur Dendy’s paper “ Studies on the
Comparative Anatomy of Sponges. IV.—On the
Flagellated Chambers and Ova of Halichondria

panicea.”

Fic. 1.—Small portion of a section vertical to the surface. Zeiss, F, oc. 3.
a, b, c. Different views of three flagellated chambers. 7. 7. Inhalant lacuna.
e. l, Exhalant lacuna. e. 0. c. Exhalant opening of a chamber. sp. Spicules.
z. Protoplasmie processes connecting a collared cell with a stellate meso-
dermal cell (?). s. m. Sollas’s membrane.

Fic. 2.—An ovum suspended in a lacuna. Zeiss, F, oc. 2. env. Envelope
of the ovum. 2. Nucleus. 2. m. Nuclear membrane. 0. Nucleolus.
ped. Peduncle.

ON MEGASCOLEX O@RULEUS. 49

On Megascolex ceruleus, Templeton, from
Ceylon; together with a Theory of the Course
of the Blood in Earthworms.

By

Alfred Gibbs Bourne, D.Sc.Lond., C.M.Z.S., F.L.S.,

Fellow of University College, London; Fellow of the Madras University ;
Professor of Biology in the Presidency College, Madras.

With Plates VI—IX.

INDEX.

PAGE PAGE
Introduction . ; 5 . 49 | Course of the blood . ; pear i)
Historical and systematic . . 50 | Nephridia. eels: : . 81
Habitat. : f : . 51 | Nervous system ; : . 82
External characters . s - 51 | Generative system . ; sn 52
Body-cavity . Berens . 55 | Bibliography . aallts . 84
Alimentary tract : ‘ . 57 | Explanation of plates ‘ . 85
Vascular system : : . 59 |

INTRODUCTION.

Durine the summer of 1889 I visited Ceylon, with the
view of determining the relation of the Earthworm Fauna of
the island to that of Southern India.

I obtained an introduction from the Madras Government,
and express here my great indebtedness to them, as well as
to Sir E. Noél Walker, Colonial Secretary, to H. W. Green,
Esq., Director of Public Instruction (Ceylon), and to
T. C. Huxley, Esq., of Peradeniya, for their assistance in
facilitating my work. To Mr. Huxley I am specially indebted
for all my specimens of Megascolex ceruleus.

I obtained no fewer than thirty-eight! different species of

1 Of these thirty-eight species I have only found seven in India, and at the
present moment I know of about twenty-nine Indian species which I did not

VOL. XXXII, PART I.—NEW SER. D

50 ALFRED GIBBS BOURNE.

earthworms, twenty-three of which are species of Peri-
cheetide, and with very few exceptions are species which I
have not found in India. As a long time must certainly
elapse before I shall be able to publish complete accounts of
all these forms, and the general views to which their study
has led me, I publish here a special account of some structural
features of Megascolex ceruleus, especially the details of
its circulatory system, which its great size has enabled me to
work out. Where I have dealt with matters other than those
relating to Megascolex I have, as a rule, embodied them in
foot-notes.
HistoRIcaAL AND SYSTEMATIC.

There cannot be the slightest doubt but that the worm here
described belongs to the same genus as the worm described by
Beddard (1)! as Pleurocheta Moseleyi.

This author (2), after an examination of the type specimens
of M. ceruleus in the British Museum and in Edinburgh,
and subsequently to the publication of the paper above
referred to, came to the conclusion that his genus Pleuro-
cheeta was identical with the genus Megascolex established in
1845 by Templeton (14). I am convinced that the species
also are identical, and that the name Pleurocheta
Moseleyi must be considered as cancelled. It is perfectly
true that, owing to the loss of the ‘‘ Memnon,”’ by which ship
Templeton despatched to England his original memoir, the
existing description is very scanty ; but on the one hand we
have Beddard’s examination of the type specimens, and on the
other the worm is very large (the largest known Perichete) and
of a striking colour, and from numerous inquiries which I made
on the spot is, it appears, well known to Europeans and natives,
and believed to be the only worm of that size in the island.

The question of the identity of M. ceruleus with any of
find in Ceylon, making the amazing total of about sixty-seven species; and I
have no reason to suppose that the field is by any means exhausted. In the
town of Madras there are four species: Pericheta armata, two new species

of Acanthodrilus, and a minute species of Moniligaster.
1 These numbers refer to the bibliography at the end of this paper.

ON MEGASCOLEX C@RULEUS. 51

Schmarda’s species is much more difficult. That Schmarda
himself was led away by a mistaken interpretation of Temple-
ton’s description has often been pointed out. Templeton
wrote, ‘ Each ring is dilated in the middle of its length into a
ridge, which carries on it, except in the mesial line of the back,
minute conical mammillz, two in number, each surmounted by
a minute bristle ;” while Schmarda (18) speaks of “ the ridges
which in Megascolex were stated to be found only on the
dorsal side, continued in our specimens also on to the ventral
side.”

It is just possible that the Pericheta leucocycla of
Schmarda may be the same worm ; I am unable to identify it
with any other species I found in Ceylon.

As to the best use of Schmarda’s generic term Pericheta I
shall speak in a subsequent paper. I agree entirely with
Rosa (11, a) in his classification of the Perichetide as far as
it goes, but am convinced that it will be necessary to create
several other genera.

HaBITAt.

All my specimens of Megascolex ceruleus came from
the neighbourhood of Kandy and Peradeniya and the hills
near, that is to say, from an elevation of about 1700 feet.
I have no reliable information as to its occurrence at any
higher or lower level. I was not fortunate enough to obtain
a specimen myself, and so be enabled to note any peculiarities
as to its habitat.

EXTERNAL CHARACTERS.

Colour and Size.—The general appearance, colour, and
size may be judged from figs. 1 and 2.!

1 Tam indebted to my wife for coloured external views of all the worms which
I have procured in a living state, and shall not give any further description.
Probably no two observers would agree in describing the-colour of an earth-
worm, and an accurate figure of a full-grown living specimen gives a better
idea of the size than any measurements. In no case can we rely absolutely
upon measurements of length, as the worm is constantly changing its amount

52 ALFRED GIBBS BOURNE.

The pigment changes very slightly, at any rate for some
months, when the worm is preserved in spirit ; but the reddish
portions shown in the figures, which are due to absence of
pigment and great blood supply, naturally disappear when the
specimen is placed in spirit. When the worm is alive these
red portions are very noticeable, and indicate the greatly in-
creased blood supply in the prostomium and round the mouth,
and in the neighbourhood of all the genital apertures
(fig. 2).

Number of Segments.—I have counted the number of
the segments in specimens varying from 10 inches to 32
inches in length, and have found it to vary between 250 and
290, being usually about 270. In fig. 1, 286 segments are
shown.

Prostomium.—This is, as is often the case, of a blood-red
colour, and is broad and blunt at the extremity; there is a
short transverse groove along its line of junction with the
peristomial (buccal) segment, and both it and the latter seg-
ment present in spirit-preserved specimens numerous longi-
tudinal grooves.!

Sete.—lIn shape the setz are, as compared with those of
many perichete worms, short, stout, and only slightly
curved.

As to their position, they are of course arranged in rings,
of contraction, and a contraction which produces a considerable alteration in
length produces a barely measurable increase of thickness.

It is interesting to note that worms of some species possess a much greater
power of contraction than others. P. mirabilis, for instance, contracts and
extends itself to but a very slight degree, and has a movement like a Nematoid,

while Moniligaster grandis can contract itself to about a quarter of its
fully extended length.

1 We have at present few data to enable us to make much use of the pro-
stomium in classification. It will be necessary to compare its structure in a
large series of living worms. In preserved specimens it becomes contracted
to a large and very variable extent. Moreover, it never has when the worm
is alive the same shape for two seconds together. Many worms have a habit
of constantly protruding a large portion of the pharyngeal region, and there is
no line of demarcation between the prostomium on its ventral side and this
latter region,

ON MEGASCOLEX C@RULEUS. 58

and there is a small gap in the ring in the median dorsal and
ventral lines. The dorsal and ventral gaps are equal in the
greater part of the body to the interspaces between three or
four of the neighbouring sete. The seta gaps are throughout
smaller in the ventral than in the lateral and dorsal regions.

In the most anterior segments the gaps, especially in the
lateral and dorsal regions, and above all the median dorsal
gap, are much larger than in the posterior segments. The
number of the sete is greater than in any other recorded
Perichete. There are 120 to 140 in most segments, but con-
siderably fewer in the most anterior segments. I have found
as few as thirty-six in segment v.!

The most ventrally placed sete, especially in the neighbour-
hood of the male pores, are a little larger than usual, but I
should not speak of them as “ modified” sete. There are no
setz in segment xvii11 between the male pores.?

I have never found the sete absent from the clitellum, as

1 I am in the habit of counting the sete in segments v, Ix, and xxv, the
latter segment serving as a type for the rest of the body, and I find that the
relation of these numbers to one another varies with other important characters
rather than the actual numbers themselves.

The method I adopt in examining the arrangement of the sete is to cut
open the freshly killed worm on one side of the dorsal median line, to scrape
out the viscera, to flatten the body-wall of the most anterior twenty-five
segments between two glass slips, and to allow it to harden in spirit; subse-
quently to treat this piece of skin with potash followed by glycerine, and
then to mount it.

This process renders it possible to see the follicles, even where the actual
seta has fallen out.

The shape of the sete varies so slightly in different species, that except in
special cases it is of little use for classificatory purposes. In a preparation
made as above, modified sete in the neighbourhood of the spermathecal aper-
tures or male genital pores should always be looked for.

? The presence or absence of sete in segment xv1r between the male
pores is a most important character among the Perichetide. In all worms
which I think on other grounds it will be advisable to place in the genus
Pericheta s. str. such sete are present. They are also present in certain
other worms which have other special peculiarities, but in a very large
number of species, e.g. P. armata, and the majority of Fletcher’s Australian
species, they are absent,

54 ALFRED GIBBS BOURNE.

recorded by Beddard (1), but then in none of my specimens
was the clitellum very well developed.’

Clitellum.—The clitellum is not shown in my coloured
figure, as I was unable to secure a live worm in which it was
developed, but I have seen it in spirit specimens collected at
some other time of year.

I have seen it developed upon the posterior portion of seg-
ment x11, and upon every succeeding segment down to and in-
cluding segment xx1; it is, however, always deficient ventrally
from segment xvii onwards, so that the male pores do not
actually open through clitellar substance.”

Genital Apertures.—The spermathecal apertures are
placed between segments vil . viri and viti . 1x, or rather just
on the anterior margins (which is the usual arrangement) of the
hinder of those segments in each case. They are all placed
equally near the median ventral line, and about in a line with
seta 9 (i.e. the ninth seta from the median ventral line).

The two oviducal apertures are placed very close together, and
very slightly in front of the seta ring in segment xtv’° (fig. 2).

1 With regard to the presence or absence of setz on the clitellum a good
deal of unnecessary confusion exists. In all young perichete worms which I
have examined setee are present on the clitellar segments, but when the
clitellum develops they may remain projecting, or they may become buried
in the clitellar substance, or any or all of them may actually drop out. I am
inclined to think that all the species of one genus behave in the same way in
this respect, but am not sure of this.

* [ know of no perichete worm in which the male pores do thus open. We
shall have to distinguish between worms in which the clitellum forms a definite
girdle, strictly limited in normal individuals to certain segments, and those in
which it always shows a tendency to spread somewhat irregularly. Even if
we abandon, as Vejdovsky (15) and Rosa (11, A) do, and as we certainly must,
Perrier’s classification (8) of earthworms into Anteclitellians, Intraclitellians,
and Postclitellians, we may still use the character of the clitellum in classifi-
cation.

3 I know of no Perichete in which there are any sete between the oviducal
pores ; the latter always lie either in the ventral seta gap or slightly in front
of the seta ring altogether. It is often a very difficult matter to ascertain
whether the oviducal pores are paired or single and median.

T am now convinced that P. saletensis and P. bivaginata, which I

ON MEGASCOLEX C(ERULEUS. 5d

The male pores lie immediately ventrad of the most ventrally
placed seta in segment xvi11. There are, as stated by Beddard,
two other pairs of apertures in this region, though not exactly
in the position stated by him. The one pair lies between seg-
ments xvit.xvii1, and the other between segments xvIit. xIx.
These pores are the apertures of the glands described below
(p. 83).

All these six apertures lie about equally near the median
ventral line and opposite sete 7 (figs. 2 and 18).

In the living worm there are no papille and no depressions
in this region, but when the worm is killed and contraction of
the body-wall takes place a depression is produced, as shown
in fig. 18, and the pores lie at the edges of this depression, the
male pores upon papille.1

Dorsal Pores.—The most anterior dorsal pore lies between
segments vi and vit, and other dorsal pores occur between
every two segments, including the clitellar segments.”

Bopy-cavity.

This is as usual incompletely divided into separate chambers
by the septa.®

separated chiefly because I believed them to differ in this respect, are the
same species, and, indeed, the same as P. armata.

1 There can be little doubt that the majority of papille: which have been
described in the region of the genital apertures of so many earthworms are
not permanent structures, but produced by contractions of the body-wall ;
they are regions doubtless in which some portion of the muscular layers are
absent, so that when general muscular contraction takes place they stand up
as papille; and when a worm is so killed as to become greatly contracted
these papilla are seen, but if the same worm had been gradually killed
they would not appear. Great care should consequently be exercised in
using such papille as classificatory characters.

2 These dorsal pores are important in classification. In some genera they
are absent (Hudrilus, Teleudrilus, Thamnodrilus, Moniligaster, &c.). The
position of the most anterior pore varies in differeut species. Sometimes
they are absent from between every two clitellar segments.

* The body-cavity of any segment is ordinarily bounded by two septa. The
anterior septum of a segment N may be conveniently termed septum N —1.N,

56 ALFRED GIBBS BOURNE,

The most anterior septum which is well marked is septum
1v.v. Ihave not been able to satisfy myself as to the presence
or absence of any septa anterior to this. The two or three
most anterior septa are usually stated to be absent, but if
present they are doubtless very thin and much obscured by
the muscular fibres which radiate from the pharyngeal to the
body-wall. I am inclined to think, both from dissection of
this big worm and from sections of smaller worms, that these
septa are usually present, and that the pharynx really lies in
segment I.

Septa v. vi and vi. vir are very thin.

Septum vit. viit is also thin.

Septum viii. 1x is thicker.

Septa 1x.x, X. XI, XI. XII, XII. x111 are very thick.

The three succeeding septa are less so, and the septa behind
these are all very thin. From all the thick septa arise nume-
rous muscles, some of which are shown in fig. 3. These pass

backwards through one or two succeeding septa to be attached
to the body-wall.

These muscles, and indeed the thickened septa, must, as has
been suggested, give greater strength to the anterior end of

and the posterior one septum N.N+1. In stating that a certain organ lies in
a certain segment great care is necessary with regard to two matters. In the
first place, the septum may be attached to the body-wall at some spot more
or less widely removed from the intersegmental groove. ‘There is great
variation in this matter in different genera; it is particularly noticeable in
Moniligaster. In all such cases which I have examined by means of longi-
tudinal sections the muscular fibres of the septum penetrate the body-wall at
the right place, but have by a growth of connective tissue become adherent to
the body-wall over some region behind the groove. In the second place,
certain septa are at times undoubtedly absent or quite rudimentary. Perrier
(9) states that two septa are absent in Urocheta. Beddard (8) also states that
septum 1x. x is almost absent in a species of this genus, and I have observed
this to be the case in specimens of Urocheta obtained from both South India
and Ceylon. The septa viir.1x and 1x.x are absent, I believe, in all the
species of one genus of Perichetide.
The two or three most anterior septa are, moreover, absent in all worms,

or so feebly developed as to have escaped notice, a fact which adds to the
difficulty of enumeration,

ON MEGASCOLEX C@RULEUS. 57

the worm and assist it in burrowing. The conical shape of
these anterior septa may be gathered from fig. 3.

ALIMENTARY TRACT.

I have little to add in this respect to Beddard’s account for
‘ Pleurocheta.” I differ from him slightly with regard to the
enumeration of the segments.

The arrangement of the anterior portion of the alimentary
canal is shown in fig. 3.

The pharynx is bounded posteriorly by septumiv.v. This
statement is not intended to exclude the possibility of there
being thin, more anterior septa passing behind it. Numerous,
doubtless unicellular glands lie outside on the walls of the
pharynx, and open into it in the dorsal region.

The gizzard consists of an anterior thin-walled portion and
a posterior thick-walled portion. Both portions are contained
in the same segment. The former bears, projecting into its
lumen, numerous hair-like processes which are lined by chitin
and doubtless serve as a strainer. I am not aware that any
such arrangement has been described in any other worm.
The latter has a very thick muscular wall, and this is lined
internally by thick, hard chitin. Following the gizzard is a
narrow portion of the cesophagus, which extends from segment
VE toir:)

There is no specially large blood supply in the walls of the
portion of the canal above described.

Segments x to xv contain the calciferous glands. In each
of these segments the canal is swollen out into a bulb-like
form, so that there is a series of such swellings, and between
them in the region of the septa there are constrictions. I
call these dilated portions of the cesophagus, calciferous glands,
because there may usually be found in them smaller and
larger concretions of carbonate of lime.?

1 Tuse for the present the term cesophagus for any portion of the alimen-
tary tract which lies anteriorly to the large intestine, and is not designated by
any special name—as, for instance, pharynx or gizzard. r

* I sent a number of these concretions to Professor W. D. Halliburton, of

58 ALFRED GIBBS BOURNE.

The inner wall is much plicated, being raised into ridges
and papille, and is excessively vascular.

The two most posteriorly placed glands are smaller than the
other four.

It is beyond the scope of the present paper to discuss the
relation of these glands to those found in other worms. I
will merely remark that this is a very common type of calci-
ferous gland among the Perichetide, though by no means the
only one. It is, moreover, the simplest.

Beddard mentions no such glands in “ Pleurocheta.”’

In segments xvi and xvii there is a further portion of the
narrow cesophagus, and in the latter of these this suddenly
widens out into the intestine.

The intestine is, as Beddard has pointed out, for “ Pleuro-
cheta”’ very complicated.

The anterior portion may be called the typhlosolar region,
and the posterior the post-typhlosolar region.

The typhlosole is in a very rudimentary condition. It isa
mere ridge of the intestine projecting into the lumen along
the median dorsal line. As stated below (pp. 61 and 70),
there is no continuous longitudinal blood-vessel running along
it, but the capillary network becomes so exceedingly dense here
as to form a longitudinally placed blood lacuna.

This typhlosole extends as far back as segment cxxxv.

In the anterior portion of the typhlosolar region I find the
intestinal wall forming the pouches described by Beddard (1,
p. 492). In segments xvi to xxxv there are the large simple
pouches, and in segments xxxvi to xui1 are the seven pairs of
sacculated pouches which correspond very closely to Beddard’s
description.

Posteriorly to these sacculated pouches the intestinal wall
is not protruded to any great extent into pouches.
the Physiological Laboratory, University College, London, and am indebted
to him for the information that they undoubtedly consist almost entirely of
carbonate of lime. But he says, “A few shreds of organic matter remain
undissolved. When the concretions are treated with acid, these prove to he
of proteid nature,”’

ON MEGASCOLEX C@RULEUS. 59

In the post-typhlosolar region the intestinal wall becomes
much thicker. In the hinder portion of the typhlosolar region
lie the remarkable ‘‘ kidney-shaped glands.” I find twenty-
two pairs of these glands, a pair in each segment from cx11 to
exxxi1I1. Beddard describes only fifteen pairs, lying in seg-
ments Lxxxvi to cr (‘for thereabouts”). With regard to
structure I can confirm in every particular Beddard’s account.
In my sections these glands present very much the appearance
which Beddard figures, and each undoubtedly opens into the
intestine. When the intestine is opened and its dorsal wall
examined from the inside, the two rows of apertures are per-
fectly clearly seen. The glands in the middle of the series
are the largest, and they get smaller and smaller at either end
of the series. As I have stated elsewhere (circulatory system),
the typhlosole, the anterior dorsal intestinal vessel, and the
series of kidney-shaped glands all cease to exist at about the
same spot.

I am unable to throw any further light upon the function
of these glands.

I give no account of the histology of any portion of the ali-
mentary tract. We still want information as to the histology
of the alimentary canal in earthworms, but it should be in the
form of a comparative account dealing with a series of typical
genera.

VASCULAR SYSTEM.

It will be convenient to describe firstly the blood-vessels,
and secondly the probable course of the blood-flow.

The red blood differs in no way from that fluid in other
worms.

The Blood-vessels.

Dorsal Vessel (figs. 5 to 11, p.v.).—The dorsal vessel
extends from the anterior portion of the pharynx to the last
segment of the body. It lies above the alimentary canal, but
is not adherent to the wall of the latter in any portion of its

60 ALFRED GIBBS BOURNE.

course. Its walls are muscular throughout its length, but
most so in segments vir to x11I, in which segments the vessel
is much dilated. In the intersegmental (i.e. septal) regions,
where the vessel is narrowest, there are in this portion of the
vessel, and in all the portion posterior to it, valves which can
doubtless entirely, or almost entirely, shut off the lumen of the
vessel in each segment from that in the other segments. These
valves are thick, semicircular ridges of connective tissue
attached to the wall of the vessel across its ventral half, and
presenting an irregular free edge (fig. 12).

I have never seen the dorsal vessel double in any portion of
its course. I am surprised at this, as Beddard (1) distinctly
states that in his Pleurocheta Moseleyi “it bifurcates no
less than five times in the first eight segments, the bifurcations
always coalescing again directly.”’!

Anteriorly the dorsal vessel bifurcates—that is to say, it
gives off a pair of branches which behave in a manner similar
to the other dorso-tegumentary vessels (see p. 71), and
are to be considered as the most anterior pair of such vessels
(fig. 8).

Posteriorly the dorsal vessel ends abruptly, as shown in
fig. 11.

Ventral Vessel (figs. 4, 6 to 11, v.v.).—This is also
known as the subintestinal or supra-neural vessel. Its walls
chiefly consist of connective tissue, doubtless elastic, with
circularly disposed bands of muscle in the regions of the septa.
These bands would serve to secure a specially great blood-flow
in the branches of the ventral vessel in any particular region
of the body. It is of very uniform calibre throughout its
entire length. At its anterior and posterior extremities it

1 To prevent any confusion upon this point I may note that I am well
acquainted with the “double” condition of the dorsal vessel which has now
been several times recorded; it obtains in a perichete worm which attains
nearly the same size as Megascolex ceruleus, and which I discovered on
the 24th May, 1887, on the Nilgiri Hills in South India. In this worm the
dorsal vessel bifurcates in each segment from vir to xvi. I have noticed a

sort of longitudinal folding in of the dorsal wall in certain segments of M.
ceruleus, but the vessel is not double.

ON MEGASCOLEX C@RULEUS. 61

terminates by bifurcating, and thus giving off its most anterior
and posterior branches, which are described under the heading
Ventro-tegumentary Vessels (see p. 72).

Supra-intestinal Vessels (figs. 4 to 6).—There are, as
Beddard states, two of these vessels lying side by side, but
widely separated, on the wall of the cesophagus in segments
ix to xu. The two are joined by commissural vessels in
segments x to x11. In segment 1x they lose themselves in the
intestinal capillary network, and in segment xiv they join, and
a very small median supra-intestinal vessel runs on into seg-
ment xvi, where it bifurcates and joins the dorso-intestinal
vessels of that segment (fig. 5). It is not continued into the
region of the intestine properly so called ; there is consequently
no typhlosolar trunk.

The supra-intestinal vessels are, as usual, closely adherent
to the intestinal (cesophageal, &c.) walls.

Subneural and Latero-neural Vessels are absent.’

Intestino-tegumentary Vessels (figs. 4, 6 to 10, 1.7.).
—This name was given by Perrier (9) to a pair of symmetrical
longitudinal vessels which are connected at either end with a
network of capillaries, the one network being in connection
with some part of the cesophageal wall (i.e. some region in

1 In many worms the supra-intestinal vessel is said to be prolonged back-
wards as the typhlosolar vessel. I should not be surprised to find that the
typhlosolar vessel is much more frequently absent than is supposed, if not
altogether so. Benham (4, p. 286) speaks of it as an “ill-defined vessel ”
in Microcheta rappi. Jaquet (6, p. 346) speaks of the circulation in the
typhlosole as being “‘trés difficile 4 poursuivre.” Vejdovsky (15, p. 110)
seems a little doubtful about it, and I have frequently seen in sections blood
in that region which I should have said at once lay in the typhlosolar vessel,
when further examination of the series of sections has convinced me that
there was only a special development of the internal intestinal capillary net-
work, a remnant doubtless of the sinus around the intestine of many
Cheetopoda.

2 Lankester (7) calls the subneural vessel the ventral vessel, but I prefer to
use this latter term for the subintestinal vessel, the latter being constant in
the Oligocheta, while the subneural vessel is absent in all the simpler and
many of the more complicated forms, e.g. many, if not all, Perichetide, Pon-
todrilus, and Microcheta.

62 ALFRED GIBBS BOURNE.

front of the large intestine) ; the other network (set of net-
works it really is) being in connection with the pharyngeal
wall, the septa, and the body-wall. Benham speaks of them
as the “ lateral longitudinal” vessels. I shall show that they
are only the much-enlarged representatives in the anterior
region of the body of a series of similar vessels which occur, a
pair in every segment, in all the remaining portion of the
body. There is, in fact, in Megascolex, and I expect in
many other earthworms, a vessel on each side in every
segment—in all except certain anteriorly placed modified
segments—a vessel which communicates with capillaries at
either extremity, the one network of capillaries being in con-
nection with the intestinal wall, and the other with the body-
wall, and the blood in circulating either passes from the in-
testinal capillaries laden with nutriment to the cutaneous and
other capillaries for the nourishment of the tissues and for its
own aération, or is collected in an aérated condition from the
cutaneous capillaries to pass to the intestinal capillaries for
the absorption of nutritive matters.

A discussion as to which of these two courses it takes
follows the description of the vessels. There is only a single
pair of such vessels in the anterior modified region, and it is
this pair of vessels which has been called intestino-tegumen-
tary vessels by Perrier. I propose to apply the term intestino-
tegumentary to the whole series, and to call anterior intestino-
tegumentary vessels the large anteriorly piaced modified pair,
and intestino-tegumentary vessels of such and such a segment
those in the posterior region.

The anterior intestino-tegumentary vessels have relations
with the twenty or so most anterior segments, and are, I be-
lieve, in connection with the intestino-tegumentary vessels of
the following segment, and these again with those that follow,
and so on, by means of capillaries or very minute vessels,
The relations of the large anterior pair have been dealt with
by Perrier, Horst (5), Benham, Beddard, Jaquet, and others.!

1 These vessels, or at any rate some vessels having somewhat similar
relations, have been stated to communicate directly with the dorsal vessel in

ON MEGASCOLEX C@RULEUS. 63

I merely describe here what occurs in Megascolex, without any
further detailed discussion of their views.’ The main trunks
of the anterior pair of intestino-tegumentary vessels run from
the sides of the pharynx, lie freely in the body-cavity in the
region of the gizzard, and then gradually take up a more
ventral position, passing to the inside of the hearts, without
being connected with them, till they become adherent to the
ventral wall of the cesophageal (calciferous) glands.” The
anterior extremity of each is connected with a network of
capillaries on the pharyngeal wall. This network is connected
with the network into which the most anterior branches of the
dorsal and ventral vessels break up in this region (fig. 8).
Passing backwards they are joined by various branches in the
regions of the septa. An inspection of figs. 4 and 8 will show
that these branches are segmentally arranged, and are con-
nected with capillary networks on the septa and in the body-
wall, which communicate on the one hand with either the
ventro-tegumentary vessels or with branches of the most
anterior heart, and so also with the ventral vessel. One
specially large branch, which has also connections with the
hearts of segments vi, vil, and vIII, communicates with a
special network on the gizzard; another with the networks
which are also connected with the four most anterior pairs of
ventro-tegumentary vessels, and so on (see fig. 4). In seg-
ments x to x1I1, i.e. those segments in which occur the four
large cesophageal glands, the arrangement is somewhat dif-
ferent, and is almost exactly repeated in each of these seg-
ments. One branch is connected with the peripheral capillary
networks, and two or three branches are connected with the

Lumbricus. In the absence of specimens of that genus I am unable to discuss
this fact, nor its bearing upon my theory as to the course of the blood.

1 It will be seen how little my account differs from that of Perrier, whose
memoirs on Urocheta and Pontodrilus are most masterly pieces of work,
and to whom I here express my many obligations.

* In fig. 4 this vessel is shown as lying at some distance below these
glands. The drawing is diagrammatic to better elucidate the relations of the
branches of this vessel.

64: ALFRED GIBBS BOURNE.

“intestinal” networks, i.e. with those networks in the walls
of the cesophageal glands.

The other intestino-tegumentary vessels have relations which
are precisely similar to one another (figs. 4, 7, 9, 10). The
main portion of the vessel in each case lies closely adherent
to the body-wall just behind a septum, i.e. in the anterior
portion of a segment; the ventral end of it is connected by
several small branches (fig. 7) with the external intestinal
capillary network, while from the trunk numerous branches,
especially near the dorsal region, pass to the peripheral net-
works. The branches of the intestino-tegumentary vessels
are thus of two kinds, peripheral branches and intestinal
branches. All the intestino-tegumentary vessels place the
peripheral and intestinal capillary networks in communication
with one another, a relationship discovered by Perrier for the
large anterior pair, which led him to give them the name of
intestino-tegumentary vessels, and to compare them to portal
vessels. The relationship, or existence even, of the posterior
intestino-tegumentary vessels does not appear to have been
hitherto described. They are connected with one another,
the pair of one segment with that of the adjoining segments,
and the one of one side with that of the other, in the intestinal
wall, and, I believe, also in the body-wall (figs. 7 and 10).
The longitudinal connections in the intestinal wall constitute
doubtless the infra-intestinal vessel which is figured by Howes
in the ‘ Atlas of Biology ’ (Lumbricus), and is stated by Benham
(4, p. 253) to have been observed by Beddard in Acantho-
drilus.

Hearts.1—There are eight pairs of rhythmically contractile

1 T use this term for all rhythmically contractile, circularly disposed vessels,
thus including, for reasons stated below, certain anterior branches of the
dorsal vessel which do not join the ventral vessel.

These hearts are either—(1) all connected with the dorsal vessel, or (2)
some only are so connected, while others are connected with the supra-
intestinal vessel only, or (3) some are connected with the dorsal vessel only,
and some with both the dorsal and supra-intestinal vessels (in Pontodrilus
and Titanus [see Perrier, 10], in Megascolides [see Spencer, 18], in Megas-
colex, suggested as a possible arrangement by Beddard and figured in this

ON MEGASCOLEX C@RULEUS. 65

branches of the dorsal vessel; none of the other branches of
the dorsal vessel, either those anterior or those posterior to
these eight, are contractile.

The three anterior pairs lying in segments v1, vil, and vit
are lateral hearts ; while the five posterior pairs lying in seg-
ments 1x to x11 are latero-intestinal hearts, i.e. they are
connected equally with the dorsal and the supra-intestinal
vessels of their respective sides. An examination of fig. 5 will
show that the dorso-intestinal vessels of segments xiv to xvi
have similar connections, but they do not appear to be rhyth-
mically contractile, nor have they the peculiar sphincter
muscle at their distal extremities that the hearts have.!

The hearts of segment vi arise immediately in front of
septum vi. vii (each pair of hearts arises immediately in front
of the septum which divides the segment in which they lie
from the segment next following). They extend only fora
short distance on the wall of the gizzard, and terminate in a
muscular bulb (figs. 4 and 6, f), a sphincter which is shut
during the diastole, and opens at the systole of the heart.
This heart is not connected with the ventral vessel. Two
branches arise at its extremity, just beyond the muscular bulb.
No branches arise directly from any of the hearts, i.e. between
the point of connection with the dorsal vessel or dorsal and
supra-intestinal vessels, as the case may be, and these muscular
bulbs. The two branches above mentioned break up into
capillaries on the walls of the gizzard, the network being con-
nected, on the other hand, with branches of the anterior

memoir, and observed by myself in many other Perichetide). In all recorded
cases in which there are some hearts which do not communicate alone with
the dorsal vessel, such hearts are the posterior ones of the series.

I adopt Perrier’s term lateral hearts for those which communicate with
the dorsal vessel only, and his term intestinal hearts for those which com-
municate with the supra-intestinal vessel only, and shall use the term latero-
intestinal hearts for those which have the dual connection to indicate tha
they correspond to both lateral and intestinal hearts.

1 These peculiar relationships show how useless it is for classificatory
purposes to record simply the number of pairs of “hearts,” as is often done,
without detailed account of what vessels are so named.

VOL. XXXII, PART I.—NEW SER. E

66 ALFRED GIBBS BOURNE.

intestino-tegumentary vessels. The hinder branch comes
into relation in this way with that branch of the anterior
intestino-tegumentary vessel which lies on septum VI. VII.

The hearts of segment vit are of nearly twice the length of
those of segment v1. They also give off two branches from
the extremity beyond the muscular bulb; the one branch has
relations similar to those described above with the branch of
the intestino-tegumentary vessel which lies in septum viI.
vit, and the other joins the similar branch from the heart of
segment vi1r and goes to the gizzard, where it, like the cor-
responding branch of the heart of segment v1, is in communi-
cation by means of a capillary network with the intestino-
tegumentary vessel.

The hearts of segment viti are directly connected with the
ventral vessel, but between the muscular bulb and this point of
connection give off two branches, one of which is mentioned
above, and the other has corresponding relations with the
branch of the intestino-tegumentary vessel which lies on
septum VIII. Ix.

The five succeeding pairs of hearts are all latero-intestinal.
Those of segment 1x join the ventral vessel; but, as in the
case of the hearts of the preceding segment, there is a certain
length of vessel between the end of the heart, as marked by
the muscular bulb and the ventral vessel ; from this portion
arises one branch only, which has the usual relations with a
branch of the intestino-tegumentary vessel—that which lies on’
septum 1x.x. There is no branch to the gizzard-wall; we
are now behind that region. All these arrangements are shown
in fig. 4.

All the segmentally arranged capillary networks with which
the hearts and intestino-tegumentary vessels communicate, as
described above, extend over the body-wall in their own
neighbourhood.

The hearts of ségments x—x111 have relations precisely
similar to one another. Each is connected at its proximal
extremity by a vessel much narrower than itself with the dorsal
vessel, and by another narrow vessel with the supra-intestinal

ON MEGASCOLEX C@RULEUS. 67

vessel of its own side (figs. 4—6). These four pairs of hearts
are much larger than any of the other hearts; they are moni-
lated, and the muscular bulbs at their distal extremities are
placed at the junction with the ventral vessel in each case;
there are consequently no branches, for, as I have stated above,
no branch ever arises from a heart proper. (Note in this con-
nection the different behaviour of the intestino-tegumentary
vessels in segments x to x11; see fig. 4.)

Capillary Networks.'—It will be most convenient to
consider these before speaking further of the vessels which
put them in connection with the above-mentioned trunks.

We can, I think, recognise two groups of capillary networks
only—peripheral networks and intestinal networks. I mean
that there are no such things as special commissural networks
placing any of the great trunks in communication with one
another. If such special networks as have been described by
various authors exist, placing the big longitudinal trunks in
communication with one another, they may be suitably termed
commissural networks. The most important of these which
has been described is the network into which the dorsal vessel
breaks up at the anterior extremity of the body, and which
comes into relation with a similar network arising from the
ventral vessel. But in Megascolex, at any rate, I see no reason
why this should not be grouped with the peripheral networks.
I wish, indeed, to bring into prominence the fact that we have,
iu the most anterior region of the body, only a series of seg-
mentally arranged networks—nothing, in short, which differs
from what obtains in other segments of the body. It must,
however, be borne in mind that my investigations have been
made upon a worm devoid of subneural and latero-neural
vessels, the presence of which may entail other variations.

The networks which connect the dorsal vessel with the sub-

1 Perrier (9, p. 466, foot-note) very justly points out that the term ‘‘ capil-
lary’ is here used in a special sense, as there is nothing like the difference
between a so-called capillary and a small vessel that obtains in a vertebrate
animal; but the same remark applies with equal force to the use of the term
in all animals other than vertebrates.

68 ALFRED GIBBS BOURNE.

neural, described by Perrier (9) as occurring in segments
vill, x, and x11, in Urocheta, may be special commissural
networks.

Another capillary network, which, if it exists in any worm,
would be a commissural network, is that connecting the dorsal
and ventral vessels at their posterior extremities. From
observations which I have made upon Megascolex, as well as
upon some small worms, mounted alive in a compressorium, I
believe that there is no such special network, but that the
terminal branches of these vessels behave just like any other
of their branches (fig. 11). Jaquet (6), however, speaking of
Lumbricus, says that the dorsal and ventral vessels “se
mettent en relation par des anastomoses réciproques” at their
posterior extremities.

Peripheral Networks.—I would give this name to all
the capillary networks in the skin in which the blood under-
goes aération, also those in the septa—in fact, all those which
do not belong to the intestinal system; and I think we are
justified in grouping all these together, as they all have
similar relations with the large vessels, and are all meta-
merically arranged.

In Megascolex, at any rate, these always establish communi-
cation between the dorsal, ventral, and intestino-tegumentary
vessels. They le for the most part in the superficial region
of the body-wall, but they are also to be found in all the
various tissues and viscera, excepting only the alimentary
canal, and even here the exception does not extend to the
pharyngeal and gizzard region. I have not had sufficient
material to work out their exact arrangement in the nerve-
cord, nephridia, generative organs, or walls of the large blood-
vessels, in which latter they are to be found as vasa vasorum,
but I am satisfied that in all these cases they have relations
similar to those of the networks found in the body-wall. I
believe that in all cases branches of the dorsal, ventral, and
intestino-tegumentary vessels enter into connection with them.
They are certainly continuous across the dorsal median line,
and I believe also from segment to segment, though not of

ON MEGASCOLEX C@RULEUS. 69

course equally dense throughout, and it is therefore possible
to speak of separate networks segmentally arranged.

The most anterior (fig. 8) is connected with the two most
anterior branches of the dorsal vessel, those of the ventral
vessel, and also the most anterior ramifications of the anterior
intestino-tegumentary vessels.

Following these there is a series of similarly arranged net-
works throughout the body, the only exceptions being the
slight ones about to be mentioned, and these are due to the
presence of the hearts. In segments vi1r to x111 the connec-
tions with the ventral vessel are wanting ; in segments VI to Ix
there are no direct connections with the dorsal vessel, their
place being taken by connections with the hearts of those
segments, or rather with the vessels which are the distal
connections of those hearts.

In all other segments there are connections with the dorsal
vessel by means of dorso-tegumentary vessels; of the exact
origin of these vessels in segments x to x11 I have no note.
In all the segments as far back as x11, and probably in the
next four or five segments also, there are connections with
anterior intestino-tegumentary vessels; in all the succeeding
segments, and perhaps in the above-mentioned four or five
segments, there are connections with the intestino-tegumentary
vessels of the various segments.

Intestinal Networks.—All the capillary networks in the
walls of the alimentary canal, excepting in its most anterior
region, where the capillaries are more superficial and belong to
the peripheral networks, fall naturally into one group, and are
described below in detail for Megascolex.

These networks, as I have found them in Megascolex, differ
considerably from those described as occurring in other genera.
I select for detailed description one of the segments x11 to
cxxxv (fig. 10).

There are two capillary networks in the alimentary canal
wall—an internal deep-lying network and an external more
superficial one.

The internal network (fig. 10), which corresponds to the

70 ALFRED GIBBS BOURNE.

“ quadrillage” of Perrier (9, p. 491) and Jaquet (6, p. 345),
is so dense a network that it may be regarded as a blood sinus
interrupted at certain spots; the interspaces are in certain
places even smaller than the vessels which surround them.
The meshes are not so regularly rectangular as they appear to
be in some other genera; they are not equally so, moreover,
in all regions of the intestinal wall. Near the typhlosole and
also along the intersegmental lines the longitudinal portions
of the network are specially developed, and the meshes fairly
rectangular ; in other regions they are less regular. The net-
work is continuous from segment to segment, and across the
dorsal and ventral median lines in each segment. The vessels
are largest and the interspaces smallest over the typhlosolar
region. There is indeed so much blood in this region that
when the intestine is opened the typhlosole strikes the eye at
once as a longitudinal band of red colour, noticeable even when
the whole surface appears red. I have seen the vessels so dis-
tended that the interspaces were hardly recognisable. With
the slight exceptions mentioned above, there is not very much
difference in calibre among the vessels composing the network.
All the vessels in any particular region may be distended or
the reverse, but that is all. This internal network is directly
connected with the dorso-intestinal vessels.

The external network is very different. The vessels are of
very various calibres. There are large vessels which divide
into small branches, and these subdivide and so on; the
smallest branches form a complete network. These networks
are arranged segmentally, and are not continuous, as networks,
that in one segment with that in another, as is the case with
the internal network. The network in each segment is con-
tinuous over the ventral median line, but not over the dorsal
region. At very numerous spots little branches from the
external network penetrate the intestinal wall and open into the
internal network. The vessels of the external network have
always clearly defined but very thin walls, do not seem capable
of distension, and do not form anything like so close a net-
work as the internal one, ‘The vessels of the external network

ON MEGASCOLEX C@RULEUS. 71

give one the impression of an agency for distributing blood to
the lacuna-like vessels of the internal network, as indeed, I
believe, they are.

The external network is formed by branches of the intestino-
tegumentary vessels.

In the hinder region of the intestine precisely similar
arrangements obtain, but neither network is nearly so dense,
and the internal network quite loses its lacunar character.

In the anterior region it is, owing to the pouches, difficult
to make a flat preparation, and so to see these networks, but
from what I have seen I conclude that the differences between
the arrangements obtaining in this region and those above
described are only slight.

Dorso-tegumentary Vessels (figs. 4 to 7, 9, p. T.).—
These are branches of the dorsal vessel connecting it with the
peripheral networks. In Lumbricus, according to Jaquet (6),
each divides into a “ branche tegumentaire ” and a “ branche
dorso-sous-nervien,” which latter is connected with the nephri-
dial capillaries. According to Perrier (9) a similar arrange-
ment obtains in Urocheta; while in Pontodrilus, which is devoid
of a subneural vessel, the ‘“‘ branche dorso-sous-nervien” is
suppressed ; this is also the case in Megascolex.

All the branches of the dorsal vessel anterior to the hearts,
and one pair of those branches in each segment posterior to
them, belong to this category.

In the most anterior portion of the dorsal vessel they arise
from this latter slightly irregularly, i.e. unsymmetrically
(fig. 8).

In the region of the three pairs of lateral hearts and the
most anterior pair of latero-intestinal hearts there are no such
vessels.

In the region of the four posterior pairs of latero-intestinal
hearts there are such branches, but I have no note as to their
exact place of origin. It may be some months before I obtain
any fresh material, and it is a point of such minor importance
that I leave it undetermined.

In all other segments they arise, regularly, from the dorsal

ip ALFRED GIBBS BOURNE.

vessel immediately posteriorly to the septum which forms the
anterior boundary of the segment in which they lie.

Ventro-tegumentary Vessels (figs. 4, 7 to 9, v. T.).—
These are branches of the ventral vessel connecting it with the
peripheral network. There is a pair of these vessels in every
segment of the body except the first, in which there is a
branch of that belonging to the second segment (fig. 8) on
each side, and except in those segments in which the ventral
vessel is joined by hearts, viz. v111 to XIII.

As mentioned above, the hearts of segments vi and vir do
not join the ventral vessel, and in these segments there is, as
usual, a pair of ventro-tegumentary vessels.

In the last segment of the body the ventral vessel simply
comes to an end by giving off two of these branches (fig. 11).

Dorso-intestinal Vessels (figs. 4 to 7, 10, p.1.).—
These are branches of the dorsal vessel placing it in connection
with the intestinal capillary networks. In segments 1 to Ix
there are no such vessels. In segments x to xr their place
is taken by vessels opening into the supra-intestinal vessels, of
which there are two pairs in each segment. These may be
called supra-intestino-intestinal vessels. In segments xv to
xvi there is a single pair in each segment, connected, as are the
latero-intestinal hearts, with both the dorsal and supra-intestinal
vessel (see fig. 5). In segments xvir to cxxxv there are two
pairs in each segment, the anterior one being always smaller
than the posterior (fig. 10). In segment cxxxvi and the fol-
lowing segments to the end of the worm there is only a single
pair in each segment, the pair which corresponds to the
posterior pair of segments possessing two pairs.

These dorso-intestinal vessels are usually covered by the
yellowish-brown ccelomic epithelium cells which are so con-
stantly found in the dorsal region of the alimentary canal.

The vessels in the segments anterior to the large intestine
soon penetrate the intestinal wall ; of those in the region of the
large intestine, where there are two to the segment, the ante-
rior one always passes round to the ventral region before
penetrating the wall, while the posterior one, after having

ON MNGASCOLEX C@RULEUS 73

received a number of little branches (fig. 7), soon penetrates
the intestinal wall. After penetrating the wall both vessels
pass on towards the ventral line, receiving numerous branches
from the internal capillary network.

All these dorso-intestinal vessels are formed by the lacunar
network, the anterior pair rather nearer the ventral median
line than the posterior pair (fig. 10).

I have now described all the important vessels in Megas-
colex.!

CouRSE OF THE BLOoop.

Having described all the principal vessels and their relations
with one another, I shall now discuss the probable course of
the blood, and put forward a theory which is on the whole
simpler than any which has been hitherto propounded.

The reader should bear in mind throughout the description
that, according to my theory, so long as the modified anterior
extremity, about the first twenty segments, remain intact, and
the thereby injured extremities of the longitudinal vessels
shrink so as to prevent bleeding, it is possible to remove any or
all of the succeeding segments without interfering at all with
the circulation—a most important condition of any tenable
theory, and, moreover, a state of things which indicates the
metamerically segmented character of the vascular system,
always excepting the cephalisation in the anterior region.”

There appears to be entire agreement among previous ob-
servers as to two points only—the forward direction of the

1 Other special vessels have been described in various genera. In worms
possessing a subneural trunk there are of course branches connected witli it.
These branches connect it, I believe, directly with the dorsal vessel, and
indirectly with the intestino-tegumentary vessels (see Jaquet, 6). Where
large nephridia occur the capillary network in connection with them would
come under the group of peripheral networks, and communicate with branches
of the ventro-tegumentary vessels on the one hand, and with branches from
the subneural or (? and) intestino-tegumentary vessels on the other.

2 T shall use the term cephalised region in the succeeding paragraphs as
designating that region of the body in which the vascular apparatus is not
similarly repeated in each segment,

74 ALFRED GIBBS BOURNE.

blood-current in the dorsal vessel, and the downward direction
of that in the hearts.

Of the blood which is brought forwards to the cephalised
region in the dorsal vessel the greater portion goes into the
hearts. Further, some or all of the blood going to some of the
hearts may be derived from the supra-intestinal vessels—some
in worms possessing both lateral and latero-intestinal hearts,
all in worms possessing both lateral and intestinal hearts; the
other and most anterior hearts receive all their blood from the
dorsal vessel.

When we come to the questions—l. Whence comes the
blood into the dorsal vessel? 2. Does any blood leave the
dorsal vessel other than in the cephalised region ?—we find
great difference of opinion. There is in Megascolex, and pro-
bably in other worms, no great inflow at the posterior ex-
tremity ; and when the dorsal vessel is filling, it fills simul-
taneously along the greater part of its length. According to
Perrier (9, p. 504) and Benham (4, p. 255), blood enters the
dorsal vessel from what I term the dorso-tegumentary vessels.
I do not believe this to be the case. Vejdovsky (15, p. 115)
states, though without giving any reason, that the blood flows
from the intestinal capillaries into the dorsal vessel, and in this
I agree with him.

The problem may be stated as follows:

A large quantity of blood leaves the dorsal vessel in the
cephalised region. In some worms some of that blood may
come to the dorsal vessel, in that region, from the supra-
intestinal vessels, i.e. from the intestinal capillaries ; but what
we want to determine is, does any of it come in all worms
from branches connected with the dorsal vessel in the posterior
regions of the body? The vessels which are so connected seem
always to be of two kinds, dorso-tegumentary vessels and dorso-
intestinal vessels ; and there can be, I think, no doubt that, in
order to constantly replenish the dorsal vessel along its whole
length, blood must come from one of these two kinds of
vessels. Does it come from both kinds, or only from one?
If the latter, from which? I think from the dorso-intestinal

ON MEGASCOLEX C@RULEUS. 79

vessels only. Perrier (9, p. 504) having observed the dorso-
intestinal vessels full when the dorso-tegumentary vessels were
empty, and vice versa, comes to the conclusion that these
two kinds of vessels play opposite réles, and decided on other
grounds that the blood flows from the dorsal vessel to the
intestinal capillaries, and towards the dorsal vessel in the
dorso-tegumentary vessels. Benham (4, p. 283), following in
Perrier’s footsteps, brought forward the arrangement of the
valves at the points of junction of these various vessels with
the dorsal vessel in Microcheeta in support of the theory ;
while Vejdovsky, as I have stated above, takes the opposite
view with regard to the direction of the blood-flow in the dorso-
intestinal vessels at any rate.

I can confirm Perrier’s observations that the two kinds of
vessels in question play opposite réles. JI have made this
observation in several recently killed and opened worms of
large size, and also in small worms mounted whole in a live-
box ; further, in a large Megascolex recently killed and opened
I have emptied all these various vessels in one region by gentle
pressure with the finger, and then watched them refill them-
selves; and, moreover, I have cut a vessel of each kind and
watched to discover from which of the cut ends blood flows;
and, lastly, I describe below an arrangement of valves slightly
different from that described by Benham for Microcheta, and
having, I believe, an exactly opposite function.

Observations made in all these various ways have convinced
me that the dorso-intestinal and dorso-tegumentary vessels do
play opposite roles, but in the reverse way to that imagined by
Perrier.

The blood enters the dorsal vessel in each posterior segment
through dorso-intestinal vessels, and leaves it by dorso-tegu-
mentary vessels. The single pair of dorso-tegumentary vessels
are always small as compared with the dorso-intestinal vessels,
and there are in many worms (e. g. Megascolex) two pairs of
these latter in many segments, so that doubtless more blood
enters the dorsal vessel than leaves it in each posterior seg-
ment, and the excess is passed forward to be sent out in the

76 ALFRED GIBBS BOURNE.

cephalised region. With regard to the arrangement of valves,
in Megascolex (fig. 12) there are valves, as Perrier has de-
scribed, in the dorsal vessel between every two segments, and
there is also a valve at the junction of each dorso-intestinal
vessel with the dorsal vessel. These latter valves consist of a
soft-looking tissue which projects as a circular ridge into the
dorsal vessel, and in the centre of this is the aperture of the
vessel, and when the walls of the dorsal vessel contract the
effect must be to occlude the apertures of the dorso-intestinal
vessels. The dorso-tegumentary vessels present no such valves
at their openings into the dorsal vessel, and are placed, more-
over, in the anterior portion of each segment of the dorsal
vessel just posterior to the valve lying in the dorsal vessel
itself, so that the effect of a forward peristaltic contraction of
the latter must be to force blood into these dorso-tegumentary
vessels.

I cannot help thinking that the arrangement of valves de-
scribed by Benham (4, p. 283) for Microcheta needs con-
firmation. His theory that such a valve as he describes at the
entrance to each dorso-intestinal vessel serves to direct the
blood into that vessel does not seem to me to be based upon
sound hydrodynamical principles.

Two additional sets of facts—the relations of the two sets of
intestinal capillaries to one another, and the relations of the
intestino-tegumentary vessels—lend support to my view as to
the direction of the blood in the branches of the dorsal vessel ;
and, moreover, as there is no doubt that in a certain region
blood flows from the intestinal capillaries into the supra-
intestinal vessel or vessels, and so into the dorsal vessel or
into some of the hearts, the opposite view presents this
anomaly, that the direction of the blood-flow in the intestinal
vessels varies according to it in different regions ; in other
words, that some of the dorso-intestinal (or it may be supra-
intestino-intestinal) vessels are efferent intestinal vessels and
others afferent; according to my view they are all efferent,
and further, the flow of blood always takes place from the
dorsal vessel to the various peripheral networks, both in the

ON MEGASCOLEX C@RULEUS. i

region of the body in front of the hearts and in that behind
them.

The ventral vessel has two kinds of vessels connected with
it—hearts and ventro-tegumentary vessels—which communi-
cate with peripheral capillary networks.

The theory with regard to the flow of blood in the ventral
vessel universally current is that it flows backwards along its
whole length. I do not believe that this is the case. By far
the largest amount of (I believe the only) blood coming into
the ventral vessel comes through the hearts, and enters, owing
to the forcible contraction of the latter, at considerable pres-
sure. Why should it flow backwards only? What pressure
can there be in the anterior portion of the ventral vessel to
resist any flow in that direction? The only pressure which
would tend to have this effect would be caused by the flow of
blood from the anterior branches of the dorsal vessel; and if
this blood flow into the ventral vessel it is probable that there
is not a sufficient quantity of it to fill also the intestino-tegu-
mentary vessels, and that the blood in these vessels also flows
forwards. There are other reasons which render this unlikely ;
but even supposing it were the case, we have on the one hand
pressure caused by the flow of blood from the dorsal vessel,
which passes through the dorso-tegumentary branches and
through peripheral capillary networks, and added to this
pressure caused by the flow of blood which has (according to
the assumption) passed through intestinal capillaries, through
the intestino-tegumentary trunks, and finally through peri-
pheral networks; while on the other hand we have pressure
caused by the simultaneous contraction of all the largest and
most powerful hearts. There can be no doubt which of these
two pressures would be the greater—the latter. I conclude,
therefore, that with regard to the ventral vessel, all the blood
which enters it comes from the hearts, and that all the ventro-
tegumentary branches—those anterior to the hearts, as well
as those posterior to them—are efferent vessels. So far as
the ventral vessel itself is concerned, they carry blood away
from it.

78 ALFRED GIBBS BOURNE.

We must now consider the intestino-tegumentary vessels,
and we shall find that the conclusion we have just arrived at
simplifies matters enormously with regard to these vessels.
There has hitherto existed considerable uncertainty as to the
direction taken by the blood in these vessels. Perrier (9, pp.
496, &c.), after an admirable discussion of the subject, in
which he was evidently tempted, on account of the connection
between the capillaries at the anterior extremities of the only
pair of these vessels known to him and those at the anterior
extremity of the dorsal vessel, to believe that there was in the
vessels in question a backward flow, a conclusion to which he
afterwards came for Pontodrilus, states that the blood flows
forwards in them, in Urocheta. If, as I have asserted on in-
dependent grounds, the blood flows forwards in the portion of
the ventral vessel anterior to the hearts, all difficulty disap-
pears, the blood flows from the dorso-tegumentary and ventro-
tegumentary vessels into peripheral networks, and from these
into the intestino-tegumeutary vessels, and from these again
into the intestinal capillary networks. So that the afferent
vessels of these latter are the intestino-tegumentary vessels,
which accords with the theory that I have advocated above
that the dorso-intestinal vessels are their efferent vessels.
These arrangements obtain not merely with respect to the
large anterior pair of intestino-tegumentary vessels of the
cephalised region, but also with respect to those which occur
in every other segment of the body. Yet one more point with
regard to the subject of the preceding paragraphs. According
to previous theories the blood at the anterior region in such
a worm as Megascolex either flows forwards in three trunks
and backwards in one, or backwards in three trunks and for-
ward in one; if what I have said is true, the blood flows
forwards in two trunks and backwards in the other two,
which, as all four trunks are of approximately the same calibre
when filled, seems a more probable arrangement.

Further, a reference to the various memoirs will show what
doubt there has always been as to any special peripharyngeal
vessel, i, e, pair of commissural vessels uniting the dorsal and

ON MEGASCOLEX C@RULEUS. “9

ventral vessels at their anterior extremities. The existence of a
peripharyngeal vessel would be, of course, inimical to my theory.
Even Jaquet (6, p. 340, and fig. 35), who asserts its existence,
figures it as becoming capillary at one portion of its course.
I am certain that it does not exist in Megascolex ; in fact, the
capillaries of the most anterior branches of the dorsal and ventral
vessels are not connected from a functional point of view with
one another, but all with those of the intestino-tegumentary
vessels.

It follows from what I have said that, with regard to the
capillary networks, the afferent vessels of the peripheral net-
works are in all cases branches of the dorsal and ventral vessels,
while their efferent vessels are branches of intestino-tegu-
mentary vessels, and the afferent branches of the intestinal
networks are branches of the intestino-tegumentary trunks, and
their efferent vessels are branches of either the typhlosolar, the
supra-intestinal, or the dorsal vessel, so that blood coming from
them is driven either into the hearts or into the dorsal vessel
at its anterior extremity, and thus in either case into peri-
pheral networks, so into the intestino-tegumentary system, and
once more into the intestinal capillaries.

Every observation which I have made in Megascolex tends
to bear out this theory of the circulation. The theory has, as
I have implied above, this undoubted merit, that it exhibits
the vascular system as a perfectly metamerically segmented
organ, that portion of it contained in the cephalized region
representing, as a whole, almost exactly the portion contained
in any other segment of the body ; the former has undergone,
in fact, a synthesis, and certain additional structures, the
hearts, have become developed in this region.

I may add a word or two about certain special points.

The narrow portions which join the heart to the dorsal or
supra-intestinal vessels, or to both, found in so many worms,
possess no little interest. In the most complicated case, that
of a latero-intestinal heart, blood flows into the heart from
the dorsal and from the supra-intestinal vessel, and fills it ; the
muscles at the point where the heart swells then act like a

80 ALFRED GIBBS BOURNE,

sphincter (they appear to have such a structure), and when the
heart contracts blood cannot flow back into either vessel.
Again, the muscular bulbs at the distal extremities of the
hearts probably act like sphincters, and ensure distension of
the hearts during their diastole, so that the systole has greater
effect ; and after the systole their contraction prevents regurgi-
tation from the ventral vessel. Again, it is interesting to recall
here what is stated above with regard to the intestinal capillary
networks. Blood flows from the external network into the
lacunar spaces, forming the internal network, and at very low
pressure—a circumstance favorable, doubtless, to intestinal
absorption ; thence it drains away gradually into the dorsal
vessel ; indeed, I expect that there is some slight pumping
action exerted by the latter. Moreover, the anatomical
arrangements of these intestinal networks indicate in some
slight degree the probable direction of the blood-flow; the
external network looks like an agency for distributing the blood
to the lacune, and in so far as this is the case it bears out what
is stated above with regard to the direction of the flow.

The peripheral networks deserve a special note in respect of
their triple connections. They are always supplied with blood
by two vessels in which there is some pressure, and so the
blood is pushed on into the third, whichis always some branch
of an intestino-tegumentary vessel, and which thus always con-
veys the blood to the intestinal wall. The arrangement of such
vessels as those shown in fig. 4, g, passing from periphery net-
works in the region of the calciferous glands to the intestino-
tegumentary vessels, once struck me as presenting a little
difficulty. I mean that, as it must according to my theory,
blood should be flowing towards the main trunk of the intestino-
tegumentary vessel in them, whilst it is flowing away from it
in branches which open close by them and go to the intestine.
But it must be remembered that the whole question is one of
relative pressure. In no part of the intestino-tegumentary
system can the pressure be very great, as it is only connected
with the contractile vessels by means of capillaries; but the
vessels in question form part of a set of branches through

ON MEGASCOLEX C@RULEUS. 81

which blood is entering the intestino-tegumentary system at a
certain pressure, which, however slight, would be greater than
that in the intestinal capillaries ; into these latter the blood
would consequently flow.

Summary.—The vascular system consists of a portion in the
cephalized region, and of other portions metamerically repeated
in all succeeding segments.

The cephalized portion differs only from that occurring in
any other segment in having undergone a synthesis, and also
in the presence of contractile hearts.

Throughout the body blood is forced from the contractile
vessels into peripheral networks; thence it is conveyed by a
system of intestino-tegumentary vessels to intestinal capillaries,
and from these it returns to the contractile vessels.

NEPHRIDIA.

Nephridia are present in the form of minute scattered
tubules, and may be seen over almost the entire extent of the
body-wall. There are no large tufts of tubules.

I have not at present worked out the structure of these
Nephridia ; they present peculiar difficulties in that they are
most minute (actually smaller than in any Perichete known
to me), and at the same time the body-wall, as might be ex-
pected from the great size of the worm, is exceedingly thick.

1 The theory, based upon observations on Megascolex, which I have put
forward with regard to the circulation, brings us to such a plausible generalisa-
tion, and is borne out by so many structural details, that I cannot help
thinking it will be found to have a very general bearing among earthworms,
and it may be worth while to speculate fora moment as to the position in
the scheme of the subneural vessels possessed by so many worms. So far as
I can gather from the descriptions of the relations of such vessels they are in
direct connection with the system of contractile vessels, and probably in
indirect connection by means of peripheral networks with intestino-tegu-
mentary vessels (see Jaquet, 6, p. 340). In this case blood passes into
them from the contractile vessels, and ultimately finds its way into intestino-
tegumentary vessels and thence to the intestinal capillaries, and they do not
thus affect the generalisations made above.

VOL. XXXII, PART I.—-NEW SER. F

82 ALFRED GIBBS BOURNE.

NERvows SystTEM.

I have nothing to add to Beddard’s account of this struc-
ture. We have at present very few criteria for any useful
comparison of the nervous system in different worms. It is,
however, well known that there is less variation in the nervous
system as it occurs in various worms than in almost any other
system of organs. This fact—the constancy in structure of
the nervous system in groups where most other organs vary
greatly—so marked among earthworms and leeches, for in-
stance, and so different from what occurs in planarians and
nemertines, will doubtless acquire great interest when we

know more as to the causes of variation in these lower groups
of animals.

GENERATIVE SYSTEM.

It would be going altogether beyond the scope of this paper
to discuss in any detail the relation of this system as it occurs
in Megascolex to that of other earthworms. I give a brief
account only of the arrangements obtaining in Megascolex.

As the only points in which the structures in question differ
greatly from those described by Beddard in ‘“ Pleurocheta”
are such as anyone acquainted with the additions which have
been made to our knowledge of this system of organs since the
publication of Beddard’s paper would have predicted, I shall
not compare my account with Beddard’s in detail.

Testes.—The testes (fig. 3) occur in segments x and x1,
and are attached to the septa bounding these segments ante-
riorly, as is usually the case.

Seminal Funnels.—The seminal funnels lie of course in
the same segments, and are attached to the septa bounding
these segments posteriorly.

Vasa Deferentia.—The vasa deferentia, which are of
course connected with the funnels, are exceedingly minute.
The two on each side soon join together, and, running back-
wards, embedded in the longitudinal muscular layer, open into
the muscular duct of the prostatic gland on each side, close to

ON MEGASCOLEX C@RULEUS. 83

where this latter opens to the exterior. They are ciliated in-
ternally.

Prostates.—The prostates are small relatively to the size
of the worm and very compact, and each is provided with a
very short muscular duct.

Seminal Reservoirs.—There is a single pair of seminal
reservoirs (fig. 3) which lie in segment x1r._ It is very unusual
to find a single pair only among the Perichetide.

Ovaries.—The ovaries lie as usual in segment x111, attached
to the anterior’ septum bounding that segment. They are
very large.

Oviducts.—The oviducts are very small, and open in-
ternally in segment x111 near the nerve-cord; they then pass
through septum x111.xIv and penetrate the body-wall, and
open to the exterior by the pores described above in seg-
ment XIv.

Spermathecx.—There are two pairs of spermathecz
which are of the same size and shape. They are pear-shaped,
and lie in segments vi1t and 1x.! Each possesses a small
cecum, which might entirely escape observation in a dissection,
as it is completely embedded in the wall of the spermatheca
itself at its basal region. It is very obvious in sections, and
contains spermatozoa, while the spermatheca itself is empty in
my specimen. The existence of these cecal diverticula bears
out the views recently expressed by Beddard on this subject.

Accessory Glands.—There only remain to be mentioned
the two pairs of small glands which open between segments
xviI and xvii and xviii and xIx.

These I have ascertained to be small solid glands composed
of clear-looking cells which are not stained by borax-carmine.

1 There has occasionally been some confusion as to the segment in which
the spermathece lie. They usually open in the groove between two seg-
ments and belong to the posterior of these two segments, but the septum is
deficient just here, and they may occasionally be found pushed forward into
the anterior of the two segments; but whenever I have found such to be
the case I have pulled them back through the aperture in the septum, and it
has become evident that they really belong to the more posterior segment,

84, ALFRED GIBBS BOURNE.

These glands do not lie in the body-cavity, but are embedded
in the muscular wall of the body in the spots where they
severally open to the exterior. No trace of them can there-
fore, as Beddard says, be seen in an ordinary dissection of the
worm. I cannot say what their function may be.

BIBLIOGRAPHY.

1 Bepparpv.— On the Anatomy and Histology of Pleurocheta Mose-
leyi,” ‘ Trans. Roy. Soc. Edin.,’ vol. xxx, Part 2, 1883.

2, Bepparp.—‘ On the Genus Megascolex of Templeton,” ‘Ann. Mag.
Nat. Hist.,’? May, 1884.

3. Bepparp.—* Observations on the Structural Characters of certain new
or little-known Earthworms,” ‘Proc. Roy. Soc. Edin.,’ 1887,

4, BenHam.— Studies on Earthworms,’’ ‘Quart. Journ. Micr. Sci.,’ vol.
xxvi, N.S., 1886.

5. Horst.— Ueber eine neue Pericheta von Java,’ ‘ Niederl. Arch. fiir
Zool.,’ Bd. iv, 1878.

6. Jaquet.— Recherches sur le systéme Vasculaire des Annelides,” ‘ Mitth,
Zool. Stat. Neap.,’ Bd. vi, 1885-6.

7. LanKEsTER.—“ The Anatomy of the Earthworm,” pt. iii, ‘Quart. Journ.
Mier. Sci.,’ vol. v, N.S., 1865.

8, Perrier.— Rech, pour servir & l’hist. des Lombriciens terrestres,”
Nouv. Arch, du Mus, d’hist. Nat. de Paris,’ 1872.

9. Perrier. —“ Etudes sur l’organisation des Lombriciens terrestres ”
(Urocheeta), ‘ Arch. Zool. Exp.,’ t. ili, 1874.

10, Perrinr.—* Etudes sur Vorganisation des Lombriciens terrestres ”
(Pontodrilus), ‘Arch. Zool. Exp.,’ t. ix, 1881.

11, Rosa.—“ Nuova classificazione dei Terricoli,’” ‘ Bolletino dei Mus. Zool.
ed. Anat, Comp. Torino.,’ 1888,

lla. Rosa.—* Perichetidi di Birmania,” Viaggio di Leonardo Fea in Birmania
e regioni Vicine, ‘ Ann. Mus. Civico.,’ Genova, 1888.

12, Scumarpa.—* Neue Wirbellose Thiere,” Leipzig, 1861.

13, Spencer.—‘ The Anatomy of Megascolides australis,” ‘Trans,
Roy. Soc. Victoria,’ vol, i, pt. i, 1888,

14, TemeLeton.—“‘On Megascolex ceruleus,” ‘ Proc. Zool. Soc. Lond.,’
vol, xii, 1844 (printed also in ‘Ann. Mag. Nat. Hist.,’ vol. xv).

15, Vespovsky.—‘ System und Morphologie der Oligocheten,” Prag. 1884,

ON MEGASCOLEX C@RULEUS. 85

EXPLANATION OF PLATES VI—IX,

Illustrating Professor A. G. Bourne’s Memoir “ On Megas-
colex ccruleus, Templeton, from Ceylon; together
with a Theory of the Course of the Blood in Earth-
worms.”

Reference Letters to Blood-vessels.

D. V. Dorsal vessel, V. V. Ventral vessel. J. 7. Intestino-tegumentary
vessels. D. 7, Dorso-tegumentary vessels, D, 7. Dorso-intestinal vessels.
V. 7, Ventro-tegumentary vessels, H, Hearts. jer. cap. Peripheral capil-
lary networks. zt. cap. Intestinal capillary networks. §. 7, Supra-intes-
tinal vessels, J. J. J. Supra-intestino-intestinal vessels. a. Vessels passing
from a supra-intestinal vessel to a heart. 4. Vessels passing from the dorsal
vessel toa heart, c, Commissural vessels joining the two supra-intestinal
vessels. d, Anterior extremities of the supra-intestinal vessels, e, Posterior
extremity of the supra-intestinal vessels. # Muscular bulbs at the distal
extremities of the hearts, g. Branches joining intestino-tegumentary vessel,
with periphery capillary networks. 4%, Vessels from the intestinal capillary
networks joining dorso-intestinal vessels. j. Branches joining intestino-tegu-
mentary vessels, with intestinal capillary networks, 4%. Vessel in the intes-
tinal wall (infra-intestinal), passing from the intestino-tegumentary system of
one segment to that of another, /. Origin of dorso-intestinal vessels from
intestinal capillary networks.

PLATE VI,

Fic. 1.—The entire worm drawn from life by Mrs. A. G. Bourne, Natural
size.

Fic. 2.—Ventral view of the twenty-three most anterior segments. m,
Mouth. sp.’ and sp.? Spermathecopores (VII. VIII and viII.Ix). ov. Ovi-
ducal pores. 6. Male pores. g/.! and gi.” Apertures of accessory glands,

PLATE VII,

Fie. 3.—The anterior portion of the worm dissected from the dorsal sure
face. The alimentary tract is represented in horizontal longitudinal section
The septa have been cut in such a way as to render visible as much of the
organisation as possible. Some of the generative organs are shown on the
one side, and some on the other. The segments are numbered I—xIx. m
Mouth. ph. Pharynx. giz,’ Anterior portion of the gizzard, with straining

86 . ALFRED GIBBS BOURNE.

hairs. giz. Posterior portion of the gizzard, showing the thick muscular and
chitinous walls. ca. gl.! The most anterior calciferous gland. ca. gi.® The
most posterior calciferous gland. iz¢. Intestine. m. rad. Radiating muscle.
t.\ and ¢.2 The anterior and posterior testes of the left side. These have
been completely pulled away from the septal wall to render them visible.
fun and fun2 The seminal funnels corresponding to the two testes drawn.
sem. res. Seminal reservoir. ov. Ovary. pr. prostate,

PLATE VIII.

Fic. 4.—Slightly diagrammatic side view of the blood-vessels of the anterior
portion of the body. The intestino-tegumentary vessels and their branches
‘are represented by a dark colour in’ this and the succeeding figures in which
they occur, the other vessels by lighter colour. The greater portions of the
hearts in Segments x—xu have been removed. The numbers I—xtiI are
placed close to the branches of the dorsal vessel belonging to those segments
respectively. The lines numbered tv .v—xviII. xIx mark fairly exactly the
position of the septa at the level of the vessels drawn. Portions of the
alimentary canal are marked as in fig. 3. The various peripheral networks
are shown diagrammatically. a. Vessel passing from the supra-intestinal
vessel to a heart. &. Vessel passing from the dorsal vessel to a heart. ff.
Muscular bulbs at the distal extremities of the hearts. yg. g. Branches of
the intestino-tegumentary vessels bringing blood from peripheral networks.
‘D. I. Anterior, and D. J? posterior dorso-intestinal vessel of one segment.

Fic. 5.—View from the dorsal side of portions of the dorsal vessel
and its connections in Segments vi—xvil1. The dorsal vessel is shown
cut in several places, and the cut ends turned backwards. The hearts
and dorso-tegumentary vessels are shown cut short. a. 4. As in fig. 4.
c. Commissural vessels joining the two supra-intestinal vessels. d@. Anterior
termination of the supra-intestinal vessels. e. Posterior termination of the
supra-intestinal vessels.

PLATE IX.

Fic. 6.—Diagram of the vessels in a segment containing a calciferous
gland. With regard to the origin of the dorso-tegumentary vessel in this

region, see p. 023. B.W. Body-wall. B.C. Body-cavity. Other letters as
before.

Fic. 7.—Diagram of the vessels in a segment in the typhlosolar region.
The arrows denote the direction of the blood flow. J. W. Intestinal wall.
typh. Typhlosole. INZ'. Intestine. 4. Vessels from the intestinal capillaries
joining the dorso-intestinal vessel. 7. Vessels passing from the main intes-

ON MEGASCOLEX C@RULEUS. 87

tino-tegumentary vessel to the external intestinal capillaries. z#¢. cap.
Intestinal capillaries. Other letters as before.

Fic. 8.—Vessels at the anterior extremity of the body. The dorsal and
ventral vessels are shown, but their branches are shown on one side or the
other only. The intestino-tegumentary vessels are shown on the left side
only. cer. Cerebral ganglia. NV. Nerve-cord. The peripheral capillary net-
works belonging to the first five segments are diagrammatically shown and
numbered. per.cap. 1—per. cap. 5. The branches of the dorsal vessel (dorso-
tegumentary vessels) are numbered i—y. ‘The septa are indicated by a line,
and marked Iv. v and V. VI.

Fic. 9.—Slightly diagrammatic view of the vessels in the body-wall, which
is supposed to have been laid open by a cut a little to one side of the dorsal
median line. The object of this figure is to show the relations of the peri-
pheral networks. 7.7. are the branches of the intestino-tegumentary vessels
which pass to the intestinal wall. A comparison of this figure with figs. 7
and 10 will show the whole course and distribution of one of the posterior
intestino-tegumentary vessels.

Fie. 10.—A view from the inside of a portion of the intestinal wall laid
open by a lateral cut to show the intestinal capillaries. The dark coloured
vessels and capillaries are connected by the branch marked 7., and other
similar branches not shown, with the intestino-tegumentary vessel. These
brauches are supposed to be seen through by transparency ; they really lie on
the outside of the intestinal wall. Both capillary networks are in reality a
little finer and denser than shown in the figure. The origin of the two pairs
of dorso-intestinal vessels from the internal lacunar network is shown, and
marked /./. #&. is the small vessel by means of which the intestino-tegu-
mentary system of one segment communicates with that of another in the
intestinal wall, the small cut branches shown connected with this pass to some
portion of the peripheral capillary networks.

Fic. 11.—View of the posterior extremities of the dorsal and ventral
vessels.

Fic. 12.—View of a portion of the dorsal vessel cut open along its
median dorsal line. Sep¢. Septa. 4.0. Valves in the septal regions; the
anterior one shown entire, the posterior one partially cut away. WD. 7.
Apertures of the dorso-tegumentary vessels devoid of any valves. D. I,
D. I. Apertures of the anterior and posterior dorso-intestinal vessels sur-
rounded by the circular valves. 2B. Transverse section through the same
piece of vessel in the region of the apertures of a pair of dorso-intestinal
vessels to show the valves 4’. W. The wall of the vessel.

Fic. 13,—External view of the region of the male pores from a spirit-
preserved specimen to show the median pit. g/.! and g/.? Apertures of the
accessory glands. 4. Male pores.

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ON A LITTLE-KNOWN_ SENSE-ORGAN IN SALPA. 89

On a Little-known Sense-organ in Salpa,
By
Arthur Bolles Lee,

Assistant in the Russian Laboratory of Zoology at Viilefranche-sur-Mer
(Nice).

With Plate X.

THIs organ was mentioned and figured in 1876 by Ussow,
in the Memoirs of the Imperial Society of Naturalists of
Moscow (T. xvit1, Bd. iti, 2, Moscow, 1876). Since then it
has not been mentioned by any author, so far as I have been
able to discover; none of the text-books, at any rate, make
any allusion to it. Ussow’s figure (pl. vii, fig. 49, of the
memoir quoted) is very imperfect, missing all of what appear
to me to be the most interesting points of the subject. His
text is inaccessible to me, being in Russian. I shall do well,
I think, to describe the organ as I find it, with only such
further reference to the work of the eminent Russian naturalist
as may appear useful.

There are, in Salpa mucronata, two of these organs.
They are end-organs of a recurrent twig of the third nerve,
They are symmetrically placed, one on either side of the Salp,
on the median frontal plane, at a point about halfway between
the large muscle that surrounds the anterior orifice of the
mantle, and the most anterior of the three muscle-bands that
form the well-known cruciform figure on the central region of
the animal. Or, in other words, they are situated on the sides
of the Salp, on a transverse plane passing somewhat anteriorly

90 ARTHUR BOLLES LEE.

to the ciliated fossa. These points are shown in fig. 1 (s. 0.,
s.o., the two sense-organs).

By patient manipulation a living Salpa may be so placed
that an immersion lens may be brought to bear on one of the
sense-organs, and the points shown in fig. 2 may then, with
some care, be made out. The organ consists of a stem ter-
minating in a bulb, which is surmounted by a delicate hyaline
claviform appendage. The stem is seen to be a cellular
tube formed by a process of the inner mantle. (Ussow’s figure,
on the contrary, shows clearly that the author considered that
the inner tunica did not penetrate into the tube.) This tube
traverses the entire thickness of the hyaline outer mantle or
tunica, and swells at its distal end into the bulb. The bulb
seems, in favorable specimens, to lie simply in a cup-shaped
depression of the tunica. The lumen of the tube, in which the
nerve is frequently very clearly shown, is seen to be continued
for some distance into the bulb, and to expand there into a
cavity or vestibule (frequently somewhat more voluminous
relatively to the lumen of the tube than that shown in the
figure), in which the nerve breaks up into a delicate arbo-
rescence whose twigs terminate by means of small conical
swellings on the roof of the cavity (v. in the figure). From
the distal surface of the bulb delicate tapering hairs are seen
to arise by means of a conical base, and to be continued for
some distance into the claviform appendage. In the most
favorable cases they may be followed for about one third of
the length of this appendage, never further in the living state
(unless the specimens have been stained intra vitam with
gentian violet or the like). The appendage itself presents
but a single contour, is absolutely hyaline, and gives the
impression of being an extremely thin-walled tube or vesicle.
This is the view apparently expressed in Ussow’s figure, but
we shall find that it does not answer to the reality. With
great care this single contour may be followed to the bottom
of the cup-shaped depression of the tunica spoken of above,
but is seen to be continuous there with the wall of this cup;
so that the appendage is seen to be a papilla of the hyaline

ON A LITTLE-KNOWN SENSE-ORGAN IN SALPA. 91

tunica, and the bulb is seen not to lie strictly speaking in a
hollow of the tunica (for it nowhere breaks through the surface
of the tunica), but correctly speaking to be surrounded by a
deep and narrow annular depression or moat excavated in the
substance of the tunica around the bulb, but. at a little dis-
tance from it (am. in the figure). The moat is not figured by
Ussow. Little more can be made out. by study of the living
object.

It may be added here that the shape of the appendage is
somewhat variable. It is club-shaped, inasmuch as it is
always somewhat thicker at the tip than lower down; but it
may swell out very gradually towards the tip, as in fig. 2, or
suddenly, as in fig. 3. These two figures represent extreme
cases of the relative degrees of swelling that are found. The
axis is normally straight, not curved as in Ussow’s figure,
which I take to represent a pathological state.

In fixed and stained preparations the appearances are very
different. As to the stem, it will be sufficient to note that the
lining epithelium of the tube is composed of cells similar
to those of the inner tunica, of which it is a process—clear
cells with somewhat loose-textured nuclei—nuclei in which the
chromatic “ mitome” is readily discernible as such.

The bulb, in good preparations, may be analysed into two
constituent elements—a centrally placed tuft of sense-cells, and
a surrounding cup-shaped wall or calyx of supporting cells.
These two groups of cells are readily distinguished by their
different optical aspect.

The cells of the calyx are clear cells, in the sense in which
that appellative was just now given to the cells of the stem;
whilst the sense-cells are dense, stain more deeply, and have
compact nuclei in which a normal chromatic mitome can only
be made out by careful examination with the very highest
powers. Fig. 3 shows these points in a sufficiently realistic
manner.

The structure of the calyx is the most difficult point to eluci-
date in the whole of the organ. In shape the calyx is variable.
In Salps of not more than 38 mm. in length it is generally regu-

92 ARTHUR BOLLES LEE.

larly ovoid, or even almost globular (fig. 3). In Salps of from
5 to 7 mm, in length it is more elongated, and presents a
contour of an elegant double curvature, the basal portion con-
taining the vestibule being much more drawn out than in earlier
stages (see fig. 4). It is composed of clear, flattened, squa-
mous cells, thicker at their proximal margins than at their
distal margins, Their nuclei, loose-textured like those of the
epithelium of the stem, occupy the proximal region of the
cells.

There are two layers of these cells (evt. ca, and int. ca. in
figs. 3 and 4). By reference to fig. 3 it will be seen that doubt
is permissible as to the proper way of describing the position
of these two layers, The cells marked int. ca. appear to be
internal with regard to the cells marked eat. ca.; but they
might also fittingly be described as “upper” or “ distal” cells.
And seeing that the calyx is evidently only an expanded portion
of the tube of the stem, which is lined by only a single layer
of epithelium cells, this may appear the more fitting view. I
incline, however, to the former view. ‘The impression left on
my mind by a most careful study of this object is that the
present arrangement is the result of the invagination of the tip
of a primitive hollow outgrowth of the inner tunica, the inva-
ginated cells forming an inner layer to the cup thus produced,
and the sense-cells being differentiated from the cells at the
fundus of the cup, or, if previously formed at the tip of the
outgrowth, being invaginated with the rest. It should be added
that [have many times distinctly observed the nuclei of the
outer cells to be placed on their outer wall, whilst the nuclei
of the inner cells were placed on their inner wall, which is the
place they would occupy if the latter are only an invaginated
state of the former,

Attempts to elucidate this point by direct observation of the
genesis of the organ have failed, as it was found that, owing to
the very precocious development of the organ, an arduous study
of serial sections of the developing stolon would be necessary,
and the case does not seem to warrant the expenditure of so
much labour.

ON A LITTLE-KNOWN SENSE-ORGAN IN SALPA. 93

At the base of the calyx its walls are relatively thick.
Towards its mouth they thin down, and terminate in a
beautifully delicate, thin, hyaline, incurved lip (/. ca. in figs.
3 and 4). According to my view, both layers of cells must
concur in the formation of this lip, but are there indistin-
guishably fused together.

Ussow does not seem to have seen the calyx at all.

The nerve runs down the greater part of the length of the
stem without dividing. At ora little before its entrance into
the vestibule it begins to break up, first into about three
branches, then into about six or seven, and finally, by what
appears to me a strictly dichotomous mode of division, into as
many twigs as there are sense-cells. This last division always
takes place quite close to the sense-cells.

The sense-cells are spindles of about 15u in length and
5u in breadth. They have an oval nucleus, of very compact
texture, somewhat basally placed, with its longer axis parallel
to the longer axisofthe bulb. They, or at least the more peri-
pherally placed ones, are curved spindles, their outlines follow-
ing in the main the lines of the calyx, and their distal ends
especially being sharply bent inwards, so as to bring them
within the orifice of the calyx (which is frequently much nar-
rower than in the figures). I believe that their number is
constant, and that there are just fourteen of them in all, They
can be counted with considerable certainty in optical transverse
sections (see fig. 5).

The sense-cells are embedded in a finely granular matter,
which may often be seen to project in a flattened dome out of
the mouth of the calyx (gr. in figs, 3and 4). I would suggest
that this granular matter is perhaps secreted by the inner cells
of the calyx. It is not figured by Ussow.

Proximally these cells taper down gradually into their
respective nerve-twigs. JDistally they taper down equally
gradually, and without the intervention of any “ granule” or
other apparatus of the sort, into delicate hairs. These hairs
are very pale, almost invisible in the living state, smooth or
very delicately mottled and corrugated. They stain with great

94 ARTHUR BOLLES LEE.

readiness with certain stains, namely, methyl green, dahlia,
gentian, methylen blue, and hematoxylin. The stain obtained
by the first three of these reagents is not of the normal colour ;
itis areddish purple. It takes effect with the greatest energy on
the distal portions of the hairs, the basal portions being hardly
affected by it (except in the case of an over-stain with hema-
toxylin). The hairs are from 054 to lu thick, and from
120u to 150u long.

The aspect of the claviform appendage is totally changed in
stained preparations. The fine outer contour described as
visible in living specimens is in many examination media quite
invisible. If visible in balsam or damar preparations it is in-
variably greatly shrunken, generally to a far greater extent
than is shown in fig. 8. On the other hand, such preparations
reveal the presence of a lumen much narrower than could be
suspected from the examination of living specimens. It is of
approximately equal diameter throughout the length of the
appendage, measuring about 44 in general; so that the walls
of the appendage, instead of being almost immeasurably thin,
have a thickness, in the living state, of 4 to 5u or more. In
very shrunken specimens they are practically indemonstrable,
and the whole appendage appears reduced to the state ofa
mere whip, consisting of the sense-hairs and the boundary of
the lumen.

The lumen does not pierce the wall of the appendage at its
tip; there is no trace of any terminal pore.

The sense-hairs are inserted into the wall of the appendage
at the extremity of the lumen. They generally merge insen-
sibly into the substance of the wall, though a slight cone or
swelling may sometimes be detected at the point of insertion
(¢.s. A. in fig. 3).

Ussow does not seem to have seen the true lumen, and seems
to conceive of the sense-hairs as floating freely in a thin-walled
vesicle represented by the outer contour of the appendage.

I have not been able to discover the organ either in the
solitary form—S. democratica, or in any other Salpa.

In the explanation of his figure Ussow calls the organ

ON A LITTLE-KNOWN SENSE-ORGAN IN SALPA. 95

a tactile organ. I am altogether disinclined to admit this
interpretation. The organ seems to me to have nothing speci-
fically tactile about it. The claviform appendage is wanting in
the most essential quality of a mediator of tactile impressions
—stiffness. There is nothing hard about it, it is eminently
soft and yielding. Compare it with the hard chitinised tactile
hair of an Arthropod, say the larva of a Dipteron, and this
difference of mechanical principle becomes conspicuous at
once. Inthe Arthropod a stiff appendage, often arranged as
a lever—here, the softest of papille, a mere jelly !

Looking to the mere essential morphology of the organ, the
nature and arrangement of its constituent cells, a different
point of view suggests itself. Fusiform sense-cells, with
terminal hairs, surrounded by supporting cells, and sunk in a
depression of the ectoderm, that is the schema of a taste-bulb.
And if the organ of Salpa mucronata were not surmounted
by the claviform appendage, or if even this appendage were
pierced by the smallest pore at its extremity, I suppose no one
would be inclined to object to its being considered a taste-
bulb. But the presence of the closed appendage does not scem
an insuperable objection to this view. The walls of the
appendage, though, as I have shown, they are relatively
thick, are after all thin structures enough, and that they must
be highly permeable is shown by the readiness, above noted,
with which the terminal hairs stain with certain reagents.
This reaction can be readily obtained in the living state by
means of gentian violet or dahlia dissolved in seawater. The pene-
tration is by no means instantaneous, by no means rapid enough
to make the organ useful in the selection or rejection of sapid
food-substances (which of course, from its position, cannot be
its function), but it is perhaps rapid enough to allow it to be
useful as an indicator of the chemical quality of the water in
which the animal swims.

These considerations appear to me to give much plausibility
to the view that the organ is either a taste-bulb, or at least
was one once. But there is yet another possible function
for which there is perhaps even more to be said—a mode of

96 ARTHUR BOLLES LEE.

action according to which, as it seems to me, this mechanism
cannot help acting. The cellulose or “ tunicin’’ mantle of
Salpa is a fairly tough but highly watery and highly hygro-
metricjelly. Givena papilla such as this claviform appendage,
with sensory hairs inserted into its tip, if the density of the
ambient water be changed, the shape and dimensions of the
papilla must be changed in consequence, it must elongate or
shorten, and the hairs must be stretched or slackened. If the
animal pass into less dense sea water, that is, a medium richer
in pure H,O, water will be taken up, the appendage will swell
and elongate, and the hairs will be pulled. If the animal be
carried into water richer in salts the claviform appendage will
shrink, and the hairs must be relaxed.

And it seems to me that I have evidence that these actions
do actually take place. I have noted above that the shape of
the appendage is variable. I have to add here that its surface
is typically smooth, but very frequently corrugated or wrinkled
into lines of shrinkage. This is especially the case with
specimens that have been kept for some time in captivity, and
with such as have been treated with chloral hydrate or solutions
of intra-vitam staining agents. Even a little methylen blue
added to the sea water, in so small a proportion as not perceptibly
to stain the hairs, may produce this effect. And in such
specimens the hairs themselves, which in perfectly fresh
specimens have a smooth and regular contour, are seen to have
a markedly corrugated outline, and sometimes a very great
degree of curl or twist near the tip.

And, all things considered, I incline to regard the organ asa
sensory areometer or hydrometric apparatus.

ON A LITTLE-KNOWN SENSE-ORGAN IN SALPA. 97

EXPLANATION OF PLATE X,

Illustrating Mr. Arthur Bolles Lee’s paper ‘On a Little-
known Sense-organ in Salpa.”

Fie. 1.—Anterior half of a Salpa mucronata. ¢w. The tunicin mantle.
g. The ganglion. ci. The ciliated fossa. mo. “ Mouth” of the mantle.
$8. 0., 8. 0. The two sense-organs. X 20.

Fic, 2.—A sense-organ in the living state. c/. The outer contour of the
claviform appendage. az. The annular depression. v. The vestibule of the
bulb, in which the terminal arborescence of the nerve is well seen. Optical
section. x 1100.

Fre. 3.—A sense-organ after preparation (solution of Flemming, Henneguy’s
acetic-acid alum carmine, damar). c/. The outer contour of the claviform
appendage. JZ. Itslumen. J/. ca. Lip of the calyx. zt. ca. Internal cells of
the calyx. ext. ca. External cells of the calyx. gv. Granular mass in which
the sense-cells areembedded. ¢.s./. Termination of sense-hairs. az. Annular
depression of the tunicin mantle. v. Vestibule of the bulb. Optical section.
x 1000.

Fic. 4.—A similar preparation without the claviform appendage. The
letters as before. Optical section. x 1600.

Fic. 5.—A similar preparation seen in optical transverse section. ca. The
cells of the calyx. s.c. The sense-cells, the letters pointing to a fourteenth
cell, which is indistinctly seen. x 1100.

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IMMUNITY AGAINST MICROBES. 99

Immunity against Microbes.
By

M. Armand Ruffer, M.A., M.D.(Oxon.)

Parr: —— Ay

WHEN examining the microscopic world actively growing in
various infusions, we are often able, says Metschnikoff,! “to
follow with our eyes the struggle taking place between the
representatives of the Flora and Fauna of microbes. Many
unicellular animals, such as Amcebe and other Rhizopoda, as
well as flagellated and ciliated Infusoria, feed on various
bacteria, devour them in great numbers, and take them into
their protoplasm in order to extract from them the necessary
nutritive material. Small Monads in the course of a few
minutes take into their interior Leptothrix filaments ten times
longer than they are themselves.” It is a very old observation
that Amcebe and other Protozoa seize dead particles, and,
after extracting the nutritive material contained in these, reject
the remainder.

According to Metschnikoff,? there is no proof that the cells
forming the ectoderm of some lower animals—sponges for in-
stance—have the power of taking solid substances into their
interior. The same observer, however, has shown that the
ectodermic cells of other Metazoa have that power. If, for
instance, a small quantity of powdered carmine or indigo be
added to water containing Plumularia (Plumularia setacea
or others), some of the carmine or indigo finds its way into the
ectoderm of the Nematocalyces. These organs send out

1 «Ann. de l'Institut Pasteur,’ 1887, p. 321.
2 © Arbeiten aus dem Zool. Institut in Wien u. Triest,’ vol. v, 1884.

100 M. ARMAND RUFFER.

pseudopodia of various shapes, which attach themselves to a
calyx or to its contained polyp, or more frequently flatten
themselves out around the stem so as almost to encircle it.
The ectodermic cells of the free extremities fuse into a common
protoplasmic mass, which sends out some few pseudopodia.
The slow creeping movements of the ends of the Nematocalyces
serve to clean the neighbouring polyps—a function accounting
for the frequent presence of foreign particles in their ectoderm.
The Nematocalyces of Plumularia, moreover, eat up three parts
of the animal which has died—the polyp heads, for instance—
and are frequently crammed with dead particles which they have
ingested. Nematocalyces, therefore, have a double function ;
namely, that of scavenging the outside of the animal by
creeping along it, and that of eating up useless tissues. The
ectodermic cells of the larve of Setinz also absorb solid mate-
rial after enclosing it with their pseudopodia.

Intracellular digestion by ectodermic cells, however, is not
observed frequently, whereas the absorption and digestion of
solid particles by mesodermic structures are easily demon-
strated. The nutrition of fresh-water sponges, for instance, is
carried out by wandering cells, which correspond to the meso-
dermic cells of higher animals.!

Hackel? noticed the fact that particles of indigo when
injected into one of the Mollusca (Thetys) are absorbed by
the corpuscles of the blood, but, although this contribution to
the knowledge of the functions of mesodermic cells is impor-
tant, still more may be learnt from the study of processes
taking place during the destruction of useless organs in the
period of metamorphosis of Invertebrata.

Taking Auricularia® as an example, we find that resorption-
phenomena occur during two stages of its life history, namely,
during the assumption of the pupa-stage, when a part of the
longitudinal ring of cilia is lost—that is, is disintegrated and

1 Compare, Metschnikoff, ‘Zeitschr. f. wiss. Zool.,’ vol. xxiv, p. 10, and
T. E. Schulze, ibidem, vol. xxv, p. 258.

2 Radiolarien, 1862, p. 104.

3 Metschnikoff, ‘Quart. Journ. of Mier. Sci.,’ vol. xxiv, p. 10.

IMMUNITY AGAINST MICROBES. 101

devoured by the mesoderm—and, secondly, during the meta-
morphosis of the pupa into a young Synepta, when mesodermic
cells become active again, collect beneath the ciliated rings,
and eat up the products of disintegration. Every amceboid
cell during both these stages is generally loaded with an
enormous mass of débris-granules contained in the interior of
clear vacuoles.

In Asterides also, whole segments of larve are absorbed by
mesodermic cells during the metamorphosis. This process, in
fact, appears to be the rule in all Echinoderms.

Mesodermic cells were supposed by Ganin! to play an active
part in the histolysis of muscular fibres during the metamor-
phosis of flies. More lately, Kowalewsky? showed that Ganin’s
supposition was correct, for he proved that large numbers of
wandering cells penetrate between the muscular fibres, take
into their interior and digest the latter.

Mesodermic cells are active not only during the period of
metamorphosis, but under many other circumstances. Schneider?
noticed that the resorption of the genital organs of Hirudo is
carried out by ameeboid cells greatly resembling blood-
corpuscles, and Metschnikoff described the same phenomenon
as taking placein the Aureliaaurita.* The developing nemer-
tine in a pilidium also, if the latter is left for some length of
time in a glass vessel, atrophies and is devoured by amceboid
mesodermic cells. Some of the mesodermic cells of transparent
salt-water animals taken fresh from the sea are filled with
foreign matter, and Metschnikoff assumes that such foreign
bodies gain an entrance by piercing through the body-wall,
being ultimately swallowed by ameeboid cells. Particles of
carmine suspended in water not only penetrate into the cells
of the entoderm, but into those of the mesoderm also.

The following experiments throw much light on the pro-
cess of intra-cellular digestion by mesodermic cells. When

1 Ganin, ‘ Beitrage, &c.,’ 1876, p. 40.

* Kowalewsky, ‘ Zeitschr. f. wiss. Zool.,’ vol. xlv, 1887.
> Schneider, ‘Zool. Anzeiger,’ 1880, p. 19.

4 Metschnikoff, loc. cit., p. 89.

102 M, ARMAND RUFFER.

carmine or indigo powder suspended in water is injected under
the ectoderm of Bipinnaria asterigera or PhyllirrhGée
bucephalum, the coloured powder is soon taken into the in-
terior of the mesodermic cells. In Bipinnaria the carmine is
absorbed by both large and small cells ; in Phylirrhée, on the
other hand, the powder is ingested by the small cells, the
larger ones containing dissolved carmine ouly. The small
particles of carmine are actually in the interior of cells, but
the larger particles of the same matter become surrounded by
numerous small cells which form plasmodia resembling the
giant-cells found in the pathological structures of vertebrata.
Similar plasmodia formed by mesodermic cells are formed
when a few drops of human blood are introduced under the
cuticle of Bipinnaria asterigera; the nuclei of such plas-
modia lie at the periphery, whilst the centre of the plas-
modium contains a mass of red blood-corpuscles in different
stages of degeneration.

Migrating cells also form a barrier around foreign bodies,
such as glass, cellulose, &c., which are introduced under the
skin of these animals; but, in many cases, no plasmodia are
seen, so that the formation of the latter structures does not
necessarily follow on the introduction of foreign bodies into
these animals.

Blood-vessels, if present, do not necessarily play a part in
the process of inflammation, for no transudation takes place
from them, provided the foreign body be introduced without
wounding a blood-vessel. Cohnheim’s dictum, therefore—
‘‘ Ohne Gefasse keine Entztindung”—is not altogether correct,
or, as Metschnikoff brilliantly says, “Inflammation is, genea-
logically speaking, of a much older date than the formation of
vessels, and exudation is a comparatively late phenomenon.”

Mesodermic cells, however, do not eat everything they
come across, but have the power of exerting a choice. Thus
the mesodermic cells of Phyllirrhée make no attempt to destroy
the fresh eggs of Spherechinus granularis injected under
the skin of the animal. Nay, more; the eggs retain their
vitality, so that they may be fecundated artificially whilst

IMMUNITY AGAINST MICROBES. 103

living in the animal’s tissues, and may form normal blastule.
Contrariwise, the living spermatozoa of Spherechinus
granularis introduced under the skin of Phyllirrhde soon
fall a prey to mesodermic cells.

The mesodermic cells of Bipinnaria asterigera also eat
and destroy the necrosed parts of the animal.

The function of ameeboid cells! is not limited to the absorp-
tion of weakened or dead tissues; but the same structures
take an active part in the fight between the animal organism
and the surrounding microbes. When a solution crowded
with micro-organisms is injected under the skin of Bipin-
naria or Phyllirrhde, the parasites, whether mobile or not,
are soon taken into the interior of the protoplasm and
vacuoles of mesodermic cells, and gradually digested by the
latter. The tunica of Botryllus, for instance, even in an
animal taken fresh from the sea, always contains colonies of
micro-organisms. The latter are actively pursued by the wan-
dering cells of the tunica, which digest them after swallowing
them up. The struggle is not a one-sided one, however, for
the victory often remains with the micro-organisms, as is
proved by the presence of dead ameeboid cells containing a
number of dead bacterial filaments radiating out of them.

Similar facts have been observed in Daphnia by Metsch-
nikoff.2. These small fresh-water crustaceans are frequently
invaded by a fungus belonging to the yeast family (Mono-
spora bicuspidata). The long needle-like spores of the
parasite penetrate with the food into the alimentary canal, pass
into the body-cavity after perforating the walls of the intestine,
quickly invade the entire body, and often kill the animal.
Whilst the disease is in active progress the leucocytes still
strive to fight, and absorb numbers of conidia. The latter,
however, rapidly multiply, and destroy the ameboid cells, the
victory ultimately falling to the parasites.

The study of the development and of the life-history of
Vertebrata affords many examples showing that the meso-

1 «Annal. de l'Institut Pasteur,’ 1887, p. 323.
2 Thid., p. 325.

104. M. ARMAND RUFFER.

dermic structures of higher animals have similar functions.
A few instances will suffice to illustrate this.

When the tadpole passes into the frog stage, the muscles and
nerves of the tail gradually disappear as they become a prey
to amoeboid cells, which surround them and eat up the tissue
which still presents the structure of normal muscular fibres.
This process is precisely similar to that which Kowalevsky
has observed during the metamorphosis of flies.

Again, amceboid mesodermic cells collect round the muscles
of the tail of living, uninjured Bombinator larve! at the be-
ginning of the metamorphosis, and gradually devour these
structures. During the progress of the atrophy of the gills
also, the presence of large fully-laden mesodermic cells may be
easily demonstrated.

The mesodermic cells of higher Vertebrata have similar
functions. The resorption of osseous substance’ constantly
taking place in the shaft of long bones is essentially dependent
on the presence of large multinucleated cells (Osteoclasts,
Myeloplaxes of Robin), which excavate small shallow pits
(foveole) in the part which is undergoing absorption.

Large mono-nucleated cells containing red blood-corpuscles
in their interior are also met with in the spleen.® These intra-
cellular red blood-corpuscles may be normal in appearance,
but more frequently they appear to be disintegrated and
digested in the interior of these large spleen-cells, a few
pigment-granules representing all that remains of them. Many
observers have succeeded in watching the process of intra-
cellular digestion of red blood-corpuscles actually taking place.

Mesodermic cells not only devour other structures, but prey
on one another also. According to Heidenhein, the walls of
the intestinal canal of certain animals,‘ more especially guinea-
pigs, are lined by epithelium cells, between which large
mesodermic cells find their way to the surface and absorb

' Metschnikoff, ‘Quart. Journ. of Micr. Sci.,’ vol. xxiv, p. 112.

2 Kolliker, ‘ Die normale usorption des Knochengewebes,’ Leipzig, 1873.
3 Bardach, ‘Ann. de l'Institut Pasteur,’ Dec., 1889, p. 599.

4 Heidenhein, ‘ Pfliiger’s Archiv,’ vol. xlvii, 1888,

IMMUNITY AGAINST MIOROBES. 105

smaller lymphocytes, which are ultimately digested in the
interior of the larger cells.

The writer! found in the lymphoid structures (tonsils,
Peyer’s patches, mesenteric glands) of the alimentary canal of
many animals large wandering cells containing two, three, or
more lymphocytes in their interior, and described all the stages
of the intracellular digestion of the latter.

More lately, the writer? has discovered that the so-called epi-
thelioid cells of the spleen-pulp have the power of taking into
their interior and of digesting smaller amceboid cells. The same
cells also swallow inert substances such as vermilion when this is
injected into the blood-stream, this fact showing that they are
amceboid structures—a supposition suggested by their irregular
shape.

Before proceeding further, it is necessary to discuss the
origin of these cells—Phagocytes, as Metschnikoff has called
them. “The function of Phagocytes,’ says Metschnikoff,?
is usually the property of two kinds of cells. Small cells (mi-
grating cells) possessing one or many nuclei—leucocytes in
the narrower sense of the term—are scattered through all
tissues and concentrated in the lymphatic and blood-system,
but emigrate in case of need to any part of the body which is
invaded by parasites. I give to these cells the comprehensive
name of Microphages. On the other hand, I give the name of
Macrophages to the fixed cells of connective tissue, the epithe-
hoid cells of pulmonary alveoli, in fact, to all kinds of structure
which possess the power of taking solid bodies into their interior,
and which are provided with a single large nucleus, less easy
to stain than the nuclei of Microphages.” This definition,
although clear, is, as will be seen later on, not quite adequate.

Since Metschnikoff’s first paper appeared it has become
evident that the Macrophages are often, if not always, derived
from Microphages. Heidenhein,* describing the Macrophages

1 Armand Ruffer, ‘ Quart Journ, of Mier. Sci.,’? February, 1890.

? The writer gave a short account of these researches before the British
Medical Association, August, 1890.

% Metschnikoff, ‘Ann. de l'Institut Pasteur,’ 1887, p. 324.
4 Heidenhein, loc. cit,

106 M. ARMAND RUFFER.

found in the intestinal canal, says, “Now there does not
appear a doubt that these large giant-cells are developed from
the ordinary lymphocytes,” and proceeds to give figures
showing their gradual development. In the writer’s paper on
the “ Phagocytes of the Alimentary Canal,’’! he showed that
the Macrophages of the Peyer’s patches and tonsils are derived
from small lymphocytes, and he described the mode of deve-
lopment of these enormous cells. In the spleen-pulp and
venous sinuses of the same organ also the writer has been
able to trace the development of the Macrophages from Micro-
phages, for ali intermediate stages between them can easily be
observed.

It has been shown that the mesodermic cells of invertebrata
not unfrequently come to the surface, scavenge the outside of
the animal’s body, and remove any dead particles which they
may come across. In vertebrata the same process takes place in
the mucous membranes lining the cavities of the body which
are in contact with the external world. Eberth®, as early as
1864, noticed that wandering cells force their way between the
epithelium cells covering the intestinal canal. Ph. Stohr* then
drew general attention to the fact that this emigration takes
place along the whole length of the alimentary tract. The
writer has shown that the leucocytes of dogs wander to the
free surface of the tonsils, take particles of dust, charcoal, &¢.,
into their interior, and carry them into the depths of these
organs. Here the Microphages and their contents frequently
fall a prey to the Macrophages present in these structures. This
fact proves that the mesoblastic cells of vertebrata have func-
tions similar to those of invertebrata, but evidence far more
conclusive can be obtained by observing what takes place in
the lungs of various animals.

Particles of charcoal, dust, &c., are to be met with in the
epithelioid cells (Saub zellen) of the lungs of most animals.
The precise nature of these cells remained uncertain until the

1 Armand Ruffer, loc. cit.

2 Bberth, ‘ Hiirzburger Naturwissenschaft. Zeitschrift,’ 1864, p. 23.
3 Ph, Stébr, see Heidenhein, loc. cit.

IMMUNITY AGAINST MICROBES, 107

experiments of Tschistowitsch! demonstrated the fact that they
belong to the group of Phagocytes, and are therefore probably
derived from ordinarylymphocytes. These cells remove particles
of foreign matter which have entered the air-passages, and
carry them to the neighbouring lymphatic glands, where they
are again arrested and often remain permanently. In the
bronchial glands of London cats, for instance, which always
contain a large quantity of charcoal, the particles of charcoal,
dust, &c., are contained in the interior of Jarge mono-nucleated
epithelioid cells which are real Macrophages.

The free surface of vertebrata, like the free surface of in-
vertebrata, is everywhere in contact with a large number of
micro-organisms. Nevertheless, the internal organs of the
body, the liver, kidneys, spleen, &c., contain no parasites, and
it follows that there must be some mechanism preventing the
entrance of micro-organisms into a healthy animal’s tissues, or
some means of destroying them should they gain admittance.

The skin of all animals is an impenetrable barrier to the
entrance of micro-organisms, and, to all appearances, the latter
do not make an attempt to penetrate through the same into
the tissues. It is plain that micro-organisms cannot easily
force their way through several layers of hard epithelium cells,
such as go to form the outer skin of most mammals or cover
the surface of the lips and tongue; but it is evident also that
micro-organisms cannot penetrate through a single layer of
epithelium cells. In sections passing through the walls and
contents of the intestines of any animal, an appalling number
of microbes are seen lining the free border of the layer of
epithelium cells, but none are found in or between the latter.
The epithelium cells therefore, together with the cementing
substance between them, form a satisfactory barrier to the
entrance of microbes.

Matters, however, are not quite so simple as they appear, for
in some organs, of healthy animals even, a struggle takes place
during every hour of the day between micro-organisms and
mesodermic cells.

1 Tschistowitsch, ‘ Ann. de l'Institut Pasteur,’ July, 1889.

108 M. ARMAND RUFFER.

In 1887, Metschnikoff found that the leucocytes present in the
mucus covering the tonsils of healthy people frequently contained
micro-organisms. In 1885, Ribbert and Bizozzero examining
the Peyer’s patches and vermiform appendix of healthy rabbits,
showed that the walls of these organs are crammed with an
enormous number of degenerated micro-organisms; both these
observers attributing the death of the parasites to the action of
the cells of the part. In 1890, the writer! demonstrated the
fact that the leucocytes which wander out between the epithe-
lium cells to the surface of the iymphoid organs of various
animals—more especially rabbits—are frequently filled with
micro-organisms. The leucocytes(microphages) carry their prey
into the interior of the tissues, but, being weakened by the
secretions of the microbes they contain, are frequently eaten
up by larger cells (macrophages), which resemble in all particu-
lars the macrophages described by Metschnikoff in pathological
specimens. The writer has shown that the same process takes
place in the tonsils also. The conclusion to be drawn from
these researches is, that the mesodermic cells, of healthy animals
even, have the power of absorbing, destroying and digesting
micro-organisms.

A little consideration shows that the lungs and intestinal
tract, which are the organs most likely to be attacked by micro-
organisms, are provided with a large number of mesodermic
cells. Attention has been drawn to the fact that mesodermic
cells proceed to the surface along the whole length of the ali-
mentary canal and destroy micro-organisms. Moreover,
immediately below the layer of epithelium cells and along the
whole length of the intestinal tract, a layer of adenoid tissue is
spread out like a lymphatic gland. Every micro-organism,
therefore, which, by any chance, passes the first barrier of epi-
thelium cells must run the gauntlet of this adenoid layer and
most probably be destroyed.

In the lungs, similar macrophages line the alveoli and remove
dust, particles of charcoal, &c., which, by gradual accumula-
tion, might choke the air-passages. Surgeons know—and Sir

1 Armand Ruffer, loc. cit.

IMMUNITY AGAINST MICROBES. 109

Joseph Lister was the first to draw attention to that fact—that,
should the skin remain intact, a wound of the lung rarely becomes
septic. Moreover, the elegant researches of Tyndal, Gunning,
Straus and H. Dubreul, and Straus!, prove that expired air
contains no microbes—that is, has been filtered in the lungs.
Other investigations have demonstrated the fact that the germs
stick to the moist surfaces of the upper air-passages, and that
very few only are carried into the lungs. Doubtless some are
carried into the alveoli, just as carbon-particles are carried
there.

M. Tschistowitsch? has lately shown what becomes of micro-
organisms which have been introduced into the lungs. He
found that micro-organisms which are non-pathogenic for the
animal used, are at once attacked by the phagocytes of the lung
which swallow, digest, and destroy them; the safety of the
animal being therefore dependant on the efficient action of
these cells. It follows that the bacilli which are carried by
the stream of air into the lungs, must at once be seized upon
and destroyed by these phagocytes. ‘The fact that the lung
consists of a number of small aseptic cavities can thus be readily
explained.

Having now studied the functions of wandering mesodermic
cells in healthy animals, let us see how these same cells react
when micro-organisms find their way or are artificially intro-
duced into an animal’s tissues.

1 For the literature on this subject see Straus, ‘ Ann. de l’Institut Pas-
teur,’ 1888, p. 181.

2 Tschistowitsch, loc. cit.

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ON A NEW SPECIES OF PHYMOSOMA. ttt

On a New Species of Phymosoma, with a Sy-
nopsis of the Genus and some Account of
its Geographical Distribution.

By
Arthur E, Shipley, ME.Ae, F.L.S.,

Fellow and Lecturer of Christ’s College, Cambridge, and Demonstrator of
Comparative Anatomy in the University.

With Plate XI.

Durine a visit to the Bahama Islands in 1887, Professor
Weldon made an excursion to the neighbouring island of
Bimini. Whilst investigating the fauna of the lagoon in that
island he came across a few specimens of what appeared to him
to be a new species of Phymosoma. One or two of these he
dissected on the spot; the remaining three he brought to
England, and was good enough to give them to me for descrip-
tion at the time when he handed over to me his specimens of
Phymosoma varians which form the subject of a previous
memoir.

I have divided this paper into the following sections:

(i) A general description of the new species.

(ii) A detailed description of those organs which differ
markedly from the similar organs in Ph. varians.

(ii1) A synopsis of the genus.

(iv) A short account of the geographical distribution of the
genus.

VOL. XXXII, PART IJ].—NEW SER. H

Eg ARTHUR E. SHIPLEY.

Panr 4:
Genus Phymosoma.

Phymosoma Weldonii, n. sp.

The length of the body is 3°5 cm. in the largest specimen,
3°25 cm. in the second, and 3 cm. inthe smallest. The greatest
width is 1] em. At the base of the introvert, which was in each
specimen retracted, the width is 2mm. The body has a plump
appearance and is slightly curved (fig. 1).

The ground colour of the preserved specimens is light buff,
which is modified by the presence of dark brown papille ;
these are so numerous as to give the animal as a whole a dark
brown colour. The papille are of two kinds (figs. 2 and 3).

No hooks or traces of hooks can be detected on the introvert.

The mouth is surrounded by a vascular lower lip, which in
the dorsal middle line is continuous with the outer limbs of the
lophophore. The latter structure is in the shape of a double
horseshoe, the outer semicircle of tentacles corresponding to
the lophophore of Ph. varians. In the ventral middle line
this outer limb is bent dorsalwards, and thus the second horse-
shoe is produced (fig.5). The lophophore bears a great number
of long tentacles, seventy or eighty (figs. 5 and 8).

Behind the head is a well-developed collar, pigmented on the
anterior surface (fig. 5).

The alimentary canal is tightly coiled, the number of twists
being twelve or thirteen (fig. 4). It is supported by a well-
developed spindle-muscle, which passes up the axis of the coil,
and is attached at one end to the longitudinal muscles of the
body-wall in the neighbourhood of the anus, and at the other
to the posterior end of the body-wall. The anus is situated
at the line of junction of the two kinds of papille.

The longitudinal muscles are arranged in ten or twelve
bundles in the anterior half of the body ; in the middle of the
body there are about twice that number, as each bundle splits
into two. These fuse together again at the posterior end of
the body. At the base of the introvert the usual inversion of

ON A NEW SPECIES OF PHYMOSOMA. 1138

layers occurs, the longitudinal muscles fusing into a continuous
sheath, the circular muscles becoming broken up into bundles.

The retractor muscles are two in number, a right and a left ;
they arise about the level of the junction of the anterior two
thirds with the posterior third of the body. They embrace the
cesophagus, forming a semicircular band of muscle-fibres which
are wanting only in the dorsal middle line where the heart lies.

The heart is provided with very numerous cecal diverticula
(figs. 4 and 7).

The external aperture of the kidneys lies on a level a little
behind that of the anus.

Habitat: the lagoon, Bimini Island, the Bahamas.

Part IT.
The Papille.

The papille of the skin are of two kinds, those on the body
and those on the introvert. In the middle of the trunk the
papille have an oblong outline, and are arranged in very
regular rings (figs. 1 and 3) ; near the posterior end of the body,
and also at the base of the introvert, the papille are so crowded
together as to lose their rectangular outline. These papillee
are of a dark brown colour, and in those regions where they are
crowded together the buff colour of the rest of the skin is
completely obliterated. The trunk papille form only low
elevations above the general level of the skin; each has a
central pore, surrounded by a number of brown horny plates,
which are modifications of the cuticle. These plates show a
faintly laminated structure; they are represented in section in
figs. 11 and 12.

Between these brown plates are placed a number of deeply
pigmented granules of a dark brown, almost black colour.
These give the dark brown colour to the papille (figs. 9 and 12).
Neither the plates nor the pigment granules show any trace of
being connected with any special cells; they seem to be modifi-
cations of thecuticle. The papille, like those of Ph. varians,
are formed by the ectoderm-cells rising up and invaginating to
form a double cup. The outer wall of the cup is formed by

114 ARTHUR E. SHIPLEY.

ectodermal cells which have kept their ordinary character, and
the inner wall is composed of a few cells of enormous size,
which all but obliterate the cavity of the cup. These cells are
wedge-shaped, and their broader ends are crowded with small
spherical concretions, which do not dissolve in alcohol,
chloroform, or benzine. Towards their outer, narrower ends
these cells become free from these concretions and stain
uniformly, or else they contain from two to five or six large
star-shaped aggregations of crystals (fig. 9) Nothing like
these were seen in the papille of Ph. varians.

The papille on the introvert stand out much farther from
the level of the skin than those of the trunk (fig. 2). They are
conical in shape, with a narrow basis. At their apex is a pore,
and microscopic sections show that this is surrounded by a
number of minute horny plates similar to those in the papille
of the trunk, though much smaller, and not visible like the
latter on the surface view. No crystals were found in
the papille of the introvert, but the large wedge-shaped cells
were in other respects similar to those described in the trunk
papillee.

The Head.

The head is represented diagrammatically in fig. 5. Part
of the outer limb of the double horseshoe-shaped lophophore,
involving eight or ten tentacles, has been cut out in order to
show the pigmented region in the hollow of the original lopho-
phore, and to display more clearly the arrangement of the
tentacles. The thin transparent collar is extended over the
head, a condition in which it is usually found when the intro-
vert is retracted. The crescentiform mouth is shown sur-
rounded by the lower lip, and at the base of the pigmented re-
gion on the dorsal side lies the brain, directly continuous at
each side with the epidermis.

The tentacles are very numerous, seventy to ninety. Like
those of Phymosoma varians, they are roughly triangular
in section. One side, that directed towards the mouth, is
grooved, and the groove is lined with cilia; the grooves of the
various tentacles tend to fuse together near their lower ends,

ON A NEW SPECIES OF PHYMOSOMA. 115

and are directly continuous with the ciliated grooves on the
wall of the cesophagus (fig. 8). A nerve runs along the base of
the groove, and on each side of the nerve is a blood-vessel ; the
third blood-vessel occupies the angle opposite the side bearing
the groove: all these three vessels anastomose occasionally, and
communicate below with a large blood-sinus at their base.

In the diagram the plane of the tentacular crown is too flat ;
instead of being at right angles to the long axis of the body it
should be raised up, and in a manner overhanging the mouth :
in other respects the figure represents the disposition of the
parts, although rather diagrammatically.

The sides of the tentacles which are directed away from the
mouth are deeply pigmented, and the pigment is continued
into a hollow at their base (figs. 5 and 8). This hollow lies
partly between the two horseshoes of the tentacles, and is
therefore itself horseshoe-shaped ; at its dorsal end the depres-
sion becomes deeper, and lodges the brain.

The most important difference between the tentacles of Ph.
Weldonii and Ph. varians lies in the absence of the rows of
those skeletal cells which formed so interesting a feature of
the latter species. Their place is occupied by a well-developed
fibrous connective tissue, which passes down into the base of the
lophophore, and is then continuous with the connective tissue
which surrounds the cesophagus, and which serves as a point
of attachment to the retractor muscles. The lower lip is also
devoid of any skeletal structures ; it is, however, very vascular ;
its inner surface is ciliated, the cilia being continued down the
cesophagus ; its outer surface is pigmented.

The area between the lower lip and the collar, as well as
the inner surface of the latter, is also pigmented, although it
has not been possible to represent this in the diagram. This
continuous lining of pigment ceases at the edge of the collar ;
its outer surface is not pigmented. The collar is represented
in fig. 5 completely expanded, and covering in the head ; it is
usually found in this condition when the introvert is retracted.
It would be interesting to know whether it is ever expanded
in this way when the head is extended during life. I have

116 ARTHUR E. SHIPLEY.

never seen it in this condition in those specimens of the un-
armed Gephyrea which I have been able to examine alive.
The absence of hooks on the introvert is a marked feature of
this species of Phymosoma. Only five other species of the
genus are devoid of these characteristic chitinous structures.

The Vascular System.

The vascular system of the unarmed Gephyrea consists of a
closed space which has no capillaries in connection with it.
The system is distributed in the various parts of the head, and
its chief function would seem to be that of distending the ten-
tacles and lower lip. In the tentacular blood-vessels the blood
is separated from the surrounding water by a thin layer of
tissue, and it is very probable that it becomes aérated during
its passage through these organs. Its function as a carrier of
oxygen cannot be of very great importance, since the system
is entirely confined to one small part of the body. Probably
those organs outside the head are dependent for their aération
on the corpuscles in the perivisceral fluid, though it is not easy
to see where these can get their supply of oxygen and elimi-
nate their waste matter.

The large vessel which lies on the dorsal surface of the
cesophagus, and which is usually known as the heart, acts as a
reservoir into which the blood retires when the tentacles are
retracted (figs. 4. and 7). In Ph. varians, where there were
few tentacles, the heart was a straight blind sac about ‘5 cm.
long, extending along the dorsal side of the cesophagus ; but in
Ph. Weldonii the number of tentacles is much greater, and
the reservoir is correspondingly increased. The heart is much
longer, being continued along the dorsal side of the esophagus
for a centimetre or two, and thus becoming involved in the
twisting of the alimentary canal (fig. 4). Its capacity is also
much increased by numerous small diverticula, which project
as finger-like processes, and give the heart a very characteristic
appearance. The walls of the heart and its diverticula are
thin, with few muscle-fibres in its substance, Similar diver-
ticula occur in the species Ph. antillarum, Ph. pelma, and

ON A NEW SPECIES OF PHYMOSOMA. ila Wy

Ph. asser; and Ph. nigrescens has smaller diverticula of the
same kind.

The corpuscles contained in this closed system are of two
kinds—large clear cells with a well-developed outline, a well-
stained nucleus which lies at one side of the cell, and apparently
no cell-contents ; the other corpuscles are smaller, with a proto-
plasmic body which stains well, and a nucleus in the centre.

In addition to the fluid of this closed system of spaces, the
general fluid of the body-cavity contains corpuscles and the ova
or sperm morule.

The Brain.

The nervous system has the same arrangement of nerve-
fibres and ganglion-cells as that which I described in Ph.
varians. The brain occupies the same relative position,
situated at thedorsal side of the lophophore at the base of aslight
depression. This depression is much smaller than the similar one
in Ph. varians; its area being encroached upon by the bend-
ing back of the tentacular crown to form the inner horseshoe,
the space is thus rendered rather slit-lke and slightly curved
(figs. 5and 8). The depression is lined by a curiously crumpled
and deeply pigmented epidermis, with which the brain is in
direct continuity in two places. The shape of the brain is
different from that of the other species, and this difference
corresponds with the alteration in shape of the pigmented area
in which it lies. It is bilobed, but the grooves between the
lobes are very slight. Each lobe of the brain is smaller in
transverse section than those of Ph. varians; on the other
hand, their long axis is much longer, so that each lobe is slimmer
and more elongated. The narrow outer end of the lobe bifur-
cates into two stout nerves, one of which passes round on each
side in the connective tissue surrounding the walls of the
cesophagus, and fuses with the similar one of the other side to
form the ventral nerve-cord. The other passes up into the
lophophore in the middle dorsal line, and then turns outwards
and runs along the base of the tentacles, giving off a branch to
each. There is a second lophophoral pair of nerves of small

118 ARTHUR E. SHIPLEY.

size, which run into the outer end of the crown of tentacles
where it fuses with the dorsal ends of the lower lip. I could
not make out very satisfactorily whether these nerves supply
branches to the tentacles of this region, but I am inclined to
think that they do.

A pair of very minute nerves leave the brain close to the
middle line ; these run to the pigmented tissue of the depres-
sion at the bottom of which the brain lies. At about the point
of the greatest circumference of each lobe the ganglion-cells
of the brain are in continuity with this pigmented epithelium,
and at one spot this epithelium is involuted into the substance
of the brain, its cells become enlarged and crowded with pig-
ment granules of dense black. The lumen of the involution is
practically occluded; these pigmented involutions form the
eyes.

The ganglion-cells form a cap which wraps round the fibres
except for about a quarter of the circumference, where they
come to the surface; this fibrous portion is ventral and posterior
in position. The whole brain is half surrounded by the large
blood-sinus into which the dorsal vessel opens anteriorly, and
which gives off the vessels to the tentacles.

The arrangement of the fibres and ganglion-cells in the
ventral cord is the same as that of Ph. varians (fig. 10).

The remaining organs of Ph. Weldonii resemble those of
Ph. varians so closely as to render any detailed account
superfluous. The nephridia are two in number (figs. 4 and 7).
The relationship of their external and ciliated internal open-
ings is shown in fig. 11. The outer wall of the nephridium is,
as this figure shows, continuous with the body-wall, but this is
for a short distance only. The organ soon becomes free, and
stretches back to the end of the body, and then in its most
extended condition may be bent back again.

In its histological details the structure of the nephridium is
similar to that which I described in my former paper. The
inner surface is broken up into a series of crypts which are
lined by large glandular cells. Outside these is a meshwork
of muscle-fibres which I have endeavoured to depict in fig. 6,

ON A NEW SPECIES OF PHYMOSOMA. 119

and covering these again is the layer of flat peritoneal epi-
thelium.

The very curious mode by which the glandular cells of the
nephridium in Ph. varians excrete their waste products, by
casting off vesicles into the lumen of the organ, is repeated in
this species. These vesicles contain granules. Their method
of formation and of breaking off from the free end of the
secreting cell is paralleled by the secreting cells which line the
mammary glands of Mammalia, or by the cells which give
rise to the secretion of the liver in Astacus.

The generative organs consist of a band of modified peri-
toneal epithelium which lies at the base of the retractor
muscles. The generative products split off into the peritoneal
cavity.

Part III.
A Synopsis of the Genus Phymosoma.

In the admirable systematic monograph! on the Sipunculide,
in which Prof. Selenka, with the assistance of Dr. de Man and
Dr. Biilow, published the results of their work on the Gephyrea
collected by Prof. Semper in the Philippine Archipelago, a
synoptical table of the genus Phymosoma, which included at
that time eighteen species, is published. Since 1883, the year
of the publication of the above-mentioned work, nine new
species have been added to the genns. Eight of these are due
to the energy of Dr. Sluiter,? who has done so much to increase
our knowledge of the marine fauna of the Malay Archipelago.
The ninth was found by Prof. Weldon in the Bahamas. It will
thus be seen that since the publication of Selenka’s synopsis
the number of described species of Phy mos oma has increased
by half the original. I have, therefore, prepared the fol-
lowing table, which is largely founded on Selenka’s, but
which includes those new species of the genus which have been
described since the publication of his monograph.

1 In a note in my previous paper on Ph. varians this work was inad-
vertently attributed to Prof. Spengel.

2 « Beitrage zu der Kenntniss der Gephyreén aus dem Malayischen Archipel,’
von Dr. C, Ph. Sluiter.

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Nae Se ee ee

‘OL | sourddymyg *yAoAOxZUT Jo F/VyY Ao1taysod Jo oovsans jestop uo sourds osivy JO Mor WV

JLOAOTJUT TO sourds yous ON

oe ee Eee

soqny UMOIG TO UINTNII4AEATP ON

‘sossooord Arepuodses ynoyyIM SOO

SLOJOVAJOI MOT “TIT

quasoad syooyyT

122 ARTHUR KE. SHIPLEY.

Part LV.
Geographical Distribution.

The genus Phymosoma contains considerably more species
than any other genus of the unarmed Gephyreans with the
exception of Phascolosoma. Including the new species
described by Sluiter, and by Selenka in his report upon the
Gephyreacollected by the “Challenger,” the former genus com-
prises twenty-seven species, the latter twenty-five. Next to
these comes Aspidosiphon with seventeen, and Sipunculus
with sixteen.

Of the twenty-seven speciesof Phymosomawhich havebeen
described, seventeen are found in the Malay Archipelago ; of
these seventeen, thirteen have been found there alone, whilst
four have a wider distribution. Three species are found in the
West Indies, of which two are found nowhere else ; five species
in the Red Sea, of which two are peculiar ; and four species in
the Mauritius, all of which occur elsewhere.

It will thus be seen that the Malay Archipelago is the
headquarters of the genus, nearly two thirds of the number of
species composing the genus being found there, and nearly
one half of the whole number being confined to that region.
This is very possibly partly due to the fact that this region of the
world is much visited by collectors, and its shore fauna is pro-
bably better known than that of any other considerable area
within the tropics. On the other hand, the great predominance
of the species in these seas is undoubtedly striking.

The following four species have a somewhat remarkable
distribution :

(i) Ph. japonicum.—This extends along the Japanese
coast, and is again met with in the Fiji Islands and off the
coast of Australia. It was one of the two species brought
home by the “Challenger,” and was found by that expedition at
Port Jackson. It thus has a considerable north and south dis-
tribution. On the other side of the Pacific we find another
species,—

ON A NEW SPECIES OF PHYMOSOMA. 123

(11) Ph. Agassizii, which, while it occurs as far north as
the former species, reaches very much farther south. This
species stretches from Vancouver’s Island down the west coast
of America as far as Puntarenas in the Straits of Magellan,
and has been found at the intermediate points of San Francisco
and Panama. The third species, with a somewhat unusual
distribution, is—

(iii) Ph. Lovénii, which is found only in the Bergen
Fiord. This is still further removed from the equator than the
southernmost point reached by Ph. Agassizii, but it must be
remembered that the Gulf Stream keeps the water on the
west coast of Norway comparatively warm. Finally, we find
one species,

(iv) Ph. granulatum, inhabiting the Mediterranean, and
stretching out into the Atlantic as far as the Azores.

If we except the four species whose geographical distribu-
tion is described above, the whole genus is confined, with the
exception of Ph. antillarum, which extends to Puntarenas,
between the tropics, or only ventures just beyond them.

The species just mentioned has a somewhat curious distri-
bution ; it occurs all round the West Indian Islands, as well as
at Surinam and Puerto Cabello; it then crosses over the
Isthmus of Panama, and is found along the coast of Chili and
Puntarenas. Another West Indian species, Ph. pectinatum,
is found on the west coast of America, and turns up again at
Mauritius, but has not been described from anywhere else.
Finally, Ph. pacificum has a wide range, stretching from the
Red Sea by the Mauritius and India to the Malay Archipelago,
and thence to the Philippines and the Fiji Islands; and Ph.
scolops has a very similar range, occurring in the Red Sea, at
Singapore, at the Philippines, and also off the Mozambique
coast.

With regard to the bathymetrical distribution of the members
of this genus there is little to say ; they all live in shallow water,
and the greatest depth which I have seen mentioned in con-
nection with them is fifty fathoms.

It is not possible to arrive at any very satisfactory results

124 ARTHUR HE. SHIPLEY.

from the scanty material at our disposal, with reference to the
geographical distribution of this Gephyrean. Nevertheless, so
little has been done with regard to the distribution of the
lower marine invertebrates, that it seemed to me to be worth
while to put together what is known about the occurrence in
space of the genus I have been lately working at. The most
striking deductions from the facts before us are—(i) the import-
ance of the Malay Archipelago as the headquarters of the
genus, but this is possibly more apparent than real; (ii) the
restriction, with few exceptions, of the genus to tropical seas ;
and (iii) their preference for shallow waters. The last two
generalisations are obviously connected with the fact that the
animals only flourish in comparatively warm water.

In conclusion, attention may be drawn to the association of
these animals with coral islands. This may be accidental, and
due to conditions of temperature only ; but, on the other hand,
several species make their homes in tubular holes burrowed
out in the soft coral rock,

Tur MorrHo.tocicaAL LaBoratory ;
CamMBRIDGE, July, 1890.

ON A NEW SPECIES OF PHYMOSOMA. 125

DESCRIPTION OF PLATE XI,

Illustrating Mr. Arthur E. Shipley’s paper ‘‘ On a New Species
of Phymosoma, with a Synopsis of the Genus, and some
Account of its Geographical Distribution.”

Fig. 1.—A view of Phymosoma Weldonii, enlarged 3 diameters. The
introvert with its conical papille is slightly protruded.

Fie. 2.—A conical papilla from the introvert, seen from the side and from
above.

Fic. 3.—Some of the depressed oblong papille from the trunk, enlarged to
show the pore and the cuticular plates.

Fic. 4.—A view of the animal cut open just to the right of the middle
ventral line. The introvert is retracted. The ventral nerve-cord is seen
running up the introvert and back close to the cut edge; the right and
left retractor muscles and the two kidneys lie on each side of the coiled intes-
tine. The kidneys are much elongated, and show irregular swellings. The
heart with its diverticula is seen in places. The longitudinal muscles of the
body-wall are not indicated. Enlarged 23 diameters.

Fic. 5.—A diagrammatic view of the head of Phymosoma Weldonii.
The collar is completely expanded, and surrounds the head. Part of the outer
limb of the lophophore involving about ten tentacles has been removed in order
to show the pigmented area within tie lophophore, and the inner circlet of ten-
tacles. The lower lip surrounds the mouth, and at its dorsal end fuses with
the end of the lophophore. The dorsal side of the tentacles, which are fully
expanded, is pigmented. The lophophore is represented too flat; it should
be oblique and overhanging the mouth.

Fie. 6.—A surface view of a piece of the wall of the kidney, showing the
glandular areas—crypts—separated from one another by muscle-fibres.

Fic. 7.—An enlarged view of the end of the coiled intestine, with the heart
partially dissected out. The spindle muscle running up the axis of the coil is
shown near its termination by the anus. The anterior ends of the kidneys are
seen right and left.

Fic. 8.—A transverse section through the base of the lophophore, showing
the lower lip, the mouth, some isolated tentacles, the fused bases of others,
and their blood-vessels and nerves. The fusion of the dorsal ends of the
lower lip and of the lophophore, and the distribution of the pigmented and
ciliated epithelia are seen.

Fic. 9.—A transverse section through a trunk papilla. This shows some
circular muscle-fibres, the ectodermic epithelium passing into the gigantic

126 ARTHUR E. SHIPLEY.

excretory cells. Some of the latter contain crystals, others Jarge granules.
Only that part of the cuticle which is modified to form the horny plates is
shown. Between the plates and round the pore are pigment granules.

Fic. 10.—A transverse section through the skin of the introvert and the
ventral nerve-cord. The introvert is retracted so that the outer surface is
concave, the inner convex. The section shows the conical papille, the thick
cuticle with pigment granules, the single layer of ectoderm-cells, the continuous
layer of circular and longitudinal muscles, the latter broken only for the
insertion of the mesentery supporting the ventral nerve-cord; and the peri-
toneal epithelium. The nerve-cord shows the dorsal disposition of the nerve-
fibres and the ventral ganglion-cells. Some of the secondary nerves are cut
as they leave the cord and traverse the mesentery.

Fie, 11.—A transverse section through both the external and internal
openings of the nephridium. ‘The structure of the skin is shown, and four of
the circular nerves arising from the ventral nerve-cord are seen. The outer
wall of the nephridium is fused with the integument, but becomes free
posteriorly. ‘The section does not show the whole of either opening, as they
do not lie wholly in one plane.

Fic. 12.—A section through the integument.

Mr. Wilson, of the Cambridge Scientific Instrument Company, has drawn
Figs. 1 and 5, and Figs. 4and 7 are drawn from sketches made by Prof.
Weldon in Bimini.

ON THE BRITISH SPECIES OF ORISIA. 127

On the British Species of Crisia.
By

Sidmey F. Harmer, M.A., B.Sc.,
Fellow and Lecturer of King’s College, Cambridge.

With Plate XII.

Tuts paper will be followed by a further memoir, which will
treat of the development of the ovicells and of the embryos in
Crisia. I have already published a preliminary note? on this
subject, and I hope to be able to complete the preparation of
the more detailed paper without much delay.

It has often been pointed out that the subdivision of the
Cyclostomatous Polyzoa into genera and species is attended
with peculiar difficulties. The character of the zocecia remains
remarkably constant throughout this group, the systematic
study of which is not facilitated by the presence of subsidiary
structures, such as the opercula, avicularia, and vibracula,
which in the Cheilostomata form so valuable a means of
distinguishing the species.

‘The task of finding satisfactory specific characters within the
limits of the genus Crisia is not less difficult than in other
genera of Cyclostomata, as is seen clearly enough by
examining the numerous works which have already been
devoted to this genus. Smitt, for instance, in his critical
analysis of the Scandinavian forms, has asserted that the
delicate C. geniculata is connected by a continuous series of
intermediate forms with the coarse form which he calls C. denti-
culata, and which he regards as the extreme point which has

1 «Proce. Cambridge Philosoph. Soc.,’ vol. vii, part 2, 1890, p. 48.

VOL. XXXII, PART 1].—-NEW SER I

128 SIDNEY F. HARMER.

been reached in the evolution of the genus. In his later works
he consistently refers to the latter “species”? as Crisia
eburnea, forma denticulata, and is of opinion that the
“ species ”? recognised by other authors are, for the most part,
merely partially fixed points in acontinuous series. Most other
writers, on the contrary, regard these forms as so many distinct
species.

In many of the characters used for distinguishing the several
species of Crisia from one another—such as the mode of
branching, the number of zocecia in an internode and their
individual shape, the position of the ovicells, &c.—each species
may vary within wide limits about a certain average. The
most satisfactory specific characters appear to me to be fur-
nished by the ovicells ; and in this respect I am only confirming
the results previously arrived at by Waters! for other Cyclo-
stomata. Indeed, I believe that in many cases the species
cannot be certainly identified unless ovicells are present.
Unfortunately, in the great majority of works referring to
Crisia, the information given with regard to these structures
is of the most unsatisfactory character. Many writers, for
instance, have mentioned the existence of “ pear-shaped”
ovicells in certain species; but this character is of generic
much more than of specific importance, and the same remark
might be made with regard to many of the other characters
which have beeu ascribed to the different species.

The importance of the form of the aperture of the ovicell,
as a specific character, has almost entirely escaped the notice
of previous writers.” Busk® has merely stated that the existence
of a tubular aperture on the ovicell is a generic character of
Crisia.

1 “Qvicells of Cyclostomatous Bryozoa,”’ ‘Linn. Soc. Journ. Zool.,’
vol. xx, p. 275, and in other places.

2 Waters has, however, called attention to the importance of this character
in several works. See ‘Quart. Journ. Geol. Soc.,’ vol. xl (Nov., 1884),
p. 676.

’ «Report on the Polyzoa,” Second Part, ‘ ‘Challenger’? Rep., Zool.,’
vol. xvii, part 50, p. 2.

ON THE BRITISH SPECIES OF CRISIA. 129

A comparison of the ovicells (and especially of their aper-
tures) of various forms of Crisia has led me to the conclusion
that the British fauna includes more species of that genus than
are usually recognised. Although the constant occurrence of
a particular form of ovicell might possibly be explained by the
assumption of a definite correlation between the variations of
the zoccia and of the ovicells (the ovicell being regarded as a
modified zocecium), I do not think that this would give a suffi-
cient explanation of the facts. I find, indeed, that the essential
characters of the ovicells are extremely constant, in spite of
the occurrence of variations of no inconsiderable magnitude in
other parts of the colony.

The following specific diagnoses, which are necessitated by
the results which I have arrived at, have been drawn up
on the model of those given by Hincks in his well-known
‘ History of the British Marine Polyzoa.’ New lists of synonyms
appear to me to be also necessary, in spite of the recent appear-
ance of Miss Jelly’s admirable catalogue,! to which I must
express my great indebtedness. My lists do not profess to be
more than a selection of those works in which particular
species have been described or figured in sufficient detail to
make their identification fairly probable. In many cases I have
been obliged to give up the attempt to identify the species to
which the description refers.

C. denticulata, Lamarck. Plate XII, figs. 1—8.

Zoarium large, erect, of rather straggling habit; the
average height of well-grown colonies about one inch; the
branches well separated from one another, and with very
little tendency to curve inwards. Internodes broad and
flattened, but usually with a slight convexity running longi-
tudinally along their anterior face, frequently with a double
curve of such a character that if the lower part of the
internode is convex towards the right side (e. g.) the upper

1 B.C. Jelly, ‘A Synonymic Catalogue of the Recent Marine Bryozoa,’
London, 1889.

130 SIDNEY F. HARMER.

part is convex towards the left side; in most cases with an
odd number of zowcia, the dominant number of which
appears to be 11. Branches arising fairly high in the inter-
node, usually from the 3rd, 4th, or 5th zocecium of either side ;
nearly always given off in perfectly regular alternation on
opposite sides of the axis. Each internode with an odd number
of zocecia is normally provided with a single branch, while the
even-numbered internodes are, with rare exceptions, branch-
less. Joints of the zoarium and of the rootlets nearly always
jet-black, except in the youngest parts of the colony. Basis
rami situated very low down on the zocecium, and appearing as
if wedged in between the zocecium which bears it and the next
zocecium below it on the same side. Zocecia entirely adnate
with the exception of a short portion, of variable length, which
bears the aperture, and which is bent forwards; a pointed pro-
jection sometimes occurring at the outer and upper angle of
the aperture. Ovicell large, always high in the internode,
usually near the end of a branch, and, like the zoccia, more
thickly covered with pores than in the other British species ;
its aperture inconspicuous, not borne on a prominent tube.
Rootlets usually with black joints, which occur at more fre-
quent intervals than in C. ramosa. (See also measurements
on p. 177.)

C. luxata—
(1) Fremine.—‘ Hist, of Brit. Animals,’ Edinburgh, 1828, p. 540.
(2) Coucu.—‘ Cornish Fauna,’ part iii, Truro, 1844, p. 99, pl. xviii, fig. 3.
C. denticulata.—
(83) H. Mitne-Epwarps.—“ Mém. sur les Crisies,” ‘Aun. Sci. Nat.,’
Qe sér., * Zool.,” tome ix, 1838, pl. vii, fig. 1.
(4) Jounston.—‘ Brit. Zoophytes,’ 2nd ed., London, 1847, p. 284, pl. 1,
figs. 5, 6.
(5) Carus.—‘ Prodromus Faune Mediterranee,’ vol. ii, Stuttgart, 1889,
p. 39.

C. denticulata (pars).—
(6) Busx.—‘Cat. Marine Polyzoa Brit. Museum,’ part 3, 1875, pl. iv,
figs. 1—4.
(7) Hrvcxs.—‘ Hist. Brit. Marine Polyzoa,’ London, 1880, p. 422,
pl. lvi, figs. 7, 7a.

ON THE BRITISH SPECIES OF CRISIA. 131

C.eburnea, Linn. Plate XII, fig. 6.

Zoarium forming dense tufts, usually attached by a single
stem, the base of which does not, in most cases, develop many
rootlets ; the average height of well-grown colonies from } to
2 inch ; the branches characteristically curved inwards. Inter-
nodes usually short, somewhat flattened ; in most cases with
an odd number of zocecia, the dominant numbers being 5 and
7. Branches generally arising from the lowest zocecium in
an internode, sometimes higher up; one branch is normally
developed from each odd-numbered internode, even-numbered
internodes being ordinarily branchless. On the main stem
or the principal branches, the branches come off in regular
alternation on opposite sides: nearer the growing-points, they
are arranged in compound helicoid cymes, of the formula!—

(@ - r,)) +
= (# +17,) +
=(*+n7) +
&e.

Joints yellow, or colourless near the growing-points, some-
times becoming dark brown in the older parts of the colony.
Basis rami short, not wedged in between two zocecia. Zocwcia
almost entirely adnate, the upper portion, which bears the
aperture, free, bent forwards nearly at right angles to the lower
part; frequently a conspicuous pointed process on the outer
side of the aperture. Ovicell large, curved inwards, usually
replacing the second, or, less often, the third zocecium of an
internode; its aperture conspicuous, elongated from side to
side, borne on a very distinct tube, which is wider at its base
than atitssummit. Rootlets usually developed in very small
numbers. (See also measurements on p. 177.)

C. eburnea.—

(4) Jounston.—P. 283, pl. |, figs. 3, 4.
(5) Carus.—P. 38.

1 This method of representing the branching is explained on p. 146. In
the above formula z would usually be 5, less often 7 or higher numbers.

132 SIDNEY F. HARMER.

(8) Surrr.—‘ Om Hafs-Bryozoernas utveckling och fettkroppar,”
‘()fvers. af K. Vet.-Akad. Foérhandl.,’ 1865, No.1, p. 9, pl. i,
figs. 15—18.

C. eburnea (pars).—
(9) Smirr.— Krit. forteckn. ofver Skandinaviens Hafs-Bryozoer,” I,
© (fvers. af K. Vet.-Akad. Forhandl.,’ 1865, No. 2, pl. xvi, figs.
10, 11, 13—19.
(6) Busx.—Pl. ii, figs. 1, 2; pl. v, figs. 1, 2.
(7) Hincxs.—P. 420, fig. 21 (p. 416).

C. eburnea, forma eburnea.—

(10) Smrtr.— Bryozoa marina in regionibus arcticis,” ‘ Ofvers. af K.
Vet.-Akad. Forhandl.,’ 1867, No. 6, pp. 444, 461.

(11) Surrr.— Recensio Syst. Bryozoorum Novaja Semlja,” ibid., 1878,
No. 3, p. 12.

(12) Smirt.—‘ Recensio Bry. e mari arctico,” ibid., 1878, No. 7, p. 23.

(18) Freese.— Beschr. Ostsee Bryozoen,” ‘Arch. f. Naturg.,’ 54,
Jahrg., Bd. i, 1888, p. 31, pl. ii, fig. 18.

C. aculeata, Hassall. PI. XII, fig. 4.

Zoarium of very delicate habit, resembling that of the next
species, from which it may be distinguished by its much
slenderer appearance; the average height of well-grown
colonies from 4 to 2 inch, the branches with very little ten-
dency to curve inwards. Internodes usually short, often
consisting of five or seven zocecia; but much longer internodes,
with more numerous zocecia, may occur, especially at the ends
of the branches. Branches usually arising from the lst or
Qnd zocecium of either side of an internode, but sometimes
(especially in the case of internodes near the ends of the
branches) higher up: an internode (especially a peripheral one)
may bear two or more branches. In nearly all colonies, in
addition to the ordinary branches, some of the internodes bear
long, jointed spines, which are curved inwards over the anterior
side of the branch ; these spines are most often developed from
the lower zoccia of an internode, or at the apices of the
terminal internodes. Joints yellow, or colourless near the
growing-points. Basis rami usually short, not wedged in
between two zocecia. Zocecia with a conspicuous, free, tubular
portion, bearing the aperture; this portion is curved forwards,

ON THE BRITISH SPECIES OF GCRISIA., 138

but not as in the last species; it is usually lost in the zowcia of
the lower parts of the colony. Aperture circular, with no pro-
jection on its outer side. Ovicell small, fairly high in the
internode, prominent near its upper end, and falling away very
suddenly to the aperture, which lies on the surface of the
zocecium next above the ovicell on the same side of the
internode; this zocecium curves round the back of the ovicell,
and always acquiresacharacteristic relation to the aperture of the
latter ; this aperture is inconspicuous, and is never borne on
a distinct tube. Rootlets resembling those of C. ramosa.
(See also measurements on p. 177.)

C. aculeata.—

(14) Hassatt.—* Cat. of Irish Zoophytes,” ‘Ann. and Mag. Nat. Hist.,’
vol. vi, 1841, p. 170, pl. vii, figs. 3, 4.

(15) Supp. to ‘ Cat.,’ ibid., vol. vii, 1841, p. 366.

(4) Jonnston.—P. 285.

(16) Smirv.— Bidr. till kann. om Hafs-Bryozoernas utveckling,”
‘Upsala Univ. Arsskrift,’ 1863, p. 3.

Smitt agrees with van Beneden (20) in stating that the ovicells are
completely closed.

(17) Jot1rt.—“ Cont. a Vhist. Bryozoaires, Cotes de France,” ‘ Arch.

Zool. Exp. et Gén.,’ vol. vi, 1877, p. 286.

C. eburnea, var. aculeata.—
(6) Busx.—P. 4.
(7) Hincxs.—P. 421, pl. lvi, figs. 5, 6.
(18) Jutuien.—“ Liste des Bry. rec. 4 Etretat,” ‘Bull. Soc. Zool.
France,’ t. vi, 1881, p. 14.
(19) Vinz.—“ Rep. on Recent Marine Polyzoa,” ‘Brit. Association Re-
port,’ Aberdeen Meeting, 1885, p. 588.

C. eburnea.—
(8) Mitnz-Epwarps.—Pl. vi, fig. 2.
?(20) P. J. van Bunzepen.— Rech. sur PAnat..... Bryozoaires
. . . . Ostende,” ‘ Nouv. Mém. de |’Acad. de Bruxelles,’ t. xviii,
1845, pl. iii, figs, 12—16.
Van Beneden states that the ovicells are closed on all sides; and
this statement is more likely to have been made of C. aculeata (in
which the aperture of the ovicell is very inconspicuous) than of C,
eburnea. The specimens figured are by no means unlike C. aculeata,
but they have no spines.

134. SIDNEY F. HARMER.

? C. eburnea (pars).—
(9) Smirt.—PI. xvi, figs. 12a, 12d.

On p. 185 of Smitt’s paper it is explained that these figures repre-
sent young ovicells (of C. eburnea), without tubular apertures.
It may, however, be remarked that an ovicell, with the contents shown
in fig. 124, would probably have had a well-developed tubular aper-
ture if it had really belonged to C. eburnea; and, further, that there
is evidence (see above, No. 16) that Smitt has worked at the ovicells
of C. aculeata. Ibid., table of formule, Nos. 2, 3 (see explanation
of the formule), and probably some of the later formule.

? C. denticulata (pars).—
(6) Busx.—PI. iii, figs. 1—6.
Notice the spine in fig. 6. In the other figures, the character of
the branching and of the basal internodes, and the small number of the
pores appear to me to prove that this plate does not refer to C.
denticulata, and that it probably refers to C. aculeata.

C. ramosa,n.sp. Pl. XII, figs. 10, 11.

Zoarium erect, often of rather straggling habit; the
average height of well-grown colonies about } inch; the
branches (in well-grown specimens) arranged in fan-shaped
systems, owing to the large number of branches given off by
the terminal internodes, and with little or no tendency to curve
inwards. Internodes often much flattened, of very variable
length ; often very long, and consisting of numerous zocecia ; in
this case often with a well-marked double curve, as in
C. denticulata (and, to a less extent, in other species
Branches developed in greater numbers than in any of the
other British species ; even in the lower parts of the colony the
internodes commonly bear two branches, while the terminal
internodes, and especially those which bear ovicells, may give
rise to as many as four or five branches, which do not neces-
sarily come off alternately on opposite sides of the stem. The
lowest branch of an internode very commonly comes off from
the second zocecium of one side; if the lowest branch arises
from the first zocecium of the internode, the next branch is
usually given off by the third zocecium of the opposite side.
Joints yellow, or colourless near the growing-points, never
black. Basis rami long, usually reaching the aperture of the

ON THE BRITISH SPECIES OF ORISIA. ta

zocecium next below it on the same side, unless it is borne by
the lowest zocecium of an internode. Zoccia usually with a
long, free, tubular portion bearing the aperture ; this portion is
distinctly curved forwards, but is usually lost in the older parts
of the colony; in other cases this tubular portion is not
developed to more than avery slight extent. Aperture circular,
without any pointed projection on its outer side. Ovicell
very large, and more regularly pear-shaped than in any of the
other species; usually a little higher in the internode than in
C. aculeata, but in some cases it may occupy as low a position
as that of the fourth member of the internode; it is perhaps
most commonly in the position of the 6th—8th member ; its
aperture circular, borne on a long and very conspicuous
funnel-shaped tube, which is considerably wider at its summit
than at its base. Rootlets often developed in considerable
numbers, sometimes attaining a great length (nearly an inch),
and composed, for the most part, of long segments, separated by
yellow or colourless joints. (See also measurements on p. 177.)

(The following list includes references to several forms of
Crisia which, as explained below, I do net believe to be
identical with C. ramosa.)

? C. cribraria.—
(21) Srrmpson.—“ Synopsis of the Marine Invertebrata of Grand Manan
[Bay of Fundy],” ‘Smithsonian Conts. to Knowledge,’ vol. vi,
1854.

This species may be identical with C. ramosa, in which case my
own specific name will have to be given up. The zoccia are described
as being “so crowded as to form often two or three longitudinal rows,
in which they are usually opposite” (p. 18). I do not see how such
a statement could be made of C, ramosa. The figures given (pl. i,
figs. 8a—c), although not unlike that species, are not drawn with
sufficient care to enable a satisfactory conclusion to be arrived at.

2 C. arctica—
(22) M. Sars.—‘ Geol. og Zool. Jagtt. anst. p. en Reise Trondhjems
Stift.,”? Christiania, 1863.

The zoarium of this form is said to reach the height of 30 mm.;
the branches and the zocecia are straight, or nearly straight; the inter-
node possesses, on each side, two to three, often eight to twelve, rarely
twenty to twenty-one zowcia. The species is said to resemble C,

136 SIDNEY F. HARMER.

denticulata and C. cribraria. It differs, according to Sars’
description, from my own specimens in the following respects.

The zocecia are fused with one another along their whole length, so
that the upper part, with the aperture, is not free. The outer and
upper angle of the young zocecia may bear a small knob (never observed
in C. ramosa). The joints are usually uncoloured, but sometimes
brown-grey in the older branches (usually yellowin C. ramosa). The
ovicells are always in the axils of the branches, and they are not
described as having an aperture (which can hardly be overlooked in
C. ramosa).

On the whole, Sars’ description suggests a form like C. denticu-
lata or C. elongata, M.-Edw. It is, perhaps, the form figured by
Smitt (9) in pl. xvi, fig. 20, The “basis rami’’ in this figure is unlike
anything I have ever seen in C. denticulata, although resembling
that of C. ramosa.

C. eburnea (pars).—
(2) Coucu.—P. 99.

Some of the larger specimens mentioned by Couch probably belonged
to this species: the ovicells are ‘somewhat urn-shaped with narrow
tubular necks, which are not placed in the centre.” This description
probably refers to C. ramosa, although the “young specimens” in
which the branches ‘all arch inwards” doubtless belonged to C.
eburnea. The magnified figure (pl. xvili, fig. 2), which is not good,
may be identified as C. aculeata by the presence of a spine; and
the figure next to it (natural size) is probably either that species or
C. ramosa.

P C. eburnea (pars).—
(9) Surrr.—Pl. xvi, fig. 9, and p. 135 (fig. 6). (These figs. may refer
to C. aculeata.)

? C. eburnea, var.— ‘
(6) Busx.—PI. v, figs. 5—10.
? C. denticulata (pars).—

(6) Busx.—PI. ii, figs. 3, 4.

(9) Smrrr.—‘ Table of Formule,’ Nos. 14—17, and probably some of
the earlier numbers (e.g. 12 and 13), which are said to belong
either to C. eburnea or to be transitional from this form to
C. denticulata. It is hardly possible that a form with so many
branches arising from the same internode as in No. 17, for in-
stance, was really C. denticulata.

(23) Smirr.— Floridan Bryozoa,” Part 1, ‘ Kongl. Svenska Vet.-Akad.
Handl.,’ B. x, No. 11, 1872, pl. i, figs. 1—65.

I do not feel certain that the form described in (28) is really iden-
tical with C. ramosa, although it can hardly be regarded as C,

ON THE BRITISH SPECIES OF CRISIA. 137

denticulata. The form of the zocecia is very similar to that found
in C. ramosa; but, on the contrary, the ovicells do not agree with
those of the latter. If the left side of fig.5 represents a young
ovicell (probably somewhat broken), the ovicells are even less like
those of C. ramosa in their early than in their fully developed con-
dition. Is this form possibly identical with the one described by
Stimpson (21) under the name of C. cribraria?
(7) Hincxs.—P. 423.

The statement that the ovicells of C. denticulata have “a
tubular orifice at the top’ was possibly made after an examination of
C. ramosa; especially as, on the same page of Hincks’s work, occurs
C. denticulata, var. a (to which pl. lvi, fig. 9, presumably
belongs) ; and there can be little doubt that this is really C. ramosa.

C. denticulata, var. tenuis.—
(24) VicrLtus.—“ Cat. of the Polyzoa ... Willem Barents,” ‘Nied. Arch.
f, Zool. Supplementb.,’ i, 1881-2.

This form is said to correspond closely with Hincks’s unnamed variety
just referred to. It is, however, impossible to accept tenuis as a
specific name, since the name C. tenuis had been applied by Mac-
Gillivray to an Australian species before the appearance of the paper
by Vigelius (see F. McCoy, ‘‘ Prodromus of the Zool. of Victoria,”

‘Decade’ iv, pl. xxxix, Melbourne, 1879.

? C. fistulosa.-—
(6) Busk (non Heller).—P. 5, pl. vi 4, figs. 1, 2.

Even if this form is identical with the species under consideration
it is better to drop Busk’s name, since the specific name fistulosa
was originally applied by Heller to a form which is clearly not the one
described by Busk (see Waters, No. 25).

Through the kindness of Mr. R. Kirkpatrick I have been enabled to
refer, at the British Museum, to a specimen of the form described by
Busk ; and I have also to thank Mr. Kirkpatrick for having subsequently
given me further information on the same subject. The specimen in
question is labelled “C. fistulosa, Hell., locality unknown. Lesina?”
T am informed by Mr. Kirkpatrick that the label is in Mr. Busk’s hand-
writing, with the possible exception of the last word; and that the
specimen is probably really from the Mediterranean.

The specimen in the British Museum is even more like my own
species than is obvious from Busk’s description, which, in Mr. Kirk-
patrick’s opinion, was probably taken from that specimen. As many as
five branches may come off from the same internode, and some of them
higher than the sixth zocecium, which, according to Busk, is their upper
limit. The ovicells, of which only two could be satisfactorily
examined, agree fairly well with those of the Plymouth form. Their

138 SIDNEY F. HARMER.

diameter is about ‘45 mm. Mr. Kirkpatrick further informs me that
the distance from aperture to aperture is ‘4 mm., and that the total
length of the zocecium is about ‘7 mm. Although these numbers are
distinctly smaller than the average measurements of corresponding
structures in C. ramosa, I am inclined to believe that my own
specimens belong to the same species as the one in the British Museum.

Waters (25), in ‘Ann. and Mag. Nat. Hist.,’ 5 ser., vol. iii, 1879,
p. 269, pl. xxiii, fig. 4 (‘‘ Bryozoa of the Bay of Naples”), identifies
C. fistulosa, Busk, with what he calls C. elongata, var. angustata.
I cannot, however, believe that C. ramosa is identical with the form
described by Waters. Although the number of zocecia in the internode
in C. ramosa may be large, this species could hardly be characterised
as having fourteen to twenty-six zocecia in the internode; nor does
the description, ‘‘ branches arising usually from the fifth to eighth
zocecium of a branch, and at about the same distance a fresh branch
grows on the other side,” correspond with the branching of C. ramosa.
As Mr. Kirkpatrick has pointed out to me, Waters’ statement that
the zocecia are ‘04 mm. apart was no doubt due to an oversight.

For further remarks on C. fistulosa, Busk, see Vine (19), p. 589.

The characters of the ovicell are so constant in my specimens
that, taken in conjunction with other facts, I cannot resist the
conclusion that this form deserves recognition as a species.
Although it is obviously alluded to in some of the works just
quoted, I cannot identify it with certainty with any form which
has hitherto received a specific name ; and I therefore suggest
for it the name C. ramosa, in allusion to the large number
of the branches given off by a single internode.

C. ramosa has been found in large numbers at Plymouth,
where it is certainly the commonest of all the forms of Crisia.

While the identification of fully developed colonies of Crisia
—in those cases at least where ovicells are present—cannot often
be a matter of doubt, it may be extremely difficult to identify
the species to which a small fragment of a colony or a young
zoarium belongs. The greatest difficulty is found, in these
cases, in distinguishing C. eburnea from C. aculeata, or
the latter species from C. ramosa. The characters of the
several species can be best brought out by a careful comparison,
under a series of distinct heads, of their more obvious external
features,

ON THE BRITISH SPECIES OF ORISIA. 139

Habit of Zoarium at Different Seasons ; Regeneration.

A very slight acquaintance with the British forms of Crisia
enables one to distinguish at a glance, in most cases, the
species to which a given specimen belongs. C.denticulata
is characterised by the coarseness of its general habit ; by the
regular dichotomous appearance of the branching, as seen by
the naked eye; and by the fact that the branches diverge from
one another to such an extent that they are separated from one
another by considerable interspaces at their ends. In C.
eburnea the branches are inflected towards the axis of the
colony, and are so closely massed together that it is impossible
to study the exact character of the branching without first dis-
entangling the branches. On flattening the specimen out ona
slide the cymose character of the branching is at once appa-
rent. C. aculeata possesses a characteristic delicacy of habit
(“of a slenderer habit than C. eburnea, which the species
closely resembles ’’') ; and it may be compared, in external
form, to a C. eburnea which has become of much laxer and
slenderer habit than usual, and in which comparatively few
branches have been developed. ‘The branches are much
straighter than in C. eburnea. C. ramosa is extremely
similar, in general appearance, to C. aculeata, but is of dis-
tinctly coarser habit ; the branches are very straight, and the
number of the branches to which the internodes near the grow-
ing-points give rise results, in actively growing colonies, in the
formation of fan-like systems of branches. The long tubular
apertures of the zowcia (if developed) give a characteristic
appearance to the species, which cannot, however, in all cases
be distinguished by the naked eye from C. aculeata.

The above remarks apply especially to colonies in their fully
developed condition; but the appearance of any species de-
pends largely on the time of year at which it was found.
Many of the specimens of C. eburnea found in the early
spring are provided with numerous ovicells, the ultimate fate
of which seems to have hitherto attracted no attention, although

1 Johnston, G., ‘ A Hist. of the British Zoophytes,’ ed. 2, p, 286.

140 SIDNEY F. HARMER.

there can be no doubt that these structures disappear after the
end of the breeding season. I have looked in vain for any signs
of the absorption of the ovicells in Crisia; and the following
facts probably imply that they are simply thrown off from the
colony after the liberation of the embryos which have been pro-
duced in them.

The typical spring form of C. eburnea possesses a con-
spicuous main stem, which forms an obvious central axis, from
which the rest of the colony comes off as a series of branches,
developed in regular alternation on opposite sides, and decreas-
ing in size fairly regularly from the base to the summit of the
colony. The main stem consists of perhaps eleven or twelve
internodes, each of which normally gives rise to a branch;
and the branches themselves are usually provided with a
profusion of ovicells, many of which are still in process of
development, and most of which are near the ends of the
branches.

In a colony of the same species found in May most of the
ovicells were at some distance from the ends of the branches,
owing to the development of several (7—8) zoccia above the
ovicells; and the branches which bore ovicells had, in most
cases, completely finished their growth: very few ovicells, and
these of a weakly appearance, were being developed. Most of
the branches ended in slender internodes, in which growth was
no longer taking place, as was shown by the fact that no grow-
ing-points were left. The exhaustion of the colony was further
shown by the fact that some of the terminal internodes con-
sisted of no more than two or three zoccia, with no growing-
points.

In the summer (August) large and highly branched colonies
with active growing-points are found, but they are nor-
mally without any trace of ovicells. In many of these cases
it may be noticed that the main stem has been broken, and is
merely represented by its basal portion. The rest of the colony
will, in this case, probably consist of a small number of large
branches given off from the remains of the stem or of its
lateral branches, and in many cases the sharp contrast between

ON THE BRITISH SPECIES OF CRISIA. 141

the clean white appearance of these highly branched parts of
the colony and the dirty-brown appearance of the stump of the
main stem, covered as it is by foreign growths of various kinds,
will give rise to the suspicion that the former have been de-
veloped at a later period than the latter, and that the latter
are the remains of colonies which developed ovicells at an
earlier period of the year.

Smitt! has called attention to the fact that the free tubular
portion of the zocecium of C. geniculata is sometimes rela-
tively transparent, and that it is separated by a sharp line from
the basal, more highly calcified part, and he suggests that this
transparent portion has in these cases been regenerated. He
further points out? that in Aetea argillacea (=Aetea
truncata, forma abyssicola elongata?) this process of re-
generation seems to be periodic, since a zocecium consisting of
portions of three different ages was in one case observed by him ;
and that in Farrella fusca (=Vesicularia fusca‘) the
zocecium may attain twice its normal length by the occurrence
of this regenerative process.° Muilne-Hdwards® had previously
pointed out that the zocecia were able to form rootlets at an
advanced period of their existence.

There can be little doubt that Smitt’s suggestion is a correct
one. InC.eburnea the older parts of the colony are fre-
quently covered with an encrusting red seaweed, the presence
of which has no doubt been responsible for the ‘“rose-red”
colour which has been mentioned by Johnston’ and others as a
feature which sometimes characterises the species. In certain
specimens found in April the basal parts of the colonies were
completely covered by this encrusting growth, while in various

1 ¢ Ofvers. af K. Vet.-Akad. Forhandl.,’ 1865, No. 2, p. 128.
2 «Om Hafs-Bryozoernas utveckling och fettkroppar,” ‘ Ofvers.,’ &e., 1865,
. 1, pp. 29, 30.

‘ Ofvers.,’ &c., 1867, No. 5, p. 280.

‘ Ofvers.,’ &c., 1866, pp. 502, 505.

Ibid., pl. xiii, fig. 39, and explanation of figure.

‘Ann. Sci. Nat.,’ 2e sér., “ Zool.,” tom. ix, 1838, p. 196.

‘British Zoophytes,’ ed. 2, p. 284.

Sana npn wo O

142 SIDNEY F. HARMER.

parts perfectly white growing points or new apertures were
_making their appearance. Similar phenomena of regeneration
have been repeatedly observed in all the species which I have
examined. Thus the first glance at an ordinary colony of C.
ramosa will suffice to show that the tubular ends, so charac-
teristic of the young zoecia, are absent in the lower parts of the
colony, where they have been either broken off or absorbed. The
zocecia which are in this condition are closed by an obliquely
placed diaphragm, as described in Crisia and other Cyclo-
stomata by Waters,! Pergens,® and others. On staining a
specimen of C. ramosa without decalcification, it is at once
obvious that these diaphragms are used for the closure of
zocecia which contain brown bodies but no functional polypides.
They are placed at the point where the zoccium normally
becomes free from the internode, aud the free portion becomes
gradually broken away or absorbed down to the point where
the diaphragm is situated. In the younger parts of the colony,
where the zocecia possess free tubular ends, no diaphragms are
present, and functional polypides or obvious buds, together
with the brown bodies formed by the death of the last poly-
pides, are found in nearly all the zocecia.

The individual life of the zocecium has not, however, neces-
sarily come to an end with the formation of one of these
diaphragms, as may be easily proved by the examination of
suitable spring colonies which have been stained with borax
carmine without decalcification. Whilst zocecia in which no
regeneration is taking place are closed by a diaphragm and
appear perfectly unstained, the red colour of the regenerating
parts is obvious at the first glance. The first indication of the
renewed activity of a zoccium is given by the fact that some
of the cells below the diaphragm have acquired the power of

1 A. W. Waters, “Closure of the Cyclostomatous Bryozoa,” ‘Linn. Soe.
Journ. Zool.,’ vol. xvii, 1884, p. 400; ‘ Fossil Cyclostomatous Bryozoa from
Australia,” ‘Quart. Journ. Geol. Soe.,’ vol. xl, 1884, p. 675.

2 Hd. Pergens, “ Revision des Bryozoaires du Crétacé figurés par d’Orbigny,”
le partie, “ Cyclostomata,” ‘ Bull. de la Soc. Belge de Géol., &c., tome iii,
1889, p. 317.

ON THE BRITISH SPHCIES OF CRISIA. 143

taking up colouring matters; slightly later a young polypide
bud is seen below the diaphragm, which is then absorbed, the
zocecium growing out (in C. ramosa) into a long tubular
portion, at the end of which is the aperture! In C. ramosa
the free portions of regenerated zocecia are sometimes con-
siderably longer than the normal length of the tubular portion.
In one case the regenerated portion, which was completely free
from the branch, was ‘69 mm. long.

It is well known that new stems are given off from various
parts of the rootlets.2. These rootlets are usually developed
from the backs or sides of the zocecia, especially of those near
the base of the colony. But in cases where regeneration is
actively taking place the tip of a branch may grow out into a
rootlet, or a rootlet may take the place formerly occupied bya
zocecium, usually one of the terminal zocecia of an internode in
this case. The rootlet thus formed may grow for a considerable
distance, and finally produce a new stem as a lateral branch ;
or the new stem may be the actual prolongation of the rootlet,
which, after a longer or shorter course, assumes the characters
of astem. In other cases a new growing-point is formed
from an old joint at the point where a lateral branch or an axial
internode has previously been lost; or it may be formed from
the apex of an internode in which the fracture has taken place
across the middle of the internode, instead of at an axial joint.
The result of this is that it is very common to observe an old
brown stem from which start new internodes (lateral or axial),
which are shown, by reason of the perfectly white appearance
of their ectocyst, to have been formed at a much later period
than the brown part of the stem. In one or two cases a
growing-point had started from the proximal side of a broken
joint, and had then given rise to a stem which grew in a direc-
tion directly opposite to that of the internode from which it
was developed. These cases are somewhat analogous to the

1 This is the process which was observed by Smitt in C. geniculata.

2 Cf. Smitt, “ Krit. Fort.,” i, ‘Ofvers.,’ &c., 1865, p. 122.

5 These statements refer, for the most part, to C. eburnea and to C.
ramosa.

VOL. XXXII, PART II.—NEW SER. K

144, SIDNEY F. HARMER.

one described by Smitt,! in which a “basis rami” had given
rise to a normal branch, and also to a growing-point directed
straight downwards from its base, which was formed by the
proximal end of the “ basis rami,” from which it was separated
by a joint.

Although regenerated lateral branches may start from the
old lateral joints, it is not uncommon to find that they are
given off from near the end of the old internode, instead of in
their normal position lower down ; this is due to the fact that
the aperture of an old zocecium has become a growing-point.

In colonies in which the process of regeneration is com-
mencing, it is frequently noticed that the young growing-
points are appreciably smaller than the normal ones. These
small growing-points naturally give rise to slender zocecia and
branches, which, however, as they grow longer, acquire fresh
strength, and soon regain their normal diameter. The regene-
rated parts of a colony are, consequently, often joined to the
older parts by slender bases, in which, moreover, the basal
internodes may consist of an unusually small number of zoecia.
In both these respects they resemble colonies which are deve-
loped directly from the larva, or from a growing-point which
starts from the rootlet of an old colony.

It is important to notice that, so far as my observations go,
the regenerated parts of a colony always retain the same specific
characters as the older parts. I have looked in vain for any
indications which might have been given by regenerating
colonies that the forms of Crisia described above as distinct
species might be merely different phases of the same species.

The general life-cycle of C. eburnea may probably be sum-
marised as follows :—The breeding season is at its height in
April and May ; and at about this period it is not difficult to
find young individuals which consist of a single zoccium
attached by a disc-like base, and which have resulted from the
metamorphosis of a free larva; small colonies are soon formed
by these primary zoccia. At first, rootlets may be altogether
absent, and in many colonies they are developed very sparingly ;

1 Loc. cit., p. 125.

ON THE BRITISH SPECIES OF ORISIA. 145

but when formed, some of them give rise to fresh stems, which
are the starting-points of new colonies. Or, again, some of the
specimens found in the summer have resulted from colonies
which developed ovicells in the earlier part of the year, and
which, after losing these structures, again burst out into
renewed growth ; in some cases leaving a single ovicell on the
colony as some indication of their past history. In the spring,
ovicells, when present at all, are found in large numbers, and
those well-developed colonies which do not possess them at
this period are probably in most cases of the male sex. Thus,
in order to find spermatozoa in April, it was generally quite
sufficient to select any colony in which there were no ovicells,
while spermatozoa were not discovered in any of the cases in
which ovicells were present.

In the early spring the discoloured stumps of colonies which
grew during the preceding year are found; from various parts
of these, new growing-points are developed, and give rise to the
colonies found at a slightly later period. The production of the
enormous number of embryos then developed seems to exhaust
the energy of the colony, whose growth practically ceases for a
time, many of the branches being thrown off. After a period
of rest, growth recommences with great vigour, and by the
middle of the summer large and highly branched colonies are
again found, although now, as a rule, with no ovicells.

Number of Zocecia in the Internode, Mode of Branching, &c.

Most of the previous accounts of Crisia merely mention the
limits between which the number of zocecia in the internode
may vary in the several species. Thus Hincks! says of
C. eburnea, “‘ 3—9 cells in an internode ;” while Johnston? is
a little more explicit, stating that ‘there are from two to
five, sometimes seven, and very rarely even nine, cells in each
internodial space ” in the same species. It appears to me that
it is quite impossible to define accurately the several species

1 « British Marine Polyzoa,’ p. 421.
* «British Zoophytes,’ ed. 2, p. 284,

146 SIDNEY F. HARMER.

without paying careful attention both to the number of zoccia
in the individual internodes and to the character of the
branching. Smitt! is the only writer who has done this in a
thoroughly satisfactory manner, and it will be convenient to
make use of a modification of a graphic method of repre-
senting the characters of the colony which he was the first to
introduce.”

In this method the limits of each internode are indicated by
brackets, in which is a number denoting the number of zocecia
in the internode; 7 indicates a branch, the position of which
is further shown by means of a number: thus ,7, e. g., indicates
a branch given off from the first (lowest) zocecium of an
internode on the left side, while 7, indicates a branch given
off from the second zoecium on the right side of the inter-
node; finally 2 indicates the growing-point, and Ov. an ovicell.
A key to the method may be obtained by referring to fig. 6,
the formula of which is included on the formula on p. 154, as
explained on that page.

Thus many of the characteristic features of C.denticulata
(figs. 2, 3) may be represented by the formula—

(8) + 9+47) + (10) 40-7) + 9+ y) + AL +r) +1 +4) + (13473) + (949) +842)
Leas+.)+d0)+tr) +44.)

|

=(5+2)
=(11+5r)+ (8+ 00.+5+7,+2)
=(2)

=(11+44r)+(2)

=(4
=(11+75;)+(10)+(11+47)+(8)+(2)
=(5 4 Ov.42+y+2)

342)
=(10)+(9+7,)+(5+0v.+5+2)

og Ov.+6+7,+2)
=(8+z2)

1 «Ofvers. af-K. Vet.-Akad. Férhandl.,’ 1865, p. 115.
2 Loe. cit., p. 139.

ON THE BRITISH SPECIES OF ORISTA. 147

—representing the axis of a branch given off by the main stem
of acolony, together with all the ramifications of two of its
secondary branches.

It will be noticed that nearly every internode develops a
single branch, and that the branches come off in regular
alternation on opposite sides of consecutive internodes of
every axis. Although the number of zoccia in the internode is
very variable, eleven may be regarded as the number most cha-
racteristic of the species.

The above formula further shows that every branch-bearing
internode whose development is complete possesses an odd
number of zocecia, while in completely developed internodes
which bear no branch the number is even. Although this rule
is not quite absolute, it is difficult to find any exception to the
striking rule that a branchless internode has an even number
of zoccia; or, conversely, that an internode with an even
number (whether this number is large or small) of zocecia
bears no branch. It may be pointed out that even if the
branches have been broken off, their previous presence can be
ascertained by the existence of the basal articulation from
which they formerly sprang.

It may further be noted that a lateral branch is, with very
rare exceptions, produced on the side of the basal zocecium of
the internode (Pl. XII, fig. 2).

The regular alternation of the zoccia of the axis of any
branch is not disturbed by the development of an axial joint;
and the last zocecium of the internode below the joint nearly
always projects beyond the penultimate zocecium (which
belongs to the other side) in the form of a free tube (fig. 2).
Since the branch-bearing internode has an odd number of
zoecia, and since the branch is developed on the side of the
basal zocecium, it follows that the last zocwicum, which is
produced into a free tube, will also be on the same side as
the branch. A moment’s consideration will show that the
basal zocecium, the branch, and the terminal (“ produced ”’) '
zocecium, in any internode, will normally be on the opposite
side to that on which these structures are situated, both in

148 SIDNEY F. HARMER.

the internode below it and in the internode above it on the
stem.

But if an even-numbered internode is developed (fig. 2),
its last zocecium will of course be on the same side of the stem
as the last zoccium of the preceding internode; and conse-
quently the basal zocecium and the branch of the internode
above it will be on the same side as its own basal zocecium,
and on the opposite side to the branch next below it; or, illus-
trating this by a formula, we shall, as a rule, find cases like
(138+7;) +(8) + (7+,7r), as shown in fig. 2.

Thus, stating the same fact in another way, an even-
numbered and branchless internode may be intercalated in the
stem without disturbing the alternate origin of the branches
on opposite sides. The same is true of those cases where two
even-numbered internodes occur consecutively on the same
axis.

The more closely one investigates unusual methods of
branching in this species, the more obvious does it become
that the growth of the colony is regulated by some well-defined
law, which finds one of its expressions in the preceding rule.

Thus it will be seen, by reference to figs. 2, 4,6, and 11,
that the basal zocecium of a lateral branch is on the abaxial
side of the latter in all the four species referred to; and
further, that the branch given off by the basal internode of an
axis is also on the abaxial side. This is obvious enough for
C. denticulata, from the formula on p. 146, where it will
be noticed that, in the one case in which the basal internode
has an even number of zocecia, the second internode develops
the first branch, and that that branch (and of course the basal
zocecium) is on the abaxial side.

On two occasions abnormal branching of the type (9 + 4r) +
(13) + (11 +,7) was noticed. Here an odd-numbered branch-
less internode occurs; but such cases seem to be rare. Since
the number of zocwcia in the branchless internode is odd, it

‘ follows that the basal zocecium, and consequently the branch,
of the third internode will be on the same side as in the first
internode.

ON THE BRITISH SPECIES OF ORISIA. 149

In another case the formula (15 + 7,) + (9) + (10) + (9+7%)
was obtained, and it is obvious that this is a further illustration
of the same principle.

In an axis, part of the formula of which was (13 + r,) +
(14 + gr + 75) + (11 + yr), one of the internodes had two
branches, the first developed on the side of the basal zocecium,
and the second on the opposite side. Since the number of
zoccia in this internode was even, the regular alternation of
the branches was not disturbed. Cases of this kind appear to
be extremely rare.

Another abnormal case, from a young colony, had the
formula—

+(5+72)+
=(7) + (6 +72) + (8) +(6)+(2+2)

Here it is obvious that the whole of the lateral branch shown
is very abnormal; the first branch is developed by the second
internode, which has an even number of zoccia; and it is on
the same side as the basal zocecium of the first internode, and
on the opposite side to that of its own internode. Here it must
be supposed that the tendency to produce the first branch on
the abaxial side has prevailed over the tendency to produce a
branch on the side of the basal zocecium of an internode.

In one case observed, in which the base of an old colony was
regenerating fresh branches, two small growing-points were
seen to have been formed, almost exactly opposite one another,
from the same internode. If this growth had proceeded some-
what further, it might not have been obvious that the abnormal
character of the branching was due to the occurrence of regene-
rative processes, in which the regularity which characterises the
normal branching does not seem to be so marked.

Some of these remarkable relations are obvious enough in
the figures given by previous authors, none of whom seem,
however, to have been struck with the general rule illustrated
by these cases. Thus Milne-Edwards,' in pl. vu, fig. 1 6
(C. denticulata), shows a portion of a colony in which two

1 ¢ Ann. Sci. Nat.,’ 2¢ sér., ‘ Zool.,”’ tome ix.

150 SIDNEY F. HARMER.

internodes have an odd number of zoccia, and in which,
further, the branch and the basal zocecium are, in each inter-
node, on the same side. The third internode which is com-
pletely figured has 10 zoccia, and possesses no branch. It must,
however, be pointed out that in his fig. 1 @ (C. denticulata,
under slight magnification) Milne-Edwards represents most of
the internodes as having an even number of zoccia; but it
may probably be assumed that in this figure, which gives an
excellent representation of the general appearance of the species,
sufficient attention has not been paid to the details of the
arrangement of the zoecia. Again, Busk! figures, in the same
species, two complete internodes, one of which has thirteen zocecia
and a branch, and the next has twelve zocwcia and no branch.

The relations above described are perhaps capable of being,
to some extent, explained in the following manner. In the
species of Crisia which I have examined, and, I have very
little doubt, throughout the genus, the base of an internode,
whether axial or lateral, is simply the basal part of the lowest
zocecium of that internode, that part having been separated by
the development of the joint from its upper or distal part.
This will be intelligible on referring to Pl. XII, fig. 1, repre-
senting an axial internode in which only two zoccia are
completely separated from the growing-point. The lowest
zocecium of the internode is seen to be divided into two parts
by the horny joint ; and the lower of these two parts forms the
articulation to which the younger internode is attached. In
examining the formation of the joint (whether axial or lateral)
in stained specimens it is at once obvious that the alimentary
canal of the youngest zocecium of the internode at first extends,
through the tubular joint, into this lower portion ; confirming
the statement made above with regard to the morphology of
the base of the internode.

It is thus clear that the occurrence of an axial joint in
no ways disturbs the alternation of the zowcia (see any of
the figures). The last zocecium of the older internode would

1 «Cat. of Mar. Pol. in Brit. Museum,’ Part ILI, “ Cyclostomata,” pl. iv,
fig. 2.

ON THE BRITISH SPECIES OF CRISIA. 151

overlap, and be fused with the next zocecium higher up on the
same side if it were not for the development of the joint ; which
is, however, formed in such a position across a zocecium as to
leave the preceding zocecium in the characteristic “ produced ”
condition which has already been described. In the ordinary
type of branching, where successive internodes produce branches
in regular alternation on opposite sides, the number of zocecia
must be odd if the branch is to be produced on the side of the
basal zocecium in each internode. The formation of a new
axial internode practically amounts to the transverse division
of a zoccium, while the formation of a branch may be ex-
pressed as due to the longitudinal division of a zocecium (at
the growing-point). Suppose that the right side of an axis
bears a branch (as in the lowest internode shown in fig. 2).
The tendency of the growing-point to produce new branches
alternately on opposite sides would normally result in the pro-
duction of a branch from the left side of the next youngest
internode; but if a lateral branch has not been produced
by the time that a new axial joint is to be formed, that axial
joint would be, as a matter of fact, normally developed from a
zocecium of the left side, as at the base of the third internode
in fig. 2; and this implies the existence of an even number
of zocecia in the second internode. The production of an even-
numbered internode may thus be regarded as due to the alter-
nate predominance of the two sides of the growing-point. The
development of a lateral branch on the right side (e.g.) has
apparently the effect of leaving the left side of the growing-point
with an excess of vigour; so that when a new internode is
formed—whether by the transverse division of a zocecium to
form an axial joint, or by its longitudinal division to form a
lateral branch—it is the left side (in this particular case) of the
growing-point by which this division is effected. Division in
the transverse direction results in the formation of an even-
numbered internode, while the production of a lateral branch
on the left side of the next succeeding internode restores the
function of producing another axial joint to the right side of
the growing-point.

152 SIDNEY F. HARMER.

That there are exceptions to this rule has been shown above
by the description of odd-numbered branchless internodes ; but
it must be remembered that these cases are rare.

From what has already been said of the laws which regulate
the growth of Crisia, it is obvious that a representation of a
colony can easily be reconstructed from a formula of the cha-
racter introduced by Smitt; and no further justification is
required for the use of such formule.

In some specimens of C. denticulata the average number
of zocecia in an internode may be higher than in the one de-
scribed; and the numbers 13, 15, 17, and even 19 are by no
meansuncommon. It may often be noticed that, although inter-
nodes consisting of any given number of zoccia do not seem
to be arranged in any definite order in the colony, an indivi-
dual colony may be characterised by the frequent occurrence of
internodes with that number of zoccia. Thus if the dominant
number, in any particular case, be 11—and this seems to me
the most common case—variations in the number of zoccia in
the internodes of that colony will apparently take place about
the number 11 as a mean; so that, although internodes of 9 or
13 zocecia are common, there may be none of so many as 15
zoeecia. But if the colony have many internodes of 15 zocecia,
for instance, then it will probably be found that some of the
other internodes have 17 or 19.

If the growth of the branch be complete, so that no more
axial joints are to be formed, the terminal internodes, and
especially those which have produced ovicells, may have a
larger number of zocecia than the internodes of the rest of
the colony; and the number of zocecia formed before the
growing-point exhausts its activity does not appear to be
regulated by the laws which govern those internodes which
are not terminal. But even the terminal internodes normally
produce no more than a single branch (ef. C. aculeata and
C. ramosa), the cases mentioned on p. 149 being the only
ones in which two branches were noted to come off from the
same internode.

The articulations of the lateral branches of this species are

ON THE BRITISH SPECIES OF CRISIA. 153

alone sufficient to distinguish C. denticulata from the other
British forms. They are situated at a very low level on the
zocecia which bear them, and each “basis rami” (Smitt)
appears to he wedged in between two consecutive zoecia (fig.
3), instead of being, as in other species, distinctly apposed to
the outer side of one zoccium (figs. 4, 6, 11). The branches
usually originate from z;,! z,, or z;; less commonly from 2,
or from 2.

The joints, both of the zoarium and of the rootlets, of this
species are in nearly all cases of a jet-black colour, as recog-
nised by most of the previous writers.” The young joints are,
as in other species, uncoloured; but the black colour is in
almost all cases very speedily acquired. Smitt and Busk do
not mention this as a specific character, no doubt because they
have given wider limits to the species than are accepted by
most writers.

The ovicell* in all species replaces an ordinary zoccium,
and in this particular species it is usually borne on a lateral
branch, and in most cases is situated at some distance above a
joint. In the instances given in the formula on p. 146 the ovi-
cell replaces the 4th, the 6th, or the 10th zocecium of an in-
ternode. I have never seen it lower than 4th nor higher
than 13th. Itis usually very near the end of a branch, and
this feature is well shown in pl. iv, figs. 2 and 4, of Busk’s
British Museum Catalogue (Part III). In one of my cases,
however, thirteen zocecia occurred above the ovicell, and eleven
below it, and very rarely a joint may be developed above it.
If the ovicell- bearing internode develops a branch, that branch
is very seldom given off from a position higher than the
zocecium which corresponds to the ovicell on the opposite side
of the branch.

1 T.e. from the third zoccium of either right or left side: the side from
which a branch comes off has no significance unless considered in relation to
other characters.

2 Cf. Fleming, J., ‘Hist. Brit. An.,’ p. 540; Johnston, A., ‘ British
Zoophytes,’ 2nd ed., p. 284; Hincks, T., ‘ Brit. Mar. Polyzoa,’ p. 423; &e.

3 See also p. 169.

154 SIDNEY F. HARMER.

The calcareous matter of the ectocyst is considerably thicker
in C. denticulata than in any of the other British species.

C.eburnea. Fig. 6.

Although this species is not likely to be mistaken for C.
denticulata, I believe that it is more nearly allied to that
species than are any of the other British forms. This is shown
by the flatness of the internodes, by the fact that each inter-
node has normally one branch, and by the characters of the
apertures of the zoccia.

The branching may be illustrated by the formula!—

(5+) + (5+) +(7 + ar) + (7472) + (7 +9) + (7 +12) + (7 + 7) + (2)
Lett ta HOEN)E(TE)-+66 +7ry)+ (7 +17) +(2)

=(54yr)+ |
| =(5+7,)+
=(5+,7)+ |
| =(5+n)+ (5+ yr) + (5+) +(54yr+2)
=(5-+4r)+
——(0 +r,)+
=(5+,r)+ |

'=(1+ Ov.+10+7,+7,+2)
=(1+ 0v.+9+,7+2) | |
| | ‘'=(4-+2)
=(¢) =(8+7,4+2)
=(3+2)
— representing the partial formula of a branch of a colony found
in April, in which ovicells were very numerous. The ovicell-
bearing internode on the right side of the formula is the one
which has been represented in Pl. XII, fig. 6.

The tendency of the branches of this species to arrange
themselves as unilateral sympodes is here most marked ; and
the formation of these helicoid cymes—again borrowing a
botanical term—is one of the most characteristic features of
C.eburnea. This was recognised by Johnston,? who says of
this species, ‘‘ Polypidom much branched, the primary divisions
aiternate, spreading ; the secondary from one side only.” It
will further be noticed that in parts of a colony in which this

1 See explanation of this graphic method given on p. 146.
2 «Brit. Zoophytes,’ ed. 2, p. 284.

ON THE BRITISH SPECIES OF ORISIA. 155

method of branching is well developed, the internodes compos-
ing the sympode are usually made up of five zocecia, and that,
although the branching may, in other parts of the colony, take
place from z, (or rarely from z, or z,), well-developed helicoid
cymes are invariably composed of internodes in which the
branching takes place from 2.

These helicoid cymes do not, however, agree with the method
of branching defined under that term in text-books of botany,
in that the main axes of the parts of the sympode are by no
means suppressed. ‘This is obvious enough from the formula,
in which the first internode on the left side forms the basal
member of a helicoid cyme developed on the left side of the
branch; but it is, at the same time, the basal member of a
long axis, which develops new cymes alternately on opposite
sides; and the same is true of the other constituents of the
sympodes. ‘Thus each of the branches indicated in the formula,
with the exception of those which are quite near to the grow-
ing-points, is again the basal member of a helicoid cyme ; and
these cymes are consequently given off alternately on opposite
sides, not only by the internodes of the main stem, but
also by the internodes of its branches of the second, third,
and other orders. The number of members of which these
helicoid cymes are composed decreases fairly regularly in a
centrifugal direction.

Each internode is typically provided with one branch, and
at the same time is composed of an odd number of zoecia,
just asin C. denticulata. It is not uncommon, however, to
find branchless internodes, whose position in the colony may
be illustrated by the formula—

(boss va) se TO Se rea silk

and, just as in C. denticulata, these branchless internodes
nearly always consist of an even number of zoccia, most
commonly of four or six, less often of two or eight. Ex-
ceptions to this rule are somewhat less rare than in C. den-
ticulata, which this species so closely resembles in its method
of branching. The exceptions are more common at the base

156 SIDNEY F. HARMER.

of the colony than elsewhere. The branch is developed on the
same side as the basal zocecium of an internode, and the last
zocecium is usually somewhat produced. In very rare cases,
of which the specimen represented in fig. 6 is an example, two
branches may be developed from the same internode.

The articulations which bear the lateral branches are rela-
tively short ; even when the branch is developed from z, or z;
the “ basis rami” is never wedged in between two zoccia, as
in the last species; and the joint which bears the branch is
nearer to the aperture of the zocecium which has developed
it than in C. denticulata.

The number of zoecia is typically five or seven, the former
number being especially characteristic of the members of a
helicoid cyme. As in C. denticulata, the definiteness with
which the colony grows is frequently indicated by the regular
repetition of the same forms of internode ina branch. Thus
the greater part of the main axis of the branch whose formula
is given on p. 154 is composed of internodes of the type (7 +75) ;
in the main axis of the branch given off by the second internode
of that stem, (5+ 7) alternates regularly with (7 +,7) until
the end of the axis is nearly reached; while in the next line
but one will be seen the formula of a branch composed of units
of the type (5+7,). The regular repetition of internodes of the
type (5 +7) in the formation of most of the helicoid cymes is
a further illustration of the same thing. In many other cases,
however, no such regularity of arrangement was noticed. In
a colony found at Plymouth in August, the dominant number
of zocecia was seven, although internodes with five zocecia were
not uncommon. But, in correlation with this increase in the
normal number of zocecia, it was found that several ititernodes
of nine zocecia occurred, and two of eleven.

In the terminal internodes the number of zocecia may be
larger; in one case observed it was as high as twenty, no
growing-point being left.

The ovicell most commonly replaces the second zocecium of
a lateral branch (fig. 6), and is consequently the basal member
of its own (axial) side; in other cases, however, the ovicell may

ON THE BRITISH SPECIES OF CRISIA. 157

replace the third zocecium above a joint (and it is then abaxial),
but it is very rarely found higher in the internode. A branch
is never given off by an ovicell.

As the age of the ovicell increases, fresh zocecia continue to
be added above it up to a certain point. The old ovicell seems
to be always surmounted by a considerable number of zoccia;
in the specimen shown in fig. 6 there are, in addition to two
incompletely formed zocecia—the last that this branch would
have produced—ten zocecia above the ovicell. It must be noted
that in this and other similar cases all zoccia which are
further from the joint than the ovicell are described as being
above the latter. The second zoccium of the right side in
fig. 6 may not, at first sight, appear to be in this position,
although an examination of the lower end of the ovicell at once
shows its real place in the series.

A joint is seldom developed above the ovicell, and the growing-
point usually completely exhausts its power of developing fresh
zocecia after a certain period.

The joints of this species are pale-coloured, or more usually
yellow. In old parts of the colony the joints may become very
dark, or almost black ; this is especially true of those parts
which form the starting-point for the regeneration of fresh
branches. The joints are probably never so dark as they are
normally in C. denticulata.

Smitt, in his valuable paper on Crisia,! gives a series of
formule illustrative of the branching, &c., of the forms of this
genus, and many of these formule illustrate in a most instructive
manner the tendency of some at least of the species of Crisia
to develop even-numbered internodes without branches. In his
explanation to No. 8 of this series Smitt expressly points out
that, in Nos. 4—8, shorter branchless internodes may alternate
with longer internodes which have developed branches. It isa
noteworthy fact that the greater number of the branchless
internodes shown in these formule have an even number of
zocecia, and that the number is odd in most of those internodes
which have developed branches. ‘This fact seems, however, to

1 © Krit. Férteckn.,” I, ‘ Ofvers. af K. Vet.-Akad. Forhandl.,’ 1865.

158 SIDNEY F. HARMER.

have escaped Smitt’s attention. It must further be pointed
out that some of the exceptions to the rule which has been so
much insisted on above are probably due to the fact that some
of the formule refer to C. aculeata, as is admitted by Smitt
in two of the cases.

A very interesting abnormality of C. eburnea is figured in
Pl. XII, fig. 5. The internode in question was the penultimate
internode of a branch of a thoroughly characteristic colony, in
which no other abnormalities were detected. In addition to
bearing two lateral branches in a very unusual position, at its
upper end, this internode distinguished itself by producing
three zocecia arranged in a row along the middle of its front
surface, giving it, when seen from this side, an appearance very
much like an Entalophora, for instance. The back of this
internode appeared normal, and it was not obvious that any of
the three growing-points borne by the internode was constructed
in such a manner that it would have reproduced the abnormality
in the next following internodes.

C.aculeata. Fig. 4.

I believe this form, which is in many respects intermediate
between C. eburnea and C. ramosa, and which was originally
distinguished as a species by Hassall,' to be a perfectly good
species. Nearly all recent authors have regarded it as a variety
of C. eburnea; this view is taken, for instance, by Hincks,?
Busk,® Smitt,* &c. Even Johnston,’ although inserting it as
a distinct species, adds that he cannot persuade himself that it
is more than a variety of C. eburnea.

My belief in the specific distinctness of C. aculeata rests
mainly on the characters of the ovicell; since a particular form
of ovicell (shown in fig. 4) is invariably found on colonies of

1 Hassall, A. H., ‘Ann. and Mag. of Nat. Hist.,’ vol. vi, 1841], p. 170.
2 ¢ Brit. Mar. Polyzoa,’ p. 421.

3 *Cat. of Marine Polyzoa in Brit. Museum,’ part iii, p. 4.

4 Loc. cit.

5 * British Zoophytes,’ ed. 2, p. 285.

ON THE BRITISH SPECIES OF ORISIA. 159

the aculeata type, and never occurs in any other type of
colony.

The method of branching and the number of zoccia in an
internode are far less constant than in the last two species.
Some of the characters of C. aculeata may, however, be
illustrated by a formula, using the letter s to indicate the
position of a spine, in addition to the symbols which have been
employed in other cases.

‘a
(1) +(4) + (8) + (4438) + (9+ 47-5, +13) H(7 Hy) + (5 $7) + (7417) + (T+7) +@)

=(5+ Ov.+10+5,+7,+ 57) +(2)
=(8+)8)+(5+,7)+(04+744+37)+(1 +2)

| =B8+yr+e)
=(z)

=(5+7,+ 0v.+2)
——————

|

=(1+ 72)

=(6+ 0v.+5+,7+7,+2)

=(1+2)
=(6+)7+2)

=2+2)

The formula represents the whole of a main stem, given off
from a rootlet, together with the whole of one of its lateral
branches and a portion of another. In several important respects

’ this formula differs from those which have been given of the pre-
ceding species. Itis by no means uncommon to find completely
formed internodes which have an even number of zowcia; and
these even-numbered internodes do not conform to the denti-
culata or eburnea type by being usually without branches.
In some cases, indeed, an even-numbered internode has no
branch; in other cases it has two branches, one on each side;
in others again it may have one branch, or it may have a spine,
which, as has often been pointed out, is a structure which may
be regarded as a suppressed branch: like the true branches,
the spine has horny joints at intervals, and is attached to the

VOL. XXXII, PART IIl.—NEW SER, L

160 SIDNEY F. HARMER.

zocecium by means of a basal piece which is quite similar to
that of a normal branch.

The spines shown in fig. 4 have been artificially bent back-
wards; in their normal position they curve over the front of
the branches.

The number of spines developed on a colony is extremely
variable ; in a few cases spines are altogether absent, and the
species could then hardly be distinguished with certainty from
C. ramosa, were it not for the presence of the characteristic
ovicells. Although, in one or two cases observed, an internode
had developed spines on all or nearly all its zocecia, it is not usual
to find more than one or two spines on a single internode,
while a large proportion of the internodes of a colony do not
develop any of these structures. The spines most commonly
occur on the lower zocecia of an internode, and are commonly
in the position—

(7 + 8 + 7)
or (7 + 8 + 8. + 73)3

being found on the abaxial side if the internode is, as is often
the case, the basal member of a branch.

The spines may, however, be developed in other positions ;
thus it often happens that the last structure developed at the
apex of a branch, before the growing-point ceases to grow,
is a spine,’ which is situated on the axial side of the last
zocecium, and is consequently in the position of the terminal
zocecium of the branch to the right of the ovicell in fig. 4.

In the specimens (most of them from Plymouth or Roscoff)
which have come under my notice the presence of a single spine
on a colony has been quite sufficient to enable the species to be
identified with certainty as C. aculeata. It is perfectly true
that the lower parts of the zoarium may have an eburnea-
like appearance ; but the colony, if well grown, seems always
to acquire the aculeata form of the zoccia and internodes
towards the ends of the branches.

I have in no case found an ovicell of the type shown in fig. 6

1 Cf, C. acuminata, Busk, ‘ “Challenger” Rep.,’ part 50, pl. iii, fig. 1.

ON THE BRITISH SPECIES OF CRISIA. 161

(C. eburnea) on a colony which, from the presence of spines
or from other characters, was found to belong to C. aculeata.
I cannot admit that there is sufficient evidence to show
that this form is merely a variety of C. eburnea. Both
in the form of its zocecia and in its method of branching
it is totally unlike this form, although it is sometimes with
difficulty distinguished from C. ramosa.

The branches of C. aculeata are usually slightly incurved,
but not nearly to the same extent as in C. eburnea; and it
does not possess the well-developed helicoid cymes of the
latter species. Many of the internodes bear two branches,
usually on opposite sides, but more rarely on the same side.
In well-developed colonies the terminal internodes, and espe-
cially those which possess ovicells, are commonly provided
with two branches.

The number of zocecia in an internode is extremely variable ;
it is usually small in the lower internodes of a stem, such
numbers as 1, 2,3, and 4 being common in this position. The
next parts of the stem, and the basal parts of the lateral
branches given off by it often assume an eburnea-like appear-
ance, the internodes consisting of 5 or 7 zocecia. At the ends
of well-developed branches the number usually becomes
higher ; a terminal internode with 22 zocecia has been observed,
although this number is higher than is usually the case.
When the terminal internodes have many zoccia they usually
bear two branches; but if the number of zoccia is still larger
the number of branches may increase to as many as five.

The position of the branches is another very variable
feature. In the lower parts of a colony the branching takes
place commonly from z,; while higher up, although some of
the branches still come off from z,, others are given off quite as
commonly from z,, and in many of these cases the zocecium
below the branch, and on the same side, bears a spine.
Branching may, however, take place from the higher zocecia of
an internode, as from z, or z,; and when several branches
come off from the same internode, those which are last
formed have a very high position in the internode, The

162 SIDNEY F. HARMER.

most striking case observed illustrating this point had the
formula—
7+ 0.+1+7+ 7+ 75 + & + 0 + 2).

It cannot fail to be remarked that the character of the
branching is much more variable in this species than in
C. eburnea.

The joints are usually yellow; the articulations which bear
the branches are usually short, and are then very similar to
those of C. eburnea; in some cases, however, they acquire
the form characteristic of C. ramosa.

Near the ends of the branches, where most of the zoewcia
have polypides, the ends of the zocecia are, in most cases, long
free tubes, and are thus strikingly different from those of
C. eburnea. The free portions of the zowcia are either
gradually bent forwards from the point where they leave the
branch, or they may be bent forwards at a distinct angle from
this point. The curvature of the zoccia is, in either case,
different from that of C. eburnea. The zoecia are dis-
tinctly longer and more “ loosely aggregated” than in that
species; and the branches are usually of slenderer habit (as
recognised by Johnston!).

The ovicell is nearly always higher in the internode than in
C. eburnea. In the average of a considerable number of
observed cases the ovicell was in the position of the 5th—6th
member of the internode above the joint, and thus replaced
z, or 3. In one case the ovicell replaced the 8th zoccium,
and in another it was the 3rd unit of the internode; in no
case was it found lower.

The stem is not usually jointed above the ovicell ; and fig. 4
is, consequently, a somewhat exceptional case. As in the pre-
ceding species, the ovicell is normally borne by a terminal
internode ; and a considerable number of zocecia may be added
above the ovicell.

1 «Brit. Zoophytes,’ ed. 2, p. 286,

ON THE BRITISH SPECIES OF CRISIA. 163

C.ramosa,n.sp. Fig. 11.
Some of the characteristics of this species are well exhibited
by the formula!—
(7+ yr+r,)+(11+57)+(10+7.4+-57)+(5 +7242)

=(12+ 7r+7r4)+
=(l2+y+r)+
— (5 +0v.47+yrt+ytr;+ +2)
=(6+.r+2)

=(z)

| =(S+urt75+2)

=(9+ 00.47 4+r.+yrt+rotirtz) | | (2)

—=(z

=(3+2)

=(9+.7+73+2)

=(1+-2)
=(4+.7+72)

=(10+7,+2) |

=(1+2)

The branching is seen to be very similar to that of C. acu-
leata, but the tendency, already manifested by that species, to
develop more than one branch from an internode, is here
carried much further, so that a considerable proportion of the
internodes have two branches each, while the terminal inter-
nodes, if the colony is well grown, will be found to have at
least two each.

The rule relating to odd- and even-numbered internodes, so
characteristic of C. denticulata and of C. eburnea, here
breaks down altogether, as, indeed, was the case to a consider-
able extent in C. aculeata. Odd-numbered internodes are
not much commoner than even-numbered ones, and either kind
may produce one or more branches, or be altogether branchless.
The first branch of an internode is, however—as in other
species—nearly always developed on the side of the basal
zocecium, and the last zocecium of an internode is very often

=(2)

1 It is obvious that, in the case of the ovicell-bearing internodes, some of
the branches are given off above the ovicells. For the purposes of the
formula, however, the ovicell is counted as an ordinary member of the inter-
node. A branch is probably never developed from the ovicell itself.

164. SIDNEY F. HARMER.

situated on the side on which the last branch is developed,
thus causing the position of the basal zocecium of the next
internode, and consequently of the first branch of that inter-
node, to be on the opposite side.

Just as C. eburnea is, on the whole, characterised by
branching from 2,, so this species may be said to branch nor-
mally from z,, or to produce branches on the type (7% +n+27);
where ” usually represents 2, or 2}.

The extent to which this must be taken as a general rule
may be understood by the following analysis of the complete

formula of a well-developed colony :
Number of Cases.

(1) Internodes with one branch, originating from 2, . o Da
(2) Branches arranged on the type (7,+47) . 4 : eb
(3) » ” 3 (7,+37) or (73+57) - 5
(4) 2? ” ” (ro+or) 7 1
(5) 33 32 I) (72+37) 5
(6) » 29 3 (7, +27) 1
(7) 39 2? ” (73+47) is 2
(8) One branch only, originating from z, : : E a) ote
(9) 2» 2 9 Z3 8
(10) ” ” ” 4 7
(11) 39 33 ”) a5 3
Total number of internodes which had developed branches 63

Only eight of the completely developed internodes were
branchless. Thus in this particular colony, in which no ovi-
cells were present, and in which no internode possessed more
than two branches—

30 p.c. of the branching internodes bore two branches ;

38 p.¢. . a5 », one branch, originating from z,.
32 p. Cc. Pr 33 “3 FA 5 from other
zocecia,
100 p. c.

Or, adding together Nos, 1 to 4, the cases in which the branches
come off from z, or in which the second branch is two zoccia
higher than the first, we find that these cases amount to 55°5
per cent. of the total number of branching internodes, and this
may be taken as a case which does not exaggerate this feature
of the branching.

ON THE BRITISH SPECIES OF CRISIA. 165

Since some of the internodes which bore branches were
immature, and had not had time to develop more than one
branch, the figures would have been slightly different if the
growth of the colony had been complete.

The symmetrical character of the branching noticed in other
species is also found in C. ramosa. Thus the branches
originating from an internode whose formula was (14+.37+7;)
developed altogether five internodes from which new branches
were given off; in two of these cases the branch was borne by
23, in two more by z,, and in the last case by 2, and other cases
of the same kind may easily be found.

The symmetry of the branching comes out with special
clearness in the case of some abnormalities, of which the
formula given on p. 163 is anexample. Two consecutive inter-
nodes of the main stem represented in the formula give off
internodes whose formule are identical. One of these gives off,
on its right side, a lateral branch consisting of a single long
internode bearing an ovicell, and the other gives off a precisely
similar branch on its left side. Both of these ovicells have the
same deformity, having developed a constriction at a particular
point near their upper end; and it will further be noticed that
the symmetry extends, to some extent, to the branches given
off by the internodes which bear these ovicells.

Fig. 12 represents an abnormal ovicell or zocecium of a type
found in more than one colony. The growing-point appears to
have started with the intention of developing an ovicell, and
then to have altered its original purpose, and to have developed
the incipient ovicell into an abnormal zoccium. This alteration
of purpose may have been due to the failure of the young
ovicell to develop the egg! which is normally found in the
immature ovicell.

The point which immediately concerns us at present is that
each of two consecutive internodes of the same stem of this
particular colony developed lateral branches, one on each side
of the stem, and that each of these lateral branches bore an
ovicell of this peculiar ‘‘suppressed”’ form. The same colony

1 ¢ Proc. Cambridge Philosoph. Soe.,’ vol. vii, p. 48.

166 SIDNEY F. HARMER,

possessed two more of these suppressed ovicells, two ovicells
showing other abnormalities, and several normal ovicells.

In another colony a normal ovicell, with a “ suppressed ”’
ovicell on a lateral branch on each side of it, was noticed.

In another case (fig. 13) a single internode bore no less than
four ovicells, and the colony to which this belonged possessed,
in different parts, four internodes, in each of which two ovicells
had been developed. It may be noted that the occurrence of
two ovicells, side by side, in the same internode, is deseribed
by @Orbigny! in C. patagonica, apparently as a normal
feature of the species.

These cases, and the general remarks which have been made
with regard to the branching of various species of Crisia,
show that the growth of the colony is even more definite in its
character than would appear from a superficial examination,
and that in each particular species the tendency to vary is sub-
ordinated to certain principles of growth, which give rise to the
special symmetry which characterises the species.

The number of zocecia which compose an internode is even
more variable in C. ramosa than in any of the other species.
Generally speaking, the number is smaller near the base of the
colony, and larger near its periphery, although this rule is by
no means absolute. The length of the internode depends
mainly on the number of zocecia it possesses. The longest
which was measured was a terminal internode in which growth
had ceased, and which consisted of 28 units, of which the 10th
was an ovicell; its total length being slightly more than 7
millimetres. These long internodes usually show a well-marked
double curve, like a much elongated S, just as was remarked
in C. denticulata; and they commonly bear 3, 4, or even 5
branches.

The appearance of the internodes depends greatly on the
condition of the zocecia. Near the ends of the branches the
zocecia generally have very long tubular mouths, and, for the
most part, contain a functional polypide. This is especially

1 D’Orbigny, A., ‘ Voyage dans l’Amérique méridionale,’ tome v, 4° partie,
1839 and 1846, p. 7, pl. i, fig. 1.

ON THE BRITISH SPECIES OF CRISIA. 167

true of actively growing colonies found early in the year: ata
later period, when growth is less energetic, the apertures may be
much less prolonged, and in many cases they are not more
prominent than in some specimens of C. denticulata, even
in the case of those zocecia which are not closed by a diaphragm.
Lower in the stem the tubular mouths are, in most cases, lost ;
the zocecium is closed by an oblique diaphragm, and no poly-
pide is present. The internode is then a flattened structure,
in which the apertures of the zocecia project even less than in
C. denticulata.

In an interesting abnormality found in August, two promi-
nent tubular apertures occurred, side by side; the extra
zocecium being in the position which would normally have been
occupied by a basis rami.

Busk! has made rather a point of the fact that in C. conferta
the free tubular portion of the zoccium is not a mere produc-
tion of the peristome, but presents “ the same puncturation as
is seen on the rest of the cell.” This is certainly the case in
C. ramosa and in C. aculeata, and to a less extent in C.
denticulata. C. eburnea is, in fact, the only species I have
examined in which the tubular portion is usually merely a thin
prolongation of the peristome.

The average position of the ovicell is somewhat higher than
in C. aculeata; but that it varies greatly in position is ob-
vious from the formula—

(44+ 0v.419+7,+ortyrtrs)+

=(13+ 00.+9+.7+7,+6r+7.)+
It has never been noted to be lower than 4th in the internode ;
but it is seldom so low as this. The branch may be jointed
above the ovicell, although most commonly the ovicell is borne
by a terminal internode, which usually possesses at least two
branches. When several branches are developed, two of them
are usually developed not far above the ovicell, one on each
side of the internode, whilst the other branches are developed
from the lower parts of the internode. This is the case in

1 “Catalogue .... Brit. Museum,’ part iii, p. 7.

168 SIDNEY F. HARMER.

fig. 11, and in both the above-cited formule in which the ovi-
cell replaces z, and z, respectively.

The joints of this species are yellow, or more rarely brown.
They are never black. Rootlets are very freely developed from
the base of the stem, and they may attain a great length.
They usually originate rather low on the zocecia and from their
lateral edges. As in other species, they become very firmly
attached to stones and other objects, and form creeping stolons,
from which (as well as from rootlets which are not attached in
this way) fresh stems may originate. The colonies do not so
often consist of a single main stem as in C. eburnea. It is
frequently remarked that the longest and most branched parts
of the colony are lateral branches, and not parts of the main
stems.

Ovicells—In C. ramosa (figs. 10, 11) the ovicells are consi-
derably larger than in any of the other species (see figures, all of
which are drawn to the same scale, and table of measurements
on p.177). They are regularly pear-shaped, their main axis
being straight; they are much inflated above, their curvature
diminishing gradually in all directions from their most pro-
minent portion. The aperture is in the form of a distinct
funnel-shaped tube, which is considerably smaller at its base
than at its mouth; and the mouth of the funnel, the actual
aperture of the ovicell, is more or less circular. In the shape
of the aperture this species differs from all the other British
forms.

The tubular aperture is of course not present in incompletely
developed ovicells: an account of the development of the ovi-
cells will be given in a forthcoming paper. It must also be
noted that the aperture is liable to be broken away in old ovi-
cells, and that in many cases, where the ovicells or their con-
tents are not normally developed, the tube itself is not formed.
In normal and completely developed ovicells the shape of the
aperture is, however, a perfectly characteristic and constant
feature.

C. aculeata (fig. 4), which in some other respects is so
similar to C. ramosa, has a much smaller ovicell than that

ON THE BRITISH SPECIES OF GORISIA. 169

species (compare fig. 4 with fig.11).! Its shape is very charac-
teristic, its most prominent portion being considerably nearer
its distal end than in C.ramosa. From this point the ovicell
slopes off very suddenly towards its aperture, and more gradu-
ally towards its base, although this latter slope is steeper than
in C.ramosa. The aperture is not borne on a distinct tube,
but it lies in a characteristic position on the zocecium next
above the ovicell, on the same side of the internode. This
zocecium curves forwards round the back of the ovicell, the
aperture of which is situated on it at the point where it makes
its appearance above the ovicell.

In C. eburnea (fig. 6) the ovicell is large—considerably
larger than in C. aculeata. Since the base of the internode
which bears it is distinctly curved inwards, the ovicell itself
has the same curvature at its base, as is best seen when the
ovicell is looked at from the side. The ovicell is well inflated,
and slopes away more gradually from its most prominent point
than in C. aculeata. The aperture is quite characteristic ;
it is borne on a tube-like structure, distinctly broader at its
base than at its free end, and instead of being circular, as in
C. ramosa, it is transversely elongated, its lower border being
often slightly convex towards the centre of the aperture.

In C. denticulata (fig. 3) the ovicell is fairly large, and
usually becomes level with the flat surface of the internode
near its base, the distal portion of the ovicell being very pro-
minent. The aperture is not situated on a well-developed
tube ; it is not, however, on the surface of a zocecium, as in
C.aculeata, but is situated between two zoccia, and it is
very nearly sessile on the top of the ovicell, as was the case in
C. aculeata.

In all four species the aperture is connected with the top of
the ovicell at the point where the latter joins the front surface
of the internode.

The importance of the form of the aperture appears to have

1 It must, however, be pointed out that the ovicell of C. ramosa may be
smaller, and that of C. aculeata larger, than in the particular specimens
figured.

170 SIDNEY F. HARMER.

been almost completely overlooked by all previous writers on
Crisia. ‘The aperture is often said to be absent, as indeed it
may be in injured or abnormal ovicells. As will be seen from
a later communication, a normal ovicell is, according to my
observations, never without an aperture from the time when
the ovicell is first developed at the growing-point to the time
when embryos are ready to escape from the ovicell. The cal-
careous aperture is, however, throughout the development
closed by a thin cuticular membrane, and the presence of this
membrane sometimes makes it difficult to see the aperture in
those cases in which this structure is not borne on a distinct
tube.

On breaking open an ovicell (fig. 10) it will be noticed that
the aperture leads into a space partially separated from the
rest of the ovicell by a valve-like structure of calcareous
nature. This valve has very definite relations to the structures
found in the interior of the ovicell, as will be described in my
subsequent paper. It springs from the posterior wall of the
ovicell, and passes obliquely forwards, being also attached to
the lateral walls of the ovicell in such a way as to leave a more
or less oval opening connecting the main cavity of the ovicell
with the aperture of the latter. The valve is most developed
at the back of the ovicell, and gradually dies away laterally as
it passes to the front wall of the ovicell, where it no longer
forms a distinct ridge.

This valve is developed in all the four species which are
specially discussed in this paper, but it appears to be less well
developed in C. eburnea than in the other species,

Note on C. cornuta, Linn., and C. geniculata, M.-Edw.

Until quite recently I had devoted no particular attention
to these forms, the specific identity of which appeared to be
perfectly established by such statements as those of Smitt,' to
the effect that they may both occur as branches of the same
stem. But, having recently found some ovicells of C. geni-
culata, I cannot help believing that the two forms are speci-

1 « Ofvers.,’ &c., 1865, No. 2, p. 128.

ON THE BRITISH SPECIES OF ORISIA. 171

fically distinct, and a more careful examination of C. cornuta
has convinced me that Smitt’s statement may be explained
in such a way that it is unnecessary to follow him in his con-
clusion.

Even if the two forms are not really distinct, it appears to
me worth while to call attention to what would then be an in-
teresting case of a definite variation of the ovicells correlated
with the presence or absence of spines on the zowcia.

C. geniculata consists typically of a series of internodes,
each of which is composed of a single zocecium ; from opposite
sides of this zocecium arise a pair of branches which are not quite
at the same level. An excellent figure of this form is given in
Busk’s ‘ British Museum Catalogue’ (part 3), pl. i, fig. 2. On
comparing this figure with fig. 7 on the same plate (representing
C. cornuta), it will be seenthat C. cornutaexactly resembles
C. geniculata as far as those zoecia which bear two branches
are concerned; but that, as Busk points out (p. 3), one of the
branches is usually replaced by a jointed spine.

It is obvious that spines will be absent in C. cornuta if
two branches are developed from each zocecium. Further, the
spines are very readily broken off, and a close examination is
then sometimes necessary to discover the small basis with which
the spine articulates. From one or other of these causes I have
several times observed branches of normal coloniesof C. cornuta
having a close resemblance to C. geniculata; and this may
be the explanation of Smitt’s statement referred to above.

In every case in which I have observed the ovicells—although
I must add that I have not obtained many ovicells of C. geni-
culata—I have noticed the following characteristic differences
between the two forms ; a reference to Busk’s figs. 2 and 10
(1. c., pl. i) shows that the forms examined by Busk were
similar to those which I have myself found. Busk’s fig. 4 (C.
geniculata) does not, however, quite agree with the specimens
which I have examined.

The ovicell of C. cornuta (fig. 9) is the basal and only
member of its own internode ; it bears a lateral branch on each
side, these branches originating at not exactly the same level.

172 SIDNEY F. HARMER.

After the branches have been given off, the ovicell becomes
perfectly free, and is in this part considerably inflated. The
tubular aperture arises near the back of the ovicell, and is
usually bent somewhat backwards from its point of origin ; so
that, in looking at the branch from its “front”? surface, the
base of the tubular aperture is nearer to the observer than its
distal end.

In C. geniculata, on the contrary, a common arrangement
is as follows:—The basal member of the internode is an ordinary
zocecium (figs. 7 and 8), which gives rise to the ovicell as the
second member of the internode. Immediately above the
ovicell is another zocecium, which gives off a lateral branch
near the level of the upper end of the ovicell. The basal
zocecium itself gives off a branch on the opposite side to the
ovicell. The internode may thus be represented as—

ee
iz r, (counting the ovicell as the basal member of its own side).

The ovicell itself is distinctly smaller than in C. cornuta, and
is not much inflated at its upper end. Its tubular aperture is
most distinctly bent forwards from its base, sometimes at a
very sharp angle, and the actual aperture is smaller than in C.
cornuta. Moreover, the ovicell is not free, as in the latter
species ; the upper zocecium of the internode being closely
attached to its back along the greater part of its course. This
zocecium ultimately becomes free from the ovicell, and curves
forwards above the upper end of the latter.

In other cases the ovicell may be the third member of the
internode, each of the two zocecia below it giving off a branch ;
and two zocecia, each bearing a branch, may occur above the
ovicell. The commonest arrangement seems to be either that
given in the above formula, or the occurrence of three branch-
bearing zoccia, one of which is below the ovicell and the other
two above it (cf. Busk’s fig. 2). The ovicellis, in any case, not
the lowest member of the internode, and one or two zoccia
are always attached to its back.

C. geniculata is a slenderer and more delicate form than

ON THE BRITISH SPECIES OF ORISIA. Lz

C. cornuta, its zowcia being distinctly longer and thinner than
in that species; and spines seem to be never developed. As
already remarked, however, parts of the colonies of C. cornuta
may be devoid of spines.

Breeding Period and Occurrence of Species.

The specimens from which the following statements are made
have been received at various periods from February to the end
of August. I have to offer my best thanks to those who have
most kindly assisted me by supplying me with material; and
especially to the staff of the Marine Biological Association,
Prof. H. de Lacaze-Duthiers, and Mr. J. Sinel.

The dominant species at Plymouth is certainly C. ramosa;
although, strangely enough, I have not been able to obtain this
form from any other place, except from a bottle found in the
Morphological Laboratory at Cambridge. The contents of
this bottle came either from the Channel Islands or from Arran !

At Plymouth, C. ramosa is found commonly at depths from
4 to 30 fathoms. It is particularly fond of growing on stones,
but is found on other objects—e. g. glass bottles, shells, red
seaweeds, Cellaria, and sponges. When it grows in the last
position it appears to be in danger of being killed by the sponge,
which grows over its branches. The Crisia is, however, gene-
rally able to keep pace with the growth of the sponge, so that
only the basal parts of itscolonies are killed, or at least prevented
from having functional polypides by the sponge. The speci-
mens growing at 4—6 fathoms were usually much more luxu-
riantly branched than the few specimens which I received
from 20—380 fathoms ; and ovicells were obtained only from
those growing under the first set of conditions.

C. eburnea is also common at Plymouth, but is almost
always found on red seaweeds or on Sertularia. ‘The restric-
tion of various species of Crisia to particular seaweeds, &c.,
has been often noted by previous observers.

Winther! has stated that, in the Danish forms of this species,

1 G. Winther, “ Fortegnelse over de i Danmark hidtil fundne Hav-Bryozoer,”
‘Naturh. Tidsskrift’ (Kjpbenhavn), 3 Raekke, vol, xi, 1877-8, p. 7.

174 SIDNEY F. HARMER.

the effects of the brackish water of the Baltic can be easily ob-
served by comparing colonies found in different localities.
Those which are nearest to the Baltic are said to have inter-
nodes consisting typically of three zoccia; in those most
exposed to the North Sea the internodes have seven zowcia ;
and in colonies from intermediate localities they have five
zocecia. I have not been able to observe any definite correla-
tion between the character of the colony and the conditions
under which it was growing.

C. aculeata is less common at Plymouth than either of the
preceding species; it was found on stones, red seaweeds, and
sponges, usually from 4—5 fathoms.

C. cornuta was fairly common, mostly on red seaweeds ;
while C. denticulata was seldom found at Plymouth.

By the kindness of Prof. de Lacaze-Duthiers [ received a
large supply of Crisia from Roscoff in June. The species
most common at Roscoff appear to be C. aculeata, C. denti-
culata, and C. cornuta. C. geniculata was less common,
and only a few fragments of C. eburnea were found.

From Mr. J. Sinel I have received numerous specimens of
C. denticulata and C. cornuta found at Jersey ; and a smaller
supply of C. eburnea, C. geniculata, and C.aculeata. The
Jersey specimens of C. denticulata were found between tide-
marks, while Smitt! states that C. denticulata is pre-eminently
a deep-water form. It is, however, probable that Smitt’s speci-
mens did not really belong to this species.

I have also obtained specimens of several species from
Guernsey and the Scilly Islands.

Smitt’s valuable contributions to our knowledge of the Poly-
zoa have been devoted, so far as they concern the genus Crisia,
to showing that the “ species” which have been distinguished
in this genus are in reality “ forms” of a single species.

In one of his later papers? Smitt remarks, speaking of the
forms of Crisia discovered in the expedition to which his paper

1 ¢Ofvers.,’ &c., 1865, No. 2, p. 188.

2 “ Recensio, syst. .... Bryozoorum .... Novaja Semlja, &c.,”
‘ Ofversigt. af-K. Vet.-Akad. Férhandlingar,’ 1878, No. 3, p. 12.

ON THE BRITISH SPECIES OF ORISIA. 175

refers, that he has united all these forms in a single species ;
and adds, “ Auctores vero si sequi volumus, unamquamque fere
coloniam speciem distinctam habebimus.”

Although not in the least denying the difficulty of finding
satisfactory specific characters other than those derived from
the ovicells, the result of my investigation has been to con-
vince me that Smitt has gone too far in denying the specific
value of certain of the forms of Crisia. My results may
possibly have to be explained by a suggestion which Smitt
himself throws out, to the effect that although the series repre-
sented by the various forms of Crisia living in different locali-
ties is one in which practically none of the stages in the evolu-
tion of the species have been lost, “vivit multis in locis altera
vel altera forma tam constans, ut species bene distincta facile
censeatur.”! Unfortunately, all the localities from which I
have been able to obtain Crisia are comparatively close to one
another ; but I have found no essential differences between the
forms of the same species from different localities.

Until it can be shown that any two or more of these forms
can be developed as branches from the same stem, or as stems
from the same rootlet, or at least that they can be produced as
descendants of the same form, it appears to me that it will be
impossible to deny to them the rank of species.

C. eburnea begins to breed at Plymouth as early as
February; ovicells are present in great numbers during
March, April, and May. Towards the end of the latter month
they disappear, and are not normally present on colonies
found in the summer. March and April appear to be the
months when oviceils are most common.

C. ramosa commences to breed (at Plymouth) in April; a
few of the colonies found at this period have young ovicells.
In May young ovicells are very common ; and the breeding
period continues from this time until August at any rate.
Immature ovicells may be found even at this time, but they
are then becoming uncommon. The breeding season is pro-
bably at its height in May and June.

1 « Ofvers.,’ &¢., 1867, No. 6, p. 461.
VOL, XXXII, PART I1.—NEW SER. M

176 SIDNEY F. HARMER.

C. aculeata.—A large proportion of the specimens found
at Roscoff in June possessed numerous ovicells. At Plymouth
ovicells were also found in April and May. The breeding
season is probably much the same as in the last species.

C. denticulata.—The only specimens obtained in which
ovicells were common were found in Guernsey and Jersey in
the summer (June to August), which may probably be regarded
as the period at which the breeding season is at its height.

C. cornuta.—The ovicells appear to be commonest in
April and May.

C. geniculata.—The only specimens in which I have found
ovicells were obtained in the summer (June to August). I
have net found this species at Plymouth.

ti

ON THE BRITISH SPECIES OF ORISIA.

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178 SIDNEY F. HARMER.

In A,B, and C of the above table the length of the internode
is measured from joint to joint, or from joint to apex of
branch if the internode is a terminal one. The ‘“ average
length” of the zoccia is obtained by multiplying the length of
the internode by two, and dividing by the total number of
zoecia. It is obvious that this does not give the total length
of the zocecium, since the zoccia overlap one another; but an
approximation to the distance from mouth to mouth of the
zocecia is obtained. This gives a more accurate average than
any single measurement of this distance would give, since the
distance is variable in connection with the extent to which the
tubular apertures of the zoecia are developed, and with other
circumstances. For the purposes of this calculation an ovicell
is counted as an ordinary zoecium. If the length of the inter-
node of five zocecia shown in fig. 6 (C. eburnea) be compared
with that of a corresponding number of units, beginning at the
the base, of the ovicell-bearing internode in the same figure, it
will be seen that the presence of the ovicell does not affect the
result so much as would be expected at first sight, and the
error due to counting the ovicell as a zocecium is further
lessened by the fact that this structure is nearly always borne
on an internode which consists of many zoccia.

In D the length is measured from any point of an aperture
to the corresponding point of the aperture of the next zocecium
on the same side of the internode, and in making this measure-
ment two zoccia whose tubular mouths were about equally
developed were always chosen.

E gives the total length of zocecia with well-developed tubular
apertures from the point where their cavity disappears at their
proximal end to the furthest point of their apertures, the measure-
ments being made from transparent (Canada balsam) speci-
mens.

G is measured immediately above the aperture of a zocecium.

H gives the length of the base with which a lateral branch
articulates.

I. The length of the ovicell is estimated by drawing an
imaginary line joining the point where the zoccium next

ON THE BRITISH SPECIES OF ORISIA. 79

below the ovicell on the same side becomes free from the
internode (if any tubular aperture is developed), with the corre-
sponding point on the zocecium next above that zocecium on the
other side of the internode. The “length” of the ovicell is
the distance of the middle point of the line drawn as above to
the uppermost point of the ovicell, exclusive of its tubular
aperture, if any.

This measurement, on the whole, gives the most constant
results. The relation of the zocecia below the ovicell to the
ovicell itself is very variable, and it is impossible to take one of
these zocecia as a fixed point, a more definite result being
obtained by taking two as described above. In C. eburnea,
if the ovicell is the second member of an internode the
imaginary line is drawn from the base of the tubular aperture
of the first member of the internode in a direction parallel to
that connecting the apertures of the upper pairs of zoccia in
the internode (a method which usually gives more satisfactory
results than might be supposed from fig. 6).

The numbers given in the table do not profess to give the
total limits within which a given part may vary, but the limits
given will probably be found to include nearly all the variations
which are observed in the ordinary forms of colonies.

In cases where it seemed to me possible to define the normal
size of any part, I have put this down as the “ average ”’ size.

An examination of this table shows that C. eburnea,
C. aculeata, and C. ramosa form an almost continuous
series, as Smitt indeed has stated. C. denticulata can hardly
be confused with any of the other forms, amongst which
C. eburnea is the one that resembles it the most.

It will be seen that in every single measurement given
C. ramosa is the largest of the four species, although in the
size of the entire colony C. denticulata surpasses it. In
many of the measurements, however, the upper limit of
C. aculeata overlaps the lower limit of C. ramosa. As
these are the two species which most closely resemble one
another, and which are probably very closely allied to one
another, it seems to me that it is not possible, in all cases, to

180 SIDNEY F. HARMER.

distinguish between a colony of C. aculeata without spines
and a small variety of C. ramosa unless ovicells are present.

C. ramosa further appears to me to be much the most
variable of all the species I have examined, while C. eburnea,
and, next to this, C. denticulata, are the least variable. The
greater variability of C. ramosa comes out perfectly clearly by
subtracting the minimum from the maximum measurements
given in the table, and comparing together the results thus
obtained for the different species. If this be done for D—J, it
will be found that, in every case except F (in which the limits
are the same for C. ramosa and C. denticulata), the greater
variability of C. ramosa comes out, and usually with striking
distinctness.

It will be noticed that the largest variations are in the length
of the zocecia, and, correlated with this, in the size of the
ovicells. But though the ovicells are thus by no means exempt
from variation, their principal specific character (the shape of
their aperture) is retained throughout without material alte-
ration.

The fact that a colony of C. eburnea, on which some
unusually small ovicells were present, has been taken into
account in the table, makes these structures appear much
more variable than they are in normal cases.

ON THE BRITISH SPECIES OF ORISIA. 181

EXPLANATION OF PLATE XII,

Illustrating Mr. Sidney F. Harmer’s paper “On the British
Species of Crisia.”

(All the figures were drawn with a camera lucida under a Zeiss a objective,
and were subsequently reduced 24 diameters.)

Fie. 1.—C. denticulata.—Young internode, with growing-point, seen
from the back (Canada balsam preparation).

Fie. 2.—C. denticulata.—lllustrating the relations of an even-numbered
internode (pp. 147, 148).

Fie. 3.—C. denticulata.—Ovicell (p. 169, &.).
Fie. 4.—C. aculeata.—Ovicell (p. 168, &c.).
Fic. 5.—C. eburnea.—Abnormality (p. 158).

Fic. 6.—C. eburnea.—Ovicell (p. 169, &c.). The internode which bears
the ovicell has two branches, an unusual arrangement.

Fie. 7.—C. geniculata.—Internode with ovicell ; back view (p. 172).

Fic. 8.—C. geniculata.—Another ovicell, seen from the front (p. 172).
Greatest diameter of ovicell = °208 mm.

Fic. 9.—C. cornuta.—Ovicell (p. 171).

Fie. 10.—C. ramosa.—Ovicell broken open to show the valve (p. 170).
Fie. 11.—C. ramosa.—Branch with ovicell (pp. 163, 168, &c.).

Fic. 12.—C. ramosa.—“ Suppressed’”’ ovicell (p. 165).

Fie. 18.—C. ramosa.—Internode which has abnormally developed four
ovicells (p. 166).

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LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 183

The Later Larval Development of Amphioxus.
By

Arthur Willey, B.Sc.
(Of University College, London).

With Plates XIJI—XV.

In the account of the development of the atrial chamber of
Amphioxus which appeared in this journal last August, under
the joint names of Professor Lankester and myself, it was
stated that we had not been able to trace the progressive steps
in the history of the gill-slits, during their passage from the
unilateral position which they hold in the larva, to the sym-
metrical condition which we find in the adult; nor the trans-
lation of the lateral mouth of the larva to the anterior median
position of the mouth in the adult.

We added that we had taken steps to obtain the critical
stages in the living condition during the summer of last year
(1890). I accordingly went down to Messina, for the second
time, in July, to undertake the study and to make drawings of
the living larve—being enabled to do so by the continued kind-
ness of Professor Lankester, to whom for this, as also for invalu-
able assistance and advice since received, I am deeply indebted.

According to Professor Lankester’s instructions I spent the
months of July and August at Faro, daily fishing in the
Pantano, and concentrating my attention on larve of the
particular stages which were required. To obtain a large
number of individuals in the desired condition of transition,
from the extreme asymmetry of larval life to the nearly sym-
metrical condition which is reached with scarcely appreciable in-
crease of size, was no easy matter. By examining daily a great
number of larve I succeeded in obtaining an ample series of the

184 ARTHUR WILLEY.

transitional forms, which were carefully drawn and described as
observed in the living condition. These observations form the
substance of the present memoir. I may possibly find it desir-
able to add to this hereafter an account of some of the struc-
tures involved, as determined by means of sections. For this
determination I have an ample supply of preserved material.

Hasits oF THE LARVA.

The conditions under which the material was obtained last
year differed in a curious way from those of the preceding year.
In 1889 the larvee of Amphioxus were present in the lake at Faro
in great numbers, especially during the months of July and
August; while last year, during the same months, they were
comparatively rare, and their place seemed to have been taken
by an incredible host of Doliola of the first or larval generation.

Exactly in what way the presence of vast numbers of
Doliola affected the larve of Amphioxus I was not able to
make out, because the spawning of Amphioxus was prolific in
the extreme. Possibly the breeding of Amphioxus as a
whole commenced rather earlier last year, and the larve may
have taken to the sand mainly before July. I found numerous
Doliola with ova and gastrulz of Sagitta in their pharynx, some-
times as many as nine ova in one Doliolum. This might lead
to the death of the embryos in question, as it appears to cause
the death of the Doliolum. I did not, however, find the ova of
Amphioxus in this position, or possibly only in one or two doubt-
ful cases. At all events, the scarcity of the larve of Amphioxus
at this particular time of the year was evidently in correlation
with the great predominance of Doliolum.

Professor Kleinenberg informed me of an analogous fact,
namely, that Salpze are sometimes very numerous in the harbour
of Messina, to the exclusion of other small pelagic organisms.

When placed in glasses containing fairly clean water the
healthy larvee of Amphioxus are seen to be suspended, appa-
rently motionless in the water, in a highly characteristic
vertical position. The suspension is no doubt effected by the
movement of the long cilia with which the epidermis is provided.

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 185

It will be remembered as a curious circumstance that the adult
habitually assumes a similar vertical position in the sand.
The time of life at which the larva seeks its home in the sand
varies greatly. While, on the one hand, I have dredged all the
stages described below pelagically, I have, on the other hand,
found larve which had only reached the third stage of the later
larval development (see below ; also Pl. XIII, fig. 4) in the sand.

RésuME oF THE Entire DeveLopmMEent or AMPHIOXUS.

The various phases in the development of Amphioxus may
be conveniently arranged in the following way :

I. Tue Pertop or Empryonic DEvELopMENt comprising the
first thirty-two hours. It commences with the segmentation
of the ovum, and ends with the formation of the mouth and
first gill-cleft. According to the habit of the embryo, this
period may be subdivided into—

(a) The time—namely, the first eight hours—during which
the rapidly developing embryo is confined within the vitelline
membrane, the successive stages being marked by the progress
of the segmentation and gastrulation, the commencement of
the formation of the myoceelomic pouches, the differentiation of
the medullary plate, and the formation of the neurenteric canal.

(6) The time which elapses between the emergence of the
ciliated embryo from the vitelline membrane, and the appearance
of the mouth, first gill-cleft, and anus—the stages being marked
by the successive formation of myocelomic or archenteric
pouches to the number of fourteen pairs. The myotomes

1 The accounts of Amphioxus given by Rathke (K6nigsberg, 1841) and J.
Miller (Berlin, 1844) have given rise to the impression that the usual status
quo of Amphioxus is, to be lying on its side on the sand. As stated above,
this is not the case. If it were so, it might be supposed to offer a simple
explanation of the one-sided character of the larva—not that I think it would,
however. J. Miiller, indeed, says that Amphioxus is fond of burying the caudal
half of its trunk in the sand, as an occasional divertissement. The lying
on one side, however, is what is occasionally done; and that is the result,
not so much of a subtle inherited tendency to assume that position of rest, as
of a gross incapacity on the part of Amphioxus to maintain its equilibrium in
any other way when out of the sand.

186 ARTHUR WILLEY.

which are added after this period never communicate with the
intestine (Hatschek).

II. Toe Pertop or Earty Larvat DrveLorment during
which fresh gill-slits appear one after the other—i. e. meta-
merically—slightly to the right side of the median line (sub-
sequently passing well up to the right side), to the number of
twelve to fifteen. Towards the close of this period the longi-
tudinal metapleural folds appear, and the closure of the atrium
commences behind by the fusion of the small subatrial ridges
which are developed on the inner faces of the metapleura.

III. Tue Periop or Later Larvat Devetopment during
which the second row of gill-slits is formed on the right side ;
the first or primary row of slits passes across to the left side,
the mouth assumes an anterior median and vertical position,
the preoral cirri appear, and the endostyle is developed from
its pre-existing rudiment.

IV. Tue Avotescent PEeRiop when the young Amphioxus,
having now attained most of the essential features of the adult
structure, has definitely ceased to lead a pelagic life, and has
taken up its abode in the sand, where its further growth in size
and maturity is accomplished.

Of the above four main periods in the development of
Amphioxus, the first has been studied by Kowalewsky (2) and
more thoroughly by Hatschek (8) ; the second has been treated
by Professor Lankester and myself (8), and the third forms the
subject of the present paper. The fourth entails the considera-
tion of the adult anatomy, for which Professor Lankester’s
paper (5) may be consulted.

The only observations recorded as to the course of events in
the third period were made by Kowalewsky (1), whose all too
brief account was considered so extraordinary that Balfour! was
“tempted to suppose that his observations were made on patho-
logical specimens.”

Kowalewsky confined himself wholly to the question of the
origin of the second row of gill-slits, and with reference to
that he said that six disc-like thickenings appear together on

1 *Comparative Embryology,’ vol. ii, 1885.

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 187

the right side above the row of primary slits. The secondary
thickenings then become perforated and increase in size, and
as they do so the primary slits pass under the pharynx, and so
to the left side.

In his figures he places the first secondary slit immediately
over the second primary. This is not correct. He did not
describe the formation of additional secondary slits, and he did
not observe the closure of any of the primary slits.

He was right as to the number of secondary slits which first
appear together, and as to their formation on the right side
above the primary slits, with the consequent moving of the
latter round to the left side.

My observations on the gill-slits, therefore, in the first place
confirm, in the second place correct, and in the third place add
to Kowalewsky’s description. Over and above this, the forma-
tion of the velum, the origin of the buccal or preoral cirri, the
fate of the club-shaped gland, and the development of the
endostyle will be found fully described below.

In the following account the later larval development will,
for convenience, be divided into eight stages. The description
of the separate stages will be followed by a brief summary of
the stages, and this by a further summary of the facts relating
to the various organs; the paper will be concluded with a few
general considerations.

Stage I.—Figs. 1, 2, 20, and 21.

This is the stage immediately succeeding the older of the
two stages figured in the paper referred to above (8), and the
primary gill-slits and metapleurs present very much the cha-
racters there described. Thus there are fourteen primary
slits (fig. 1), most of which open directly to the exterior ; but
the last three or four discharge into the atrial tube, which,
while closed behind, is widely open in front, where the right
metapleur is seen to overhang the anterior slits.

The new feature with which we have now to deal is the
appearance of a continuous ridge in the right wall of the
pharynx above the row of primary slits.

188 ARTHUR WILLEY.

On this ridge are developed typically six oval thickenings or
enlargements, alternating with the primary slits and appearing
simultaneously. They consist of a fusion of the pharyngeal
wall with the body-wall at six different points, the first point
lying above and between the third and fourth primary slits.
They are the forecast of the second row of slits. Between the
ridge with its nodal thickenings and the row of primary slits
a longitudinal blood-vessel is seen to hold its course; this
becomes eventually the ventral or subintestinal vessel which
lies beneath the endostyle (see Lankester, 5).

The first and last thickenings, as Kowalewsky also noticed,
are usually rather smaller than the intervening ones.

Shght deviations from the above mode of procedure occur.
One of these is shown in fig. 2, where only five secondary
thickenings have appeared at once, the first of them occurring
between the fourth and fifth primary slits, while the one which
usually has the first place at this stage has been somewhat
retarded in its development.

In other cases, in which there are also only five thickenings,
it may be the sixth that is late in appearing.

In the larva represented in fig. 2 there were only twelve
primary slits. As mentioned above, the number of primary
unpaired slits which are formed in the larve varies from twelve
to fifteen. The most usual number is perhaps fourteen, while
fifteen is exceptional.

In front of the gill-slits is seen the club-shaped gland,
closely apposed to the remarkable patch of modified hypo-
blastic epithelium which, in the paper by Professor Lankester
and myself (8) describing the larva, was termed the glandular
tract. The nature of this organ has been enigmatical up to
the present time ; but I may as well at once call it the
“ endostyle,”’ since it certainly becomes that organ.

The opening of the club-shaped gland into the cavity of the
mouth—i. e. the intra-buccal orifice—lies at this stage slightly
dorsal, and at a later stage quite dorsal, to the endostyle.
This position of the internal aperture of the gland is important,
as will appear in the sequel.

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 189

On the left side (figs. 20, 21) the large lateral mouth is seen
with its margin ciliated. A ciliated groove leads from the
preoral pit to the antero-dorsal margin of the mouth. Ante-
riorly the mouth ends in a sharp point; but in fig. 21, on close
inspection, a very slight protuberance can be seen in the edge
of the mouth projecting into the ciliated groove. This minute
protuberance is the first indication of the combined forward
and transverse growth by which the mouth moves from a
lateral to an anterior median position.

Just below the mouth in front is the external orifice of the
club-shaped gland, and some way behind this (fig. 20) is seen
a small circular piece of homogeneous tissue, which is really
a differentiation in the mesoblast of the lower lip of the mouth,
showing through the outer integument, and is, in fact, the
first element of those peculiar cartilaginoid structures which
form the skeleton of the preoral cirri. In fig. 21 there are
two such elements, and they go on increasing in number by
additions at both ends.

Ruuuing below the mouth, and then bending up to assume
a dorsal position, is a ciliated band composed of columnar
hypoblastic cells, forming a slight ridge in the pharyngeal
wall. It is continuous in front with the base of the lower arm
of the endostyle. On the other side there is a similar band
running from the upper arm of the endostyle, but it cannot be
seen in surface views at this stage.

In these, and all drawings of the left side, the anterior
aperture of the nerve-tube described by Hatschek (3 and 4) is
shown very clearly.

Stage II.—Figs. 3, 22, 23, and 24.

The distinction of this stage lies in the perforation of the
secondary thickenings. The second to the fifth inclusive of
the thickenings become perforated, as a rule, before the first
and sixth; and, again, the first is usually perforated rather
sooner than the sixth. This order, however, is subject to
frequent exceptions. In fig. 3, for instance, the sixth secondary
slit is open, but the first is not.

190 ARTHUR WILLEY.

The apertures of the secondary slits are at first extremely
small, and appear as dark spots, with transmitted light, in
which long cilia are to be seen working.

In fig. 3 it will be noticed that a seventh secondary
thickening has been added behind, but it frequently does not
appear before the next stage.

In the same figure there are fourteen primary slits, but the
fourteenth is only indicated in side view by a median de-
pression in the floor of the pharynx. This means that the
slit isin process of closure, as I found by occasional ven-
tral views, which were obtained through the struggling of the
animal when placed between slide and cover-glass. In this
way the larva frequently got on its back, and became fixed in
that position by the slight pressure of the cover-glass.

The endostyle and club-shaped gland present the same
features as in the preceding stage.

The metapleura are also in much the same condition; but it
may be noticed that while in fig. 1 the right metapleur con-
curs with the left about the region of the ninth and tenth
primary slits, in fig. 3 it does so in the region of the seventh
primary slit, thus indicating that the closure of the atrium
has advanced forwards.

A view of the left or oral aspect of the larva (figs. 22—24)
shows that in this stage the anterior extremity of the mouth
becomes no longer pointed. This is due to a continuation of
that hunching up of the antero-dorsal margin of the mouth
into the ciliated groove, of which we saw the first indication in
Stage I. It is ultimately carried to such an extent as to
entirely change the original shape of the mouth. Meanwhile
the anterior portion of the mouth sinks inwards deeper and
deeper towards the other side of the body, so that it has to be
examined with a deeper focus than in Stage I.

The upper part of the oral hood commences to form simply by
the continued growth downwards of the upper margin of the pree-
oral pit and ciliated groove ; while the lower part, in which alone
the buccal cirri have their origin, arises independently beneath
the under lip of the mouth. Figs. 22—24, besides showing the

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 191

gradual change in the shape and level of the anterior half of the
mouth, show also the progressive increase in the number of the
elements of the buccal cirri—which, as they grow in number,
grow also in size, and soon becoming irregular in their contour,
show signs of growing out into tentacle-like processes.

I append a list of half a dozen observations on different larvee
at this stage, to show the nature and extent of the variations in
the condition of the secondary slits. In all cases the first slit
or thickening was above and between the third and fourth
primary slits.

Number of secondary Number of Thickenings not
slits or thickenings. open slits. perforated.
@)-< ‘ 6 : » Nos. 2 ta 5 -:. . Nos. 1 and 6
(ii) . 6 EG »» land 2
Giese Ge me eT ees No. 6
(iv) . : 7 : She gel et ea Nos. 1, 6, and 7
(v) . 7 ITI 'G » land7
(vi) . 5 lee B No. 1

In the second row of the above table the numbers are
inclusive.

It is worth noting here that the first primary slit is distinctly
smaller than the second and following slits. If reference be
made to the figure of the larva with three gill-slits given in the
paper previously quoted (8) it will be found that the first slit
is larger if anything than those which follow it. Ata certain
stage its growth is arrested, and later still, ina most remarkable
way, it not only becomes relatively but actually smaller in size.

Stage III.—Figs. 4 and 25.

In a larva of this stage the closure of the atrium has extended
forwards so as to leave only a small portion unclosed in front, so
that in this and all subsequent stages the gill-slits are entirely
seen through the transparent wall of the atrium.

In fig. 4 there are twelve primary slits ; the usual number
at this stage is, however, thirteen. The secondary slits are
seven in number; the first is circular in outline, and has just
opened ; while the seventh is only present as a thickening not
yet perforated, and is also circular. The other secondary slits

VOL. XXXII, PART II.—NEW SER. N

192 ARTHUR WILLEY.

are more or less elliptical in shape; but it will be noticed that
the third and fourth, which are the largest, are shaped like a
plano-convex lens, their tops being flattened.

The increase in size of the secondary slits is accompanied by,
and in fact is evidence of, a transverse growth by which the
primary slits are gradually taken round to the left side.

The hindermost primary slits are always bent under the
ventral wall of the pharynx; but in fig. 4 this bending under
extends to the more anterior gill-slits, namely, up to and
including the fourth primary slit. This should be compared
with figs. 1, 2, and 3.

The first primary slit is now smaller than we have hitherto
seen it. The twelfth is very small, but when seen in ventral
view does not yet show the definite signs of closing which will
be described later.

The anterior wall of the mouth (fig. 25) has now sunk in or
bent round so far that it can be easily seen through the right
body-wall at this stage (fig. 4). This part of the mouth becomes
the right half of the oral sphincter (velum of Huxley and
Hatschek). As the mouth sinks towards the right side in this
way, so also does the preoral pit; and the latter gradually
becomes flattened out as the development of the oral hood pro-
ceeds, and eventually becomes simply a ciliated tract on the
under side of the oral hood, known in the adult as the “ Rader-
organ,” which has been identified by Hatschek with the preoral
pit of the larva.

The club-shaped gland and endostyle are approximately in
the same condition as in the preceding stages, but in some
larve of this stage I detected signs of the backward growth of
the endostyle, in that the club-shaped gland was frequently
seen to overlap the posterior edge of the endostyle.

Of the six secondary slits which have been described above
as‘appearing at the same time, the first, which nearly always
lies between the third and fourth primary slits, does not
eventually become the first secondary slit, but it becomes the
second, a new one being formed in front of it. The latter,
however, occasionally appears as soon as the others. Thus

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 193

when all the secondary slits! are established, the one that stands
first in position is not the first formed, but arises rather later
than the six slits which follow it. The usually late appearance
of the first secondary slit is possibly correlated in some way
with the complicated growths which are taking place in the
anterior region, and is not of any special significance ; while, on
the other hand, the simultaneous appearance of the greater
number of the secondary slits is of considerable importance, as
will appear later.

To bring the complicated nature of the transformation which
is being effected vividly before the mind, it may be noted that,
while the mouth is moving forwards, the endostyle is growing
backwards, and the primary gill-slits are crossing bodily over to
the left side.

The oral aspect of a larva at this stage (fig. 25) presents no
particularly new features, but merely an extension of the pro-
cesses described in the preceding stage as being in operation ;
thus the anterior portion of the mouth is seen at a still
deeper focus, the apparent length of the aperture of the mouth
when seen from the side has decreased, and the buccal
skeleton has further advanced in development.

The following table comprises a few observations chosen out
from a number to show some of the variations that are met
with in this stage :—

| ; eee
Ieeegadary.alite eke PeleLouines Position of first secondary slit
| or thickenings. open slits. perforated. or thickening.
6 Nos. 1 to 6 — Between 3rd and 4th primary.
ay 6 2) 1 ” 5 No. 6 9 ”
Ml) 5 »” bl ”» 4 ” 0 ” 9
iv) 6 256 wal » 2ndand 8rd primary.
% if pte ee x0 Nos. 1 and 7 es ss
vi 9 Spee ee ok BUG AUE gee Le say 8 » 3
vil 5 so asad — » 4thand 5th primary.

1 The primary slits form, as has been said, to the maximum number of
fifteen. The secondary slits never normally exceed the number of nine. By
far the greater number of the gill-slits of an adult Amphioxus must, there-
fore, be distinguished by the name of tertiary slits. The reason of this will
become clear as we proceed.

194, ARTHUR WILLEY.

This table will be intelligible if one compares what has been
said above with reference to the first secondary slit with the
position of the latter as given in the fourth column. In
No. 7 it will be noticed that the first two secondary slits
were late in appearing. No. 6 in the above table was
obviously aberrant, but is interesting as exhibiting a hastening
of the development of the full number of secondary slits. It
was also slightly abnormal, since, although the ninth slit was
well open, the eighth was only present as a thickening.

In all cases the eventual first secondary slit arises above and
between the second and third primary slits.

The trifling confusion which may attend this description is
unavoidable. The straightforward course of events, unham-
pered with variations, will be given in the summary.

Stage 1V.—Figs. 5, 6, and 26.

This is a well-marked stage, characterised by a general
increase in size of the secondary slits, accompanied by the
dipping downwards or bending inwards of the dorsal wall in
the largest of them—namely, Nos. 3 to 5 inclusive. The
fusion of the down-growth from the dorsal wall with a small
up-growth from the ventral wall of the slit (which, however,
does not occur in this stage) results, as is well known from
the figures of _Kowalewsky and others, in the formation of the
so-called tongue-bar of the slit.

The bending under of the primary slits from the right to
the left side has now proceeded much farther; and the first
primary slit is very much reduced in size. In the next stage
we shall see that it closes up and ultimately disappears without
leaving a trace.

In fig. 5 the thirteenth primary slit is seen in course of
closure—that is to say, in a ventral view it would present the
appearance shown in the case of the twelfth slit in figs. 9 and
10. There are seven secondary slits, the first being between
the second and third primary slits.

The atrium is now completely closed anteriorly.

The most striking and unforeseen characteristic of this

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 195

stage is that the patch of modified epithelium which we have
already spoken of as the endostyle has definitely commenced
to push its way past the club-shaped gland, so that the latter
now lies upon instead of behind it. This can also be expressed
by saying that the endostyle has begun to fall away from its
previously oblique position, so as ultimately to assume a longi-
tudinal horizontal and ventral position. The incipient change
of position of the endostyle is not accompanied in this stage
by any growth in its length. The movement in bulk of a patch
of modified epithelium from one position to another is suffi-
ciently remarkable. It is probably, however, effected by the
same transverse growth which affects the primary gill-slits.
A consideration of the figures will show that it would be quite
possible for a growth of this kind to have the effect of
dragging down the at first very oblique endostyle.

It is not so much the primary obliquity of the endostyle, as
its primary anterior position in the region of the first myo-
tome, which should be especially noted.

A view from the left side of a larva of this stage is given in
fig. 6, the object being chiefly to show to what extent the
primary slits have come round to this side. The first primary
slit is hardly recognisable from this side, but can just be seen
with a median focus. In this larva there were fourteen
primary slits; but the more usual number for this stage is
perhaps thirteen, and, moreover, the thirteenth is usually in
course of closure.

The oral hood, both as to its dorsal and ventral halves, is
now well marked, and the cartilaginoid elements of the buccal
cirri, of which there are six, are growing out into distinct ten-
tacles, which give a crenate margin to the lower half of the
oral hood. The external orifice of the club-shaped gland is
undoubtedly present, but is covered over by the buccal cirri.

The apparent length of the mouth is rather greater in fig. 6
than is usual at this stage, and in this respect the condition
of the mouth represented in fig. 26 is much more typical; a
glance at this drawing will make it possible to understand
that, while the contour of the mouth undergoes a complete

196 ARTHUR WILLEY.

alteration in form, the actual size of the mouth does not
materially change; though in the adult, of course, the relative
size of the mouth or velum is much less than in the larva.

The variations in the condition of the secondary slits at this
stage are not sufficiently striking to make it necessary to
tabulate them. Their number, as a rule, varies between seven
and eight; and almost invariably the first secondary slit
appears at this stage between the second and third primary
slits (fig. 5).

Stage V.—Figs. 7—10 and 18.

There is a considerable difference between the condition of
a larva at the beginning of this stage and at the end of it, as
will be seen by comparing fig. 7 with fig. 10. The constant
peculiarity for the stage is, that although the ‘primary slits
have not quite attained their final position on the left side,
yet the tongue-bars have commenced to grow down from the
dorsal borders of the slits, but do not meet the ventral borders
during this stage. There is also a feature which is more
obvious in the preceding stages, and which we see for the last
time in the present stage. I refer to the fact that the long
axis of the secondary slits is at this and the foregoing stages
parallel to the long axis of the body, and at right angles to
the long axis of the several primary slits. This is the case
even in fig. 10, where, however, one would probably fail to see
it at a glance. It is important in that it is characteristic of
the earlier stages in this period of the development.

In fig. 7 a view is given of the right side of a larva which
had just entered on this stage. There are eight secondary
gill-slits, the first, as one would expect, being between the
second and third primary slits. The tongue-bars, as we have
already seen, are in course of formation, and that of the third
secondary slit has actually fused with the ventral border of
the slit. Unfortunately in the larva here figured (fig. 7) the
endostyle is not typical for this stage, being in the condition
described in Stage IV. A characteristic endostyle is, however,
shown in fig. 18. It has become much more horizontal in

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 197

position, and has grown a long way past the club-shaped
gland. The latter is still present with its large intra-buccal
orifice, but when seen in the living animal it seems to present
signs of disintegration, the constituent cells assuming a
loosely aggregated and turgid appearance preparatory to dis-
solution. The intra-buccal orifice is apparently the most
persistent part of the gland. In fact, before the end of this
stage the club-shaped gland atrophies altogether. From
observations which I made on numerous larve belonging to
this stage, I am inclined to believe that the cells of the gland
break away from each other and pass into the alimentary
canal, where they are possibly absorbed.

The ciliated hyperpharyngeal band of the right side, which
has been referred to above, can now be seen proceeding from
the upper arm of the endostyle ; while the corresponding band
of the left side proceeds from the lower arm of the endostyle.
These upper and lower portions become, at a later stage,
respectively the right and left halves of the endostyle; and,
indeed, the latter—namely, the lower half—when looked at
from the right side, is found to lie at a deeper focus than the
right or upper half.

One of the most curious events of this stage is the closure
of the first primary slit, which occurs very shortly after, if not
at the same time as, the atrophy of the club-shaped gland.
In fig. 7, which is a drawing of a younger larva than those
represented in figs. 8, 9, and 10, the first primary slit can just
be seen in side view ; on the other hand, in the larva of which
a three-quarter ventral view is given in fig. 9 it could not
have been distinguished at all in side view ; but a ventral view
showed it in the condition of a minute aperture with cilia
working in it, surrounded by cells in such a state of aggrega-
tion as to present a coarsely granular and dark appearance,
which, combined with the loss of a sharp outline to its wall, is
the distinctive feature of a closing slit, and enables it to be
placed in marked contrast to a newly formed slit, of which the
wall is clear and refringent.

In fig. 10 there is the merest trace of the first primary slit

198 ARTHUR WILLEY.

just in front of the one marked “second primary slit.” In
fig. 8 there are indications of fourteen primary slits; the
twelfth is, however, very small, and the thirteenth and four-
teenth are on the verge of closure.

In both fig. 9 and fig. 10 the twelfth primary slit is shown
in course of closure.

Fig. 10 is quite an exceptional view of a larva from the
ventral or ventro-lateral aspect, obtained by the larva getting
fixed on its back between slide and cover-glass, as a result of
its struggles to get free. Such a complete view as is here
represented can only be got very rarely, and when it does
happen it lasts but a very few minutes, as the larva either
speedily rights itself or dies.

The shape of the primary and secondary slits has been
already described. The tongue-bars of most of the latter have
fused with the ventral borders of the slits, but not one has
done so in the case of the primary slits. It should be noticed,
also, how the row of primary slits curves round to the middle
ventral line behind ; the twelfth is nearly closed, the eleventh
would close later on, while the tenth might or might not close
later: judging from the shape and size of it, one may say
that the balance of probability is in favour of its not closing,
In some of the primary slits there is a small up-growth from
the ventral border of the slit, pointing towards and subse-
quently destined to fuse with the down-growing tongue-bar.
All the slits are of course seen through the transparent
atrial wall. The endostyle is seen anteriorly between the
primary and secondary slits, and the buccal cirri are also well
shown.

The mouth of this stage is shown in fig. 8. The lower por-
tion of the oral hood, carrying the buccal cirri, is now more
or less continuous with the upper portion into which the cirri
have not yet extended; but, at the point of junction, a ridge
is formed under which the cartilaginoid tissue eventually forces
its way (cf. figs. 12, 13). The upper and lower portions of the
oral hood become respectively left and right.

Thus, while the right and left halves of the oral hood are

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 199

independent of one another in their origin, the elements of the
buccal skeleton arise entirely unilaterally from a single differen-
tiation of mesoblast, which grows at both ends, and is situated
primarily in the right or lower half alone of the oral hood, and
secondarily continues itself into the left or upper half.

The lower edge of the oral hood is prolonged anteriorly as a
ridge, into which the buccal skeleton is subsequently con-
tinued.

With regard to the variations met with in this stage, it is
only necessary to say that the number of secondary slits varies
from seven to nine ; and as to the primary slits, it is usual to
find the twelfth, sometimes also the thirteenth, on the point of
closing ; but I have found an instance in which, although there
were nine secondary slits and the club-shaped gland was
beginning to atrophy, yet there were fifteen primary slits, all
in a healthy condition, with the exception of the first, which
was closing.

Stage VI.—Figs. 11, 12, and 19.

The constant characteristic for this stage is that the primary
and secondary gill-slits are of equal size and similar shape for
the most part ; and, excluding the first two and the last two on
each side, they are approximately as broad as they are long.
Further, the tongue-bars in most of them have fused with the
ventral borders of their respective slits.

The first primary slit has entirely gone, and it is usual to
find the eleventh primary slit, together with the twelfth and
sometimes also the thirteenth (figs. 11 and 12), in a state
bordering on closure. All the slits of the left side are now, as
a rule, entirely confined to that side throughout their whole
extent (cf. fig. 8), with the exception of the ninth, which
bends under ventrally. The first slit of the left side in this
and all succeeding stages is the original second primary slit ;
and similarly the ninth slit of the left side is the original tenth
primary, and so on for the other primary slits.

Owing to the atrophy of the first primary slit, the first slit
of the right side—i. e. the first secondary slit—comes to lie

200 ARTHUR WILLEY.

opposite to the boundary between the first and second slits of
the left side—and this is its final position.

Except when the above-mentioned ninth slit is found in an
undoubtedly rudimentary condition, it is impossible to predict
whether it would eventually close or not. My observations
show that in the majority of cases it does close, but that, on
the other hand, it frequently does not. I have never found a
less number of primary than of secondary slits, so that when
nine of the latter are formed (of which instances have been
given above) it is certain that nine of the primary slits will
persist.

On comparing fig. 12 with fig. 11 it will be found that the
gill-slits of the former are not quite so far advanced as those of
the latter, while the buccal cirri in the former have reached a
higher stage of development than those of the latter. This is
a slight variation.

The junction of the originally independent upper and lower
portions of the oral hood is very distinct.

The anterior or right wall of the oral sphincter or velum has
now passed so thoroughly round to the right side that it
cannot be inserted in a drawing of the left side. Four velar
tentacles make their appearance during this stage; they are
much clearer, however, in the next stage.

The left ciliated (hyperpharyngeal) band is seen to join the
left or lower half of the endostyle ; the right or upper half of
the endostyle is not seen from this side naturally.

In all the drawings of larvz belonging to previous stages,
from the left side, there is shown a peculiar structure, de-
scribed as a nephridium by Hatschek (4), and placed just
below and to the left side of the notochord in the region
between the preeoral pit and the mouth.

It presented the appearance represented in the figures, but
I could not certainly detect cilia in it, and in fact was unable
to understand its import. It seems to possess a superficial
resemblance to the head-kidney of Annelid larve (trocho-
spheres), but I can form no opinion as to the reality of any
such resemblance. Hatschek discovered that it opened at one

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 201

end into the cavity of the mouth. (See also 8, pl. xxx,
figs. 2—4..)

It requires, and would probably repay, further study. From
the present stage onwards I could not recognise this so-called
nephridium in the larve. The structure marked “ 2” in the
figures is probably part of the ciliated tract of the preoral pit,
seen through the oral hood.

In fig. 19 a view of the endostyle is given. It does not
differ markedly from its condition in Stage V, extending about
the same distance backwards between the anterior secondary
and primary slits, but the absence of the club-shaped gland (of
which feeble remnants are still shown) has to be noticed.

The number of secondary slits in this stage varies again
from seven to nine.

Stage VII.—Figs. 13, 13a, and 14.

The slits—primary and secondary—have now begun to elon-
gate in a direction at right angles to the long axis of the body.

A view of the left side of a larva is given in fig. 13. The
first slit on this side (i.e. the original second primary slit) is
simple and remains simple in the adult, while all the others
become doubled by the formation of the tongue-bars.

The ninth slit of this side is very small and lies at a rather
deeper focus than the other slits, so that in this case it would
in all probability close. Behind the ninth (i.e. the tenth
primary) is seen a rudimentary indication of the eleventh
primary slit.

A separate view of the velum with the four velar tentacles
is given in fig. 13a. Fresh tentacles arise between the four
primary ones at a much later stage, when the gill-slits are
about three times as long as they are now, and the endostyle
twice as long. The final number of velar tentacles is twelve.

The right wall of the velum is not quite opposite to the left
wall even yet: it is slightly in front of the left wall, and the
extent to which it is so corresponds to what has been hitherto
spoken of as the apparent length of the aperture of the mouth.

On the right side (fig. 14) it is usual to find eight slits. The

202 ARTHUR WILLEY.

ciliated tract of the preoral pit is now beginning to alter its
shape—becoming constricted in the middle. This constriction
is carried still further in the next stage (fig. 15), and, as is
already known, the ciliated tract subsequently grows out into
several lobes, which give its characteristic appearance to the
Rader-organ.

The most important feature in fig. 141s the endostyle, which
has increased considerably in length, reaching to the fourth
slit. Its double nature is very apparent; the left (lower) half
lying, as before mentioned, at a much deeper focus than the
right (upper) half, so that the former only is seen from the
left side (fig. 13).

In the larva represented in fig. 14 there were eight slits on
the left side, and the ninth was seen to be in course of closure
in the mid-ventral line.

We are now approaching the period in the larval develop-
ment in which there are an equal number of slits on both sides
of the body, all those on the left side being primary and all
those on the right side secondary.

After a long interval, during which the gill-slits increase in
vertical height and the endostyle extends further backwards,
fresh tertiary gill-slits form behind on each side in a manner
characteristic of later growth, and continue to do so through-
out life.

The stage, then, with an equal number—seven to nine pairs
—of primary and secondary slits, and before the formation of
any tertiary slits, may well be called ‘‘the Critical Stage.”!

As a general rule the critical stage is characterised by the
presence of eight pairs of slits, namely, eight primary slits on
the left side and eight secondary slits on the right.

Stage VIII.—Figs. 15—17.

During this stage the critical stage, as defined above, is
firmly established, and in fact may be said to commence as soon

1 The remarkable fact that the number of pairs of gill-slits in the critical
stage agrees approximately with the full typical number of the craniate gill-
clefts should not be lost sight of.

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 203

as the fate of the tenth primary slit has been decided. In
fig. 16 the ninth slit has not perfectly assumed its position
on the left side, and so its permanence cannot be considered
as being beyond question, although its size and appear-
ance would lead one to think that it would persist and not
close.

The length or vertical height of the gill-slits has further
increased, and has been taken as the distinguishing feature of
this stage.

Fig. 17 represents the last (seventh) pair of slits of a larva,
in which not only had the tenth primary slit closed, but the
ninth was alsoon the point ofclosing. The seventh slit of the
right side is seen with a surface focus, the ninth rudimentary
primary slit is seen with a median focus, and the seventh slit
of the left side, which is doubling in, is seen with a deep focus.

The mouth or velum (fig. 16) is now anterior and median in
position ; the right and left walls of it are quite opposite to one
another, and thus the apparent length of the mouth in side
view is now reduced to zero.

The oral hood and buccal cirri have not yet, however, attained
their final condition.

The endostyle (fig. 16) has greatly increased in length,
extending to the fifth gill-slit on this side.

The right side of another larva of this stage is shown in
fig. 15, chiefly for the sake of the endostyle. This is the last
stage at which its double origin can be recognised. The right
half of the endostyle is now seen to lie at the side of, and not
above, the left half.

The ciliated hyperpharyngeal band appears to die out behind
the pharynx. On this side there are eight (secondary) slits:
on the left side of this larva there were also eight (primary)
slits, and no indication of any more, so that it is a typical
example of a larva which has entered upon the critical
period.

It is important to emphasise the fact that the critical period,
in which the larva has usually eight pairs of slits, has a con-
siderable duration, in the course of which the only changes

204. ARTHUR WILLEY.

that take place are the completion of the oral hood and the
increase in the size of the gill-slits and endostyle, but no fresh
slits are formed.

The close of the critical period may, therefore, be taken as the
close of the entire larval phase of development. Up to this
point all the gill-slits of the left side are primary—that is to
say, they appeared at first as asingle series on the actual right
side of the larva, and were pushed across to theleft side by the
developing secondary slits. We shall find reason to suppose
that the actual right side of the larva is not precisely identical
with the morphological right side.

Summary of the Stages.

In the following summary it will be found desirable to give
a few measurements, which, however, have only an average
value, the unit employed being a hundredth of a millimetre.
It is also intended, for the most part, to ignore the variations
which have been detailed above, and to state only what
appears to be the usual condition of things, as inferred from
observations on numerous larve.

Stage I. Right Side.—Fourteen primary slits, none
closing ; six secondary thickenings above the primary slits ;
endostyle in front of club-shaped gland, and therefore in front
of all the gill-slits ; atrium widely open anteriorly.

Left Side.—Large lateral mouth ; apparent length (in this
case the actual length), 45 units; 1 to 2 elements of buccal
skeleton.

Stage II. Right Side.—Fourteen primary slits, four-
teenth closing ; secondary thickenings perforated; endostyle
unaltered ; atrium somewhat less widely open anteriorly.

Left Side.—Lateral mouth beginning to bend round in
front ; apparent length, 36; 3 to 5 elements of buccal skeleton.

Stage III. Right Side.—Thirteen primary slits; se-
condary slits simple ; endostyle unaltered ; atrium slightly open
anteriorly.

Left Side.—Mouth still further bent round in front;
apparent length ranges from 28 to 12, average about 25; 5 to6

LATER LARVAL DEVELOPMENT OF AMPHIOXOUS. 205

elements of buccal skeleton commencing to grow out as
tentacles or cirri.

Stage IV. Right Side.—Thirteen primary slits, all bend-
ing under the pharynx, first very small and thirteenth closing ;
seven secondary slits, the new one appearing in front; the
larger secondary slits commencing to double in, i.e. to form
tongue-bars; endostyle extending a short way beyond the
club-shaped gland; length of endostyle, 24; atrium closed
anteriorly.

Left Side.—Primary slits are now partly on this side;
average apparent length of mouth, 17 ; oral hood containing in
its lower half incipient cirri.

Stage V. Right Side.—Twelve primary slits just visible
at base of pharynx, twelfth closing; the first primary slit
undergoes atrophy ; eight secondary slits with complete tongue-
bars in the larger ones; endostyle extends a long way past the
club-shaped gland ; length of endostyle, 28; club-shaped gland
undergoes atrophy.

Left Side——Primary slits not fully arrived on this side
yet; tongue-bars commencing in the primary slits; average
apparent length of mouth, 11 ; junction of upper and lower por-
tions of oral hood.

Stage VI. Right Side—Eight secondary slits, length
equal to the breadth, measuring about 1] units, both in a direc-
tion parallel to long axis of body and in a direction at right
angles to it; length of endostyle, about 34.

Left Side.—Ten primary slits (i.e. Nos. 2 to 11 inclusive),
the eleventh closing ; tongue-bars complete in the larger slits,
which are equal in size to the corresponding secondary slits ;
mouth same as in Stage V; buccal skeleton commencing to
penetrate the upper portion of the oral hood.

Stage VII.—Slits on both sides commencing to elongate
in a vertical direction ; average length of the central slits, 14.

Right Side.—LHight slits; length of endostyle, 39.

Left Side.—Nine slits, ninth (i.e. tenth primary) usually
closing ; apparent length of mouth or velum, 4; four velar
tentacles.

206 ARTHUR WILLEY.

Stage VIII.—Average length of central slits, 21.

Right Side.—Hight slits; length of endostyle, 45. The
two halves of the endostyle are now definitely right and left.

Left Side.—Hight slits ; apparent length of velum, 0.

Of the above stages, the fifth is certainly the most eventful.
With regard to the last stage, it is well to be reminded that it
inaugurates the critical period; that at the commencement of
it there is often a rudimentary or doubtful primary slit behind ;
and, finally, that the presence of eight pairs of slits is not a
universal rule, as there are sometimes only seven and some-
times nine pairs, but never normally more than nine.

The central feature, perhaps, of the foregoing observations
is the development of the endostyle ; next in importance comes
the atrophy of the club-shaped gland and the first primary
gill-slit during Stage V, and of almost equal importance and
novelty is the atrophy of from four to six of the hinder slits of
the primary series.

It goes without saying that there is no hard and fast line
between any two consecutive stages. The division into stages
is entirely for convenience, all the processes of development
and transformation being gradual.

SUMMARY OF THE HiusToRY OF THE INDIVIDUAL STRUCTURES.
1. The Gill-slits.

The primitive or ancestral left row of gill-slits has, by a
process which will be discussed later, been made to assume a
position on the right side of the larva, where slits form to the
number usually of fourteen, but occasionally fifteen.

The actual perforation in the case of each slit occurs at first
at the base of the pharynx, near the mid-ventral line, and then
extends upwards on the right side; the hindermost slits, how-
ever, retain their ventral position all through.

The slits which thus appear at first in a single unpaired
series are called the primary slits.

The right row of gill-slits, which originally corresponded to
the primary slits before the latter had forsaken their primitive

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 207

situation on the left side, has suffered retardation of develop-
ment. When these retarded slits do appear, they are called,
arbitrarily, the secondary slits.

The secondary slits form to the number usually of eight
(sometimes seven and sometimes nine), above and alternating
with the primary slits on the right side, appearing long before
the completion of the atrium in front. The first secondary
slit always lies between the second and third primary slits, and
usually arises somewhat later than the succeeding six secon-
dary slits, which develop simultaneously. It will be shown
later that the simultaneous appearance of the secondary slits is
only due to a masking of the metameric mode of formation.

As the secondary slits grow in size the primary slits gradu-
ally pass round to the left side, and meanwhile certain of them
commence to atrophy. It is especially noteworthy that after
Stage I the first primary slit becomes progressively smaller and
smaller, until by the end of Stage V it has disappeared alto-
gether. The view will be supported below that the closing of
the first primary slit has a profounder significance than the
closing of the hinder slits, or at any rate a different meaning.

By the closing of some of the primary slits their number is
reduced to the same figure as that of the secondary slits,
whether it be seven, eight, or nine. The primary slits which
atrophy are, therefore, the following :

a. When 7 primary slits persist—Nos. 1 and 9, 10, 11, 12, 13, 14 close.
B. 3D 8 33 ” 32 1 > 10, nie 12, 13, 14 RK
Y: ” 9 ” ” ” iL ” 1a 12, 13; 14 oe

It is during the passage of the primary slits from the right
to the left side that the completion of the atrium is effected.

The long axis of the secondary slits is at first parallel to the
long axis of the body, while at the same time that of the
primary slits is at right angles to the long axis of the body.
The axes are equalised as soon as the two series of slits are
properly adjusted on their respective sides, and then the slits
on each side commence to elongate in a direction at right
angles to the long axis of the body.

VOL. XXXII, PART II.—NEW SER. )

208 ARTHUR WILLEY.

The formation of the tongue-bars commences very early in
the secondary slits, but not in the primary slits until they are
well round to the left side. The first slit on each side remains
simple, and does not form a tongue-bar.

As soon as all the primary slits that are going to close
have done so, the larva enters upon the so-called critical
period. At the end of this period fresh slits begin to form
normally on both sides, and continue to do so throughout life ;
these are called the tertiary slits in order to distinguish them.

Thus the critical period may be defined as the period in
which the slits are arranged regularly—an equal number of
pairs—in two rows, right and left; but all those on the left
side are primary (in the sense in which the term is here
employed), and all those on the right side are secondary.

2. The Endostyle.

The forecast of the endostyle appears in the embryo at a
very early stage, namely, shortly after the formation of the
club-shaped gland in front of which it lies. It consists of a
patch of modified columnar hypoblastic epithelium, bending
obliquely backwards and folded forwards upon itself, and
lying on the right side of the mouth-cavity in the region of
the first myotome.

The upper arm of the endostyle is at first much shorter than
the lower arm; the former becomes the right half of the adult
endostyle, and the latter becomes the left half.

The endostyle retains its original shape and its anterior posi-
tion until some time after the first appearance of the secondary
slits.

At Stage IV it begins to fall away, as it were, from its
oblique position—the evidence of this being found in the fact
that in this stage it extends a short distance beyond the club-
shaped gland, so that the latter, instead of lying behind it, lies
upon it (fig. 5). Although the endostyle has thus grown
backwards, it has done so in bulk—that is to say, it has bodily
shifted its position without increasing in length.

In Stage V, in addition to a further change of position, it

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 209

has commenced to grow in length (fig. 18), and now extends
between two or three of the anterior secondary and primary
slits.

In Stage VII (fig. 14) it has greatly increased in length, and
is quite horizontal and ventral in position ; but the upper and
lower (i.e. right and left) halves are still perfectly distin-
guishable.

In Stage VIII the two halves of the endostyle, which have
hitherto been in the relation to one another of upper and lower
—though the lower has been in the later stages at a deeper
focus than the upper—come to lie alongside of one another,
and so become definitely right and left respectively.

In fig. 16 the two halves can just be recognised in front
for the last time.

From the anterior extremity of each half of the endostyle a
ciliated ridge composed of the pharyngeal epithelium passes
dorsalwards, and becomes on each side the hyperpharyngeal
band.

The anterior portion of the ciliated bands, where they curve
dorsalwards from the endostyle, is undoubtedly to be compared
with the pericoronal ridges of Ascidians which pass from the
front end of the endostyle round the mouth to the dorsal lamina.

The following table shows the proportionate average increase
in the length of the endostyle, together with the concomitant
growth in length or vertical height of the gill-slits during the
critical period. The unit of measurement, as before is =4,
millimetre.

Length of central slits. Length of endostyle.
1 27 units : : : 52 units.
2 Sau ss : - 5Gns;
See ge Se . OU ORI “Gene
an : : ANE: : . 7Ga 3

The endostyle does not reach the posterior end of the pharynx
before the close of the critical period.

3. The Club-shaped Gland.
The development of this gland was made out by Hatschek (8).
To his description of it has been added the discovery of the

210 ARTHUR WILLEY.

intra-buccal orifice (8), and in the present paper the account
of its atrophy. It arises in an embryo with nine to ten pairs
of myoccelomic pouches, as a ventral transverse fold of the ali-
mentary canal at a point intermediate between the first and
second myotomes.

The fold extends up to the right side on the one hand, and
ventralwards round to the left side on the other. It then
becomes separated throughout its whole length from the
alimentary canal, and develops an opening to the exterior
on the left side, just below the lower margin of the mouth,
Its intra-buccal orifice does not appear until a rather later
period.

In striking contrast to all the other organs which adjoin it,
the club-shaped gland does not undergo any change of position
during what has been elsewhere called the provers of symmetri-
sation (cf. figs. 4, 5, and 18).

During Stage V theclub-shaped gland commences to atrophy,
and towards the end of that stage it disappears altogether, the
intra-buccal orifice being the last to go.

Considering their close proximity to one another, it would
be natural to suspect that intimate relations existed between
the club-shaped gland and the endostyle.

I made a few observations which seemed to show that the
endostylar epithelium, at least before it has assumed its ventral
position, is extremely delicate and liable to disrupture.

When a larva had been confined for a very short time between
slide and cover-glass, the epithelium of the endostyle would
commence to give off numerous small round homogeneous
ciliated bodies, which passed into the alimentary canal, and were
there apparently absorbed. .

This happened sometimes so soon after the larva had been
placed upon a slide that I did not recognise it at first as a
pathological phenomenon, as of course it was.

The fact that the endostyle is such a highly specialised and
delicate (while young) structure suggests the idea that possibly
a collateral organ like the club-shaped gland (concerning the
morphology of which see below) might function as a sort of

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. peal

special outlet for any nauseous substances entering the mouth,
which might otherwise act injuriously on the endostyle. But
the histological structure of the cells and the physiological
properties of the gland require further study before any safe
conclusion can be arrived at as to its function. The cells of
the gland are ciliated, and the external orifice is surrounded by
cilia.
4. The Mouth or Velum.

The mouth arises in an embryo of about thirty-two hours as
a minute circular aperture on the left side in the region of the
first myotome (see Hatschek, loc. cit.). It then grows rapidly
in size, and becomes a large lateral lens-shaped orifice, more
or less pointed at both ends. From Stage I onwards it begins
to grow gradually round the anterior extremity of the alimentary
canal, the change in position being accompanied by great alte-
ration in shape, consisting chiefly in the gradual equalisation
of its diameters, which are at first remarkably unequal, the
longer diameter being four or five times the length of the
shorter one. The bending round of the mouth commences by
a curious hunching up of the antero-dorsal margin of the mouth
into the ciliated groove which runs from the preoral pit to the
dorsal edge of the mouth.

By this means the previously pointed anterior extremity of
the mouth becomes lost, and this part of the mouth then pre-
sents a truncated appearance. Meanwhile the anterior half of
the mouth begins to bend inwards towards the right side, and,
in fact, it eventually becomes the right halfof the velum. The
gradual attainment by the mouth of an anterior median and
vertical position is fully illustrated in the plates at the end of
this paper. The mouth is relatively much smaller in the adult
than in the larva, but not actually so.

The edge of the larval mouth is provided with two kinds of
cilia, namely, (1) a set of evenly distributed small cilia, and (ii)
at a rather deeper focus much longer cilia, which hang together
in groups at equal intervals. At the anterior extremity of the
mouth in the earlier stages there is a group of very long cilia,

212 ARTHUR WILLEY.

seen with quite a surface focus ; and the whole external surface
of the lower wall of the mouth is beset with small cilia (figs.
21 and 23).

Four velar tentacles are present in Stage VII, and this
number continues all through the critical period. Eventually
fresh tentacles arise and make up the number which occurs in
the adult, viz. twelve.

5. Oral Hood and Buccal Cirri.

The right and left (lower and upper) halves of the oral hood
develop independently of one another, and are also entirely
independent of the metapleural or atrial folds, as will be readily
gathered from the figures. It will be seen that the oral hood
is formed mainly after the completion of the atrium. Professor
Lankester has suggested (5) that “the oral hood is the preoral
portion of the epipleural folds.” This, of course, is now seen
not to be the case.

The oral hood develops concurrently with the change of
position of the mouth.

The upper portion of the oral hood arises between Stage II
and Stage III as a fold of the integument overhanging the
preoral pit and ciliated groove. Beyond the latter it merges
into the upper margin of the mouth. This can be seen in
section by referring to the paper which has been already
quoted several times (8, pl. xxxi, figs. 13 and 14a). As
the fold becomes larger it extends to the posterior extremity
of the mouth, where it is met by the lower fold. Anteriorly
the upper fold of the oral hood is continued somewhat beyond
the limit of the preoral pit, and ends as a small ridge on the
left side of the head region, just below the level of the
notochord.

The formation of the buccal cirri is coincident with the
development of the lower portion of the oral hood. In the
adult the right and left halves of the oral hood are equally pro-
vided with cirri. It is, therefore, a most curious fact that the
buccal cirri take their prime origin in the lower or right half
only of the oral hood, and, after reaching a considerable stage

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 213

of development while still confined to the lower portion, they
then grow into the upper portion also.

Thus the oral hood itself has a double origin; it may be
said to arise from two independent ébauches (to employ a
French term which has not, when used in this connection, a
satisfactory English equivalent, “‘ forecast’”’ being the nearest
approach), while the system of the buccal cirri arises from a
single ébauche.

The formation of the cartilaginoid elements which form the
skeletal or supporting tissue of the buccal cirri, and which,
taken together, may be referred to as the buccal skeleton,
commences in Stage I. In this stage we find first one and
then two minute circular and homogeneous differentiations in
the mesoblast of the lower lip of the mouth, which can be seen
through the outer integument. They appear some distance
behind the external orifice of the club-shaped gland, just below
the mouth. They commence to form before the mouth has
begun to turn round in front.

In the succeeding stages fresh elements are added at both
ends—anterior and posterior—so that the central elements are
always the oldest ; and they soon begin to get irregular in out-
line, indicating a tendency to send out processes. Eventually
each piece gives rise to one process.

As the differentiation of the elements of the future buccal
skeleton proceeds the superjacent integument becomes raised
up into a fold, which is the lower or right fold of the oral hood.
About Stage IV the above-mentioned processes, which grow
out from the separate elements, attain such a size that the in-
tegumentary fold over them is thrown into a series of pleats
(fig. 6, &c.). By an extension of this pleating the buccal cirri
of the adult are formed.

From Stage V onwards the lower fold of the oral hood meets
the upper fold, and the point of junction is very plainly marked
by a ridge, under which the buccal skeleton penetrates into
the upper fold (figs. 8, 12, 13, and 16).

The lower fold is continued anteriorly some distance beyond
the buccal skeleton as a ridge. Eventually the cirri extend

214 ARTHUR WILLEY.

along the whole length of this ridge, as they do also along the
upper fold.

It has been stated above more than once that the lower fold
of the oral hood in the larva becomes the right half of the
oral hood in the adult ; and this half is, as Professor Lankester
lias pointed out (5), continuous in front round the extreme
point of the body with the dorsal fin.

A little reflection will show that this is the condition that
might be expected from the development of the lower fold, as
shown in the drawings accompanying this paper.

It is important to note that the buccal skeleton grows at
each end only, and that fresh elements are not formed inter-
stitially. In the adult the median cirri are smaller than the
others; and one would at first naturally suppose that these
were the youngest, and that this was the point at which fresh
cirri would be formed; as a matter of fact, however, the small
size of the median ventral cirri of the adult is deceptive, for
they are the oldest cirri, and new ones are only added at the
free extremities, right and left, of the buccal skeleton.

GENERAL CONSIDERATIONS.

It is my intention to confine myself mainly to the attempt
to give some explanation (or at least suggest one) of the three
most prominent features in the larva of Amphioxus, namely :

1. The asymmetry.
2. The endostyle.
3. The club-shaped gland.

The Asymmetry of the Larva.

With regard, then, first to the asymmetry of the larva, we
must begin by assuming—and the assumption will be sup-
ported later on—that in the primitive type from which the
Cephalochorda diverged the notochord did not extend to the
anterior end of the body, but, on the contrary, that the forward
extension of the notochord in Amphioxus is secondary and
adaptive; and further, that the primitive position for the

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 215

mouth was dorsal, as itis in the Ascidian tadpole.! The last as-
sumption may appear to be the most arbitrary, but it is justified
by the close affinities which exist between Amphioxus and the
Ascidians, and which will be referred to again. It may as well
be stated at once that the view I am about to put forward
depends essentially on this way of regarding the forward exten-
sion of the notochord ; and the immediate reasons for supposing
it to be secondary are—

i. That it does not extend to the front end of the body at its
first origin (see Woodcut, fig. 2).

i. That its forward extension is an obvious advantage to
Amphioxus when burrowing in the sand, which is its constant
occupation after a certain period of development.

il. The analogy of the Ascidian tadpole.

It is clear, then, that as the notochord pushed itself forwards
beyond the anterior limit of the nerve-tube, the mouth would
be compelled to forsake its primitive dorsal position—indeed,
it would be forced, as it were, to one side, just as the anterior
opening of the nerve-tube, which is at first actually dorsal, is
made to assume a lateral position through the development of the
dorsal fin; or, again, as the anus is similarly displaced by the
caudal fin. Itisasingular coincidence that the anterior neural
pore, the mouth, and the anus are all deflected to the left side.

The lateral position of the mouth is thus supposed to be due
to—or to occur in correlation with—the forward extension of
the notochord ; and it is evident that the same process which
caused the mouth to rotate from a dorsal position to its pre-
sent position on the left side in the larva would also by
implication cause structures which were originally on the left
side, or ventral, to pass across to the right side. There can
hardly be any doubt that a semi-rotation of the ancestral
pharynx has virtually taken place; the only question is,
whether we are right in correlating it with the forward exten-
sion of the notochord. Supposing the ancestral mouth to have
been small and circular, as it is in the Ascidian tadpole and in
the very young larva of Amphioxus, it is possible that this

1 See Baliour, ‘Comparative Embryology,’ vol. ii.

216 ARTHUR WILLEY.

rotation (which, as above indicated, need only be a virtual
phenomenon) may only have affected at first the most anterior
portion of the pharynx, namely, the portion containing the
endostyle, club-shaped gland, and first gill-slit.

In the ventral view shown in fig. 10 it will be noticed how
the primary slits curve round posteriorly to the ventral middle
line. It may be that formerly this tendency of the hinder
slits to lie in the direction of the left side, though now only
reaching the middle line, was more accentuated, and extended
to the more anterior slits. The position of the primary slits
which succeed the first on the right side has thus to be accounted
for, and this is done by assigning the cause of it to the further
adaptation which led to the huge size of the lateral mouth.

Having got on to the left side in the way above described,
it was found advantageous to have as large a mouth as circum-
stances would permit, and accordingly the capacious mouth of
the larva was established by the action of natural selection.
A glance at the figures should carry with it the conviction that
where the mouth is, there the gill-slits could not be.

Consequently the gill-slits which succeed the first develop,
like it, on the right side, though, as we have seen, behind the
region of the mouth they continue to retain a ventral position.

The lateral position of the mouth, then, is correlated with the
forward extension of the notochord, and the unilateral position
of the primary gill-slits with the lateral position of the mouth.

The rotation of the original left side gill-slits on to the right
side has led, as might have been expected, to the temporary
obliteration of the slits which properly belong to the right
side, but it is not so much an obliteration as a retardation in
development that has been effected. Thus, in a larva with
apparently only a single series of gill-slits, there is potentially
a double series; but, owing to the force of conditions which
have been secondarily induced, one of the two rows is some-
what late in putting itself in evidence.

That we have to do here with a retardation in the develop-
ment of a whole series of gill-slits is certainly proved by the
fact that when the secondary slits do appear, they form mainly

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. DAI

not one after the other, as is usual with metamerised organs,
but simultaneously. The successive or metameric mode of
formation, in fact, obtains theoretically with the secondary
slits of the larva of Amphioxus, only in this case it is latent,
with the natural result that the slits appear simultaneously.
The usually but not invariably late appearance of the first
secondary slit is due to a further slight retardation, and is
probably of no great significance.

The morphological mid-ventral line of the larva is indicated
by the blood-vessel which lies dorsal to the primary gill-slits
(fig. 1, &c.).

While the lateral mouth is serviceable to the pelagic larva,
it would plainly be awkward to the animal during its career as
a dweller in the sand—hence the necessity for the transforma-
tion which has been described above.

I have now given a possible explanation of the asymmetry
of the larva of Amphioxus, namely, that it can be traced ulti-
mately to the adaptive forward extension of the notochord. It
is thus a purely ontogenetic phenomenon, and is not an
ancestral character. It is only a temporarily superinduced
condition, and is destined to give place after a brief existence to
the inherent but latent bilateral symmetry of the organism.

AMPHIOXUS AND THE ASCIDIANS.

To pave the way for considering the significance of the endo-
style and club-shaped gland of the larva of Amphioxus, we
must institute a comparison between the embryo and larva of
an Ascidian and of Amphioxus. The remarkable researches
of Professors Ed. van Beneden and Charles Julin on the
morphology of the Tunicates (6) have rendered it possible to
do this the more effectually.

The peculiarities of an Ascidian embryo as described by the
above-named authors are as follows (see Woodcut, fig. 1):

The enteric cavity at first of two portions, viz. a pre-
chordal and a subchordal portion ; the greater part of the lumen
of the latter portion disappears at a very early period—it has
been identified with the post-anal gut of higher Vertebrates

218 ARTHUR WILLEY.

(see Balfour, ‘Comp. Emb.,’ vol. i1), into which the neurenteric
canal opens; but there is every probability that there is no
more post-anal gut in the Ascidian embryo than there is in
the embryo of Amphioxus.

The mesoblast consists of two longitudinal bands differen-
tiated from the primitive hypoblast.

These bands were formerly thought to be quite solid, but van
Beneden discovered (loc. cit.) that anteriorly they occur asa pair
of archenteric pouches (see Woodcut, fig. 1). The mesoblastic
bands in the region of the so-called tail are solid, and consist of
only asingle layer of cells continuous ventrally with the remains
of the hypoblast; but in front they consist of several layers of
cells, which at first surround a short lumen which communi-
cates, as stated above, with the enteric cavity on each side.

This single anterior pair of openings into the alimentary
canal is the only indication in the Ascidians of archenteric
pouches, and it is regarded by the authors as equivalent to the
first pair of somites of Amphioxus. ‘The rest of the mesoblast
in Ascidians is unsegmented, and the general absence of meso-
blastic pouches, with the exception of the first pair, is con-
nected with the early atrophy of the posterior or subchordal
portion of the alimentary canal, and is therefore probably
a product of degeneration, and not an ancestral character.

In the embryo of Amphioxus, at a roughly corresponding
stage (Woodcut, fig. 2) the notochord does not reach the
anterior end of the body, but even at this early period it extends
beyond the anterior opening of the nerve-tube (cf. Woodcut,
fig. 1). Partly in front of the notochord and partly below it
is a portion of the alimentary canal which lies in front of all
the mesoblastic somites. This anterior chamber consists of a
median portion and two lateral horns. The latter become con-
stricted off from the chamber, and form the anterior intestinal
diverticula, whose entire development has been fully described
by Hatschek (8); while the median portion of the chamber
simply merges with the anterior extremity of the alimentary
canal, and ceases to be recognisable asa distinct chamber. As
is well known, the left anterior diverticulum opens to the exterior

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 219

as the preoral pit, while the right one becomes the expanded
head-cavity of the proboscis-like end of the body.
The anterior chamber with its two lateral horns, together

yi

as
Tt

BiGh 2

EXPLANATION oF THE WoopcuTs.

Frc. 1.—Diagrammatic dorsal view of an embryo of Clavelina Rissoana
(modified after van Beneden and Julin). The mesoblast is represented
in an extremely diagrammatic way. It consists really of large polyhedral
cells.

a.p. Anterior neural pore. z. ¢. Nerve-tube, lying on the hypoblast
in front and on the notochord behind. p.v. Prechordal vesicle. J/. d.
Lateral diverticulum (paired) of prechordal vesicle. mes. p. Mesoblastic
or archenteric pouch : only one pair, equivalent to the first pair of pouches
in Amphioxus. mes. Mesoblast. ch. Notochord.

Fic. 2.—Dorsal view of an embryo of Amphioxus of a corresponding
age (after Hatschek).

Letters as above. The nerve-tube lies entirely on the notochord.
There are numerous pairs of mesoblastic pouches.

220 ARTHUR WILLEY.

with the region of the first myotome, is held by van Beneden
and Julin to be homologous with the przchordal portion of the
alimentary canal in the Ascidian tadpole. In the latter case,
also, two horn-like outgrowths are developed from this part of
the enteric cavity. They are subsequently met by two inva-
ginations—right and left—of the epidermis, with which they
fuse to form the so-called primary branchial canals. They
become the right and left peribranchial cavities, and consist
partly of epiblast and partly of hypoblast, and are considered
to be homologous with the right and left anterior intestinal
diverticula of Amphioxus, in which, however, the epiblastic
element is wanting.

The peribranchial cavities then expand, and, growing round
dorsally, fuse together to form the atrium. Meanwhile the
characteristic gill-slits or stigmata have commenced to form,
leading from the branchial to the peribranchial cavity, and
finally the left portion of the latter cavity alone communicates
to the exterior by the atriopore, the orifice of invagination of
the right portion of the cavity having been lost. It is stated
that probably the original communication of the primary
branchial canals with the enteric cavity is lost, and that all the
stigmata proper are secondary.

Thus the visceral wall of the Ascidian atrium is derived mainly
from the hypoblast, while the peripheral wall is formed from the
epiblast, and therefore the part of the atrium which leads to
the exterior on the left side by means of the atriopore consists
of epiblast; so that the preoral pit of Amphioxus, which is
composed entirely of hypoblast, does not exactly correspond to
the atrium of Ascidians, but is represented in the latter
group by the visceral wall of the atrium, while the actual
opening to the exterior of the przoral pit corresponds to the
junction of the hypoblastic and epiblastic elements in the
Ascidian atrium.

Neglecting their subsequent modifications, and supposing
the epiblastic involutions to have been suppressed or reduced
to zero in Amphioxus, the anterior intestinal diverticula of
the embryo of the latter do actually coincide in every essential

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 221

respect with the primary paired atrial cavities, or, as they have
been called, branchial canals of the Ascidian embryo; and the
final opening to the exterior in both cases is connected with
the original left diverticulum.

The two embryos represented in the woodcuts are thus
regarded as practically identical with one another in all mor-
phological respects. There is, in fact, no positive difference
between them except in the relative proportion of parts of the
body; the general absence of mesoblastic somites in the
Ascidian being, as before stated, secondary and correlated
with the atrophy of the subchordal portion of the archen-
teron.

It will possibly have been noticed that the forward exten-
sion of the notochord in Amphioxus commences at a remark-
ably early stage, months before the larva takes to the sand.
This can be accounted for as a hastening of the development,
a familiar phenomenon.

The divergence, then, between the two embryonic types
consists in the great development of the prechordal region in
the Ascidian embryo, and in the reduction of the same region
in the embryo of Amphioxus.

It will have been inferred from what has been said that the
tail of the Ascidian tadpole is regarded as the equivalent of the
trunk of Amphioxus.

In the Ascidian the expanded portion of the prechordal
region after giving rise to the peribranchial diverticula, becomes
the branchial sac or pharynx, the floor of which is specialised
as a glandular organ known as the endostyle.

The narrow portion of the prechordal vesicle, which also
extends a very short distance beneath the anterior extremity
of the notochord, gives rise to the esophagus and stomach,
while the intestine arises as a lateral outgrowth from the latter;
but the extraordinary feature of it is that the intestine grows
out from the stomach to the right of the mid-ventral line,
and then passes across to open into the cloacal vesicle on the
left side.

There can be little doubt that the ancestor of the Ascidians,

222 ARTHUR WILLEY.

before the atrophy of the subchordal portion of the intestine,
had an anus at the posterior extremity, as is the case with
Amphioxus. This primitive outlet has, therefore, been sup-
planted by the development of the outgrowth from the stomach
mentioned above, which van Beneden and Julin call a collateral
diverticulum, and at the same time make the suggestion,
which my observations tend to support, that the intestine of
the Ascidian, arising as above described, is homologous with
the club-shaped gland of Amphioxus.

THe ENDOSTYLE AND CLUB-SHAPED GLAND OF AMPHIOXUS.

We must now define the position of the endostyle in the
larva of Amphioxus in order to grasp the situation of the club-
shaped gland.

It has been already implied that in all probability the
ancestor of Amphioxus was provided with a prechordal region
and was symmetrical; and, in fact, resembled an Ascidian tad-
pole before the atrophy of the subchordal portion of the
intestine.

In the larva of Amphioxus the endostyle lies at the extreme
front end of the alimentary canal, which, as we have said
above, represents the median portion of the anterior or ‘ pre-
chordal’ chamber, the lateral portions of which formed the
anterior intestinal diverticula. It is further placed in the
region of the first myotome ; its peculiar oblique and lateral
position has been already dwelt upon and explained. It also
lies immediately in front of the club-shaped gland.

It results, first, from the analogy of the Ascidian tadpole,
and secondly, from the position of the endostyle in the larva of
Amphioxus, that it (the endostyle) was originally differentiated
as a mid-ventral groove in the prechordal region. This primi-
tive anterior position for the endostyle had been previously
surmised by van Beneden and Julin, so that my account of
its development in Amphioxus harmonises well with their con-
clusions.

The transverse growth which affected the anterior region

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 223

of the alimentary canal of Amphioxus at the time of the
forward extension of the notochord naturally rotated the
originally median ventral endostyle on to the right side in
company with the collateral organ, now known as the club-
shaped gland.

A comparison of the figures accompanying this paper, illus-
trating the development of the endostyle, shows clearly and
beyond question that everything dorsal to the endostyle is on
the right side of the morphological middle line, while every-
thing below it is on the morphological left side. Thus the
intra-buccal orifice of the club-shaped gland is on the right
side, and the first and succeeding primary slits are on the left
side—morphologically. At this juncture I want especially to
emphasise the relative positions of the intra-buccal orifice of
the gland and of the first primary slit.

The csophagus and stomach of the Ascidian, which arise,
ontogenetically, from the preechordal vesicle, are practically un-
represented in Amphioxus—the part of the vesicle from which
they arise being suppressed. In fact, in their respective
capacities as cesophagus and stomach they are not even
theoretically present. Bearing this in mind, we can now pro-
ceed to compare the position of the club-shaped gland and its
apertures with the position of the intestine in the young
Ascidian.

In the larva of Amphioxus the club-shaped gland is situated
behind the endostyle, i.e. at the posterior extremity of the
primitive preechordal vesicle—in this respect agreeing with the
intestine of an Ascidian. In both cases the aperture of the
collateral diverticulum (whether gland or intestine) into the
alimentary canal is immediately to the right of the mid-ventral
line—in the one case actually, and in the other case morpho-
logically ; and in both cases the external aperture is on the
left side. The point of origin of the club-shaped gland between
the first and second myotomes also corresponds more or less
closely with that of the Ascidian intestine. There is, therefore,
a fairly strong case in favour of the homology of the two struc-
tures. The question still remains as to whether the club-

VOL. XXXII, PART Il,—NEW SER. P

224, ARTHUR WILLEY.

shaped gland is a rudimentary structure which formerly served
as an intestine, or whether it is what van Beneden and Julin
would call a primitive collateral organ, which in the Ascidians
has come to function as an intestine. A great many con-
siderations render it only reasonable to suppose that the intes-
tine and anus of Amphioxus are primitive, and hence the
club-shaped gland must be regarded as an ancestral organ
occurring in company with a posterior anus.

Assuming, then, that the ancestor of Amphioxus possessed a
collateral organ of unknown function situated behind the
primitive median endostyle, with internal and external orifices
as described, it is difficult to conceive how it can have become
evolved, except by the modification of some pre-existing organ.
The spectacle of an unpaired hypoblastic gland opening to the
exterior on the one hand, and into the alimentary canal on the
other, without having a morphological equivalent in the whole
animal kingdom, is certainly too much.

There are thus two points to settle:

1. What was the nature of the pre-existing organ referred
to above ?

2. Was it paired or unpaired ?

It may be remembered that the intra-buccal orifice of the
club-shaped gland, which was described and figured for the
first time in the paper by Professor Lankester and myself quoted
several times above (8), is not present in the very young larve,
but forms later. This is probably another instance of a slight
and unimportant retardation in development.

Before becoming acquainted with the views of van Beneden and
Julin I had come to the conclusion, on grounds which seemed
sufficient, that the club-shaped gland was a modified gill-slit ;
and, as I am aware of no other attempts to account for the mor-
phology of the gland, I will give reasons for so regarding it.

In the second place, there are reasons for supposing that it
was originally one of a pair of gill-slits, and further, that the
original pair actually exists in the larva of Amphioxus—the
members of the pair being the club-shaped gland and the first
primary gill-slit.

LATER LARVAL DEVELOPMENT OF AMPHAIOXUS. 22d

If, then, the club-shaped gland is shown to be a modified
gill-slit on the one hand, and is homologous with the intestine
of the Ascidians on the other, one is led to the apparently pre-
posterous conclusion that the intestine of Ascidians is the
morphological equivalent of a gill-slit. ‘The secondary intes-
tine of Ascidians is certainly not homologous with the intestine
of Amphioxus. No other homology was thought of before
van Beneden and Julin suggested that of the club-shaped
gland of Amphioxus. My observations do not run counter to
that suggestion, except possibly in so far as they show that
the club-shaped gland is a modified gill-slit. The question is,
can it be both?

Reasons for regarding the Club-shaped Gland asa
Modified Gill-slit belonging tothe Right Side—
the Corresponding Slit of the Left Side being re-
presented by the First Primary Slit.

1. The club-shaped gland communicates with the exterior
at one end and with the alimentary canal at the other end.

2. Its internal aperture, being dorsal to the larval endostyle,
is therefore to the right of the morphological middle line.
What was originally a simple branchial passage has thus been
virtually drawn out into a long tube, and the external aperture
has been shifted from the right to the left side.

3. The first primary slit, being below the endostyle, is there-
fore to the left of the morphological middle line, and the
appearance which it presents of being distinctly behind the
gland is superficial and secondary. It lies in the region of the
second myotome.

4. The club-shaped gland arises simultaneously with, and in
a similar manner to, the first primary slit, as a ventral pouch of
the alimentary canal (see Hatschek, 3); and though it arises
clearly in front of the first slit, this may be due partly to the
remarkable tubular form that it has attained as a result of its
evolution. I am not attempting to explain why it passes
across to open on the left side, but as it does so it must pass

226 ARTHUR WILLEY.

either in front of or behind the unmodified slit with which it
was primitively paired. As a matter of fact, of course, it
passes in front of that slit, and so it has come—possibly in some
measure owing to the curious torsion which has affected this
region—to arise in front of the first gill-slit, with which it
really on this view corresponds as a pair. It thus belongs
morphologically to the region of the second myotome.

5. The atrophy of the club-shaped gland occurs simulta-
neously—or nearly so—with the atrophy of the first primary
slit. This fact is worthy of the utmost emphasis.

6. In the formation of the secondary slits the first of them
pairs with the second primary slit, and no true secondary slit
ever appears to correspond with the first primary slit.

7. After the simultaneous formation of the club-shaped
gland and the first primary slit (the mouth and anus being also
present) there is a prolonged interval, during which no further
formation of gill-slits occurs.

This might indicate a distinction between the first primary
slit and the succeeding ones.

These are the reasons, drawn from facts, all of which, except
Nos. 4 and 7, are illustrated by figures accompanying the
present paper.

Deduction.—The club-shaped gland and first primary gill-
slit represent an ancestral pair of gill-slits, which atrophies
in the course of larval development.

How or for what special object a gill-slit could be modified
into such a tubular collateral organ is quite another matter,
which I do not now propose to consider further (vide p. 210).
Suffice it if the reasons given above are enough to lead us to
suppose that such a thing has indeed happened in the present
case.

The closure of the hinder primary slits presents an interest
different from that of the first, and is the decisive step in what
I have before called the “‘ symmetrization ”’ of the larva. The
transverse growth which carries the more anterior slits from
the right to the left side does not extend back so far as the
hinder slits, so that they are left in the middle ventral line,

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 227

where the endostyle will eventually come to lie; hence the
necessity for their closure.!

MorrHonocicaL ImMPpoRTANCE OF THE DEVELOPMENT OF THE
ENpostyLE or AMPHIOXUS.

1. Its anterior position in the larva in the part of the alimen-
tary canal which is equivalent to the median portion of the
* preechordal”’ vesicle, shows its entire homology with the endo-
style of Ascidians, which is also situated in the median portion
of the prechordal vesicle.

2. Its anterior position also supports the view that the endo-
style was primitively differentiated in and confined to the mid-
ventral line of the ancestral prechordal vesicle.

3. Its oblique position on the right side is secondary, and is
due to the rotatory growth which affected the anterior region
of the primitive alimentary tract, in correlation with the
adaptive forward extension of the notochord.

4. It enables the morphological median line of the larva to
be fixed with absolute precision, and shows that it, viz. the
median line, has come to occupy a secondary and temporary
position, high up on the actual right side of the larva.

5. The important fact can thus be established that the intra-
buccai orifice of the club-shaped gland is situated to the right
of the morphological middle line.

6. The endostyle of the larva of Amphioxus possesses two
distinct halves, fused together posteriorly, but with free anterior
extremities. At first the right or upper half is much smaller
than the lower portion, and this is only natural when one re-
members that most of the other structures belonging to the
primitive right side are wholly retarded in their development.

7. In the larva it is confined to the region of the first myo-
tome which is destitute of gill-slits.

8. The backward growth of the endostyle of Amphioxus has
been acquired concurrently with the multiplication of gill-slits
in the segmented region of the trunk.

* It must be confessed, however, that the meaning of the first appearance
and subsequent atrophy of these slits is not quite clear.

228 ARTHUR WILLEY.

9. Since, as will be shown below, the pharynx of Amphioxus
is not homologous, in the true sense of the word, with the
branchial sac of Ascidians, the adult position of the endostyle
of Amphioxus cannot be homologous with that of the Ascidian
endostyle. The observations recorded in this paper, however,
prove unquestionably that the position of the endostyle in the
adult Amphioxus is secondary, and that in its origin it is, as
stated above, perfectly homologous with the endostyle of the
Ascidians.

Further Homologies.

It will have been inferred, from what has been stated above,
that the pharynx of Amphioxus, which is part of the trunk, is
not truly homologous with the branchial sac of Ascidians, which
is part of the prechordal vesicle; so that we have to face the
paradox that while a highly specialised organ of (speaking
generally) identical structure and relations, and extending the
whole length of the pharynx, is present in both cases, yet the
pharynx taken as a whole of the one is not homologous with
that of the other. This, no doubt, sounds at first like a
reductio ad absurdum of a morphological problem, but
the figures in the plates accompanying this paper, showing the
primary and secondary positions assumed by the endostyle of
Amphioxus, should assist in the realisation of this at first
startling paradox.

The gill-slits or branchial stigmata of the Ascidians are
homologous with those of Amphioxus only in the sense that
they are distinct vertebrate structures, but they are not
homologous in their origin, position, or relations. There is
thus a sort of group homology existing between them.

In Amphioxus the slits form metamerically in the segmented
region of the trunk. In Ascidians they form in front of the
(however imperfectly) segmented region, and, moreover, van
Beneden and Julin point out that they do not arise metameri-
cally one after the other, but irregularly ; for instance, the first-
formed slit will be the fourth of the series eventually, and so on.

If these conclusions are as sound as they appear to be, it is

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 229

interesting to note that there are two kinds of edie homo-
logies, which may be called—

1. Individual homology, the one more commonly met with.
This is a morphological factor.

2. Group homology, a physiological factor, but entirely
distinct from homoplasy.

It also follows that the peribranchial cavity of Ascidians is
in no sense homologous with the atrium of Amphioxus ; in fact,
this is a pure instance of homoplasy.

Bateson (9) points out the probable homology of the proboscis
cavity and proboscis pore of Balanoglossus with the anterior
“ prechordal ” vesicle and preoral pit respectively of Amphi-
oxus; and also he urges the homology between the gill-slits in
both cases. These identifications are likely enough to be
accurate, but I need not recapitulate the reasons for regarding
Amphioxus as more closely related to the Ascidians than to
Balanoglossus, though, since it forms a considerable feature in
the present paper, the absence of a true endostyle in Balano-
glossus might be pointed out. The forward position of the rudi-
ment of a notochord in Balanoglossus has probably no relation
whatever with the forward extension of the notochord in
Amphioxus.

As to the affinities of Amphioxus with Ammoceetes, I will
only refer to the homology existing between the endostyle of
Amphioxus and the thyroid gland of Ammocecetes, which was
advocated first by W. Muller and then by Dohrn (7), who worked
out the development of the latter structure. It is well known
that Dohrn regards the thyroid gland, and therefore the endo-
style, as representing a pair of gill-slits. This is of course aside
issue, and cannot be fully gone into here ; but it may be pointed
out that while the thyroid gland gives no reliable indication
of a double origin, the endostyle of Amphioxus is composed
of two distinct halves, which are at first upper and lower re-
spectively, but eventually become right and left—morphologi-
cally of course their relation to one another is always right
and left.

As for the gill-slits in the two cases, attention may be drawn

230 ARTHUR WILLEY.

to the pair of rudimentary gill-slits, called branchial diverticula,
in front of the first pair of true slits, of which Dohrn established
the existence in the young Ammoccetes, and which never
actually open to the exterior; and this circumstance should
be remembered in connection with the atrophy of the first pair
of gill-slits (i.e. the club-shaped gland and first primary slit)
which occurs at a certain stage in the larval development of
Amphioxus.
Further, the fact that the larva of Amphioxus passes through
a prolonged critical period, during which it possesses only eight
or at most nine pairs of gill-slits, is at least suggestive.

ADDENDUM.

The common ancestor of the Ascidians and Amphioxus
cannot be properly imagined until further knowledge is forth-
coming as to the significance of the primary pair of diverticula
of the przchordal vesicle and the function of the club-shaped
gland.

Gill-slits were probably present in the segmented region of
the trunk, which have been lost by the existing Ascidians,
but whether one pair or several pairs is a question.

If the club-shaped gland is admitted to be a modified gill-
slit, as I hope to have shown that it is, there must have been
at least one pair of such slits present.

LITERATURE REFERRED TO.

=

Kowatewsky.—-‘ Mém. Acad. Imp. des Sciences de St. Petersburg,’ 7th

series, vol. xi, 1867.

. Kowatewsky.— Archiv fiir mikro. Anat.,’? Bd. xiii, 1877.

. HatscHeK.—‘ Claus’s Arbeiten,’ 1881.

. HatscneK.—‘ Zoologischer Anzeiger,’ 1884.

. LANKESTER.—‘ Quart. Journ. Micr. Sci.,’ vol. xxix, 1889.

. Van BEnEDEN AND JuLIn.—‘ Archives de Biologie,’ vol. vi, 1887.

. Donryn.—Studien No, XII, ‘ Mittheilungen aus Zool. Stat. zu Neapel,’
Bd. vii, 1887.

. LANKESTER AND WILLEY.—‘ Quart. Journ. Mier. Sci.,’ vol. xxx, 1890.

. Bateson.—‘ Quart. Journ. Micr. Sci.,’ vol. xxv, 1885.

Ioa pr © WY

© @

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 231

DESCRIPTION OF PLATES XIII—XV,

Illustrating Mr. Arthur Willey’s paper on “ The Later Larval
Development of Amphioxus.”

All the figures, without exception, were drawn from the living animal, and
each figure is from a separate larva. The drawings are made to scale. From
Stage IIT onwards all the slits are seen through the transparent atrial or
body-wall—i. e. they open into the atrium, and not directly to the exterior.

N.B.—The long cilia on the surface of the body are not represented in the
figures, nor in those of the memoir by Professor Lankester and myself (8).
Their character and position are well shown by Hatschek in his memoir in
Claus’s ‘ Arbeiten,’ 1881.

Fic. 1.—Stage I. Right side. Fourteen primary slits—notice small size of
the first. Six secondary thickenings occurring in a continuous ridge above
the primary slits, the first of them being above and between the third and
fourth primary slits. The preoral pit of course opens on the left side, and is
only seen by transparency on this side.

Fic, 2.—Stage I. Right side. Twelve primary slits. Five secondary
thickenings, the last beg much smaller than the others, and the first being
situated between the fourth and fifth primary slits. The second as well as
the first secondary thickening has in this case been slightly retarded; thus
showing that the retardation of the first secondary slit, which has been stated
in the text to be usual, is not specially important. Of course, the general
retardation of development which affects the whole series of secondary slits
is extremely important. Notice the endostyle and club-shaped gland.

Fic. 3.—Stage II. Right side. Fourteen primary slits, the fourteenth
appearing in side view merely asa pit in the floor of the pharynx; this shows
that it is on the way to close up. Seven secondary slits, one having been
added behind; the first still lies between the third and fourth primary slits ;
a new one is subsequently formed in front, so that what appears to be the first
slit or thickening at this stage eventually becomes the second secondary slit.
In this larva the second to the sixth of the secondary thickenings have just
been perforated. The appearance of a seventh thickening behind is rather
early to happen at this stage.

Fic, 4.—Stage II]. Right side. Twelve primary slits; the first of them is
getting smaller. The fin-chambers seem to form about this stage. There is
a small anterior portion of the atrium still unclosed. The right wall of the
future velum is now seen from this side, owing to the great extent to which

232 ARTHUR WILLEY.

the mouth has bent round the anterior extremity of the alimentary canal.
There are seven secondary slits, the seventh not yet perforated ; the first is
still above and between the third and fourth primary slits. Notice the
flattened tops of the larger secondary slits. All the primary slits behind and
including the fourth now bend under the pharynx towards the left side. Notice
the endostyle and club-shaped gland.

Fig. 5.—Stage IV. Right side. Thirteen primary slits, the thirteenth
closing. Seven secondary slits, the first and last having only just opened.
The true first secondary slit has now appeared, and lies above and between
the second and third primary slits. The larger secondary slits have begun to
double in to form the tongue-bars. Notice the direction of the long axis of
the secondary slits. All the primary slits are now bending under the pharynx,
and the first is extremely small. The endostyle has definitely commenced to
grow beyond the club-shaped gland. This should be noticed carefully and
compared with the condition of the two structures in the preceding stages.
The atrium is quite closed in front.

Fic. 6.—Stage IV. Left side. This is to show the extent to which the
primary slits have come round to the left side at this stage. The number—
fourteen—of primary slits is not typical for this stage (see Fig. 5). The right
or anterior half of the velum is seen with a deep focus. There are six
elements of the buccal cirri in the lower half of the oral hood, growing out
into tentacles and covering up the external orifice of the club-shaped gland.
The upper half of the oral hood is hanging over the upper wall of the mouth
or velum and over the preoral pit.

Fic. 7.—Stage V. Right side. This is the very commencement of Stage V;
and the first primary slit, though very small, is still to be seen in side view.
The secondary slits, of which there are eight, have become larger ; and in the
third slit the tongue-bar has actually fused with the ventral wall of the slit.
The endostyle and club-shaped gland have the same relations as in Fig. 5.
The cecum is commencing to grow out; there is no fixed stage at which this
occurs. The primary slits have nearly disappeared from this side, but are still
seen below the secondary slits.

Fic. 8.—Stage V. Left side. The primary slits are not fully round to the
left side yet, but the larger of them are commencing to form tongue-bars.
The twelfth primary slit is very small, and the thirteenth and fourteenth are
closing. Notice the point of junction of the upper and lower portions of the
oral hood; the latter is prolonged anteriorly as a ridge. ‘The right wall of the
velum is deeply focussed.

Fic. 9.—Stage V. Three-quarter ventral view. This shows the first
primary slit in a condition of approaching atrophy. The twelfth is also closing.
The bases of the secondary slits can just be seen.

Fic. 10.—Stage V. Ventral view. This should attract particular attention.

LATER LARVAL DEVELOPMENT OF AMPHIOXUS. 233

Feeble traces of the first primary slit can just be detected in front. The
twelfth primary slit is again seen to be closing. Notice the series of primary
slits curving round to the mid-ventral line behind. The tongue-bars are
complete in most of the secondary slits, but in none of the primary slits.

Fic. 11.—Stage VI. Left side. Nine (primary) slits, the first being of
course originally the second primary ; but it has come to be the first in position
in consequence of the atrophy of the true first. Behind there are traces of the
eleventh and twelfth primary slits. Only the left half of the velum can now
be seen from this side. I do not know what has become of the nephridium
of Hatschek; 2 is probably part of the ciliated tract of the preoral pit.
Notice the left half of the endostyle which extends under three slits ; the right
half is out of focus. In this larva there were eight secondary slits on the
right side.

Fic. 12.—Stage VI. Left side. This shows the twelfth aad thirteenth
primary slits in course of closure. The condition of the gill-slits is not quite
so far advanced as in the preceding figure, but the buccal cirri are farther
advanced, There were seven (secondary) slits on the right side.

Fic. 13.—Stage VII. Left side. In this larva the ninth slit (= tenth
primary) would probably have closed. The eleventh primary slit is represented
by arudiment. Czcum is commencing to bulge in the region of the twentieth
myotome. Notice the length of the endostyle. The buccal skeleton is
beginning to grow into the upper portion of the oral hood.

Fig. 13a.—Separate view of the velum of the same larva, showing the
four primary velar tentacles. The right and left halves of the velum are seen
to be not quite opposite to one another.

Fie. 14.—Stage VII. Right side. The slits are commencing to elongate in
a direction at right angles to long axis of body. ‘There are eight (secondary)
slits on this side. On the left side of this larva there were eight (primary)
slits, and there was also a median rudimentary slit on the point of closure,
viz. the tenth primary, which sometimes persists, as explained above, as the
ninth slit of the left side. The view of the endostyle is very important,
showing its double nature—upper and lower halves. It has grown consi-
derably in length. The right metapleur is omitted. The floor of the atrium
consists of the expanded subatrial ridges.

Fig. 15.—Stage VIII. Right side. In this larva there were eight slits on
each side and no rudimentary one behind, so that this is typical of the critical
period. The view of the endostyle is again very important. The two halves
have now assumed a definite right and left position. Its double origin can be
detected here for the last time. Notice in this and the preceding figures the
hyperpharyngeal band. Right metapleur is omitted.

Fie. 16.—Stage VIII. Left side. Nine gill-slits ; the ninth has not fully
come round to this side, but bends under the pharynx. There were eight slits

234, ARTHUR WILLEY.

on the right side. Notice the ccelom just in front of the cecum. ‘The blood-
vessel in this region is contractile. The endostyle is farther advanced than in
the preceding figure. The right and left walls of the velum are quite opposite
to one another.

Fic. 17.—Stage VIII. Combination of three focussings, showing the last
(seventh) pair of slits of a larva in which the ninth primary slit was ina
rudimentary condition in the mid-ventral line.

Fic. 18.—Stage V. Right side (anterior portion only), to show the endo-
style, which has grown a long way past the club-shaped gland between the
primary and secondary slits.

Fic. 19.—Stage VI. Similar view of right side, to show that the club-
shaped gland has’ now atrophied, its remains being still visible on the
endostyle.

Fie, 20.—Stage I. Left or oral aspect of a larva. Note pointed anterior
extremity of mouth, ciliated groove, external orifice of club-sliaped gland, and
one element of buccal skeleton.

Fie. 21.—Stage I. Similar view. Antero-dorsal margin of mouth, com-
mencing to hunch up into the ciliated groove. Two elements of buccal
skeleton.

Fic. 22.—Stage II. Similar view. Hunching up and sinking inwards of
the anterior portion of the mouth continued. Three elements of buccal
skeleton.

Fic. 23.—Stage I. Similar view. Four elements of buccal skeleton,
commencing to grow out as tentacles. When the very young cirri are seen
end-on they present a circular appearance.

Fic. 24.—Stage II. Similar view. Five elements of buccal skeleton.

Fie, 25.—Stage III. Similar view, to show the anterior half of the mouth
bent at a deep focus towards the right side. Note also the two portions—
upper and lower—of the oral hood, the latter containing the buccal skeleton.

Fie. 26.—Stage IV. Similar view. This is an important view of the mouth,
showing its change in shape but not diminution in size. The external orifice
of the club-shaped gland is covered over by the buccal cirri.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 285

On the Structure of Two New Genera of Earth-
worms belonging to the Eudrilide, and some
Remarks on Nemertodrilus.

By

Frank E. Beddard, M.A.,
Prosector to the Zoological Society of London.

With Plates XVI—XX.

Tue worms which form the subject of the following paper
were obtained in a living state from Kew Gardens, along with
a number of others whose structure I propose to investigate
later.

I had applied to Mr. Thiselton Dyer for leave to sift soil
coming from tropical countries in the Wardian cases which
are generally used for transmitting plants in pots. This Mr,
Dyer very kindly permitted me to do, and in his absence Mr.
Morris was so good as to put me in the way of carrying out
my wishes. To both these gentlemen my thanks are tendered.
In the earth surrounding a number of pots containing plants
from Lagos, West Africa, I found about half a dozen worms,
which proved on examination to belong to the family
Eudrilide.

Until quite recently one genus—Eudrilus itself—was the
only known representative of the family; three years ago Dr.
Rosa described from Scioa, Africa, another genus—Teleu-
drilus; and quite recently Dr. Michaelsen has received from
Zanzibar and the mainland opposite, and from the mouth of the
Zambesi River, a number of species, all belonging to this family,
and referable to four new genera: the family is, therefore, in the
present state of our knowledge, characteristically Ethiopian,

236 FRANK HE. BEDDARD.

though the type genus Eudrilus has not yet been met with
from the African continent.!

The Eudrilide which I obtained from Kew belong apparently
to two distinct species, which are also generically distinct.
Four specimens I refer to a new genus, for which I propose
the name of Hyperiodrilus. The fifth specimen is a species
of another genus—Heliodrilus, nov. gen. Thesixth is a very
minute worm, measuring barely an inch in length, and being
sexually immature was indeterminable.

I.—Hyperiodrilus africanus, nov. gen., sp. nov.

The worms are of different sizes; the largest specimen
measures (after preservation in spirit) about five inches;
during life the length was rather greater. Their form is very
slender, and their movements were active, though they did
not show any power of jumping, such as is shown by
Pericheta. While moving the buccal cavity is to a certain
extent everted, and is made use of as a sucker for attaching
the front end of the body ; the eversion of the buccal cavity is
not nearly so pronounced as in Pericheta,

The colour of this species is pinkish, and the posterior
segments have a distinctly ringed appearance. There does not
seem, however, to be much pigment in the skin; the colour is
entirely due to the enclosed viscera, particularly, of course,
to the blood-vessels—even the ringed appearance of the poste-
rior segments is due to the same cause; the blood-vessels
ramifying over the septa produce the appearance of bands of
red pigment corresponding to the segments: the clitellum is
yellowish.

§ External Characters.

An examination of the external characters at once shows
that this worm is referable to the Eudrilide, but that it cannot
be included in any known genus of that family, with the
possible exception of Stuhlmannia.

1 Since the above was written Drs. Horst and Michaelsen have received
from Africa specimens of Hudrilus.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 237

The prostomium does not completely divide the peri-
stomial segment; but it sends back (see fig. 6) a very narrow
prolongation, which is embedded in the peristomial segment
up toa point not very far distant from its posterior border.
Such a narrow prolongation of the prostomium appears to be
characteristic of the HEudrilide ; it is specially mentioned by
Rosa (10) in the case of Teleudrilus, and is figured by
Michaelsen (7, pl. 111, fig. 17) in Nemertodrilus.

The dorsal sets are in couples; of the ventral sete, the
individual sete of each couple are some little way apart, as
illustrated in figs. 9 and 12.

The clitellum is developed all round the body, and
occupies four segments, Nos. 14, 15, 16, and 17, as in
other Eudrilide, where four segments is also the usual extent
of the clitellum, though sometimes, as in Polytoreutus, ex-
ceeded. Only one specimen, however, had the clitellum
developed upon the 14th segment, and here it was incomplete,
extending only over the dorsal surface of that segment. In
two other specimens the clitellum only occupied Segments 15,
163417.

Nephridiopores were obvious upon most of the segments
of the body, particularly upon the clitellum, where the smooth
swollen integument rendered them very easily visible. They
lie almost in the intersegmental furrow in front of the dorsal
sete. It may be noted that they occur, as shown in fig. 23 of
Pl. XVIII, in the 14th and 17th segments, where the ducts of
the generative organs open.

Dorsal pores could not be detected in any part of the
body.

The apertures of the oviducts are upon the 14th seg-
ment near to its posterior border: each has the appearance of
a minute hemispherical projection, which cannot be confounded
with a nephridiopore; besides, as already mentioned, the
14th as well as the 15th segment has its nephridiopores.

The oviducal papillz, as they may be more accurately spoken
of, are situated behind the dorsal seta of the 14th segment on
each side; in one case, as shown in fig. 23 of Pl. XVIII, they

238 FRANK E. BEDDARD.

were placed just in front of the nephridiopore of the segment
behind ; more usually, however, they are a little dorsal of the
nephridiopore.

The aperture of the sperm-ducts is, as in Stuhl-
mannia and Teleudrilus, single and median; it lies at the
end of the clitellum, between the 17th and 18th segments. It
does not, however, present the appearance of an orifice, but of
a prominent hemispherical papilla, as in the case of the ovi-
ducal pore; the papilla is in this case considerably larger.

In one specimen (that which had four clitellar segments)
this papilla was connected by two grooves running obliquely
forwards with a pair of conspicuous rounded papille, placed
near to the boundary line between the 17th and 16th segments ;
the two grooves diverged from each other at an angle of about
40°; each ran along a slight mound-like elevation, as shown
in fig. 24 of Pl. XVIII, and more highly magnified in fig. 19
of the same plate. The two papille are unsymmetrically dis-
posed, as also shown in the figure ; the left-hand one is placed
just behind the groove separating the 17th from the 16th
segment ; the right-hand one at about the middle of the 17th
segment.

In a second specimen (Pl. XVIII, fig. 22) the median papilla
was quite invisible, owing probably to the worm having died
with the part in question retracted.

In the only other specimen belonging to this species, which
was nearly mature, a prominent glandular swelling occupied
the middle ventral line between Segments 17 and 18 (Pl. XVIII,
fig. 20). A careful examination showed that the apparently
single protuberance is not to be confounded with the median
papilla described in the first of the three specimens ; it really
represents the two anterior papillz of the 17th segment closely
fused; the groove issuing from each may be detected, and the
median papilla itself is visible at the point where the two
grooves nearly come into contact.

In Stuhlmannia variabilis (Michaelsen) the papillz
connected with the male efferent apparatus show certain re-
semblances to those of the present species.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 239

There is a median aperture (as in Teleudrilus and other
Eudrilide) of the “ prostate glands” upon the 17th segment ;
this is spoken of as a slit-like orifice; but the apparent
difference in this particular from Hyperiodrilus africanus
may be a mere question of the state of contraction of the
worm’s body. From the aperture upon the 17th segment a
deep furrow runs forward to a process which bears the outlet
of a peculiar gland; this process is median, but slightly in-
clined to the right side. This papilla, however, appears from
Michaelsen’s description to lie upon the 13th segment, near to
the ventral unpaired orifice of the spermatheca. As already
mentioned, the lateral processes, though varying somewhat in
their position upon the segment, are always on Segment 17.

On the 13th segment is a median aperture (see fig. 24)
corresponding to the male generative orifice upon the 18th
segment, though situated at about the middle of the segment,
rather nearer to the anterior than to the posterior boundary.
This aperture was not at all conspicuous upon any of the
specimens examined, and might very easily be overlooked.

§ Integument.

The layers of the body-wall are as in other earthworms,
except for the presence in the epidermis of certain peculiar
organs, which appear to be met with in all Eudrilide except
Nemertodrilus; but this genus is in other respects (see p.
266) a very aberrant member of the family. These structures
are described later.

The muscular layers of the body-wall show no noteworthy
peculiarities as regards the structure and arrangement of the
fibres. The longitudinal coat does not exhibit the bipinnate
character which is frequently met with in earthworms, par-
ticularly in the Lumbricide.

The muscular coats contain numerous irregular spaces
filled with coelomic corpuscles (see fig. 3) ; besides the ordinary
corpuscles large multinucleate bodies are met with, which
may be pathological formations. I believe that Kihenthal
(18) was the first to specially call attention to the spaces

VOL. XXXII, PART Il.—NEW SER. Q

240 FRANK E. BEDDARD.

in question. They are unusually abundant in both Hyperio-
drilus and Heliodrilus—more so than I have observed in
any other earthworm. In places each individual fibre was
separated from its neighbours by quite large spaces filled with
corpuscles.

§ Sete.

The arrangement of the sete is somewhat peculiar ; the more
dorsally situated pair, which are in reality lateral, and not
dorsal, are closely approximated ; the ventral couple (fig. 12)
are at some distance apart; the distance separating the 2nd
from the 3rd seta! is twice that which separates the Ist from
the 2nd. The arrangement of the setz is such as to produce the
impression that there are only three on each side of the body.
It appears to me possible that Kinberg’s genus Tritogenia
has the setz arranged in this way, having thus led Kinberg to
make the statement that it possessed only six sete per somite.
Perrier, who has (9) re-examined this genus, has discovered
that there are eight sete in each segment; the position of the
male generative pores of Tritogenia between the 16th and
17th segments (according to Kinberg) is another point of
similarity between the two genera, which are very likely
identical.

No other known genus of Eudrilide has the sete arranged
in this way ; there appears, however, to be a slight difference
in Nemertodrilus griseus in the distance which separates
the individual seta of each couple; this is a step in the
direction of Hy periodrilus.

§ Epidermal Sensory (?) Organs.

The epidermis of this worm is furnished with certain curious
structures of doubtful nature, identical with those which I was
the first to describe in Eudrilus.

As Rosa has indicated the presence of these structures in
Teleudrilus, and as I have found them in Heliodrilus,
they may be regarded as characteristic of the Eudrilide,
and, as far as our present knowledge goes, confined to that

1 The first seta is that nearest to the nerve-cord.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS, 241

family as defined by myself.! This, although a small point, is
an additional argument in favour of retaining that family
within the limits which I have proposed for it.

The structures in question are visible in the epidermis
when the body-wall is examined as a flat preparation in
glycerine ; and they may be observed to be scattered irregularly
over the segments, thus affording an example of another
system cf organs which have no perceptible relation to the
metamerism of the body. In a preparation of this kind the
sensory organs appear as longitudinal furrows, longer than
broad, arranged after no system that I could discover, save
that they were absent upon the intersegmental furrows. In
transverse sections of the body-wall (cf. fig. 3) the sensory
organs are seen to lie in the epidermis, but not to reach its
surface ; they generally cause the membrane which separates
the epidermis from the underlying layer of transverse muscles
to be bulged out towards the latter. Above each body is a
row of short epidermic cells which divide it from the cuticle.
The real form of the sensory bodies is better seen in longitu-
dinal sections, for they lie for the most part parallel with the
long axis of the body of the worm.

In such a section each sensory body is seen (see fig. 2)
to consist of a central cylindrical core faintly stained by
borax carmine, in which are embedded a variable number of
large oval nuclei. Round the axis are a series of coats like
those of an onion, which seem to be composed of an elastic
membrane ; in such sections, and in the transverse sections
also, these coats appear as highly refracting fibres. In frag-
ments of the skin mounted entire and viewed from above, the
membranes present the appearance of a series of fine striz
surrounding the axis of the body. Between the several mem-
branous coats are darkly staining nuclei which are quite
distinguishable from those which lie in the axis by their very
much smaller size ; the central nuclei are fully twice the size
of the peripheral nuclei.

1 In a forthcoming résumé of the classification and distribution of Harth-
worms to be published in the ‘ Proceedings of the Royal Physical Society.’

242 FRANK E. BEDDARD.

These bodies have a striking resemblance to certain “‘ end
organs” which are found among the Vertebrata; they are
particularly like the Pacinian bodies, having the same con-
centric lamelle surrounding a central sheath. It is this
resemblance which makes me believe that the structures in
question are of a nervous nature, for I must confess to having
found no unmistakable evidence of nerve-fibres connected
with them. It is, however, not easy to trace the ramifica-
tions of nerves in the skin of earthworms which have been
preserved with alcohol.!

§ Alimentary Canal.

In its main features the alimentary canal agrees with that
of Heliodrilus, to be presently described.

The pharynx is large, and extends back to the 6th or 7th
segment; there is no anteriorly situated gizzard, but a series
of five or six, each occupying a single segment at the junction
of esophagus and intestine.

The cesophagus is lined with a thin chitinous cuticle as far
back as the opening of the calciferous glands; beyond this
point its walls are ciliated.

§ Calciferous Glands.

Asin Eudrilus, and apparently other Eudrilide, the pre-
sent genus is furnished with two kinds of calciferous glands.

(1) A pair of voluminous glands are attached to the ceso-
phagus in Segment 13, the cavity of which they largely fill.
The cesophagus itself is very narrow in this region, scarcely
wider than the dorsal blood-vessel, which like it is completely
hidden by the large glands. These glands appear of a reddish-
purple colour in the spirit-preserved specimen, the colour
being of course due to the abundant blood-spaces interspersed
among the tissue of the glands. The glands have a trifid
appearance as in Eudrilus, and a large blood-vessel passes
over each.

In transverse sections these paired calciferous glands are

' Since the above was written Dr. Horst in a paper cited on p, 252, foot-
note, has suggested the sensory nature of the problematical structures.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 243

seen to consist of numerous folds of epithelium with blood
lacunee lying between the epithelial layers; the secretion of
the glands has the form of small spherical particles.

(2) The present genus, like other Eudrilide, possesses
impaired ventral diverticula of the cesophagus.

The occurrence of these structures was first put on record
by myself in Eudrilus sylvicola (1), where they are simple
diverticula with the lining epithelium thrown into a few lon-
gitudinally running folds. Rosa (10) mentions the presence
of three such glands in Segments 9, 10, 11, in Teleudrilus,
but gives no special description of them.

To Michaelsen (7) our chief knowledge of these peculiar
glands is due. He terms them “ Chylustaschen,” and com-
pares them with certain glandular diverticula in the Enchy-
treide. The function of these structures is considered by
Michaelsen to be not analogous to that of the calciferous
glands ; instead of secreting calcareous granules like the calci-
ferous glands, or producing any other kind of secretion, it is
supposed that they serve for the absorption of food—hence the
term ‘Chylustaschen.” It must be admitted that no cal-
careous spherules have been found in these pouches, although
their structure recalls that of the calciferous glands; so far
this is evidence, if not in favour of the suggestion of Michael-
sen, at any rate of their performing a different function from
that of the calciferous glands in the economy of the worms.

In Pygmeodrilus the 9th segment contains a pair of lateral
forwardly directed diverticula; it may be that the impaired
glands of Eudrilus are formed by the fusion of a pair which
get to be more and more approximated ventrally : on the other
hand, the diverticula of Pygmeodrilus rather suggest the
calciferous glands of Urocheta, which really secrete calci-
ferous particles.

No such structures are mentioned in Eudriloides or
Nemertodrilus.

In Polytoreutus there are median impaired pouches as in
Hyperiodrilus, three in number, besides the paired calci-
ferous glands; the folds in the interior of the glands are so

244 FRANK E. BEDDARD.

~ complicated as to present the appearance of a bundle of
longitudinally running vessels.

In Hyperiodrilus the ventral cesophageal pouches are in
certain respects more remarkable. There are three of them in
Segments 9, 10, and 11—one pouch to each segment.

The pouch appears to communicate with the cesophagus a
little to the right of the ventral median line. The orifice is
narrow, and the cells are at first identical with those which
constitute the epithelial lining of the csophagus; the cells
are tall and columnar, and between their bases are spherical
or pear-shaped darkly staining cells (fig. 31) like those of
the cesophagus. Tracing the pouch back, the cells are seen
to alter their character, and to become low and quadrangular
in form, while the epithelium is so folded as to give the appear-
ance of a series of tubes running approximately parallel to
each other (fig. 25). At the point of origin of the pouch, as
shown in fig. 81,the muscular wall of the cesophagus and the
peritoneal covering becomes reflected over the gland in such a
way as to leave a wide space between itself and the pouch;
further back this space is obliterated by the coalescence of the
muscular and peritoneal layers with the outer layer of the gland.

Further from the point of the opening of the gland into the
intestine the appearance of parallel running tubes is increased,
and they have become at the same time of smaller calibre.
The cells which form the lining epithelium are broad and
somewhat flat, though quadrangular in form; the nuclei are
thus, owing to the large size of the individual cells, very far
apart, and the cell outlines are not distinguishable. As the
folding gets more and more complicated the “tubes” present
more and more the appearance of having an intra-cellular
lumen; this actually does take place at the extremity of the
pouch. One of the last sections through a pouch is illustrated
in fig. 26: it will be seen there that three comparatively wide
tubules end in a complicated meshwork of fine capillary tubes ;
the section presents the strongest possible resemblance to a
portion of a nephridial network such as I have figured in
Acanthodrilus multiporus. Even the three large tubes

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 245

figured in that drawing appear to be excavated in the substance
of cells, while there can be no possible question about the
ramification of minute tubes.

One is tempted to regard every ductule excavated in the
substance of cells as of nephridial nature, but the origin of the
blood-capillaries in the leech by direct canalisation of cells,
discovered by Lankester and confirmed by Lang, as well as
unicellular glands with a central duct, shows that this conclu-
sion cannot be always drawn.

The study of these cesophageal pouches in Hyperiodrilus
is instructive as showing how folds may gradually acquire the
character of a system of tubes, and how the subdivision of the
tubes, without a corresponding decrease in size of the epithe-
lium, may ultimately lead to an intra-cellular network. This
series of facts shows also how irrational is the distinction,
which some have attempted to draw, between nephridia with
an intercellular duct and nephridia with an intra-cellular duct,
e.g. between those of the Polycheta and Oligocheta.

The tubes are everywhere separated by an abundant plexus
of blood-capillaries, which seems to form a continuous sinus.

In Bucholzia there is a dorsal diverticulum of the ceso-
phagus, which Michaelsen has compared with the ventral
pouches of Eudrilus; and in this Enchytreid the diverticulum
is formed of a nuwber of tubules with intra-cellular lumina.

§ Generative Organs.

1. Male Generative Organs.—I could only find a single
pair of testes; as these were extremely smali, it is possible
that I may have overlooked the second pair. The pair found
were in the 11th segment; each is attached to the vas deferens
just where it issues from the septum separating this segment
from the one in front. As will be seen later, after the other
organs of the male reproductive system have been described,
the peculiar arrangement of the vasa deferentia, which are
bent upon themselves near to their celomic opening, suggests
that the missing pair of testes, if they are really present, will
be found in the 12th segment.

The testes of Hyperiodrilus are not enclosed in a special

2.46 FRANK E. BEDDARD.

sac, as they are in Heliodrilus. The seminal cells must
therefore trust to accident to find their way into the interior of
the sperm-sacs.

There are two pairs of sperm-sacs (fig. 44) in Segments 11
and 12. Hach depends from the anterior of the two septa which
bound the segment by which it is contained. The sperm-sacs
are not very large, and are perfectly independent of each other.
The sperm-sacs are shaped something like a bean, the hilum
being the point of attachment to the septum by means
of a short pedicle. The interior of each sperm-sac is divided
up by numerous trabecule into a series of very small cavities,
which contain decidedly more gregarines than developing
spermatozoa.

The vasa deferentia present close resemblances to those of
Teleudrilus (Rosa, 10). The funnel opens into the sperm-
sac, and therefore traverses the septum twice, since the sperm-
sacs lie on the posterior aspect of the septa separating
Segments 11, 12, and 10, 11 respectively. This is precisely
what occurs in Teleudrilus, and I have recently pointed out
that in a species of Moniligaster there must be something of
the same kind, inasmuch as the funnel projects into the sperm-
sac which is attached to the front wall of its segment. Gene-
rally when the sperm-sacs are attached in this way the funnel
of the vas deferens is not in direct continuity with them, but
projects freely into the interior of the segment a little way in
front of the posterior septum of the segment.

The vasa deferentia open in the way that has been
described by four rather small funnels completely concealed
within the four sperm-sacs. On leaving the sperm-sac the vas
deferens is at first a somewhat narrow tube, lined by numerous
small quadrangular cells, which are of course ciliated; the
peritoneal covering is slight, and there is no such conspicuous
muscular coat as I have figured and described in Eudrilus.
Almost immediately the vas deferens widens out exactly as in
Teleudrilus, and is sharply bent upon itself, and again
traverses the septum; directly it has passed through the
septum it narrows. The wide U-shaped portion of the vas

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 24,7

deferens is lined by a tall columnar epithelium, and the cilia
are very long. The narrow portion lying behind the septum
is composed of an epithelial layer of Jow quadrangular cells
comparatively few in number; the two vasa deferentia of
each side pass down the body just covered by the peritoneum,
and on a level with seta No. 3; they are accompanied by a
blood-vessel which supplies them with capillary branches.

In the 17th segment is situated the terminal apparatus of
the male reproductive organs, which consist of two large
“prostate” glands or atria, opening on to the exterior by
means of the protrusible penis.

Each atrium is furnished with a muscular duct which leads
to the exterior.

The entire gland is sausage-shaped, and recalls the corre-
sponding structure in Acanthodrilus; it has the same form
as in that genus, and the same opaque white appearance.
We do not find the nacreous appearance of these organs in
Eudrilus; the reason for this difference is to be found in .
the absence from Hy periodrilus of the thick muscular coats
composed of longitudinal and circular fibres which are charac-
teristic of Eudrilus. In Hyperiodrilus the muscular
layer is indeed present, but it is reduced, as shown in fig. 88,
to a very thin layer. This figure may be compared with
that illustrating a transverse section through the atrium of
Eudrilus (Beddard, 1, pl. xxx, figs. 8—10).

The interior of the distal part of the prostate is formed of a
compact mass of cells loaded with darkly staining granules,
I could not distinguish (see fig. 42) two layers of cells, such as
appear to be met with in the prostates of all other earthworms
in which those glands have the tubular form which they
exhibit in the present genus; that this, however, is due to the
obliteration of the distinction into two layers by the immense
quantity of secretion present is shown by another specimen
(fig. 38), in which the inner layer of columnar cells was quite
plainly visible.

The vasa deferentia, which retain their distinctness up to
the very point of opening, open into the glandular part of the

248 FRANK E. BEDDARD.

prostate nearer to the distal than to the proximal end. The
direct communication between the vasa deferentia and the
“‘ prostate ” appears to be the rule with Eudrilide ; and this led
me to compare the so-called prostate in the terrestrial Oligo-
cheeta to the atrium of the Limicolous forms. Quite recently
Benham (6) has expressed himself against this identification,
but admits that “‘a portion of the prostate in Pericheta,
Eudrilus, and other genera, in which the sperm-duct and
prostate form is probably the homologue of the ‘atrium’ of
Tubifex.’’ In this case half of the glandular portion of the
terminal organ of the vasa deferentia in Eudrilus, Hype-
riodrilus, &c., will be the equivalent of the “ atrium,” while
the other half will be comparable to the “ cementdriise ” of
Tubifex! This will hardly do, for there is no structural
distinction between the two halves; they form a continuous
whole. I need not recapitulate here the various intermediate
conditions which unite the Eudrilide with other genera; it
appears to me impossible to draw a line between the “ prostate”
glands of Pericheta and those of Eudrilide; they are
obviously homologous structures. The name that is applied
to one must be applied to the other.

There are no penial set present.

2. Female Generative Organs.—These organs seem to
be most like those of Stuhlmannia variabilis, which have
been briefly described by Michaelsen as follows :— “ The
orifice of the spermatheca situated in the middle ventral line
of Segment 13 leads into a wide atrium. From this is reached
an unpaired long sac-like crinkled spermatheca. From the
atrium arises on each side another spermatheca-like broad
canal. These two canals extend upwards and fuse together
above the gut, forming in this way a single short sac, which
communicates with the atrium by a ring-like canal surrounding
the gut. Two greatly coiled oviducts, each furnished with a
receptaculum ovorum, open laterally on Segment 14, On the
other side they communicate with the spermatheca. The two
ovaries lie anteriorly in Segment 13. (They are connected by
narrow canals with the oviducts ?) ”’

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 249

On dissecting a specimen of Hyperiodrilus, the only part
of the female reproductive system that could be at first detected
(fig. 47) was a longish oval spermatheca lying upon the dorsal
wall of the gut, in the 13th segment, and directed backwards.
After carefully removing the calciferous glands, to which the
spermatheca and neighbouring part of the reproductive organs
are closely attached, the spermatheca was seen to divide into
two thick-walled tubes; these (see fig. 1) are placed close to the
septum which divides the 13th from the 12th segment, and
which is conspicuous on account of its being the last of the
specially thickened septa. The two tubes, asin Stuhlmannia,
completely encircle the gut, and meet below in the small
atrium which opens on to the exterior by the median pore
already referred to as existing upon the 13th segment. Each
of these two tubes is provided on the outer side (fig. 8) with
a small prominence, which looks like a diverticulum of it, and
which corresponds to the structure termed by Michaelsen
“yeceptaculum ovorum” in Stuhlmannia; close to this
arises on either side the oviduct, which passes straight to its
opening on the 14th segment.

The atrium is not furnished with a second diverticulum
corresponding to the “ spermatheca” of Stuhlmannia.
Fig. 47 represents the parts described as seen on the dissection
of the worm. In fig. 8 is a more diagrammatic sketch of the
same parts, in order to display their mutual relations, and
their position with regard to the cesophagus and the dorsal
blood-vessel.

An investigation by means of longitudinal and transverse
sections shows that these structures, which are alone clearly
visible on a dissection, and which form a continuous whole
easily to be separated and mounted on a slide, do not represent
the entire female reproductive system.

The ovaries are paired structures, lying as usual in the 13th
segment; they are very compact bodies, not frayed out into
numerous processes. Instead oflying freely inthe cavity
of the 13th segment, each ovary isenclosedinaspecial
celomic sac of a globular form; this sac also includesa

250 FRANK E. BEDDARD.

portion of the nephridium of the 13th segment (fig. 10) ;
its walls are made up of muscular fibrils, and it has a coating as
well as a lining of peritoneal cells. Each sac is situated near
to the anterior wall of the 13th segment; traced backward by
means of a continuous series of transverse sections, the sac
abruptly diminishes in calibre, and forms a narrow tube which
is continuous with the tube formed by the narrowing of the
ovarian sac of the opposite side of the body, A section which
illustrates these relations is illustrated in fig. 51.

The bursa copulatrix, which is spherical in transverse
section, opens on to the exterior by the median pore of Segment
13. From the bursa copulatrix arises a single blind pouch,
which may be regarded asthe spermatheca. This has a lining
of tall epithelial cells of a glandular appearance, and very thick
muscular walls. The spermatheca directly it leaves the bursa
becomes enveloped in a ccelomic sac, as shown in fig. 11; this
coelomic sac is not the connecting tube between the two
ovarian sacs shown in fig. 51, but it is continuous with
the sacinvolving the ovary of its own side; the sperma-
theca of each side runs up the side of the esophagus for a
short way, terminating blindly at about the middle of the dorso-
ventral diameter. The sac in which the spermatheca is con-
tained passes right round the cesophagus, and, fusing with its
fellow of the opposite side of the body, is prolonged backwards as
an unpaired median sac lying above the cesophagus ; this struc-
ture is that which is illustrated in figs. 5, 11, and lettered sp’,

It must be noted, therefore, that what appears on dissec-
tion to be an unpaired spermatheca, lying above the’
esophagus and connected with the bursa by a ring
round the esophagus, is really a celomic sac contain-
ing the true spermatheca, and does not communicate
directly with the exterior through the bursa copula-
trix.

This coelomic space comes into close relations with the ovi-
ducal funnel which seems to open into the receptaculum
ovorum (figs. 1, 5, 8, 11 7. 0.), but does not involve the recep-
taculum or the oviduct.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 2051

The receptaculum, or egg-sac, as it may be more simply
termed, is of considerable size, and is divided up into numerous
compartments, which lodge the developing ova, by trabecule ;
it is closely attached to the pericesophageal ccelomic sac, as
shown in the figure (fig. 10), and in all probability opens into
it, though I confess to not having been able to find the actual
orifice of communication.

The oviduct is a short, straight tube, which passes directly
from its opening into the egg-sac to the external aperture upon
Segment 14. It is lined by a columnar ciliated epithelium,
and it has strongly developed muscular walls.

The celomic sac involving the ovaries, and continuous with
the ring round the cesophagus, facilitates the passage ef ova
from the ovary to the receptaculum and to the oviduct; but
the extension of the sac beyond the limits necessary for that
purpose is a little difficult to understand.

I could not find any spermatozoa in the dorsal sac, nor, for
the matter of that, in the spermatheca; and if impregnation
takes place by the bursa copulatrix, there seems to me to be no
way by which the spermatozoa could reach the interior of the
sac. I could detect no orifice leading from the spermatheca
to the sac which involves them, and such a connection seems
hardly likely to occur.

In nearly all the Eudrilide the ovary is contained in a
special ccelomic sac, which communicates indirectly with the
exterior. The principal exception is Nemertodrilus; but
here the reduction in size of the cavity of the 13th segment
seems to guide the ova to the aperture on to the exterior of
the body, and there is thus no need of the formation ofa special
tube (see p. 266).

The remarkable genitalia of Polytoreutus are perhaps
rendered more intelligible by the facts which I have been able
to make out in Hyperiodrilus. I have little doubt that the
median sac, communicating on the one hand with the
“ spermatheca,” and on the other with the oviduct, will prove
to be a celomic space perhaps surrounding the true sperma-
theca. It seems to be possible that the female organs of

252 FRANK E. BEDDARD.

Teleudrilus will bear reinvestigating from this point of view.
The two tubes which are described by Rosa (10) as diverti-
cula of the spermatheca have a certain likeness to the narrow
celomic sac of Hyperiodrilus, which puts into communica-
tion the ovarian sacs and the space surrounding the sperma-
theca; and they approach each other in the middle line, which
suggests the possibility of their being really fused.

The mature ova have the characters illustrated in fig. 49.
The most remarkable feature is a thick, darkly staining mem-
brane, which completely surrounds the ovum; this membrane
is traversed by numerous pores.

I have described something of the same kind in Eudrilus
(2), where, however, it forms a cap at one end only of the ovum.
After examining Hyperiodrilus it seemed possible that I
was mistaken in considering that the membrane in question
was limited to half of the ovum in Eudrilus. I cannot, how-
ever, find that I have made any mistake in this matter in my
description of Eudrilus.! This being so, it does not seem very
likely that the cap of darkly staining cubical bodies which
cover one pole of the ovum in Eudrilus are comparable toa
radiately striated egg-membrane; such a membrane would
surely be produced round the whole of the periphery of the
ovum at once. There are, therefore, still reasons for adhering
to the opinion which I expressed in the paper dealing with the
structure of that ovum, viz. that the columnar layer is in reality
a product of the follicular cells, being formed by the metamor-
phosis of a certainnumber ofthem. This opinion is considered
by Vejdovsky (12) to be probably true.

The resemblance between this structure in Eudrilus and
the complete membrane which surrounds the ovum of
Hyperiodrilus is close; but I do not feel sure that they
actually correspond ; a membrane surrounding one half of the
ovum seems to be exceedingly anomalous. The ova within

* My description has been confirmed in a paper by Dr. Horst which I
received after the present memoir was sent to Professor Lankester, ‘Sur
quelques Lombriciens exotiques appartenant au genre Dudrilus,” ‘ Mém. Soc.
Zool, de France,’ t. iii, p. 223 (ef. fig. 11 of plate).

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 253

the compartment of the egg-sac are associated with a few
germinal cells, which are generally closely attached to the
ovum, and frequently show signs of degeneration. The masses
of immature germinal cells and ova, in all stages of develop-
ment, which I have figured and described in Eudrilus (2)
are not to be found in the present genus.

II].—Heliodrilus lagosensis, nov. gen., n. sp.}

Among a number of earthworms which arrived in a Wardian
case at Kew Gardens from Lagos, West Africa, was a single
specimen which I refer to a new genus, belonging, like Hyper 1-
odrilus, to the family Eudrilide.

§ External Characters.

It is of about the same size and colour as Hyperiodrilus,
but the external characters of the specimen when killed and
preserved are quite unlike those of the former species.

The prostomium is of the same form as that of Hy perio-
drilus.

The sete have precisely the same arrangement as in
Hy periodrilus—that is to say, the ventral couple are at some
little distance from each other, while the lateral couple are very
closely approximated.

The clitellum was not very distinctly marked, but appeared
to comprise four segments, 14—17.

Dorsal pores could not be detected.

The nephridiopores are placed in front of the lateral setz.

The oviducal pores, as in all Eudrilide, are lateral in
position ; they are upon the 14th segment, and are quite con-
spicuous, owing to their being surrounded by a slightly raised
margin.

The male generative pore is unpaired and median in
position ; it lies upon the border-line between Segments 17 and

18: the pore is situated upon the summit of a prominent
elevation.

1 The generic name might be confused with Helodrilus if there were
any chance of that problematical form ever being properly identified.

254, FRANK E. BEDDARD.

The spermathecal orifice was not visible upon the
exterior; by means of transverse sections it was found to be
upon Segment 11.

The most characteristic external mark of this species is
afforded by a series of sucker-like structures. There are
six of these, one to each of Segments 10O—15. The last three
are accurately median and ventral in position, and are situated
on about the middle of their segment. The two in front are
placed considerably to the left of the middle line, as shown in
fig. 21; the first is nearer to the middle line.

§ Integument.

The epidermis of Heliodrilus agrees in every particular
with that of Hyperiodrilus, as does also the structure of
the clitellum. The limits of this modified region of the
body-wall could not be ascertained with certainty by an inspec-
tion of the worm; transverse sections show that it extends over
Segments 14—17.

The peculiar sensory organs which are so characteristic of
the Eudrilide occur in Heliodrilus; their structure calls for
no remark, as they resemble in every particular those of
Hyperiodrilus, which have been already described on p. 236.

The muscular layers of the integument are also identical in
every respect with those of Hyperiodrilus.

§ Alimentary Tract.

The buccal cavity occupies the first three segments.

The pharynx extends back to the 6th segment.

The esophagusis very narrow, and passes back without any
change as far as the 10th segment ; here it becomes narrower,
and the lining epithelium is thrown into a series of regular
longitudinal rugze ; at the mesentery before the 11th segment,
as in the case of the subsequent segments, it becomes a trifle
wider, recurring to its former dimensions, which are between
one third and one fourth of the diameter of the body of the
worm in this region. The first of the two ventral esophageal
pouches lies in this segment, and is directed forwards, having
anteriorly no connection with the wall of the cesophagus;

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 255

further back it is suspended by two closely approximated and
very delicate mesenteries, which later become continuous with
a bridge of tissue, along which blood-capillaries pass from the
pericesophageal blood-sinus to the vessels of the pouch. This
bridge forms the walls of the aperture of communication
between the cesophagus and the pouch. The minute structure
of the pouch is as in Hyperiodrilus, but there is no splitting
of the muscular layer of the cesophagus at the origin of the
pouch, such as I have figured and described in that earthworm.

The second pouch, which lies in the 12th segment, has a
precisely similar origin from the cesophagus, and is identical in
all respects with the first.

In the next segment is a third cesophageal pouch, which is
very much smaller than either of the other two ; its interior is
not so subdivided by the development of folds, and the aperture
into the cesophagus is distinctly larger ; it has the characters
rather of a folded-off portion of the esophageal tube than of a
diverticulum.

After this the tube remains narrower for some distance, with
the epithelium longitudinally folded; it is here, as throughout
its whole extent up to this point, lined by a thin chitinous
layer; there is, however, no gizzard upon the eso-
phagus, and no special thickening of its muscular
coat which could be compared to a gizzard.

In the 14th segment the cesophagus becomes wider, and
receives the ducts of the calciferous glands; these ducts have
exactly the same structure as the cesophagus, and are not of a
very greatly inferior calibre. Their epithelium is dis-
tinctly ciliated; each duct opens by a wide aperture on to
the side of the esophagus. After the opening of the ducts of
the calciferous glands the epithelium of the esophagus
alters its character and becomes ciliated. The dia-
meter of the tube is at the same time larger, and the plexus of
blood-vessels more richly developed. Commencing with the
18th segment the alimentary canal is provided with six gizzards,
one to the 18th and to each of the five foliowing segments,
conuected by sections of thin-walled intestine,

VOL. XXXII, PART II1.—NEW SER. R

256 FRANK E. BEDDARD.

§ Body-cavity.

As is usual in earthworms, the first few segments are not
separated from each internally by regular septa; irregular
fasciculi of muscles attach the buccal cavity and the pharynx
to the parietes. The 5th segment is separated from the 6th
by the first regular intersegmental septum; this and the 7th
which follow are specially thickened, and consist of several
layers of muscular fibres. These septa (fig. 48) fit into each
other like a series of cups ; they arise from points which corre-
spond accurately with the intersegmental furrows.

The ccelom in this worm is further broken up into the com-
partments which lodge the testes and extremities of the vasa
deferentia, and into those which lodge the ovaries and the
spermatheca ; finally a special sac encloses the supra-intestinal
blood-vessel. All these coelomic chambers are specially
described under the different organs which they enclose.

As in other Oligocheta (cf. Kiikenthal 13), the coelomic cor-
puscles are of two kinds.

§ Vascular System.

The supra-intestinal vessel exists in Heliodrilus, and
is connected with the hearts, as Perrier was the first to point
out in some other genera.

In Heliodrilus this vessel is small, and lies in a special
coelomic space above the gut. A transverse section through
the supra-intestinal vessel and this perihzmal sac is illustrated
in fig. 46. The sac has exceedingly thin walls, which are con-
nected with the walls of the blood-vessels by irregularly dis-
posed trabeculz ; the interspaces are largely filled with cor-
puscles which have the characters shown in the figure; these
cells are quite similar to those which occur in the walls of the
intestine. The inclusion of the supra-intestinal vessel in a sac
recalls the similar periheemal space which I have described in
Deinodrilus (4) as surrounding the dorsal blood-vessel.

The large “hearts” of Segments 11—13 communicate
with the supra-intestinal as well as with the dorsal vessel. The

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS, 257

supra-intestinal vessel does not, as it does in Eudrilus, become
double above each of the ventral cesophageal pouches.

§ Generative Organs.

(1) Female Organs.—As in most of the genera belonging
to the family Eudrilide, there is a complicated system of
coelomic spaces developed in connection with the ovaries and
the other organs belonging to the reproductive system.

The ovaries are paired, and in Segment 13. Each ovary is
enclosed in a sac which it almost completely fills; a narrow
tube running dorsal to the nerve-cord connects the ovarian
sacs of the two sides of the body ; there is further a communi-
cation between the ovarian sac and the egg-sac of its own
side, as in Teleudrilus and Hyperiodrilus: this communi-
cation is effected by a celomic tube which is at first very
narrow ; as it approaches the egg-sacs it becomes wider, and
finally forms a somewhat oval sac enclosing the funnel of the
oviduct and communicating with the egg-sacs, into which the
oviducal funnel also opens. So far as I can make out from a
complete series of transverse sections the arrangement is, so
far, very like that which has been figured and described by
Rosa in Teleudrilus (10); but Heliodrilus apparently
differs from Teleudrilus, and certainly agrees with Hy perio-
drilus in the communication between the right and left
ovarian sacs. I found it quite easy to trace the course of the
tube which connects the ovarian sac with the considerable
space surrounding the funnel of the oviduct ; but any doubt as
to the reality of this connection is removed in the present in-
stance by the occurrence of ova floating freely in the wide
space round the funnel; for the most part these ova were to
be observed singly, each surrounded by a follicular layer of
flattened cells, of which the nuclei alone were conspicuous: in
a few cases the ova were also surrounded or partially surrounded
by groups of germinal cells, as a rule comparatively few in
number. The ova in the ovary, as well as those which I found
in the sinus surrounding the funnel of the oviduct, had a well-
developed vitelline membrane, but showed no traces of the

258 FRANK E. BEDDARD.

remarkable striated membrane which I shall refer to directly
in describing the ova within the egg-sacs.

The oviduct has been incidentally referred to in the fore-
going description ; it opensinto the egg-sac and intothe ceelomic
space continuous with the perigonadial sac; it is a short tube,
and passes straight to its opening upon the 14th segment ; it is
not twisted upon itself, as is the oviduct of Eudrilus. The
oviduct has fairly thick muscular walls, the fibres of which are
for the most part arranged in a series of rings round the tube,
and a lining of columnar-ciliated cells. The calibre of the
oviduct diminishes gradually from the funnel to the external
aperture.

The egg-sacs are also situated in the 14th segment: the
septum dividing this segment from the one in front is entirely
or largely absent; but the position of the egg-sacs within the
14th segment suggests that they lie near to where the anterior
wall of that segment should be. The interior of the egg-sacs
is divided up by trabecule anastomosing with each other into
a series of very small compartments, only just broad enough
to contain a single ripe ovum; the compartments, as in other
earthworms, are lined with small peritoneal cells (see fig. 29).

The mature ova do not present any noteworthy differences
from those of Hyperiodrilus.

Spermatheca.—As in Hyperiodrilus, there is only a
single spermatheca present, which lies on the right side of the
body—the opposite side, therefore, to that which the sperma-
theca occupies in Hyperiodrilus. ‘The spermatheca in
Heliodrilus is a large conspicuous organ, which can be seen,
on a dissection of the worm, to reach on to the dorsal side of
the gut; it contrasts, therefore, with the very small sperma-
theca of Hyperiodrilus. As in the latter worm, the appa-
rent bulk of the spermatheca is increased by a prolongation of
the perigonadial sinus which partially surrounds it; but the
arrangement of this sinus in Heliodrilus is very curious, and
quite unlike that of Hyperiodrilus. But before describing
the sinus I will direct attention to the characters of the
spermatheca itself, which differs in certain points from the

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 259

spermatheca of any other Eudrilid, or in fact any other earth-
worm at present known.

It is a large oval sac lined by columnar cells; a portion of
one of the walls is represented highly magnified in fig. 37:
below the layer of columnar cells are some smaller cells, the
contours of which are not very clear, though their nuclei are;
outside these are a few muscular fibres, which make up a layer
of no great thickness. The interior of the spermatheca con-
tains a granular substance which appears to be formed by the
columnar cells. The calibre of the spermatheca (fig. 41)
gradually diminishes towards the apex and towards the ventral
side of the body ; here the cells lose their glandular character,
and become at the same time considerably shorter, so that the
muscular coat appears to acquire an additional thickness.
The narrow duct of the spermatheca does not open upon the
13th segment as in Hyperiodrilus, but bends under the
nerve-cord and runs forwards, always lying beneath the nerve-
cord, as far as the 11th segment; throughout the whole of its
course beneath the nerve-cord it is a narrow tube with thick
muscular walls, and a lining of short columnar cells, which, it
is perhaps unnecessary to remark, show no traces anywhere of
cilia. The diameter of the spermathecal tube in these seg-
ments is about equal to that of the nerve-cord. In the 11th
segment the spermathecal tube perforates the body-wall, and
opens on to the exterior by an inconspicuous orifice which is
situated on the median ventral line. The ventral sucker-like
organ of this segment is pushed to one side, as shown in fig.
21, and does not therefore interfere with the accurately median
position of the spermathecal pore.

The number of segments occupied by the spermatheca is
thus considerably in excess of that which is found in any other
earthworm.

If the spermatheca is developed in the Eudrilide as in the
Lumbricide by an inpushing of the epidermis, the point of
opening will fix the morphological position of the organ ; hence
Heliodrilus serves in this respect to connect the Eudrilide
with other earthworms, for the spermatheca opens in front of

260 FRANK E. BEDDARD.

the ovarian segment, and yet the main part of the organ lies in
that segment.

The apex of the spermatheca cannot be seen on a dissection
of the worm, for the reason that it lies embedded in a ccelomic
sac. Figs. 32—34 represent a series of sections showing the
relations of the spermatheca to this sac. The sections are part
of a series running from behind forwards; towards the poste-
rior end of Segment 14 the end of the spermatheca is
seen to lie between two cclomic spaces, which are really
continuous, and envelop the apex of the spermatheca as seen
a few sections later; in this section the extremity of the
spermatheca is seen in transverse section to lie in the middle
of the coelomic sac, which is incomplete dorsally: this is,
however, merely due to an accidental cut; the sac is really
closed. Round the spermatheca is a mass of tissue which is
seen in a later section to be the wall of a second sac lying
within the first. The spermatheca is pushed against the
wall of this, driving it before it for a little way, but it hardly
enters the second sac: fig. 34 is therefore a little exaggerated in
this particular ; the cavity of the spermatheca does not appear
to be continuous with that of the second sac, although I should
have preferred longitudinal sections to decide the point ; in any
case the character of the lining cells is absolutely different.
This second sac which lies within the first is also closed ;
it has the same general structure, consisting of two layers
of peritoneum, between which are a few fibres of what
appears to be muscular tissue; but the lining peritoneum, as
shown in fig. 33, is very much thicker, and the cells are larger
and rounded.

In both sacs masses of corpuscles lie here and there within
the lumen.

The outer coelomic sac gradually narrows ventrally, and ulti-
mately becomes an extremely narrow tube, which is attached to
the spermatheca by the mesentery; it finally becomes continuous
with the perigonadial sinus. The general disposition of the
female reproductive organs and of these coelomic sacs connected
with them is shown in asemi-diagrammatic form in fig. 41. In

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 261

reconstructing this figure from the transverse sections I have
put in the intersegmental septa between Segments 12,
18, and 13, 14, which I have not actually observed; I
imagine that they will be found to be partially absent, as in
Hyperiodrilus. The position of the coelomic sacs is also
not quite as in nature; they have been slightly altered to
permit of everything being seen in one figure : but these altera-
tions do not affect the mutual connection of the various parts ;
these are, I trust, accurately displayed in the figure, and also
in fig. 36, which represents the apex of the spermatheca (sp.).
Comparing the arrangement of these parts in Heliodrilus
with that of Hyperiodrilus we find an increase in size of the
spermatheca, and a decrease in size of the ccelomic sac involving
the spermatheca. The latter, instead of forming a complete
ring round the csophagus completely enclosing the sperma-
theca on one side, is only developed on one side of the body,
and surrounds only the extremity of the spermatheca. The
second sac, lying within the dorsal dilated part of the sac which
surrounds the extremity of the spermatheca, is peculiar to
Heliodrilus, and is an extraordinary structure, concerning
the meaning or function of which I can offer no suggestion.
The enormous development of the lining cells of the circum-
cesophageal sacs of Hyperiodrilus into the similitude of a
glandular epithelium is not found in Heliodrilus. It is pos-
sible, however, that it is a periodical occurrence which does not
happen to have taken place at the time when the only specimen
of Heliodrilus that I possess was killed. The reduction of
the coelomic sac surrounding the spermatheca not only in size,
but also in the extent to which it involves the spermatheca,
culminates in Eudrilus, where, as far as I can see, there is no
vestige of any perispermathecal sac left. Ishould like to make
this point quite certain, but in any case it is evident that if
such a sac does exist, there must be merely traces of it. The
question is whether Eudrilus represents the last term in this
series of modifications, or whether Hyperiodrilus does;
from what we know of the anatomy of the Oligochzeta it seems
more reasonable to suppose that the development of these

262 FRANK E. BEDDARD.

ceelomic spaces is secondary. Accordingly, as regards the
Eudrilide, Eudrilus represents perhaps the most archaic form.
This is also, it may beremarked, in accordance with the fact of its
geographical distribution. It occurs in South America, the
West Indies, St. Helena, New Caledonia, and New Zealand,
being therefore one of the most widely spread of earthworms.
It seems, therefore, permissible to argue from Eudrilus to
the other Eudrilide; and this I have attempted to do in the
case of the ovarian oviducts and atria (see p. 268). It would
be interesting to ascertain how far the spermathecal coelomic
spaces are represented in Teleudrilus; the connection
between the perigonial sinus and the egg-sac is developed in
that genus as in Heliodrilus and Hyperiodrilus, but not
Eudrilus.

2. Male Generative Organs.—The testes are, asusual,
paired structures which lie in the 10th and 11th segments.

Each testis has a number of processes of unequal sizes, as
is so very generally the case with earthworms. The testes,
however, do not conform to the general rule in their position ;
they lie near to the posterior septum of their segment, as in
Acanthodrilus annectens (4), alone among earthworms at
present known.

Furthermore, each testis, instead of lying freely in the
celom, is surrounded by a small sac, which is only large
enough to contain the testis (see fig. 15); this sac is attached
to the lateral parietes some way above the nerve-cord by a thin
mesentery (fig. 15, mes.), and directly to the septum which
divides its segment from the one following. This sac has for
its size tolerably thick muscular walls, and of course a lining
as well as a coating of peritoneal cells.

The vasa deferentia have the same curious arrangement
that Rosa was the first to describe (10) in Teleudrilus, and
which I have already mentioned as occurring in Hyperio-
drilus; each vas deferens perforates septa 10, 11, or 11, 12,
and then, passing back, again perforates them to reach the in-
terior of one of the sperm-sacs which depend from the BuHveiey
surface of these septa.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 2638

But in Heliodrilus there are some differences of detail.
The expanded portion of the vas deferens, which lies in front
of septa 10, 11, and 11, 12, is proportionately much larger,
while the narrow neck by which it is connected with its funnel
is much longer than in Hyperiodrilus.

Furthermore, the expanded portion lies in the coelomic space
which contains the testis, or rather it is closely invested by a
narrow space (figs. 11, 16, and 18, ccelom), which is perfectly
continuous with that enclosing the testis. Sections show that
there is no real demarcation between these spaces, which form
on each side a pair of sacs—one larger enclosing the vas
deferens, and one smaller containing the testis. It should be
mentioned that the vas deferens itself on the posterior side of
each of the septa lies quite freely in the ceelom.

The dilated region of the vas deferens is quite similar in
structure to the rest of the tube; it is lined by a layer of
low columnar ciliated cells, and is invested by a sheath con-
taining a few muscular fibres. The interior, as in Eudrilus,
was filled with loosely lying spermatozoa, not compacted
together in any way.

The proportions of this dilated sac to the cesophagus are
shown in fig. 11, which is copied from a drawing made by
the help of the camera lucida. The details of structure and
the proportions of the investing sac are more clearly shown in
fig. 35.

Atria.—I term the structures in question “ atria’’ rather
than “ prostates” for the reason that I have given elsewhere (4),
and recapitulated briefly on p. 248 of the present paper. They
form a pair of long tubes, which were disposed as follows in
my specimen. On the right side of the body the atrium, which
opens, as already stated, on to the 17th segment, passed straight
back as far as the 24th segment; it was then sharply bent
back upon itself, and again reached the 17th segment, where it
ended blindly. On the left side the atrium was entirely con-
tained within the 17th, 18th, and 19th segments, being much
folded (fig. 18).

A dissection of the worm did not show any differentiation of

264 FRANK FE. BEDDARD.

the atrium into a glandular and an efferent section, such as
is found in Hyperiodrilus andin other earthworms. There
is no such sharp demarcation between the highly glandular
more distal part of the atrium and the duct which perforates
the body-wall, and is lined with simple columnar epithelium.

The glandular part of the atrium has the typical structure,
although the demarcation between the two layers of cells was
obscured by the great abundance of secretion; the inner layer
conld be only detected by the presence of a regular row of
nuclei. Towards the external aperture the character of the
lining epithelinm gradually alters: this alteration affects, in
the first place, the thickness of layers; they become gradually
thinner, until not far from the external orifice there is but a
single layer of cells. The cells in this layer (fig. 39) are of
two kinds—large swollen glandular cells with a scanty amount
of protoplasm lie embedded in a mass of narrow columnar cells ;
when the tube enters the body-wall on its way to the exterior
the glandular cells disappear, and there are only columnar cells
present of a non-glandular character.

The vasa deferentia, as in Hyperiodrilus and
Eudrilus, open into the glandular portion of the atrium,
which is, I should remark, not divided into two chambers
bound up in a common sheath, as it is in Eudrilus. The
two vasa deferentia retain their distinctness, and open into
the atrium (fig. 42) at some little distance from each other.

The glandular part of the atrium has a very thin muscular
sheath; this becomes thicker towards the external orifice,
where it is plainly divisible into two layers—an outer layer of
circular fibres, and an inner layer of longitudinally running
fibres. The layers, however, are even here very thin, and do
not consist of more than two rows of fibres. The two atria
traverse the body-wall independently of each other, and unite
at the bottom of a tubular depression which communicates
directly with the exterior. The structure of the parts is such
that it does not appear to be capable of eversion as a penis,
There are no penial sete present. The body-wall is consider-
ably thickened in the region of the atria for some distance on

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 265

either side of the ventral line; the thickening ceases at the
first seta.

A curious fact in relation to the atria is the connection of
the nephridia with these organs. In the other segments of the
body the nephridia open in front of the lateral sete; but in
the 17th segment the duct of the nephridium traverses
the ventral body-wall in an oblique direction, and
joins the atrial duct just before its opening into the
external depression already spoken of (fig. 40). As this
occurred on both sides of the body it does not seem likely to
be an abnormal condition characterising the individual only.

§ Summary of more important facts in the structure of Helio-
drilus and Hyperiodrilus.

(1) The epidermis is furnished as in other Eudrilide, and in
that family alone, with peculiar organs possibly of a sensory
nature ; these consist of a central nucleated core surrounded
by many nucleated sheaths; the organs have a certain resem-
blance to Pacinian bodies of Vertebrates, and are scattered
irregularly over the surface of all the segments save the first.

(2) The alimentary canal has a single pair of large lobed
calciferous glands in the 13th or 14th segment; in each of
Segments 10, 11, 12, 1s a median diverticulum of the ceso-
phagus, of which the epithelium is much folded, so that it
presents the appearance of a series of parallel tubes ; peri-
pherally the cells themselves are excavated, and form a rami-
fying system of ductules ; the pouch of Segment 10 is smaller,
and the foldings are much simpler and do not anastomose.
There is no anterior gizzard, but six gizzards, one to each
segment, at the junction of the esophagus with the intestine.

(3) The supra-intestinal blood-vessel in the cesophageal
region is enclosed in a special coelomic compartment, which is
almost filled by deeply staining nucleated corpuscles.

(4) The male genital pore is single and median upon
Segment 17. The “atria” are glandular and very long; in
Heliodrilus the vasa deferentia open into them in both
genera. In neither are there any penial sete, but in Hype-

266 FRANK E. BEDDARD.

riodrilus there is a penis, which is a hollow process of the
body-wall. In both genera the vasa deferentia open in the
11th and 12th segments into the interior of the sperm-sacs ;
each vas deferens perforates the septum, from which the sperm-
sac depends, twice.

(5) The ovaries are enclosed in special ecelomic sacs which
communicate with the egg-sac, and are prolonged dorsally so
as to entirely (Hyperiodrilus) or partially (Heliodrilus)
enclose the single spermatheca which opens on the middle line
of the 138th (Hyperiodrilus) or llth (Heliodrilus) seg-
ment; in the latter case the spermatheca itself lies in the 13th
segment, and has a long duct. In Hyperiodrilus the peri-
gonadial sacs form a ring round the cesophagus, and are con-
nected with a dorsal unpaired sac.

III.—Some Notes upon Nemertodrilus griseus, Mich.

The principal points in the anatomy of this Eudrilid have
been already made out by Michaelsen; there are, however, a
few facts of minor importance which I have been able to note
down from the examination of specimens which Dr. Michaelsen
was so good as to place at my disposal. With regard to the
female reproductive organs, I can only confirm the accurate
description given by Michaelsen; but the structure of these parts
suggests certain reflections concerning the homologies of the
various organs which constitute the female reproductive system
in the Eudrilide. As Michaelsen has stated, the cavity of
Segment 13 is greatly reduced, so that the ovaries are enclosed
in a narrow chamber ; the receptaculum ovorum communicates
with this segment, and is also connected with a large pouch
extending on each side of the body through several segments.

Michaelsen suggests that these large pouches may be possibly
the equivalents of the receptacula and the distal part of the
spermatheca. ‘The proximal part of the spermatheca on this
supposition is represented by a pair of orifices, of which more
later, opening into the exterior of the body from the reduced
cavity of the 13th segment. Dr. Michaelsen decides, however,

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS, 267

that most probably the sacs in question are simply extensions
of the egg-sacs, inasmuch as there is no definite break between
them and the egg-sacs; the trabecule which divide up the
interior of the egg-sac into a series of more or less isolated
compartments, as in other earthworms, get gradually less and
less until the cavity becomes perfectly smooth.

The paired orifices upon the 13th segment are considered
by Michaelsen to represent the rudiments of spermathecee
which open into this segment in other Eudrilide.

These orifices are fringed with numerous frayed-out cellular
processes, which would appear to be of the nature of prolifera-
tions of the peritoneum.

The effect of these must be analogous to the twigs of a
lobster-trap ; they would prevent egress from the interior of
the segment, but would permit the penis to be thrust in. It
seems most likely that these orifices are used in copulation ;
the sperm can then readily find its way into the interior of the
egg-sacs; and as a matter of fact I can fully confirm
Michaelsen’s statement that bundles of spermatozoa are found
in the interior of these sacs.

The large sacs which extend from the 14th to the 17th seg-
ment seem to me to be in all probability equivalent to the
coelomic sacs which I have just described in Hyperiodrilus
as encircling the cesophagus and fusing above it to form a
large unpaired sac. I consider that Michaelsen is quite right
in deciding that they do not represent a portion of the two
spermathece cut off from the duct leading to the exterior;
but, on the other hand, I regard my own identification
of these openings as a little more probable than that of
Michaelsen.

The relationship of the egg-sacs to the ccelomic sacs in this
genus and in Hyperiodrilus is something like that of the
sperm-sacs in Dichogaster and other worms to certain sacs
connected with the testes and the funnels of the vasa deferentia.
It is, perhaps, worth remarking that the connection of these
sacs above the intestine is curiously paralleled by the connection
of the ovarian sacs in Hyperiodrilus. Next, with regard to

268 FRANK E. BEDDARD.

the paired orifices upon Segment 18, Michaelsen considers
these to be the remains of the spermathece which open on to
this segment in Eudriloides, Teleudrilus, and other
Eudrilids. This identification cannot be regarded at present
as being anything more than possible. The spermathece of
Lumbricus are developed as involutions of the epiblast, and
if the course of development of the spermathece in the Eudri-
lide is the same, it is not likely that they could ever come to
be represented by pores.

Considering the matter necessarily in the absence of any
knowledge of the development of the parts in question, it
seems possible to regard these pores as the rudimen-
tary equivalents of oviducts.

In describing the structure of Eudrilus I have pointed out
that there are apparently two pairs of oviducts present in that
worm. One pair, represented in all other earthworms, open
on to the 14th segment; the other pair are short tubes con-
nected with the sac involving the ovary in Segment 13; they
Open in common with the spermatheca and the other oviducts
on to the 14th segment.

These peculiarly modified organs in Nemertodrilus are
quite intelligible on the hypothesis that they have been
derived from the corresponding organs of Eudrilus.

I pointed out that there was some evidence in favour of
regarding the oviducts of Segment 13 as being in course of
degeneration ; they are very short, with feebly developed walls,
and the lining epithelium is not ciliated. This reduction is
carried further in Nemertodrilus; the oviducts are reduced
to the condition of the oviducts in the Enchytreide, where there
is little more than a pair of orifices. At the same time the
coelomic cavity of the segment is greatly reduced ; this renders
it easy for the ova to reach the exterior through the oviducts
of the 14th segment, which are apparently as well developed
as in Eudrilus.

In Teleudrilus there is no trace (?) of the oviduct of Seg-
ment 13, and, except for the continuity of ovarian sac with
the receptaculum, the female reproductive organs of this genus

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 269

are not far removed from those of the more typical Oligo-
cheeta.

It is perhaps possible to regard the pores in Nemerto-
drilus as the earlier condition. Observations upon the abdo-
minal pores and oviducts of certain Ganoids and Teleosteans
seemed at one time to indicate that the pores were the
primitive structures, and that a groove in the peritoneum,
converted later into a tube, connected these pores with their
respective gonad. But if Jungersen! is right in looking upon
the oviducts of Teleostei as being after all Miillerian ducts,
this view must fall to the ground. Sedgwick? brings forward
many facts towards proving that in Peripatus the genital
ducts are coelomic sacs communicating with the exterior by
pores. The ovary in Nemertodrilus, closely invested by the
ccelom which opens on to the exterior by a pore, is comparable
to the gonad and its duct in Peripatus; it is possible, as I
have already suggested, that in Eudrilus the oviduct is only
a portion of the ceelom connected with a pore; but I am more
disposed to hold that Nemertodrilus is in these points a
degenerate form of Eudrilus; the reduction of the ccelom of
Segment 13, and the disappearance of the spermathecz bears
out this view.

In any case two pairs of oviducts seem to imply two
pairs of ovaries, which are reduced to one by the disappear-
ance of the anterior oviducts. And we thus arrive at the
normal condition of the female reproductive organs in earth-
worms.

In Teleudrilus the complete disappearance of one pair of
oviducts is correlated with the disappearance of the second
pair of ovaries.

In short, the new facts discovered by Dr. Michaelsen lend
support to the conclusions which I formulated in the paper
referred to above, apart altogether from the question as to the

1 “Beitrage zur Kenntniss der Entwickelung der geschlechtsorgane bei
den Knochenfischen,” ‘ Arb. Zool. Zoot. Inst.,’ Wiirzburg, 1890.

2A Monograph of the Development of Peripatus capensis,”
‘Studies Morph. Lab. Cambridge,’ vol. iv, part 1.

270 FRANK E. BEDDARD.

primitive or non-primitive condition of the reproductive system
in Eudrilus.

The atria present the appearance of the corresponding
organs in Acanthodrilus or Pontodrilus—that is to say,
they form two somewhat bent tubes of an opaque white
colour; they differ, however, in the fact that it is impossible
to distinguish a muscular and a glandular portion: in the two
genera mentioned, and in many other forms in which the atria
are tubular, the organ communicates with the exterior by a
narrow duct; this duct is lined with an epithelium which is
not in the least glandular, and is surrounded by a tolerably
thick muscular coat. In Nemertodrilus no such duct is
present; the organ is identical in structure throughout with
the glandular part in Acanthodrilus: its epithelium is of
two kinds; the innermost layer is formed by a single row of
unusually short columnar cells; beneath these are the usual
layers of pyriform gland-cells, each with a long slender pro-
longation which reaches, or nearly reaches, the lumen of the
gland.

As Perrier (8) first remarked, the atrium of Eudrilus is
remarkable on account of its nacreous appearance and per-
fectly straight course. The nacreous appearance is due to an
enormously thick muscular coat, which I figured in transverse
section in a paper dealing with the structure of Eudrilus
sylvicola (1). .

In Nemertodrilus, as I have already implied by compar-
ing the appearance of the organs to that presented by the
atria of Acanthodrilus and Pontodrilus, the nacreous
appearance is entirely wanting. Sections of the atrium,
however, show that the muscular coat itself is not absent,
but is greatly reduced as compared with Eudrilus and, ac-
cording to Rosa’s observations, Teleudrilus. The whole
organ, in fact, is a little more degenerate than that of Eu-
drilus. Considering the absence of the duct, which is
so universal a feature of the atrium among earthworms, I
should be disposed to regard the atrium of Nemertodrilus
as having been derived from that of Hudrilus by reduction,

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 271

and not vice versa. There is at any rate nothing in the facts
which is opposed to this view, though the converse might be
asserted with some probability.

As Dr. Michaelsen has pointed out, the two vasa deferentia
of each side retain their distinctness, but are accurately super-
imposed, thus giving rise to the impression that but one tube
is present.

A curious peculiarity in the vasa deferentia of Hudrilus,
which appears to be confined to that genus and to other
Eudrilide, was first pointed out by myself. In those genera
each vas deferens has, like the atrium into which it opens, a
well-developed muscular tunic; each vas deferens, moreover,
in those very aberrant earthworms commences with a very
wide dilatation immediately connected with the funnels.

In both these points Nemertodrilus differs from its two
allies—the vas deferens has neither the oval or spherical di-
latation nor the muscular coat; it conforms, in fact, in every
particular to the usual type met with among earthworms.

Michaelsen mentions that the two vasa deferentia of each
side, maintaining their distinctness to the very last, become
lost in the body-wall just in front of the atria.

This is undoubtedly the impression which a dissection of
the Annelid produces, but it is not perfectly accurate. The
point is one of some little importance as touching the affinities
of Nemertodrilus to Eudrilus.

In the latter genus I showed that the vasa deferentia opened
into the atrium at about the middle of its length. In Teleu-
drilus Rosa has stated that the vasa deferentia also open into
the atrium.

In Nemertodrilus a series of longitudinal sections shows
that the two vasa deferentia cross the atrium close to its ex-
ternal aperture ; they then traverse the muscular coat exactly
as in Eudrilus, and each may be recognised still preserving
its independence as a ciliated tube lying between the epithelial
lining and the muscular coat. They finally open into the
interior of the atrium.

Michaelsen has pointed out two characters in which Nemer-

VOL, XXXII, PART Il.—NEW SER. 8

272 FRANK E. BEDDARD.

todrilus differs from the Eudrilide: the first is the position
of the nephridiopores, which are situated in front of the ven-
tral instead of the lateral setze; the second character is the
absence of any glandular diverticula to the alimentary tract.

To these I can add a third character—the absence of those
integumental bodies which occur in all other Eudrilide that
have been sufficiently well examined.

The structure of Nemertodrilus shows that it is deci-
dedly an aberrant member of the family Eudrilide. But the
alterations in structure from the typical Eudrilide do not de-
finitely point in the direction of the Cryptodrilidz, with which
family, as I distinguishit, Rosa unites the Eudrilide.

The only reasons for referring this genus to the Eudrilide
are (1) the absence of spermathecz lying in front of the testes,
(2) the completely separate vasa deferentia opening into the
interior of the atrium, (3) the large ccelomic sacs connected
with the egg-sacs, (4) the muscular oviduct.

The absence of any specialisation into the atrium, of dilata-
tion upon the vasa deferentia, of integumental organs, are
perhaps indications of degeneration. The fact that the ovary
is not, as is the rule among the Eudrilide, enclosed in a special
compartment of the celom, might be used as an argument for
the primitive position of Nemertodrilus among the Eudri-
lidee were it not for the reduction of the cavity of the thirteenth
segment. The reduction of this segment renders the develop-
ment of any such sacs unnecessary, though of course it does
not necessarily follow that they were originally present and
have been lost. On the other hand, the pores upon Segment 13,
whether Michaelsen’s explanation of them or mine be correct,

seem to be in all probability rudimentary structures of some
kind.

Lonpon; September 9th, 1890.

12.
18.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 273

List oF MEMOIRS REFERRED TO.

. Bepparp, I, E.—‘ Contributions to the Anatomy of Harthworms.”

1. “On the Structure of Eudrilus sylvicola,” ‘ Proc. Zool. Soc.,’
1887, p. 372.

. Bepparp, F. E.—“‘ Note on the Structure and Development of the

Ovum in an Annelid (Eudrilus),” ‘ Journ. Anat. and Phys.,’ October,
1887, p. 9.

. Bepparp, F, E.—“ Contributions to the Anatomy of Earthworms, with

Descriptions of some New Species,” ‘Quart. Journ. Micr. Sci.,’
vol. xxx, p. 421.

. Bepparp, F. E.— On the Structure of Three New Species of Earth-

worms, with Remarks on Certain Points in the Morphology of the
Oligocheta,” ‘Quart. Journ. Micr. Sci.,’ vol. xxix, p. 101.

. BEppARD, F. E.—‘‘ On the Classification and Distribution of Earth-

worms,” ‘Proc. Roy. Phys. Soc,’ (to appear shortly).

. Bennam, W. B.—“ ‘Atrium’ or ‘Prostate’ ?’’ ‘Zool. Anz. Jahrg.,’ xiii,

p. 368.

. Micuartsen, W.— Beschreibung der von Herrn Dr. Franz Stuhlmann

im Miindungs gebiet des Sambesi gesammelten Terricolen,’’ ‘Jahrb.
Hamb. wiss. Anst.,’ vii.

. Perrier, E.— Recherches pour servir & Vhistoire des Lombriciens

terrestres,” ‘ Nouv. Arch. Mus.,’ t. viii.

. PERRIER, E.—‘ Comptes rendus ’ for 1886.
. Rosa, D.—‘* Lombrichi dello Scioa,” ‘Ann. Mus. civ. Genova,’ 1889,

p. 571.

. Kiypere.—“ Annulata Nova,” ‘Oefv. k. Svensk. Akad. Handl.,’ 1866,

pe oT.
Vespovsky, F.—‘ Entwickelungsgeschichtliche Untersuchungen,’ Heft i.

Kitxentuat, W.— “ Ueber die Lymphoid-zellen der Anneliden,” ‘ Jen.
Zeitschr.,’ Bd. xviii, p. 319.

274 FRANK E. BEDDARD.

EXPLANATION OF PLATES XVI—XX,

Illustrating Mr. Frank E. Beddard’s memoir “ On the Struc-
ture of Two New Genera of Earthworms belonging to the
Eudrilidx, and some Remarks on Nemertodrilus.”

PLATE XVI.
Hyperiodrilus africanus.

Fic. 1.—Female reproductive apparatus, seen after opening body-wall from
above and removing one of the calciferous glands. ca. Calciferous gland.
sp’. Large ccelomic pouch, connected with a periesophageal ring. 7. 0. Re-
ceptaculum ovorum. od. Oviduct. «@s. Gsophagus. d.v. Dorsal vessel.
s. Intersegmental septum.

Fic. 2.—Integumental sense (?) organs in longitudinal section.

Fie. 3.—The same in situ. Transverse section. This drawing also
shows the lymph-spaces between the fibres of the circular muscular coat.

Fic. 4.—Diagram of structure of the same.

Fic. 5.—Diagrammatic lateral view of the different parts of the female
reproductive apparatus. The segments are numbered. sp. Spermatheca.
ov. Ovary. Other letters as in Fig. 1. The walls of the coelomic spaces are
here and there cut away so as to display the contained viscera.

Fie. 6.—Anterior segments from above, to show form of prostomium,

Fie. 7.—A portion of integument, highly magnified, to show relationship
of nephridiopore (zp.) to seta (s.) of dorsal pair.

Fic. 8.—Female reproductive apparatus removed from body, but with its
relations to other organs indicated. 6. Bursa. Other letters as in
Figs. 1 and 5.

Fic. 9.—Transverse section to show arrangement of sete.

Fie. 10.—Transverse section through a portion of perigonadial sac and the
neighbouring body-wall. «. Nephridium. Other lettering as in Fig. 8.

Fie. 11.—Diagrammatic view of female reproductive organs. The peri-
cesophageal sinuses and the unpaired dorsal sac in which they meet are pushed
back ; the cesophagus is cut away, but the nerve-cord is left. On the left
side the anterior wall of perigonadial sinus and pericsophageal sinus is
removed in order to display enclosed viscera. The segments are numbered,
Lettering as in Figs. 1, 5, and 8.

Fic. 12.—A few segments pressed out to show arrangement of sete (s.)
and position of nephridiopores (w.) by lateral sete.

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 275

PLATE XVII.
Hyperiodrilus africanus and Heliodrilus lagosensis.

Fic. 13.—Diagrammatic transverse section of Hyperiodrilus in neigh-
bourhood of the female reproductive organs, to be compared with

Ftc. 14, representing a corresponding section through Heliodrilus,

Fic. 15.—Heliodrilus. The sac, containing testis and its attachment
to the body-wall. mes. Mesentery. /m. Longitudinal muscle-fibres.

Fic, 16.—Heliodrilus. Dissection of ccelomic space, containing testis
and first part of vas deferens. The wall of the sperm-sac is partly removed
to show funnel.

Fic. 17.—Transverse section through alimentary canal (cz¢.) and the
dilated part of the vas deferens (v. d.) of Heliodrilus.

Fic. 18.—Dissection to show male reproductive system of Heliodrilus.
ves. sem. Sperm-sacs. atv. Atrium uncoiled; on the opposite side it is left
coiled up.

PLATE XVIII.
Hyperiodrilus africanus and Heliodrilus lagosensis.

Fic. 19.—Hyperiodrilus. Magnified view of the ventral surface of a
portion of Segment 17 to show the papille (pap.) connected by grooves with
the penis. The left papilla is on the boundary line between Segments 16
and 17; the right hand one in the latter segment. s. Sete.

Fic. 20.—17th and neighbouring segments of another individual.

Fic. 21.—Heliodrilus. Ventral aspect of anterior segments to show
position of male pore ( ¢), spermathecal pore ( ?), and papilla (pap.).

Fie. 22.—Hyperiodrilus, A third individual, 17th and neighbouring
segments. In this specimen the papille are symmetrical and upon the 17th
segment.

Fic. 23.—Hyperiodrilus. Lateral view of genital segments, showing
oviducal pores (2), nephridiopores (z.), and male pore ( ¢).

Fie, 24.—Hyperiodrilus. Ventral view of 17th and neighbouring seg-
ments of same individual as that represented in Fig. 19. p. Penis, slightly
protruding from male pore. @ Spermathecal pore. pap. Papille.

Figs. 25 and 26.—Hyperiodrilus, Sections through ventral esophageal
pouch at two points; for explanation see text.

Fic. 27.—Hyperiodrilus. Vasa deferentia, with their peritoneal coat.

Fic. 28.—H yperiodrilus. Dilated portion of vas deferens.

Fie. 29.—Heliodrilus. A portion of a section through the receptaculum

276 FRANK E. BEDDARD.

ovorum. ov. Mature ovum with membrane. germ. Germinal cells attached
to ovum.

Fic, 30.—Hyperiodrilus. Transverse section through unpaired cesopha-
geal pouch, to show blood-spaces (4/ack) and lumina between folds of
epithelium.

Fic. 31—Hyperiodrilus. Origin of unpaired cesophageal pouch (ca.).
d. Glandular (?) cells found among the ordinary epithelial cells. p. Peri-
toneum.

PLATE XIX.
Heliodrilus lagosensis.

Fics. 32—34.—A series of sections to show the investment of the apex of
the spermatheca by cclomic spaces.

Fig. 32. The apex of the spermatheca (sperm.) lies between two out-
growths of the cclomic sac (c@.).

Fig. 33. The apex of spermatheca (sp.) is now enclosed within the
coelomic sac, and the commencement of a second ccelomic space
within the first is just visible.

Fig. 34. This section lies beyond the apex of spermatheca, which just
reaches the interior of the sac (ce/'.), lying within that (cwlom) which
is continuous with the perigonadial sac.

Fig, 35.—Diagrammatic representation of the extremity of the sperma-
theca (sp.) enclosed within a sac, which is itself enclosed within an extension
of the perigonadial sac; the narrow canal leading from the latter to the ovary
is seen. Portions of the walls of the different sacs are removed to show
their contents.

Fie. 36.—Section through dilated portion of vas deferens with surrounding
coelomic sac. vd.e. Ciliated epithelium of vas deferens. yp. Peritoneum
covering ccelomic sacs ; in its interior are seen clumps of cells.

Fic. 37.—Section showing structure of spermatheca. ep. Epithelium.
gv. Granular matter thrown off from cells into the interior of the spermatheca.
m. Muscular coat with blood-vessels, shown black.

Fic, 38.—Section showing structure of distal part of atrium. msc. Mus-
cular coat. . Nuclei of innermost epithelial layer.

Fie. 39.—Section showing structure of atrium, near to its external orifice.
Large glandular cells (g/.) are seen, separated by interstitial cells. muse.
Muscular coat.

Fie. 40.—Section through common orifice of atrium and nephridium.

Fic. 41.—Diagram of female reproductive system. sperm. Spermatheca.
sp. 0. Its external orifice. ov. Ovarian sac, represented as being cut open on

STRUCTURE OF TWO NEW GENERA OF EARTHWORMS. 277

the left side to show the contained ovary. sp. sac. Coelomic sac involving a
second sac, in which lies apex of spermatheca; the walls are represented
as partly cut away. This part is shown more highly magnified in Fig. 35.
r. 0. Receptaculum ovorum, represented as being cut open on left side to show
funnel of oviduct (od.). The nerve-cord is removed for the greater part to
show the underlying narrow duct of the spermatheca.

Fic. 42.—Section through atrium at the point where the two vasa deferentia
open.

Fic. 43.—Section through a portion of one of the ventral cesophageal
pouches. 4/. Blood-vessels, coloured black. sf. Peritoneal covering. x.
Intra-cellular part of lumen.

PLATE XX.

Hyperiodrilus africanus, Heliodrilus lagosensis, Nemertodrilus
griseus, Teleudrilus ragazzii, and Eudrilus.

Fic. 44.—Hyperiodrilus. Diagram of male reproductive organs. s. ¢.
Sperm-sac. v.d. Vas deferens. Atria and testes also shown.

Fie. 45.—Hyperiodrilus. Longitudinal section through duct, leading
from egg-sac. c. Outer. c’. Inner peritoneum.

Fie. 46.—Hyperiodrilus. Supra-intestinal vessel, enclosed within a
peritoneal space, which is divided by trabecule. In the interstices lie clumps
of cells. The perihemal sac is connected to the dorsal vessel (d. vess.) by a
mesentery.

Fie. 47.—Hyperiodrilus. Dissection of the anterior end of the worm,
sp'. Anterior. sp. Posterior of thickened intersegmental septa. @s. sac.
(sophageal pouches. ca. Calciferous glands. sp. Coelomic sac, connected
with pericesophageal ring, seen anteriorly. v.d. Vas deferens. at. Atria.
g- First of posteriorly-situated gizzards. The segments are numbered from
the 6th to the 17th.

Fie, 48.—Heliodrilus. A few of anterior segments to show overlapping
and interconnection of thickened intersegmental septa.

Fic. 49.—Hyperiodrilus. MRipe (4) and nearly ripe (@) ovum. 2.7,
Zona radiata. v. Vitelline membrane.

Fic. 50. Hyperiodrilus. Germinal cells from ovary.

Fie. 51.—Hyperiodrilus. Section through ovarian sacs, to show their
interconnection. v. 4. Ventral blood-vessel.

Fie. 52.—Heliodrilus. Gizzards and commencement of intestine.

Fig. 53.—Hudrilus. Diagram of female reproductive system. ov. Ovary,
surrounded by a sac continuous with a duct (od.). ov’. Second ovary, sur-

278 FRANK E. BEDDARD.

rounded by a sac (receptaculum ovorum) continuous with oviduct (od'.). sp.
Spermatheca. 9. Female pore.

Fic. 54.—Nemertodrilus. A similar diagram. Lettering as above.
od. is simply a pore.

Fie. 55.—Teleudrilus. A similar diagram. Letters as above.

Fic. 56.—Hyperiodrilus. Terminal portion of atria, and their connec-
tion with penis. ».d. Vas deferens. pr. Glandular part of atrium. m,
Muscular part of ditto. yp. Penis.

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 279

The Renal Organs of Certain Decapod
Crustacea.

By

w. F. R. Weldon, M.A, F.R.S.,
Jodrell Professor of Zoology and Comparative Anatomy in University College,
London ; Fellow of St. John’s College, Cambridge.

With Plates XXI & XXII.

In a paper published during the summer of 1889,! I gave an
account of the excretory apparatus of Palemon serratus,
in which I drew attention to the enormous development of
the bladder, which extends dorsally from the level of the green
glands to the anterior boundary of the pericardium.

The existence of a well-developed nephro-peritoneal sac,
having many of the relations of the “‘ celomic”’ body-cavity
of other types, had not, at the time of publication of my
paper, been recognised in any Crustacean. I have since
observed the occurrence of a similar sac in a large number of
Decapoda; and M. P. Marchal,? who has evidently worked
without any knowledge of my observations, has described
similar structures in a considerable number of genera.

The object of the present communication is to give an
account of the renal organs of certain Caridide (Pandalus,
Virbius, and Crangon) in which the structure of the green
gland itself is modified in a very remarkable manner.

Before proceeding to a description of the genera which it is
proposed more particularly to consider, it may be well to give

1 «Journal of the Marine Biological Association,’ N.S., vol. i, p. 162.
2 «Comptes Rendus,’ cxi, 12, and cxi, 16.

VOL. XXXII, PART III.—NEW SER. Ad

280 W. F. R. WELDON.

a short summary of the results already arrived at concerning
the structure of the excretory apparatus in the Prawn.

The nephro-peritoneal sac of Palemon is a median,
unpaired structure, lying in the cephalothorax, dorsal to the
alimentary canal, and ventral to the ophthalmic artery and to
the median dorsal blood-sinus. Its walls are composed of a
single layer of flattened excretory epithelium, which has the
power of absorbing indigo carmine and similar substances
when these are injected into the blood of the living animal.
Posteriorly the nephro-peritoneal sac is in close contact with
the anterior extremity of the generative gland ; anteriorly it
gives off on each side a narrow tube, which passes vertically
downwards beside the cesophagus, and, after passing under the
cesophageal nerve-commissure, bends outwards to open into
the urinary bladder of its own side. The ducts of opposite
sides communicate with one another, not only dorsally, through
the cavity of the nephro-peritoneal sac, but also vertically, by
means of a transverse commissure which passes in front of the
cesophagus, and bears a conspicuous dilatation in the cavity of
the upper lip.

A diagram of the whole arrangement is given in Pl. XXII,
fig. 9; the relations to the other systems of organs, as seen in
transverse section, may be gathered from figs. 1 and 2.

The bladder, besides receiving the openings of the nephro-
peritoneal ducts, gives off on the one hand the ureter, and on
the other the system of excretory tubules of the green gland.
These last form a complex mass of branching tubules, which
are in close contact one with another, and which form the
glandular substance of the green gland. In my former paper
I pointed out the existence, in this glandular plexus, of several
excretory tubules, an observation confirmed by M. Marchal.
I was unable last year, and have still been unable, to convince
myself that the various tubules anastomose freely with one
another in the way described by this observer. Be this as it
may, however, the tubules, after a tortuous passage through
the substance of the green gland, unite to open by a common
aperture into the “ end-sac,’? whose structure has been re-

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 281

peatedly described, especially by Grobben, Marchal, and
myself.

In my former paper I described only the arrangement
found in P. serratus, and the observations of M. Marchal,
which are in complete agreement with my own, relate also
to this species. I have since satisfied myself that the ex-
cretory system of P. squilla, and that of Palemonetes
varians, are practically identical with that of the species
described.

In the genera Virbius, Pandalus, and Crangon, a
series of modifications may be observed, resulting in the dis-
appearance of the whole tubular portion of the green gland,
and the hypertrophy and specialisation of the end-sac. The
nephro-peritoneal sacs are also arranged in a manner strikingly
different from that which obtains in the Prawns.

The nephro-peritoneal sacs are nearly identical in
arrangement in the three genera in question. Their relations
will be readily understood from the diagram, fig. 10, and the
transverse section of Pandalus brevirostris, fig. 3. The
bladder gives off from its internal aspect a duct, which is both
wider and shorter than the corresponding duct of Palemon,
the duct from each bladder passing inwards under the
cesophageal commissure of its own side, and opening into a
large sac, which does not communicate with its fellow of the
opposite side, and which does not extend to the middle dorsal
line above the alimentary canal. The median wall of each
sac is closely applied partly to the lateral and ventral wall
of the stomach, and partly to the median face of the sac of the
opposite side, so that the stomach appears in section to be
supported by a well-marked ventral mesentery, which exists
beneath the part of that organ which projects in front of the
cesophagus, as well as beneath its post-oral region. Both
bladder and sac give off numerous processes, which ramify
in the base of the antenne and among the organs of the
thorax. The epithelium forming the walls of these organs is
in all cases striated and excretory, being very closely similar
to the corresponding epithelium of the Prawns. In none of

282 WwW. F. R. WELDON.

the three genera is there any connection between the wall of
the nephro-peritoneal sacs and the generative glands.

The structure of the “ green gland” differs in each of the
three genera.

In Virbius (varians) a horizontal section through the
bladder and “ green gland ” has the appearance represented in
fig. 4. The bladder itself is bounded by a single layer of
epithelial cells, whose inner margins frequently project irre-
gularly into the cavity of the organ. These cells exhibit,
especially in their peripheral portions, the well-marked longi-
tudinal striation which is so constantly noticed in excretory
tissues ; their internal portions are, however, frequently vacuo-
lated, and their free internal borders are ragged and indefinite.
When this condition prevails the cavity of the bladder is seen
(in stained sections) to contain an irregular, granular coagulum,
which absorbs hematoxylin with readiness. The nuclei of
the bladder-cells are oval, and of moderate size; they stain
deeply, and do not, in preparations preserved in corrosive sub-
limate, exhibit any very evident reticulum. The cells com-
posing that portion of the wall of the bladder which invests
the end-sac are flatter and more regular than the others ; they
stain more deeply with hematoxylin, and the longitudinal
striation is perhaps more evident in these than in the other
bladder-cells.

The renal tubule is single, and has a much wider lumen
than any of the corresponding tubules of Palemon. It leaves
the bladder at the postero-external margin of that organ, and
is at first directed nearly horizontally outwards. After passing
outwards for a very short distance, however, the tubule turns
backwards and then inwards, so that it becomes U-shaped, and
opens into the end-sac. The general direction of the single
renal tube is that just described, but it does not lie in one
plane with such accuracy as to enable it to be included ina
single section. In the section figured (Pl. XXI, fig. 4) the
two ends of the tube only are seen, the one leaving the
bladder, the other entering the end-sac. By examination of
the following sections, the course of the tube was determined

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 283
as that shown by the dotted lines in the figure. [The deter-
mination of such a point is so easy that it has not seemed
worth while to publish the figures of the sections upon which
it rests. |

It is evident that the single, wide, U-shaped tube, running
from the bladder to the end-sac, is the only representative in
Virbius of the complicated plexus which goes to make up the
mass of the green gland in the Prawn. In structure, its walls
somewhat resemble those of the bladder already described.
They are, however, higher and more regular, their internal
extremities of the component cells being less given to the
exhibition of vacuoles and irregular processes. ‘The longi-
tudinal striation is more evident, and the nuclei are larger
and stain more deeply with hematoxylin. Before opening
into the end-sac, the lumen of the renal tubule contracts con-
siderably, so that the orifice, by which the cavities of the two
structures communicate, is very small. Owing to its size and
position, it is exceedingly difficult to demonstrate this opening
in transverse sections, but in carefully adjusted horizontal
longitudinal sections it is, as will be evident from the figure,
easily to be seen.

The end-sac itself is completely enveloped by a layer of
bladder epithelium, the wall of the bladder being invaginated
by it. The epithelium of the bladder is, however, not in
absolute contact with that of the end-sac, the two being sepa-
rated by a blood-space (left white in the figure). I have not
been able to detect any epithelium bounding this blood-space ;
and I am inclined to believe, after careful examination, that
no such epithelium exists. The account given by Grobben of
the end-sac of Palemon treillianus leads to the belief
that the renal vessels of this species end in lacunz which are
devoid of epithelial lining; and I have been unable to demon-
strate an epithelium in the smaller blood-spaces of the kidney
of Pandalus (annulicornis and brevirostris). Professors
Claus! and Lankester® have, as is well known, arrived inde-

1 ¢ Arb. Zool. Inst. Wien,’ Bd. v, Heft 3 (1884),
? This Journal, vol. xxv, p. 518,

284, W. F. BR. WELDON.

pendently at the conclusion that the blood system of the
Decapod Crustacea is everywhere closed; but I am not aware
that either of these observers has paid special attention to the
blood-supply of the renal organs. Nevertheless, the positive
testimony of two such accomplished anatomists makes me
hesitate to lay undue stress upon my own failure to demon-
strate an epithelial lining to the particular spaces under con-
sideration.

The epithelium of the end-sac, like that of the rest of the
excretory system, is everywhere one cell thick. The indi-
vidual cells stain more deeply than do those of the bladder
and renal tubule; their protoplasm is crowded with granules,
which are, however, not arranged in regular rows, so that the
cells do not exhibit a longitudinal striation. The nuclei stain
very deeply, and exhibit, in specimens preserved in corrosive
sublimate, one or two large nucleoli, with no recognisable
trace of a chromatin reticulum. The inner extremities of
these cells are much vacuolated, and are very irregular; the
vacuoles frequently containing spherical concretions of a homo-
geneous material, which stains slightly with hematoxylin,
less readily with borax carmine. The cavity of the end-sac is
generally found to contain a greater or less quantity of granular
clotted material, which appears in sections as a deeply staining,
finely granular reticulum.

In Pandalus (annulicornis and brevirostris) a further
deviation from the tubular type of nephridium occurs, the
renal tubule being in the adult condition entirely absent,
while the whole body of the “‘ green gland” is built up of the
curiously modified end-sac and the associated portion of the
wall of the bladder.

I have not followed the early phenomena of the development
of the kidney ; but in the late ‘‘ Mysis ” stage the relations of
kidney and bladder present a striking resemblance to those
which are permanent throughout life in Virbius. The
appearance presented at this stage in horizontal longitudinal
section is shown in Pl. XX, fig. 5.

The bladder is large compared with the size of the renal

RENAL ORGANS OF CERTAIN DECAPOD ORUSTACEA. 285

tubule and end-sac; its cells are, for the most part, pale and
uniformly granular, having no trace of longitudinal striation ;
their nuclei are rounded, and do not stain deeply. At this
stage there is little difference in appearance between that por-
tion of the bladder-wall which is immediately adjacent to the
end-sac and the remainder. The cells adjoining the end-sac
are slightly smaller than the rest, and their margins slightly
more regular, but in other ways the characters of the bladder-
cells are everywhere the same; and the sac, though closely
applied to the wall of the bladder, causes hardly any invagina-
tion of that structure.

The renal tubule (Pl. XXI, fig. 5) is very short, being
simply represented by the curved neck which connects the
end-sac with the bladder. Its walls are very similar in struc-
ture to those of the bladder itself, and its lumen is very narrow.

The end-sac (Pl. XXI, fig. 5) is small, and bounded by
finely granular or nearly homogeneous cells, the nuclei of
which are clear and vesicular in appearance. Between the
end-sac and the wall of the bladder is a lacunar blood-space ;
and both here and on the outer side of the end-sac are groups
of connective-tissue cells.

In a very young Pandalus annulicornis, which had
apparently only just acquired the adult characters, the appear-
ance presented by a transverse section through the excretory
organs is shown in fig. 6. [I have only obtained a single
individual of this age, and before the present investigation
was commenced I had prepared transverse sections of the
specimen. I have therefore been unable to figure a section
in the same plane as those above described. ]

The wall of the bladder exhibits already a distinct specialisa-
tion into two regions, one lying beside the end-sac, the other
having no relation with that organ. In the latter portion,
which of course includes the greater part of the bladder, the
cells exhibit distinct indications of longitudinal striation in
their peripheral parts, and their inner extremities are often
vacuolated. ‘The portion which adjoins the end-sac consists of
cells which are more columnar than those of the general

286 W. F. R. WELDON.

surface of the bladder, which stain more deeply, and are more
crowded with granules. The nuclei of these cells exhibit
frequeut indications of division.

The end-sac is by this time more closely applied to the
bladder, the wall of which it invaginates to such an extent
that about half its surface is invested by a layer of bladder-
cells. The epithelium of the end-sac is made up on the side
next the bladder of small cubical cells, and on the opposite
side of cells which are somewhat flattened. The protoplasm
of all these cells is pale and coarsely granular, as it remains
throughout life. The connective tissue, which was noticed in
the last stage, has slightly increased in amount, and the blood-
spaces between end-sac and bladder are much better developed
than during the “ Mysis” stage. The renal tubule has
ceased to be distinguishable as a separate region, and the end-sac
now opens by its posterior extremity directly into the bladder.

The stages in the further development of the bladder I have
been unable to observe, my remaining material consisting
entirely of adult individuals. It is evident, however, that the
end-sac becomes completely surrounded by the wall of the
bladder, so that it finally projects freely into the cavity of
that organ, being attached to the bladder-wall by a narrow
stalk, on which is situated the communication between the
cavities of the two structures.

While the process of enclosure of the end-sac by the bladder
is going on, the wall of the first-named organ becomes pro-
duced into a number of complicated branched papille, which
project into the cavity of the bladder, each being, of course,
covered by a layer of bladder epithelium. At the same time
the whole organ increases in size till it becomes about as large
as the whole “ green gland” of Palemon.

A section through the adult end-sac is drawn in fig. 7,
where the extent of the papilliform prolongations into which
it is produced is rather below the average. The epithelium is
flattened and irregular, composed of a pale, granular proto-
plasm. The cells do not exhibit vacuoles, and I have not
observed the presence of concretions in any of my specimens.

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 287

The nuclei are small, rounded, and pale, with scattered, deeply
staining chromatin granules. The communication between
the cavity of the organ and that of the bladder is seen to lie
at one side of the stalk, and to be quite direct, without the
intervention of anything which can be held to represent the
system of tubules of Palemon, or even the simple U-shaped
tubule of Virbius.

Between the wall of the end-sac and the investing portion
of the bladder there is a certain quantity of connective tissue,
which in places forms fairly conspicuous masses, the characters
of which tissue will be gathered from the figure. Besides
the connective tissue a system of blood-vessels ramifies between
end-sac and bladder, consisting in part of larger vessels, in
which it is easy to recognise an epithelial lining, and in part
of smaller, apparently lacunar spaces. These vessels are sup-
plied by one or two main trunks which pass along the neck of
the end-sac. Two of these vessels, cut transversely before
their entrance into the space referred to, are seen in fig. 7
lying outside the wall of the bladder.

The epithelium of the bladder itself is everywhere the same.
The cells are columnar, and very regular in outline, showing no
trace of vacuolation or of production into irregular processes.
They exhibit an exceedingly well-marked longitudinal striation,
and stain deeply both with hematoxylin and with borax
carmine. The internal border of the bladder-cells seems always
to be darker and more homogeneous than the rest, but there
is no indication of the existence of a definite cuticle. The
nuclei are rounded and granular, and stain fairly deeply.

The youngest individuals of Crangon vulgaris which
were examined had already attained the external characters of
the adult, although they were scarcely larger than the oldest
« Mysis ” larve.

These specimens correspond in age to the second stage in
the development of Pandalus, as above described, and the
condition of the excretory system is practically identical in
the two species.

The bladder- wall in the young C. vulgaris (see Pl. XXI,

288 W. F. R. WELDON.

fig. 8) exhibits already the characteristic longitudinal striation,
and as in the young Pandalus it surrounds half the end-sac.
The communication between its cavity and that of the end-sac
is direct, there being no trace of the renal tubule.

The end-sac is bounded by a layer of pale, finely granular
cells, the protoplasm of which exhibits the well-known “ ground-
glass” appearance. The epithelium of that half of the sac
which is enclosed by the bladder is more columnar, that of
the unenclosed portion being flatter. The cells of both regions
exhibit numerous vacuoles at their inner margins, but no con-
cretions were observed.

Between the end-sac and the bladder is a well-developed,
apparently lacunar blood-space, and outside the end-sac is a
layer of connective tissue.

The excretory system of an adult shrimp resembles that of
Pandalus in the direct communication between end-sac and
bladder, and in the formation of papille upon that surface of
the end-sac which projects into the bladder. The whole sac is
not, however, enveloped by the bladder so completely as it is
in Pandalus.

It is evident from what has been said that the excretory
system of the Decapoda is much more varied in its structure
than has hitherto been supposed. The observations here
recorded, together with those of M. Marchal already referred
to, enable us to divide the modifications into groups as follows :

In the Schizopods (Mysis) the whole excretory system
appears, according to Grobben,! to consist of a single coiled
tubule, opening by one extremity to the exterior, and by the
other to an irregular end-sac, whose walls are composed of an
irregular epithelium, and are not apparently very highly
specialised. The single renal tubule may dilate into a small
bladder near its external opening, but there is no indication of
the extension into the thorax of a nephro-peritoneal sac.

In all the Decapods proper the end-sac has become more
highly specialised, possessing a lining epithelium of definite
characters, a well-defined system of blood-vessels, and so on;

1 *Arb. Zool. Inst. Wien,’ iii, 1881.

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 289

but the characters of the remaining portions of the excretory
system vary greatly.

In the Thalassinide (Axius and Gebia) the number of
the tubules, and the complexity of their arrangement, have
increased, so that several tubules lead from the ureter to the
end-sac; but there is no vesicular dilatation of any part of
the system.

In the Astacide (Astacus, Homarus, and Nephrops)
several tubules run from the ureter to the end-sac ; but by the
dilatation of one of these, which receives the openings of the
others, a bladder is constituted; and this bladder intervenes
between the plexus of tubules, forming the mass of the “ green
gland,” and the ureter. In this group the bladder has no
considerable extension into the thorax.

In the Loricata (Palinurus) the bladder has the same
relations as those described in the Astacide; but the
number and complexity of the tubules forming the green
gland is very largely increased.

In the Carididz the structure of the excretory system
varies greatly, the four modifications described in the previous
section of this paper all occurring within the limits of the
group.

In the “Anomura” there is a well-marked nephro-
peritoneal system, which in Pagurus forms a median dorsal
sac, situated far back in the abdomen. The end-sac is fre-
quently produced into papille, which are surrounded, not
by a modified portion of the bladder, as in Pandalus and
Crangon, but by a layer of renal tubules.

In the Brachyura the nephro-peritoneal sac sends well-
developed prolongations into the thorax ; there are, at least
frequently, two such prolongations on each side, one dorsal
and one ventral. The end-sac is frequently produced into
papille, which are covered by the epithelium of the short,
wide renal tubule, the cavity of this tubule being frequently
broken up by a system of trabecule.1

1 In this sketch of the various Decapods, the account of the Thalassinide
and of the Brachyura is taken directly from Marchal’s paper.

290 W. F. R. WELDON.

It appears, from the foregoing statements, that the nephro-
peritoneal sacs of the Decapoda should be regarded rather as
enlarged portions of a tubular system, such as that found in
Mysis and in the Thalassinide, than as persistent rem-
nants of a “ccelomic” body-cavity, into which tubular
nephridia open. ©

The presence of coxal glands in Nebalia,! and of tubular
nephridia in the zocea of Eriphya,? gives much interest to the
search for an embryonic celom in these animals, which may
be expected to behave like the coelom of Peripatus.

EXPLANATION OF PLATES XXI & XXII,

Illustrating Professor W. F. R. Weldon’s paper on “ The
Renal Organs of Certain Decapod Crustacea.”’

List of Reference Letters.

Bil. Bladder. Br. Brain. B.v. Blood-vessel (or lacuna). Comm. Circum-
cesophageal nerve-commissure. Z#. s. End-sac. Zdr. Labrum. J. p. Nephro-
peritoneal sac. (C#¥s. isophagus. S%. Stomach. Zw. Tubular portion of
“oreen gland.” U. Ureter.

Fic. 1.—Transverse section through the head of a Palemon serratus,
showing the connection between the dorsal nephro-peritoneal sac and the
bladder.

Fic. 2.—Transverse section through the same specimen, rather behind Fig.
1, showing the dilated median ventral portion of the neprho-peritoneal
system lying in the upper lip, and the dorsal portion lying above the stomach.

Fic. 3.—Transverse section through the head of Pandalus brevirostris,
just in front of the cesophagus, showing the ventral mesentery formed by the
two nephro-peritoneal sacs.

Fic. 4.—Horizontal section through the bladder and end-sac of an adult
Virbius. The dotted lines indicate the course of the single renal tubule.

1 Claus, ‘Arb. Zool. Inst. Wien,’ Bd. viii, Heft 1, 1888.
2 Ebedinski, ‘ Biol. Centralbl.,’ x, p. 178.

RENAL ORGANS OF CERTAIN DECAPOD CRUSTACEA. 291

Fic. 5.—Horizontal section through the bladder and “ green gland” of
Pandalus annulicornis in the Mysis stage.

Fic. 6.—Transverse section through the kidney of Pandalus annuli-
cornis, just after the final metamorphosis.

Fic. 7.—Section through the end-sac and associated wall of the bladder
in an adult Pandalus. The letters BV. lie in the cavity of the bladder.

Fie. 8.—Horizontal section through the kidney of a young Crangon vul-
garis, just after the final metamorphosis.

Fic. 9.—Diagram of the excretory system of Palemon.
Fic. 10.—Diagram of the excretory system of Pandalus.

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THE NEPHRIDIUM OF LUMBRICUS. 293

The Nephridium of Lumbricus and its Blood-
supply; with Remarks on the Nephridia in
other Cheetopoda.

By

W, Blaxland Benham, D.Sc. (Lond.),
Anatomical Department, Oxford.

With Plates XXIII—XXV.

Tue nephridium of the common earthworm has recently
been the subject of a contribution by Dr. Goehlich (20), in
which he remarks on various statements made by Gegenbaur
in his classical paper on the subject, published in 1853 (19),
in which he corrected the then prevailing view that the seg-
mentally arranged tubes were respiratory, and suggested their
excretory function.

As several of Goehlich’s statements are at variance with those
of Gegenbaur I was led to look into the details of the organ,
and I then found that on those points the more recent writer
was in error, and that Gegenbaur’s description is much more
nearly correct than Goehlich’s. In fact, I have not very much
to add to the description of the earlier writer beyond matters
of histological detail, and more especially the blood-supply,
which Professor Lankester suggested that I should work out.

The work was carried out in the Zoological Laboratory of
University College during the past eighteen months.

294 W. BLAXLAND BENHAM.

ConTENTS OF PAPER.

1. Nomenclature of the parts of the nephridium, and the course of the
various regions.

2. Histology of the various regions, and suggestions as to their function.

3. Comparison with the nephridium in other genera.

4, The nephrostome of Pericheta malamaniensis, n. sp., and other
genera,

5. Circulation in the nephridium,

6. The nephridium of Arenicola.

1. NoMENCLATURE OF THE Parts AND CoURSE OF THE
Various REGIonNs.

The long winding tube which constitutes the excretory
organ of Lumbricus is for the most part embedded in a
coating of vesicular cells, which are in their turn surrounded
by the pavement-cells of the coelomic epithelium. The tube
is, in its adult condition, as in its development, divisible into
two portions: (I) a preseptal portion, consisting of the in-
ternal funnel or nephrostome and a short ciliated tube; and
(II) a much more extensive post-septal portion. The latter is
readily distinguishable into four regions: (1) the very long
but narrow tube in continuity with the preseptal tube; (2)
the short brownish ciliated middle tube; (8) the wide large
tube; and (4) the muscular tube or duct, which opens to
the exterior. The first three of these tubes (1, 2, 3) are twined
about in rather a complicated but quite constant manner, and
are bound together by the coat of vesicular cells in such a way
as to form two great “ loops,” visible fairly distinctly with the
naked eye when an opened worm is covered by spirit. Another
“loop” is formed by the muscular duct (4). This last I will
call the “first loop” (x, figs. 1, 2, 3); the ‘‘ second loop”
(Fr) contains the greater part of the narrow tube (1), and of
the large tube (3) ; finally, the ‘third loop” (eG) consists of a
part of the narrow tube, a part of the large tube, and the
whole of the middle tube (2).

The course taken by these various regions, from the funnel
to the external aperture, and the relations of the tubes to the

THE NEPHRIDIUM OF LUMBRICUS. 295

loops, will be readily seen by a glance at fig. 2, which will
convey more information than any verbal description. I will,
therefore, pass on to the histology of the various parts.

I will premise that, with the exception of the funnel itself,
or rather a part of the funnel, together with probably the mus-
cular duct, the canal of the nephridium is “ intra-cellular,”
i. e. the nephridium consists of a large number of perforated
or “drain-pipe” cells, placed end to end (see Pl. XXIV, figs.
30, 31, 32). The size of the cell and the relative proportion
occupied by the lumen constitute the main morphological
differences observable in the various regions; but the character
of the protoplasm is of very great physiological importance,
although we do not fully know the exact function of each
region.

2. THe SrructTuRE oF THE VARIOUS REGIONS.

I. The Preseptal Portion.—The nephrostome or funnel
is carried at the end of the short portion of the “ narrow tube ”
which passes through the anterior septum bounding a given
somite. The character of this portion of the tube will be
described below. This preseptal portion of the tube is sur-
rounded by a mass of vesicular cells, the outlines of which are
readily seen in the living condition, and frequently in stained
preparations. These cells resemble those coating the post-
septal portion, and appear to be similar to those described and
figured by Kukenthal round the vessels and nephridia of
Tubifex (24), and not improbably they have the same
meaning. The superficial cells are usually flattened, and
form a celomic epithelium continuous with that covering the
septum. |

The funnel (Pl. XXIII, figs. 4—7, and 32) has recently
been figured by Goehlich (20) in some detail, but he has alto-
gether misunderstood the appearance represented. As is well
known, the funnel consists of a number of very long, narrow,
somewhat wedge-shaped cells—which I shall call “ marginal
cells”—arranged around a central point. The nucleus in
each is placed near the outer end of the cell, which is rounded

VOL. XXXII, PART III.—NEW SER. U

296 W. BLAXLAND BENHAM.

Fic. 1.—A nephridium treated with nitrate of silver, to
show the pavement ccelomic epithelium covering the vesicular
tissue, and forming in parts a suspensory fenestrated mem-
brane, which carries blood-vessels. H. First loop. F F.
Secondloop. GG. Thirdloop. N. Nerve-cord. S. Septum.
SN. Subneural blood-vessel. V. Subintestinal blood-vessel.
a. Somatic branch from subintestinal blood-vessel. d. Ventral
part of commissural vessel. /, h, h. Suspensory membranes,
carrying and covering the nephridial blood-vessels. 7. The
group of ccelomic epithelial cells around the przseptal region
of the narrow tube.

298 W. BLAXLAND BENHAM.

off peripherally, and ciliated all over one surface. Goehlich
describes and figures one or two rows of specially long cilia
radiating from the centre. These I have never seen, although
I have had under observation hundreds of nephridial funnels
in demonstrating to my students, and have examined carefully
dozens forthis very purpose. The appearance figured by Goehlich
is due to a crumpling of the funnel—an action to which it is
liable on being covered by a glass slip,—so that there will be
caused radial folds, the cilia of which will then be seen side-
ways instead of from above, will therefore be more distinct,
and will appear longer. There are normally no such longer
cilia or such radiating lines.

Another phenomenon, which, indeed, renders the interpreta-
tion of the central portion of the funnel difficult, is the col-
lection of a mass of coelomic corpuscles in the funnel. This
gives rise to the appearance represented in Goehlich’s figure,
and which was considered by him to form part of the wall of
the funnel ; but a careful examination, both of fresh specimens
and stained preparations, has convinced me that these cells do
not belong to the funnel, but are cells of the ceelomic fluid, in
some cases dead or dying, which will probably be carried to the
exterior, as Kukenthal has suggested. In the fresh state this
group of cells or “‘ débris ” appears darker and less transparent
than the cells of the funnel. Stained with borax carmine the
nuclei have an appearance distinct from those of the nephridial
cells, being brighter, more granular, and much smaller. In
fact, many have the appearance of cells which have died, and
have begun to disintegrate.

This mass of cells is shown in figs. 5 and 6. In one case
(that represented by fig. 6), in which the funnel was examined
twenty-four hours after the worm had been killed, not only
were the cilia still active, but some of the cells forming the
débris were still amcboid, one or two short pointed pseudo-
podia being present (fig. 6, a). Another case (fig. 5), where
the cells of the débris are relatively few, shows the shape of
the real mouth of the funnel, which is marked out by the
aggregation of the cells into a crescentic mass, It was this

THE NEPHRIDIUM OF LUMBRICUS. 299

specimen that first led me to a better understanding of the real
nature of the funnel, for no existing drawings or description
give any clue to the real mode of communication between the
celom and the nephridial tube.

I will now proceed to describe the true structure of the
nephrostome.

If we follow the “narrow tube ” forwards from the septum
to the funnel, we see in optical section the finely granular
wall on each side, with the nuclei of the component cells
alternately on this side and on that. Arrived at the centre of
the funnel, or thereabouts, the two walls suddenly diverge,
each bending outwards, and then sharply backwards nearly
parallel to its former course (fig. 4). The true “ drain-pipe ”
cells cease at this point of divergence. The backwardly
directed, or ‘ centrifugal cells,’ as they may be termed, are
merely grooved! (fig. 32), but otherwise resemble the drain-
pipe cells in structure, being granular and ciliated. These
grooved or “ gutter” cells, in reality, lie in a different plane
from the narrow tube, a higher plane when the funnel is
viewed from in front; and they frequently hide the wall of the
tube, as is the case in fig. 5.

After traversing a distance equal to about half the radius of
the funnel the gutter-cells suddenly cease, and become con-
tinuous with cells having quite a different appearance (fig. 4) ;
they are clearer and more distinctly marked off from one
another. They are, in fact, similar to the marginal cells, with
which they are continuous. The marginal cells are arranged
in such a way as to form what at first sight looks like a
circle, and has been figured as such; but what in reality is a
horseshoe, the ends of which are bent inwards, the in-turned
ends being very close to one another. The cells forming these
ends may be called the “ centripetal” marginal cells; and,
as above mentioned, they meet the “ centrifugal ”’ gutter-cells
(fig. 4).

The marginal cells are elongated, almost free from granules

1 T.e. the lumen of the drain-pipe cell has enlarged; the wall on one
side thins out till it parts, and the lumen is walled in only on one side.

300 W. BLAXLAND BENHAM.

Fic. 2.—The complete nephridium as seen (for the most
part) in optical section, without any blood-vessels. It shows
the character of the tubule in the different regions (note espe-
cially the ciliated tracts), the course of the tubule, and the
arrangement of the constrictions, so as to form loops. C.
The ampulla.

E., The muscular duct in the first “loop.” FF. Thesecond
“loop.” GG. The third “loop.” J. The preseptal region,
with the nephrostome. a, 0, c, d, e, f,g, A point to different
parts of the “ narrow tube.” O. The muscular duct, dipping
into body-wall to open externally.

a—b. The first ciliated tract of the ‘‘ narrow tube.” c—c’
(at the apex of the second “ loop”) is the second ciliated tract
of the narrow tube. d—e is the narrow tube in the third
“Joop.” e—e’. The third ciliated tract in the recurrent part
of the narrow tube. f to g, to A. The remainder of the recur-
rent part of the narrow tube lying in second “ loop.”

h—j. The ciliated “ middle” tube in the third “ loop.”

k—l. Part of the “ wide” tube in the third “loop.” J—m.
The ‘‘ wide” tube in the second ‘loop.’ m—n. The “ wide ”
tube in the third “loop.” 2 points tothe junction of the wide
tube with the muscular duct, where the tube is no longer seen
in section, but in surface view. 7. Nucleus of one of the cells
covering muscular duct. s. A muscle-fibre, part of the net-
work on the muscular duct. ¢¢. Vesicular connective tissue,

in which the tubule is embedded for the greater part of its
course.

PB.

Bese 5 canner
Seber ys

paper
Bae ¢

Fire. 2.

302 W. BLAXLAND BENHAM.

(hence are clear in the living state), with an oval nucleus ;
the cell boundaries are distinct, the outer free edge being
slightly rounded, and the whole of that surface of the cell
which is directed towards the funnel is ciliated. ‘The centri-
petal marginals gradually decrease in size as the centre is
approached, and become cubical, while the nucleus becomes
round.

The inner ends of the marginal cells do not reach to the
gutter-cells; and this space is usually figured, as by Goehlich
(20), by d’Udekem (81, pl. iii, fig. 6), by Howes (in the ‘ Bio-
logical Atlas’), and others, as being occupied by numerous
small rounded cells, considered as constituents of the funnel ;
these cells are, in truth, the ‘ débris”! to which I have
referred above, and have no structural relation to the funnel.

A very fortunate preparation shows the true structure of
the back of the funnel (fig. 4) ; the space between the central
ends of the marginal cells and the gutter-cells is occupied by
one large crescent-shaped cell, with a particularly large
oval nucleus containing a distinct nucleolus. This “‘ central”
cell is extremely difficult to see, for it is concealed by the
débris ; but having seen it in one funnel in which no débris
was present, I have been able to detect it in other cases in
which the débris was but slight; but generally it is completely
concealed. In sections it can readily be recognised, as
shown in fig. 7 (Ce.), which is the middle one of a series of
sections through a funnel in a young worm.

The actual mouth of the funnel in communication between
the ccelom and the nephridial tube is, therefore, placed
between the inner edge of the central cell on the one side (the
fine line a in fig. 4), and the outer edge of the grooved centri-
fugal cells of the other (the fine line 6). The marginal cells
and the general shape of the funnel are secondary, and are no
doubt developed in order to influence by the ciliary current a
greater extent of the celom. As to the probable steps in the

1 Beddard has figured such a mass of cells in the funnel of Urocheta (3,

pl. v, figs. 6 and 8; pl. xxiii, fig. 5), and has indicated it by “pg.;” he
believes them to be ccelomic cells.

THE NEPHRIDIUM OF LUMBRICUS. 303

evolution of the funnel of Lumbricus, &c., I shall have a
few words to say later on, in section 4.

The back of the funnel, as Vejdovsky has figured for A llo-
lobophora rubida (82, pl. xvi, figs. 15, 16), is covered by
a flat coelomic epithelium, the nuclei of which are readily dis-
tinguishable from those of the nephridial cells by their smaller
size, and denser character of their chromatin network (fig. 7).

II. The Post-septal Portion.—(1) The “ narrow tube,”
which is the longest portion of the whole nephridium, pos-
sesses a thin transparent wall, the lumen being relatively to
the diameter of the cell enormous (figs. 9, 10, 15). The pro-
toplasm is finely granular; the nuclei, which can be seen
readily in spirit or in moribund specimens at intervals on
alternate sides along the tube, are fairly large, oval, and present
adistinct nucleolus. This, in fact, is true of the nucleus of all
the cells entering into the formation of the nephridium, as
well as the tube itself, as in the funnel; the size and looser
nature of the chromatin coil render them quite distinct from
those of the surrounding connective-tissue cells.

For the greater part of its course the diameter of the lumen
of the “ narrow tube” is constant, but here and there certain
portions have a very irregular lumen (figs. 8, 8a). In fact,
in these cases, each cell, instead of being simply perforated by
a straight tube, has a slightly branched lumen, a condition
which is more marked in the nephridium of Microcheta
rappi, as I have shown (8, pl. xvi d7s, figs. 31, 33), as well
as in other earthworms, and suggesting the complicated con-
dition met with in Clepsine, Hirudo, &c.

Gegenbaur observed the fact that the cilia are not uni-
formly distributed along the whole course of the nephridial
canal, but are limited to certain regions of the “narrow tube”
and elsewhere. But in the exact limitations of these ciliated
tracts he seems to have been in error.

Now Goehlich contradicts Gegenbaur, and maintains that
the whole of the narrow tube is ciliated.! That this state-

1 This is also implied by Hatchett Jackson in ‘ Forms of Animal Life,’ 2nd
edit., p. 204.

304 W. BLAXLAND BENHAM.

ment is absolutely wrong—at least for the nephridium of
Lumbricus herculeus, Savigny (= L. agricola, Hoff-
meister, = L. terrestris, Linnzus)—a careful examina-
tion of a living nephridium will readily show. How far the
description of the excretory organ holds for all species of
Lumbricus I am not yet in a position to say; but from
examination of those of Allolobophora, sp., Criodrilus,
and other genera, I can confidently state that the cilia are
confined to certain limited tracts of the “ narrow tube ”
(and to the whole of the “ middle tube ”’).

There are three such ciliated tracts in the narrow tube:
the first extends from the funnel to just beyond the point
labelled “ 5” (in fig. 2)—it reaches, that is to say, just round
the bend of the tube where the latter enters the second loop ;
this tract was quite correctly described by Gegenbaur. The
second ciliated tract is at the apex of the middle loop (F),
between the letters c,c’ (fig. 2). I have observed that the
extent of this tract varies to a very slight degree; but the
cilia are always here, in the outer bend of the tube. The
third tract of cilia lies in loop G; it commences (at e) where
the narrow tube bends sharply upon itself, and extends about
halfway along this loop, ceasing at e’ (fig. 2).

Gegenbaur thought that the cilia recommenced at this point
e, and extended right along to the end of the narrow tube.

It is comparatively easy to satisfy oneself as to the existence
of these isolated tracts of cilia, either by observation on
the living nephridium, or in series of sections ; for if the whole
tube were ciliated we should expect the cilia to be visible in
every section of every tube if they were visible in any one
section, but such is not the case; not only in Lumbricus,
but in other genera of earthworms, I have sections in con-
secutive series which show cilia only in certain parts
(fig. 15).}

Wherever these cilia occur they are arranged in a spiral

1 Howes (‘Biological Atlas’), in his figure of a portion of the third loop,

shows correctly the structure of the different tubes, and represents one part
of the ‘‘ narrow tube” ciliated, and the parallel portion non-ciliated.

THE NEPHRIDIUM OF LUMBRICUS. 305

fashion, as Gegenbaur pointed out; not, however, along a
single spiral line, but along two spiral lines. This can be seen
in the fresh state, and in sections (figs. 9, 15); the cilia are
directed outwards, i.e. from the funnel towards the muscular
duct and external pore. D’Udekem (31) wrongly represents
the cilia directed in the reverse way (pl. iv, fig. 9).

(2) The “ middle tube” is quite short, occupying only one
side of the loop G. The recurrent portion of the narrow tube,
after following its previous course, arrives at the base of the
second loop (F, fig. 2), and here enters a suddenly dilated
region (at h), the commencement of the ‘‘ middle tube,” which
extends only to the apex of the third loop (G).

The width of the lumen and of the wall of this region is a
good deal greater than that of the narrow tube, but decreases
slightly in size after its commencement. In the living state this
middle tube has a brownish, semi-opaque appearance, and cilia
can be seen actively moving within ; under a high power small
pale yellowish spherules can be seen in the protoplasm of the
cells (fig. 11), and it is to these spherules that this middle
tube owes its opacity. These spherules are of different sizes,
the larger ones being nearer to the lumen. In the lumen
itself, being carried onwards by the cilia, are numerous
spherules, some smaller, others larger than those in the proto-
plasm, but otherwise similar to them (fig. 11). In transverse
section, too (fig. 15), this region is readily distinguished from
the other parts of the nephridium by the character of the wall,
i.e. by the spherular products of the protoplasm.

Gegenbaur mentions these spherules as occurring in this
and the following portion of the nephridium.

The whole of this tube is ciliated, and, as before, the cilia
are arranged along two lines! (fig. 15).

(3) The “wide tube” commences at the apex of the loop
G, and has the course shown in fig. 2.

1 In Marshall and Hurst’s ‘ Practical Zoology,’ 2nd edit., the statement is
made on p. 65 that this and the following regions of the tube have “ proper
cellular walls,’ implying that the lumen is intercellular. This is certainly not
the case; the lumen is distinctly “ intra-cellular.”’

306 W. BLAXLAND BENHAM.

At the apex of the loop the middle tube opens into a bladder-
like enlargement (C’) of the tube, called by Gegenbaur the
‘“‘ampulla;” this is the commencement of the “ wide tube.”
The ampulla is bent sharply upon itself, and gradually nar-
rows in diameter till it reaches about twice that of the middle
tube, which it retains throughout its course. There are no
cilia in this part of the tube, notwithstanding Goehlich’s
statement that it is ciliated.

The wall of the ampulla is very different from that of the
other regions. The cells are relatively very large, but are still
‘‘drain-pipe”’ in nature; their general arrangement and their
shapes are shown in a figure taken from a living though mori-
bund specimen (figs. 12, 30). The substance of the cell is
divisible into two well-marked parts, a central and a peri-
pheral portion (see figs. 13, 15), easily recognisable in a living
nephridium. So marked is the distinction between the two
parts, that in section it appears as if the perforated cells were
surrounded by others, as Beddard (4) suggested might be the
case in his description of Allurus; but such is not the case.
A series of sections and examinations of living nephridia show
that this appearance is really due to the modification of the
protoplasm bordering the lumen.

The central portion of the protoplasm, which forms a thin
lining to the lumen, is made up of numerous small granules
arranged radially, giving the impression of a radial striation
(figs. 18, 15). In preparations stained with borax carmine
these granules take the dye much more strongly than the other
parts of the nephridium, and give this ampulla a very charac-
teristic appearance. ‘The peripheral portion, which surrounds
the central portion, forms a much greater bulk of the cell, and
possesses much smaller granules, amongst which the nucleus
is situated. The granules here also are arranged in rows,
though less distinctly than in the central portion. A peculiar
appearance, as of nuclei, is presented sometimes by this peri-
pheral portion of the protoplasm: deeply stained, usually
irregular, though sometimes regular, masses of different sizes
are present (fig. 15). As a rule each mass, or sometimes a

THE NEPHRIDIUM OF LUMBRICUS. 307

group of small bodies, is surrounded by a clear space or vacuole.
In the living nephridium the ampulla is seen to be filled with
larger and smaller vesicles (fig. 13), which are probably the
excretory products of the protoplasm of the tube; some, no
doubt, pass into the ampulla from the preceding region—the
“middle tube;’’ while others, and perhaps the larger globules,
are, I believe, formed by the protoplasm of the cells of the
ampulla, and the deeply staining masses in the peripheral
portion of the protoplasm seen in sections are the above-
mentioned globules or excretory products in situ, the sub-
stance being precipitated by the alcohol, stained.

Passing away from the ampulla, the central and peripheral
portions of the protoplasm become gradually less distinct, till
finally no such difference is to be observed; at the same time
the wall of the lumen becomes relatively thinner. The granules
in the protoplasm still retain a radial arrangement, though this,
too, becomes less and less marked further from the ampulla.

In the greater part of the wide tube the wall is compara-
tively thin, but is not of uniform thinness; here and there it
widens ; in fact, each cell possesses a zone of wider protoplasm
somewhere near its middle, in which the nucleus is situated ;
hence the margin of the wide tube is wavy (fig. 2). This
was figured by Horst in his paper on the anatomy of L.
terrestris.

(4) The muscular duct of the nephridium is considerably
developed in Lumbricus; it is much wider than any of the
preceding regions, and differs from these in the presence of
abundant muscle-cells forming a trellis-work around the
enclosed duct (£).

The lining epithelium of this region is very difficult to make
out. It is usually stated that the lumen is here intercellular,
and we should expect, if this be the case, to find a number of
cells, with their nuclei fairly close together, arranged around
the tube ; but in Lumbricus we do not see this. After exa-
mining several preparations and series of sections in various
directions, and treated in different ways, I still feel uncertain
as to the true arrangement,

A

308 W. BLAXLAND BENHAM.

Frequently in a section, or even in several consecutive
sections, no nucleus appears ; then perhaps a section will show
only one, or others show two, and very rarely three (fig. 16) ;
these nuclei are quite similar to those in other parts of the
nephridium—oval, with a nucleolus; and they appear to be
embedded in a finely granular though very thin layer of proto-
plasm, showing a feebly reticular structure (fig. 16, ep.); in
which, however, I can detect no cell boundaries.

Immediately outside this layer of protoplasm comes the
muscular coat (musc.), which consists of cells arranged in
various directions (fig. 16) ; some are longitudinal, others cir-
cular; still others are oblique, in relation to the long axis of
the duct. These muscles appear also to be embedded in finely
granular protoplasm (cep.), in which small round nuclei are
scattered. It is difficult to make out a continuous layer of
flattened ccelomic epithelial cells outside the muscular coat,
either in section or in nephridia treated with nitrate of silver.
From the appearance presented in many sections this granular
protoplasm seems to represent the vesicular connective-tissue
cells plus ccelomic epithelium of the other parts of the tube.

The muscular duct penetrates the body-wall, the muscles
appearing to be continuous with those of the circular layer ;
a slight invagination of the epidermis meets the nephridial
lumen, and puts the latter into communication with the
exterior.

With a view to elucidating the character of the epithelium
lining this part of the tube I have examined sections through
other earthworms, and amongst these Microcheta rappi, in
which the muscular duct is of enormous size (see my paper
[8], pl. xvi, fig. 21). In this worm the lining of this region of
the nephridium consists of very numerous vesicular cells, quite
unlike what we have in Lumbricus; they are small, with
distinct boundaries, and appear to be but loosely attached to
the wall. This layer of cells can be traced through the body-
wall up to the circular layer of muscles, where it meets the
epidermic pit. I think, therefore, that we may conclude that
in Lumbricus, too, the lumen of this region is intercellular,

THE NEPHRIDIUM OF LUMBRICUS. 309

although the cells are of very great size and relatively few in
number, whereas in Microcheta they are small but very
numerous: in fact, we have perhaps a stage in the transition
between perforated cells and a more definite epithelium.

3. CoMPARISON WITH OTHER GENERA.

Although we have, in numerous recent contributions, outline
figures of the complete nephridium with the funnel of various
Oligochzeta, yet we have very few details as to the histological
structure of the different parts of the tube.

Amongst the majority of the Microdrili (in the “ limi-
colous” Oligochetes) there appears to be no distinction
of the nephridial] tube into more than two regions : for example,
amongst the Enchytreide (see Michaelsen’s and Vejdovsky’s
works) we find (a) the post-septal portion consists of a more
or less extensively convoluted “narrow tube” embedded in
a mass of vesicular cells; and (0) a preseptal portion, con-
sisting of the simply constructed funnel and a short ciliated
tube leading to the post-septal tube (see Pl. XXIV, fig. 27).

In Tubifex, however, where the coils of the nephridium are
free, Vejdovsky (82) figures (a) a narrow region, (4) a middle
or glandular region, and (c) a terminal or wide region, in addi-
tion to (d) the preseptal region. As in Lumbricus, the tube
is surrounded by vesicular cells (see also Kukenthal, 24).

In the literature of the earthworms or Macrodrili, also, the
detailed information as to the character of the tube of the
nephridium is very scanty. Beddard (6) describes the different
calibres of the tube in Pericheta aspergillum and P.
armata, where three regions are evidently present, cor-
responding to the “‘ narrow,” “ middle ciliated,” and ‘‘ wide”
tubes; the last has striations in its wall. In Ac. multi-
porus a similar variation in calibre is noted (loc. cit., fig. 14),
but only two regions are apparently distinguishable. Beddard

1 Michaelsen, in his description of Pachydrilus sphagnetorum (‘ Arch.
f. Mikr. Anat.,’ xxxi, p. 491), states the whole of the tube is ciliated; and in
Enchytreus Mobii and others, he figures this tube as slightly dilated just
before the external aperture.

310 W. BLAXLAND BENHAM.

has also figured similar different regions in Allurus (4, pl. xxv,
figs. 11, 12).

Spencer (30) describes and figures (pl. vi, fig. 26) for Megas-
colides the funnel and ciliated preseptal region of the large
posterior nephridia, as well as the “ wide” intra-cellular tube
which passes into the body-wall as far as the circular muscles.
At the junction of this layer with the epidermis a group of
muscle-cells, arranged as a sphincter around the nephridial
tube, is shown, and has a great resemblance to the condition
found in Criodrilus(Pl. XXIV, fig. 17). Spencer also gives
some details as to the opening of the small nephridia to the ex-
terior, in which there is a short intercellular portion interposed
between the epidermic pore and the intra-cellular tube.

Perrier some years ago (26, pl. xvi, figs. 38, 39) described
with some detail the nephridium of Urocheta. He seems to
imply that the whole tube is ciliated, with the exception of a
very small muscular duct. More recently Beddard (3 and 6)
has paid more attention to this genus. He indicates at least
two regions (the ‘‘ narrow” and “ middle”’ tubes) ; he does not,
however, represent the cilia, which I believe are present only
in certain tracts, and not throughout.

Pontodrilus was also investigated by Perrier (27), who
describes and figures (pl. xiv, figs. 10, 11, 13) the nephridia.
The anterior nephridia (in somites 14, 15, 16, 17) differ
slightly from the remainder, which have very abundant vesi-
cular cells around the convolutions of the tube, much as in
Enchytreids. The tube appears from his description to be
similar throughout, for he speaks of it as a ‘‘ sinuous glandular
tube, ciliated internally throughout” its length. His figure
of a section closely resembles that of the “ middle” tube in
Lumbricus.

In my first ‘Studies on Earthworms’ (8) I figured and
described certain differences in the character of the tube in
the nephridium of Microcheta rappi; and this genus I
have re-examined, and can confirm my statements.

I believe the above are the only earthworms in which any
attempt at histological detail has been made, and no author has

THE NEPHRIDIUM OF LUMBRIOUS. 311

made any observations as to the different characters presented
by different parts of the tube except Beddard. I have, therefore,
examined sections through a number of earthworms of different
genera, in order to ascertain how far there is any general agree-
ment between their nephridia; and I find that such an agree-
ment is present, with the exception that the muscular region
exhibits a great variability, sometimes being entirely absent
(as in Criodrilus), sometimes being enormously developed (as
in Microcheta), and in other cases presenting intermediate
conditions.

The nephridium of the undermentioned worms presents the
same three regions that I have described for Lumbricus,
viz. (1) a non-ciliated narrow tube ; (2) a ciliated middle tube ;
and (3) a non-ciliated wide tube, which has frequently a wall
of irregular thickness. In all cases the whole of these three
regions is intra-cellular, and this may dip into the body-wall
directly, or enter (4) an intercellular muscular duct as in
Lumbricus. The funnel, however, presents a good deal of
variety in structure, as I will describe below in section 4.

The worms I have examined! are Criodrilus (foradrawing
of funnel see Vejdovsky, 32).

Allurus (see Beddard,4, pl. xxv, figs. 11, 12, and Vejdovsky,
32).

Microcheta (see Benham, 8, pl. xvi, figs. 21 to 26; pl. xvi
bis, figs. 31, 32, 33).

Urobenus (see Benham, 9, pl. viii, figs. 18, 19).

Brachydrilus.

Rhinodrilus (see Beddard, 1a, pp. 160, 161; and Horst,
21, fig. 8).

Urocheta (see Perrier, 26, pl. xvi, figs. 88, 89; Beddard,
3, pl. v, figs. 5 to 9; Beddard, 6, pl. xxiii).

Diachzta (see Benham, 9, pl. ix, figs. 27, 28).

Perionyx.

1 It may be worth while to add references to the figures of complete nephri-
dia, but which give no histological detail of other genera :
Hormogaster, Rosa, 28, fig. 8.
Moniligaster, Horst, 21, fig. 2.

VOL. XXXII, PART III.—-NEW SER, x

312 W. BLAXLAND BENHAM.

Eudrilus (see Beddard, 2, pl. xxxiil, fig. 17; and Horst,
22, figs. 3, 4).

Some of these deserve further mention.

In Criodrilus the narrow tube is very much more extensive,
and undergoes a greater amount of coiling than in Lum-
bricus, and herein rather agrees with the Microdrili. There is
no muscular duct, the wide tube penetrating the body-wall as
far as the circular muscles (Pl. XXIV, fig. 17). It is sur-
rounded by a very loose connective tissue as it passes through
the longitudinal muscles ; and this connective tissue is modified
just before the tube reaches the circular muscles. Here can be
seen seven or eight, or perhaps a few more, nuclei belonging to
certain cells, which are elongated transversely to the long axis
of the tube (fig. 17,y) ; they resemble, in position and arrange-
ment, the “sphincter muscle-cells ” figured by Spencer around
the tube of the large nephridia of Megascolides; but in
Criodrilus they are certainly not muscle-cells.

At this point, when the tube has reached the circular
muscles, it becomes difficult to be sure of the exact nature of
the wall—i.e. whether it is formed by perforation of cells
or by an epithelium ; for the nephridial tube undergoes a slight
flexure here, in order to reach the deep but narrow pit formed
by invaginated epidermis, and constituting the nephridiopore.

A study of several series of consecutive sections through this
region, however, gives me the impression that there is a very
short intercellular tube between the wide tube and the epi-
dermic pit: this then would correspond to the duct or muscular
region of Lumbricus deprived of muscle-cells, the contrac-
tion of this part being, when necessary, brought about by the
muscles of the body-wall. This anatomical difference between
Lum bricus and Criodrilus seemed, when I first observed it,
to agree with the different descriptions of the development of the
nephridia given by various authors; some affirming the exist-
ence of an epiblastic invagination to meet the mesoblastic tube,
others denying this invagination, e. g. Bergh in Criodrilus
(12), where, indeed, we should not expect it to occur; but
Bergh has quite recently (13) emphatically denied it also

THE NEPHRIDIUM OF LUMBRICUS. 313

in certain species of Lumbricus and Allolobophora, where
he states that, as in Criodrilus, the wholenephridium is derived
from mesoblastic cells which grow outwards, and ultimately
pass between the epiblast-cells to reach the exterior.

A very interesting peculiarity in the “narrow tube” was
figured by me in 1886 for Microcheta rappi (8), where I
show the “smaller lumina ” (/’, pl. xvi bis, figs. 31, 33), as I
called this part of the tube, branching and anastomosing
so as to form a network around the two other regions of the
tube (J. /’.).

No notice has been taken of this network in the recent dis-
cussions by Beddard and Spencer, as to the derivation of the
nephridia of earthworms from a “ plectonephric” condition,
such as occurs in Acanthodrilus, Megascolides, and
many others ; and yet it seems to me not without interest.

Quite recently a similar condition of things has been men-
tioned by Rosa (29) in the nephridium of Desmogaster,
one of the Moniligastride; Vejdovsky, too (82), figures the
branching of the lumen in the case of Chetogaster.

Again, in Brachydrilus, where there are two pairs of
nephridia in each segment, each having the same structure as
that of Lumbricus, a similar network is formed by the
“narrow tube” (see Pl. XXIV, fig. 26), and which is almost
identical with that of Pericheta, &c.

Now what meaning is to be attached to this network in
relation to the “‘ plectonephric” condition? (1) Are the two
networks homoplastic, i.e. has each been developed indepen-
dently, or are they homogenetic? and if the latter, (2) is the
“plectonephric” condition derived from the “ meganephric ”
condition (of Lumbricus, Urobenus, Eudrilus, &c.) by the
branching of the tubule; or (8) is it a remnant of a plec-
tonephric condition still retained by the meganephric genera ?

(1) The suggestion that the network in Brachydrilus and
Microcheta (and no doubt this network will be found else-
where) is homoplastic with that of Acanthodrilus might
be held; but if the tubule can branch independently of any
ancestral network, there is no reason to regard the plecto-

314 W. BLAXLAND BENHAM.

nephric condition of Acanthodrilus as related to that of
Pericheta and other worms; and it would have no syste-
matic meaning at all, but might have been developed in any
genus, and at any period in its history. This assumption
would, no doubt, explain many difficulties, such as the pre-
sence of a network in Pericheta, and of large nephridia in
Perionyx, or the differences in different species of Acan-
thodrilus. In fact, the character of the nephridium would
be no guide as to the affinities of worms.

(2) The assumption that the meganephric condition is ar-
chaic would, I think, plunge us into a sea of difficulties ; for
if a large nephridium is the more primitive, then we have to
explain how it comes about that in some genera and species
the paired nephridia are in relation to the inner couple of
sete, and that in others they are in relation to the outer
couple; how is it that in some worms, e.g. Perionyx
saltans and Ac. nove-zealandiz, the position of the ne-
phridiopores alternates from segment to segment? and finally
how can we explain the presence of two pairs of nephridia per
segment in Brachydrilus? For if a large nephridium is
ancestral, was there but a single pair, or two pairs, or four
pairs? The last assumption would of course explain all the
above queries.

If the plectonephric condition is derived from the branching
of one pair of large nephridia per segment, then the double
pair in Brachydrilus would have to be regarded as a secon-
dary condensation of tubules after the appearance of a plec-
tonephric coudition ; and the possibility would then arise that
a large nephridium might be either primitive or secondary.

Ontogeny has not yet revealed to us any sufficient evidence
on this point; the only plectonephric form studied is Acan-
thodrilus. According to Beddard (‘ Proc. Roy. Soc.,’ 1890)
there is at first a single pair of large nephridia in each seg-
ment, and these by branching give rise to the characteristic
network. This looks like strong evidence in favour of Horst’s
view of an ancestral paired meganephric condition. But, as
just stated, we should then have to meet the above difficulties

THE NEPHRIDIUM OF LUMBRICUS. 315

by supposing the branching and anastomosing of four pairs
of large nephridia in order to give rise to the condition found
in Ac. multiporus, where we have eight groups of tubules
per segment ; a further extension of the anastomosis leads to
the more complete plectonephric condition of Pericheta
aspergillum.

Megascolides would be explained by one of the four
pairs remaining simple (in the posterior part of the body), the
rest branching ; still further, reduction to two pairs leads to
Brachydrilus, and again to Lumbricus, Urocheta, &c.

But it seems to me that this is a strained hypothesis. For,
after all, the occurrence in the ontogeny of Ac. multi-
porus of the paired nephridia may be merely ccenogenetic,
and have no meaning of an ancestral nature; it would come in
the same category as the formation of the heart in mammalia
from a double rudiment.

The paired nephridia of Acanthodrilus! may, therefore,
be merely provisional, like the embryonic nephridia described
by Vejdovsky in Rhynchelmis and in Lumbricus, and
those of embryo Pulmonates and other animals.

(3) It therefore remains for us to regard the branching tubes
of Brachydrilus and other “ meganephric ” genera, as well
as of the strictly ‘‘ plectonephric genera,” as homogenetic,
and the plectonephric condition as the more archaic. This
is the position taken up by Beddard and by Baldwin Spencer,
although there is some difference as to details in the two
theories.

Spencer has pointed out the parallel series of stages in
Oligocheta and in Hirudinea ; and it is interesting to com-
pare the nephridium of Hirudo (see 14, p. 485) with its net-
work of fine intra-cellular tubes around the main intra-cellular
tube with that in Brachydrilus; and the less branched and
non-anastomosing tubule in Clepsine (14, p. 483) with the
condition met with in Lumbricus and other earthworms.

1 If this be the case, then it would appear that the genital ducts in Acan-

thodrilus (and perhaps in other worms) arise, in part at least, by a modifi-
cation of the embryonic provisional nephridia.

316 W. BLAXLAND BENHAM.

Again, one species of Dinophilus has been described as
having a network of nephridial tubules; and another species,
by Harmer, as having simple unbranched nephridia.

It appears possible that the meganephric condition may
have arisen, as Spencer suggests (30, p. 43), either (a) by en-
largement and elaboration of one tubule of the network, or
(5) by an aggregation of a portion of the network.

Spencer does not state which alternative he believes to
obtain in Megascolides, but probably the former, as he men-
tions no network in the large hinder nephridia, which he
implies are simple unbranched tubules. He remarks that it
would be impossible to map out the course of the tubules by
means of sections: this might, however, be done by examina-
tion of the whole nephridium, as I have done for Micro-
cheta; and I am inclined to regard the second alternative—
aggregation of a portion of the network—as the probable
mode of origin of the large nephridium (Pl. XXYV, fig. 42).
The tubule might become enlarged, lose some of its branches,
and thus present a looser network; it would then become
differentiated into “ wide” and “narrow” regions, and ulti-
mately lose all connecting tubules, so as to give rise to the
complicatedly coiled tube of Lumbricus, &c.; while
Microcheta and Brachydrilus would represent the inter-
mediate stages with still some of the primitive network in
part of the course.

Another intermediate condition appears to be presented by
the “ peptonephridium” of Urocheta, which retains some of
the branches and three funnels, but has only one external
aperture.

4. Tus FUNNEL oF PERICHAZTA MALAMANIENSIS, N. SP.,
AND SOME OTHER GENERA.

In my earliest contribution to lumbricological literature
(8) I stated that a certain species of Pericheta from Mala-
mani, amongst the Philippines, possessed numerous small
nephridia in each segment with several funnels. I believe I
was the first zoologist to mention this multiplicity of nephridial

THE NEPHRIDIUM OF LUMBRICUS. 317

funnels in Oligocheeta, a character which, with the rapid exten-
sion of our knowledge during the last five years, has received
considerable attention, owing to the careful researches of
Beddard, and, more recently, of Spencer.

The species, of which I hope to give a more detailed account
in a future paper, appears to be a new one, to which I give
the above name. It agrees with P. quadrigenaria, Perrier,
P. elongata, Perrier, and P. exigua, Fletcher, in possessing
only one pair of spermathece ; but it differs from each of these
three species in various anatomical features.

As the funnel of the nephridium, which has the typical plec-
tonephric character, differs from that figured by Beddard (6,
pl. xxiii, fig. 10) for P. aspergillum, it appears worth while
to give a figure of it (see Pl. XXIV, fig. 25,a@ and 6). It con-
sists of eight or nine marginal cells set in a circle around the
terminal aperture of the tubule. All the cells are equal in size,
and each is ciliated over the whole of the centrally directed face,
the other face being covered by a few ccelomic epithelial cells.
There is no “central cell,” nor any centrifugal gutter-cells.
In P. aspergillum, however, the marginal cells are arranged
in a horseshoe fashion, as in Lumbricus (see Beddard’s
drawing), but without the marked incurving of that genus,
although a few smaller cells figured may represent either the
completion of the circle or the “ centripetal cells.” However
that may be, the funnel in this species, having a greater
number of cells (some fifteen in number), which vary in
size, is at a higher stage of development than that of P.
malamaniensis.

I will add a few remarks and suggestions as to the funnel in
various earthworms ; for although there are but scanty mate-
rials for generalising, yet I think that, even with the few
careful descriptions and figures, we can make out something
of the evolution of the nephrostome in Oligocheta.

More or less careful figures of the funnel of Pericheta,
Megascolides, Urocheta, Microcheta, Urobenus,
Criodrilus, Allolobophora, and Pontodrilus occur in
the literature (see above); but the majority of authors have

318 W. BLAXLAND BENHAM.

contented themselves with a brief mention of the fact of its
presence, or given only a rough sketch of it. It appears to me
that, just as the character of the nephridium itself in various
genera is of interest and importance as representing to some
extent probable stages in the phylogeny of the organ, so the
nephrostome will indicate such steps.

Amongst the leeches, we know from Bourne’s researches that
Clepsine has a funnel composed of only two cells (14, fig. 52) ;
other leeches show an increase in number of these cells; and
Hirudo, with its more complicated nephridium, possesses a
very large number of funnel-cells, each perforated or partially
perforated by a branch of the nephridial tubule.

Amongst the Microdrili, such as Tubifex and Enchy-
treus, two is the normal number of cells composing the
funnel. In Rhynchelmis there are eight such cells. In
the plectonephric earthworms, such as Acanthodrilus, and
the smaller tubules of Megascolides no funnels are present.
In Pericheta from eight to fifteen marginal cells form the
funnel; in the large tubules of Megascolides a larger
number of cells are present, which from Spencer’s drawing
(30, pl. vi, fig. 26) appear to be arranged in a complete
circle round the aperture, as in P. malamaniensis. In
the higher meganephric condition we come across larger
funnels, as a rule, with a great number of marginal cells,
which form an incomplete circle, and differ in size according
to their position.!

In my figure illustrating this anatomical point of Uro-
benus (9, pl. vii, fig. 19) I have drawn a number of cells
curving inwards from the ends of the marginal cells, which
are arranged in a horseshoe. These cells are no doubt the
‘centripetal marginals”? and ‘‘centrifugal gutter-cells” of
Lumbricus, although when I made the drawing I had a very
imperfect idea as to their meaning. For Urocheta, Perrier
(26, pl. xvi, fig. 42) gives a large drawing of the funnel, which
presents a very wide aperture to the celom; the margin of

1 An exception is to be noted in the case of Microcheta rappi, where
the funnel seems peculiar, and may perhaps be degenerate.

THE NEPHRIDIUM OF LUMBRIOUS. 319

the funnel is deeply incised at the front, and the cells forming
this part are apparently quite continuous with the other mar-
ginal cells; but I think that it is very probable that the funnel
of Urocheta is formed on the same plan as in Lumbricus.
Perrier was unable to distinguish any nuclei in the wall of the
widely dilated portion of the tube immediately below the margin
of the funnel—between, that is, the nuclei of the marginal
cells and those of the intra-cellular tube itself,—and suggests
that the marginal cells are extremely elongated.

Beddard (3 and 6) regards this dilated region as being intra-
cellular. In the former paper he gives figures of sections
through the funnel of the ordinary nephridia, in which there
is—contrary to Perrier’s figure—no very marked dilatation of
the tube; but the funnel appears to consist (fig. 7) of several
rows of non-perforated cells (although such is, perhaps, not
intended, and probably is not the case). Of the peptonephri-
dium, Beddard (6) figures the funnel, and there he shows the
perforated cells (? = “ grooved cells”) diverging from one
another, so as to give rise to avery wide “ intra-cellular” tube
before the actual aperture is reached. The whole margin is
formed by one row of marginal cells, and he makes no
mention of the incurving of the margin represented by
Perrier.

For Pontodrilus, Perrier (27, pl. xiv, fig. 12) figures only
the side of a nephrostome covered by celomic epithelium, and
gives little detail in the text ; from it I cannot be sure whether
the perforated cells reach to the marginal cells simply as in
Pericheta, or whether there is any widening of the tube, as
in Urocheta.

In Rhinodrilus (Thamnodrilus), Beddard (1a, p. 161)
gives a figure of a complete nephridium,and describes the funnel
as an “elongated, folded membrane, composed of ciliated
columnar cells.” I have removed a nephridium from a speci- -
men of the same species (Rh. gulielmi) ; and I can confirm
this statement, but will add that the funnel is quite similar to
that of Lumbricus (with marginal cells, centripetal and cen-
trifugal cells) if we imagine it to be, not circular, but drawn

320 W. BLAXLAND BENHAM.

out on each side to a very considerable extent, so as to be two
or three times as wide from side to side as from front to back.
Even with such scanty details as we have, I think it is
possible to trace a few stages in the evolution of the compli-
cated funnel of Lumbricus, Rhinodrilus, &c. Starting
with the nephrostome of the Enchytrzidz, for example, we
have a funnel formed merely by the terminal perforated cell
without accessory marginal cells (see Pl. XXIV, figs. 27, 28,
29, a). By the subdivision of this cell, as in the ontogeny of
Stylaria, the lip of the aperture is formed by two cells (as is
the case also in Clepsine amongst the leeches) ; in Tubifex
also two or a few more (a similar increase is observed in
Pontobdella). In Rhynchelmis—one of the nearest of
the waterworms to earthworms—eight marginal cells have
become formed, as is the case, too, in Per. malamaniensis
(Pl. XXIV, fig. 25); now a differentiation occurs amongst
the cells, as in P. aspergillum. In Urocheta, in the
peptonephrostome, not only have the marginal cells greatly
increased in number, but the intra-cellular tube is dilated (see
diagram, Pl. XXIV, fig. 25, 0), so as to increase the effect of
the ciliated margin. Suppose this dilatation were carried
further, so that the sides diverge, the ‘perforated cells”
would become “ grooved cells,’ and would lie between the
intra-cellular tube and the marginal cells (diagram, fig. 25, c).
I am not sure that any of the earthworms studied correspond
to this stage. A little further differentiation, by a still further
divergence and an outward curve of the “‘ grooved cells,” leads
on to the stage presented to us by Rhinodrilus and Lum-
bricus (cf. diagram, fig. 25, d, with Pl. XXIII, fig. 4).
These large nephrostomes replace the numerous small ones of
Pericheta, such being evidently more efficient for the removal
of phagocytes than is the smaller funnel of the latter genus.

5. Tue Vascutar Suppty or tHE NEPHRIDIUM.
The living nephridium of an earthworm is a remarkably
beautiful object when viewed under a moderately high power ;
and this is due to the delicate and complex network of blood-

THE NEPHRIDIUM OF LUMBRICUS. 321

vessels which twine amongst themselves and between and
around the tube of the nephridium (fig. 3).

Although the existence of this elaborate blood-supply has
long been known (for instance, Morren describes it), and was
indeed adopted by Claparéde as one of the distinctive cha-
racters of earthworms as opposed to water-worms, yet no
drawing or detailed description appears to have been pub-
lished.

The remarkable dilatations of the blood-vessels here and
there were observed by Williams long years ago; they re-
ceived further attention from Lankester and from d’Udekem,
who figured them (81, pl. iu, fig. 27), and a little later from
Claparéde (16). In Pontodrilus, Perrier (27, pl. xvii, fig.
35) observed them; and in Diacheta, I pictured them (9,
pl. ix, figs. 35, 36).

It is to be noted that these dilatations are by no means
constantly present in every earthworm. I have frequently
examined dozens of nephridia from one specimen of Lum-
bricus, and seen no dilatations whatever (as fig. 22) ; whilst
in other specimens examined at the same time, from the
same locality, and apparently in the same sexual condition, I
have seen these dilatations in abundance. But, again, not
every nephridium in the same worm presents this character ;
in some they are rare, in other nephridia they are altogether
absent.

As is well known, they occur not only on the nephridia, but
on the septa ; and are, as Claparéde figured, usually filled with
corpuscles (as in Pl. XXIV, fig. 18).

The cause of their appearance and the use of them to the
worm is a matter, at present, of speculation. Gegenbaur (19)
noted them and their inconstancy, and suggested that they
are present when the gonads are fully ripe; but they seem to
be independent of this condition.

Another peculiar phenomenon is sometimes to be seen in
the blood-vessels in the nephridium ; the colour of the blood,
instead of being pale yellow, as is normally the case, is more
distinctly red—a pinkish red—comparable to the colour of

322 W. BLAXLAND BENHAM.

Fic. 3.—The vascular supply of the nephridium, the loops
of which are represented in outline. H#. The first loop. FF.
The second loop. GG. The third loop. N. Nerve-cord. S.
Septum. SN. Subneural blood-vessel. V. Subintestinal
blood-vessel. @. Somatic vessel from subintestinal vessel. 0.
Its septal branch. c. Its nephridial (afferent) branch. c’ c’.
The further subdivision of this. d. Ventral portion of a com-
missural vessel, connecting the dorsal vessel with the subneural.
e. Its septal branch. jf. Its nephridial (efferent) branch.
SJ f'. Its further subdivisions to third loop. gg’. Dilatations
in the capillary vessels.

324 W. BLAXLAND BENHAM.

venous as opposed to arterial blood; and in fact, as Professor
Lankester suggested to me, this is probably due to the reduced
condition of the hemoglobin in the blood.

The vessels supplying the nephridium, as shown in the dia-
gram (fig. 3), are connected with the subintestinal vessel (V)
on the one hand, and with the dorsal vessel and subneural vessel
(SN) through the commissural vessel (@) on the other.

It is a difficult matter to decide what course is taken by the
blood outside the large median trunks, where it flows forwards
in the dorsally, and backwards in the ventrally situated ves-
sels; it also passes downwards from the dorsal to the sub-
intestinal vessel in the lateral hearts ; but what is its course in
the commissural vessels in the segments behind the gizzard ?

A couple of years ago, when studying the arrangement of
vessels in Lumbricus, I succeeded after considerable trouble
in injecting the dorsal vessel in two specimens; the blue
injection passed forward readily, but only in one or two seg-
ments did it pass into the commissural vessels. I have a note
that “internal intestinal plexus and typhlosolar sinus received
injection,” but that only “in a few cases did injection appear
to pass on to the wall of intestine.” By observation of the
narcotised worm it can readily be seen that blood flows into
the dorsal vessel from the commissural vessels, and this is
confirmed by the arrangement of the valves at the entrance.

The course of the blood appears to be different in Micro-
cheta, Urocheta, and Megascolex.'

I believe from the above experiments and observations (which
I intend to pursue further) that the blood usually passes from
the subneural (SN) into the dorsal vessel by way of the com-
missural vessels (d) ; the latter receives, amongst other branches,
one from the body-wall (e), another from the nephridium (/) ;
the blood is therefore aérated in the body-wall and denitro-
genised in the nephridium before it reaches the dorsal vessel.

The blood, in passing backwards along the subintestinal
trunk (V), passes outwards through the lateral (a) branches
to the nephridium (c), and to the body-wall (6). The former

1 A, G. Bourne, ‘ Quart. Journ. Micr. Sci.,’ 1891, February.

THE NEPHRIDIUM OF LUMBRICUS. 325

divides up into branches to the three loops of the nephri-
dium, and these branches subdivide; the ultimate capillaries
wind in and out around the individual tubes, or around the
whole bundle of tubes in a loop, as seen in figs. 19—24.

The blood-vessels to and from the nephridium pass along
fenestrated membranes as shown in fig. 3, at h, which thus
serve also to suspend the nephridium. In some cases I have
seen a number of small twigs given off from a vessel in the
septum, passing to the nephridial vascular network, in addi-
tion to the main supply (fig. 23).

An examination of the various figures will, I think, suffice
to explain the relations and variations of the elaborate and
delicate vascular network, and will render further remarks
unnecessary.

6. Tur NEPHRIDIUM oF ARENICOLA.

While I was working in the laboratory of the Marine Bio-
logical Association at Plymouth, during a short time in last
summer, I took the opportunity of examining the nephridia of
some of the readily obtainable Polycheta, amongst them
Nereis, Aphrodite, and Arenicola, and I here introduce
a figure of the nephridium of the last worm for comparison
with that of Lumbricus.

The figure (Pl. X XV, fig. 33) is drawn under the low power,
and exhibits, amongst other points, the elaborate vascular
network, whilst the sections demonstrate the intercellular
character of the wide lumen of the organ.

The nephridium of Arenicola has recently been described
by two authors, namely, Cosmovici in 1879 (17), and Cun-
ningham in 1887 (18); and I have, indeed, nothing to add to
their descriptions. Cosmovici figures the nephridium in situ
(pl. xx, fig. 10), and gives various drawings illustrative of
the minute anatomy; though, as Cunningham has pointed
out, his description of the lining epithelium is incorrect, inas-
much as he considered it to be stratified, which is not the
case,

326 W. BLAXLAND BENHAM.

Both the above-named authors figure and describe the net-
work of blood-vessels on the funnel and the body of the
nephridium, but neither mentions the blind dilated termina-
tions and irregularities in the capillary vessels (Pl. XXV, fig.
34), recalling those in Lumbricus ; Cosmovici, however, does
mention these as occurring elsewhere in Arenicola.

Notwithstanding the general accuracy of the descriptions
given by these authors, I have thought it worth while to
introduce another figure of the whole organ, which to my
mind represents the form and relations of the parts more
clearly than either of those already published.

The nephridium of A. piscatorum (Pl. XXV, fig. 33)
consists of (a) a large, wide, tapering tube or “ body,” lined by
a single layer of ciliated vesicular cells (fig. 40) ; (0) of a very
wide funnel attached to the broad end of the “body” by a
narrow “ neck ;” (c) of a dilated, more muscular region, open-
ing to the exterior and separated from the narrow part of the
body by a well-marked constriction.

A very noticeable contrast to the nephridium of Oligo-
cheta is presented by that of Arenicola and other Poly-
cheta, e.g. Polynoé (A. G. Bourne, 15, pl. xxvi, figs. 22,
23, 24), in that the lumen is intercellular throughout ; and
this point deserves mention because Baldwin Spencer (80,
p. 44) has argued against the theory that the genital ducts of
Oligocheta are modified nephridia, because, whilst the lumen
of the latter is mainly intra-cellular, that of the former is
wholly intercellular; yet, I imagine, he would not deny that
the nephridia in the two groups are homogeneous; and they
appear to serve as genital ducts in some Polycheta.

The wide “body”? contains no convoluted tube, as is the
case in Nereis and other genera, but consists of one very
wide, simple tube, as is readily to be seen in sections through
the organ (Pl. XXV, figs. 35, 36, 37). The epithelium is one
cell deep, is ciliated throughout (figs. 40, 41), and for the
most part contains brown concretions (fig. 41), giving the
whole organ a yellowish colour. In the lumen of the gland,
in section, is a granular mass, frequently having the appear-

THE NEPHRIDIUM OF LUMBRICUS. 327

ance of a network ; this is probably the coagulated excretory
product.

The structure of the “ bladder,” or posterior terminal dilated
portion, is similar to that of the “ body,” but the concretions
are fewer, and as the external aperture is approached are
almost absent ; the walls of this region are, too, provided with
muscles.

The anteriorly directed funnel is very large, and consists of
neck and lips. The two lips are dorsally and ventrally
situated with regard to one another. The dorsal lip carries a
large number of delicate processes, sometimes simple, some-
times bifid or multifid, and even arborescent. The ventral
lip, on the other hand, is entire; the slight notches, &c., in
the drawing represent the folds resulting from compression.

The cells forming the edges of the two lips differ ; the dorsal
lip is lined by ciliated vesicular cells (fig. 40), similar to the
general lining epithelium of the body, but with numerous cilia,
and without concretions.

The cells lining the ventral lip, on the other hand, are
short, columnar, pigmented, and ciliated (fig. 39), whilst at the
extreme edge the cells become more cubical, large, and are
deprived of pigment granules (fig. 38).

I hope, in the near future, to examine chemically the pro-
ducts of excretion in Arenicola and other Polychetes.

I have (fig. 36) drawn one of many sections showing ova
in the neck of the nephridium ; however, I found none in the
body or in the “bladder,” and it may possibly be that the
funnel merely retains the ova for a period, and does not serve
as a genital duct. I think it probable that they really pass
to the exterior by the nephridium.

Cosmovici and Cunningham have given some details of the
vessels supplying the nephridium. The nephridial vessel is a
branch of the branchial vessel which is given off by the sub-
intestinal vessel, much in the same way as in Lumbricus
(Cosmovici, loc. cit., figs. 2—10).

The network on the nephridium lies between the excretory
epithelium within and the flat coelomic epithelium outside.

VOL. XXXII, PART III.—NEW SER. Y

328 W. BLAXLAND BENHAM.

The funnel itself is particularly well supplied with blood,
more especially the dorsal lip, which is traversed by one of the
main branches of the nephridial vessel, as shown in the figure ;
the other branch (a’) passes along the gonad, which is not
figured, and enters (at 6) the general network in the
nephridium.

A portion of this network is exhibited at fig. 34, which is an
enlarged view of the vessels on the neck of the funnel.

This network is present in many Polycheta, as Cosmovici
and others have pictured.

BIBLIOGRAPHY.

1. Bepparp.—‘ On the Specific Character and Structure of Certain New
Zealand Earthworms,” ‘ Proc. Zool. Soc.,’ 1888, p. 810.

la. Brpparp.—‘‘ On the Structure of a New Genus of Lumbricide,”
‘Proc. Zool. Soc.,’ 1887, p. 154.

2. Brpparp.—‘ Contributions to the Anatomy of Earthworms. On the
Structure of Eudrilus sylvicola,” ‘ Proc. Zool. Soc.,’ 1887, p. 372.

8. Bepparp.—“ Observations on the Structural Characters of Certain
New or Little-known Harthworms,” ‘Proc. Roy. Soc. Edinburgh,’
1887, p. 160.

4, Bepparp.—“On the Anatomy of Allurus tetraedrus,” ‘ Quart.
Journ. Mier. Sci.,’ xxviii, p. 365.

5. Brpparp.—‘‘ On the Occurrence of Numerous Nephridia in the same
Segment, &e.,”’ ‘Quart. Journ. Micr. Sci.,’ xxviii, p. 397.

6. Bepparp.—“ On Certain Points in the Structure of Urocheta, &.,”
‘Quart. Journ. Micr. Sci.,’ xxix, p. 235.

7. Bepparp.—‘ Development of Nephridia and Genital Ducts in Acantho-
drilus,” ‘Proc. Roy. Soc.,’ xlviii, 1890, p. 453.

8. BennamM.— Studies on Harthworms, I.” (Microcheta), ‘ Quart. Journ.
Micr. Sci.,’ xxvi, p. 267.

9. Bennam.—“ Studies on Earthworms, Il.” (Urobenus, Diacheta,
Trigaster), ‘Quart. Journ. Micr. Sci.,’ xxvii, p. 77.

10. Bennam.—“ Ona New Earthworm” (Brachydrilus), ‘ Zool. Anzeig.,’
1888, No. 271.

11. Bennam.—“‘ An Attempt to Classify Earthworms,” ‘Quart. Journ.
Micr. Sci.,’ xxxi.

12.

13.

14.

15.

16.

ET.

18.

19.

20.

21.

22.

23.

24.
25.

26.

27.

28.

29.

30.

THE NEPHRIDIUM OF LUMBRICUS. 329

Bercu.—* Zur Bildungsges. d. Exkretionsorganes bei Criodrilus,”
“Arb. Zool. Inst. Wirzburg,’ viii, p. 228.

Brreu.—* Neue Beit. z. Embryol. d. Anneliden. I. Zur Entwick. und
Different. d. Keimstreifen von Lumbricus,” ‘ Zeit. f. wiss. Zool.,’
], 1890.

Bourne, A. G.— Contributions to the Anatomy of Hirudinea,”
‘Quart. Journ. Mier. Sci.,’ xxiv.

Bourne, A. G.—Certain Points in the Anatomy of Polynoina,”
‘Trans. Linn. Soc.,’ 1883.

CLAPAREDE.—“ Histolog. Untersuch. d. Regenwiirmes,” ‘Zeit. f. wiss.
Zool.,’ xix, 1869, p. 217.

Cosmovict.—‘ Glandes Genitales et org. segmentaires des Annél. Poly-
chétes,” ‘Arch. d. Zool. Exp. et Gen.,’ viii, 1879.

CunnincHam.—*On some Points in the Anatomy of Polychetes,”’
‘Quart. Journ. Mier. Sci.,’ xxviii, p. 239.

GEGENBAUR.—“ Ueber d. Sogennan. Respirationsorgane des Regen-
wirmes,” ‘ Zeit. f. wiss. Zool.,’ iv, 1853, p. 221.

GorHLico.—* Lumbricus terrestris,” ‘Schneider’s Zool. Beit.,’ ii,
p. 133.

Horst.—“ Description of Earthworms, IJ,” ‘Notes from the Leyden
Museum,’ ix, 1887, p. 97 (Moniligaster and Rhinodrilus).

Horst.—* Sur quelques Lombrici exotiques, &.’’ (Eudrilus), ‘ Extr.
d. mémoires d. ]. Soc. Zool. d. France,’ iii, 1890, p. 228.

JacauEet.—“ Syst. Vascul. d. Annélides,” ‘ Mitth. Zool. St. Neapel,’ vi,
1885, p. 330.

KuxentHaL.— Lymphoidzellen d. Anneliden,” ‘Jen. Zeit.,? 1885.

Prerrier.—“ Sur un nouv. gen. de Lum. terr.” (Plutellus), ‘Arch. d.
Zool. Expér.,’ ii, 1873.

Perrier.— Etudes s. organisation d. Lum. terr.” (Urocheta),
‘Arch. d. Zool. Expér.,’ iii, 1874, p. 331.

Perrrer.— Etudes s. organ. d. Lum. terr. (Pontodrilus),” ‘Arch.
d. Zool. Expér.,’ ix, 1881, p. 175.

Rosa.— Sulla Strutt. d. Hormogaster redii,”’ ‘Mem. d. Reale
Accad. d. Sci. di Torino,’ xxxix, 1888.

Rosa.—* Lombrichi di Birmania,” ‘ Ann. d. Mus. Civ. d. Stor, Nat. d.
Genova,’ 2nd ser., t. ix, 1890.

Spencer, BALDWIN.—“On the Anatomy of Megascolides australis,”
‘Trans. Roy. Soc. Victoria,’ i, 1888.

330 W. BLAXLAND BENHAM.

31. p’Upexem.—“ Mém. s. les Lombriciens,” ‘Mém. d. |’Acad. Roy. de
Belge,’ xxxv, 1863.

32. VEspovsKy.—‘ System und Morph. d. Oligochaeten.’

38. Vuspovsky.—* Das larvale und definitiv Exkretionssystems,” ‘ Zool.
Anzeig.,’ 1887, No. 260.

EXPLANATION OF PLATES XXIII—XXV,

Illustrating Dr. W. Blaxland Benham’s paper on “ The
Nephridium of Lumbricus and its Blood-supply ; with
Remarks on the Nephridia in other Cheetopoda.”’

(Where not otherwise stated, the drawings refer to Lumbricus
herculeus, Sav.)

Fic. 4.—The nephrostome or internal funnel, seen front in front and repre-
seuted as slightly flattened, so as to show distinctly the various parts. Cilia
are omitted for the sake of clearness. The figure is fully named. The
central cell is left white. The vesicular connective-tissue cells (ves.) around
the preseptal portion of the tube are represented only by their nuclei.
n. cel. ep. is the flattened ccelomic epithelium. The two fine concentric
lines near the centre of the funnel represent the boundaries of the actual
aperture, which places the tube in communication with the ccelom, and which is
further shown by the arrow. The linea is the inner limit of the large “ central
cell,’ its outer limit being formed by the inner ends of the “‘ marginal cells.”
The line 4 is the thin outer free edge of the series of grooved centrifugal
cells, which are essentially drain-pipe cells, open along one side, or gutter-
cells. ‘These two lines require not only a high power (Zeiss, 3, F), but a
particularly good clear preparation for their detection.

Fic. 5.—A nephrostome drawn during life, to show the collection of leuco-
cytes or “‘débris’’ blocking the actual aperture of the funnel. These have
misled previous observers (see text). ‘The cilia on the marginal cells are to
some extent diagrammatically represented, as only a few rows are drawn; in
reality they cover the whole of the inner surface of the cells. The “ gutter-
cells” are hidden by the superposed centripetal marginals.

Fic. 6,—A funnel in profile, from a living specimen, showing leucocytes or
débris.”
Fic. 6a.—A few of these leucocytes, showing pseudopodia.

Fic. 7.—A longitudinal section through a funnel, from a series of sections
through a young worm [? sp.] (which were cut by my friend Mr. E. Goodrich).
The figure is fully explained.

THE NEPHRIDIUM OF LUMBRICUS. 331

Fic. 8.—A portion of the apex of the second loop, from a living nephri-
dium (Zeiss, 4, B), in order to show the second ciliated tract, c to ¢’ (see also
Fig. 1), in the outer narrow tube; together with the adjacent non-ciliated
portions. ves. Vesicular connective-tissue cells.

Fic. 8a.—A portion of another nephridium, to show the irregular character
of the lumen (? remnants of branchings) of the narrow tube.

Fic. 9.—A portion of Fig. 8 under a higher power (Zeiss, 4, E), showing
the character of the wall.

Fic. 10.—A portion of Fig. 9—a living nephridium—still further enlarged,
to show the termination (c) of the ciliated tract at the apex of the second
loop. This tube at ¢ is seen in optical cross-section owing to its sudden bend.

Fic, 11.—A small portion of the “middle tube” in optical section,
during life (Zeiss, 4, E). It shows the character of the protoplasm, the limits
of the cells, the excretory products in the protoplasm and in the lumen, and
the double row of cilia. The arrow points towards the ampulla (see Fig. 1).

Fic. 12.—The surface of a portion of the ampulla (drawn from a living
nephridium under Zeiss, B). It shows the shape and boundaries of the
constituent perforated cells.

Fic. 13.—An optical section of a small portion of the ampulla (drawn from
a living nephridium with Zeiss, HE). The differentiation of the protoplasm of
the cells into “central” and “ peripheral’ regions and the radial arrange-
ment of the granules are shown. ‘The circles in the lumen represent the
spherical excretory products.

Fic, 14.—An optical section of the “wide tube,” drawn from a living
nephridium (under Zeiss, E).

Fig. 15.—A transverse section across the third loop, near its apex, at
about the level of y in Fig. 1 (from a series stained in borax carmine; drawn
with Zeiss, 4, E). It is fully explained in the Plate. The middle tube shows
the characteristic spherular character of the protoplasm of its wall, and the
two rows of cilia; the granules in the lumen are the remains of excretory
products. 4. v. Blood-vessels. x. Nuclei of their endothelium. c. ¢. Vesi-
cular connective-tissue cells, the limits of the cells not being visible in
the sections. zc. Their nuclei. c. ep. Nuclei of cclomic epithelium. 2. A
curious fibrous appearance sometimes visible in sections.

Fic. 16.—A transverse section through the muscular duct (from the same
series as Fig. 15, under Zeiss, 3, F). ep. The lining of the duct. mus. e.
Muscle-cell. 2. mus. Nucleus of muscle-cell. ¢. ep. represents ceelomic
epithelium.

Fic. 17.—A portion of a transverse section through the body-wall of
Criodrilus lacuum, in the clitellar region, in order to show the absence of
a muscular duct in this genus, and the consequent perforation of the longi-
tudinal muscles by the “ wide tube.” y is a group of connective-tissue cells,

352 W. BLAXLAND BENHAM,

which hide the small intercellular part of the nephridium intervening between
the ‘wide tube” and the epidermic pit forming the nephridiopore. ci. Cli-
tellar cells. ep. Epidermis. cow. tis. Connective tissue. circ. Circular
muscles. /g. mus. Longitudinal muscles. xeph. pore. Epidermal pit, consti-
tuting the nephridial pore (which is deeper in the neighbouring sections),

Fic. 18.—A portion of the nephridial loop of Lumbricus in longitudinal
section, passing through some of the dilatations of the blood-vessels, in which
are groups of blood-corpuscles.

Fic. 19.—A transverse section through a part of the nephridium on one
half of the second loop, showing the blood-vessels ramifying between the
different parts of the tube.

Fic. 20.—A similar section through the tip of the third loop, to show
blood-vessels ramifying between the different parts of the tube.

Fic. 21.—The apex of the third loop, to show blood-vessels passing among
and around the various parts of the tube. It is slightly diagrammatic.
Those portions of the blood-vessels which pass behind the tubes are repre-
sented only in outline.

Fic. 22.—A portion of a nephridium, drawn from a living specimen under
a high power, to show the complexity of the coils and branchings of the capil-
laries; there were no dilatations in this specimen. ‘The outlines, merely, of
the nephridial tube are indicated.

Fic. 23.—Portion of a loop of a nephridium of another worm, in which the
dilatations of the blood-vessels were numerous (drawn under a low power from
a living specimen). Many of the finer vessels are not shown, nor are the
outlines of the nephridial tube. A portion of the septum is shown, with
small blood-vessels passing across from it to the nephridial network.

Fie. 24.—A portion of the same network greatly enlarged, to exhibit the
varying size and shape of the dilatations.

Fic. 25.—The nephrostome of Pericheta malamaniensis, n.sp. a.
In longitudinal section. &. Fromin front. These drawings are taken from the
same transverse section through the worm, and the two funnels are quite close
together, some five or six occurring in the whole section. The drawing is
introduced to show the simple character of the nephrostome when compared
with that of Lumbricus. ma. Marginal cells. ap. Communication between
nephridial lumen and the celom.

Fic. 26.—A portion of a section through a nephridium of Brachydrilus,
to exhibit the branching of the narrow tube which wraps round the
wide tube. The blood-vessels are also shown in black or in grey, according
to the relative depth in the section.

Fic. 27.—A nephridium of an Enchytreid, to demonstrate the simple
character of the nephrostome (s¢.), the uniform diameter, &c., of the tube
(¢.), which is embedded in one mass of vesicular cells (c.¢.). The external
aperture is shown at p.

THE NEPHRIDIUM OF LUMBRICUS. 333

Fic. 28.—The nephrostome of an Enchytreid (modified from Michaelsen)
in surface view, to show the nephrostome formed by the terminal per-
forated cell.

Fic. 29.—A series of nephridial funnels in optical section, diagrammatically
represented to exhibit the possible evolution of that of Lumbricus and
others (@) from that of an Enchytreid (@). The intermediate form (4) is met
with in Urochexta; that represented by c is, as far as I know, hypothetical.
Between a and 6 that of Per. malamaniensis may be inserted.

Fics. 30—32.—Diagrammatic views of constituent cells of a nephridium.

Fig. 80. A perforated (drain-pipe) cell, as met with in the ampulla.
Fig. 31. A drain-pipe cell from the middle tube.

Fig. 31a. The same in longitudinal section.

Fig. 32. A gutter-cell from the nephrostome of Lumbricus.

Arenicola piscatorum.

Fic. 33.—A complete nephridium. Surface view, under a low power.
The specimen was removed from the body, stained in borax carmine, and
mounted in Canada balsam. The natural size is shown by the line (z. s.)
underneath. The nephridium is seen from its ventral side, and the gonad,
which in life is closely connected with it, has not been drawn. The network
of blood-vessels, in black, is drawn as far as I could make it out; no doubt
it is more complicated and extensive than is shown. a is a branch of the
branchial vessel, passing to the nephridium. a’ is a branch to the gonad,
passing behind the nephridium. @ is the other end of this (cut) after passing
through gonad to join the network in the nephridium, mes, is a small piece
of mesentery, which suspends the funnel to the ventro-lateral muscle.

Fie. 34.—A portion of the network in the “neck” of the nephridium,
drawn under a higher power, and showing the cecal dilated terminations (di.).
The ccelomic epithelium (cel. ep.) is drawn in part of the figure.

Fie. 35.—A longitudinal section through the body of the nephridium, with
the ovary in situ. The blood-vessel marked 2 is intermediate between a’
and @ in Fig. 33.

Fic. 36.—A section through the neck of the funnel, with a couple of ova
within.

Fic. 37.—A section along the funnel and neck, with neighbouring parts of
the body-wall and ventro-lateral muscle (mzs.). The letter p points to the
relative position of the nephridiopore, which occurs several sections further
back. o. is an ovum.

Fic. 38.—A portion of the epithelium on the edge of the ventral lip of
the funnel (from Fig. 36, under high power). 2. Nucleus.

Fre. 89.—A portion of the epithelium lining the ventral lip (under high
power). 2. Nucleus.

Fic. 40.—Similar section through dorsal lip.

Fic. 41.—A portion of the lining epithelium of the body of the nephridium

304 W. BLAXLAND BENHAM.

(under high power). 4. v. Blood-vessels. cal. ep. Nucleus of flat ccelomic
epithelium. cone. Coloured concretions in the vesicular renal cells (epith.).
n. Nucleus of these cells. exc. pr. A granular coagulum, probably excretory
product in nephridium.

Fie. 42.—Diagram to illustrate the suggested evolution of a single nephri-
dial tube from an aggregation of a group of plectonephric tubules. a. A
portion of primitive network of a segment. &. One of the many funnels
occurring in each segment (as in Pericheta). c. One of the numerous
ducts passing to exterior. 4’. One of the funnels which persists and has
become increased in size by addition of marginal cells. c’. The ultimate
single duct. e. The tube (darkly shaded) of meganephric condition. fA A
few of the anastomoses (dotted) remaining, as in Brachydrilus, &c.

NOTES ON THE NAIDIFORM OLIGOCHATA. 335

Notes on the Naidiform Oligocheta; contain-
ing a Description of New Species of the
Genera Pristina and Pterostylarides, and
Remarks upon Cephalization and Gemma-
tion as Generic and Specific Characters in
the Group.

By
Alfred Gibbs Bourne, D.Sc.(Lond.), C.M.Z.S., F.L.S.,

Fellow of University College, London; Fellow of the Madras University ;
Professor of Biology in the Presidency College, Madras.

With Plates XXVI and XXVII.

THESE notes were commenced at the instigation of Professor
Lankester in 1882, to whom the late Mr. Thomas Bolton
had forwarded specimens of a Naid which Professor Lankester
identified as the Nais littoralis of O. F. Miiller, together
with another Annelid which I subsequently described as
Haplobranchus estuarinus.

Professor Lankester kindly placed in my hands all his
drawings and notes relating to Naids; and those on Nais
(Paranais) littoralis and Pterostylarides macro-
cheta, to each of which he had devoted considerable atten-
tion, are made use of in this paper.

To Mr. Bolton I was indebted for a very large number of
Naids collected in all parts of England, and I had intended to
prepare a monograph on the British species of this group,
but I left England with a series of scattered notes and
half-finished drawings. Vejdovsky, in his magnificent
‘System und Morphologie der Oligochaeten,’ brought to-
gether his own researches and a summary of our knowledge

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NOTES ON THE NAIDIFORM OLIGOCHATA. 307

of the group; and Mr. E. C. Bousfield! has still more recently
in a valuable paper given a systematic account of the various
species of the genus Dero at present known.

I am induced, even after this lapse of time, to publish some
of my uncompleted notes, partly because no one has since
described the new species which were discovered by Professor
Lankester and myself,? and partly because no writer on the
group has adequately dealt with the importance of the number
of cephalized segments as a generic character.

A monograph of the British species is still a desideratum.
I received from Mr. Bolton on several occasions specimens
other than those herein described, which were certainly not
referable to any existing species, but for which my notes are
insufficient to warrant the creation of new species. It has
frequently been pointed out, but very little stress has been
laid upon the fact even by Vejdovsky,? that the dorsal setz
are often wanting in several of the anterior segments of the
body, while ventral setz are present in these segments. It is
this character which chiefly marks what I term, at Professor
Lankester’s suggestion, a cephalization. There is almost
always, if not always, a certain amount of cephalization in the
Oligocheta; that is to say, there is in the anterior region
a segment or number of segments which differ in their
organization from the segments which follow, these latter
being usually similarly developed throughout the remainder of
the worm. This may be exhibited by peculiarities of the
alimentary canal, the circulatory system, the arrangement of
septa, the absence of nephridia from the most anterior
segments, and soon. In most if not all Oligocheta there is a
peristomial segment which is devoid of any sete, and in many
Naids the dorsal setz are absent from three, four, or six of the

1 «Journ. Linn. Soc. Zoology,’ vol. xx, 1887, p. 91.

* Professor Lankester referred to Pterostylarides macrocheta (under
the name of Pterygonais macrocheta) and exhibited my drawing of this
worm, which is here published, at the meeting of the British Association at
Southport in 1883.

8 *Oligochaeten,’ 1884.

338 ALFRED GIBBS BOURNE.

segments immediately following the peristomial segment ; and,
moreover, the number of segments thus modified (cephalized)
appears to be constant in all the species of a genus.! The
accompanying woodcut shows the various generic types.

The laws which govern gemmation or budding were worked
out by Semper. I have added the following remarks upon the
subject with the view of making this somewhat complicated
matter clearer to English readers, in the hope that some
microscopist may be induced to make further observations on
the subject in species where we still lack information as
to the budding individuals. There are many such, but there
are an even greater number of species in which the sexual
individuals have never been seen.

In the following diagram—
Cc
B ‘a
C

1 The only recorded exception to this which I have found is given by
Bousfield (1. c.), who states that in D. furcata there are only four segments
destitute of dorsal sete, while in all other species there are five such seg-
ments. I may, however, point out that D. furcata differs in another
marked character from all other known species of Dero in the possession of
palpi,” and should, I expect, be therefore placed in another genus.

NOTES ON THE NAIDIFORM OLIGOCHATA. 339

A is a primary zodid developed from the egg. A will give
rise to two secondary zodids, B and C, in the following manner :
—after its nth segment (m is a number characteristic probably
of each species) a bud, Z, will form; this will divide into two
regions, z and z': z will consist of an indefinite number of
segments, and form the tail of B; while z’ will consist of a
definite number of segments (a number characteristic of each
genus), and will become the head of C.

B and C may now separate, or may remain joined together
until two new budding regions have appeared.

In either case, after the nth segment of B another bud, Z,
may form, which will behave in the same way as the first bud
behaved, and divide into z and z’'; and z will become the tail of
a tertiary zodid B; and z’ the head of another tertiary zodid, c;
and a similar process will take place after the nth segment of
C, giving rise to two other tertiary zodids, c! and c*. The chain
of four zodids represented by the letters B, c, cl, c?, may now
separate into two chains of two zodids each, Bc and c'c*, or so
far as we know may remain joined together until, by the
formation of four new budding regions, a chain of eight
zooids, b, c, cl, c*, c3, c*, c®, c®, has developed. As a matter
of fact I believe that in no Naid do more than four zodids
hang together in a chain.

Among a mass of Naids where asexual reproduction was rife,
specimens might occur which could be represented thus :—A, B,
G, SC, 8;.c7c!).c*, Bc, cc? bye, 16",1¢7,,.c°,_C*,.6) eo. be, Ce.
cect, c'c®, bec!e”, c8ctc®c®, &c.

I am inclined to believe that the forms which do occur are
A, BC, sc, cc, Bcc!c”, becte’, c3ctc®c®, &c.; and among these
we may theoretically recognise three types:—(1) forms A,
developed from the egg; (2) forms BC, scc!c’, bec!c?, &ec., in
which the original head end is retained; and (3) forms c!c?,
c®ctc*c®, &c., in which the head is a budded region; and it is
possible that there are forms C, which must be distinguished
from c'c?, c®ctc’c®, &c., as they possess the tail end of a form
A, while c'c? and c®ctc°’c® must consist entirely of segments
formed by budding.

340 ALFRED GIBBS BOURNE.

I am not aware that any of these forms can be distinguished
from one another by observation of their structure, but it
would be most interesting to discover whether all or some of
them only can become sexual individuals.

I believe that the number of segments in the sexual indi-
vidual is constant for the species, while that in asexual indi-
viduals is indefinite; but that » is a number constant for the
species, and that the number of segments in z’ is constant for
the genus, » and z' being used with the signification above
described.

The segments represented by z’ are referred to in the
generic definitions as the cephalized segments: in most cases
all these cephalized segments remain marked in the adult; in
all cases the most anterior of them, the peristomial segment, is
obvious, while the others are usually marked by the non-deve-
lopment of dorsal seta bundles; but in Pristina, at any rate,
the bud shows that there are really seven cephalized segments
(i.e. z consists of seven segments), a fact which could not be
ascertained by an inspection of the well-grown head region.

The following table represents six zodids or chains of zodids
belonging to the same species, an imaginary species in which
I have supposed x= 10, and z’ to consist of three segments ; all
the other letters have the signification above described.
Instead of actually drawing the segments I have represented
them by some numeral.

341

NOTES ON THE NAIDIFORM OLIGOCHATA.

‘on )
TIAX
TAX
‘on | AX
SG ATX
CG Tix ¢ 0
13 11x
sere
i
8ST f py IG
11 | Hl
f [Z Be 7, 3 ff "Op |
ns AL
1, a8 | | I
| {2 81
{ 4 1G /VL
Z\ 0 | |
61 (OL
SI eA
AI ot + &
91
ert? 8
FL y
eI 9
ra G
IL P
1 g
J :
T
SI} SIU}

oyu MOIS TIM O OJUL MOIS TIM gf

LI
9L
cI
FL
elie
SL
i
m1
( [Z< It
| I
‘om ae
TIAX 74 TIAX
TAX | TAX
AX AX
AIX L a ATX “OM |
TIX TIX SI
1x | 11x rl
"Ow 1x L = ii
LT OT OL ta or |
9I 6 é :
ST 8 ty
FL L ie j2
SI 9 9 9 |
ZL G c c
IL P F | 7 |
ut ¢ g g
1 G G %
I T cL tJ
Oo pue 2 :s1y} o7UT MorS IM 7 * 880 OTH

moi pedopaasp
{enprarpar sMLL

—oyur oyzredos
feu ureyo siyy,

342 ALFRED GIBBS BOURNE.

SysTeM OF THE NAIDOMORPHA.

The Naidomorpha are Oligocheta in which the central
nervous system presents cerebral ganglia, pharyngeal com-
missures, and a ventral cord. The cerebral ganglia are always
separated from the epiblast.

In addition to sexual, asexual reproduction by means of
gemmation and subsequent fission occurs. A clitellum de-
velops in sexual individuals at the breeding season.

The setze are placed in four rows. They are capillary, spear-
shaped, or crotchet-shaped.*

There are, as a rule, more than two sete in each bundle.

A stomachal enlargement of the alimentary canal occurs in
segment vil or vill. They are all aquatic, living in fresh or
sea water. Branchial processes may be present or absent.

To this definition must be added, when information is
available, the position of the various generative organs. The
position of the testes and ovaries given by Vejdovsky does not
agree with my observations on N. barbata and P. littoralis.

According to Vejdovsky, the following genera form the
family of Naidomorpha :—

Aulophorus, Schmarda.
Dero, Oken.

Nais, s. str., O. F. Miller.
Bohemilla, Vejdovsky.
Ophidonais, Gervais.

1 I do not use these three terms to correspond exactly to Haar-borsten,
Spalt-borsten, and Haken-borsten. The capillary sete are the long hair-like
sete, which may be serrated as in Bohemilla; the crotchet-shaped sete
have the well-known /-shape, with the forked free extremity ; while what I
have called spear-shaped sete are such sete as present characters to some
extent intermediate between those of the foregoing types. They may be
straight with a sharp-pointed end, or straight and bifurcated at the end, or
somewhat crook-shaped.

NOTES ON THE NAIDIFORM OLIGOCHATA.

Slavina, Vejdovsky.
Stylaria, Ehrenberg.
Pristina, Ehrenberg.
Naidium, O. Schmidt.

345

I think it is desirable to add to this list Pterostylarides,
Czerniavsky, for P. parasita (syn. Stylaria parasita,
O. Schmidt) and P. macrocheta, sp. n., mihi; Paranais,
Czerniavsky, for P. littoralis (syn. N. littoralis, O. F.
and ? P. uncinata, and Chetobranchus, for
C. semperi, mihi.

Miller)

>
> PP

Saeco oe tae

og ee eS

GENERA AND Species or NAIDOMORPHA.

Aulophorus, Schmarda.

. discocephalus, Schmarda; Kingston, Jamaica.
. oxycephalus, Schmarda; Galle, Ceylon.
. vagus, Leidy; Philadelphia, N. America.

Dero, Oken (see woodcut).

. latissima, Bousfield.

. Perrieri, Bousfield.

. obtusa, D’Udekem.

. limosa, Leidy.

. philippinensis, Semper.
. acuta, Bousfield.

. furcata, Oken (syn. D. palpigera, Grebnitzky ;

Dy. SeieaecG. Semper).

Nais, s. str., O. F. Miiller (see woodcut).

Four cephalized segments in addition to the peristomial
segment remain well marked in the fully grown head.

There is no prostomial tentacle.

Some or all of the dorsal setz are capillary.

There are no branchial processes.

Eyes may be present or absent.

VOL, XXXII, PART II]. —NEW SER. Z

344. ALFRED GIBBS BOURNE.

1. N. elinguis, O. F. Miller.
This species is common in England.

2. N. barbata, O. F. Miiller.

This species is common in England. According to Semper!
the budding zone appears after segment xvii in this species
(i.e. n, as used on page 3389, is equal to 17).

3. N. josineg, Vejdovsky.

I have not seen this species.

4, N. fusca, Carter.

This species is recorded from India, but is not sufficiently
characterised.

5. N. scotica, Johnston; England.

Not sufficiently characterised.

Bohemilla, Vejdovsky (see woodcut).

Three cephalized segments in addition to the peristomial
segment remain well marked in the fully grown head.

There is no prostomial tentacle.

The capillary dorsal setz are serrated.

There are no branchial processes.

Eyes are present in the only known species.

1. B. comata, Vejdovsky.
(Syn. Nais hamata, Timm.)

I received numerous specimens of Bohemilla from Mr. Bol-
ton. It has not been, I believe, hitherto recorded from England.
The specific characters agree very closely with those given by
Vejdovsky and Timm. I never found a specimen without eyes ;
Vejdovsky makes no mention in his large book of their
presence or absence, but Timm states that the eyes are not
always present. I have noticed that the pigment in the
alimentary canal commences in the region of the first dorsal
seta bundle in this and many other species of Naids.

The nephridia are not present in all the segments. I
observed nephridia in segments VIII, XI, XVI, XVIII, xx, and
xx1, each with a funnel opening into the segment in front.

1 © Arb, zool. zoot. Institut. Wurzburg,’ Bd. iv, 1877.

NOTES ON THE NAIDIFORM OLIGOCHATA. 345

One of the most interesting features in the anatomy of this
worm is the condition of the sete in the cephalized segments.
The first pair of ventral seta bundles, viz. those belonging to
segment 11, are larger than those in the two following segments.
Those in segment tv are very small, and seldom contain more
than two setee—indeed, they are frequently altogether absent ;
but I have always found them in a newly budded head, so
that when they are absent it is doubtless because they have
dropped out. I have unfortunately no other notes with regard
to the budding.

Ophidonais, Gervais (see woodcut).

Four cephalized segments in addition to the peristomial seg-
ment are well marked in the fully grown head.

There is no prostomial tentacle.

There are no capillary sete.

There are no branchial processes.

Eyes are present in the only known species.

The only other known geuus of Naids in which capillary setz
are absent is Paranais.

1. O. serpentina.
This species is well known in England; all the dorsal setz
are spear-shaped and straight.

Slavina, Vejdovsky (see woodcut).
Four cephalic segments in addition to the peristomial segment
are well marked in the fully grown head.
There is no prostomial tentacle.
The dorsal setze are capillary.
There is a girdle of papille in the middle of each segment.

1. S. appendiculata, Vejdovsky.

I received a few specimens of a species of Slavina from
Mr. Bolton, but have no notes which enable me to offer an
opinion as to the identity or otherwise of the English species
with S. appendiculata, Vejdovsky.

According to Bousfield' the English species is identical with
the Nais lurida of Timm, and he calls it S. lurida, and

1 «Journ. Linn. Soc. Zoology,’ vol. xix, 1886, p. 264.

346 ALFRED GIBBS BOURNE.

maintains that this species is not identical with S. appendi-
culata, as Vejdovsky believes it to be. As Stolc’ has pointed
out, this is very probable, and the difference is to be found in
the arrangement of the tactile papille ; but Lagree with Stole,
that Bousfield has fallen into a great error in associating
Ophidonais serpentina with the genus Slavina. The
absence of capillary sete distinctly marks Ophidonais as a
separate genus.

Stylaria, Lamarck (see woodcut).

Four cephalized segments in addition to the peristomial
segment are present in the bud, and remain well marked in
the fully grown head.

The prostomial tentacle is very long.

The dorsal setz are capillary.

There are no branchial processes.

Eyes are present in the only known species.

1. S. lacustris.

A full synonymy of this species, which is perhaps most
widely known as Nais proboscidea, is given by Vejdovsky.

I have occasionally observed specimens in which dorsal
setze were present in the one or two most posteriorly placed of
the usually cephalized segments. I can only suggest that these
were abnormal individuals, but it isa matter worthy of further
investigation. I described the process of budding in this
worm at the meeting of the British Association at Aberdeen,
in 1885. I quote here the substance of the note published in
the report of that meeting. I have altered the numbering of
the segments to make it accord with the system adopted in the
present memoir, in which the peristomial segment is called
segment I.

«* When budding is about to commence, a slight thickening
of one of the septa which separate one ccelomic segment from
another occurs. This thickening increases, the body-wall in
the region thickens, and an actual bud is here formed. This

1 * Zoologischer Anzeiger,’ vol, ix, 1886, p. 502.

NOTES ON THE NAIDIFORM OLIGOCHATA. 347

new region elongates, and presents a solid appearance. The
alimentary canal grows in this region, but the newly formed
portion is at first unpigmented, and may still be detected at a
much later period by its lighter colour. Its lumen remains,
however, all the time, and a continuous line of fecal matter
may be observed (Pl. XXVII, fig. 9, fec.). This budding region
divides into two portions. The anterior portion develops
numerous sete, and gives rise to an indefinite number of
segments which form the tail of the old worm. The posterior
portion develops four pairs of ventral sete, this development
taking place from before backwards; and subsequently at its
anterior region the peristomial segment and the characteristic
proboscis are developed, and the two individuals separate. The
budding region usually forms between segments xxvir and
XXxviII, so that segment xxvii becomes segment vi of the
posterior daughter worm : the five anterior segments of this
worm never present dorsal seta.”

What is described above as taking place is clearly shown in
Pl. XXVII, figs. 7—9, which represent various stages of the
process.

The division of the budding zone into a new tail on the one
hand and a new head on the other was clearly stated by
Semper, whose paper on the subject I had not seen when
writing the above.

It will be seen that, according to the nomenclature I have
adopted, = 26, and z’ consists of five segments in this
species.

Pterostylarides, Czerniavsky (see woodcut).

Four cephalized segments in addition to the peristomial
segment are indicated in the fully grown worm. The two
hinder of these are devoid of ventral as well as of dorsal setz.

The prostomial tentacle is of medium length.

The dorsal sete are capillary.

There are no branchial processes.

Hyes are present.

Vejdovsky associates this genus with Stylaria, but I think

' © Arb. zool, zoot. Instit. Wurzburg,’ Bd. iv, 1877.

348 ALFRED GIBBS BOURNE.

that the peculiar character of the cephalization is a feature
amply sufficient to warrant the separate genus.

Pterostylarides differs from all the other genera in the
constant possession of cephalic segments other than the peri-
stomial which are devoid of all sete. Itis, of course, an assump-
tion that the region between the second and third seta-
bearing segments does represent two segments. Vejdovsky
in his “ Oligocheta” assumes that this is the case; but I
have not seen his paper on ‘‘ Thierische Organismen der Zum-
menwasser von Prag, &c.” In Pl. XXVI, fig. 1, which was
drawn from nature, there is but the very slightest external
indication of segments in this region, and I made no observa-
tions upon the septa.

As stated above, in Bohemilla comata the sete in the
most posterior cephalized segment are few in number and very
small, and may disappear; when this occurs we have an undeni-
able instance of a cephalized segment devoid of all sete. Such
absence of all sete is, I think, to be regarded as extreme
cephalization.!

In the following description of P. parasita IL have selected
from Vejdovsky’s description what seem the important points
in distinguishing this species from P. macrocheta.

1. P. parasita, O. Schmidt.

Prostomial tentacle of about the same length as the peri-
stomial segment (“ Mundsegment ”’).

The eyes are on the dorsal side.

The ventral setz of the second and third segments are curved
and crotchet-shaped, and about a third longer, and with longer
teeth than the ventral sete of the other segments. There
are five or six sete in these bundles, and seven or eight in
those of the other segments.

The sete in the three most anterior dorsal bundles are

1 There can be no doubt that Chetogaster, which in my opinion ought to
be classed with the Naidomorpha, presents a region corresponding to two or
even three imperfectly developed segments devoid of sete between the first
and second pairs of bristle bundles.—E, R. L.

NOTES ON THE NAIDIFORM OLIGOCHATA. 349

capillary, and longer than the first five segments. There are
twelve to fifteen in the bundle. The other dorsal sete are capil-
lary, and of about one fifth the length of the anterior ones.

2. P. macrocheta, mihi (Pl. XXVI, fig. 1).

The prostomial tentacle is much longer than the peristomial
segment.

Eyes as in P. parasita.

The ventral seta bundles of the second and third ‘segments
consist of two to three sete, and those of the other segments
of two to five sete.

The sete in the three most anterior dorsal bundles are all
capillary; two to five of these are very long, longer even than the
long setz in P. parasita, while the other four to six are shorter
than the other dorsal sete. The remaining dorsal seta bundles
usually contain two setz, one rather longer than the other.

I found no individuals with generative organs, and none
exhibiting gemmation.

[The long setz are frequently found thrown forward so as to
partly encase and protect the head when the worm forms for
itself a temporary tube. They are also used to strike the water
in swimming.—E. R. L.]

Paranais, Czerniavsky (see woodcut).

Three cephalized segments in addition to the peristomial
segment remain well marked in the fully grown head,

There is no prostomial tentacle.

There are no capillary sete; all the sete (dorsal as well as
ventral) are crotchet-shaped.

There are no branchial processes.

Eyes may be present or absent.

1. P. uncinata, Oersted.
According to Czerniavsky the Ophidonais uncinata of
Oersted belongs to the genus Paranais.
2. P. littoralis, O. F. Miller (Pls. XXVI and XXVII,
figs. 2 to 6).
This species is by no means well known. It is recorded by

350 ALFRED GIBBS BOURNE.

Oersted, and more recently mentioned by Czerniavsky, who
founded the genus Paranais for it and the above-mentioned
species. Czerniavsky found it in Abchasia.

Professor Lankester received specimens of this worm from
the great mud-banks at Sheerness, at first through Mr. Bolton;
but we afterwards obtained it in quantity from Mr. W. H.
Shrubsole, who found it living in enormous numbers along
with Haplobranchus and Hemitubifex. It is one of the
few Oligocheta which are known to inhabit salt water.

I am able to give only such notes of its structure as were
made at the time by Professor Lankester and myself.

The prostomium is blunt and rounded.

The sete present an arrangement the main features of which
are doubtless characteristic of the genus, and are given above
in the generic definition.

Sete are absent, as is universally the case, from the peristo-
mial segment (segment 1).

In segments 11, 111, and tv, ventral seta bundles alone are
present, while in all other segments dorsal and ventral seta
bundles are present. The usual number of setz to the bundles
is as follows:

Dorsal. Ventral.
Segment 1 : . _—. ; : 5 a
9 iI = 5
Ay ll — 3
» ‘IV am 3
sp tw WN ; : ainmiite é ‘ : 3
we VI F : peo ere . : : 3

and soon. In the dorsal bundles there are occasionally two
setze only ; and in the budding region there are, as a rule, in
the early stages of the bud two sete only in each bundle, one
longer than the other.

Theordinary sete vary but little in shape throughout the body.

In no other genus of Naids have all the dorsal sete the
crotchet shape, as is here the case (Pl. XX VII, fig. 6).

In Ophidonais none of the dorsal sete are capillary, but are
spear-shaped setz, and unlike the ventral sete of that genus.

In P. littoralis the sete of the cephalized segments are

NOTES ON THE NAIDIFORM OLIGOCHATA. 351

longer and thinner than those which follow, and the ventral
setee of the majority of the segments are a little thicker and
shorter than the dorsal sete (Pl. XX VII, fig. 5).

Modified genital setz are present in sexual individuals, They
are the ventral sete of segment v. There is no evidence that
they represent any interpolated segment. The ordinary sete
doubtless drop out from the ventral bundles in that segment
during the breeding season, and are replaced by the modified
genital sete. There are usually three inthe bundle. They are
very stout, and longer than the ordinary sete, and they possess
a mere rudiment of the crotchet at the free extremity.

In the budding individual the arrangement of the setz in a
well-advanced bud is shown in Pl. XXVI, fig. 2. In the new tail
the series of seta bundles are seen to fade away gradually as one
passes backwards—that is to say, the most posterior bundle is
the youngest. In the new head there are three pairs of ventral
seta bundles, only the new head assumes from the first its
adult character (compare this with Pristina).

The pharynx occupies about three segments, and at its sides
and posterior to it occur two large glandular masses (Pls.
XXVI, XXVII, figs. 2 and 3, ph. gil.), which are‘ possibly
groups of unicellular salivary glands. The “stomach” is well
marked, and lies in segment viit; the intestine is narrow in the
following segment, and then widens out.

The dorsal and ventral vessels are clearly defined through-
out the whole length of the worm. They communicate with
one another by three pairs of branching lateral vessels in seg-
ments 11 to iv, and by three pairs of larger unbranched
‘‘ hearts ” in segments v to VII.

It is exceedingly interesting to note état in the newly
budded regions and other regions of active growth, as in all
tail regions, the dorsal and ventral vessels are joined by com-
missural vessels in every segment, and that this is therefore
doubtless the primitive arrangement ; and the suppression of
the commissural vessels in all except certain anterior segments
indicates a particular kind of cephalization (Pl. XXVI, fig. 2).

I was unable, even after repeated examination, to discover

352 ALFRED GIBBS BOURNE.

any nephridia ; and their absence, if they are really absent, is
a very remarkable character, which I should be glad to see
verified.

I found numerous individuals in which the generative organs
were well developed (Pl. XX VII, fig. 3).

The testes occur in segments viii and 1x, and the ovaries in
segment x.

The spermathece lie in segment v, and open near the modi-
fied genital setz, between segments v and v1.

I was able to make a few observations on the asexual repro-
duction.

The most complicated chain of zodids which I obtained ex-
hibited two regions of active growth. Such a chain would be
represented, according to the nomenclature which I adopt,
thus :—BC, or sc, or clc®, &c. According to the same nomen-
clature, »=17 in P. littoralis, and z consists of four
segments.

Pristina, Ehrenberg (see woodcut).

Seven cephalized segments—of which only one, the peristomial
segment, is recognisable in the fully grown head region—may
be distinguished in the newly budded head region.

The prostomial tentacle is short.

The dorsal sete are capillary or capillary and spear-shaped.

Branchial processes are absent.

Eyes are absent in all the known species.

1. P. longiseta, Ehrenberg.

This species has been recently re-described by Vejdovsky. I
have not seen it, nor has it, so far as I know, ever been recorded
from England.

2. P. equiseta, sp. n.

I found this species in great quantity inthe Victoria regia
tank in the gardens of the Royal Botanic Society at Regent’s
Park, London.

It differs from P. longiseta only in the non-development of
the long sete to which that species owes its name.

NOTES ON THE NAIDIFORM OLIGOCHATA. 353

It is very small, about 8 mm. long.

The majority of my specimens in which there were no genera-
tive organs, and which were not budding, consisted of eighteen
to twenty-one segments.

I frequently found the ventral sete of the fourth segment in
non-budding individuals to be much larger and stouter than
the other ventral sete. These were probably modified genital
sete, but I never obtained specimens showing any further
sexual developments. The dorsal sete are all capillary and of
similar lengths throughout the body. There are usually two in
the bundle, one longer than the other; but those of the third
segment are not extra long, as is the case in P. longiseta.

The ‘‘ stomach ” lies in segment VIII.

The blood is yellowish.

I observed a single pair only, of commissural vessels.

The most anterior nephridia lie in segment 1x, and not in
segment x, as in P. longiseta.

The coelomic corpuscles are very large, and greenish in colour.

I observed numerous budding individuals, but in all cases
there was only a single budding region in any particular speci-
men, from which I infer (though not with certainty) that as
soon as a chain of two zodids is formed a separation takes
place.

The budding takes place after segment x11I (i.e. 2 = 18),
and there are seven segments in the new head (i.e. z’ consists
of seven segments). The most anterior of these becomes the
peristomial segment, and the remaining six develop dorsal and
ventral sete; so that in the adult they are indistinguishable, so
far as the setz are concerned, from those which follow.

3. P. breviseta, sp. n. (Pl. XXVII, figs. 11—15).

This is a species which I found in enormous numbers in one
or two localities in Madras; and although I hope to describe it
and other Indian Naids in detail at some future date, I refer
to it now because, from observations made upon it, I have been
enabled to verify the law of budding described in the foregoing
portion of these notes.

354 ALFRED GIBBS BOURNE.

P. breviseta grows to a larger size than the other species
of Pristina, and contains a greater number of segments,
and differs from them besides in two or three well-marked
characters.
The dorsal setz are of two kinds (Pl. XX VII, figs. 14. and 15),
capillary setee and spear-shaped sete: the latter are peculiarly
shaped, and are bifurcated at the distal extremity. (Compare
with these Vejdovsky’s figures of the dorsal sete in Naidium
luteum, and those figured by myself for Chetobranchus).
The capillary sete are of about the same length throughout
the body, except that in the second and third segments they
are shorter (this is the character referred to by the name of
the species).
Those of segment 11 are about half, and those of segment 111
about three quarters as long as those that follow.
The coelomic corpuscles are black and very noticeable.
The most anterior nephridia are in segment Ix.
I have not hitherto found sexual individuals. The budding
takes place, as arule, after segment xxii (i.e. m= 22), and
the newly budded head consists, as in P. equiseta, of seven
segments (i.e. z’ consists of seven segments in the genus Pris-
tina). (See Pl. XXVII, fig. 12.) It has been largely the
comparison of these two species of Pristina which has led
me to believe that, as stated on p. 338, mis constant (with a
few individual variations) for the species, while the number
of segments in the new budded head is constant in the genus.
I give below a few examples, selected from the very large
number of specimens in which I have counted the segments.
1. More than thirty segments, no budding.
2. More than forty segments, no budding.
3. More than forty-six segments, no budding.
4, More than fifty segments, with a budding zone after seg-
ment xx.

5. More than fifty-five segments, with a budding zone after
segment XXII.

6. More than fifty-six segments, with a budding zone after
segment XXII.

NOTES ON THE NAIDIFORM OLIGOCHATA. 355

7. More than seventy-one segments, with a budding zone
after segment XxII.

8. More than seventy-eight segments, with a budding zone
after segment xx11, and another budding zone after
the eighteenth segment beyond the new head of the
anterior budding zone.

I say “more than” so many segments in all these cases
because it is impossible to count the actual number of segments
in the tail region.

P. inequalis, Ehrenberg, is insufficiently characterised, and
P. flagellum, Leidy, does not belong, according to Vejdov-
sky, to the genus Pristina, but comes somewhere between
Dero and Aulophorus.

Naidium, O. Schmidt (see woodcut).

One cephalized segment only recognisable in the fully grown
worm, and no information as to the bud.

There is no prostomial tentacle.

The dorsal sete are capillary and spear-shaped.

There are no branchial processes.

HKyes absent in the only known species.

1. N. luteum, O. Schmidt.

Chetobranchus, mihi (see woodcut).

One cephalized segment only, recognisable in the fully grown
worm, and insufficient information as to the bud.

There is no prostomial tentacle.

The dorsal setz are capillary and spear-shaped. Branchial
processes enclosing some or all of the setz of the dorsal bundles
present in all the anterior segments, with the exception of the
peristomial segment.

In the shape and arrangement of the sete this genus very
closely approaches Naidium, from which genus it differs in
the remarkable branchial processes.

1. C. semperi, mihi.!

' «Quart. Journ. Mier. Sci.,’ vol. xxxi, 1890.

356 ALFRED GIBBS BOURNE.

DESCRIPTION OF PLATES XXVI & XXVII,

Illustrating Professor Alfred Gibbs Bourne’s ‘“ Notes on the
Naidiform Oligocheta,” &c.

PLATE XXVI.

Fic. 1.—Surface view of Pterostylarides macrocheta. The seg-
ments are numbered 1—28. m. Mouth.

Fic. 2.—View of Paranais littoralis. The Arabic numerals indicate
segments of the parent worm; the Roman numerals are attached to segments
in the budding zone. m. Mouth. ph. Pharynx. pd. gl. Pharyngeal glands.
st. Stomach. d. v. Dorsal vessel. v.. Ventral vessel. zé¢. Intestine.
a. Nerve-cord.

PLATE XXVII.

Fic. 3.—Anterior extremity of a sexual individual of Paranais lit-
toralis. ph. gl. Pharyngeal glands. gem. se¢. Genital seta bundles. sp.
Spermatheca. ¢. Testis. ov. Ovary.

Fic. 4.—Ventral seta from segment 1, 111, or Iv, of Paranais littoralis.

Fie. 5.—Modified genital seta of Paranais littoralis.

Fic. 6.—Dorsal seta from Paranais littoralis.

Fies. 7, 8, and 9.—Views of the budding zone in Stylaria lacustris in
successive stages of growth. 72. Budding zone. <z. Newly budded tail.
z'. Newly budded head. pr. New prostomial tentacle. The Arabic numerals
indicate segments of the parent worm; the Roman numerals indicate newly
budded segments. Fac. Feces, showing how they pass through the newly
budded region.

Fre. 10.--Enlarged view of the seta bundles of one side of the new head
from the same specimen as Fig. 9.

?

Fic. 11.—Diagram of the anterior extremity of Pristina breviseta.

Fic. 12.—Diagram of the budding zone of Pristina breviseta. Letters
and numerals as in Fig. 9. The animal is drawn from the side to show the
dorsal and ventral seta bundles.

Fic. 13.—Ventral seta of Pristina breviseta.

Fics. 14 and 15.—Capillary and spear-shaped dorsal sete of Pristina
breviseta.

ON PELOMYXA VIRIDIS. 357

On Pelomyxa viridis, sp. n., and on the Vesi-
cular Nature of Protoplasm.

By

Alfred Gibbs Bourne, D.Sc.(Lond.,), C.M.Z.S., F.L.S.,

Fellow of University College, London; Fellow of Madras University ;
Professor of Biology in the Presidency College, Madras.

With Plate XXVIII.

My assistant, M.R.Ry. A. Sambasivan, B.A. (Madras),
during the course of an exhaustive examination of the fauna of a
small tank (pond) in the neighbourhood of the Presidency
College discovered and drew my attention to numerous green
“‘egg-like ” bodies, of about ;4, inch in diameter, which were
to be seen on the surface of the finely divided mud.

These bodies proved upon examination to be Rhizopods be-
longing to the remarkable genus Pelomyxa, and forming a
new species of that genus, but differing in some important
particulars from all hitherto described Rhizopods. I propose
to call the species P. viridis.

The species is peculiarly interesting not only on account of
its great size (it is larger than any known form of the Lobosa),
but also on account of the presence of chlorophyll and symbiotic
bacteria,’ and the assistance which this renders in the study of
its protoplasm.

The rod-like bodies here regarded as bacteria were described as crystals
of unknown composition by Greeff in his account of Pelomyxa palustris.
Leidy describes them as exhibiting transverse striations. The symbiosis of
bacteria and ameboid Protozoa has a special importance at the present
moment in connection with Metschnikoff’s doctrine of Phagocytosis.

358 ALFRED GIBBS BOURNE.

Habitat and General Description.

I have only found P. viridis in one tank, and this a tank
which has not been known to dry up for many years.

When the mud from this tank is placed in a dish with a
little water and allowed to stand, several individuals of P.
viridis may generally be found close to the surface of the
mud ; if these are removed and the mud stirred up and again
allowed to stand, it is not usual to find many other specimens.
So that I believe that this Rhizopod lives close to the surface of
the mud. I have never seen it crawling on the sides of the
dish or on any other object ; nor have I ever seen it much
spread out while in the mud, but when mounted on a slide with a
cover-glass it readily spreads itself out and becomes very flat,
and soon commences to exhibit active amoeboid movements.
Fig. 1 represents a mounted specimen, partially spread out
and magnified about four diameters. It is difficult to convey
any idea of the size to which such an organism may attain, as
so much depends upon the extent to which it is spread out. I
have seen specimens so much spread out that they would
average as much as + inch in diameter. The specimens are of
very varying sizes, but I have never found very small speci-
mens ; I have never, in fact, been able to find a specimen by the
aid of the microscope which I had not previously picked out
with the naked eye, although I have searched many slides of
the mud for this purpose. It may be that small individuals
are to be found at some other time of year, or it may be that
I have not, in spite of my search, happened to alight upon a
small specimen.

When viewed by reflected light individuals vary a good
deal in colour, from the rich transparent green of a gelatinous
lichen to that of a faded leaf. I presume that the chlorophyll
undergoes some degeneration. I have never found any
approach to the red colour found in Euglena or Hemato-
coccus. There is often a whitish, more opaque-looking spot,
such as has been shown in the middle of fig. 1. This is caused
by au aggregation of sand particles, and it is often more clearly
defined than in my figure. When seen by transmitted light

ON PELOMYXA VIRIDIS. 359

with a low power such regions uvaturally appear dark, but
with polarised light they appear as bright spots, owing to the
doubly refracting character of the sand particles. A number
of these particles is always present, but at times their number
is so enormous that there is almost as much sand as proto-
plasm. I have watched an individual so filled collect the
greater number of these particles at one spot, and then simply
pour them out from the protoplasm. An individual may thus
get rid of hundreds of these particles in a minute or two, but
the process of collecting them together preparatory to their
extrusion is slow. I believe that there is a periodical whole-
sale extrusion of these particles. These particles have all the
appearance of sand ; they are of very varying size and of very
irregular shape, and are insoluble in all ordinary reagents ;
they are crystalline in nature, and, as stated above, are doubly
refracting.

The presence of these particles has not, I fancy, any special
significance, they occur in great numbers in the mud and
are taken in with the food ; the animal, indeed, seems to exercise
no discretion as to what it takes in. They become collected
together as a mechanical result of slight movements, and when
the animal makes some more extensive movements are thrown
out in great numbers.

Structure of the Protoplasm.

Viewed with an ordinary high power of the microscope the
main mass of the organism appears of a green colour, while at
the periphery, from time to time, perfectly colourless regions
may be seen. One’s first impression is that there is a green
endoplasm and a colourless exoplasm (figs. 2 and 3). With
the same magnifying power, in addition to the various proto-
plasmic contents described elsewhere, what appear to be
granules may be recognised abundantly distributed through
the greater mass of the protoplasm. The colourless peripheral
regions above mentioned are usually devoid of these granules.
The granules all prove upon further examination to be bacteria,
and but for their presence the protoplasm itself would appear

VOL. XXXII, PART III.—NEW SER. AA

360 ALFRED GIBBS BOURNE.

absolutely non-granular. Exceedingly careful and repeated
observations with a Powell and Lealand’s ;4 inch oil immersion
objective, Abbé’s sub-stage condenser, and Engelmann’s dark
chamber, which last is invaluable in the glare usually pre-
valent here, have enabled me to ascertain with great accuracy
and clearness the real structure of the body substance.

The protoplasm is throughout a perfectly colourless and
apparently homogeneous substance, and this substance, except
in the peripherally placed portions which flow out from time to
time to be sooner or later reabsorbed into the central mass, is
densely packed with spherules of a semi-fluid stroma, im-
pregnated with what I have ascertained to be chlorophyll, and
to these chlorophyll-bearing spherules the green colour of the
organism is due.

In these spherules we have not to do with chlorophyll-
corpuscles, as they exhibit no phenomena of division, and are,
moreover, more fluid than chlorophyll-corpuscles.

Their fluid character often becomes very evident after the
death of the organism, when some of them may be usually
observed to run together into larger masses. Where these
spherules are packed closely together they assume the form of
regular polyhedrons. They seem about as fluid as the stroma
of human red blood-corpuscles. I have come to the con-
clusion that they correspond to the vacuoles or, as they are
better termed, to distinguish them from other vacuoles,
vesicles, which occur in so many specimens of protoplasm
and give to such specimens a vesicular character.

The fact that they contain in P. viridis a substance im-
pregnated with chlorophyll, a state of things hitherto un-
observed, has enabled me to throw some new light upon their
nature and upon the organization of the body substance.

The word protoplasm is used above in the sense in which
Biitschli’ uses the word plasma. It designates the substance
which Leydig speaks of as the spongioplasma, as distinguished
from hyaloplasma (chylema, Strasburger). In other words,
I consider that the substance within the vesicles is to be

‘ Bronn, ‘ Protozoa,’ p. 1892.

ON PELOMYXA VIRIDIS. 361

regarded as an entoplastic product of the protoplasmic activity
rather than as any portion of the protoplasm itself. The
chlorophyllogenous stroma occupying the vesicles of P.
viridis is no more entitled to be regarded as a portion of the
protoplasm than are chlorophyll bodies or oil-globules, or any
other structures ordinarily spoken of as protoplasmic contents.
It is, of course, the presence of the chlorophyll which leads me
to this conclusion. If I am correct in regarding these chloro-
phyllogenous structures as the homologues of the vesicles
found in other protoplasms, the condition of P. viridis lends
strong support to Biitschli’s theory that the plasma is the sub-
stance which forms the envelopes of these vesicles only, or,
as Bitschli has termed it, the substance forming the scaffold-
ing of a honeycomb (Substanz des Wabengerusts), and does not
include their contents (Wabeninhalt).

The structure of P. viridis is, on the other hand, rather
opposed to Bitschli’s most recent views with regard to proto-
plasmic movements. If I understand rightly from the abstract
available to me, Biitschli believes that a movement starts by the
bursting of some vesicle, a phenomenon which would naturally
escape observation in ordinary protoplasm, but of which, if
it is constantly occurring, I should surely have seen something
in a protoplasm whose vesicular contents are brightly coloured.

With regard to the homology which I have assumed to exist
between the vesicles of ordinary protoplasm and those of
P. viridis with their chlorophyllogenous contents, it is quite
clear, in the first place, that these are the structures which
render the protoplasm of P. viridis vesicular. There exist,
itis true, in addition to them a number of vacuoles of varying
sizes, but of these there are after all comparatively few, and if
they alone were present the protoplasm could not be called
vesicular. There is no evidence that these vacuoles contain
any substance other than water.

We can undoubtedly distinguish in P. viridis between two
varieties of non-contractile fluid-containing spaces (nicht con-
tractile Flussig-Keitsriume), the vacuoles containing water
and the vesicles having chlorophyllogenous contents.

362 ALFRED GIBBS BOURNE.

The vesicles vary very little in size, being always about
sooo Inch in diameter.

The vacuoles, on the other hand, vary greatly in size, some
being almost as small as the vesicles, while others attain five
or six times that diameter.

No distinction has, as far as I am aware, been hitherto drawn
between vacuole and vesicle in any so-called “vesicular” or
** vacuolised ” protoplasm.

Bitschli,* writing ten years ago, says of non-contractile
vacuoles, “ Seltener hingegen wird ihre Zahl so betrachtlich,
dass das sie trennende Plasma uur noch ein Maschenwerk von
Scheidewinden zwischen ihnen herstellt, das Plasma eine
schaumige oder alveolire Beschaffenheit annimmt. LEin
derartiges Verhalten begegnet uns z. b. gewdhnlich bei
Pelomyxa, auch bei gewissen Amében tritt ahnliches mehr
oder weniger deutlich hervor.”

Of Biitschli’s most recent views I can only judge from M.
Yves Delage’s’ note on the models of protoplasm with which
Biitschli has been recently experimenting, and if I understand
rightly in, for instance, the soap and xylol model, the plasma
(Biitschli) is represented by the soap and the chylema (Stras-
burger) by the xylol. So that the contents of both vacuole
and vesicle would be termed chylema, but there is no word
as to the possibility of the chylema being other than a single
substance.

Now, in P. viridis, two distinct substances take the place
of the xylol of Biitschli’s model, the contents of the vacuoles
and those of the vesicles, and were it not for the presence of
the chlorophyll in one of them these would not be optically
distinguishable one from the other.

Two species of Pelomyxa have been hitherto described,
P. palustris, Greef,> and P. villosa(Ameba villosa),
Leidy.* The protoplasm of both these species has been de-

' Bronn, ‘ Protozoa,’ p. 101.

* «Arch. Zool. Experimentale,’ 1889.

% Arch. f. mikr. Anatomie,’ Bd. x, 1874.

* *Fresh-water Rhizopods of North America,’ Washington, 1879.

ON PELOMYXA VIRIDIS. 363

scribed as highly vesicular, and Gruber,’ in speaking of
P. villosa, which he found in Germany, says, ‘‘ Zunachst
fallen die zahlreichen Fliissigkeitsvacuolen ins Auge, welcher
den gréssten Bestandtheil des Korpers ausmachen und dem-
selben ein Schaumiges Aussehen verleihen. Diese Vacuolen,
die von wechselnden Umfange Sind, liegen eingebettet in
dem homogenen Plasma, welches mehr oder weniger feine
Scheidewander zwischen ihnen bildet, ahnlich eine Intercellu-
larsubstanz zwischen den einzelnen Zellen eines Gewebes.”

I think it probable that in these forms we have both vacuoles
and vesicles, but as the vesicles are filled with colourless
substance no distinction has been drawn between them. It
will be interesting to re-examine these forms to ascertain if
from the relative size and frequency any distinction can be
drawn. ‘The importance of the question lies in this, that the
vesicles have probably an intimate relation to the structure of
the protoplasm, and possibly to the production of amoeboid
movements ; while the vacuoles vary from time to time in size
and number, and have nothing to do with the ultimate structure
of the protoplasm.

Fig. 6 represents a fragment of the protoplasm in a specimen
which was deeply stained with osmic acid. At 2 is seen pro-
jecting at the surface a small portion of the hyaline protoplasm,
devoid of all contents, stained brown and rendered granular by
the osmic acid. The rest of the drawing shows the vesicles
once occupied by the chlorophyllogenous substance bounded
by the protoplasm. The chlorophyll has been dissolved, but
whether any substance now remains in the vesicles I cannot
determine; there is nothing which is stained by osmic acid.
With all other fixing reagents which I have used the vesicular
structure disappears.

The protoplasm of P. viridis appears then to be perfectly
homogeneous, and small portions of it may at times be observed
at the periphery of the organism free from all contents, but the
great mass of it forms a mere scaffolding for the numerous
vesicles, and is, moreover, densely packed with bacteria

1 ¢ Zeit. f. w. Zool.’ Bd. xli, p. 189.

364. ALFRED GIBBS BOURNE.

(‘‘ crystals’ of Greef), to say nothing of its various other con-
tents. The vesicles contain a fluid substance impregnated
with chlorophyll. The vesicles and the bacteria are to be
regarded as bodies contained in the protoplasm, and the latter
may flow out leaving all its contents behind. When the proto-
plasm does flow out in this way some of the bacteria soon follow,
and may then be seen to start an active movement; and if the out-
flow continues, the superficial vesicles leave the central mass
and may be seen isolated in the hyaline protoplasm (fig. 5).

Protoplasmic Movements, Pseudopodia.

Specimens of P. viridis will often remain for a long time
absolutely quiescent; in such a condition the animal is, if not
flattened by acover-glass, fairly spherical in shape, and the green
vesicles extend close up to the periphery. They always remain
embedded in the protoplasm, so that they do not actually come
to the surface ; and, indeed, there is at the surface something
more than the mere envelopes of the most superficially placed
vesicule, or else the margin of an optical section would pre-
sent a sinuous curve, the sinuosities of which corresponded to
a row of vesicles, which is not the case.

In specimens where movements are taking place these move-
ments are usually very active, and most so at the periphery,
although they are by no means confined to that region.
In this matter P. viridis agrees with other species of
Pelomyxa.!

Long-continued movements often take place without the
protrusion of any pseudopodia. Owing to its great size, P.
viridis, and, indeed, other species of Pelomyxa, move about
from place to place, at any rate while under observation, less
frequently than smaller amceboid organisms.

When the movements, which result neither in the protrusion
of a well-marked pseudopodium nor in a translation of the
organism, are taking place, the margin of an optical section
consists of a hyaline layer, the movements in which have a
wave-like, undulating appearance. It looks as though the

1 Bronn, ‘ Protozoa,’ p. 97, foot-note.

ON PELOMYXA VIRIDIS. 365

hyaline protoplasm protruded at some one spot, leaving all its
contents behind, and then spread out laterally, while a sharply
defined line seems to mark the original boundary; this line
afterwards disappears, giving one at first the impression that
the protoplasm, which has, so to speak, overflowed the original
boundary but remained in contact with it, is separated from the
rest of the protoplasm, except at the spot whence it actually
protruded, by a sort of pellicle, which is subsequently dissolved.
Gruber! has figured such an appearance for P. villosa, and
says, “ Wenn an einem ausgetretenen Protoplasmafortsatz
die Vacuolen und die Kornchen einige Zeit durch eine scharfe
-Linie von der Sarkode getrennt bleiben, so beruht dies darauf
dass das Pseudopodium durch Zusammenfliessen des Plasmas
ueber diese Stelle hinentstand, und dass die feine peripherische
Schicht von Protoplasma, welche vorher die Grenze gegen das
umgebende Medium bildet, und welche jetzt von der Masse
des Pseudopodiums umflossen wird noch eine Zeit lang er-
halten bleibt ;” and regards this as further evidence of the
existence of a relatively hard cuticular layer or pellicle formed
by the action of the water, a view which the same author put
forward on a previous occasion.”

I cannot accept this interpretation of the facts. I do not
believe that the hyaline protoplasm is protruded at one spot
only, and that there is lateral displacement of its particles ;
the undulating appearance is chiefly caused, like other wave
movements, by extrusions and retractions in a radial direc-
tion. The appearance of the boundary line may be explained
by careful focussing, the line being at some slightly different
level from the protruding protoplasm, and its disappearance
being due to a subsequent protrusion at that particular level.
I am not at all sure how far the existence of any such pellicle
as Gruber describes is in accordance with the views as to the
relation of protoplasm to imbibition water.®

If the density of the surrounding medium is increased, as

1 * Zeit. f. w. Zool.,’ Bd. xli, 1884, pl. xiii, fig. 2, and p. 190.

? ‘Zeit. f. w. Zool.’ Bd. xxxvi, 1882, pp. 457—469.

° See Engelmann on “ Protoplasmic Movement,” in this Journal, vol. xxiv,
1884, p. 390.

366 ALFRED GIBBS BOURNE.

by the addition of a 1 to 2 per cent. solution of common salt,
the protoplasm shrinks, owing to withdrawal of imbibition
water. This might, of course, take place through the pellicle,
but this is not probable, as such a layer must be impermeable,
or difficultly permeable, to water, if it have the function of
protecting the central protoplasm from the action of water ;
and, in any case, when a sudden shrinking took place, one
might expect to see some crenation of the surface, such as
is exhibited by a human red corpuscle when it is exposed to a
medium denser than its own plasma. No such crenation
takes place; but, on the other hand, some portions of the
protoplasm usually stick to the cover-glass and become torn
off, while some of the protoplasmic contents escape into the
water and are carried away by the current.

I have at times observed a similar phenomenon when a long
pseudopodium was quickly withdrawn. A specimen of P.
viridis, which has been moving about under a cover-glass,
habitually leaves some portions of itself sticking to the glass.
Appearances, such as are shown in fig. 2, constantly occur
where a pseudopodium is retreating, and are due to the stick-
ing of the superficial protoplasm to the glass.

I have also constantly seen villi, such as Gruber! has
figured for P. villosa, formed in the same manner.

Such phenomena seem. to me to render the existence of
any pellicle unlikely. Moreover, as will be described below,
when a specimen of P. viridis is torn into pieces with
needles the water seems to penetrate and cause almost instant
disintegration. Now, if the protoplasm of these forms is in
the habit of forming a pellicle directly any new surface is
exposed to the water, why does it not do so in this case? It
appears more probable that so long as the protoplasm is alive
the amount of imbibition water which can be taken in is
regulated, and that in the instance above cited the protoplasm
is killed by a shock caused by the teasing, so that excess of
water can at once penetrate and cause disintegration.

The pseudopodia are usually very blunt and lobose, and

1 « Zeit. f. w. Zool.,’ Bd. xli, pl. xiii, fig. 3.

ON PELOMYXA VIRIDIS. 367

resemble those figured for P. palustris. When one of these
large pseudopodia is about to be extruded the movements
affect very deep-lying portions of the protoplasm; a wave-like
bulging occurs, which gradually pushes out in a radial direc-
tion; an active current passes outward along its centre; all
the protoplasmic contents in the neighbourhood are carried
outwards and, having arrived at the extremity, pass back again
in the peripheral portions of the pseudopodium, which thus
narrows while it is elongating. The whole process gives one
strongly the impression that the impulse for the extrusion is
central and deep-lying in its origin. It is not that a small
pseudopodium is gradually increased in size, but the mass of
protoplasm extruded is actually greatest at first and then
gradually diminishes. Even when fully extruded the back-
ward peripheral currents still continue, especially in the
proximal portion of the pseudopodium, which thus becomes
flask-shaped, joined to the main portion of the organism by a
narrow neck.

All this time no peripherally placed colourless layer ap-
pears, the vesicles and other protoplasmic contents being
pushed on up to the surface; but when this pseudopodium
commences to retreat the contents pass backward faster than
the protoplasm itself, and a colourless superficial layer is left.
Fig. 3 represents the distal extremity of a pseudopodium
which has just commenced to retreat. If this withdrawal is
rapidly continued some of the colourless protoplasm will be
left sticking to the glass.

Besides these large pseudopodia, small ones are often found
which seem to take their origin in the superficial layer. In
these latter the hyaline protoplasm is first protruded, and into
this a number of the bacteria appear to burst out, followed
almost at once by some of the vesicles, which often sepa-
rate a good deal from one another and appear as so many
isolated spherical green droplets; these are sometimes fol-
lowed by the other protoplasmic contents (fig. 5). These
pseudopodia are never prolonged to any extent.

The rate at which the large pseudopodia are protruded is

368 ALFRED GIBBS BOURNE.

considerable. I have found it to be about 0°75 mm. a
minute. Engelmann! states that a velocity of 0°5 mm. a
minute is sometimes attained by an Ameeba, and may be con-
sidered as exceptionally rapid. I have made my observations
at the ordinary temperature of the room; but this is about
30° C. at this time of year, and is thus not far below the
optimum temperature for protoplasmic movement.

Nuclei.

The number of nuclei present is enormous. I have en-
deavoured to estimate the number on stained preparations.
I have made these preparations in various ways; one of the
most satisfactory is to fix and harden on the slide with
chromic acid (2 per cent. solution), followed by water and
alcohol of increasing strengths; stain with picro-carmine,
wash, and again treat with alcohol of increasing strengths ;
dehydrate, clarify, and mount in Canada balsam.

I put down the number of nuclei present in a large indi-
vidual at 10,000.

They vary a little in size, but average, when living, about
0:03 mm. in diameter.

It has struck me that there may be some connection
between the bulk of nuclear matter and the bulk of proto-
plasm connected with it.

I calculate that in P. viridis all the nuclei taken together
occupy ;; of the total bulk of the organism. I have not
many data at hand for comparison with this calculation, but
in a mammalian ovarian ovum the nucleus occupies about
gy of the bulk of the ovum. So that 10,000 nuclei in P.
viridis give relatively about the same bulk of nuclear mat-
ter as is found in a mammalian ovum. The nuclei closely
resemble the nuclei of other species of Pelomyxa.

In a fresh condition they are spherical, and consist of a
nuclear membrane containing a perfectly clear and trans-
parent fluid nuclear substance, in which are seen from nine to
twelve highly refracting spherical nucleoli (fig. 7).

1 Lc. p. 379.

ON PELOMYXA VIRIDIS. 369

I have never seen these enlarged or containing anything like
a vacuole, a condition described by Greef for P. palustris;
nor have I ever seen anything in the protoplasm of P. viridis
resembling the “ glanz-kérper ” of that author, or which might
be nucleoli escaped from a nucleus.

I have frequently seen nuclei extruded from the protoplasm,
usually along with food débris or sand particles; when so
extruded they remain for some time unacted upon by the
water, but in time the water seems to penetrate the nuclear
membrane, and the nucleoli exhibit Brownian movements,
and after some time come out into the water (fig. 9); but
although I have watched them for a long time I have not ob-
served them to undergo any change.

I have been unable by the use of any preservative or stain-
ing process to make these nucleoli behave as does the single
nucleolus of so many Amcebe. This latter when treated with
osmic, chromic, or picric acids, and subsequently with picro-
carmine or alum carmine, stains deeply, while the nuclear
substance remains clear and transparent, almost unstained.
But when any one of these acids or even acetic acid is allowed
to act upon the nuclei of P. viridis, the nuclear substance
becomes opaque and granular, and the nucleoli disappear from
view on account of the now opaque character of the nuclear
substance. When stained, either by osmic acid or by picro-
carmine or alum carmine (preceded by fixing and hardening
reagents), and mounted in Canada balsam, they appear as
quite different bodies from the fresh nuclei. To such an
extent is this the case, that until I had actually watched a
single nucleus throughout the entire process I was not satis-
fied that the bodies which I had taken to be nuclei in the
fresh state were actually such bodies. What really happens
is, according to the views of Pfitzner and others, that the
chromatin substance, which is something distinct altogether
from the nucleoli, is brought into view by the staining process,
and obscures to a certain extent the nucleoli. The nucleus
stains diffusely all over, while very numerous minute granules
appear to stain more deeply.

370 ALFRED GIBBS BOURNE.

I have even after most careful examination of specimens,
mouuted whole and of sections, been unable to find any cases
of nuclear division.

Other Protoplasmic Contents.

I have already spoken of the chlorophyllogenous substance,
the permanent vacuoles, and the sand particles.

In addition to these, I have observed in many individuals
small globular refractive bodies, about ;,4,, inch in diameter,
which, from the fact that they are blackened by osmic acid, I
suspect to be of a fatty nature. They are much more nume-
rous at some times than at others.

I have on two or three occasions found all the specimens
then examined crowded with large (about 54, inch in dia-
meter) disc or lens-shaped, fairly transparent, but not highly
refracting bodies, which, from the fact that all these specimens
had been feeding on some special substance, I believe to be
nutritive bodies of some kind. I have very little doubt from
the appearance of the débris that the food on these occasions
had been Spirogyra filaments, but I was never fortunate
enough to see them in the protoplasm in an undigested state.
The protoplasm in all these individuals was simply crammed
with the food débris. It is probable that these specimens had
got hold of some Spirogyra, of which there was a little on the
surface of the mud, and that it had proved a very easily digest-
ible food, and that a great deal of nutriment had become
stored up in the protoplasm. The bodies under discussion
stain a rich deep colour with iodine, but do not lose the
colour on warming. They are probably some amyloid sub-
stance.

P. viridis takes in plenty of solid food of all kinds, Daph-
nize, Ostracods, Diatoms, Naids, &c. I have also found
specimens which had taken in two or three or more plants of
what I believe to be W olffia arrhiza (one of the Lemnacez),
and which swarms in most of the tanks in Madras, and is by-
the-bye often eaten, curiously enough, in enormous quantities
by the tank frogs.

ON PELOMYXA VIRIDIS. 371

The food sometimes lies in food vacuoles, while at other
times the surface of the food particles is actually in contact
with the protoplasm.

Occasionally I have observed, when a large piece has lain in
a large vacuole, pseudopodia protruded from the vacuolar wall
and reflected over the surface of the food particle.

Physiology.

I have tried but without success to demonstrate the activity
of the chlorophyll. The animal does not readily allow of
much experiment, as it almost always dies after any manipula-
tion. Treatment with iodine has, moreover, failed to demon-
strate the presence of starch. The enormous number of the
symbiotic bacteria is correlated doubtless with the activity of
the chlorophyll.

I have tried by putting them together under a cover-glass
to get two individuals to fuse together, but without success.
On the other hand, I have often observed two pseudopodia of
the same individual form and fuse together at their distal
extremities, so as to leave for a time a hole right through the
animal. Further, I have several times cut a specimen in half
with a sharp scalpel, and kept the two halves living for some
time, and by placing the two halves close together have
induced them to fuse together again.

I have also teased specimens into Getiente with a pair of
needles, but in that case the fragments have at once com-
menced to die.

No information which we at present possess seems to me to
explain in any way why the halves of a divided individual or
the peripheral extremities of long pseudopodia protruded by
any one individual should freely reunite, while separate indi-
viduals will do nothing of the kind.

Gulliver’s Views.—Since writing the above my attention
has been drawn to a note on the minute structure of Pelo-
myxa palustris by Dr. G. Gulliver.! The author believes, in
the first place, that the exoplasm is permanently differentiated

* Journal of the Royal Microscopical Society,’ 1888, p. 11.

372 ALFRED GIBBS BOURNE.

from the endoplasm; and in the second place, that the
“endoplasm ” is composed of a number of cells. Both these
are very startling views, and while I feel sure that the conclu-
sions are erroneous, I have observed certain phenomena which
may have given rise to them.

Gulliver writes, “In the process of hardening this layer
(the exoplasm) readily separates from the subjacent softer
endoplasm.”? I have noticed in hardening specimens in a
watch-glass for section-cutting, and also under the cover-glass
for subsequent staining and mounting as whole preparations,
that the peripheral portion which is first attacked by the
hardening reagent often becomes separated from the still
living and shrinking central mass, another layer is then
attacked, and so the whole structure may harden in irregular
layers. The hardened portions break away with extreme
readiness from the unhardened ones.

Very curious appearances are sometimes thus produced, but
I am convinced that the phenomenon is a mere accident in
the hardening process. The examination of living specimens
can leave no doubt as to the absence of any permanent differ-
entiation of the body substance into exoplasm and endoplasm.
With regard to the structures which Gulliver takes to be cells
building up the “ endoplasm,” and which Lankester has sug-
gested may be swarm-spores, I have formed a less definite
opinion. In a living specimen one can see no such structure,
but in hardened specimens one does undoubtedly find small
rounded portions of the protoplasm lying in and yet separated
from the rest of the protoplasm. These bodies have, however,
no definite and constant characters, they differ considerably
in size, sometimes include one nucleus, sometimes more than
one, and frequently none at all. It is difficult to detail the
precise appearances which lead me to the conclusion ; but I
am much inclined to think that we have here merely an acci-
dental rounding off of portions of the protoplasm, which takes
place during the hardening process. I have seen similar
phenomena in specimens which have died from having been
kept mounted for a long time under a cover-glass. The body

ON PELOMYXA VIRIDIS. one

substance in these large Pelomyxe forms a considerable mass,
and this breaks up very readily after death into small
droplets.

I find nothing in the structure of P. viridis which would
lead one to suppose that these organisms have any affinities,
other than those usually assigned to them. Pelomyxa
belongs, as Biitschli says, to the family Amcebea lobosa.

In spite of repeated endeavours I am unable to throw any
light upon the reproductive processes which may obtain in
Pelomyxa.

EXPLANATION OF PLATE XXVIII,

Illustrating Professor Alfred Gibbs Bourne’s paper on
“ Pelomyxa viridis.”

Fic. 1.—An entire specimen, partially spread out under a cover-glass,
magnified about four diameters. In the left-hand lower corner is a Daphnia
shell, which the animal had just extruded. The opacity of the central region
is caused by a great accumulation of mud or sand particles.

Fic. 2.—Extremity of a blunt pseudopodium, which is retreating. Small
droplets of the colourless non-granular protoplasm are being left sticking to
the slide ; some of these will become detached as the pseudopodium retreats
further. The magnification is not sufficient to allow of the nuclei and vesicles
being distinguished, but a few of the larger vacuoles, v. v., are seen.

Fic. 3.—A long thin pseudopodium in the act of retreating, drawn to an
even smaller scale than the preceding figure.

Fic. 4.—Extremity of a blunt pseudopodium, more highly magnified. The
existence of the peripheral layer of colourless protoplasm indicates that the
pseudopodium is not being actively protruded, while its small extent and
comparative regularity shows that the pseudopodium is not being rapidly
withdrawn. 2.2. Nuclei. fv. Food vacuole, containing food débris. — s.
Naid seta. m.m. Sand or mud particles. v.v. Vacuoles. The small circles
which are shown all over the figure, except in the colourless peripheral portion,
represent the vesicles with their chiorophyllogenous contents. In places

374 ALFRED GIBBS BOURNE.

where, as over the food vacuole, marked /. v., there is only a single layer of
these vesicles, the green coloration is very faint.

Fic. 5.—Portion of the periphery of an individual where there has just been
a slight outflow of colourless protoplasm, and which will be withdrawn without
the protrusion of any pseudopodium. The bacteria (? crystals) seen coming
out into the protruded protoplasm exhibit active (? Brownian) movements. The
green bodies represent the vesicles; two of these are seen coming out into
the protruded protoplasm.

Fig. 6.—View of a region precisely similar to that drawn in the preceding
figure, in an individual which has been treated with osmic acid. The colour-
less protoplasm has stained; while the vesicular contents, now no longer
green, are not to be seen. It is seldom that the vesicular structure is so well
seen; usually, upon the application of reagents, the vesicles seem to collapse,
and so no such network as is here shown can be seen.

Fic. 7.—View of a small portion of the central region of an individual,
showing the same structures as in Fig. 4, with the addition of the bacteria.
Three nuclei are drawn with their contained nucleoli.

Fie. 8.—A nucleus with an unusually large number of nucleoli.

Fic. 9.—A nucleus which has been extruded with food débris, and which
has swollen under the action of the water, while its nucleoli have burst out.

N.B.—Figs. 5—9 are drawn to about the same scale as the line A B at the
bottom of the plate, which represents +3,, of an inch.

THE MEDUSA OF MILLEPORA MURRAYI. 375

The Meduse of Millepora murrayi and the
Gonophores of Allopora and Distichopora.

By

Sydney J. Hickson, M.A., D.Se., &c.,
Fellow of Downing College, Cambridge.

With Plates XXIX and XXX.

I. Toe Mepus# or MILLEPORA MURRAYI.

In 1884, Quelch (11), while examining the structure of the
hard parts of a new species of Millepora (M. murrayi), dis-
covered a number of small cavities which he supposed to be
the receptacles of the ova or embryos like the ampullz of the
Stylasteride. .

Professor Haddon has recently placed in my hands some
excellently preserved specimens of a species of Millepora
that he collected on the reefs of one of the islands in Torres
Straits. This species seems to be closely allied to Quelch’s
Millepora murrayi, but the identification is a matter of
some difficulty, as the pieces at my disposal are small.

On making a series of sections through a portion of a decal-
cified branch I discovered a number of medusiform structures,
each bearing a large saucer-shaped spermarium. They are
situated immediately beneath the surface, and covered by an
operculum of modified ectoderm cells.

Sections made by von Koch’s method of grinding hard and
soft parts together in solid Canada balsam show further that
these medusx exactly fit into the ampullar cavities of the
skeleton, and form the only explanation of their presence.

VOL. XXXII, PART III,—NEW SER. BB

8376 SYDNEY J. HICKSON.

The eggs of this species are, as in Millepora plicata (6),
very small and contain no yolk, and I have seen no embryos and
no parasites that could cause or fit into these cavities.
Quelch’s ampulle, then, are the cavities that contain male
medusz.

The Structure of the Meduse.—The meduse may be
found in all stages of development in the different parts of the
same branch. They are very irregularly distributed, and it is
difficult at present for me to give any hints to guide natural-
ists in the search for them. They are never found, so far as
my experience goes, close to the free extremities of the
branches. In my specimens they were found in greatest
abundance at a distance of three quarters of an inch to one
inch from the free extremity, but a few specimens were found
quite close to the attached base of the colony. Some branches
appear to be devoid of them.

All the stages of development may be found with care and
patience, but the stage represented in fig. 10 is the most fre-
quent in my preparations.

A central ManuBRIuM (Man.) hangs in the sub-umbrella
cavity bearing the large spermarium (Sperm.). It is composed
of irregular endodermal cells, and contains a considerable
cavity continuous with the cavity of the subjacent ccenosarcal
canals.

The sPERMARIUM appears to be double in section, but is
really saucer-shaped. It contains a large number of spherical
spermoblasts lying in a homogeneous fluid (?). It is covered
by a very thin coat of flattened ectoderm cells continuous with
the inner ectodermic lining of the umbrella.

The uMBRELLA is composed of three layers: a median layer
of solid endoderm continuous with the endoderm of the manu-
brium, and an inner and outer sheath of ectoderm continuous
with one another at the free rim of the umbrella.

The inner sheath of ectoderm is, as mentioned above, con-
tinuous at its proximal side with the thin coat of ectoderm
covering the spermarium. The outer sheath is continuous
with a sheath of ectoderm (Gon.) lining the cavity of the

THE MEDUSA OF MILLEPORA MURRAYI. OV7

ampulla; and this again is continuous with the superficial
ectoderm of the colony.

At the margin of the umbrella both ectoderm and endoderm
are thicker than they are elsewhere, and the medusa presents
in consequence a thickened rim at its free border. There are
no radial or ring canals. In meduse at this stage no cavity
is apparent between the outer wall of the umbrella and the
ectoderm lining the ampulla.

Above the codonostome (i.e. mouth of the umbrella) there
is an operculum (op.) of flattened ectoderm cells continuous
with the superficial ectoderm and the ectoderm lining the
cavity of the ampulla, which completely closes the gonan-
gium.

Different Forms of the Medusa.—The spermarium
varies immensely in size. Sometimes it is simply a thickened
ring round the manubrium, sometimes it nearly fills the cavity
of the umbrella. In consequence perhaps of this variation in
the size of the spermarium, the appearance of the manubrium
varies. In fig. 10 the manubrium is a large well-developed
structure with a considerable lumen. In fig. 9, which repre-
sents a younger stage, there is no manubrium at all apparent,
but the spermarium simply rests on an irregular mass of
vacuolated endoderm cells. Many intermediate conditions
between these two extremes may be observed. Further, the
condition of the endoderm of the manubrium presents many
variations. In some cases the cell outlines are well marked,
and the nuclei regular in position and spherical in shape. In
other cases the endoderm is a loose vacuolated tissue in which
no ceil outlines can be distinguished, and the nuclei are irre-
gular in shape and scattered through the spongy substance of
the tissue.

It is not my purpose to offer in this place any explanation
of these appearances, I wish merely to call attention to them
before passing on to other matters.

Development of the Medusa.—The medusa of Mille-
pora is a transformed zooid. It is not a zooid specially
modified from its first appearance to bear the spermarium, but

378 SYDNEY J. HICKSON.

an ordinary zooid of the colony changed into a medusa after
the migration of spermospheres into its ectoderm, and subse-
quent development there.

The evidence that supports this statement rests upon a
number of observed facts, that for convenience’ sake may be
arranged under the following heads:

1. The various stages in the transformation of the zooids
into medusz that can be observed in sections of the decalcified
corallum.

2. The absence of any structure that can be compared to
the ectodermic invagination, called the entocodon or glocken-
kern, that characterises the early stages in the development of
the medusa of the Hydroidea.

8. The position of the medusz in the colony.

4. The presence of large nematocysts in the superficial ecto-
derm above the younger forms of medusz.

1. The most important of these, and the only one upon which
much stress can be laid, is the first. The others afford the
necessary confirmation.

The earliest recognisable forms of the sperm mother-cells
are found in the canals in the immediate neighbourhood of
the zooids (Sperm. S,., fig. 1). They migrate from this po-
sition into the ectoderm of the zooids, where they collect
together to form a spermarium.

That the sperm mother-cells do actually migrate from the
germinal epithelium into the zooids seems to me to admit of
no doubt. The youngest stages of the germ-cells are never
found in any part of the zooids, and the youngest stages of
the zooids never bear either germ-cells or spermoblasts.
These two observations prove, firstly, that the germ-cells do
not arise in fully developed zooids ; and secondly, that new
zooids or medusz are not formed at the localities in the canals
where the germ-cells arise. They must, therefore, move from
the position where they are first developed to the position
they occupy in the zooid.

In a few cases I have seen two or three spermospheres
(Sperm. S,., fig. 1), or aggregations of spermospheres, lying

THE MEDUSZ OF MILLEPORA MURRAYI. 379

separately in the ectoderm of the zooids, but in the majority
of cases there is but a single cluster or aggregation (figs. 2,
3, and 4). The largest and most fully developed of these lie
at the apex of the zooids (figs. 5, 6, and 7).

The conclusions from these facts seem to be that the germ-
cells developing in the canals until they reach the stage corre-
sponding to the sperm-morula or spermosphere migrate towards
the zooids, fusing into aggregations as they doso. Having
reached the zooids they take up a position between the ecto-
derm and endoderm of their apices, and continue there the
later stages of their development.

The spermospheres are most frequently found in the dacty-
lozooids, but in a few cases I have found them in gastro-
zooids (fig. 3). They have probably no preference for
either the one form or the other; but they are found
more frequently in the dactylozooids, partly because these
forms are more numerous, and partly because the gas-
trozooids are usually more remote from the larger ccenosar-
cal canals.

The spermarium having been formed at the apex of the
zooid certain noticeable changes take place. In the first place
by a thickening of the ectoderm the pore becomes narrowed
(figs. 5,6, and 7). The tentacles become flattened out, and
the nematocysts disappear. The spermarium sinks into a cup-
shaped receptacle on the summit of the zooid, and the endo-
derm of the edge of the cup grows out, pushing before it the
ectoderm.

These changes are represented in the two figs. 6 and 7. In
the next stage the cup-shaped receptacle of the spermarium
has grown out into a bell-shaped structure (fig. 8). The sper-
marium is much larger in size, and the pore is completely
closed by ectoderm. In the later stages (figs. 9, 10, and 11)
the following changes may be noted. The operculum is formed,
shutting off all access from the cavity of the gonangium to the
exterior. The walls of the bell-shaped outgrowth become
considerably attenuated, and lie close against the ectodermic
wall of the ampulla. The manubrium is formed probably by

380 SYDNEY J. HICKSON.

a regeneration of the endodermic tissue and its growth into the
centre of the spermarium.

In the last stage I have observed the medusa is completely
separated from the canal system, and lies freely within the
cavity of the ampulla. The walls of the umbrella, except at
the margin, are extremely thin. ‘The manubrial endoderm
contains a closed cavity (fig. 11). This stage is probably the
last that occurs before the embryo escapes frem the corallum.
There are no nematocysts developed on the thickened margin
of the umbrella, there are no sensory bodies, there is no
velum, and no mouth.

2. In the development of the medusz of Millepora that
has just been described there is no structure formed at any
time that can be compared with the inner fold of ectoderm or
* glockenkern” that forms the walls of the sub-umbrella cavity
of the medusa of the Hydroidea. Had such a structure been
found, there might have been some ground for supposing that
this medusa is a bud that grows out of the degenerated tissues
of a zooid. The meduse of Millepora are, however, certainly
not formed by budding from the zooid in the sense that the
medusz of such a form as Corymorpha are budded from the
hydranth.

3. The diagrammatic figures that are frequently given of
zooids of Millepora, representing a centrally placed gastro-
zooid in a complete circle of dactylozooids, is perfectly correct
for some species of Millepora and the younger branches of
others.

In M. murrayi the zooids are scattered over the older
parts of the corallum in an irregular manner. The circular
systems can be made out, but over and above the zooids in their
regular circles there are both gastrozooids and dactylo-
zooids scattered irregularly within and between the circles
(cf. Quelch [11], p. 192).

The medusz occur both in the regular circles and irregu-
larly between them, as may be seen by reference to Woodcut 1.
When a piece of Millepora is decalcified and cleared in oil of
cloves or turpentine, and examined with a low power of the

THE MEDUS# OF MILLEPORA MURRAYI. 381

microscope, the arrangement of the zooids, meduse, and
coenosarcal canals can be very readily observed. The figure I
have given was drawn, by the help of the camera, from such a
preparation. The larger canals to which I have referred above
form a wide-meshed network immediately below the surface.
Each mesh is an irregular polygonal figure embracing the

Woopcut 1.—Diagram of the arrangement of the zooids and meduse of
Millepora murrayi. G. Gastrozooids. D. Dactylozooids. M. Me-
dus. The larger canals are represented by irregular black lines.

whole of one circular system of zooids. The medusz are
always found either upon or quite close to these large canals,
and thus they are sometimes without the circles, and some-
times in a position corresponding to that of a dactylozooid of
the regular circle.

The position of the meduse in the colony cannot be used as
an argument against my statement of their origin; in fact,
whatever bearing it may have is in its favour.

4. When a decalcified specimen of Millepora is examined
from above, a cluster of large nematocysts may be seen at the
mouths of the gastropores aud dactylopores. They may also
be seen in sections (figs. 1, 2,6,7, Nemat.). When the medusa
is formed and the pore closed by the operculum these large
nematocysts can be of little or no service, so they are shot
and no new ones take their place. In the figures of the
sections through the older meduse (9, 10, 11) the reader will
notice that none of these large nematocysts are to be seen.
Where the operculum is not completely formed (fig. 8),

382 SYDNEY J. HICKSON.

although the zooid has to all appearance changed into a
medusa, one or two of these large nematocysts remain.

Il. Toe Mate GonopHores oF ALLOPORA AND
DIsTICHOPORA.

1. Distichopora.—The male gonophores of Distichopora
may be seen in clusters on the branches of the male stocks.
They are small whitish bodies lying in the ampulle of the
coenosteum, and covered by a very thin semi-transparent wall
of lime and cenosarcal canals.

An examination of a series of sections through one of these
branches shows that the male gonophores are found only in
these superficial clusters (fig. 12). They are never found
deeply seated in the ccenosteum, nor in other places than those
indicated by external appearances.

One, two, or even three gonophores, in different stages of
development, springing from a diverticulum of the cceno-
sarcal canal system, may occupy each ampulla.

A ripe male gonophore (fig. 14,) is a spherical, oval, or
pear-shaped body, with an endodermal cell mass, representing
the trophodisec on the side turned towards the axis, and a
short conical or tubular seminal duct on the side turned
towards the periphery. The sheath of the gonophore seems
to be a simple layer of flattened ectoderm; but I am per-
suaded, after the examination of a great many sections and
the study of the development, that two layers are represented,
the inner or endodermal layer being extremely attenuated
and devoid of nuclei.

When a very young bud is examined with a high power
(figs. 13,, 14,, 15), the rudiment of the spermarium may be
seen to be a homogeneous mass of protoplasm, containing a
number of large spherical nuclei. It occupies a position
apparently between the ectoderm and endoderm of the bud,
As the spermarium increases in size the endoderm becomes
cup-shaped in the bud, and the margins of the cup are pro-
duced into a very thin sheath between the ectoderm and the

THE GONOPHORES OF DISTICHOPORA AND ALLOPORA. 383

spermarium (fig. 16). At the peripheral pole of buds that
are about half-way developed there is a thickening of the two
sheaths of the gonophores, cell outlines are well marked, and
the cells are nucleated (figs. 14 and 16). In this way the first
rudiment of the seminal duct is formed. The two layers are
from their first appearance quite distinct from one another,
and there is never any indication that the two cell layers
are formed by a splitting of the ectoderm. Just before the
spermarium becomes mature the ectoderm, and subsequently
the endoderm, are folded to form a conical cap, and this
subsequently pushes through the superficial covering of the
gonangium to form the seminal duct to the exterior (fig. 18).

Meanwhile, changes have taken place in the endoderm at
the base, i.e. on the axial side of the bud. In the early
stages of the bud there is a wide lumen in the endoderm, the
cells are cubical in shape, and their outlines well marked; in
the later stages the lumen becomes obliterated, the cells lose
their distinct outline, and the endoderm degenerates into an
irregular mass of tissue, with scattered nuclei (figs. 18, 14,
15, 16).

2. In Allopora (fig. 19) the male gonophores are scattered
irregularly in the corallum, and lie at such a distance from
the surface that there is no trace of their existence exter-
nally. I have been able to find them only in the old thick
branches. I cannot say for certain whether Allopora is her-
maphrodite or diccious. The specimens at my disposal
consisted of a number of fragments in a bottle, and I found
on the smaller and younger branches numerous female gono-
phores, and on the thicker and older branches numerous male
gonophores; but I have not found both sexes on the
same branches. I have no information whether the older
and younger branches in the bottle are fragments of the same
colony. If Allopora is not dicecious, then it is probably
protogynous, like Millepora, the female sexual cells being
formed first in the younger parts of the colony, and the male
sexual cells later in the older parts.

The male gonophores of Allopora resemble those of Dis-

384 SYDNEY J. HICKSON.

tichopora in every detail of structure except one, and that
is that the endoderm of the base is produced into the
substance of the spermarium as a club-shaped spadix or
manubrium (fig. 20, Spa.).

The spermarium is covered by a double sheath of very thin
ectoderm and endoderm, and the seminal duct is produced in
the same way that it is in Distichopora. When the sper-
marium is ripe the seminal duct perforates the superjacent
structures, and serves as the duct for the spermatozoa to
escape to the exterior. As the gonophores of Allopora are
situated much more deeply than they are in Distichopora,
the seminal ducts are considerably longer.

As a rule, only one gonophore is seen in each ampulla of
Allopora, but occasionally two (fig. 19, gonophore 2), and
very rarely three, in different stages of development may be
“seen.

Lastly, it must be observed that the fully developed male
gonophores of Allopora are much larger than those of
Distichopora.

From a large number of measurements I have obtained the
following average measurements :

Longest diameter of male gonophores of Allopora . 0°38 mm,
” ” Pe Distichopora 0°19 ,,

The male gonophores of a few species of Stylasteridz have
been described by Moseley (10).

In Sporadopora dichotoma the specimens were all males.
“They are ovoid bodies with the long axes directed at right
angles to the surface of the coral. Sometimes only one such
body is present in an ampulla; sometimes two or three. The
outer extremities of the gonophores are sometimes drawn
out into a short tail-like prolongation.” ‘This structure pro-
bably corresponds with the seminal duct of Allopora and
Distichopora. ‘ There is a cylindrical spadix in the centre.
The bases of the gonophores are continuous with large canals
of the ccenosarcal meshwork, the endoderm of the spadix
being continuous with that of these canals.”

THE GONOPHORES OF DISTICHOPORA AND ALLOPORA. 3885

In Pliobothrus symmetricus the male gonophores are
sacs containing a number of small ovoid bodies, which contain
spermatozoa, or sperm-cells, in various stages of development.
The exact structure of these smaller bodies and their relations
to the endoderm were not determined.

Only male specimens of Stylaster densicaulis were ob-
tained. Each male ampulla contains two or three ovoid gono-
phores, which are attached to large offsets of the ccenosarcal
meshwork at one end of their longer axes. They have an
internal spadix, and in finer structure seem to differ very
little from the male gonophores of Sporadopora.

Moseley also describes the male gonophores of Allopora
profunda, and remarks that they are very similar to those of
Sporadopora. He does not figure the seminal duct of this
genus.

Only one male specimen of Astylus subviridis was exa-
mined by Moseley. ‘‘ The male gonophores appear as large
rounded lobulated masses resting within the ampullar sacs, and
springing from stout offsets of the ccenosarcal meshwork,
which pass into the sacs to reach them. .... The sac as
it enlarges becomes gradually pedicellate, and, when mature,
is attached to the central mass by a narrow pedicle of some
length. The walls of the pedicle are continuous with the
ectodermal wall of the sac, which wall contains well-developed
nuclei in its substance. Within the sac of the lobule a
second sac, composed of a finer membrane, encloses the
mature or developing generative elements. The wall of this
inner sac is not prolonged into the cavity of the pedicle, but,
passing across its commencement, shuts off the main cavity of
the lobule from this latter. .. .. No rounded spadix, such
as that occurring in Allopora, is present in the interior of
the lobules.” These gonophores seem, from the figures and
the description given, to be very similar to those of Disti-
chopora.

It is not at all probable that Moseley overlooked the
spadix, for in his figure there are represented no fewer than
seven gonophores ; and he remarks that his material was in a

386 SYDNEY J. HICKSON.

good state of preservation. The “inner sac” of the gono-
phore that he mentions and figures is most probably the same
as the inner endodermic lining that I have described in both
Allopora and Distichopora. It would be certainly very
remarkable if this membrane is not attached to the endoderm
of the pedicle in Astylus, but this point can only be deter-
mined with accuracy by the examination of a continuous
series of sections.

The male gonophores of Cryptohelia pudica seem to be
similar to those of Astylus.

III. Toe Femate Gonornores or DistTIcHOPoRA.

The position of the female gonophores of Distichopora can
be readily seen on the female stocks by the prominent swell-
ings on the surface of the corallum. They are usually situated
on only one side of the thicker branches, but occasionally
there may be found in addition a small cluster on the opposite
side.

A section through one of these clusters shows the eggs and
embryos in many stages of development, from the minute im-
mature yolkless eggs in the ccenosarcal canals to well-advanced
planule (fig. 21).

The mature ova (fig. 23, ovum) are 0°3 to 04 mm. in diameter,
and contain a large number of spherical yolk-globules. The
large germinal vesicle is situated close to the peripheral border
of the egg, and is surrounded by a number of yolk-globules
much smaller in size than those of the other part of the egg.
The eggs rest in the cup-shaped trophodise (cf. Allopora,
Hickson, 7), and is covered by a thin coat of ectoderm and
endoderm. The trophodisc is similar to that of the female
gonophores of Allopora, but not so complicated in its foldings.
In transverse section it exhibits twelve pouches at its margin
(fig. 24). In vertical section it is simple (fig. 23); the inner
and outer pliets that I have described in Allopora are not found
in this genus.

When fertilisation has taken place the germinal vesicle loses

THE GONOPHORES OF DISTICHOPORA AND ALLOPORA. 3887

its sharp outline, and remarkable changes occur in the shape
and arrangement of the yolk-globules. My observations are
not yet complete of the early stages of the development, but I
hope to be able to publish shortly a separate memoir, giving
a full account of the development of this form up to the stage
when the larva escapes from the ampulla.

During the early stages of development the trophodisc
rapidly atrophies, and by the time a layer of columnar epiblast-
cells has formed round the embryo no recognisable trace of it
can be seen (figs. 22 and 25).

In the meantime young eggs are migrating from the sub-
jacent canals to the base of the ampulla, and in many cases
before the larva has escaped a new egg, borne by a new
trophodisc, occupies a considerable space in the same ampulla
(fig. 25).

The young eggs (fig. 22, ov.) are frequently seen quite deeply
situated in the canal system; those that are nearer to the
ampulle are larger in size and amoeboid in shape. As soon as
they reach the ampulla they show very minute yolk-granules,
which increase in size with the growth of the egg and the
development of the trophodisc.

The female gonophores of a few species of Stylasteridze have
already been figured and described by Moseley (10).

In Pliobothrus symmetricus “the gonophores are con-
tained in ampulle which are often sunk deep in the coenosteum.
. . . . The ova are solitary, one only being developed in each
growing ampulla. Hach ovum is developed within the cup of
a cup-shaped spadix,” i.e. trophodisc. ‘‘ As the ovum ad-
vances in development and increases in size the spadix enlarges
with it. Subsequently, however, in later stages, the spadix
appears not to increase further, and when in relation with a
nearly fully developed planula appears proportionately small.”

In Errina labiata “the female gonophores are closely
similar in structure to those of Pliobothrus symmetricus;
but there is this great difference, that whilst in Pliobothrus
the ampulle and their contained ova and planule remain
until maturity immersed in the coenosteum beneath its surface,

388 SYDNEY J. HICKSON.

in Errina the ampullz project more and more above the surface
as development proceeds.

«The spadix in Errina labiata is at first cup-shaped, the
walls of the cup being composed of a very thick layer of endo-
derm. The cavity of the cup is directed towards the surface of
the coral, and within it rests the single large ovum with its
distinct germinal vesicle and spot. Each ampulla contains
invariably only one spadix and ovum.”

Moseley gives a detailed account of the female gonophore of
Cryptohelia pudica. In a late stage the trophodise is
“complicated at its margin by subdivision of its lobes, which
form a network over one half of the surface of the ovum, ter-
minating in a fringe of numerous tentacula-like lobes.”

From these descriptions of Moseley and my own it seems
probable that the female gonophores of the various genera of
Stylasteride are very similar in general structure to one
another. Moseley does not describe nor figure an inner endo-
dermal membrane covering the egg, but in other respects his
descriptions of the female gonophores of the three genera,
Errina, Pliobothrus, and Cryptohelia, agree with mine of Allo-
pora and Distichopora. The chief point of variation among
the different genera is probably the lobulation or branching of
the margins of the cup-shaped trophodisc.

I prefer to retain the word trophodisc that I introduced in a
former paper to the word spadix used by Moseley for the cup-
shaped receptacle of the ovum. This structure cannot be con-
sidered to be strictly homologous with the spadix or manubrium
of the adelocodonic gonophore of the Hydromedusz. It seems
to me to be more probable that it is homologous with the
umbrella. .

LV. Tue GonoPHoRES OF THE HyDROCORALLINE AND
HypDROMEDUSH COMPARED.

In the absence of a knowledge of the minute anatomy of the
gonophores of the Hydrocorallinz, the true position of this
group in the classification of the Hydrozoa has not yet been
very satisfactorily made out.

THE GONOPHORES OF THE HYDROCORALLINA. 389

The peculiar characteristics of the group, namely, the
dimorphism of the polyps and the extensive skeleton of car-
bonate of lime, have not been considered by naturalists to be of
sufficient importance by themselves to justify the separation of
the Hydrocoralline from the Hydromedusz.

Lankester (9) places them in a separate order of the sub-
class Hydromeduse.

In the classification used at Cambridge Balfour placed Mille-
pora and the Stylasteride in the sub-order Hydroidea of the
order Hydromeduse.

Claus, in his ‘ Grundzuge der Zoologie,’ makes the Hydro-
coralline the first sub-order of the order Hydromeduse.

In Jackson’s edition of Rolleston’s ‘ Forms of Animal Life’
(8) the order Hydroidea is divided into the three sub-orders
(1) Tubulariz, (2) Hydrocoralline, and (3) Campanularie.

The opinion I have come to, based upon Moseley’s researches
and my own, is that the Hydrocoralline should be placed in
an order apart from the Tubulariz and Campanularie (i. e.
Hydroidea of Balfour and Jackson).

The classification of the Hydrocoralline with the Hydroidea
was perfectly justified by the state of knowledge at the time.
Both dimorphism and skeletal structures are, comparatively
speaking, uncertain features for the purposes of classification,
and the character and structure of the polyps and their con-
necting canal systems show undoubted affinities with many
forms of Tubulariz.

Unless, then, the organs that bear the sexual products can
be shown to differ very widely from those of the Hydroidea,
and present characteristics peculiarly their own, the Hydro-
corallinee must remain in the position that is assigned to them
by some authorities in the order Hydroidea.

These considerations demand a careful and exhaustive com-
parison of the typical gonophores of the Tubulariz, and of those
Hydrocorallines that are at present known to us.

To aid in the discussion of the homologies I have given on
p- 890 diagrammatic figures representing the structure of
(Woodcut 2) a phanerocodonic medusa, (3) a medusa of Mille-

390 SYDNEY J. HICKSON.

pora, (4) an adelocodonic medusa, (5) the male gonophore of
Allopora, (6) the male gonophore of Distichopora, (7) the
female gonophore of Distichopora.

Figs. 2 and 4 are copied from Allman with this modifica-
tion, that both endodermal tissue and endodermal cavity are

Wooncuts 2—7.—The structure} of the different gonophores compared.
Diagrammatic sections of—2. A phanerocodonic gonophore. 3. The me-
dusa of Millepora. 4. An adelocodonic gonophore. 5. Male gonophore
of Allopora. 6. Male gonophore of Distichopora. 7. Female gonophore
of Distichopora. A. Manubrium. B.Gonad, C. Endoderm. D. Ecto-
derm. E. Umbrella.

represented in black. The diagrams are modified in this way,
because no important morphological distinctions can be drawn
between endodermal structures possessing a cavity and those
that do not. For example, no one would think of drawing a

THE GONOPHORES OF THE HYDROCORALLINA. 391

fine morphological distinction between the dactylozooids of
Millepora and those of Allopora because in the case of the
former there is a lumen and in the case of the latter the endo-
derm is solid.

In comparing the structure of the phanerocodonic medusa
and the medusa of Millepora a very general similarity may be
observed.

In both there is a centrally placed manubrium (a).

In both the generative elements (B) are developed between
the ectoderm and endoderm of the manubrium.

In both there is a contractile bell umbrella, from the centre
of whose coneavity the manubrium is suspended ; in both this
umbrella is composed of a centrally placed sheath of endoderm
covered by a sheath of ectoderm on both sides; and in both
the gonophore lies in a gonangial cavity of ectoderm, which,
before the medusa is set free, is continuous with the ectoderm
of the outer wall of the umbrella.

The principal points in which these two forms differ from
one another are these :

The manubrium of 2 possesses a mouth.

The manubrium of 3 does not.

There is a system of canals (longitudinal and ring) in the
umbrella of 2.

There are no canals in the umbrella of 3.

There are tentacles and sensory epithelium at the margin of
the umbrella in 2.

There are no tentacles or sensory epithelium at the margin
of the umbrella of 3.

There is a velum in 2.

There is no velum in 3.

Too much stress should not be laid upon any of these points
of difference, for it is quite possible that tentacles, eyes, or
auditory organs, a velum and a system of gastro-vascular canals,
may be subsequently developed in the medusa of Millepora
after it is set free.

It is of importance to note, however, that these organs are
not developed while the medusa is still attached to the parent

VOL. XXXII, PART 11I1.—NEW SER. co

392 SYDNEY J. HICKSON.

stock, as they are in the typical phanerocodonic medusa of the
Tubularie.

Comparing the medusa of Millepora with the adelocodonic
gonophore (fig. 4) of Hydromeduse, the following points of
difference may be observed :

There is a codonostome in the former, there is none in the
latter.

In the former the endoderm extends almost to the margin
of the umbrella, in the latter the endoderm is reduced to a
shallow cup surrounding the base of the manubrium.

In other respects the two gonophores are practically similar.

Comparing the adelocodonic gonophore (fig. 4) with the male
gonophore of Allopora (fig. 5), two points of difference may be
observed. In the first place the endoderm completely sur-
rounds the gonad in the latter, excepting at a small aperture
at the distal pole, where it forms the inner wall of a narrow
seminal duct. Secondly, there is no layer of ectoderm between
this endoderm and the gonad in Distichopora. In the adelo-
codonic gonophore there are two layers of ectoderm between
the gonad and the wall of the gonangium.

The male gonophore of Distichopora (fig. 6) resembles that
of Allopora (fig. 5) in all respects except one, namely, that in
the former there is no manubrium.

The female gonophores of the two genera of Stylasteride
resemble the male gonophores in most respects, but in the
former there is a more complicated plieting of the base to form
a nourishing dise (trophodisc), and no structure corresponding
to a manubrium can be observed.

Do these gonophores of the Hydrocoralline represent stages
in the degeneration, or do they represent stages in the evolu-
tion of the free medusiform gonophore ?

It would be more satisfactory, perhaps, to leave these
questions to be answered at a time when weare better acquainted
with the minute anatomy of the gonophores of other species
of Millepora and the other genera of the Stylasteride.

The very convincing proofs that have been brought forward
by Balfour, Weismann, and others, showing that the gonophores

THE GONOPHORES OF THE HYDROCORALLINA. 393

of the Hydroidea, however simple in structure, represent
stages in the degeneration of medusz, may lead to the con-
clusion that these gonophores of the Hydrocorallines are also
degenerate meduse ; and it is necessary to issue a warning that
this is probably not the case.

That the medusa of Millepora is not degenerate but primitive
in its simplicity must be apparent.

In the course of its development there is no abbreviation nor
any trace of organs that were at one time functional and have
since become rudimentary. Moreover, it cannot be considered
at all probable that a free-swimming medusa, bearing immature
spermatozoa, would have lost its mouth, tentacles, sensory
organs, endoderm canals, and velum; or, if it is a degenerate
medusa, that the development of these organs would be post-
poned until after its escape.

The only view that seems to me to be at all tenable is the
one that considers the medusa of Millepora to be primitive in
its simplicity.

As regards the male gonophores of Allopora and Disticho-
pora, there is without doubt a close similarity in appearance
between certain stages in the development of the male gono-
phores of both these genera and the younger stages of the
medusz of such forms as Pennaria and other Tubularians (ef.
this paper, Pl. XXX, and Weismann (12), pl. xvii, fig. 8);
and the manubrium of Aliopora is undoubtedly closely similar
in general appearance to the manubrium of the adelocodonic
gonophore of many of the Tubulariz. In fact, the gono-
phores of some of the Hydroidea, such as Clava (Allman)
and Corydendrium (Weismann), are much less like adelocodonic
medusz, even when they reach their full development, than are
these gonophores of Allopora and Distichopora.

If it could be shown that the inner membrane covering the
spermarium is derived from the ectoderm and is not endo-
dermic as I have described it, and that structures correspond-
ing to the “‘glockenkern” do occur in the development of
these gonophores, then my principal objections to the view
that they are degenerate meduse fall to the ground. A very

394. SYDNEY J. HICKSON.

careful examination of my sections of gonophores in all stages
of development convinces me that there is in these forms no
true “glockenkern,” and that the two membranes covering
the gonad are truly homologous with the two membranes
covering the ova, namely, an outer ectodermic membrane and
an inner endodermic membrane.

The manubrium of the gonophore of Allopora is, I believe,
strictly homologous with the manubrium of the medusa of
Millepora; that is to say, it is a subsequent endodermal in-
growth into the spermarium developed for the purpose of
affording increased nourishment to the rapidly increasing sper-
moblasts.

These gonophores, then, do not represent, in my opinion,
stages in the degeneration of meduse. The Stylasteridz never
possessed free-swimming medusz, I believe, although their
gonophores may indicate to us some of the stages that the
meduse of Hydroidea passed through in the course of their
phylogeny.

Before entering into a discussion of the meaning of the
gonophores of the Hydrocorallines, it is necessary to consider
briefly the principal views that have been put forward con-
cerning the primitive or ancestral form of the Hydrozoan. Is
it probable from the evidence at our command that the ances-
tral form was a fixed colonial hydroid, or was it like a
scyphistoma larva (Hydra tuba); or, lastly, was it a floating
Hydra or actinula ?

Balfour says, “‘ A condition like that of Hydra, in which the
ovum directly gives rise to a form like its parent, is no doubt
the primitive one, though it is not so certain that Hydra
itself is a primitive form. The relation of Hydra to the
Tubulariz and Campanulariz may best be conceived by
supposing that in Hydra most ordinary buds did not become
detached, so that a compound Hydra became formed ; but that
at certain periods particular buds retained their primitive
capacity of becoming detached, and subsequently developed
generative organs, while the ordinary buds lost their gene-
rative function.”

THE GONOPHORES OF THE HYDROCORALLINA. 395

Weismann’s view is similar to that of Balfour. He says,
“Die niedrigste d. h. einfachste Form der heute lebenden
Hydroiden ist wohl Hydra; es scheint mir wenigstens fur
jetzt kein Grund vorzuliegen, sie fiir eine ruckgebildete Form,
wohl aber manche Griinde sie fiir eine sehr alte Form zu
halten, wie oben schon genauer begriindet wurde, und wie es
auch so von den meisten Forschern angenommen wird”’’ (12).

Both of these authors considered that the primitive type of
Hydrozoan was a simple sessile form more or less similar to
our modern Hydra, and that the medusa originated by the
modification of individuals bearing the sexual cells that were
budded from, and set free from, the primitive simple sessile
Hydra.

Lankester says, “‘ The particular form which the proximate
ancestor of the Hydrozoa took is most nearly exhibited at the
present day in Lucernaria, and in the scyphistoma larva
(Hydra tuba) of Discomeduse. It was a hemispherical cup-
like polype with tentacles in multiples of four, with four lobes
to the wide enteric chamber. This polype, after passing a
portion of its life fixed by the aboral pole, loosened itself and
swam freely by the contractions of the circular muscular fibres
of the hypostome (sub-umbrella), and developed its ovaria and
spermaria on the inner walls of the enteric chamber. This
ancestor possessed, like its descendants, a very marked power
of multiplication, either by buds or by detached fragments of
its body. Accordingly it acquired definitely the character of
multiplying by bud formation during the earlier period of its
life; each of the buds so formed completed in the course of
time its growth into a free-swimming person. We must sup-
pose that the peculiarities of the two phases of development
became more and more distinctly developed, the earlier bud-
ding phase exhibiting a more elongated form and simple
enteric cavity (Hydra form), which subsequently became
changed in the course of ontogeny into the umbrella or disc-
like form, with the coalesced enteric walls and radial and cir-
cular surviving spaces (medusa form). And now the ancestry
took two distinct lines, which have given rise respectively to

396 SYDNEY J. HICKSON.

the two great groups into which the Hydrozoa are divided—
the Scyphomeduse and the Hydromeduse.”

Another view has been put forward by Brooks (8), who,
from a consideration of the developments of the Trachome-
dusze and Narcomeduse, comes to the conclusion that the
ancestral form was a simple solitary floating or swimming
Hydra.

It does not seem to me to be at all clear that Claus pre-
viously expressed the same view in the ‘ Grundzuge der Zoologie,’
for although he says that Hydra is certainly not a primitive
form, that the medusa is a higher form than the polype, and
that intermediate forms between the medusa and polype are
represented by the actinula of Tubularia and Tetrapteron
volitans, he does not commit himself to the view that the
ancestral Hydrozoan was a free-swimming Hydra-like larva.

Bohm (2), on the other hand, expresses his views very clearly :
‘Kine der nachsten Nachkommen der uralten Gastraea muss
als die Stammform der Zoophyten, eine nicht weit von ihr
entfernte als die der Hydromedusen angesehen werden. Bei
der hypothetischen Construction der letzteren hat man zwischen
drei Moglichkeiten zu wahlen.

“ Entweder war diese schon entschieden polypoid ihre
nachsten Nachkommen waren Polypen, und die Medusen
haben sich erst spater aus diesen entwickelt.

“ Oder sie war ganz medusoid, die Medusen die primiaren die
Polypen die secundaren Nachkommen.

** Oder schliesslich es war eine intermediare zwischen Poly-
pen und Medusen stehende Form, und Polypen wie Medusen
haben sich von ihr aus nach zwei verschiedenen Richtungen hin
entwickelt.

‘“* Die letztere Annahme schient mir manche Griinde fiir sich
zu haben. Denn der lange Weg vom wenig differentzerten
festsitzenden Polypen bis zur hochausgebildeten freischwimm-
enden Meduse wird wesentlich abgekiirzt durch die Annahme
einer Mettelform.”’

Notwithstanding the arguments of these authors, it is not
easy to believe that the free-swimming actinula represents an

THE GONOPHORES OF THE HYDROCORALLINA. 397

ancestral type of Hydromedusan. The parasitic or semi-
parasitic habits of the actinula of most of the Narcomeduse
suggest that it is an extremely modified form, and it seems to
me to be extremely hazardous on the part of Brooks to base
his phylogenetic considerations upon such a weak and slender
foundation. The views of the earlier writers that the sessile
form is the more primitive, that in those cases in which the
medusa develops directly from the egg the trophosome has
disappeared from the developmental cycle, seem to be more
probable.

It is not necessary to enter further into the discussion of
these extreme speculative questions.

I have referred to them not in the hope of adding anything
new, nor of throwing light upon them, but in order that I may
place clearly before the reader the position I take with regard
to them.

It seems to me to be more satisfactory to regard the sessile
trophosome rather than the free-swimming actinula as the
primitive type, and the medusa as a structure produced
originally by a polypoid colony for the nourishment and dis-
tribution of the gonads.

Having thus stated my opinion as to the original form of
Hydroid, it is necessary to go further and express an opinion
as to the mode in which meduse originated.

The views of Weismann and Balfour on this question are as
nearly as possible identical. They supposed that the medusa
originated by certain buds bearing the primitive sexual cells,
retaining their primitive capacity of being detached from the
parent, and that such buds became modified for a free-swimming
existence. According to these views the medusa is homologous
with a polype, it is simply a modified trophosome, or that
trophosomes and gonophores are both modifications of some
common type.

Huxley’s original view that the gonophore is a peculiar
sexual organ has in recent years been subject to a storm of
criticism, and there are very few naturalists of the present day
who would defend the position he took. “ A medusoid, though

398 SYDNEY J. HICKSON.

it feeds and maintains itself, is in a morphological sense simply
the detached generative organ of the hydrosoma on which it is
developed.”

The gonophores of the Hydrocorallinz do not seem at first
sight to throw much light upon these questions. If we
arbitrarily assume that they are degenerate medusee comparable
to the adelocodonic gonophores of the Tubulariz and Cam-
panulariz, we cannot expect to find in them any evidence to
support either the one view or the other. But there is no
reason to suppose that they are degenerate medusiform gono-
phores. Neither in Millepora, nor in Allopora and Disticho-
pora, are there any features in development that suggest rudi-
mentary structures of medusz.

If they are not degenerate structures, then, but gonophores
of a primitive type, how can we reconcile the medusa of Mille-
pora, which is a metamorphosed polype, with the gonophores
of Allopora and Distichopora, which show no trace of polypoid
or medusoid structure ?

The explanation I would suggest is briefly as follows:
When the ova or sperm-mother cells reach a certain size and
are too large to move freely in the canal system, they set up a
local stimulus or irritation, which causes a cup-shaped folding
of the adjacent canal or polype wall. ‘This cup-shaped fold
being of advantage to the sexual cells during their maturation,
by affording increased facilities for nourishment and by in-
creasing the size of the cavity by solution of its walls, has been
modified into a definite form in each species by natural selec-
tion. When the sexual cells arrive at their maturity the
nourishment afforded by these cells is no longer necessary,
and consequently the stalk of connection with the canals be-
comes constricted until the gonophore is set free in the cavity
of the ampulla. In the ancestral form of the Millepora a
ready access to the exterior was open to the separated gono-
phore by way of the dactylopore, and thus the detached gono-
phore was able to escape and lead a free-swimming existence.

It is reasonable to suppose that all the cells of the colony of
a Millepora are capable of a certain amount of contractility,

THE GONOPHORES OF THE HYDROCORALLINA. 399

and that the slight power of contractile movement that the
original free gonophore possessed being of advantage to the
species—by enabling the gonophore to keep afloat longer and
thus spread the sexual products farther—was increased by
natural selection. Similarly the rim of the gonophore cup
was produced until it assumed the size and shape of a medusa.

The whole of this hypothesis of the origin of the medusz
rests upon the supposition that the sexual cells when they
reach a certain size set up a local irritation or stimulus, causing
a cup-shaped growth of the ccenosarc in its immediate neigh-
bourhood.

[s it reasonable to suppose, in the first place, that the gonads
when they reach a certain size do produce a local stimulus or
irritation? In young immature stocks there is no trace of
ampullz or other receptacles in the ccenosteum of sufficient
capacity for the mature gonads. Nor are there found in stocks
that are bearing but few sexual organs any empty cavities in
the ccenosteum. It is almost certain, then, that the gonads,
when they reach a certain size, cause a stimulus to certain cells
to secrete an acid (?) which dissolves the lime of the coenosteum
and causes an ampulla to be formed. There can be no doubt,
then, that the sexual cells do cause one kind of stimulus to the
tissues.

But is a local irritation or stimulus likely to cause any such
modification as circumferential folding of the canals in its
neighbourhood ?

The only direction in which we can look for an answer to
this question is to the effects caused by the irritation of foreign
substances and parasites. The Hydrocorallines, like most of
the corals, are subject to the attacks of many kinds of parasites.
Worms, molluscs, barnacles, and other forms may be seen in
every specimen that is examined.

When the colony is attacked by such a form as Tetraclita,
for example, the ccenosarce at the immediate spot on which the
parasite settles is killed, but this does not cause an atrophy of
the surrounding canal system. On the contrary, a pronounced
hypertrophy of the canal system immediately surrounding the

4.00 SYDNEY J. HICKSON.

parasite takes place, and in time it grows round and over the
parasite until itis almost buried in its substance. An examina-
tion of other forms of coral will show similar examples of
parasites and other foreign bodies covered by an hypertrophied
growth of the ccenosare.

The formation of the corbula of Aglaophenia may be
accounted for by a similar explanation. The stimulus of the
growing blastostyles causes, not only an increased activity in
the growth of the lateral branchlets, but a growth in such a
manner as to enclose the blastostyles in a cup.

Similarly the various kinds of animal galls found in Hydroids
and Alcyonarians are probably caused by a circumferential
hypertrophy of the tissues surrounding the parasitic pycnogo-
nid, crab, or mollusc.

From this evidence, then, it does seem probable that a local
stimulus or local irritation of the ccenosare of these forms
causes a growth of the tissues which gradually folds over
the seat of the irritation.

If this is the case, then, the production of a very rudi-
mentary and imperfect umbrella-shaped structure is a phy-
siological result of the stimulus caused by the growth of the
sexual cells, and the medusa is simply a modification, produced
by natural selection, of such a structure.

If this view is a reasonable one, we get over the principal
difficulty in accepting the view that the ancestral Hydrozoan
was a colonial Hydra form.

One of the chief features of the higher Protozoa and of
the Coelenterata is the power they possess of forming large
colonial organizations by asexual reproduction. And it is
reasonable to suppose that when the primitive Hydrozoan
became differentiated off from its colonial Protozoan ancestry
it retained the power of forming colonies by fission or
gemmation.

It has seemed to me improbable that Hydra can be closely
related to the ancestral type, because it does not possess this
power.

If this view of the origin of medusz is correct, there is

THE GONOPHORES OF THE HYDROCORALLINE. 401

no difficulty in believing that the ancestral form was a colonial
trophosome, and that medusez of different kinds may have
originated quite independently of one another from the
Hydroid stocks.

The original position of the gonads was the centre of the
concavity of the umbrella. As they became larger and larger
in phylogeny a conical growth of the endoderm, with re-
spiratory and nutritive functions, penetrated them, and
became the manubrium. All of these stages may be seen
repeated in the ontogeny of the medusa of Millepora. When
a mouth was formed at the end of the manubrium the gonads
were in some forms (anthomeduse) restricted to the sides of
that organ; but in other forms (leptomedusz) they were
shifted to a more convenient place in the radial canals. Ac-
cording to my view, then, the manubrium of the male
gonophore of Allopora does not prove that it is a degenerate
medusa, but, rather, that it is one stage further than
Distichopora on the road that all meduse have travelled in
the early history of their phylogeny ; that is to say, a stage
with a larger spermarium, and a special process of endoderm
for its more perfect nourishment and respiration.

Another question arises in connection with the gonophores
of the Hydrocoralline that at one time would have been
considered one of vital importance.

In the description given above of the development of the
medusa of Millepora, I have shown that it is formed by a
metamorphosis of a dactylozooid. This would support the
view, then, that the medusa is a modified trophosome.

In the description of the development of the gonophores of
Allopora and Distichopora I do not mention the zooids at all.
The gonophores are not developed in these genera (figs. 12,
19) in connection with either the gastrozooids or dactylo-
zooids, they arise quite independently from the coenosarcal
canals. They have no particular relation to the systems in
which the zooids are arranged, and there is every reason to
suppose that they are quite independent of them. Further,
these gonophores are not, according to my view, degenerate

402 SYDNEY J. HICKSON.

meduse. They must, therefore, be special organs of the
colony bearing the gonads.

To those naturalists who believe that there is a sharp dis-
tinction to be drawn between the idea of the “ individual”
and the “organ” in the animal kingdom, these apparently
contradictory cases must be very puzzling. In the one case
they would say the gonophore is an “ individual;” in the
other, it is an “ organ.”’

I am not inclined, however, to believe that it is possible
to draw a sharp distinction between these two ideas. They
are relative ideas, as Claus (5) maintains, just as “cell”
and ‘‘ tissue,” ‘‘individual” and “ colony,” must be.

The stimulus of the sexual cells of a certain size would
produce the same effect if they were formed in the ccenosarcal
canals or the zooids; but natural selection has stepped in
in the case of the Hydrocorallines, so that in the case of
Millepora the gonads do not produce this effect until they
reach the zooids, and, in the case of the Stylasteride, not
until they reach certain parts of the canal system.

The two kinds of gonophores are, then, to my ideas really
homologous, although in the one case they have reached such a
stage of development as to justify us in considering them
‘individuals,’ while in the other case they cannot be con-
sidered more than sexual “ organs.”

General Conclusions.

1. In Millepora murrayi (sp. 2?) the male gonads are
borne by meduse which escape from the ampullze in which
they are developed before the spermatozoa are matured.

2. The ova of this species are, like the ova of Millepora
plicata, extremely small and alecithal. They move in an
amoeboid manner in the ccenosarcal canals, and do not ulti-
mately rest in gonophores, nor in any specialized portion of
the system.

3. The meduse of Millepora murrayi have no radial nor
ring canals in the endoderm of the umbrella, no velum, no
sensory organs, and no mouth.

THE MEDUSH OF MILLEPORA MURRAYI. 4.03

4, The meduse are formed by a metamorphosis of an
ordinary zooid; in the majority of cases dactylozooids, but
in others gastrozooids.

5. The sperm-cells originate in the ectoderm of the cceno-
sarc and wander into the ectoderm of the zooids, where they
fuse into aggregations to form a spermarium.

6. The young spermarium is formed at the distal extremity
of the dactylozooid, and when it has reached a certain size
it causes a retrograde metamorphosis of the tissues. The
tentacles flatten out and disappear, and the zooid loses all
its characteristic features.

7. A cup-shaped outgrowth next appears which forms the
umbrella of the medusa, and subsequently a conical growth
of the endoderm penetrates into the substance of the sper-
marium and forms the manubrium.

8. The male gonophores of Distichopora occur in groups of
two or three in each ampulla in different stages of develop-
ment. The gonad is supported by a small cup-shaped
trophodisc, and enclosed in a double sac of ectoderm and
endoderm. At the distal pole of the ripe gonophore there is
a short seminal duct.

9. The male gonophore of Allopora differs from that of
Distichopora, in the fact that it is provided with a club-shaped
endodermal manubrium or spadix.

10. The female gonophore of Distichopora resembles that
of Allopora described in a previous paper; but the folds of
the trophodise are not so complicated.

11. The gonophores of the MHydrocoralline are not
degenerate medusz.

404. SYDNEY J. HICKSON.

BIBLIOGRAPHY.

-

. I. M. Batrour.—‘ Comparative Embryology,’ 1880.

. R. Boum.—“ Helgolander Leptomeduse,” ‘Jen. Zeits.,’ xii, 1878.

. W. K. Brooxs.— The Life History of the Hydromeduse :” a discussion
of the origin of Meduse and the significance of Metagenesis, ‘Mem.
Bost. Soc. Nat. Hist.,’ vol. iii, No. 12, 1886.

4. Craus.—“ Ueber Halistemma tergestinum,” &c., ‘Arbeiten des Zool.
Instit. zu Wien,’ tom. i, 1878.

5. C. Cravs.— Zur Beurtheilung des Organismus der Siphonophoren und
deren phylogenetischer Ableitung,” ‘ Arbeiten Zool. Instit. zu Wien,
vill, Heft 2, p. 159; translated in the ‘Annals and Magazine of
Natural History,’ vi, 21, pp. 185—198.

6. 8. J. Hicxson.—* The Sexual Cells and the Early Stages in the Develop-
ment of Millepora plicata,” ‘ Phil. Trans.,’ vol. clxxix, 1888, B.,
pp. 193—204.

7. 8. J. Hickson.—‘‘ On the Maturation of the Ovum and the Early Stages
in the Development of Allopora,” ‘ Quart. Journ. Mier. Sci.,’ vol. xxix,
p. 579.

8. W. H. Jackson.—‘ Rolleston’s Forms of Animal Life,’ 2nd edition,
1888.

9. HE. Ray Lanxester.—Article “ Hydrozoa,” ‘Encyclopedia Britann.,’
9th edition.

10. H. N. Mosztzy.—“ Report on Certain Hydroid, Alcyonarian, and Madre-
porarian Corals,’’ ‘ ‘ Challenger”? Reports,’ vol. ii, 1881.

11. J. J. Quetcu.— On the Presence of Ampulle in Millepora murrayi,”
‘Nature,’ vol. xxx, 1884, p. 539; and in ‘ “Challenger” Reports,’
vol. xvi, ‘‘ Reef Corals.”

12. A. Weismann.—‘ Die Entstehung der Sexualzellen bei den Hydro-

medusen,’ Jena, 1883.

a bo

THE MEDUS#® OF MILLEPORA MURRAYI. 405

DESCRIPTION OF PLATES XXIX & XXX,

Illustrating Mr. Sydney J. Hickson’s paper “ The Meduse
of Millepora murrayi and the Gonophores of Allo-
pora and Distichopora.”

Cale. The calcareous skeleton or ccenosteum. Caz. The ccenosarcal canals.
In the superficial regions the canals are crowded with zooxanthelle. Het. Ecto-
derm, coloured red. Zvd. Endoderm, coloured blue. G@oz. The ectodermal
lining of the ampulla forming the wall of the gonangium. Maz. Manubrium
of the medusa. Nemat. Large nematocysts guarding the dactylopores.
op. Operculum of modified ectoderm cells covering the pore of the ampulla.
Sperm. Spermarium. Sperm. S,. Spermospheres or aggregations of spermo-
spheres in the ectoderm of the zooids. Sperm. S,. Young spermospheres in
the ectoderm of the canals. Tent. Retracted tentacles. Um. Umbrella of
the medusa, consisting of a solid endoderm covered on both sides by ectoderm.

PLATE XXIX.
Millepora murrayi.

Fie. 1.—Section through a retracted dactylozooid of Millepora murrayi,
showing a number of spermospheres (Sperm. S,.) in the ectoderm of the cceno-
sare, and in the ectoderm (Sperm. S,.) at the base of the dactylozooid.

Fic. 2.—Section through a retracted dactylozooid, showing a single small
aggregation of spermospheres (Sperm. S.) in the ectoderm at the base of the
dactylozooid.

Fic. 3.—Section through a retracted gastrozooid, showing an aggregation
of spermospheres in the ectoderm. ‘The gastrozooids may be readily dis-
tinguished from the dactylozooids by the presence of a mouth and by the
large endoderm cells, the peripheral portions of which are filled with mucus.
Just below the gastrozooids may be seen a plate of vacuolated ectoderm cells
in section, which forms the last tabula of the gastropore.

Fie, 4.—Section through a dactylozooid, showing a large aggregation of
spermospheres on its side in a condition very similar to that I have described
in Millepora plicata (6). The spermospheres have caused a very con-
siderable depression in the dactylozooid, and are partially covered by the
surrounding parts.

Fic. 5.—An aggregation of spermospheres at the peripheral extremity of a
dactylozooid. The tentacles (¢ezt.) are visible.

Fic. 6.—An aggregation of spermospheres (Sperm. S,.) at the peripheral

4.06 SYDNEY J. HICKSON.

extremity of a dactylozooid, sunk in a cup-shaped receptacle. At Umb. may
be seen the first trace of the formation of the umbrella by the growth of the
endoderm. The position of the tentacles is still indicated by the rows of
small nematocysts.

Fic. 7.—Section through another dactylozooid, showing a still further
growth of the folds forming the umbrella. All trace of the tentacles has
disappeared.

Fic. 8.—Section through a young medusa of Millepora. The form of the
dactylozooid is completely lost. ‘The endoderm of the umbrella is solid, and
much thicker than it is in later stages. The opening of the dactylopore can
still be traced, although it is blocked with the thickened ectoderm cells. The
pore is guarded by nematocysts (emat.).

Fie. 9.—Section through another medusa. The umbrella is not completely
developed, but the endoderm is much thinner than it is in Fig. 8. The sper-
marium is much larger, but there is no trace of a manubrium. ‘The dactylo-
pore is completely closed by an operculum (op.) formed by flattened strap-
shaped ectoderm cells.

Fic. 10.—Section through another medusa, with a well-developed manu-
brium (maz.), containing a cavity continuous with a Jarge canal. The umbrella

walls are much thinner than they are in the specimens drawn in Figs. 8 and 9,
except at the margin.

Fig. 11.—Section through a medusa that lies freely in the gonangium. It
is not connected organically with the colony at any point. It is probably
ready to escape. The umbrella (Umd.) is extremely thin, except at the
margins. ‘There is a small cavity in the endoderm, but there is no mouth.
There are no tentacles, velum, nor sensory bodies on the margin of the
umbrella. Between the codonostome and the superficial ectoderm there is a
layer of mucus.

PLATE XXX.

Fic. 12.—Transverse section through a decalcified branch of Distichopora,
showing the male gonophores lying in the ampulle. One, two, or three
gonophores occur in each ampulla. At the edges of the branch are situated
the rows of dactylozooids (Dact. Z.) and gastrozooids (Gas¢. Z.).

Fic. 13.—Section through an ampulla of Distichopora, containing two
young male gonophores. Each of these is supported by its own trophodisce
containing a large lumen.

Fie. 14.—Section through an ampulla of Distichopora, containing three
male gonophores in different stages of development. The largest of these
(1) contains ripe spermatozoa, and shows on its distal pole a conical cap of

THE MEDUSM OF MILLEPORA MURRAYI. 407

cells, the undeveloped seminal duct. The trophodise (¢roph.) is reduced to an
irregular mass of endoderm cells.

Fie, 15.—Section through a very young male gonophore of Distichopora.
The young spermarium (sperm.) lies apparently between the ectoderm and
endoderm of the bud, but the endoderm is cup-shaped, and the margins of the
cup project between the ectoderm and the proximal hemisphere of the
spermarium.

Fic. 16.—Section through an older male gonophore of Distichopora,
showing the spermarium covered by the two membranes, a thin nucleated
ectoderm and a thinner non-nucleated endoderm, which is continuous with
the endoderm of the trophodisc.

Fie. 17.—Section through the earliest stage I have found of the formation
of the seminal duct. The ectodermic and endodermic elements are from the
very first quite distinct from one another.

Fie. 18.—Section through a seminal duct of a ripe male gonophore, open
to the exterior.

Fic. 19.—Section through a portion of a decalcified branch of Allopora,
showing three male gonophores lying in their ampulle. As a rule, only one
gonophore is found in each ampulla; but one case is figured (gonophore 2) in
which a large gonophore and a very young bud occur in the same ampulla.

Fie. 20.—Section through a nearly ripe male gonophore of Allopora,
showing the club-shaped endodermal spadix, and the two membranes (Ze¢. and
End.) surrounding the spermarium.

Fic. 21.—Section through a portion of a decalcified branch of a female
stock of Distichopora, showing a number of ova and planule in various
stages of development lying in their ampulle.

Fig. 22.—A portion of the same as Fig. 21, more highly magnified. The
ampullz are occupied by planule. Below the ampulle there may be seen in
the endoderm of the canals some very young eggs, containing no yolk-granules
and showing blunt amceboid processes,

Fie. 23.—An ovum of Distichopora that is nearly mature, as seen in
section. The germinal vesicle (Germ. Ves.) lies near the superficial side of
the egg, and is surrounded by small yolk-granules. The trophodisc is simple,
in vertical section, and contains a pronounced lumen.

Fre. 24.—Transverse section through an ovum and trophodise of Disti-
chopora in the plane represented by the line x a’ in Fig. 23, showing the
twelve pouches of its margin.

Fic. 25.—Section through an ampulla of Distichopora, containing a planula,
and below it a young ovum in a young trophodise.

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ON A RED PIGMENT-FORMING ORGANISM. 409

On a Red Pigment-forming Organism,
B. corallinus (?).

By

Charles Slater, M.B.(Cantab.).
With Plate XXXI.

THe number of micro-organisms known which form red
pigments is already considerable, and include the M. prodi-
giosus, B. indicus, B. ruber, and B. rouge de Kiel. The
organism described below differs from any of these in morpho-
logy, cultural characteristics, and tint of pigment produced.
It occurred as a coral-red, slow-growing, circular, non-lique-
factive colony on a gelatine plate which had been used in an
examination of the tap water of the laboratory. As the plate
had been withdrawn from the moist chambers and examined
at least once before the colony appeared, it is impossible to say
whether it was derived from the water or was an air contami-
nation. It was most probably the latter.

The colony was found to consist of short, thick bacilli with
very rounded ends. Their breadth, which is very constant, is
about 1 mu, and the average length of the individual cells from
2 to3mu. Very frequently two cells are joined end to end, but
itis unusual for more than two fully formed cells to remain in
apposition. The organism is motile, the movements being of
a rolling recurving character, with but slow motion of transla-
tion, except in the case of certain apparently young short cells.
The character of the motion recalls that of B. megaterium,
but is somewhat more active. The apparent curving and

410 CHARLES SLATER.

straightening movements seem to be due tothe rolling motion
of a slightly curved organism round its longitudinal axis.

By far the most noticeable characteristic is the highly re-
fringent nature of the poles of the cells. This refringence is
noticed in a very large proportion of the cells of a culture
examined at any stage of growth, and will be again re-
ferred to.

On gelatine peptone growth takes place easily, though not
very rapidly, and does not produce liquefaction of the medium.
The growth forms a regular, raised, moist-looking, somewhat
glistening streak on the surface of the gelatine, and is fre-
quently surrounded by a white opalescence due to a deposit of
oxalate of calcium. The colour is pinkish or coral red, and
though this tint deepens during the first week the coloured
colonies are not preceded by any definite uncoloured stage, as
is the case in the Prodigiosus, &c.

Ina stab culture the growth takes place very scantily along
the needle track, and remains colourless. On the upper sur-
face the growth spreads from the point of inoculation, and
colours well. The organism is distinctly aérobic.

On agar the growth is a little slower, and is rather more
vermilion in colour. The addition of carbohydrates to the
agar seems to make no difference in either the rapidity or
colour of the growth. In the case of the B. rouge de Kiel,
M. Laurent (Ann. Inst. Past., iv, 465) has shown that the
colour is not produced in the presence of carbohydrates. The
addition of glycerine to the medium is, however, inhibitory to
the growth of the bacillus now described. At the most there
is a slight pink growth at the margin of the fluid which always
collects at the bottom of these tubes.

In liquid media the growth is never copious, and the colour
extremely ill-developed. The media tried were ordinary
bouillon, bouillon with carbohydrates, and Pasteur’s fluid with
and without sugar. The addition of sugar both to the bouil-
lon and the Pasteur’s fluid certainly increases the growth and
assists the colour formation. The growth in these media
tends to collect at the edge of the fluid and to attach itself to

ON A RED PIGMENT-FORMING ORGANISM. 411

any extraneous body, and forms delicate, slightly resistant,
gelatinous films. The organism in these films does not appear
to have any distinct morphological peculiarities. There is a
decided tendency to the formation of a gelatinous material
round the cells in the cultures on solid matter, especially on
potato.

On potato the growth is copious and the colour well de-
veloped. The organism forms a raised irregular waxy-looking
coating with a bright reflecting surface. The growth is gela-
tinous, adherent to the surface of the potato, and the colour
is much the strongest in the superficial layers. A greyish
blue discoloration of the potato occurs round the growth,
appearing early but subsequent to the development of the red
pigment, and obviously connected with the growth of the
organism. The potato acquires within the zone of grey pig-
ment a slight alkaline reaction, but no distinct odour is de-
veloped, as in the case of M. prodigiosus. In old potato
tube cultures the pigment becomes a pale chocolate-brown.

Temperature.—The organism grows well at the ordinary
temperature, and has its optimum between 20° and 23°. Above
this the growth is slower, and at 37° ceases, but the cultures
grow rapidly again on removal to a lower temperature.
Exposure for one hour to a temperature of 60° kills the
organism. Simple drying at 37° on a cover-glass does not
impair the vitality.

Pigment.—The pigment in this organism is largely con-
tained in the cells, and is not an excretion. It cannot be ex-
tracted by simple shaking with water, ether, alcohol, or chloro-
form, but requires to be first liberated by disruption of the
cells by boiling. It is set free by boiling in water, but is not
dissolved. Alcohol and chloroform dissolve it easily, but
ether does not. It has not been obtained ina crystalline form.
Wheu acted oun by alkalies it is turned a yellowish brown, and
this is probably the reason of the change of colour in old potato
cultures. Acids restore the colour affected by alkalies, but do
not themselves cause any change in the original pigment.
With the spectroscope no absorption bands could be detected,

412 CHARLES SLATER.

the reddish solution simply cutting off the blue end of the
spectrum.

The production of the pigment is unaffected by light,
taking place as well in the dark as in diffused light.

The pigment-producing power is remarkably constant, and
in no case have any colourless colonies been obtained when
free access of air is possible. In stab cultures, and in the
depths of liquid media, the colour is absent.

If recent cultures of the organism on gelatine or potato
are examined, the growth is found to consist chiefly of cells
scarcely twice as long as broad, and so rounded at the ends
as to appear oval. These small cells are actively motile, and
the protoplasm appears to be collected as a refringent mass at
each pole. These cells are not unfrequently united in pairs,
connected by a thin filament. The bicellular organisms are
actively motile, and present the appearance of a flagellated
organism. They are especially noticeable in bouillon cultures.
Besides those two forms which seem to be characteristic of
young and active growth, there occur cells which are three
to four times as long as broad, some showing signs of division
into the bicellular stage, and others no trace of this separa-
tion.

The organism increases by fission, which is somewhat slow,
a cell dividing at the temperature of 15° C. in about twelve
hours. The most noticeable feature in this organism at what-
ever stage examined is, as mentioned above, the irregular
refractive power of its protoplasm, and its tendency to collect
in masses, having various positions in the organism, but very
frequently to be found at the poles of the cells. Stained
specimens present, also, a very great variety of appearances,
owing to their extremely irregular staining. A stained pre-
paration of this bacillus recalls, very strongly, a culture of the
Bacillus typhosus when in the condition which shows the
‘clear space’ and the so-called spores, and the appearances
presented by this pigment-forming organism are not without
interest when compared with those of the pathogenic microbe.
Certain cells show, both when stained and unstained, caps of

ON A RED PIGMENT-FORMING ORGANISM. 413

protoplasm, with an intermediate hyaline scarcely stained cell
body. These resemble the clear space cells of the B.
typhosus. In other cells the refractive mass is gathered
into a distinct rounded spore-like mass at each pole, and in
stained preparations a very common appearance is that repre-
sented in fig. 3, where a strongly stained central mass is
flanked by two oval unstained bodies, apparently corresponding
to the refringent masses, and having the appearance of
terminal spores.

Buchner has shown that the refringent terminal mass, de-
scribed as a spore in the case of B. typhosus, was really a
collection of protoplasm, and stained deeply, and did not
correspond to the unstained oval space in stained specimens.
Similarly, by direct staining on the slide, it may be seen
that the rounded refractive polar masses are protoplasmic
and stain deeply, and that the two forms a and 6 in figs.
1 and 2 are not corresponding stained and unstained appear-
ances, but the forms a and c. That the oval unstained spaces
seen in fig. 3 are not spores is indicated by their rather
irregular shapes and somewhat faint outline, which suggest
rather the space left by the withdrawal of the central pro-
toplasm from that part of the cell. Further, no attempts at
differential staining or any of the usual methods of spore
staining have been successful, and no free spores can be
demonstrated in the cultures. The cultures containing these
forms are sterilised by one hour’s heating to 60° C.

Regarding the irregularity of staining as due to irregu-
larities in the distribution of the cell contents, the question
arises as to whether the forms noticed are produced artificially
in preparation, or are the expression of normal changes in the
cells, or are the results of degenerative processes. That the
various forms are not produced artificially is shown by the
fact that similar results are obtained whether the specimens
are fixed by heat, alcohol, or simply drying, and that a
parallel irregularity is seen in fresh specimens, and in pre-
parations made by staining directly on the slide without
fixation.

414 CHARLES SLATER.

Biitschli has advanced the theory that the cell contents of
bacteria represent the nuclei of other cells, while the proto-
plasm is reduced to an extremely thin layer, in many cases
only represented by the cell wall. He bases his views
(‘ Biitschli ueber den Bau der Bacterien,etc.,’ Leipzig, 1890) on
the study of the large flagellated organisms found in sulphur
waters, which he shows to possess an internal cell substance,
having the structure and staining properties of nuclei, and
containing the granules of Ehrlich. The refractive portions
of the organism described in this paper are strongly stained
by the ordinary nuclear stains, such as alkaline aniline dyes,
and especially logwood. On this view the forms which have
been described as occurring in young freely growing cultures,
viz. the short oval cells with refractive poles occurring either
singly or in pairs, would apparently be the forms resulting
from direct division, and the polar collections of protoplasm
would represent the direct division of the nucleus. The suc-
cessive stages of this division would be represented (fig. 4, a)
by—l, the cell, with refringent poles ; 2,a form resembling a
large diplococcus, stained throughout, and formed by division
of stage 1—this form occurs but rarely; 8, a bicellular
organism found by the growth of stage 2. In this third stage
the cell stains at first equally throughout, but soon, either
before the cells separate or soon afterwards, the protoplasm
collects at the poles, i.e. the nucleus divides directly, and
stage 1 is reproduced. The division of the cell in stage 1, and
the formation of stage 3, have been directly observed.

Dr. Delépine, who kindly examined some of the specimens,
suggested that some of the other appearances were due to
karyokinesis, and, having regard to the columnar form of the
organism and its small size, various forms may be distinguished
which would represent the formation of a central plate and its
subsequent division and gradual separation. These forms are
represented in fig. 4, 6. Starting with a cell, whose contents
stain equally and moderately, these contents representing a
nucleus, there is a gradual gathering of the chromatin until
an intensely staining central plate is formed with unstained

ON A RED PIGMENT-FORMING ORGANISM. 415

poles. The whole cell is atthe same time larger. The central
chromatin then shows signs of division, and the two halves
gradually separate and travel away from one another towards
the poles. Division of the cell takes place between the two
halves, and the chromatin is once more equally distributed
over the cell. The appearance of constriction in fixed and
stained specimens at the central plate is partly artificial, asin
organisms examined in a drop culture containing methy-
lene blue the outer border of the cell may be made out, pre-
serving its original breadth until the division is about to take
place. It has already been mentioned that the B. typhosus
presents similar appearances to the above, and Babes (‘ Soc.
Anatomique,’ December, 1884) figures and describes very simi-
lar forms occurring in the comma bacillus of Koch. He states
that at first the poles of the cells are deeply stained, but as the
organism grows the deeply staining portions pass to the centre
and become fused ; and that then a clear space forms, dividing
the central mass into two parts, which indicates the com-
mencing cell division.

This organism, when grown under somewhat unfavourable
conditions, readily shows involution forms. When grown on
gelatine with scanty access of air the cells become three to four
times as long as broad without, in many instances, dividing,
and after some days become vacuolated. These vacuoles are
generally three in number, and symmetrically arranged. This
vacuolation explains many of the appearances seen in the
stained specimens. The cultures in fluid media, especially in
the depths and in sugar-containing fluids, show long, almost
leptothrix, forms, and there is a great tendency to the
formation of bud-like projections, which in some instances are
so prolonged as to resemble branchings. The protoplasm in
these forms also presents irregularities of distribution.

The following is a short list of the principal red pigment-
forming bacilli, with their differential characteristics ;

VOL. XXXII, PART III.—NEW SER E E

416

CHARLES SLATER.

Name.

Character of Cells.

Culture on Gelatine.

M. prodigiosus
B. ruber (Frank)

(Breunig)

seus (Babes)
B. corallinus .

.| Oval and round cells ‘5 to 1 p,
also has a bacillus form.
Motile.

.| Bacilli from 6 to 8 p» long, by
ly» broad. Very motile.

B. rouge de Kiel! Bacilli 2°5 to 5 long, by 7

to 8 broad. Motile.

B. indicus (Kocb)| Short cells with rounded ends.
B. granulatus ro-| Straight rods 3 to 4m broad,

5 to 6 times as long.
.| Oval cells and rods 1 » broad,
2 to 3 pw long.

Liquefies gelatine
quickly.

Liquefies gelatine.

Liquefies gelatine, which
is rapidly coloured
throughout.

Liquefies gelatine.

Does not liquefy gela-
tine.

Does not liquefy gela-
tine.

DESCRIPTION OF PLATE XXXI,

Illustrating Mr. Charles Slater’s paper ‘ On a Red Pigment-
forming Organism, B. corallinus.”

Fic. 1.—Organism unstained.

Fie. 2.—Organism stained alkaline methylene blue.

Fie. 3.—Organism stained logwood, showing apparent spore-like bodies,

Fic. 4.—(a) Stages indicating direct division.

karyokinesis.

6) Forms possibly due to
Pp y

Fig. 5.—Budding and branched forms occurring in liquid media.

Fic. 6.—Vacuolated forms unstained.

Fie. 7.—Involution forms—torula-like forms.

Fic. 8.—Organism from fluid, sugar-containing medium ; stained.

Fic. 9.—Organism growing in gelatine medium.
Fie, 10.—The same colonies twenty-four hours later, showing growth and

cell division.

Figs. 9 and 10 are not so highly magnified as the others.
Fic. 11.—Potato culture.

IMMUNITY AGAINST MICROBES. 417

Immunity against Microbes.
By

M. Armand Ruffer, M.A., M.D.(Oxon.).

With Plates XXXII and XXXIII.

Part II.

Tue first part of this paper, with few exceptions, was taken
up with the struggle taking place in the healthy body between
micro-organisms and ameeboid cells. It is now time to study
what happens where micro-organisms have found their way
into the tissues of animals.

Very little consideration, however, shows that the problem
to be solved is a most difficult one, on account of the many
factors which influence the clinical course and pathological
appearances of a disease.

In the first place, the clinical aspect and the pathological
appearances will vary according to the number of micro-
organisms introduced into the system. This fact was formerly
denied by competent bacteriologists ; for it was difficult to
imagine that the number of micro-organisms could be of
importance, when it was shown that 1,000,000,000,000th part
of a drop of blood from an animal dead of septicemia injected
into another animal reproduced the same disease. Lately,
however, overwhelming evidence has been brought together
proving the influence of numbers. Thus 250,000,000 staphy-
lococci injected into a rabbit will produce an abscess in this
animal,! but 1,000,000,000 of the same parasites will cause

1 Davaine.
VOL. XXXIJ, PART IV.—NEW SER. F F

418 M. ARMAND RUFFER.

death.! A few drops only of a culture of the Bacillus pyo-
cyaneus injected into the veins of a rabbit give rise to a
chronic disease; whereas a quarter of a cubic centimetre of
the same culture kills the animal with certainty.? One cubic
centimetre of an emulsion of the culture of the Bacillus
prodigiosus in beef-broth produced no effect on a rabbit ;°
whereas 3 cubic centimetres of the same emulsion proved fatal
to another animal.

Another and most important factor is the virulence of the
micro-organisms introduced into the system. Without entering
at present into the question of the means by which the viru-
lence of microbes can be raised or lowered, or into the other
question as to what is really meant by virulence, it is a well-
ascertained fact that this factor varies within very wide limits.
The same number of anthrax bacilli contained in a drop of
blood from an animal dead of anthrax, in a drop of Pasteur’s
first or of his second vaccines for the same disease, gives
rise to widely different effects if introduced into three animals
of the same species. The disease produced in rabbits by the
injection of a culture of the bacillus of fowl cholera ten days
old differs widely from that produced by the injection of a cul-
ture one day old. One might say almost that the virulence of
a given species of micro-organism is inversely proportionate to
the number of microbes necessary to kill a given animal.

The species of animal used for the experiments is, perhaps,
of still greater importance than the quality of the virus.
Rabbits inoculated with the bacilli of fowl cholera, of anthrax,
or of diphtheria, speedily succumb to these diseases, but
the injection of cultures of quarter-evil or typhoid fever has
no effect on these same animals. Guinea-pigs, on the other
hand, perish quickly when the bacillus of quarter-evil is in-
jected into them. ‘The inoculation of the bacillus of anthrax
into a number of European sheep is followed by the death of

1 Odo Bujwid’s numbers are somewhat higher, but this does not alter the
truth of the fact.

2 Charrin, ‘La Maladie Pyocyanique,’ Paris, 1889.

3 M, Armand Ruffer, unpublished observations,

IMMUNITY AGAINST MICROBES. 419

most of them, whereas Algerian sheep are not affected by the
same virus. The virus which may prove fatal to puppies proves
harmless to full-grown dogs.

The result of the inoculation is influenced to a considerable
extent by the fact of the animal having recovered from a previous
attack of the same disease, or having been rendered immune
against it by artificial means; much will also depend on the
state of the animal’s health at the time of inoculation. Fowls
which are naturally immune against anthrax take the disease
readily if their temperature be artificially lowered by immersion
into cold water. Contrariwise, frogs, which are also immune
against anthrax, inoculated with the same disease, die when
their temperature is artificially raised. Moreover, if the tem-
perature of a warm-blooded animal be artificially lowered, or if
it be submitted to long-continued fright, the micro-organisms
normally present in the intestine pass through the lining
epithelium and invade the blood.

Lastly, much will depend on the mode of introducing the
virus. Thus, a quarter of a cubic centimetre of a culture of
Bacillus pyocyaneus injected into the veins of a rabbit
proves fatal within forty-eight hours ; but the same quantity in-
jected under the skin causes death after a week only, or not at all.®

The problem is a very complex one, and in this paper, there-
fore, the writer will only make use of data which have been
obtained by actual experiments.

It will be best to discuss in the first place the cases in which
the virus produces a well-marked inflammatory reaction at the
point of inoculation, when introduced under the skin. We
will take, as examples, the cases in which guinea-pigs have
been inoculated with virulent quarter-evil (Bacillus Chau-
vei), as being the most typical example of the kind.

At the end of twelve to eighteen hours after a small quan-

1 Ch. Bouchard, “ Essai d’une Théorie de |’ Infection,” ‘ Proceedings of the
Tenth Medical Congress,’ Berlin, 1890.

2 Charrin, ‘La Maladie Pyocyanique.’ Compare also Armand Ruffer ‘On
the Nature of the Disease produced by the Inoculation of the Bacillus pyo-
cyaneus.’

420 M. ARMAND RUFFER.

tity of the virus, in the shape of a dried powder, has been
introduced under the skin, a small hard swelling is formed at
the seat of inoculation, which crepitates on pressure as if its
contents consisted of gas and fluid. The tumour increases in
size, and soon becomes harder and more resistent. The parts
surrounding it are then considerably thickened, the walls of
the abdomen, for instance, feeling twice as thick as natural.
After twenty-four hours or so the animal shows evident signs
of a general illness; it refuses food, does not run away when
touched, gets more and more drowsy, and generally dies just
within forty-eight hours after the inoculation.

On examining the tissues of the spot where the inoculation
has been made, the remains of the powder containing the virus
are found lying in a kind of cavity full of almost clear fluid.
The walls of this cavity are lined by a thin, greyish, and
fibrinous layer. The muscles surrounding the cavity are
highly cedematous, so that on cutting through them a large
quantity (10 e.c. to 20 c.c. or more) of clear, transparent,
reddish fluid may be collected.

If twelve hours after the inoculation a sharp capillary pipette
be introduced into the centre of the tumour already formed,
and a little of the transparent fluid be carefully drawn off and
examined under the microscope, an enormous number of bacilli
will be found moving about in the liquid. Here and there a
leucocyte is seen. As a rule, the wandering cells met with at
this stage are empty, but occasionally one holds two, three, or
more bacilli in its interior.

In cover-glass preparations stained in an alkaline solution
of methyl-blue, or better, gentian-violet, the bacilli floating in
the liquid are, to all appearances, perfectly normal, and,
according to the reagent used, stain of a lovely dark blue or
purple colour. Most of the bacilli contained in the cells stain
normally, but here and there an intercellular bacillus shows
signs of degeneration.

A drop of the liquid from the tumour withdrawn and ex-
amined at the end of forty-eight hours, or immediately after
the animal’s death, contains an enormous number of free

IMMUNITY AGAINST MICROBES. 421

bacilli. The fluid, as a rule, is somewhat turbid, owing to the
presence of wandering cells in fairly large numbers. These
ameeboid cells belong to the small variety (microphages), and
are either uni- or multi-nucleated. The nuclei possess hardly
any intra-nuclear network, and are surrounded by a small
amount of protoplasm only. Many of these cells contain
bacilli, some in enormous numbers, as many as ten micro-
organisms being occasionally enclosed in one cell. The micro-
organisms in the liquid show no signs of degeneration, whilst
some of the bacilli contained in the ameeboid cells have clearly
undergone a process of digestion. Floating in the liquid are
seen remnants of the dried blood used for the purpose of
inoculation.

On microscopical examination the free aspect of the abscess-
wall is covered everywhere by an enormous number of bacilli,
lying free in the coagulated exudation fluid, staining in a
perfectly normal manner, and showing to all appearances no
traces of degeneration.

The abscess-wall itself consists of an innumerable number of
small migrating cells. Some of these amceboid cells have but
one single nucleus ; others contain two, three, or four nuclei.
In many the nucleus has the triradiate aspect found in other
inflammatory conditions—for example, pneumonia. The
amount of protoplasm round the nucleus is very small as a
rule. Close to the free surface of the abscess-wall these cells
are pressed together in enormous numbers, and so closely
packed that the contours of each are scarcely to be recognised.

In the deeper regions of the abscess-wall the cells are massed
together less closely. The muscular tissue around contains
within its meshes and between its fibres a large number of
small round migrating cells. The latter are met with even in
places where not a single bacillus is to be seen. Here and
there, especially in the deeper strata of the abscess-wall, a few
cells occur which are larger, have a single, clear, vesicular
nucleus, with a well-marked intra-nuclear network, surrounded
by a large amount of coarse protoplasm, and not infrequently
contain in their interior remnants of degenerated bacilli and

422 M, ARMAND RUFFER.

leucocytes. These cells probably represent the first stage in
the development of macrophages; they do not attain their full
size owing to the early death of the animal. On the whole,
macrophages play quite a secondary part at this stage.

Many leucocytes found close to the free surface of the
abscess-wall contain in their interior a large number of charac-
teristic bacilli. Nevertheless, many micro-organisms remain
quite free between the cells in the surrounding medium, which
has been coagulated by the action of the hardening reagent ;
but whereas the latter micro-organisms do not show signs of
degeneration, many of those contained in the migrating cells
have undoubtedly undergone a process of disintegration. The
number of micro-organisms in the deeper layers of the abscess-
wall, at some distance from the free surface, gradually becomes
smaller and smaller ; and while in the upper strata some micro-
organisms are free between the cells, the bacilli in the lower
strata of the abscess-wall are always contained in the interior
of microphages. Lastly, a few millimetres away from the free
surface, although the number of migrating cells is still very
large, no bacilli are to be seen except occasionally the remnant
of a dead micro-organism contained in an ameeboid cell.

It will be convenient to speak now of the forms of degenera-
tion as seen in the micro-organisms of quarter-evil, when they
have been taken into the interior of lymphocytes. I will
describe these changes as seen in preparations examined with
a Vérick microscope, oil immersion -/;th or -1;th, ocular 1 or 3,
and Abbé’s condenser. The bacilli not contained in the cells
I shall, for brevity’s sake, call extra-cellular micro-organisms,
whilst the bacilli in the interior of leucocytes must go by the
name of intra-cellular micro-organisms.

The extra-cellular micro-organisms present in the coagulated
exuded fluid, when carefully examined, consist of rods some-
what shorter and thicker than anthrax bacilli. Spores are not
often present when the virus is inoculated into guinea-pigs.
The bacilli stain of a deep blue colour with methyl-blue, in-
tensely red with fuchsin, and purple with gentian-violet.

Though staining uniformly, the extra-cellular bacilli are not

IMMUNITY AGAINST MICROBES. 423

all of the same length and thickness; these differences in size
being due partly to an actual difference in size, partly to two
or more being joined together, and possibly to the fact that
some are placed more or less obliquely in the preparation.
None show any signs of degeneration whatever.

The appearance of the intra-cellular bacilli is very different.
A large number are, to all appearances, quite healthy, stain uni-
formly, and are of normal size or shape. Here and there, how-
ever, an intra-cellular bacillus is found which, instead of stain-
ing as deeply as the others, assumes a light purpletinge. This
tint may be perfectly uniform, but at other times part only of
the bacillus is stained of a dark purple colour, whilst other
parts of the same micro-organism assume a lighter shade, and
the most varied forms of degeneration can be observed.

Sometimes the centre of the bacillus takes up the colouring
matter badly, whilst the periphery still stains deeply. The
micro-organism then consists of a central, badly staining core,
and a dark, deeply staining sheath (see figs. B 3, 6, B 5, c).
Sometimes one edge only of the micro-organism retains the
colouring matter deeply, whilst the remainder assumes a light
purple colour (fig. B 5, 6). In another stage the whole
bacillus is of a uniform pale purple hue, owing to the gradual
breaking up and disappearance of the colouring matter in the
sheath.

In other bacilli the degeneration, marked by the loss of
power of fixing the colouring substance, begins at the peri-
phery of the micro-organism, the centre still retaining the
colouring matter, while the edges remain unstained. The
central coloured part becomes less and less in amount, so
that at last a small streak of it only is left. Later on the
central part itself breaks up, and forms a very thin, inter-
rupted, irregular streak lying in the interior of a bacillus.

The process of degeneration in other bacilli affects the whole
breadth of the bacillus uniformly. One end of the bacillus
stains normally, whilst the other is of a pale homogeneous
colour (see fig. B 4, 6). Sometimes an intra-cellular bacillus
consists of a row of dots staining darkly, and embedded in a

424, M. ARMAND RUFFER.

less darkly staining material (fig. A, a), or of dark and light
parts alternately (see fig. B 3, a).

At the same time that these changes take place in the colour-
ing of the bacilli, the latter undergo alterations in shape and
size. Whereas the bacilli present in the inflammatory fluid
are usually straight, and very rarely bent, many of the intra-
cellular bacilli are curved on themselves (see figs. A and B).
This is better seen in sections made at the point of injection
in animals inoculated with a weak virus, in which, as we have
seen, the bacilli have a tendency to join together end to end.
These intra-cellular rods are nearly always curved, and present
a highly characteristic appearance (figs. A, B).

The intra-cellular bacilli in later stages of degeneration show
a diminution in thickness. Many are much thinner than
normal (fig. B 2, a, 6); and while the diminution of thickness
in some bacilli is uniform, in others it begins at one extremity,
the bacillus looking as if one end of it were being eaten away
(fig. B 3, ¢). The contours of the intra-cellular micro-
organisms, also, instead of being sharply defined as in the
extra-cellular bacilli, are irregular, and their exact outline
is by no means easily traced out (see figs. A and B 3, a).
In a later stage the bacilli become thinner and thinner,
more and more irregular (fig. B 2, a, 0), and lose whatever
power they still possessed of retaining colouring matter, so
that finally nothing is left in the cells but small dots (figs.
A, c,d,e, and B 1, a), some of which still retain more or
- less colouring matter, whilst others appear as light, highly
refracting granules, or very pale rods, looking like the remnants
of the sheaths of dead bacilli.

Occasionally intra-cellular bacilli are met with which, instead
of taking up aniline dyes, stain with carmine or logwood.
Others stain partly with carmine and partly with aniline dyes,
looking like little dots stained purple and red alternately.

When a small dose of a weak virus is inoculated subcu-
taneously, the struggle between leucocytes and bacilli is
evident on the fifth day, or even later; but the number of
micro-organisms gradually diminishes, so that after the fourth

IMMUNITY AGAINST MICROBES. 425

day hardly any bacilli are found at the seat of inoculation.
On the fifth day the few bacilli which can still be seen are con-
tained in migrating cells, and show evident signs of the most
advanced degeneration.

The study of the initial lesion of animals in which a chronic
form of the disease has been induced by the inoculation of large
quantities of a weak virus (0°40 centigramme or more) is not
less interesting.

The animals inoculated with a strong virus perish so soon
after the introduction, that the struggle between the cells and
the micro-organisms is more or less one-sided; for although
the animal dies, most of the cells present at the point of
inoculation are normal and show no signs of degeneration.
When, however, a chronic form of the disease is produced,
many of the lymphocytes perish as the result of their struggle
with the invading bacilli.

If the animal dies on the fourth or fifth day, sections through
the exact spot where the virus has been inoculated show that
the bacilli have infiltrated the neighbouring muscles to a far
greater extent than in the acute form of the disease. Many
bacilli are extra-cellular, lie in a coagulated inflammatory
effusion, and are perfectly normal and healthy, staining well
with aniline dyes, and retaining colouring matters in a normal
fashion. A few micro-organisms are contained within ameboid
cells, and often present various forms of degeneration; but
although in the acute form of the disease most of the wander-
ing cells are healthy, many of the cells met with in the more
chronic affection are markedly degenerated. In other words,
they become true pus-cells—that is, dead cells.

The nucleus of such a cell, instead of staining deeply with
carmine or logwood, possessing a coarse intra-nuclear network,
and being marked off sharply from the surrounding protoplasm,
stains rather more diffusely, and often shows signs of breaking
up. In later stages the nucleus undergoes distinct fragmenta-
tion, three or four fragments of nuclei lying in the protoplasm
of a cell. In the last stage the nucleus disappears, nothing
remaining but a pale round mass of protoplasm, which no

426 M. ARMAND RUFFER.

longer takes up colouring matter. Not infrequently one of
these cells is contained in a larger cell, possessing a clear vesi-
cular nucleus, which is surrounded by a large amount of some-
what coarse protoplasm; in other words, it is taken into the
interior of a typical macrophage.

Some of the dead lymphocytes contain bacilli which some-
times appear healthy, but not infrequently are more or less
degenerated. It might be supposed that these cells died owing
to the entrance of bacilli into their protoplasm ; many dege-
nerated cells, however, are found in which not a single living
or dead bacillus is to be seen. These cells probably never
contained bacilli, but perished as the result of the toxic influ-
ence of the poisons secreted by the micro-organisms surround-
ing them. In the chronic form of the disease, therefore, it is
possible to follow the struggle of the bacilli and the lympho-
cytes, a struggle in which sometimes the former and sometimes
the latter perish. It is a noticeable fact that although the
animal dies from the inoculation, no micro-organisms penetrate
into the tissues ; the bacilli have all been arrested at the point
of inoculation, and this arrest is evidently due to the barrier
formed by the ameeboid cells at the seat of inoculation.

Similar facts have been observed in the erysipelas of man, or
when the disease is artificially produced in animals by inocu-
lation.

The part of the skin affected with erysipelas may be divided
into three zones. The first or peripheral zone, although show-
ing no trace of redness, nevertheless contains numerous
streptococci lodged in the lymphatic vessels. The second zone
looks rather inflamed, and contains many migrating cells, the
greater number of which are filled with streptococci. Finally,
the third or innermost zone is crowded with small round-cells,
and contains no streptococci.

The following facts were observed on seven cases of erysipelas,
Two of these cases proved fatal, and in them numerous strepto-
cocci were found in the lymphatic vessels, in the skin, and in
the layers immediately beneath, but always outside the vessels.

IMMUNITY AGAINST MICROBES. 42.7

The cocci were occasionally single, more often they formed a
more or less lengthy chain. Innumerable leucocytes had
emigrated to the spot, whilst the fixed cells of the connective
tissue showed signs of proliferation.

The microscopic appearances were very different in the cases
which ended in recovery, for many of the migrating cells were
absolutely crammed with micro-organisms, forming a mass
surrounded by a clear vacuole in the interior of the leucocyte.
The nucleus of such leucocytes varied greatly in size, and was
irregular in shape and not easy to stain. The leucocytes were
far less numerous where the skin had become gangrenous, and
nearly all the streptococci were then outside the cells.

Other ameeboid cells larger than leucocytes were also present.
Their nuclei, instead of staining deeply with methylene blue,
took the stain badly, and they further differed from those of
leucocytes by their oval and irregular contours, and by their
possessing very distinct nucleoli. These cells also belonged to
the group of phagocytes.

The same phenomena may be noticed in white mice inocu-
lated with erysipelas. If little cylinders of elder-pith, previously
soaked in a pure culture of streptococci, be placed under theskin
_ of such animals, the foreign body very soon becomes surrounded
by a mass of leucocytes. The latter are so full of strepto-
cocci that the cells stand out as black dots in sections stained
with gentian-violet. Two days afterwards almost all the
streptococci have disappeared. But in the mouse, as in man,
not a single one is to be found in the macrophages. The micro-
organisms enclosed in the leucocytes gradually lose their
regular outlines, break up and disappear, and when stained
with methylene blue, may assume any shade of colour, from an
intense blue to a reddish purple or pale violet.

Another series of researches, made with the bacillus of
“ rouget des porcs ” or swine fever,! has yielded similar results.
According to Emmerich and Mattei,’ the bacilli of swine fever

1 B. Metschnikoff, “ Etudes sur l’Immunité,” ‘Annales de l'Institut Pasteur,’
June, 1889, p. 289.
2 Emmerich and di Mattei, ‘ Vienna Congress,’ 1888,

428 M. ARMAND RUFFER.

injected into rabbits previously rendered immune against this
disease are completely destroyed within fifteen to twenty-five
minutes after their introduction, even if the quantity of culture
injected be very considerable. If this be correct, it is impos-
sible to ascribe the destruction of micro-organisms to the
action of phagocytes ; for, in order to exert their action, phago-
cytes must first accumulate round the injected culture, then
take the bacilli into their interior, and finally destroy them.
According to these authors! this destruction is accomplished
by an antiseptic liquid which is not produced by the bacilli,
but by the cells of the immune organism.

Metschnikoff, however, who controlled the facts suet
by Emmerich and di Mattei, has obtained very different results
from those published by these observers.

Metschnikoff inoculated rabbits with the bacilli of swine
fever, and after a lapse of time, varying from half an hour to
six days, removed with a capillary pipette one or several drops
of liquid which had accumulated at the point of inoculation.
This liquid was sometimes quite transparent, but more often
was slightly stained with blood, and in several cases consisted
of almost pure blood.

In four cases the liquid taken from the point of inoculation
62, 17, 19, and 26 hours after the operation, contained no
micro-organisms. In eleven other cases, in which the liquid
was withdrawn 13, 4, 5, 6, 6%, 19, 20, 24 hours, and 4 days
after the inoculation, the beef-broth into which it was placed
gave pure cultures of the bacilli of swine fever, which, when
inoculated into pigeons and mice, always proved exceedingly
virulent. The liquid collected at the point of inoculation 4,
19, 20, and 24 hours after the injection of the first vaccin into
both immune and non-immune rabbits, contained bacilli of
the first vaccin. The liquid taken 19 hours after the in-
oculation of two animals, which had been rendered immune
against swine fever by repeated injections, gave a culture ;
whilst a third sample, taken 20 hours afterwards, remained
sterile.

1 ¢ Fortschritte der Medicin,’ 1888, T. vi, p. 729.

IMMUNITY AGAINST MICROBES. 429

Further, Metschnikoff proved that the bacilli of the first
vaccin of swine fever live a long time in the anterior chamber
of the eye of rabbits, as well as in the eye of dogs, which are
naturally immune against swine fever.

One rabbit, after having been rendered immune by seven
injections of the virus, was finally inoculated into the anterior
chamber of the eye with the first vaccin. In spite of the previous
inoculations, a large number of leucocytes emigrated into the
anterior chamber. A drop of aqueous humour examined twenty-
two hours after the operation gave a pure culture of the first
vaccin, whilst another drop, taken after forty-eight hours,
contained no bacilli. The bacilli of the first vaccin were
generally contained in the interior of leucocytes which had
found their way into the anterior chamber.

A small case composed of four cover-glasses joined together
with sealing-wax, and filled with the virus of swine fever, was
introduced under the skin of a rabbit. One hour after its
introduction the case was already full of leucocytes contain-
ing bacilli, which stained quite as well as those lying in the
exudation fluid ; on cover-glasses withdrawn two and a half
hours after their introduction under the skin of a rabbit, which
had resisted six former injections of the virus, the number of
leucocytes containing bacilli in their interior was consider-
able. In all M. Metschnikoff’s experiments with cover-
glasses the bacilli were very quickly taken into the interior
of wandering cells, which gradually became more and more
numerous at the point of inoculation.

A glass-case was filled with the virus of swine fever and left
for fifty-one hours under the skin of animmune rabbit. It was
then found that almost all the bacilli were contained in the
interior of cells and showed marked signs of degeneration ;
their outlines had lost their customary clearness, stained
badly, and their contents had become granular. On the other
hand, it was found that in glass-cases placed under the skin
of non-immune rabbits the greater number of the bacilli,
although surrounded by phagocytes, appeared perfectly normal
and stained well,

430 M. ARMAND RUFFER.

In order to see exactly what does happen when micro-
organisms are inoculated into a living fluid containing no
cells, Metschnikoff! introduced the spores and bacilli of
anthrax into the anterior chamber of the eye of living animals.
In this manner he proved that even in animals like frogs,
which are naturally immune, or animals rendered artificially
immune, e. g. sheep, the aqueous humour is a very good culti-
vating material for anthrax. The micro-organisms (even those
of the first vaccin) inoculated into the anterior chamber of the
eye of immune sheep, grow and multiply rapidly until arrested
by leucocytes, whilst the same micro-organisms inoculated into
the subcutaneous tissue of the same animal do not multiply.
The normal aqueous humour of immune animals, therefore,
contains no substances harmful to the bacillus of anthrax.

Anthrax spores inoculated into the anterior chamber of the
eye of rabbits multiply abundantly, and cause the death of the
animal, In order to isolate the virus from cellular influences
it may be placed in small cylinders of elder-pith, or in a small
bag prepared from the intestines of frogs, or more simply
still, in filter-paper. The spores thus protected, when intro-
duced under the skin of frogs, grow into normal bacilli, whilst
the unprotected germs are devoured by ameeboid cells. On in-
troducing under the skin of frogs membranous bags containing
blood of guinea-pigs which had succumbed to anthrax, the
bacilli grew abundantly and preserved their virulence until
the sixth day at least; it is clear, therefore, that the living
fluids of frogs have no harmful influence on anthrax. Con-
versely, the living unprotected bacilli are eaten by the cells of
frogs.

Very interesting also are the facts observed when anthrax is
inoculated into rats. Young white rats ® succumb to anthrax
much more readily than adult ones, and by passing the virus

1 BH. Metschnikoff, ‘Sur la Maniére d’étre des Bactéries Charbonneuses
dans l’Organisme,” ‘ Virchow’s Archiv,’ Dec., 1888 (‘Annales,’ Jan., 1889,
p- 41).

2B. Metschnikoff, “Etudes sur l’Immunité,” ‘Annales de I’Institut
Pasteur,’ April, 1890, p. 199.

IMMUNITY AGAINST MICROBES. 431

through several rats it is possible to obtain an exceedingly
powerful virus, which proves fatal to very old rats in three to
six days after inoculation.

It happens occasionally that the bacilli of anthrax will not
grow in vitro in the blood of rats which have actually died of
anthrax; but, as a general rule, the spores of bacilli thrive in
the blood of rats, which, before being killed, proved immune
against anthrax. The bacilli of anthrax grew abundantly and
produced spores even in the aqueous humour of rats which
had survived an inoculation of anthrax. The same holds true
for the subcutaneous tissue, which often contains numerous
leucocytes.

Spores of anthrax inoculated into rats always germinate and
produce bacilli, no matter whether the inoculation be performed
into the anterior chamber of the eye or under the skin. One
such experiment is extremely interesting, aud may be related
here. A white rat was examined four days after having been
inoculated subcutaneously with the blood of a guinea-pig dead
of anthrax; there was considerable cedema at the point of
inoculation, but the exudation fluid contained degenerated
bacilli only. Several little silk threads loaded with aathrax
spores were introduced into the cedema, and twelve hours after-
wards a considerable number of young and perfectly normal
bacilli were found in the fluid. Eighteen hours after the in-
troduction of the threads one was withdrawn, and found to be
surrounded by an immense number of bacilli, staining easily,
and perfectly normal in every respect. The rat survived the
two inoculations, and only died three months after from the
effects of an inoculation in the eye, with moderately virulent
anthrax.

The inoculation of anthrax into the most resistent rats is
usually followed by an abundant growth of anthrax bacilli ;
but after repeated inoculations on the same animal the latter
becomes so perfectly immune that finally the spores do not
seem to grow at all. M. Metschnikoff inoculated an old rat
(female) which had withstood three previous injections of an-
thrax, by introducing under the skin of the back a silk thread

432 M. ARMAND RUFFER.

laden with spores, and wrapped in a little piece of sterilised
cotton-wool, another similar thread being placed on the cotton-
wool without being surrounded by the latter. Nineteen hours
afterwards a semi-transparent liquid exudation had gathered
round the threads, and a few short but normal bacilli could be
distinguished init. The thread lying in the cotton-wool was
surrounded by far more anthrax bacilli than its fellow outside
thread. These prove, then, that anthrax bacilli find a medium
favorable to their growth even in animals rendered immune by
previous inoculations, and that they grow best when protected
against the attacks of ameeboid cells. Further, bacilli of the first
vaccin may develop in the organism of rats which have resisted
virulent anthrax—ain the anterior chamber of the eye of a white
rat one month and a half after its recovery from virulent
anthrax, for instance.

Some of the leucocytes of white rats inoculated with anthrax
contain no, or very few, bacilli; whilst others devour them in
such numbers that it is often difficult to distinguish individual
bacilli. These greedy phagocytes are with but few exceptions
multinucleated microphages. The bacilli contained in these
cells often stain easily, and appear normal; but, on the other
hand, more or less degenerated micro-organisms are often met
with.

When filled with bacilli phagocytes burst with extraordinary
facility, and in every field the observer meets with cells which
have allowed part of their contents to escape. The presence
of free and degenerated bacilli in the exudation of white rats
inoculated with anthrax, which at once strikes the observer,
can thus be readily explained. Metschnikoff, however, does
not think that all the bacilli found dead outside the cells come
without exception from the interior of phagocytes, for, in the
most favorable cultivating media even, a certain number of
microbes die, either as a result of the sudden transportation
into a new ground, or from some similar cause.

During the first two days immediately following the inocula-
tion of anthrax into the anterior chamber of the eye of white
rats the larger number of the bacilli are found outside the cells ;

IMMUNITY AGAINST MICROBES. 438

but on the third day, should the rat recover, all of them almost
are taken into the interior of leucocytes. Amongst the intra-
cellular bacilli a great many for a time retain the property of
absorbing aniline dyes; others lose this property, and only
stain very slightly. Many of the leucocytes, however, perish,
as is proved by the granular appearance of their nuclei. Besides
the débris of the nuclei of these phagocytes, more or less de-
generated extra-cellular bacilli are sometimes found, which at
some time were undoubtedly in the interior of leucocytes. The
exudation from the anterior chamber of the eye also contains
a more or less large number of macrophages, which, however,
do not as a rule destroy many bacilli, but often enclose a large
number of leucocytes.

The following experiment shows that the bacilli are taken
into the interior of leucocytes whilst still in the living state.
A drop of exudation was removed from the eye of a rat iocu-
lated with anthrax after the death of the animal, and at atime
when the phagocytes contained a great number of rods; and
after ascertaining that all the leucocytes were really dead a little
broth was added to the exudation, and the large drop thus pro-
duced placed in the warm chamber. Some hours later it was
plain that a certain number of the bacilli were alive in the in-
terior of the leucocyte, for they had multiplied and lengthened
into filaments. A drop of this exudation introduced under
the skin of guinea-pigs produced fatal anthrax in these
animals.

Within the tissues and internal organs of white rats anthrax
bacilli are very soon taken into the interior of microphages, or,
more frequently, macrophages, e. g. the cells of the splenic
pulp, the star-shaped cell of Kupffer, and the mononuclear leu-
cocytes found in the capillaries of the liver. As the macro-
phages get more and more filled with bacilli the vacuoles
increase in number, and the protoplasm at last becomes a mere
sac with thin walls, filled with a transparent substance, evi-
dently formed by the secretions of the phagocytes and of the
parasite. The nucleus, which keeps its characteris tic shape
is found in the thickest part of the sac.

VOL, XXXII, PART IV,—NEW SER. GG

434. M. ARMAND RUFFER.

The macrophages filled with bacilli and cells, swollen in
consequence of the formation of large vacuoles, finally burst
and allow their contents to escape. The bacilli, freed in this
way, often show signs of degeneration. Thus one meets with
free bacilli, segments of which stain violet and pink alternately.

Confirmatory evidence has been obtained by Metschnikoff
by studying the processes taking place when pigeons are
inoculated with anthrax. This animal is immune against the
disease, and Baumgarten! and Czaplewski state that anthrax
bacilli introduced under the skin of pigeons perish sponta-
neously at the end of four hours, the phagocytes taking no part
in their destruction. Lubarsch also came to the same conclusion.

Nuttall, on the other hand, thought that the blood of
pigeons is a medium in which the bacilli of anthrax begin to
grow almost immediately after their introduction into it.

Metschnikoff* then proved that anthrax does not become
attenuated in pigeons, but often acquires new virulence.

The aqueous humour of pigeons introduced into capillary
tubes and inoculated with spores of anthrax proves a most
satisfactory cultivating medium, the spores giving rise to an
abundant crop of felted threads, which are perfectly normal
in appearance and ultimately produce innumerable spores.
The same result is obtained when anthrax is inoculated into
the aqueous humour of the eye of a pigeon which has been
rendered doubly immune by a previous inoculation of anthrax
from which it has recovered.

The same result can be obtained by inoculating the exudation
fluid with an exceedingly small quantity of an old culture of
the first vaccin of anthrax. The day after the inoculation the
whole drop is full of an abundant culture of normal anthrax
filaments.

The following experiment is extremely interesting :—A small
piece of silk thread loaded with spores was introduced into the
anterior chamber of the eye of an immune pigeon inoculated

1 «Centralblatt fiir klinische Medicin,’ 1888, No. 29, p. 516.

2 Metschnikoff, “Etudes sur ’Immunité: Le Charbon des Pigeons,”
‘ Annales de l’Institut Pasteur,’ 1890, vol. iv, p. 65.

IMMUNITY AGAINST MICROBES. 435

two days previously with spores of anthrax in the same eye.
Before introducing the thread it was ascertained that the
aqueous humour of the eye contained a considerable number
of leucocytes, but no free bacilli; the day after, on withdraw-
ing the thread, it was seen that a certain number of spores had
grown into bacilli composed of several (as many as six) seg-
ments. In another similar experiment the piece of silk thread
was introduced five days after the first inoculation, and ata
time when the bacilli had already entirely disappeared. Twenty-
four hours afterwards newly grown bacilli, showing as many
as seven segments, were found in the exudation from the eye.
The bacilli took a dark blue colour with methylene-blue, and
were quite normal.

Repeated subcutaneous inoculations into pigeons which had
already recovered from one attack of anthrax no longer gave
rise to serious symptoms. Nevertheless the bacilli introduced
in the state of spores fixed on silk threads grew more or less
abundantly.

Four hours only after the subcutaneous inoculation of very
virulent anthrax into pigeons a marked emigration of micro-
phages took place at the point of inoculation, and some of
these contained one or more and as many as nine bacilli.

After atime the inflammation increased, and a larger number
of phagocytes—micro- and macro-phages—accumulated at the
point of inoculation. Many phagocytes, especially those which
were crammed with bacilli, burst very easily, allowing their
contents, including micro-organisms in every stage of de-
generation, to escape. This bursting took place not as a
consequence of the mode of preparation, but also in the living
animal—as was proved by the degenerated form of some
phagocytes containing bacilli.

In pigeons which recovered from the disease the larger num-
ber of the bacilli were found in the interior of the phagocytes,
whilst in the birds which perished the majority of microbes lay
outside the cells, although in all, the ameboid cells devoured a
large number of micro-organisms. So full were the microphages
sometimes that their nuclei could hardly be made out, and when

436 M. ARMAND RUFFER.

in that state they might easily be mistaken for clusters of
bacilli.

Anthrax does not become less virulent by passing through
pigeons. Thus, a guinea-pig contracted anthrax when inocu-
lated with virus which had remained six days under the skin of
an immune pigeon; and a large rabbit died after being inocu-
lated with a drop of exudation extracted twenty-two hours
after the introduction of anthrax blood into the eye of a pigeon
which had undergone four previous inoculations. A guinea-pig
received one drop of exudation taken twenty-six hours after the
inoculation of anthrax into the eye of a pigeon ; it is noteworthy
that the number of free bacilli in this exudation fluid was already
greatly diminished, and that the leucocytes contained masses
of bacilli; nevertheless the guinea-pig contracted the disease,
and died from it sixty hours after the inoculation.

The following beautiful experiment shows that the cells take
into their interior living micro-organisms. <A drop of exuda-
tion fluid was removed from the anterior chamber of the eye of
an animal inoculated with anthrax. This, being found to con-
tain numerous cells enclosing bacilli, was mixed with a very
small quantity of beef broth, and the whole was placed in a
warm chamber. The phagocytes died at once, but the bacilli
continued to grow; and by placing under the microscope some
cells enclosing bacilli, the development of the latter could be
easily followed.

After three hours’ stay in the warm chamber it was easily
ascertained that the bacilli contained in several of the phago-
cytes had already begun growing. ‘Three of these phago-
cytes were isolated by a difficult but very neat process, and
thrown into beef broth; the development of these bacilli was
carefully watched, and it was ascertained that they were alive
and active. At the end of eight hours the bacilli in the drop
gave a culture containing filaments, and the next morning an
abundant pure growth had developed. A white mouse, a young
guinea-pig, and two full-grown rabbits were inoculated with
this culture ; the mouse died in twenty hours, and the guinea-
pig twenty-four hours after the inoculation, while one of the

IMMUNITY AGAINST MICROBES. 437

rabbits died in four days and the other in less than three. The
four animals showed the characteristic signs of anthrax (bacilli
in the blood and organs, hypertrophy of the spleen, edema of
the skin, in the rabbit and guinea-pig, &c.).

Similar facts! have been observed in fowls when these animals
which are immune against anthrax are inoculated with this
virus. When, however, these animals are placed in cold water
they readily die when inoculated with anthrax, and Wagner
has shown that the phagocytes in the latter case are unable to
cope with the bacilli, though they may make an attempt to do so.

I have shown elsewhere that in diphtheria of man a struggle
takes place in the diphtheritic membrane between the leuco-
cytes and the specific microbes, and that the disease is localised
in the upper part of the respiratory tract owing to the action of
the former. In thin sections passing through the entire false
membrane and the surrounding structures—a section of a small
bronchus, for instance, when the disease has extended into the
lungs—the most superficial part of the membrane, that is the
free edge of the lumen of the tube, is found to contain an enor-
mous number of specific bacilli, lying, for the most part, quite .
free in the exudation fluid (fig. C). The micro-organisms re-
semble short rods of the size of tubercle bacilli, and are often
aggregated together in clusters containing hundreds of microbes.
So thick is this layer of micro-organisms at times that on holding
the sections to the light a thin violet line is seen running along
the inner margin of the section tube, this line representing the
layer of microbes (fig. E, a). Scattered among these specific
bacilli, more especially in the upper air-passages, other kinds
of microbes—micrococci and bacilli of various sizes and shapes
—are also met with.

If the false membrane immediately below the superficial
layer just described be now examined, the number of micro-
organisms is found to be exceedingly small (fig. E, 6); and it
will be noticed that not a single bacillus is to be seen in the
deeper layers of the false membrane, in the mucous membrane,

1 Wagner, “ Le Charbon des Poules,” ‘ Annales de l'Institut Pasteur, Sept.,
1890, p. 578.

438 M. ARMAND RUFTER.

or in the deep layers of the tissue. The whole process, there-
fore, as far as the bacilli are concerned, takes place on the
surface of the false membrane and nowhere else.

Shortly stated, the false membrane consists of a reticulated
network of fibrin, containing within its meshes a large number
of wandering cells, some of which are healthy, whilst others
have undergone a distinct process of degeneration. I must draw
attention to the fact, however, that the wandering cells present
in the membrane are of two kinds; some smaller and mono- or
multi-nucleated (microphages),and others, less numerous, much
larger, with a single, clear, vesicular nucleus (macrophages).

Many of the leucocytes which have penetrated between the
bacilli are quite empty; but others, especially those in the
layer of the false membrane close to the thick layer of bacilli
previously described, contain one, two, or more diphtheritic
bacilli (fig. C, a, 6), as many as five or six micro-organisms
being sometimes enclosed in one cell. Some of these intra-
cellular bacilli appear to be healthy, staining and retaining
colouring matters well; whilst other micro-organisms also
contained in amceboid cells show sigus of degeneration, varying
from a mere difference in the power of retaining gentian-
violet to complete disorganisation and destruction (see fig.
Bis! Ay CA he 5f419)s

Many ameeboid cells in the membrane, more especially
those present in its superficial part, show distinct necrotic
changes, many of them losing the power of retaining colouring
matter. The nucleus breaks up, and does not stain at all,
even with strong logwood or carmine, the surrounding proto-
plasm swelling up, becoming more yellow, and not unfre-
quently vacuolated. In later stages they show distinct signs
of breaking up. Many of the degenerated cells contain bacilli
in their interior, whereas others are quite empty.

Those lying between the cells in the fibrin and coagulated
exudation fluid are, for the most part, quite normal to all
appearance. Occasionally, though very rarely, one is met
with presenting signs of degeneration similar to those found
in the bacilli contained in ameeboid cells. I do not think it

IMMUNITY AGAINST MICROBES. 439

justifiable to argue from this fact that the inflammatory liquids
present in the membrane possess a microbe-killing power, but,
judging from the number of dead leucocytes present in the
membrane, it is far more probable that these partially digested
microbes were originally contained in a leucocyte which had
died and undergone liquefaction.

Similar, but far more interesting, facts are noticed when
we examine the processes taking place in chronic infectious
diseases, such as tubercle and actinomycosis. Shortly stated,
a nodule of tubercle or actinomycosis contains, besides the
specific parasites, three kinds of ameeboid cells: Ist, small uni-
or multi-nucleated cells; 2nd, large mononucleated or epi-
thelioid cells ; and 8rd, multinucleated giant-cells.

The development of giant-cells has been well studied in the
Spermophilus guttatus by Metschnikoff.2 When a Sper-
mophilus guttatus is inoculated with a pure culture of the
tubercle bacillus no tuberculous formations are found in any
of its organs after death, but microscopical examination of the
liver, spleen, and lymphatic glands shows these organs to be
crammed with giant-cells, the development of which can be
followed with ease.

These cells are derived from isolated epithelioid cells in
the following manner:—The nucleus of an epithelioid cell
assumes a star-shaped appearance, and each of the rays so
produced is ultimately converted into one of the nuclei of the
giant-cells. A swelling then forms at the extremity of each
ray, which at first appears to be homogeneous; but later on
becomes filled by a transparent mass with irregular outlines,
the mass resembling a new nucleus. The chromatin now
slowly divides into a peripheral and a central part, and in this
way one or more nuclei are formed which are connected by a
thread only with the remainder of the old star-shaped nucleus.
These masses, which in the first stage are extremely irregular
in form and outline, finally become detached and assume the

1 I intend to discuss Baumgarten’s and Weigert’s views in another part of
this paper.

2 Metschnikoff, ‘ Virchow’s Archiv,’ vol. exili, 1888.

440 M. ARMAND RUFFER.

round or oval form of typical nuclei. During their formation
the protoplasm of the epithelioid cell increases in size and
takes the characteristic dimensions of a giant-cell. This process
is not actually caused by the presence of the tubercle bacilli,
for it occurs in empty cells as well as in cells containing bacilli.

Most of the bacilli contained in epithelioid cells appear to
be quite healthy, but in the giant-cells degenerated micro-
organisms are frequently met with. Many bacilli contained
in the giant-cells are of a faint rose colour, remain unstained,
or take up the blue colour only; whilst in a further stage of
degeneration they become surrounded by an aureole resembling
the clear space found around the pneumococcus of Friedlander.
The micro-organisms become paler, stain less deeply or may
even not stain at all, whilst their outlines become clearly
defined. Finally they seem to disappear, but outlines of the
surrounding yellowish capsule become more defined and
sausage-shaped. These sausage-shaped bodies join, fuse
together, lose their shape, and finally form an amber-coloured
mass, which in no way resembles the bacilli from which it is
derived. This process, therefore, is not a true digestive pro-
cess, since the cells convert the bacilli into a solid resistent
mass instead of liquefying them; it resembles rather the
encapsulation of some Infusoria.

Similar forms of degeneration are to be met with in the
giant-cells of tuberculous rabbits.!. But although the writer
has not unfrequently observed the formation of giant-cells by
indirect division of the nucleus of epithelioid cells, it is quite
clear that in the rabbit, guinea-pig, and man the giant-cells
arise from the fusion of two or more epithelioid cells without
any apparent transformation in the nucleus.

Lately fresh evidence has been accumulating to prove that
the function of giant-ceils formed in pathological new growths
is identical with that of giant-cells found in the normal healthy
body. Soudakewitch’ has shown that the giant-cells of lupus
in man feed on the neighbouring diseased tissue, and digest

1 Metschnikoff, loc, cit., and Armand Ruffer, unpublished observations.
2 Soudakewitch, ‘ Virchow’s Archiv,’ v. exv, p. 264.

IMMUNITY AGAINST MICROBES. 441

large quantities of elastic fibres. The writer! has proved that
in the spleen of tuberculous animals such cells not only destroy
the specific tubercle bacilli, but also devour an enormous
number of red blood-corpuscles, these cells being found to be
absolutely crammed with blood-pigment (figs. I, K).

I may here record an experiment which is extremely in-
teresting, as it illustrates the development and the functions
of giant-cells. Having inoculated a rabbit with 0:005 of first
vaccin of charbon symptomatique, enveloped in a paper bag,
I was prevented by a temporary illness from coming to the
laboratory. On the sixth day the animal appeared quite well ;
the paper bag, together with the surrounding tissue, was taken
out, hardened in absolute alcohol, and cut, and the sections
stained very deeply with carmine or M. Gerrard’s logwood and
Grain’s method. In all the sections which I examined from
various situations I was unable to find a single bacillus. The
cavity of the paper bag was still perfectly recognisable, although
it contained the débris of the powder used, and numerous
ameeboid cells. On the other hand, the outer side of the paper
was already permeated by young connective tissue which was
already vascularised.

I need not enter here into the description of the microphages
(small mononucleated or multinucleated cells), or of the
macrophages (large cells with a single clear vesicular nucleus).
Suffice it to say that, contrary to the opinion of Messrs.
Ballance and Sherrington, who were unable to find in their
experiments any intermediate forms between the leucocytes
and the plasma-cells, I maintain that every stage of develop-
ment can be shown between the leucocyte and the plasma-cell.
The formation of the plasma-cell in pathological effusions is
identical with the formation of similar cells which I have
described in normal tissues (spleen, lymphatic glands, Peyer’s
patch, tonsils, &c.).2_ Not only is their development the same,
but their functions are also identical ; for in the filter-paper
introduced subcutaneously, as well as in the normal spleen and

1 Armand Ruffer, ‘ British Medical Journal,’ 1890.
’ “On Formation of Scar Tissue,” ‘ Journal of Physiology,’ 1889, p. 558.

44,2 M. ARMAND RUFFER.

lymphatic glands, these cells were found to devour numbers
of small lymphoid cells. Lastly, all the evidence I could
gather seemed to point to the fact that the large epithelioid cells
have nothing whatever to do with the development of scar tissue.

In the filter-paper we find not only these cells, but also true
multinucleated giant-cells, resembling those of tubercle.
They are formed by the fusion of several epithelioid cells, and
it can be seen from the following facts that they have exactly
the same functions as epithelioid cells. On careful examination
many of them are found to contain peculiar glistening, homo-
geneous, irregular, yellowish bodies, which in no wise resemble
anything normally found in the animal organism. These
foreign bodies are nothing more than partially digested masses
of filter-paper which the giant-cells have absorbed. In some
cases, indeed, the cells have been fixed by the reagent at the
exact moment when they were absorbing the paper fibre; one
may then see one of these giant-cells almost completely sur-
rounding the fibre, which is still partly sticking out of the
cell. This new fact, together with those described by
Metschnikoff, Soudakewitch, and myself, absolutely proves that
the giant-cell is not a weak and diseased structure (Weigert,
Koch), but an extremely active and useful body—a fighting cell.

Appearances similar to those found in tubercle are easily
demonstrated in actinomycosis.

Professor Crookshank had already noticed the presence
of the peculiar parasite of actinomycosis in the interior of
giant-cells. More lately I have, thanks to the kindness of
my friends Lingard and Sims Woodhead, obtained a large
amount of material from cattle suffering from this disease ; and
though I must reserve details of the development of the fungi,
both in animals and man, for a future occasion, I may be allowed
here to draw attention to a few facts relating to the subject
now under consideration as observed in actinomycosis of cattle.

A nodule of actinomycosis, which has not yet undergone
caseation, consists of the central rosette formed by the parasite,
and a mass of amceboid cells of various sizes, the whole nodule
being surrounded by a strong capsule of connective tissue.

IMMUNITY AGAINST MICROBES. 443

The ameboid cells which enter into the formation of the
nodule are derived from small round mononucleated cells, which
exactly resemble ordinary leucocytes. The nucleus of such a
cell is round, stains deeply with hematoxylin or alum carmine,
and shows no trace of intra-nuclear network. The delicately
mottled protoplasm surrounding the nucleus varies somewhat
in amount, whilst the whole cell is sometimes quite round, and
occasionally slightly irregular in shape. I cannot say for
certain whether these small round cells are all derived from
emigrating leucocytes, or whether some are not derived from
pre-existing connective-tissue cells. Both modes of origin are
probably the rule, for at the periphery of the new growth—in
places where no parasites can be discovered—the connective-
tissue cells are often in a state of active proliferation. 'The
blood-vessels, their walls, and their immediate neighbourhood,
on the other hand, are frequently crammed with leucocytes.

It is quite easy to trace the development of epithelioid and
giant cells from leucocytes in the nodule of actinomycosis.
The nucleus of some of the small lymphocytes is occasionally
composed of a dark outer border, and a clear, but somewhat
swollen, centre (fig. M, a). In others the centre of the
nucleus becomes clearer, and the chromatin almost entirely
disappears in places, thus leaving a fine intra-nuclear network
possessing one or more nucleoli (fig. M, 6). Owing to the
gradual disappearance of the chromatin in the centre and
periphery, the nucleus becomes clear and bladder-like, and
possesses a beautiful intra-nuclear network (fig. M, ce); but
the protoplasm in this stage has as yet undergone no change, and
the whole cell may not be larger than an ordinary lymphocyte.

The nucleus now slowly becomes larger, the protoplasm
increases enormously in size, becomes coarser and vacuolated,
with extremely irregular contours, and the cell finally presents
the appearance characteristic of an epithelioid cell (fig. M,
d, e). The nucleus now gradually changes its place and
approaches one of the poles of the cell.

Just as in the healthy lymphoid tissues of animals the
larger ameeboid cells or macrophages take into their interior

44.4 M. ARMAND RUFFER.

and destroy the smaller ones, so do the macrophages met with
in actinomycosis destroy the microphages. It is easy to find
such macrophages containing one, two, or as many as five or
six microphages in various stages of degeneration (figs. L,
m,and M, h,j, k, 1; also fig. H). Several epithelioid cells may
then coalesce and form a giant-cell (fig. H, a, 6, c, d, e);
but I have never seen division of nuclei as in the epithelioid
cells of Spermophilus guttatus.

Near the growing edge of the tumour each rosette is sur-
rounded by a layer of epithelioid and giant cells, forming a
regular palisade around the nodule. The nuclei of such cells
are always placed at the pole situated away from the parasite.

The epithelioid cells in other places penetrate into the rosette
of actinomycosis and take into their interior huge bunches of.
these parasites (fig. F), which then undergo degeneration in the
interior of these cells. The giant-cells likewise often contain
masses of such parasites (fig. G). At the growing edge of the
tumour, however, giant-cells are few, and, as might be expected,
there is no formation of new connective tissue.

In the parts of the tumour which are older, and therefore
harder and more fibrous, the appearances are very different.
The periphery of each nodule is surrounded by a dense capsule
of fibrous tissue. The epithelioid cells, however, take no part
in the formation of this fibrous tissue, for in no case could I
see appearances justifying such an assumption. On the
contrary, the connective tissue is derived from small cells pos-
sessing a hard, darkly staining single nucleus. These cells
gradually elongate, the nucleus becomes oval and clearer at
the same time, until typical long spindle-shaped connective-
tissue cells are produced (fig. J).

The appearances of the disease in a fibrous part of the
tumour are very different from those seen near the growing
edge. The older parts generally contain a large number of
epithelioid cells and giant-cells, holding in their interior
bunches of actinomycosis, which often show signs of degenera-
tion. Instead of staining of a dark purple colour with gen-
tiau-violet the parasites do not stain at all, but become con-

IMMUNITY AGAINST MICROBES, 445

verted into a homogeneous, highly-refracting mass, consisting
of sheaths of the dead parasites. At the same time, however,
the giant-cells and epithelioid cells suffer in the fight, their
nuclei become granular, the protoplasm liquefies, and so finally
parts of the tumour consist of actinomycosis—few of which
show any signs of life—and masses of dead ameeboid cells.
In other parts everything is dead, and nothing is left but the
dead bodies of the parasites and pus-cells. Strangely enough
when this is the case, the lymphoid cells in the surrounding
parts do not appear to make the slightest attempt to take the
dead particles into their interior, but neglect them in favour
of the living leucocytes and fungi in their neighbourhood—a
proof, if any were wanting, that phagocytes do not eat up
everything they come across, but exert a distinct choice.

In actinomycosis, therefore, as in tubercle, the ameboid cells
which enter into the formation of the “‘ pathological granuloma”’
are fighting cells which actively destroy the invading parasites.

Although many facts have accumulated to give us some idea
as to what happens when micro-organisms are introduced into
the subcutaneous tissue, our knowledge of the processes fol-
lowing on their forcible introduction into the blood is still
extremely meagre.

According to Wyssokowitch! micro-organisms are not des-
troyed in the blood, but remain for a certain length of time in
the organs, more particularly in the lymphatic glands, the
medulla of bones, and the spleen. Wyssokowitch found them
in the endothelial cells of blood-vessels. Micro-organisms, ac-
cording to the same author, are not eliminated by the urine,
unless through some lesion of the kidney (diapedesis, rupture
of vessel, abscess, infarctus, &c.); Berlioz? has arrived at the
same conclusion. That this, however, is not an absolute rule
has been proved by Charrin® and myself.*

' Wyssokowitch, ‘ Zeitschrift f. Hygiene,’ 1886, T. i, Heft 1, p. 45.

* Berlioz, ‘ These de Paris,’ 1888.

3 Charrin, ‘La Maladie Pyocyanique.’

4 Armand Ruffer, ‘ Experimental Investigation into the Nature of the Dis-
ease produced by the Inoculation of the Bacillus pyocyaneus,’

4.46 M. ARMAND RUFFER.

In a series of experiments Wyssokowitch injected into the
veins of animals—l1st, simple saprophytes ; 2nd, bacilli patho-
genic for certain animals, but harmless for the animals used ;
8rd, pathogenic micro-organisms ; 4th, micro-organisms which
become pathogenic only when injected in large quantities.
Examining the blood of animals by cultivating it on gelatine
plates, he found that non-pathogenic micro-organisms disap-
peared with the greatest rapidity, so that the blood contained
none after three hours. The micro-organisms belonging to
the second class disappeared more slowly, i.e. in twenty-four
hours; whilst pathogenic micro-organisms, e. g. anthrax, di-
minish in number at first, so that none could be found after
four hours, but then multiplied rapidly, so that the blood
twenty-four hours after inoculation contained an incredible
number. The bacilli of anthrax pass into the urine only a
few hours before death when the urine contains blood; other
micro-organisms do not pass in the urine unless some lesion of
the kidney is present.

Non-pathogenic micro-organisms which form spores dis-
appear from the blood with exceeding slowness. Thus the
spores of the Bacillus subtilis, when the latter micro-
organism is injected into the blood, are met with in the liver
and spleen two or three months after the injection, even
when the animals have remained in perfect health.

Wyssokowitch has found the micro-organisms of suppura-
tion (Staphylococcus aureus and S. albus) in the cells
of the milk of women suffering from puerperal fever. Ac-
cording to Malvoz,' the passing of bacilli from the maternal
to the foetal placenta only takes place when slight vascular
ruptures are present.

Clinical facts also show that repeated examinations of the
blood in the large majority of infective diseases does not reveal
the presence of micro-organisms in that fluid. In typhoid
fever, for instance, the bacilli are never met with in the blood.
(For an excellent résumé of this question see J. Gasser, ‘Arch.

1 Malvoz, ‘Annales de l'Institut Pasteur,’ 1888, p. 121.

IMMUNITY AGAINST MICROBES. 44,7

de Médecine Expérimentale et d’Anatomie Pathologique,’ vol.
ili, No. 1, 1891, p. 119.)

Such competent observers as Pfuhl,) Merke,? Leitz, and
Lucatello,’ failed to find any micro-organisms in the blood of
patients during the third and fourth week of typhoid fever.
Neuhaus* examined the blood in nine cases of the same
disease, by inoculating it on gelatine tubes; all the tubes
remained sterile, with the exception of six, which had been
inoculated with the blood coming from the spots of six
different patients. Riitimeyer® came to similar results, and
hence we may conclude that in this disease the blood contains
the specific bacilli in an intermittent manner only, although
the spleen® always contains them. These clinical results agree
exactly with the experimental data; for it has been found?
that if a pure culture of typhoid bacilli be injected into the
veins of a rabbit, the animal killed after eighteen hours, and
particles of organs, &c., sown on gelatine plates, the number
of colonies varies with the organs used.

Thus the spleen gave ; : i . 242 colonies.
Medulla ossium : : : ; . 200 33
Liver : ; i : : ‘ ~ al A
Heart : ; 5 ‘ ; : a elk

The bacillus of tubercle in man at least has only once been
met with in the blood,’ and yet it is undoubtedly carried about
by this fluid. Thus meningeal tubercles are often closely
applied to a small artery which is obliterated by fibrin. In
the tunica intima of vessels of tubercular meninges? are found
a large number of bacilli, small cells and giant-cells; and all

1 * Deutsche Militararzte Zeitschrift,’ 1886, p. 23.

2 ‘Munich Med. Wochenschrift,’ 1880, p. 491.

3 «Bull. d’Académ. de Genova,’ ii, 3, 1887.

4 © Berlin Klin. Wochenschrift,’ 1886, p. 389.

§ «Correspondenzblatt f. Schweiz Aerzte,’ p. 397, 1887.

6 Chantemesse and Widal, ‘ Arch. de Physiologie,’ March, 1887.

7 Wyssokowitch, loc. cit.

8 Weichselbaum, ‘ Wiener Med. Wochenschr.,’ 1884, No. 12.

® Cornil, “Contribution a étude de la Tuberculose,” ‘ Journ, de l’Anatomie,’
1880,

448 M. ARMAND RUFFER.

these may be met with even in the clot obstructing the vessel.
In general miliary tuberculosis, moreover, tubercular granula-
tions of the intima of pulmonary veins, of the right endo-
cardium, or inferior vena cava, are generally found. In patients
suffering from miliary tuberculosis! bacilli have been met with
post mortem in the clots of large vessels. In experiments
in which the Bacillus pyocyaneus had been injected into
the veins, it was not unfrequently found that the blood con-
tained no micro-organisms,” when the liver, kidneys, lungs,
and even urine, were swarming with them. The blood, there-
fore, is not their usual habitat; but micro-organisms, like
inert powders, injected into the blood, are arrested wherever
the circulation is slowed by any cause; that is, in the liver,
spleen, kidney, marrow of bone, lymphatic glands, &. When
injected in large quantities, the bacilli may produce lesions
simply by their numbers. Thus most beautiful and typical
infarcts can be produced by injecting large quantities of a
culture of Bacillus pyocyaneus into an animal which has
been rendered artificially immune.’ One thing is certain,
namely, that the activity of the lymphoid cells contained in
the blood and other tissues varies exceedingly, according as
the animal possesses some degree of immunity or not. Thus,
according to Metschnikoff, when virulent anthrax is injected
into a rabbit, the bacilli in the blood are extremely numerous,
and lie free in the fluid. The same observer rendered animals
immune against anthrax by Pasteur’s method; he then inocu-
lated virulent anthrax, and examined the blood at varying
intervals after the inoculation. Sixteen hours after the inocu-
lation some bacilli were still free in the fluid, while many were
contained in white corpuscles, Twenty-two hours after the
inoculation all the bacilli were contained in lymphoid cells; or
if by any chance a number of them were found floating free
in the liquid, they were surrounded by a mass of white cor-
puscles. Three days afterwards all the bacilli had disappeared.

1 Weigert, “ Zur Lehre von der Tuberculose,” ‘ Virchow’s Archiv,’ 1882.
2 Charrin and Armand Ruffer, loc. cit.
3 Charrin and Armand Ruffer, C. R., ‘ De la Société de l’Anatomie,’ 1889.

IMMUNITY AGAINST MICROBES. 449

Hesse has arrived at similar results by injecting virulent
anthrax into the veins of animals, such as frogs and dogs,
which are naturally immune against anthrax.

In tuberculosis, also, giant-cells are met with in all tuber-
cular tissues ;! but one may say that whenever the bacilli of
tubercle invade the tissues through the blood, giant-cells are
met with in the blood-vessels of these regions. When a pure
culture of virulent tubercle bacilli is injected into the lateral
vein of the ear of a rabbit, the bacilli, becoming arrested in
the veins and capillary vessels of the liver and spleen, give rise
to small fibrinous coagula, in which they multiply up to the
fifth or seventh day. After a week the cells of the spleen and
the leucocytes proliferate actively in the vessels, and the
colonies of bacilli become surrounded by migrating cells, which
develop into epithelioid and giant cells.

Up to the present time I have only stated the microscopical
facts which may be observed under the microscope. I have
not attempted to explain why the cells migrate or not out of
the vessels. This question will be fully discussed in the next
part of the paper, when I intend to speak of the influence
exerted on amceboid cells and on the other anatomical elements
of the tissues by the chemical poisons secreted by micro-
organisms.

Norr.—In a lecture delivered before the Royal Institution
on February 20th, 1891, my friend Dr. E. Klein, F.R.S.,
vehemently attacked the whole theory of immunity being due
to the action of phagocytes. Ido not intend to break through
the plan of this work in order to answer Dr. Klein’s objections,
as the facts on which he bases his theories will be fully discussed
later on; but I take this opportunity of saying at once that I
fully maintain the truth of every one of the observations
which I have made, and every new fact which I have observed
lately confirms me in my opinion.

1 Cornil and Babes, ‘ Les Bactéries,’ vol. ii, p. 92.

VOL. XXXII, PART IV.——-NEW SER. HH

450 M. ARMAND RUFFER.

DESCRIPTION OF PLATES XXXII & XXXIII,

Illustrating Dr. M. Armand Ruffer’s paper on “ Immunity
against Microbes.”

Fic. A.—Wall of abscess, following an inoculation of the weak virus of
quarter-evil into a guinea-pig. Alum-carmine and gentian-violet. Veérick,
oc. J, x ~;th. a, 6, ce, d point to bacilli in various stages of degeneration.

Fic. B.—From the same section as the preceding. Vérick, oc. III, x 2th.
For letters see text.

Fic. C.—Sections through superficial part of the diphtheritic membrane
in man. Gentian-violet and alum-carmine. Verick, oc. I, x ~,th. a and &
point to cells containing bacilli in their interior.

Fic. D.—Section of diphtheritic membrane under a low power. a@ repre-
sents the layer of bacilli; 2 the membrane underneath.

Fic. E.—From the same section as preceding. Gentian-violet and alum-
carmine. Vérick, oc. III, x +th. Leucocytes of diphtheritic membrane
containing bacilli in various stages of degeneration.

Fic. F,—Epithelioid cells containing clubs (actinomycosis). Logwood
and fuchsine staining. Verick, oc. Ill, x 3th.

No. 1.—z. Nucleus of cell. a, 4, c, Clubs in various stages of de-
generation.

No. 2.—z. Nucleus. a, 4, c. Clubs of actinomycosis in various stages
of degeneration. 7. Débris of lymphocyte.

Nos. 3, 4, 5.—#. Nuclei. a, 0, c. Débris of clubs and vacuole.

No. 6.—z. Nucleus. a, 4, c, d, e. Débris of clubs. 7, Débris of
lymphocytes.

Fic. G.—Giant-cell, containing—a. Degenerated bunch of actinomycosis,

' Fic. H.—Giant-cells in actinomycosis.

Fie. I.—Epithelioid cells from tubercular guinea-pig’s spleen. Logwood
staining. Vérick, oc. I, x ~:th. x. Nucleus. a. Blood-pigment,

Fic. J.— Development of connective tissue at the periphery of a nodule of
actinomycosis.

Fic. K.—Giant-cell from tubercular guinea-pig’s spleen, containing blood-
pigment.

Fie. L.—Epithelioid cells from nodule of actinomycosis, containing in their
interior leucocytes in various stages of degeneration.

Fic. M.—Development of epithelioid cells from lymphocytes in nodule
of actinomycosis. Logwood staining. Vérick, oc. I, x 7th.

FORMATION AND FATE OF THE PRIMITIVE STREAK, 461

The Formation and Fate of the Primitive
Streak, with Observations on the Archen-
teron and Germinal Layers of Rana tem-
poraria.

By

Arthur Robinson, M.D.,
Senior Demonstrator of Anatomy in the Owens College ;

and

Richard Assheton, M.A.,
Demonstrator of Zoology in the Owens College.

With Plates XXXIV and XXXV.

EvEN a superficial perusal of the literature of the develop-
ment of the Amphibian ovum is sufficient to acquaint the in-
quirer with numerous contradictory statements concerning
points of weighty morphological importance. Not only are
there differences of opinion as to the interpretation of some of
the developmental features which are generally allowed to be
readily recognisable, but there are also assertions, made by
different observers, concerning merely the structural arrange-
ment and the fate of various portions of the Amphibian ovum
which are absolutely irreconcilable with each other, The con-
fusion is added to by a loose application of terms describing
changes and areas, so that it becomes difficult to decide the
exact relation that the Amphibian ovum bears to the ovum of
other Vertebrates.

In view of the chaos which already exists we would not
readily enter upon any further discussion of Amphibian

452 ARTHUR ROBINSON AND RICHARD ASSHETON,

development, were it not that we think the results of our recent
observations on the development of the Anura tend to simplify
the problem we have been studying by throwing light upon
several interesting developmental phenomena. For these
reasons alone are we induced to publish the results of a research
which tend to conclusions different from those of our prede-
cessors, whose experience is, in most cases, much more exten-
sive than our own.

At the outset it may, with advantage, be stated that our ob-
servations were directed principally to the mode of formation of
the archenteron and blastopore, the fate of the blastopore, and
the formation and fate of the primitive streak ; but incidentally
we have dealt with the separation of the germinal layers and
the relation of the mesoblast to the chorda and hypoblast.

We worked independently of each other. One of us was
led on to the investigation by the contradictions between the
current statements regarding and the obvious facts disclosed
by the developing anural ovum; whilst the other approached the
subject with the conviction, based upon theoretical grounds,
that either the descriptions of certain phases of Amphibian de-
velopment were incorrect, or that there were very peculiar and
significant differences between the Amphibian and other Verte-
brate ova. Until our conclusions had been arrived at neither
of us was aware that the other was engaged upon the subject.
Consequently our observations were made and our conclu-
sions formed independently of each other.

Methods.—Living and hardened ova were examined, but
we relied chiefly upon sections cut in the three usual planes,
horizontally, sagitally, and transversely ; and of these the hori-
zontal have proved in some respects the most useful and in-
structive, more especially in the observations upon the fate of
the primitive streak.

The ova were hardened either in Perenyi’s fluid or in
Kleinenberg’s picro-sulphuric acid solution. The former fluid
was most useful in the younger stages. After the hardening
was completed the ova intended for sections were embedded in
paraffin in the usual manner.

FORMATION AND FATE OF THE PRIMITIVE STREAK. 453

None of the ova were stained, for our experience has been
that the solutions through which the ova are passed, before the
staining is complete, alters the relations of the cells, and that
the staining hinders rather than facilitates the microscopical
examination of sections of the younger stages.

We obtained several surface views which were useful for com-
parison with the results of previous observers ; of these the one
represented in fig. 11 was drawn from a living embryo.

Fig. 23 was drawn partly from a surface view of an ovum
hardened in Kleinenberg’s fluid, and partly from a model con-
structed by putting together, in order of sequence, pieces of
cardboard cut to correspond with camera drawings of a com-
plete series of transverse sections. Our work was greatly
facilitated by Professor Marshall, who very kindly placed at
our disposal a large series of sections, and to him also our
thanks are due for much kind advice.

Before proceeding to the description of our own observations,
we must refer shortly to previous records, for by this course
alone shall we be able to indicate clearly the differences be-
tween our results and those of the observers who have pre-
ceded us, and at the same time we shall obtain an opportunity
of defining some of the terms that we shall be obliged to use in
our description.

Turning, therefore, in the first place to the consideration of
the formation of the archenteron and the blastopore (but
leaving aside for the present the concrescence theory of His
[22 and 23], so far asit concerns the formation of this cavity),
we find that upon this, as upon most other important points,
there is a distinct difference of opinion between the previous
observers. It is stated by some that after the formation of the
segmentation cavity, and when the ovum is only partially
covered by the pigmented epiblast-cells, the archenteron is
formed by an invagination, which “ first commences by an inflec-
tion of the epiblast-cells for a small are on the equatorial line
which marks the junction between the epiblastic cells and the
yolk-cells” (1, p. 102).

This preliminary invagination is also described by Perenyi

A454 ARTHUR ROBINSON AND RICHARD ASSHETON.

in Bombinator (40), O. Schultze in Rana fusca (45), and
by Scott and Osborn in the Newt (48).

Whilst it is proceeding the enclosure of the yolk has been
rapidly taking place. ‘‘It is effected by the epiblast growing
over the yolk at all points of its circumference” (1, p. 102). We
have in this latter statement of Balfour’s a very fair summary
of the general opinion that the epiblast and yolk are distinct
parts of the ovum ; indeed, Balfour speaks of the yolk-cells as
“ these large cells which are part of the primitive hypoblast”
(1, p. 101), with which, in our opinion, they cannot fairly be
compared; and we shall speak of them in the sequel merely as
yolk-cells, though probably the more correct term would be
germ segments. It is not necessary, however, to enter into a
discussion of the mode of extension of the epiblast at the pre-
sent moment, and we can proceed, therefore, to a point which
is allowed by all, namely, that the superficial extension of the
epiblast terminates at the margin of a large circular opening,
called the blastopore or the anus of Rusconi, at the margin of
which the surface epiblast becomes continuous with the cells
lining the archenteric cavity, which, on account of their posi-
tion, are spoken of as hypoblast, although those situated in the
dorsal wall of the cavity are said to be invaginated epiblast in
the Newt (48), or partly epiblast and partly differentiated
yolk-cells in the Anura (1, p. 102), whilst the lateral walls
and the floor of the archenteron are formed by modified yolk-
cells.

Houssay (26) and Moquin-Tandon (388) are, so far as we are
aware, the only authors who have combated this supposititious
mode of formation of the archenteron. Houssay examined
the Axolotl and Moquin-Tandon several Anura. They both
state that the archenteron is formed by splitting amidst the
yolk-cells, from which it follows that both in the Urodela and
Anura there is no invagination of epiblast into the yolk, and
that the archenteron is surrounded by modified yolk-cells
which eventually form a distinct hypoblastic layer, which is
continuous with the epiblast at the margin of the blastopore.

The blastopore is unanimously defined in Amphibians as an

FORMATION AND FATE OF THE PRIMITIVE STREAK, 450

opening, round the margins of which the epiblast and hypo-
blast are continuous, and which forms the passage of com-
munication between the archenteron and the exterior. But
concerning the formation and surroundings of the archenteron
there are two opposed opinions, which are—

1. That the archenteron of the Amphibia is a cavity, formed
as in Amphioxus by invagination, and is lined partly by modi-
fied yolk-cells and partly by invaginated epiblast.

2. That the archenteron is formed in situ by splitting
amongst the yolk-cells, and that it is entirely surrounded by
modified yolk-cells.

To these opinions it will be necessary to return, but in the
meantime we pass to a consideration of the various accounts
of the fate of the Amphibian blastopore.

This opening, after remaining for a time, is, according to
Balfour’s account, decreased in size by the approximation of
its lips until it forms “a narrow passage, on the dorsal side of
which the neural tube opens. ... . The external opening of
this passage gradually becomes obliterated, and the passage is
left as a narrow diverticulum leading from the hind end of the
mesenteron into the neural canal (1, p. 108). It forms the
post-anal gut, and gradually narrows and finally atrophies.”
This account is in the main conformable with Goette’s first
description of Bombinator: “ Die verengte sich vorherrschend
von beiden Seiten her, sodass sie Spaltartig wurde und ihr
Langsdurchmesser in der Medianebene des sich entwickelenden
Embryonalkorper lag” (14, p. 182). And, further, Balfour
(1, p. 108) states that the anus is formed in Rana temporaria
at an earlier period than in Bombinator, as described by Goette,
but in the same manner, that is, by the fusion of a diverti-
culum from the mesenteron with a cutaneous invagination.
Almost the same change takes place in Rana fusca (46);
Spencer, however, as a result of his study of Rana tem-
poraria, came to the conclusion that the blastopore was
transformed into the permanent anus (52); and Alice Johnson
and Lilian Sheldon (28) arrived at a similar conclusion
concerning the blastopore of Triton cristatus, which,

456 ARTHUR ROBINSON AND RICHARD ASSHETON.

according to the account of Scott and Osborn (48), is enclosed
by the neural folds.

Durham (9) describes a neurenteric canal and a blastopore
present at the same time in Rana ; and Sidebotham (61) asserts
that the neural folds do not enclose the blastopore, which is
closed subsequently to the meeting of the neural folds, and
that the anus is derived from an independent proctodeal
invagination.

Morgan’s (89) interesting observations tend to a solution
of the difficulty caused by the preceding contradictions, inas-
much as they show that in Amblystoma punctatum the
mesial portions of the lateral lips of the blastopore are first
approximated, the blastopore thus becoming hour-glass-shaped,
and then fused, so that the previously single opening is divided
into two; the anterior of the two is enclosed by the medullary
folds, and becomes the neurenteric canal; the posterior remains
as the anal orifice. Goette (15) has lately described a some-
what similar condition in Bombinator.

In Rana helecina, according to Morgan (89), there is
behind the blastopore a groove communicating posteriorly in
a dark spot, which is a later formation than the blastopore.
In describing a very similar groove in Bufo lentiginosus
he states that it is separated from the archenteron by three
embryonic layers, a point of importance in any comparison of
this region with the primitive streak of other Vertebrates.

Erlanger’s (10a) observations upon the Anura have led him
to the conclusion that the blastopore closes both from before
backwards and from behind forwards. The closure from before
backwards is associated with the formation of the primitive
streak, which lies in front of the neurenteric canal. The
closure from behind forwards gives rise, in the first instance,
to a fused mass of cells, which rapidly differentiates into layers,
after which the anus forms in the situation previously occupied
by the posterior portion of the blastopore. Therefore, so far
only as the statements of the authors to whom we have
referred are concerned, the blastopore of Amphibians may
either—

FORMATION AND FATE OF THE PRIMITIVE STREAK. 4097

Become (1) gradually constricted, being transformed into
a narrow canal, which for a time unites neural and
alimentary cavities, but eventually disappears—
Balfour (1), Schultze (46), Scott and Osborn (48) ;

Or (2) it is transformed into the anus—Alice Johnson
(27), Spencer (52) ;

Or (3) it is not enclosed in the neural folds, it does
not form the anus, but gradually disappears—Side-
botham (51) ;

Or (4) the anterior part becomes the neurenteric canal and
the posterior part the anus—Morgan (89), Schwarz
(47), Goette (15) ;

Or (5) the anterior portion becomes the primitive streak, the
middle portion the neurenteric canal, and the posterior
part, after being closed, is again reopened in a small
portion of its extent as the anus—Erlanger (104).

But this summary does not include all the fates which are
allotted to the blastoporic opening, for its history has been
intimately connected with that of the primitive streak and
the axial line of the embryo by the researches of His (22 and
23) and Rauber (42), whose observations resulted in the
“ concrescence theory” of Vertebrate development, which
Minot (86) has striven to support in his recent papers. It
is necessary to consider this theory, as the results of our ob-
servations bear upon it; but before referring further to it we
must draw attention for a moment to the structure of the lips
of the blastopore. It is almost universally allowed that in all
Vertebrates, with the exception of Amphioxus, there is in this
region a fusion of all three layers of the germ, and it follows
as a consequence, if the lateral borders of the blastopore
are approximated and united, that there should result a mesial
axial rod of fused tissue, which cannot in the first instance be
resolved into distinct layers. Such a fusion of the blastopore
lip was long ago shown to occur in Elasmobranchii by Balfour
(1, p. 51), who compared the line of fusion of the lips of the
Elasmobranch blastopore to that portion of the blastodermic
area of the Avian ovum to which the term primitive streak

458 ARTHUR ROBINSON AND RICHARD ASSHETON.

had been applied. ‘‘ The primitive streak represents the linear
streak connecting the Elasmobranch embryo with the edge of
the blastoderm after it has become removed from its previous
peripheral position, as well as the true neurenteric part of the
Elasmobranch blastopore” (1, p. 238). ... “That it is in
later stages not continued to the edge of the blastoderm, as in
Elasmobranchii, is due to its being a rudimentary organ” (1,
p. 240). In Balfour’s opinion, therefore, the primitive streak
represents a portion of the blastoporic opening situated poste-
rior to the neurenteric canal, and in conformity with his state-
ment that the anus of Rusconi in the frog is the whole of the
blastopore, which becomes gradually contracted, he does not
describe a primitive streak in Amphibia. His conclusions,
however, are not in accord with those of the upholders of the
concrescence theory, who believe that the axial portion of the
embryo is formed by the fusion of the lips of an elongated
blastopore. The fusion takes place from before backwards,
and results in the formation of an axial area, which lies in
front of the remains of the blastopore. This area is termed
the primitive streak.

If we examine the anus of Rusconi of Amphibia in the light
of Balfour’s conclusions concerning the blastopore, and then in
that of the concrescence theory, we are forced to believe in
the first case that either there is no primitive streak in
Amphibians, or that it lies somewhere posterior to the neu-
renteric canal; and, further, that it would be completely repre-
sented by the fusion of the middle part of the lateral lips of
the blastopore in Amblystoma punctatum (89), and in
Bombinator according to Goette’s description ; or, in the second
case, that the anus of Rusconi represents the posterior part of
the blastopore, and that the primitive streak is situated in
front of it.

Schwarz (47) and Goette (15) compare the “ Prostoma-
schluss” which lies between the neurenteric canal and anus in
Bombinator to the primitive streak of the higher Vertebrates.

Morgan (39), Alice Johnson (27), and Schultze (45) mention
a primitive streak in front of the blastopore. Morgan gives no

FORMATION AND FATE OF THE PRIMITIVE STREAK. 409

description either of the formation or structure of the streak.
Alice Johnson (27) describes a fusion of the germinal layers in
the region in front of the blastopore which she calls the primi-
tive streak, but she does not say whether the fusion is primary
or secondary. ‘There is, however, no doubt about the formation
of the area called primitive streak by Schultze in his de-
scription of Rana fusca, for he describes the process in the
following words:—‘‘ Wahrend auf den vorhergehender Ent-
wicklungsstadien dorsal und median das dussere Keimblatt
durch einen Spaltraum von dem mittleren Blatt getrennt war,
und beide Blatter nur an der ringformigen Zone der Blasto-
poruslippe in einander wbergingen, bildet sich gegen Ende der
Gastrulation, von der Mitte der dorsalen Urmundlippe aus, eine
- nach dem Kopfe hin vorwarts schreitende lineare Verwachsung,
des Ausseren und mittleren Blattes aus”... . “diese
lineare Verwachsung, in welcher wie in dem Primitivstreifen
des Hoheren Wirbelthiere Ektoblast und Mesoblast zusammen-
hingen, von der dorsalen Urmundlippen sich nach dem Kopfe
hin allmahlich ausdehnt, und wachst also der Primitivstreifen
auch bei den Amphibien von hinten nach vorn ” (45, p. 330).

This statement of Schultze’s tends to increase the difficulty
of the situation, for it agrees neither with Balfour’s nor with
the concrescence theory of the primitive streak. In opposition
to Balfour’s theory Schultze places the primitive streak in
front of the neurenteric canal, and he says that the streak is
formed from behind forwards, a statement incompatible with
the concrescence theory, which necessitates that “the ento-
dermal canal and primitive streak begin at the edge of the
blastoderm and grow at their posterior end away from the seg-
mentation cavity, and at the same rate the blastoderm over-
spreads the yolk ” (86, p. 511).

Neither do Erlanger’s observations (10a) tend to simplify
the problem. He associates the primitive streak with the
antero-posterior closure of the dorsal portion of the blastopore,
but according to his figures the primitive streak ultimately ex-
ceeds in length the diameter of the primitive blastopore. He
figures no sections to show the structure of the streak, and we

460 ARTHUR ROBINSON AND RICHARD ASSHETON.

are inclined to believe that he has mistaken the neural furrow
for the primitive streak. But if his conclusions are correct,
they support the concrescence theory only in part, in so far as
the fusion from before backwards of a portion of the blastopore
is concerned; whilst the fusion of the lip of the ventral por-
tion of the blastopore, which he describes as taking place
from behind forward, is at variance with the concrescence
theory.

Nevertheless, to the summary which has already been given
of the fate of the blastopore in Amphibians (p. 457), we must
add the further possibility that, according to the concrescence
theory, the posterior part of it only is to be recognised in the
anus of Rusconi, the lateral lips of the anterior part having
undergone a “ retrogressive fusion” similar to that described
by His in bony fishes (22), and that this fusion is continued
until the complete obliteration of the blastopore results.

From the statements noted in the preceding survey of the
literature of the subject we may deduce the following summary
of the presence or absence and the mode of formation of the
primitive streak in Amphibia :

1. There is no mention of a true primitive streak—Balfour
(1).

2. There is a primitive streak situated in front of the anus
of Rusconi—Schultze (45), Minot (36), Alice Johnson (27),
Erlanger (10a).

3. The fused lips of the blastopore between the neurenteric
canal and the anus represent the primitive streak—Schwarz
(47).

The primitive streak is formed—

1. By retrogressive fusion of the lips of the blastopore (con-
crescence theory).

2. By fusion of the lips of the blastopore between the neu-
renteric canal and anus—Schwarz (47).

3. By fusion of the epiblast and mesoblast from behind for-
wards in front of the anus of Rusconi—Schultze (45).

Before concluding this short survey of previous records we
wish to summarise the statements, some of which have been

FORMATION AND FATE OF THE PRIMITIVE STREAK. 461

already referred to concerning the formation of the anus in
Amphibians.

1. It is a new formation in the region below the blastopore—
Balfour (1), Sidebotham (51).

2. It is the blastopore—Sedgwick (49), Alice Johnson (27),
Spencer (52).

3. It is the posterior portion of the blastopore—Morgan
(Amblystoma punctatum) (39), Schwarz (47), Goette (15).

4, It is a secondary opening in the situation of the posterior
part of the primitive blastopore—Erlanger (10a).

The Enclosure of the Yolk by Epiblast and the
Formation of the Archenteron.

At the period generally spoken of as the end of the seg-
mentation the anural ovum is a sphere which contains an
excentrically situated segmentation cavity.

The segmentation cavity has a roof which ultimately becomes
the anterior wall of the gastrula; for the anus, which marks the
posterior end of the embryo, appears at the opposite pole of
the ovum, that is in the floor of the segmentation cavity.

The roof of the cavity is formed by two or three rows of
comparatively small pigment-bearing cells, and the floor by a
mass of ill-defined nutriment-laden cells which collectively
form the yolk. At the margin of the segmentation cavity the
two walls merge into each other by a series of insensible
gradations, so that it is impossible to say where one ends and
the other begins. Nevertheless it is certain that the cells of
the roof of the segmentation cavity are epiblast, for they ulti-
mately form a portion of the external covering of the embryo;
but it is incorrect to speak of the opposite wall, the yolk-cells,
as modified hypoblast (1, p. 103), unless it is allowed that
epiblast may be formed from modified hypoblastic cells. For
during the formation of the blastopore the epiblast does not
grow over the yolk-cells, enclosing them by a process of epibolic
invagination. If it did, it would be possible to recognise on
section a line along which the epiblast terminated. No one
has described or figured such a line of limitation except dia-

462 ARTHUR ROBINSON AND RICHARD ASSHETON.

grammatically ; and a careful examination of thin sections of
anural ova, in the stages preceding the completion of the
circular blastopore, shows clearly that the descriptions given of
the gradual extension of the epiblast over the yolk represent
theory rather than fact.

The changes observable are, firstly, that the cells of the
superficial layer of the yolk gradually become more distinctly
pigmented; and, secondly, that they thereafter divide into
smaller and more distinct segments, which arrange themselves
into two layers, a superficial layer of somewhat cubical deeply
pigmented cells, and a deeper layer of less pigmented and more
rounded cells, irregularly arranged into two or more rows: the
latter are separated by a distinct space from the subjacent
remainder of the yolk-cells. This transformation and re-
arrangement proceeds backwards towards the blastopore, stop-
ping about °352 mm. from that aperture, first on its dorsal
and then on its lateral and ventral borders, where an area of
fusion remains, which is for convenience divided into a
dorsal, a ventral, and two lateral lips or borders.

We conclude, therefore—(1) That the segmentation of the
anural ovum does not result in the formation of a vesicle, the
roof of which is epiblast and the floor modified hypoblast, but
that at the end of the segmentation the primary layers of the
ovum are ouly partially formed. The roof of the segmenta-
tion cavity is epiblast, but the floor or yolk is not modified
hypoblast. It consists of indifferentiated germ-cells, the true
characters of which are not at first recognisable.

(2) That the yolk is not enclosed by the gradual extension
over it of a previously differentiated epiblast, but that the
superficial layer of yolk-cells becomes gradually differentiated
into the two layers of epiblast, leaving a remainder of the
yolk. This consists of hypoblast and mesoblast, which are not
at first separated from each other.

The formation of epiblast from yolk-cells does not take
place over the whole surface of the yolk, for at the posterior
pole of the ovum there is a circular patch of yolk-cells from
which no epiblast is formed, The cells in question form the

FORMATION AND FATE OF THE PRIMITIVE STREAK. 463

yolk-plug, and when the epiblast formation has arrived from
all sides at their margin, the circular blastopore or anus of
Rusconi is first definitely established.

During the period of differentiation of the epiblast the
archenteron has also been forming. The first indication of
this cavity is the appearance of a curved area of pigmentation
(fig. 1 a and fig. 2 AR) of semilunar outline amidst the yolk-
cells at the posterior pole of the ovum above the equator. The
convexity of the pigmented area is directed forwards amidst
the yolk-cells, towards the segmentation cavity. The angles
are turned ventro-laterally. The centre of the concavity of
the semilunar area corresponds to the dorsal lip of the blasto-
pore. As the convexity of the area extends forwards towards
the segmentation cavity, its lateral angles travel ventrally along
the lines of the lateral lip of the blastopore until they meet in
the situation of the ventral lip of that opening, which is thus
marked out by a line of pigmentation that indicates the situa-
tion of the future cavity.

The pigmented area is produced and extended by the deposit
of pigment in the adjacent margins of a double row of yolk-
cells in the manner shown in figs. 1 and 2, which are drawings
of sections of the advancing anterior extremity of the pig-
mented area of a much more developed ovum. The pigment is
deposited in the protoplasm of the double row of cells which,
eventually, will form the boundary wall of the archenteron,
and it also radiates from this area along the adjacent margins
of the cells of each row.

It is to be understood, however, that directly after the pig-
mented area is first formed, and as it extends forwards and
ventrally, a slit-like space appears in the middle of its pos-
terior portion. This space first limits the dorsal lip of the
blastopore, and then extends forwards and ventrally, fol-
lowing the deposit of pigment, and separating the two rows
of marginally pigmented cells from each other. It is the
archenteric space. As its lateral angles extend ventrally along
the line of pigmentation they define the lateral lip of the blas-
topore; their union ventrally gives rise to the ventral lip, and

464 ARTHUR ROBINSON AND RICHARD ASSHETON.

at the same time defines the posterior limit of the ventral wall
of the archenteron (fig. 3).

A diagrammatic representation of a sagittal mesial section of
the ovum at the period of completion of the blastoporic lip is
given in fig. 4, from which it will be readily seen that, at this
period, the ventral wall of the archenteron extends from the
point a to the point 2 Ina portion of its extent the floor of
the slit-like cavity lies parallel to the superficial surface of the
ovum, but posteriorly it is projected outwards into a deficiency
in the external wall (the blastopore), forming in this region the
yolk-plug or bouchon d’Ecker. At the same stage the dorsal
wall of the archenteron is not co-extensive with the ventral. It
extends only from the point a to the point 6, which marks the
dorsal lip of the blastopore. From the dorsal to the ventral
lip of the blastopore the wall of the archenteron is deficient.
It will be shown afterwards how this portion of the wall, which
we shall call the posterior wall of the archenteron, is completed.

At present we more particularly desire to call attention to
the fact that the point x, which at this period marks the pos-
terior limit of the ventral wall of the archenteron, remains
fixed throughout the later periods, that eventually it is situated
just in front of the anal orifice, and that there is no extension
ventrally of the archenteron beyond it, such as that described
by Balfour, who, speaking of the gradually narrowing blasto-
pore, says: “ Atits front border, on the ventral side, there
may be seen a slight ventrally directed diverticulum of the
alimentary tract, which first becomes visible at a somewhat
earlier stage” (1, p. 108). On the contrary, the ventral wall
of the archenteron is only increased in front of the ventral lip
of the blastopore by the forward extension of the anterior end
of the cavity and the growth of the ovum, and is completed
immediately in front of the ventral lip, so far as we have been
able to ascertain, by the gradual withdrawal of the yolk-plug
and the rearrangement and differentiation of its constituent
cells.

It is certainly noteworthy that the slit-like archenteron does
not appear in the midst of the yolk-cells until the epiblast

FORMATION AND FATE OF THE PRIMITIVE STREAK. 465

reaches the region of the blastoporic margin ; and it is this fact,
together with a predisposition to discover, if possible, an invagi-
native process, which seems to have led to the description of
the formation of the archenteron of the Anura by invagination
of the epiblast.

A careful examination of the walls of the extending archen-
teric cavity reveals no evidence in support of this ideal invagina-
tive process ; on the contrary, it shows that even at the period
of completion of the blastoporic opening (see figs. 1 and 3)
the greater part of the cavity is surrounded by large nutri-
ment-laden, marginally pigmented ceils. More especially is
this the case in the anterior portion of the, as yet, incom-
plete cavity (fig. 2), and at its posterior extremity (fig. 3),
where the slit-like space is only just formed. In the
latter situation the only cell that can be considered as dis-
tinctly epiblastic is that marked c. The remaining cells are
undoubtedly large yolk-cells; and it is only by the division of
these cells, and the arrangement of some or all of their
descendants into a distinct layer, that the true hypoblastic
lining of the alimentary canal and its diverticula is formed.
As the completion of the definite hypoblast is intimately asso-
ciated with the separation of the mesoblast and chorda, its
further history cannot at this period be entered upon, and we
may therefore proceed to the termination of this section by a
summary of the conclusions deduced from that which has been
here set forth.

1. The blastopore is a deficiency in the posterior wall of
the archenteron.

2. The archenteron of the Anura is not formed by inva-
gination, but by a process of splitting amongst the yolk-
cells very similar to that described by Houssay (26) in the
Axolotl.

3. The situation of the archenteric cavity is first defined by
the deposition of pigment in the adjacent margins of a double
row of yolk-cells.

4. No portion of the archenteric wall is formed by invagi-
nated epiblast; on the contrary, the archenteron is surrounded

VOL. XXXII, PART 1V.—NEW SER. 11

4.66 ARTHUR ROBINSON AND RICHARD ASSHETON.

in the first phases of its development by large yolk-cells,
which eventually give rise to the definite hypoblast.

5. The ventral lip of the blastopore indicates the posterior
end of the primitive ventral wall of the archenteron.

6. There is no ventral and forward extension of the archen-
teron in front of and below the ventral lip of the blastopore
by the production of a diverticulum in that situation.

7. The ventral wall of the archenteron, in front of the ven-
tral lip of the blastopore, is completed by the extension of the
anterior end of the cavity, and by the withdrawal and modi-
fication of the cells of the yolk-plug.

Manner in which the Anus of Rusconi closes.

Having discussed the changes that take place in the ovum
up to the time of the formation of the large circular blastopore
or anus of Rusconi, we will now proceed to describe the
manner in which, according to our observations, the closure of
the blastopore or anus of Rusconi seems to be effected, and to
give in some detail an account of the structures which are
the immediate result of the method of closure about to be
described.

According to some of the former accounts, to which we have
made reference above, the anus of Rusconi has been said to
diminish in size by the gradual coming together of each portion
of the blastoporic rim simultaneously. This we believe to be
incorrect. No doubt from surface views alone the hitherto
accepted accounts receive much support.

We have, however, paid particular attention to this point,
and have come to the conclusion that the anus of Rusconi
gradually diminishes in size by the concrescence of the ventral
part of the lateral lips, as indicated in figs. 10 and 18. In
these diagrams, the space lettered BR, inclosed between two
concentric rings, represents the blastoporic rim at the time of
the completion of the circular blastopore, or anus of Rusconi;
and the shaded space, lettered BR’, represents in fig. 10
the position of the blastoporic rim soon after the anus of
Rusconi has begun to diminish in size, and in fig. 18 a later

FORMATION AND FATE OF THE PRIMITIVE STREAK. 467

stage, when the anus of Rusconi has “ contracted ”’ so much
that its diameter is now only about one third of its original
size.

Figs. 7, 8, 9 are from sections taken nearly horizontally
through an embryo in which there was no trace of neural folds,
and in which the anus of Rusconi had only slightly contracted.

Fig. 10 is a diagram of the same embryo representing the
changes which we conclude must have taken place.

The blastoporic rim, round which the three layers, epiblast,
mesoblast, and hypoblast, are all fused, which occupied, on the
completion of the circular anus of Rusconi, the position indi-
cated by the space BR between the two concentric circles, had
in the case of the embryo examined come to occupy the posi-
tion of the shaded portion lettered BR’ in the diagram. In
the same diagram the line w represents that portion of the
original anus of Rusconi which has become closed up by the
concrescence of the ventral part of the lateral portions of the
rim, as stated on p. 466.

We were unable in this specimen to find any trace of the
line of fusion 2 on the surface, and as yet there was no trace
of the neural folds. Accordingly, in order to be able to cut
our sections as near as possible in the horizontal plane as
desired, we had to dissect away the opposite pole of the
embryo, and then by observing the shape of the yolk-plug we
were able to cut our sections very nearly as required.

Fig. 7 has been drawn from a section (which is not quite
horizontal) taken through the centre of that portion of the
blastopore which was still open. At the lips of the blastopore
the three layers are seen to be fused.

The line running from ZZ’ ends at the furthest point from
the blastoporic lip to which the fusion of epiblast (HP) and meso.
blast (ME) extends on one side, and the line from ZZ ends at
the furthest point from the blastoporic lip to which the fusion
of those layers extends on the other side. It will be noticed
that on one side the fusion extends further than on the other.
No doubt this is chiefly owing to the section being not quite
horizontal, the side of the ZZ’ being the more ventral of the

468 ARTHUR ROBINSON AND RICHARD ASSHETON.

two. We have, however, taken the thicker side for the purpose
of measurement.

If the diminution in size of the blastopore has been produced
by the concrescence of the ventral portion of the lateral lips as
suggested above, and as diagrammatically shown in fig. 10, it is
clear that tlie fusion of layers ought to extend further from the
actual lip of the blastopore on the ventral border of the blasto-
pore than on the sides or dorsal border. This will be clearly
seen by reference to the diagram, fig. 10.

It is equally clear that this would not be so if the closure of
the blastopore were strictly centripetal, in which case the fusion
of layers should extend on all sides equally.

If the distance be measured across the blastopore in fig. 7
with a pair of compasses, it will be found to be about the same
distance as between the edge of the blastopore lip and the
point of furthest extension of fused epiblast and mesoblast
on the side of ZZ’.

In the series of sections from which this fig. 7 and the two
represented in figs. 8 and 9 were selected, the open blastopore
was present in forty sections.

Assuming the open blastopore to have been circular, we may
conclude that it would have required forty sections to cut
through the fused mass between the edge of the blastoporic lip
(B’) and the furthest extension of the fused mass indicated by
the line drawn from ZZ’, for we have just now seen that the
distance from B to B’ is about the same as from B’ to ZZ’,
In other words, the forty-first section ought to show no, or
but little, fusion of layers.

Fig. 9 has been drawn from the forty-first section, and
instead of there being no fusion of layers there is an extent of
fused layers about equal to the two fused blastoporic rims, or,
indeed, a little more.

At the sixtieth section the epiblast and mesoblast are still
distinctly fused, and it is not until the eightieth section below
the edge of the ventral lip of the blastopore that the layers can
be definitely said to be separate. In short, whereas by calcula-
tion the fusion of layers on the lateral lips will be cut through

FORMATION AND FATE OF THE PRIMITIVE STREAK. 469

in forty sections, it requires close upon eighty sections to cut
through the fused mass at the ventral lip.

In fig. 10 the levels of the sections (figs. 7, 8, 9) are indicated
by the lines 7, 8, 9.

Thus a sagittal section through the blastopore ought to show
a very much larger mass of fused layers at its ventral lip than
at its dorsal.

This is seen to be the case as shown in fig. 5, which is a
sagittal section through the blastopore of a rather older frog
embryo. The blastopore has closed to a considerably greater
extent than was the case in the embryo we have just been
describing. The actual extent of the closure of the blastopore
by the concrescence of the latero-ventral lips is represented, we
think, by the distance between the point # and the present
edge of the ventral lip of the blastopore.

We find by a series of measurements that at whatever stage
during the closure of the blastopore the section be taken, the
distance between point x and the edge of the dorsal lip of the
blastopore is always approximately the same, and the same as
the diameter of the blastopore at its first commencement.
We say approximately, for, owing probably to variation in
size of the egg, the measurements do not exactly agree.

Fig. 11 was drawn from a living specimen in which the
blastopore had become greatly reduced, and in which the
neural plate and neural groove were distinctly visible along the
dorsal surface.

From the lower, or ventral, lip of the open blastopore stretches
a very faint (too deeply drawn in the figure) line, which pro-
bably is the remaining trace of the line of concrescence of
latero-ventral blastoporie lips.

Fig. 13 is a section taken at right angles to this line, not of
the same embryo, but of one about the same age. In this the
line is seen to be a groove (PG) on the surface, below which
all these layers are fused.

We will at once call this groove primitive groove, and
the fused mass below it primitive streak ; for, as we hope to
prove in the sequel, there can be no reasonable doubt that

470 ARTHUR ROBINSON AND RICHARD ASSHETON.

these structures are the homologue of those parts in the chick
embryo to which these names were originally given.

A section taken dorsal to the blastopore, and dorsal to the
fusion of layers at the dorsal lip of the blastopore, is seen in
fig. 12.

Here the epiblast is seen to be quite distinct from the meso-
blast. There is a distinct groove, the neural groove, very
different in character from that of the groove ventral to the
blastopore, or primitive groove, while the epiblast is thickened
on either side to form the neural plate. The level of the re-
spective sections is marked in fig. 11 by the lines numbered
12 and 13.

Before discussing the relations between the structures, the
formation of which we have been following in the last two or three
pages, and which we have named primitive streak and primi-
tive groove, and the primitive streak and primitive groove of the
chick, we will follow its history a little further, until the stage
at which we may say it has attained its greatest development.

We must, therefore, describe the condition of the primitive
streak of a frog embryo of about 24 mm. in length ; that is to
say, an embryo in which the neural folds are on the point of
meeting, or have just met, as shown in fig. 23.

Primitive Streak of the Frog at the Time of the
Closure of the Neural Folds.

Fig. 23 is the surface view of the posterior end of a 24 mm.
frog embryo, in which the neural folds have just met.

It was drawn partly from a preserved specimen, and partly
from a model, as described on page 453.

The line of junction of neural folds (J) is marked by a deep
groove ending posteriorly in the blastopore.

The blastoporic opening, as seen from the surface, is no
longer circular, but is more or less lozenge-shaped, due to the
folding its dorsal lips have undergone (as will be described
later).

FORMATION AND FATE OF THE PRIMITIVE STREAK. 471

The lengthening of the embryo has removed from the blas-
topore all trace of the “ yolk-plug.”

Running ventrally from the blastopore is the primitive
groove, a rather narrow but now sharply defined groove near
the ventral end, which is suddenly deepened into a pit.

Beyond the pit the groove continues a short dis-
tance. This pit is the commencement of the anus, which will
shortly perforate at this spot. In fact, in the model from which
this figure was partly drawn the anus had just perforated. In
the sections, however, about to be described as representing this
stage the anus has not quite perforated. On either side of the
primitive groove a distinct ridge is visible. .

The bracket (PS) in the figure includes the whole of the
primitive streak from end to end.

The sections 14, 15, 16, 17, are horizontal sections taken
along the lines 14, 15, 16, 17, in fig. 28, and are, therefore,
taken at right angles to the longitudinal axis of the primitive
streak.

Fig. 14 is a camera drawing of a section through the blas-
topore (line 14, fig. 23), which is nearly closed in.

In this section the epiblast, mesoblast, and hypoblast are
seen to be all fused at the lips of the blastopore. It may be
noted that the mesoblast, close to the lip of the blastopore,
seems to be divided into two layers: a very dense compact
layer (IVE’) next the epiblast, in which the cells are so closely
packed as to render it quite impossible to draw each cell by
camera; while inwardly the mesoblast-cells (ME”) are much
looser, and are more deeply pigmented and easily traceable by
help of the camera. In fact, one is almost inclined to say that
the former are mesoblast-cells of epiblastic origin, while the
latter are mesoblastic cells of hypoblastic origin.

Fig. 15 is a section taken about midway between the blas-
topore and the anal pit; that is to say, across the middle of the
primitive streak. This section is directly comparable to that
just described, except that the lips of the blastopore
have met and fused, as described in the earlier part of this
paper. The surface is distinctly grooved, the edges of the

4.72 ARTHUR ROBINSON AND RICHARD ASSHETON.

groove being raised slightly into ridges. The same feature in
the mesoblast may be noticed here as in fig. 14.

Fig. 16 is taken through the anal pit.

Here the primitive groove is very much deepened, so that
not more than two or three cells prevent the completion of
the anal perforation. This pit is deeply pigmented, and
heavy deposits of pigment line the few intervening cells
between exterior and archenteron. All three layers are still
fused.

Fig. 17 represents a section taken below the anal pit; that
is to say, it was the thirteenth section after the last one
figured, fig. 16. In this, although behind the anus, there is
undoubted fusion of epiblast and mesoblast. It is altogether
ventral to the archenteron, so that there can be no fusion with
true hypoblast. There is no fusion either with the yolk-cells.
The surface is grooved, as in the sections anterior to
the anus.

The fusion of layers continues through six more sections ;
that is to say, there are about twenty sections ventral to the
anus, in which there is fusion of epiblast and mesoblast; in
other words, the anus is a perforation through the
primitive streak.

Comparison of the “ Primitive Streak” of the Frog
with the Primitive Streak of the Chick.

We have now described the history of the blastopore or
anus of Rusconi from the time of its first formation until it
has been reduced to a comparatively minute passage between
the archenteron and exterior. We have described this change
as being due to the concrescence of the lower lips of the
blastopore, as shown diagrammatically in figs. 10, 18, 19,
whereby there is produced a median streak characterised by
the fusion of layers, along the surface of which lies a groove,
stretching from the ventral lip of the remaining blastopore
ventralwards as far as the original extension of the blastopore

FORMATION AND FATE OF THE PRIMITIVE STREAK. 473

or anus of Rusconi. To this structure and to the groove upon
it we have applied the terms primitive streak and primitive
groove respectively.

If one of our sections figured, e. g. fig. 15, is compared with
the figure of a transverse section of the primitive streak of a
chick on p. 155 of Balfour’s ‘Comparative Embryology,’
vol. ii (second edition), the resemblance between the two
sections is very marked,

In each case there is an intimate fusion between epiblast and
mesoblast, and a more uncertain fusion between mesoblast and
hypoblast. In each case the surface is marked by a groove.

Nor is it at all impossible that the loose pigmented cells
noticed above, and lettered MEH” in figs. 14 and 15, may be
compared with Balfour’s “layer of stellate cells’? shown in
the figure in the ‘Comparative Embryology’ referred to. In
both cases they seem to arise from the hypoblast rather than
the epiblast. In the frog, however, they are unrecognisable a
short distance from the streak.

In comparing the two streaks with regard to their relations
to the rest of the embryo, we find that we must use the term
primitive streak in a rather wider sense than it is usually
used. In the chick the anterior limit of the primitive streak
may be said to be marked by the posterior end of the noto-
chord with which the streak is fused.

The structure we have so far called primitive streak runs
from the ventral lip of the blastopore. It is, however,
obvious that had the concrescence continued a little further, so
that the whole anus of Rusconi had been obliterated by the
fusion of the blastoporic lips, the primitive streak would
have then commenced at the posterior end of the notochord,
as in the chick; for the notochord and neural folds in the frog
are continuous with—that is to say, fused with —the dorsal lip
of the blastopore. Itis, therefore, clear that the homologue
of the primitive streak of the chick is in the frog the
whole of the blastoporic lip, whether fused or not.

4.74 ARTHUR ROBINSON AND RICHARD ASSHETON.

Relation of Anus to Blastopore.

The relation of the anus to the blastopore has been a much
controverted subject. Goette (14) and Balfour (1) described
it as an entirely separate perforation of the body-wall ventral
to the blastopore; Spencer (52) stated that the blastopore
became directly the anus; Sidebotham (51), who gave a most
careful and exact account of the facts, agreed with Goette and
Balfour on this point, and, failing to recognise the primitive
streak in the frog, described it as they had done, as a new
opening independent of the blastopore. In Erlanger’s (10a)
opinion it is a secondary opening in the situation of the poste-
rior part of the original blastopore. Those who have followed
our account so far will observe that it agrees most closely with
those of Goette, Balfour, and Sidebotham on this point; but,
since the perforation takes place within the primitive
streak, the conclusions we draw from the facts are different.
We infer, therefore, with Erlanger, that the anus of the frog,
although apparently a new perforation, is really a reopening
of a temporarily closed portion of the original blastopore.

Since the perforation occurs at the base of the diverticulum
of the archenteron (which, it will be remembered, was formed
by the closing in of the ventral portions of the lateral lips of the
blastopore), it may be supposed that it is the most ventral
end of the blastopore which, morphologically speak-
ing, persists as the anus.

The Primitive Streak of the Frog and other
Vertebrates.

The term primitive streak appears to have been first applied
to the dark line which appears in the avian blastoderm at the
posterior part of the area pellucida. This line is generally
looked upon as the optical expression of a linear thickening
and fusion of the blastodermic layers, though Kolliker (81,
p. 134) maintains that it is due to proliferation of the epiblast-
cells alone. It seems certain, however, that it is impossible,

FORMATION AND FATE OF THE PRIMITIVE STREAK. 475

during certain periods of the growth of the embryo, to distin-
guish the three germinal layers from each other in the area of
the primitive streak (Duval, 10, p. 181).

After a time differentiation proceeds in the streak ; a groove
appears on its superficial surface, and this is deepened ante-
riorly into a perforation, which ultimately becomes the neuren-
teric canal (47). The anterior wall of this perforation is
formed by a mass of cells in which the epiblast and hypoblast
are united (12). In the posterior portion of the streak the
anal membrane is developed by the gradual thickening and
apposition of the hypoblast and epiblast (18, pl. x, figs. 4 and
5, p. 299; and 47, pl. xiv, fig. 81, p. 203).

The lateral margins and posterior extremity of the streak
are continuous with the mesoblast, which appears to grow out
from them into the surrounding area.

The anterior end of the streak is continuous with the chorda
ventrally, and the central part of the neural plate dorsally.
In the region surrounding the primitive streak the three layers
of the blastoderm are distinct from each other, except in front
of the anterior extremity of the streak, where the so-called
“‘ Kopffortsatz,” the first rudiment of the chorda, is still con-
tinuous with the entoderm. The connection between the
hypoblast and the mesoblast, which exists throughout the
whole length of the streak, is first dissolved posteriorly, where
for a certain period the epiblast and mesoblast are Se but
the hypoblast forms a distinct layer.

Between the primitive streak of a bird and the frog there
are resemblances and differences of importance. In the frog
the primitive streak is formed by a concrescence of the lips of
the blastopore, which proceeds from behind forwards, and
which is only completed on the obliteration of the neurenteric
canal. In the bird the primitive streak is formed from before
backwards according to Duval (10) and Schwarz (47), from
behind forwards according to Balfour and Deighton (2) and
Koller (80) ; and the appearance is due apparently to thicken-
ing and fusion of the two primary layers—Duval (10), Balfour
and Deighton (2).

4.76 ARTHUR ROBINSON AND RICHARD ASSHETON.

In both the frog and the bird the lateral margins and poste-
rior extremity of the streak are continuous with the mesoblast,
which lies free between epiblast and hypoblast outside the
area of the streak.

In the frog the anterior wall of the neurenteric canal is
bounded by an area of fusion, in which the middle of the
neural plate and the posterior end of the chorda are united.
After the obliteration of the neurenteric canal the posterior
end of the chorda and the centre of the neural plate are con-
tinuous with the anterior end of the primitive streak.

In the bird the anterior end of the primitive streak is at
first continuous with the centre of the neural plate and the
“ Kopffortsatz,’ and after the formation of the neurenteric
canal the chorda and the neural plate are fused in the anterior
wall of the latter orifice, becoming again continuous with the
anterior end of the primitive streak after the disappearance of
the neurenteric canal.

In the frog the anus is formed in the posterior part of the
primitive streak. It is a reopening of a portion of the closed
blastoporic orifice. It is not the remains of the blastopore, as
in Amblystoma punctatum (39) and Bombinator (18).
The anus of the bird is morphologically equivalent to the anus
of the frog; and it is also formed, in all probability, by a re-
opening of a previously closed orifice (10).

In reptiles, according to Kupffer (33), there is no primitive
streak, but in the posterior part of the embryonic area the
epiblast is invaginated, and the mesoblast arises, in part at
least, from the margins of the invagination. The cavity of
invagination is the archenteron, and the superficial opening
the Urmund, which would thus entirely correspond to the anus
of Rusconi in Amphibians.

But Balfour (1, p. 168), Strahl (54), Hoffmann (24), Weldon
(56), Mitsuruki and Ishikawa (387), and Wenckebach (57)
describe a primitive streak. Balfour states that in the primi-
tive streak of lizards the epiblast and hypoblast are fused,
though the greater part of the streak consists of proliferated
epiblast. Wenckebach, however, looks upon the hypoblast as

FORMATION AND FATE OF THE PRIMITIVE STREAK. 477

a purely passive agent in the formation of the primitive streak
in lizards; and in the tortoise, according to Mitsuruki and
Ishikawa, the streak consists, in the first instance, mainly of a
mass of hypoblast or yolk, which they compare to the yolk-
plug of Amphibians. To this, however, it cannot correspond,
for we have already shown that the yolk-plug of Rana is a por-
tion of the ventral wall of the archenteron, whilst the primi-
tive streak is formed by the fusion of the lateral lips of a
deficiency in the posterior wall of the same cavity.

But, whatever the mode of its formation may be, the streak
eventually becomes perforated, both anteriorly and posteriorly :
anteriorly by the neurenteric canal—Balfour, Hoffmann,
Weldon, and Strahl;' and posteriorly by the anus—Weldon.
It is thus, in all essential respects, comparable with the primi-
tive streak of birds.

In mammals also a primitive streak is found. Itis described
as commencing in the sheep (5) and in the shrew (25) as a
knob-like swelling of the epiblast in the posterior part of the
embryonic area. The epiblastic thickening extends backwards
and terminates in a tail-swelling. According to Bonnet (5),
Hubrect (25), and Hensen (20), the two primary layers fuse in
the streak, though it does not seem certain that the hypoblast
takes any part in the thickening. Kolliker (31) denies that
the hypoblast takes part in the formation of the streak in the
rabbit. Rabl (41) agrees with Kolliker, and Fleischmann (11)
makes a similar statement concerning the cat. In the primi-
tive streak of the guinea-pig (29) there is fusion of the layers
anteriorly, but posteriorly the thickened epiblastic ridge is
separate from the hypoblast. In the mole (17) all the layers
take part in the formation of the streak, but after a time they are
fused only at its anterior and posterior ends, whilst in the middle
the hypoblast forms a distinct layer. At the anterior end of
the mammalian primitive streak more or less distinct traces of
a neurenteric canal have been found in the rabbit by Strahl

1 Strahl’s statement (54), that in L. agilis the perforation occurs in the
middle of the streak, simply means, apparently, that there is a region of fusion
between the epiblast and hypoblast in front of the opening.

478 ARTHUR ROBINSON AND RICHARD ASSHETON.

(55), in the mole by Lieberkuhn (85) and Heape (17), in the
sheep by Bonnet (5 and 6), and in the bat by Van Beneden
(3). In the anterior wall of the opening the chorda and neural
epiblast are fused, and laterally and posteriorly the mesoblast
hangs in connection with the margins of the streak.

The anal membrane is formed in front of the posterior end of
the streak in the rabbit—Strahl (55); in the mole—Heape
(17) ; in the guinea-pig—Keibel (29) ; and in the rat and mouse
—Robinson (43). In the sheep, Bonnet (7) states that the
anal membrane is situated at the posterior end of the streak ;
and Rabl (41) figures it in the same position in the rabbit.

The differences described are slightand probably unimportant,
and the main facts stand out clearly. Asin the Sauropsida, the
primitive streak is a line of fusion amidst the germinal layers.
It is continuous anteriorly with the neural epiblast and the
chorda, laterally and posteriorly with the mesoblast of the sur-
rounding areas. It becomes perforated by an evanescent neu-
renteric canal and by a permanent anal orifice.

In the Cyclostomata the posterior part of the blastopore
remains open as the anus. Inthe ventral lip of this orifice the
epiblast alone is at first differentiated, the hypoblast and the
mesoblast remaining fused until a comparatively late period—
Goette (15) and Kupffer (34). The blastopore is closed from
before backwards according to Goette’s (15) statement, and
consequently there is formed in front of the anus a mass
of cells called the ‘ Teloblast” by Kupffer (34). This mass
of cells remains for a time fused with both epiblast and
hypoblast (Goette, pl. iv, fig. 41), but afterwards it sepa-
rates from the superficial epiblast and the hypoblast (Kupffer,
pl. xxvii, fig. 28), but remains continuous in front with
the chorda and neural tube. Evidently the area from the
front of the anus to the posterior end of the chorda and
neural tube in Petromyzon corresponds closely to the anterior
portion of the primitive streak of the Sauropsida and mam-
mals. It is never perforated by a neurenteric canal, but
the absence of this passage has no important bearing, as it has
probably been suppressed.

FORMATION AND FATE OF THE PRIMITIVE STREAK. 479

In Teleostean fishes an area of fusion is found round the lips
of the blastopore (Henneguy, 18), but the fusion in front of
the blastopore is much more extensive than that on the sides
and posteriorly, and in its anterior part a vesicle appears—
“ Kupffer’s vesicle.” In front of Kupffer’s vesicle the chorda
and neural plate are fused, and the margins of the fused area
are continuous with the mesoblast. Eventually the posterior
portion of the blastopore is closed by the fusion of its lips
(18, p. 502). The position of the anus is not definitely
stated. The fused area corresponds in the Teleostei even
more closely than in the Cyclostomata with the anterior por-
tion of the primitive streak of the Sauropsida and mammals, for
it is partially perforated anteriorly by Kupffer’s vesicle, which
is evidently situated in the position of the neurenteric canal of
the higher Vertebrata.

We have before referred to Balfour’s description of the con-
dition of the posterior end of the embryo in the Elasmo-
branchii (p. 11), and to this we have only to add that at a later
period the ventral part of the blastopore also becomes closed
by fusion, and that at a subsequently later period the anus is
formed as a secondary opening in the line of fusion (Schwarz,
47).

The observations upon the Ganoids are not sufficiently com-
plete to afford any definite basis for comparison, but it appears
(see Balfour, vol. 11, pp. 84—86, and the extract in Hoffmann
and Schwalbe’s ‘ Jahresbericht’ for 1878, p. 222) that after the
segmentation and during the formation of the archenteron the
blastopore becomes distinct, first at its dorsal lip and then in
its whole circumference ; it ultimately closes, and Salensky (44)
states that the anus is produced afterwards in the situation
which was first occupied by the blastopore. There is, however,
no evidence as to whether the blastopore in Ganoids closes
from before backwards, as in Teleostei and Elasmobranchii and
Cyclostomes, or from behind forward as in the Rana tem-
poraria,

The primitive streak of the Amphibia appears during the
period of extension of the archenteron. It is formed by a fusion

480 ARTHUR ROBINSON AND RICHARD ASSHETON.

of the layers in the lips of the blastopore and by concrescence
of the lips of the blastopore, either from before backwards as in
Triton (Goette, 15), or by fusion of the lateral lips of the
blastopore between the neurenteric canal and the anus, arriv-
ing at its completion on the obliteration of the neurenteric
canal, as in Bombinator (15) and Amblystoma punctatum
(39); or it may be formed, as we have already shown in Rana
temporaria, by fusion of the lips of the blastopore from behind
forwards.

In Teleosteans and Cyclostomes it is formed by fusion of
the blastoporic lips from before backwards, also whilst the
archenteron is being formed and extended, and in Elasmo-
branchs by fusion of the lateral lip of the blastopore behind
the neurenteric canal.

In the Sauropsida and Mammalia we have no definite proof
of a primary blastoporic opening, for the aperture so named
by van Beneden in the rabbit (4), and by Selenka in the
opossum (50), is not yet definitely located ; but the primitive
streak in these Vertebrates is readily comparable with the
secondary condition of the Amphibian blastopore as formed in
Rana temporaria. It is an area of fusion of the germinal
layers which afterwards becomes perforated by the formation
of the anus. The appearance of the fused area at a compara-
tively late stage in reptiles, birds, and mammals—that is, after
the archenteron is well established (if we except Kupffer’s
views on the function of the archenteron of reptiles, according
to which the primitive gut is produced by invagination)—
throws but little difficulty in the way of the comparison, for it
is most probably a mere heterochronous displacement asso-
ciated with the precocious segregation of the hypoblast in the
higher Vertebrates.

Therefore, if we use the term primitive streak in the sense in
which it is used in the description of the avian blastoderm—
that is, as a term which signifies an area of fusion of the blas-
todermic layers which is continuous laterally and posteriorly
with the separated epiblast, mesoblast, and hypoblast, and is
in front continuous with the neural plate and chorda, and

FORMATION AND FATE OF THE PRIMITIVE STREAK. 481

which is perforated posteriorly by the anus, and may be per-
forated, more or less completely, anteriorly by the neurenteric
canal,—then we are bound to admit that this region and the
corresponding areas in mammalian and reptilian blastoderms
are the homologues of the primitive streak of Rana tempo-
raria; and conversely we are forced to conclude that, as the
primitive streak of Rana temporaria is formed by the linear
fusion of the undifferentiated area in the lips of the blastopore,
the primitive streak of the Sauropsida and Mammalia is homo-
logous with the fused lip of the blastopore of Rana.

Strictly speaking, therefore, the typical primitive streak is
an area which extends antero-posteriorly from the point at
which the fusion of layers commences, in front of the anterior
lip of the blastopore (fig. 10, Z), to the point at which the
fusion of layers terminates behind the posterior lip of the
blastopore (fig. 10, Z’) ; and laterally from the termination of
the fusion on the left lip of the blastopore (fig. 10, ZZ’), to
the termination of the fusion of the layers in the right lip of
the blastopore (fig. 10, ZZ).

If this is the case, then it is evident that the primitive streak
of Bombinator, Triton, and Petromyzon, as usually described,
is not homologous with the primitive streak of Rana tempo-
raria, the Sauropsida and Mammals, but only with that
portion of it which lies in front of the anus. And it is also
evident that the primitive streak of Teleosteans and Elasmo-
branchs, after the complete closure of the blastopore, is the
exact homologue of the typical primitive streak as formed in
birds.

With regard to the Ganoids, it is only possible to say that
they seem to closely resemble the Teleosteans. It will be
noticed that we have made no reference to Amphioxus. With
regard to this somewhat anomalous Vertebrate, we can only
observe that so far as the observations of Hatschek (16) and
Kowalevsky (82) go, there is no primitive streak formed,
unless we accept in full the concrescence theory, and look
upon the dorsal axial line as its representative; but even if
we adopt this theoretical conception, of which there is no

VOL, XXXII, PART IV.——-NEW SER. KK

482 ARTHUR ROBINSON AND RICHARD ASSHETON,

positive proof, we are still impressed by the fact that the
anus is neither the remains of the blastopore, nor is it formed
secondarily by reopening of the fused lips of the blastopore,
but it appears as a new formation below and in front of the
blastoporic opening; at least Hatschek figures it in that
situation, and his description is as follows :—“ Die Unterbre-
chung der Communication zwischen Darm, und Medullarrohr
erfolgt ungefahr gleichzeitig oder auch etwas spater als der
Durchbruch des Afters. Der After bricht ventralwarts von
dieser Communications-dffnung, die den letzten Rest des
Gastrulamundes reprisentirt” (16, p. 79).

The anus of the frog and many other Vertebrata bears a
similar relation to the neurenteric canal, but in no other Ver-
tebrate except Amphioxus is the anus a distinctly new forma-
tion ; on the contrary, it is very evidently an aperture formed
by the reopening of a temporarily closed orifice, or by perfo-
ration of the homologue of that orifice. In view of this fact,
it seems very probable that future observations will modify
Hatschek’s results, and remove the contradiction which at
present exists between Amphioxus and other Vertebrata. If,
however, this proves not to be the case, it will then have to be
decided whether the condition which is found in Amphioxus,

or that which is so general amongst the other Vertebrata, is
the more primitive.

The Fate of the Primitive Streak in the Frog.

In discussing the primitive streak of the frog it will be
convenient to consider separately—

(1) The fate of the ventral moiety.

(II) The fate of the dorsal moiety.

Goette (15) has pointed out that in Petromyzon the homo-
logue of the primitive streak must be the whole rim of the
blastopore. Apparently, however, he does not extend the
same reasoning to the case of the frog. For in that case,
according to his description, and to the figure he gives, in
which he indicates the position of the primitive streak by a
bracket, he does not include the ventral lip of the anus,

FORMATION AND FATE OF THE PRIMITIVE STREAK. 488

To quote his words, “Das Homologon des Primitifstreifs
ist dat die ventrale Halfte des Schwanzes von seiner Spitze
bis zum After” (Haut, ventrale Halfte des Schwanzdarms und
der Mesoderm platten, Hinterwand des Aftersdarms).

It is a question how far we ought to speak of the deriva-
tives of the primitive streak being “das Homologon”? of the
primitive streak. Is it quite correct to say that the adult
frog is the homologue of the egg from which it has been
developed ?

However, it is clear that Goette means that the ventral half
of the tail is derived from the primitive streak. Here we
must again differ from him.

As the result of our observations we conclude that not
only the ventral half of the tail is derived from the
primitive streak, but also the dorsal half as well,
with the exception of nearly the whole of the skin
of the tail.

This we shall hope to show while considering the fate of
the dorsal moiety of the primitive streak,

We now proceed to describe the fate of the ventral moiety
subsequent to the stage corresponding to figs. 25, 15, 16, 17.

Figs. 20, 21, 22, are camera drawings of sections of the
posterior end of a 44 mm. tadpole, taken at levels correspond-
ing to those of sections 15, 16, and 17 respectively (vide lines
20, 21, 22 in fig. 26, and lines 15, 16, 17 in fig. 23).

In each case the drawings have been made by camera, and
from unstained sections, and the natural appearance as
regards tint and shape has been reproduced as nearly as we
were able to reproduce it.

In figs. 20, 21, and 22, there is no trace of a primitive
streak, but all three germinal layers are now distinct and
separate. In fig. 22 the mesoblast WE may be said to be still
fused with the epiblast HP’, but it is, nevertheless, quite
distinct in character of cell, and in degree of pigmentation.
In the two other sections, figs. 21 and 20, the mesoblast has
become entirely separated from the epiblast.

By comparing these three sections with the sections from

484. ARTHUR ROBINSON AND RICHARD ASSHETON.

which figs. 15, 16, 17 were drawn, the actual fate of the
primitive streak in this region may be fairly easily deter-
mined.

In comparing fig. 20 with fig. 15, the change that has
taken place in the former seems to be of this nature. The
three layers, epiblast, mesoblast, hypoblast, instead of being
“fused,” are now (fig. 20) easily distinguishable and com-
pletely apart.

This fusion of layers we must most probably interpret as
indicating an area of proliferation of cells, which indeed is a
characteristic feature of a primitive streak; and when this
proliferation of cells ceases, the primitive streak may be said
to be no longer of physiological importance, though the area
of its former extension, being of morphological interest, should
be noted.

Thus, if we are regarding the primitive streak from a
physiological point of view, we must say that this portion has
now ceased to exist. If we regard it from a morphological
point of view, we may say that this portion has ceased to be
‘functional,’ but, nevertheless, includes the area in the
bracket labelled PS in fig. 26 as being within the limits of the
original extension of the primitive streak. This portion we
have attempted to render more distinct in fig. 26 by a different
method of shading and crossed lines, PS’ PS”.

The fate of the ventral moiety may be understood by
reference to fig. 26, combined with a comparison of figs. 20,
21, 22 with figs. 15, 16,17. By words we may explain its
fate by saying that the ventral moiety of the primitive streak
splits up into portions of the three germinal layers.

(I) Laterally and ventrally into (a) the posterior ex-
tremity of the mesodermal plate.

(II) In the median plane into two layers, (4) an outer or
epiblastic layer, forming part of the skin, and (c) an inner or
hypoblastic layer of large darkly pigmented cells, fig. 20, HY’,
forming the hind wall of the rectal spout.

The difference in character of cell between the posterior
wall and anterior wall of the rectal spout is very marked, as

FORMATION AND FATE OF THE PRIMITIVE STREAK, 480

shown especially well in fig. 20. This feature is not so marked
in stained specimens, where the degree of pigmentation does
not show up so well.

This description so far agrees with Goette (15), and
Schwarz (47), who follows Goette in this respect, except that
we include the ventral lip of the anus within the primitive
streak, which apparently these authors do not, though for
what reasons it is not easy to understand.

Fate of the Dorsal Moiety of the Primitive
Streak, and Development of the Tail.

In the dorsal moiety the fate of the primitive streak is
different.

Instead of splitting up, it remains as a proliferating area;
or according to our definition above, this portion of the primi-
tive streak remains for a longer time “ functional.”

The relation of the neural folds to the blastopore, that is
to say to the primitive streak, must be necessarily considered
in connection with the fate of the dorsal moiety of the
primitive streak.

Reference to fig. 14, which passes through the rapidly
closing blastopore (BL’), conclusively proves that there is here
no trace of neural folds. The epiblast shows no signs what-
ever of any thickening, the mass of cells being merely the
fused layers—that is to say, the undifferentiated cells at the
lips of the blastopore—of typical primitive streak.

There is no trace of neural folds—that is, of thickened
epiblast separate from mesoblast—until several sections an-
terior to the still open portion of the blastopore. That is to
say, that which from a surface examination appears to be
the lower end of the neural folds, is really the dorsal portion
of the lateral lips of the blastopore folded up along with the
true neural folds.

If this were not so, there would be a sudden break at the
posterior end of the neural folds; and the neural canal, if it
opened anywhere, would open to the exterior dorsal to the blas-

486 ARTHUR ROBINSON AND RICHARD ASSHETON,

topore, and not into the blastoporic canal, as is well known to
be the actual case.

This we have tried to make clear by the diagrams, fig. 18
and fig. 19.

In these diagrams the external surface of the neural plate
has been dotted NP. The space BR between the two small
concentric circles represents the position of the fused layers—
that is to say, the lips of the blastopore at the time of the first
formation of the blastopore.

The shaded space BR’ represents the position of the same
fused layers in fig. 18 at the time of the complete formation of
the neural plate, but before it has commenced to fold up; in
fig. 19 after the neural plate has become completely folded,
and its outer lateral edges are just meeting.

BL’ is that portion of the blastopore which remains open
longest.

The two asterisks in fig. 18 mark the two lines of epiblast
immediately adjoining the lateral edges of the neural plate.

When the neural plate has become folded, and when its
lateral edges have met and fused, and separated from the
adjoining epiblast, the lines of epiblast indicated by the asterisk
will have fused and made good the gap which would otherwise
be caused in the skin by the separation from it of the neural
plate.

In fig. 19 the folds are represented as having nearly met,
so that the outer surface (dotted) of the neural plate is now no
longer seen, except a narrow strip through the now rapidly
approaching lateral edges of the neural plate.

The daggers similarly mark two spots in the epiblast
adjoining the fused layers at the outer edges of the lateral
lips of the still open portion of the blastopore.

In fig. 18 the relation of the neural plate to the dorsal lip
of the blastopore—that is to say, to the dorsal end of the
primitive streak—is clearly represented.

In fig. 19 the folding up of the neural plate is nearly
complete, and with it the dorsal part of the primitive streak
—that is to say, the dorsal portions of the lateral lips of the

FORMATION AND FATE OF THE PRIMITIVE STREAK. 487

blastopore have been folded up along with the neural folds,
and the adjoining areas of epiblast marked by the daggers
will shortly meet and fuse, just as will the areas of epiblast
adjoining the lateral edges of the neural plate marked by the
asterisks.

Similarly, just as after the meeting and fusing of the
epiblast along the lateral edges of the neural plate has taken
place, and just as the neural plate—or tube, as it now will be
—separates from and lies within the skin, so also will that
portion of the primitive streak separate from the skin and
come to lie within the embryo.

The posterior or ventral portion of the primitive streak
does not become folded in this way, for by the time the folding
of the dorsal portion has been completed and the separation
from the skin has taken place, the lower or ventral portion
has, as we have described above, split up and ceased to exist
as a ‘functional ”’ primitive streak.

By this means the primitive streak loses all direct con-
nection with the surface epiblast, though the connection of the
primitive streak with the surface epiblast may be said to be in
reality retained by the direct continuity of the cells of the
primitive streak with the interior lining (i. e. epidermic
epiblast) of the neural tube.

The semi-diagrammatic sagittal sections, figs. 24, 25, and 26,
will aid in rendering clearer the above statements.

Fig. 24 is an early stage, only a little older than the stage
represented by fig. 18.

The “primitive streak,” or area of fused layer which is
cut through in these three sections, is shown by a diagonal
shading.

In fig. 24 the mass of primitive streak is seen to be much
greater ventrally than dorsally ; the extent of closure of the
original blastopore is expressed by the distance between 2 and
the inner edge of the lip bounding ventrally the still open por-
tion of the blastopore.

In fig. 25 the neural plate has become folded upon itself
and fused, but the neural tube so formed has not yet separated

488 ARTHUR ROBINSON AND RICHARD ASSHETON.

from the skin,—it is, in fact, on the point of so doing; it there-
fore represents a stage rather later than that of fig. 19.

Ventrally, the primitive streak isin the middle line dividedinto
a post-anal (PS’) and pre-anal portion (PS”’) by the deepening
of the primitive groove at one spot (A), which shortly after
completely perforates and forms the permanent anus. As we
have stated above, we regard this as practically a reopening of
the ventral part of the original blastopore. This is also shown
diagrammatically in fig. 19, A.

Dorsally, we get a portion of primitive streak (PS”) continu-
ous with the dorsal wall of the archenteron (HY), the noto-
chord (CH), and floor of the neural tube, which is the actual
dorsal lip of the blastopore, and is exactly the same as that
portion of primitive streak which bears the same relations to
the same structures—dorsal wall of archenteron (HY), noto-
chord (CH), floor of neural tube (VP)—ain fig. 24.

Externally to this and posteriorly the section cuts another
portion of primitive streak (PS*), which is the dorsal part of
the lateral lips of blastopore which have been folded up along
with the neural folds, as we have already described, and as we
have endeavoured to represent diagrammatically in fig. 19.

By this means a canal is formed leading from the archenteron
to the neural tube, known as the neurenteric canal, which
is therefore the anterior portion of the blastopore
(BC), together with the canal (PSC) bounded ventrally
by the anterior portion of the primitive streak (PS”),
and laterally and dorsally by the folded-over anterior
portions of the lateral lips of the blastopore (PS").

At this period there is still a direct passage to the exterior
from the archenteron, as well as into the neural tube; but as
the folding up of the dorsal portion of the primitive streak
progresses, combined probably with the continued concrescence
of the more ventral portion of the lateral lips of the blastopore,
this external opening—that is to say, that portion of the original
blastopore which apparently remains open for the longest
period—is entirely and finally closed from the exterior.

By the time this has occurred, or shortly after, the neural

FORMATION AND FATE OF THE PRIMITIVE STREAK. 489

tube has become separated from the skin, and with it the dorsal
portion of the primitive streak.

The primitive streak should, however, still be connected
with the skin ventrally by that portion which does not become
folded up and nipped off as does the dorsal portion, together
with the neural tube.

Possibly this connection may exist for a very short time,
but practically the separation from the skin of the dorsal
moiety takes place contemporaneously with the splitting up of
the ventral moiety, so that the space between the dorsal moiety
of the primitive streak and the skin from which it has separated,
and the space between the posterior wall of the rectal spout
and post-anal gut on the one hand, and the skin at the same
level caused by the splitting up of the primitive streak of that
area, become confluent at the point in fig. 26 just where the line
PS” crosses the space between primitive streak and skin.

Thus it comes about that the dorsal portion of the primitive
streak, which remains “ functional’’ and gives rise to the
greater part of the tail, comes to lie entirely within the
embryo.

Fig. 26 represents the stage at which the tail has definitely
begun to grow. The dorsal moiety of the primitive
streak is seen to be lying within the embryo, and it is
by proliferation of its cells that the whole of the tail
is formed with the exception of the skin. In no section
after (the stage shortly antecedent to this) have we been able
to see a fusion between the skin and the primitive streak,
though the skin lies closely over the primitive streak.

The skin would seem to grow, not at any one point, but over
its whole surface, on account of the pressure caused from within
by the growth of the main axis of the body, in response to which
the skin must either grow or rupture.

To a certain extent the skin of the middle line of the ventral
surface of the tail is derived from the primitive streak ; that is,
from the ventral moiety by the splitting up of the latter. This
portion is distinguished in fig. 26 by a different mode of shading ;
but with the exception of this, which we think extends only a

490 ARTHUR ROBINSON AND RICHARD ASSHE&TON,

short way, no part of the skin of the tail is derived from the
primitive streak.

The neurenteric canal is undoubtedly to be regarded as the
most dorsal part of the blastopore; and, although it remains
open for a short time after the commencement of the tail, it
gradually closes, as we have shown in the semi-diagrammatic
figure, fig. 26, NU. This closing may, perhaps, be not incorrectly
spoken of as the continuation and completion of the concres-
cence from behind forwards which caused the closure of the
ventral portion of the original blastopore.

As far as we have been able to observe, the post-anal gut
only exists during the persistence of the neurenteric canal.

In fig. 26 the part of primitive streak PS‘’ has been drawn
too large proportionately to the part PS”.

Conclusions as regards the Primitive Streak; its
Origin and Fate.

A structure exactly comparable to the primitive streak in
the chick, median and grooved, is formed in the frog (Rana
temporaria) by concrescence of the lips of the blastopore
from behind forwards.

The anus perforates the posterior or ventral end of the
primitive streak, being a deepening of the primitive groove.
It may, therefore, be regarded as the reopening of the most
ventral part of the blastopore.

The portion of the primitive streak with which the dorsal
wall of the archenteron, notochord, and floor of the neural
tube are continuous is the most anterior limit of the primitive
streak, and is the dorsal or most anterior lip of the blastopore.

The neurenteric canal, which is bounded anteriorly by the
dorsal lip of the blastopore, is therefore the most anterior por-
tion of the blastopore.

The ventral moiety of the primitive streak shortly after the
perforation of the anus ceases to exist, or, as we have preferred
to term it, ceases to be ‘‘ functional,” and splits up.

The dorsal moiety of the primitive streak becomes folded
upon itself like, and along with, the neural plate, and becomes

FORMATION AND FATE OF THE PRIMITIVE STREAK. 491

separated from the skin, and, remaining “ functional,” gives
rise to the whole of the tail with the exception of the greater
part of the skin.

We cannot find at any time any trace of blastopore or
primitive streak anterior to any part of the neural plate
or tube.

The Formation and Separation of the Mesoblast.

After the transformation of the superficial layer of the
yolk-cells into epiblast is completed the remainder of the
yolk may be looked upon as hypoblast and modified hypoblast,
for it eventually becomes transformed, partly into the true
hypoblastic lining of the enteric cavity, and partly into meso-
blast and chorda. It is separated from the epiblast, except at
the margins of the blastopore, and it never again, in Rana
temporaria, becomes fused with the epiblast in front of the
blastopore, after the manner described by Schultze (45) in
Rana fusca.

During the completion of the archenteron, and before the
blastopore is closed, the mesoblast, in front of the blastopore,
begins to separate from the true hypoblast along the dorso-
lateral aspects of the archenteron by a process of delamination.

A slit-like space, which rapidly distends, appears between
the hypoblast and mesoblast in these regions (fig. 12, H),
whence it gradually extends both towards the dorsal and
towards the ventral middle lines; but considerably before
these clefts reach near the mid-dorsal line two sagittal clefts
appear. The latter clefts separate the chorda in the middle
line from the mesoblastic plates laterally (fig. 12, S, and
fig. 6, S).

At this period the chorda is continuous with the hypoblast,
and the mesoblastic plates and the hypoblast are still fused just
outside the chorda, as well as on the ventral aspect.

Along the lines of attachment of the mesoblastic plates to
the hypoblast, at the sides of the chorda, slight depressions are
noticeable (fig. 6, D), which appear to indicate a continuation

492 ARTHUR ROBINSON AND RICHARD ASSHETON.

of the archenteric cavity into the mesoblast in the manner
suggested by O. Hertwig (21).

Eventually the mesoblast in front of the blastopore is entirely
separated from the hypoblast, first dorsally and then ventrally,
the ventral fusion remaining in the region of the liver for a
considerable time.

These changes do not take place simultaneously, but appear
to proceed from before backwards, so that the mesoblast is still
adherent to the chorda and the hypoblast, in the posterior
portion of the embryo, for a short time after the separation
has been completed more anteriorly.

The Primitive Streak Mesoblast.

The fusion of all the germinal layers in the lips of the blasto-
pore continues after the margins of the orifice have concresced,
and there is, therefore, behind the neurenteric canal an axial
rod of tissue, which is grooved upon its upper surface, and
which is not distinguishable into definite layers (fig. 15).

We have not been able to find in Rana temporaria,
before the formation of the anus, any separation of the con-
stituent parts of this axial rod into layers in the manner de-
scribed and figured by Erlanger (10a). On the contrary, we
find that the anus is formed as a perforation through the fused
mass. But, after the separation of mesoblast from hypoblast
has extended as far as the margins of the primitive streak, we
find that the lower cells of the mesoblast are more loosely
arranged, and that they are of more rounded form than the
more superficial cells. When these lower more rounded cells
are followed towards the streak they are seen to be continuous
with the hypoblast of the primitive streak. The upper and
denser layer of the mesoblast, traced in the same direction, is
found to be continuous with the epiblast (fig. 15).

These appearances suggest the idea that the mesoblast of
this region is formed partly from the epiblast and partly from
the hypoblast, but there is no definite proof that such a double
formation actually occurs.

FORMATION AND FATE OF THE PRIMITIVE STREAK. 493

The Chorda Dorsalis.

We have already stated that in Rana temporaria the
chorda is formed from the yolk-cells which lie beneath the
neural groove. It is unimportant whether we consider these
cells to be mesoblastic or hypoblastic, but we wish to emphasise
the fact that the chorda in Rana temporaria is not formed
from a layer of mesoblast which has previously separated from
the hypoblast. We have been unable to find in Rana tem-
poraria any appearances which would justify the conclusions
which Schultze forms from his observations upon Rana fusca,
i. e. § Dass schon auf dem Stadium der beginnenden Gastrula-
tion in der dorsalen Urdarmwand drei Keimblatter existiren”
(45).

Ata later stage, however, just about the time when the
medullary groove is only appearing in the posterior part of the
embryo, we have noted, under the low power, an apparent sepa-
ration of the mesoblast as a distinct layer across the mid-
dorsal line ; but this appearance we have only seen in ova which
have been treated with staining reagents, and on examination
with higher powers we have never been able to convince our-
selves that the line of separation was normal, for we have
invariably found in the space between the mesoblast and hypo-
blast broken fragments of cells, and we are therefore inclined
to the belief that the separation of the layers was in these cases
artificial, and that it had been produced by the reagents used
in staining.

In unstained specimens the appearances seen in the mid-
dorsal line at the time of the appearance of the medullary
furrow are represented in fig. 12. The epiblast and mesoblast
are separated by a distinct space(#). Laterally the hypoblast
and mesoblast are also distinct from each other (H), but along
the dorsal wall of the archenteron the mesoblast is neither
separated from the chorda, though there are traces of the com-
mencement of the separation on the right side (S), nor from
the hypoblast, and the chorda is fused both with mesoblast
and hypoblast,

494, ARTHUR ROBINSON AND RICHARD ASSHETON.

At a later period the mesoblastic plates become entirely
separated from the dorsal hypoblast, and afterwards the chorda
also is separated ; it then forms a distinct rod, which lies free
below the floor of the neural groove and above the dorsal hypo-
blast except at its posterior extremity, where it terminates in
the mass of cells which form the anterior wall of the neuren-
teric canal.

Before the commencement of the separation of the chorda
from the hypoblast there are, here and there, appearances
which indicate a projection of the archenteric cavity into the
mass of chorda-cells, in some cases merely as a cleft-like space,
in others as a more distinct diverticulum.

In Rana fusca O. Schultze found a peculiar secondary
fusion of the mesoblast with the epiblast in front of the dorsal
lip of the blastopore, “‘ bildet sich gegen ende der Gastrula-
tion von der Mitte der dorsalen Urmundlippe aus eine nach
dem Kopfe hin vorwirts schreitende lineare Verwachsung des
dusseren und mittleren Blattes aus” (45). This line of fusion
he compares to the primitive streak.

We have already said that in Rana temporaria there is
no true layer of mesoblast along the dorsal axial line in front
of the blastopore, and that in this situation the chorda is
formed by differentiation of the yolk-cells; it follows, there-
fore, that if the fusion occurs in Rana, it will be between the
last-mentioned cells and the epiblast.

We have examined our sections carefully with reference to
this point, and we find that the epiblast after its separation
from the yolk-cells does not again fuse with the subjacent
layer in the dorsal axial line in front of the blastopore.

It will be remembered that the separation of the epiblast
terminates at a distance of ‘352 mm. (p. 462) in front of the
dorsal lip of the blastopore, that is, at the commencement of
the primitive streak. In other words, the dorsal lip of the
blastopore has an antero-posterior length of *352 mm., and
during the early stages this length does not increase; there-
fore there is no forward extension of the primitive streak in
front of the blastopore.

FORMATION AND FATE OF THE PRIMITIVE STREAK. 495

The mere fact that in Rana temporaria the distance
from the dorsal margin of the blastopore to the point at which
the epiblast is separated as a distinct layer remains constant
throughout the earlier stages is not a proof that the blasto-
poric lips undergo no concrescence from before backwards, but
it is a proof that the epiblast after its separation does not again
fuse with the subjacent layer.

The Formation of the Celom.

Along the line of attachment of the mesoblastic plates to
the hypoblast, at the sides of the chorda, it is possible to find
at irregular intervals small evaginations from the archenteron
(fig. 6, D), but these small diverticula cannot be traced for any
great distance into the mesoblastic plates.

They remain for a time as blind diverticula, and then they
disappear. So far as we have been able to discover they do
not communicate with the celom, and the celom is not
formed by extension of the archenteric diverticula, as in
Amphioxus, but by a splitting of the -mesoblast which first
occurs laterally, and then extends dorsally and ventrally as in
the higher Vertebrata.

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496 ARTHUR ROBINSON AND RICHARD ASSHETON.

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Scuwarz, D.—“ Untersuchungen der Schwanzendes bei den Embryonen
der Wirbelthiere,” ‘Zeitschrift fiir wissenschaftliche Zoologie,’ xlviii,
1889, pp. 191—223, plates xii—xiv.

Scort, W. B., anp Osporn, H. F.—‘On some Points in the Early
Development of the Common Newt,’’ ‘Quart. Journ. Mier. Sci.,’
N. S., 1879, pp. 449—472, plates xx and xxi.

Sepcewick, A.—‘‘ On the Origin of Metameric Segmentation, and
some other Morphological Questions,” ‘ Quart. Journ. Micr. Sci.,’
N.S., 1884, pp. 24—42, plates ii and iii.

SELENKA.—‘ Studien tiber Entwickelungsgeschichte der Thiere.—Das
Opossum,’ Wiesbaden, 1886.

SipeBoTHAM, H.—‘‘ Notes on the Fate of the Blastopore in Rana
temporaria,” ‘ Quart. Journ. Mier. Sci.,’ N.8., 1888, pp. 49—54,
plate v.

FORMATION AND FATE OF THE PRIMITIVE STREAK. 499

52. Spencer, W. B.—‘‘Some Notes on the Harly Development of Rana
temporaria,” ‘Quart. Journ. Micr. Sci.,’ N. §., Supplement, 1885,
pp. 1283—137, plate x.

58. Straut, H.—“ Beitrage zur Entwickelung der Reptilien,’ ‘ Archiv
fiir Anatomie und Physiologie—Anatomische Abtheilung,’ 1883, pp.
1—48, plate i.

54. “ Beitrage zur Entwickelung von Lacerta agilis,” ‘Archiv fiir Anatomie
und Physiologie—Anatomische Abtheilung,’ 1882, pp. 242—278,
plates xiv and xv.

55. “ Zur Bildung der Cloake des Kaninchen-embryo,” ‘ Archiv fiir Anatomie
und Physiologie—Anat. Abtheil.,’ 1886, pp. 156—168, plate iv.

56. Wetpon, R.—“ Notes on the Early Development of Lacerta muralis,”
‘Quart. Journ. Micr. Sci,’ N.S., xxiii, 1883, pp. 134—150, plates
iv—vi.

57. WencxeBacu, K. F.—‘“ Der Gastrulationsprocess bei Lacerta
agilis,” ‘ Anatomischer Anzeiger,’ 1891, pp. 57—66 and 72—77.

EXPLANATION OF PLATES XXXIV & XXXV,

Illustrating Dr. Arthur Robinson’s and Mr. Richard Assheton’s
paper on “The Formation and Fate of the Primitive Streak,
with Observations on the Archenteron and Germinal
Layers of Rana temporaria.”

Alphabetical List of Reference Letters for all the Figures.

A. Anus. a. Anterior extremity of archenteron. 4 P. Post-anal gut.
AR. Archenteron. 42’, That portion of the archenteron the posterior wall of
which is formed by the closure of the blastopore. B, B’. Rim of blastopore lip.
4. Dorsal lip of blastopore. BC. Blastoporic canal. BZ. Blastopore. BL’.
Portion of original blastopore not yet closed. 2B. Original position of fused
layers, that is to say, lips of blastopore at the time of the first formation of
the circular blastopore. BR’. Position of fused layers or lips of blastopore
after concrescence to a greater or less extent of the lateral lips of the original
blastopore. BR’. The dorsal portion of the lateral lips of the blastopore folded
over with the neural plate, so that that which was their ventral or inner surface

500 ARTHUR ROBINSON AND RICHARD ASSHETON.

is now seen from without. c. Ventral lip of blastopore. CH. Notochord.
D. Diverticulum from archenteron towards mesoblast. d. Line of section of
Fig. 2. #. Space between epiblast and mesoblast. #P. Epiblast. #P’. Epi-
dermic layer of epiblast. #P”. Nervous layer of epiblast. H. Slit between
hypoblast and mesoblast. HY. Hypoblast. MY’. Primitive streak of hypo-
blast. J. Line of fusion of the folded-up edges of the neural plate. WZ.
Mesoblast. J/ 2’. Epiblastic or outer layer of mesoblast of primitive streak.
M&". Hypoblastic or inner layer of mesoblast of primitive streak. VC.
Cranial nerve. NG. Neural groove. WP. Neural plate. NS. Roof of
spinal cord. WZ. Central canal of spinal cord. MU. Neurenteric canal.
PG. Primitive groove. PS. Primitive streak. PS’. The part of the
primitive streak ventral to the anus, i.e. ventral lip of original blastopore.
PS". The part of the primitive streak which is continuous with the posterior
end of the notochord, i. e. dorsal lip of original blastopore. P.§'’. The part
of the primitive streak which lies between that portion of the blastopore
which remains open longest and the anus, i. e. the greater part of the fused
lateral lips of the original blastopore. P.S%. The part of the primitive
streak which is folded up with the neural folds and forms the posterior wall
of the neurenteric canal. PS'c. The dorsal portion of the neurenteric canal.
S. Line of separation of mesoblast from chorda. SG. Segmentation cavity.
x. The lowest extremity of that part of the archenteron which is bounded
posteriorly by the fused lips of the blastopore. Y. Yolk-cells. YP. Yolk-
plug. 27,2’, ZZ, ZZ’. Furthest points from the edge of the blastopore, at which
a distinct fusion of germinal layers is perceptible. * *. Two points of epiblast
beyond the neural plate, which meet and fuse when the folding up of the
neural plate is completed. ++. Two points of the epiblast beyond the dorsal
portion of the primitive streak, which meet and fuse when that portion of the
primitive streak is folded up with the neural plate.

Fic. 1.—Sagittal section of a portion of the ovum represented in Fig. 4,
showing the dorsal lip of the blastopore and the area of fusion in front of it.
The cavity of the archenteron, and the pigmented streak in front of it which
indicates the line of extension. x 50.

Fic, 2.—<A transverse section through the pigmented area in front of the
archenteric cavity. The section is taken in the direction of the line d in
Fig.l. x 115.

Fic. 3.—A sagittal section through the posterior lip of the blastopore of
the ovum represented in Fig. 4. x 115.

Fig, 4.—A sagittal section of an ovum at the period of completion of the
blastopore. x 23.

Fie. 5.—A sagittal section of an ovum after the disappearance of the seg-
mentation cavity, and when the blastopore is partially closed. x 23.

Fic, 6.—A portion of a transverse section of au ovum after the separation

FORMATION AND FATE OF THE PRIMITIVE STREAK. 501

of the chorda from the mesoblast, but before the complete separation of the
mesoblast and chorda from the hypoblast. x 138.

Fic. 7.-—-A nearly horizontal section through the posterior part of an
embryo in which the blastopore had only slightly diminished. The section is
taken through the centre of that part of the original blastopore which stil.
remains open. Drawn with camera, x 30,

Fic. 8.—A nearly horizontal section through the same embryo, but taken
ventrally to the existing blastopore through the ventral portion of the original
blastopore, the lateral lips of which have coalesced. x 30.

Fic. 9.—A nearly horizontal section through the same embryo, but taken
ventral to both Figs. 7 and 8. It was the forty-first section ventral to that
of Fig. 7. x 30.

Fic. 10.—A diagram to illustrate the changes that had taken place in the
shape of the blastopore, in the embryo of which Figs. 7, 8, and 9 are
sections. The lines numbered 7, 8, 9 are drawn about the levels at which
the sections bearing those numbers are taken. The space lettered BR, en-
closed by the two small concentric circles, represents the position and extent
of the fused layers of the blastoporic rim at the time of the first formation
of the circular blastopore or anus of Rusconi; the shaded space B R’ represents
the position and extent of the fused layers at the stage of the embryo of
which Figs. 7, 8, and 9 represent sections. The line lettered z represents
the extent of the closure of the ventral portion of the lateral lips up to this
stage. The letters 7, Z', ZZ, ZZ’, refer to the same spots as those letters do
in Figs. 5 and 7.

Fic. 11.—A surface view of the posterior end of an embryo, in which
the blastopore has diminished considerably, but in which it is still circular.
The neural plate, V P, is distinctly visible. The line P Gis the primitive
groove, at this period only with difficulty made out in surface view. In the
figure it is drawn too distinctly. The levels at which the sections Figs. 12
and 13 were taken are indicated by the lines numbered 12 and 13. These
sections (Figs. 12 and 13) were not taken from the same embryo from which
Fig. 11 was drawn. This drawing was made from a living embryo.

Fie. 12.—A transverse section through the posterior portion of the neural-
plate region, showing the commencing separation of the mesoblast from the
chorda. x 115.

Fic, 13.—A transverse section through the posterior part of the primitive
streak of an embryo, about the same age as that represented in Fig. 5. The
section is taken along the line z in the latter figure. x 50.

Fies. 14, 15, 16, and 17 are horizontal sections through the posterior end
of the same embryo, the stage being precisely the same as that of Fig. 23,
on which the levels of the four sections are shown by the lines 14, 15, 16,
and 17. Fig. 14 is taken through the centre of the nearly closed “ blasto-

502 ARTHUR ROBINSON AND RICHARD ASSHETON.

pore,” and the sections 15, 16, and 17 are the 16th, 32nd, and 44th sections
respectively below section 14.

Fig. 14. A horizontal section through the posterior end of the embryo,
and therefore a section transverse to the longitudinal axis of the
primitive streak. The section is taken through the centre of the small
part of the original blastopore which is still open, which, however, is
seen to be on the point of closing. The three germinal layers are
fused at the blastoporic lip, and two layers of the mesoblast may be
distinguished—a denser outer or epiblastic layer, MZ’, and a looser
inner or hypoblastic layer, MH’. The cells of the latter are further
characterised by being rounder and more deeply pigmented. Drawn
with camera, X 55.

Fig. 15. 24 mm. Tadpole. A horizontal section parallel to the preceding
section, taken through about the centre of the primitive streak. It
presents practically the same features as the preceding figure, except
that at this point the blastopore has completely closed, and the opposite
lips have fused, forming a typical primitive streak. Camera drawing,
x 55s

Fig. 16. 24 mm. Tadpole. Horizontal section through the posterior part
of the embryo. This section passes through that portion of the
primitive streak in which the anus will shortly appear. The primitive
groove is at this point very much deeper, forming that which is usually
called the proctodeum ; but complete perforation has not yet occurred.
The line of perforation is, however, foreshadowed as it were by a deep
irregular line of pigment. Camera drawing, x 55.

Fig. 17. 24 mm. Tadpole. A horizontal section of posterior end of an
embryo ventral to the anus. This is the thirteenth section behind the
anus. The primitive groove is still well marked. As there is no true
hypoblast here, there is fusion only between epiblast and mesoblast.
Drawn with camera, X 55.

Fic. 18.—A diagram to show the relation of the neural plate to the
‘blastopore” and rest of the primitive streak. The dotted area represents
the neural plate. The shaded area BR’ represents the primitive streak at
the time just prior to the commencement of the folding up of the neural
plate. The space BR between the two concentric circles represents the
original position of BR’. The asterisks denote two points in the epiblast
which will come together when the neural plate has folded up, and the daggers
denote two other points alongside the blastoporic lips which will come together
as explained in the text.

Fic. 19.—A diagram to show the relation of the neural plate to the blasto-
pore and rest of the primitive streak, and to show the way in which a portion
of primitive streak becomes folded up along with the neural folds. It will be
seen that here the folding is nearly completed; the asterisks and daggers of

FORMATION AND FATE OF THE PRIMITIVE STREAK. 503

Fig. 18 have been brought together so as to almost coincide. This figure also
diagrammatically represents the anus as a reopening of the ventral part of
the original blastopore. This diagram and the diagram of Fig. 18 represent
nearly the stages of the drawings Fig. 23 and Fig. 11 respectively.

Fie. 20.—43 mm. Tadpole. A horizontal section through the posterior end
at a level corresponding to that of Fig. 15, but of an older embryo. Instead
of the layers being fused along the middle line, they are now separated and
distinct. Camera drawing, x 55.

Fic. 21.—43 mm. Tadpole. A horizontal section through the posterior end
at a level corresponding to that of Fig. 16, that is to say, through the anus,
which is now completely formed. Camera drawing, x 55.

Fic. 22.—43 mm. Tadpole. A horizontal section through the posterior end
at a level corresponding to that of Fig. 17. These three sections, Figs. 20,
21, and 22, are from the same series. The sections are not stained, and the
drawings have been made to represent the actual shades of the three layers as
nearly as possible. Caimera drawing, x 55.

Fig, 23.—23 mm. Tadpole. A surface view of the posterior end of a
tadpole, in which the neural folds have met and are fusing. The “blastopore”
is still open, and the anus is conspicuous. The horizontal lines numbered 14,
15, 16, and 17 are drawn at the levels at which the sections were taken, from
which Figs. 14, 15, 16, and 17 were drawn. ‘The bracket, PS, includes
the dorsal and ventral limits of the primitive streak. This figure was drawn
partly from a specimen hardened in Kleinenberg’s picro-sulphuric acid, and
partly from a model constructed by pasting together in order pieces of card-
board corresponding to a complete series of camera drawings.

Fic. 24.—A sagittal section of the posterior three quarters of a frog embryo,
in which the neural plate is beginning to fold up, and in which the anus of
Rusconi has become much reduced. ‘The yolk-plug has been retracted,
and the embryo is lengthening. The extent of the primitive streak is indi-
cated by the bracket PS, and by the diagonal shading.

Fic. 25.—22 mm. Tadpole. A semi-diagrammatic sagittal section through
the posterior half of a frog embryo, of the same age as that from which Fig.
23 was drawn. ‘The extent of the primitive streak is indicated by the bracket
PS, and by the diagonal shading. The neural plate has completely folded
over, but has not yet separated from the external epiblast. This figure shows
the dorsal portion of the primitive streak folded over with the neural plate,
and with it about to be separated from the adjoining epiblast. The blasto-
pore has not quite closed in the middle line. The neurenteric canal is seen to
be formed of two portions; the ventral, being the original opening from arch-
enteron to exterior, the blastoporic canal (B.C.) or blastopore, and a dorsal,
the canal through the folded-up lateral lips of the blastopore, or

504, ARTHUR ROBINSON AND RICHARD ASSHETON.

portion of the primitive streak (PSC). The anus has not yet completely
perforated the ventral part of the primitive streak.

Fic. 26.—44 mm. Tadpole. A semi-diagrammatic sagittal section through
the posterior end of au embryo in which the tail has just begun to grow out.
The central nervous system has entirely separated from the skin, and with it
that portion of the primitive streak which was folded up with it (PS), By
this means that portion of the primitive streak (P Si’) comes to lie now within
the embryo. The ventral moiety of the primitive streak has ceased to exist
as such, and the portions derived therefrom are shown by cross-shading ; while
the dorsal portion, or still functional portion, if it may be so termed, is indicated
as in the two immediately preceding figures by diagonal shading. The bracket
PS includes the full extent of primitive streak or its derivatives.

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 505

On some Points in the Histology and Develop-
ment of Myriothela phrygia.

By

w, B. Hardy, B.Ae,

Shuttleworth Scholar of Gonville and Caius College, and Junior Demonstrator
of Physiology in the University of Cambridge.

With Plates XXXVI and XXXVII.

WuitE working at the Marine Biological Laboratory at
Plymouth during the summer of 1888 my attention was
directed to the remarkable hydroid Myriothela phrygia. I
was acquainted with Professor Allmann’s (1) memoir, and it
appeared to me that there were several points which demanded
further study—notably the development of the gonophore and
the structure of the endoderm, especially in relation to the
physiological processes taking placein the enteric cavity. My
work was continued, so far as circumstances would permit,
during the ensuing winter; but on the appearance of Korot-
neff’s account (2) of the growth of the gonophore, published in
the ‘Archives de Zoologie Expérimentale’ for that year
(1888), I discontinued it, in the hope that at some future time
I might find the opportunity of examining with something like
completeness the physiology of Myriothela. That opportunity
unfortunately has not arisen, and I am induced to publish my
results as they stand, believing that though incomplete they
contain certain points of interest.

The first complete account of the anatomy and development of

506 W. B. HARDY.

Myriothela is that of Professor Allmann, which was published
in the ‘ Phil. Transactions’ in 1875. This was followed by a
long monograph illustrated with abundant figures, and pub-
lished at Moscow (8) in 1880, unfortunately in Russian, by
Korotneff, who was the first to study Myriothela with the aid
of properly prepared sections.

In 1881 Korotneff published a further paper dealing with
the same subject (4). A copy of this I have unfortunately
not been able to obtain. The further literature of the subject
will be referred to as occasion demands in the following pages.

The work was carried out partly in the Marine Biological
Laboratory at Plymouth, and I am grateful for the many kind-
nesses experienced there. But the bulk of the work was done
in the Morphological Laboratory of Cambridge University,
and I would here thank Mr. Sedgwick for placing the resources
of his department at my service.

The general structure of Myriothela will be best learned
by reference to Allmann’s monograph, and to that I must refer
my readers, merely stating here that it is a solitary attached
hydranth. The proximal part of the body is usually bent at
right angles to the rest, is covered with a thick perisarc, and
gives origin to short processes by which it is attached to the
under side of large stones. The perisarc of the foot is repre-
sented over the rest of the body bya delicate cuticle. Follow-
ing on the foot is the middle region of the body, whence spring
numerous blastostyles. Each of these bears gonophores, male
and female, on its proximal portion, while short capitate ten-
tacles spring from the distal extremity. The blastostyles are
without a mouth.

The distal or oral region of the body of the hydranth is the
longest, and is studded with very numerous, small, capitate
tentacles to within a short distance from the mouth.

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 507

Tur Earty Staces IN THE FoRMATION OF THE GONOPHORE
AND THE ORIGIN OF THE SEXUAL ELEMENTS.

The points noticed under this heading may be regarded as
an addendum to Korotneff’s account of the development of the
gonophore (8) alluded to above. His brief and somewhat
diagrammatic account is illustrated by figures which unfortu-
nately show little attempt at histological detail.

The Structure of the Ectoderm of the Blasto-
style.—In each blastostyle two regions may be distinguished ;
a distal and shorter region bearing several small capitate ten-
tacles, and a proximal region, on the whole free from tentacles,
on which the gonophores are developed. The latter embraces
two-thirds to four-fifths of the whole length of the blasto-
style. The ectoderm of the distal portion resembles that of
the body generally, with the exception that the muscle-fibres,
which form its deepest layer, are not so strongly developed.
Its structure is shown in Pl. XXXVI, fig. 1. Like the ectoderm
of the animal generally, it is covered superficially by a well-
marked cuticle, which, when stripped off and examined with a
high power, is seen to be divided into irregular areas, doubtless
corresponding to the subjacent cell layer. This latter is com-
posed of long columnar cells, tapering at their lower end. Each
cell is about 25 long, though their dimensions possibly vary
according as the whole blastostyle is in a condition of exten-
sion or contraction. Lying here and there between the pointed
bases of these cells are scattered ganglion-cells, which are con-
nected with arich plexus of nerve-fibrils situated in the deepest
part of the ectoderm and immediately over the muscle-fibres.
The latter run longitudinally, and immediately overlie the sup-
porting lamella. Fibrils from the basal nerve plexus pass
between the columnar cells towards the surface, and, in tangen-
tial sections, are seen to form a superficial plexus between the
columuar cells.

The same type of nucleus is found throughout the whole
ectoderm. It is characterised by the presence of a distinct

508 W. B. HARDY.

nucleolus linked by scattered fibrils with irregular patches of
chromatin on their course to a deeply-staining envelope.

The cell-substance of the columnar cells is granular and
turbid, the granulation mostly being fine. Picro-carmine or
hematoxylin stains it only slightly. The ganglion-cells, on the
other hand, stain well with picro-carmine.

The ectoderm of the proximal or gonophore-bear-
ing region is vastly different from that just described. In
the first place it is much thicker and more complex, being
composed of more varied elements. The ectoderm of the
distal region is about 30 to 35 yu thick, of the body 40 to 50 n,
while that of the gonophore-bearing region varies from 50
to 70 mw in thickness. The only other region in which the
ectoderm at all approaches it in thickness is in the foot. To
this fact we will return later.

The second striking feature of the proximal ectoderm is that
its characters are not constant. It is most complex and
thickest in specimens killed in spring and early summer, while
in autumn it is not only much thinner (30 to 35 1), but also
presents the appearance of being exhausted. A comparison
of fig. 2 with fig. 4 will render this abundantly evident. The
following description applies to specimens killed in March,
April, and May.

Starting from the outside we have first a well-developed
cuticle, which overlies cells resembling the columnar cells
of the distal region, but, for the most part, shorter and
broader. They are composed of the same ill-staining granular
protoplasm, and the border between cell and cell is often so
indistinct that we might almost call this with Allmann a nucle-
ated protoplasmic layer. The proximal region is abundantly
armed with nematocysts, which, as was pointed out by Allmann,
are of two kinds. Here and there these may be seen wedged
between the columnar cells. The next following layer is a
fairly distinct one of large rounded cells engaged in the
manufacture of nematocysts. The protoplasm of these cells,
when stained with osmic acid, appears granular under
moderate magnification. With high powers (Zeiss ;';th ob.)

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 509

this is seen to be due to the presence of numberless minute
vacuoles. ach cell has embedded in its substance a hyaline
mass of uniform texture (fig. 2), which at first is deeply
placed in the cell, but as growth proceeds it is pushed to one
side (fig. 5 6), and eventually forms one of the curious rhabdite-
like nematocysts which stain so deeply with hematoxylin and
osmic acid. The first formed amorphous hyaline masses,
however, stain only lightly with osmic acid, and, whatever the
chemical change may be which alters so profoundly their
behaviour towards that reagent, it does not occur until a
relatively late period in their development. I am not at all
certain that the hyaline masses found in these cells are, in
every case, early stages in the development of nematocysts.
For many reasons I am inclined to think that they may some-
times be of the nature of reserve nutritive material. My
researches on this point are, however, incomplete, and I will
merely content myself here with noting the fact that the very
earliest change in that part of the ectoderm where a gono-
phore is about to be developed, is amongst other things a local
accumulation of the large cells bearing hyaline masses, some-
times one, sometimes two (fig. 8), while at the same time there
is a total disappearance of nematocysts from that area.

The deepest layer of the ectoderm of the gonophore-bearing
region is the most remarkable. It consists for the most part
of small rounded cells (fig. 6), either scattered about fairly
evenly, or, and more frequently, gathered into smaller or
larger clusters. If one of these clusters be examined with a
high power, the cells composing it are seen to present con-
siderable differences in size, varying from 8 uw to 12 w in
diameter. In most of the clusters, and more especially if the
blastostyle under examination be one which has scarcely
commenced to produce gonophores, few or many of the cells
will be found with two small nuclei, and the cluster will
betray other evidence of the active proliferation of its consti-
tuents (fig. 2).

In those cells with a single nucleus, that nucleus is uniformly
of about 4 mw in diameter, and, like the nuclei generally, is

510 W. B. HARDY.

characterised by the possession of an exceedingly distinct
nucleolus. The smallest of these cells are generally isolated
(fig. 6), and each consists of a nucleus surrounded by a deli-
cate pellicle of exceedingly finely granular protoplasm. The
whole cell may be only 5 w in diameter. They form the dis-
tinctive feature of the ectoderm of the proximal region of the
blastostyles, and, by their number, give to it its great thickness.
What part they play we shall see later.

The remaining constituents of the proximal ectoderm as yet
unnoticed are its nervous and muscular elements. These I
propose to mention very briefly, for they lie to a certain extent
outside the limits of the present paper. One of the most
striking features of osmic acid preparations, whether sections
or teased, are tufts of branching filaments with curious deeply-
staining matter disposed on them in irregular patches and
granules (fig. 3). In teased preparations these filaments are
seen to largely end in a thick plexus between the columnar
cells (fig. 8), but some of them also end in the cells which are
developing or have developed a nematocyst (fig. 5). Not
infrequently a filament may be seen having on its course a mass
of granular protoplasm containing a nucleus (fig. 2). Traced
downwards these filaments appear to be connected with a
deeply-placed nerve network,’ which in turn is in close relation
to the muscle-fibres which are placed immediately upon the
supporting lamella.

If the ectoderm of the gonophore-bearing region of a speci-
men killed in the autumn be examined, it will be found to
consist of externally a cuticle with columnar cells underlying
it, and then an indefinite number of ganglion-cells and cells

1 Hig. 5 is an accurate drawing made with the aid of a camera lucida
of a portion of this nerve complex, which by good fortune was isolated
in a teased osmic acid preparation. It exhibits a remarkable fact in the
arrangement of this primitive nervous system, namely, that some of the ganglion-
cells are enclosed in a fine reticulum, formed by the breaking up of filaments
derived from the general nerve network. Since this drawing was made the
remarkable researches of Golgi, Ramon y Cayal, Kolliker, and others have
demonstrated the existence of similar structures in the central nervous system
of the higher animals.

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 511

concerned in the manufacture of nematocysts (fig. 4). In
some sections scanty patches of small cells similar to those
described above may be seen, but they are now the exception
and not the rule. On the other hand, over considerable
regions the ectoderm may be even more decrepit than that de-
scribed above, for even the columnar cell layer may be imper-
fect, while the basal cells are represented largely by irregular
spaces. On the other hand, the muscular layer and supporting
lamella are proportionately better developed. Measured from
the external surface of the cuticle to the external surface of
the supporting lamella the ectoderm at this period is from 30
to 35 w in thickness.

We thus see that as the season advances and the reproductive
period comes to a close the ectoderm of the gonophore-bearing
region becomes more and more exhausted of those small cells
which are such a characteristic feature of it in the spring, and
I think there can be little doubt that their disappearance is
connected with the active formation of gonophores during the
summer months.

So far I have spoken of them merely as the small cells of the
proximal ectoderm. It is necessary to see whether these cells
are all alike, or whether they may be divided into one or more
sets differing in their function and connections. In tracing
the process of formation of the nematocysts I have said that
they first appear as a rounded hyaline mass embedded in the
protoplasm of a cell which then lies in the deeper part of the
ectoderm.

The smallest cells containing these masses may be only 10 u
in diameter ; but though they do not differ markedly from the
bulk of the small cells in point of size, they do differ in one
very important particular, namely, in the fact that they are
always connected by a delicate process with the nerve network
(fig. 5).

On the other hand, we have in addition to these small cells
which are in connection with the nerve network, and so often
contain some trace of a developing nematocyst, other still
smaller rounded cells, which appear, even with the highest

512 W. B. HARDY.

powers, to be entirely free ; and these are the elements which
occur so characteristically in little groups, the cells of which so
frequently betray signs of active proliferation. These histo-
logical facts, together with the absence of these free cells in
other parts of the body and their peculiar relation to the gono-
phores, entitle us, I think, to regard them as preformed sexual
elements.

The Earliest Stages in the Formation of the Gono-
phore and its Relation to the Process of Budding
in Myriothela.

In early spring, and before sexual reproduction has taken
place to any marked extent, specimens of Myriothela may be
found which bear buds in various stages (Pl. XX XVII, fig. 13).
These appear to be always developed just at the junction of
stolon and body. Once only have I met with a bud formed
elsewhere, namely, in the lower tentacular region. This had,
however, more the appearance of a permauent growth than of
a bud to be cast off.

The process of budding, so far as I have followed it, is a
rather remarkable one. The first stage is a modification of
the character of the ectoderm, which in the stolon and lower
part of the body is composed of very long columnar cells, re-
sembling the columnar cells of the blastostylar ectoderm in all
particulars save in their inordinate length. Lying between
the bases of these columnar cells are interstitial cells, charac-
terised by the fact that they stain more deeply with picro-
carmine. These cells appear to be partly nervous and partly
concerned in the formation of nematocysts, which, curiously
enough, are produced in limited number even under the thick
and dense perisarc of the upper part of the foot. Wherea
bud is about to be formed the ectoderm-cells lose their
defined characters, proliferate, and a bulging mass of amor-
phous tissue results. At the same time the thick supporting
lamella becomes absorbed, and the endoderm-cells likewise
proliferate and take on an amorphous character. The result
is a kind of blastema in which the limits of ectoderm and

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 513

endoderm are undistinguishable. This grows while at the
same time its elements lose their distinctness and become
highly charged with spherical masses of stored nutriment,
resembling in many particulars the nutritive spheres of the
general endoderm. As it grows it pushes the perisare before
it, and ultimately forms a rounded egg-like mass attached to
the parent body by a short thick pedicle (fig. 13). From
this the young Myriothela is developed (fig. 13). All con-
nection with the body of the parent is lost at a very early
period, almost before the bud has re-formed its ectoderm and
endoderm and enteric cavity. It remains attached to the
perisarc, however, by a sucker-like arrangement at the aboral
pole until it is fully formed.

As will be seen, the formation of a gonophore is, in its
earliest stages, essentially similar to this method of budding.
In other words, the gonophore is a true bud which, like the
other buds, is derived from a blastema formed by a fusion of
ectodermal and endodermal elements. The difference, however,
lies in the fact that in the case of the gonophore bud, after it
is a well-formed structure, a group of the primitive germ-cells
make their way into it.

The first stage in the growth of a gonophore is shown
in fig. 8. The ectoderm of the gonophore-bearing region
becomes thickened over a small surface, the increase in
thickness being due largely to an accumulation of the
primitive germ-cells, but partly to an increase in the cells
carrying hyaline masses. At the same time nematocysts dis-
appear in that region of the ectoderm, though they may occur
in their usual profusion in close proximity. In the next stage
the basement membrane is absorbed or ruptured (figs. 7 and
9), I cannot determine which, and a tongue of endoderm-
cells pushes its way into the ectoderm and through the deepest
layer. Thus the cluster of primitive germ-cells come to lie
not on its apex, but, generally, asymmetrically disposed on
one side.

The removal of the supporting lamella is, I am inclined to
think, mainly a process of solution, since scattered rounded

VOL. XXXII, PART IV.—NEW SER. M M

514. W. B. HARDY.

fragments may sometimes be seen; and further because the
muscular elements are absorbed for some little space about the
point where the rupture takes place (figs. 7 to 10).

The tongue of endoderm-cells rapidly becomes a tubular
outgrowth, and the cells at its apex lose their nutritive
spheres and become small, dense, on the whole ill-staining
cells.

At this stage it is perhaps impossible to distinguish the limit
between ectoderm and endoderm, while at the same time a
fusion of cell substance has taken place (fig. 10), so that we
have a stage closely resembling the blastema which gives rise
to the bud as described above.

Fig. 10 represents this stage, and is an accurate drawing,
made with the aid of a camera lucida, of a preparation
from a specimen killed with corrosive sublimate and stained
with picro-carmine; the whole specimen being remark-
able for the good preservation and clear definition of its
histological elements. The section figured passes rather
obliquely through the young gonophore, but the next in the
series shows that the primitive germ-cells have now travelled
in under the superficial columnar cells of the ectoderm to form
a cap for the fused ectoderm and endoderm.

The next stage to be noticed is in many respects remarkable.
By a fresh formation of supporting lamella the whole bud with
its contained germ-cells becomes separated from the maternal
tissue, while at the same time a fold of supporting lamella be-
comes formed which separates the ectodermal elements with
the primitive germ-cells from what is usually known as the
endoderm lamella (fig. 11). The endoderm lamella, there-
fore, from this time onwards, is separated from the endo-
derm of the parent by a well-defined and permanent support-
ing lamella; and it is noticeable that up to this stage and until
they degenerate the cells of the endoderm lamella are not of the
ordinary endoderm type, but resemble in every detail
the ectodermal cells of the gonophore.!

1 The columnar celis of the maternal ectoderm remain undisturbed and
unaltered by these various changes. I regard them as belonging to the

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 515

In the further history of the gonophore I am only concerned
with one point, namely, the fate of the primitive germ-cells.
I entirely agree with Allmann that in their earliest history it
is impossible to distinguish between male and female gono-
phores. Position does not help one, for, as has been recognised
before, both male and female elements are produced on the
same blastostyle, and to a certain extent indiscriminately, the
same transverse section frequently passing through both male
and female gonophores. Very soon, however, the male gono-
phores become distinguished by the rapid proliferation of their
generative elements.

In the female gonophore at some period, often relatively late,
two or three of the generative cells become larger and more
prominent than the others. The period at which this happens
does not appear to be fixed; and whatever factor it may be,
whether something inherent or accidental, that determines
which of these struggling cells shall obtain the mastery and eat
up its fellows, it sometimes does not come into play until the
gonophore has become a well-formed structure. But itis quite
late in the history of the gonophore, when that structure is
large and already swollen with yolk, before these two, three
or four cells, which, so to speak, have succeeded in attaining to
the final heat, decide who is the winner.

In the facts which have been set down above with regard to
the structure of the ectoderm of the blastostyles, and the for-
mation of the gonophores, two main points appear to me to be
of special significance. These are (1) that the gonophore
appears to be a curiously modified bud, and (2) that the gene-
rative elements pre-exist as free cells having lodgment in the
tissues of the adult, and only travel into the abortive bud,
which is their place of final development.

maternal tissues, and not to the gonophore bud. They are, therefore, not
included in the above statement.

516 W. B. HARDY.

On CrErtaIn Points IN THE STRUCTURE AND FUNCTIONS OF
THE ENDODERM OF MyRIoTHELA.

In any animal three main problems concerning the manipu-
lation of its food-stuffs present themselves. These are (1) the
disintegration and solution of its food; (2) the absorption of
the dissolved or liberated and unchanged material; and (3)
the distribution of the products of digestion. We might also
add to these the storage of prepared food-stuff. Many and
diverse reasons justify the statement that a cell is hampered
or, better, limited in the range of its activity by being loaded
with indifferent reserve nutriment; and it therefore becomes
almost the duty of a special tissue to store material, either as
a provision for some extraordinary metabolic effort, or as a
consequence of an infrequent and uncertain food-supply. In
the case of Myriothela we shall see that this last problem—
the storage of reserved nutriment—occupies a large part of
the endoderm.

The disintegration of the food in all animals not possessed
of a masticatory apparatus is a process of solution differing
only in degree from the final solution of the smaller particles.
Yet I think we are justified in speaking of the whole act as a
process of disintegration and solution, because of the very
general tendency of animals to divide the process into those
two stages, and to differentiate the alimentary tract into a
special region where disintegration of the food by solution
takes place, e. g. the stomach of Vertebrates, and another
tract where solution is completed, and where solution and
absorption go on hand in hand.

In an animal of any considerable size the distribution of the
products of digestion becomes a problem of the utmost import-
ance, not only physiologically, but also from its far-reaching
influence on the morphological characters of animals. It has
been solved in three main ways:

1. By the utilisation of the enteric space itself, which func-
tions in part as a digestive cavity, and in part as a common

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 517

space in close communication with all parts of the organism,
and containing not only the results of the solution of the food,
but also material discharged from the lining cells of one region,
and destined for the nutrition of other parts of the organism.
It is as an example of this class that I wish to consider
Myriothela.

2. By the development of a system of spaces or a common
space round the gut, into which the results of digestion can be
discharged, and from which the tissues can directly derive their
nutriment. Such a space would be the hemoccel of morpho-
logists. The physiological significance of the ccelom is still, I
think, very much under judgment.

3. By the aid of a closed vascular system. The true rela-
tions of hzemoccel and ccelome to one another and the relations
of both to the vascular system of Annelids, which appears to
be initially respiratory ; and to the vascular system of Arthro-
pods, which is in its first inception a mechanism for the cir-
culation of the fluids of the circum-enteric space, are ques-
tions which do not concern us here, but the interrelation of
cases 1 and 2 demands brief notice.

It is necessary at the outset to distinguish clearly between
the digestive functions of the enteric space, and the part it may
play in the distribution of the nutritive material. The re-
searches of Miss Greenwood on the digestive process in Hydra
justify the conclusion that in that animal the enteric space is
used mainly, if not solely, for digestion. The endoderm-cells
forming its walls absorb and largely store the products of
digestion. There our exact knowledge ends, but the uniform
character of the endoderm throughout the entire animal, the
fact that its cells everywhere, even in the tentacles, absorb and
store nutriment, renders it probable that in the physiology of
Hydra the discharge of elaborated nutritive material into the
enteric space to form a common nutritive fluid akin to the
blood of higher animals plays little part. But in the higher
Ccelenterates, in the colonial forms, in Medusz, and in Cteno-
phora especially, we have no reason to doubt that such a fluid
does exist, and that it forms the metabolic link between the

518 W. B. HARDY.

different regions of those animals in such a way that the de-
mands of one part may be met by the discharge of stored
nutritive material from other regions. Such a fluid, which
Allmann has called the ‘‘ somatic fluid,’’ would not only con-
tain the immediate results of digestion, but also elaborated
material from the store of reserve nutriment possessed by
certain cells discharged in response to any special demand
in some particular region. In other words, it is not merely a
fluid for the distribution of the immediate products of digestion ;
it is more than that, and we shall see reason to think that it is
a true metabolic link between one part of the body and another
strictly comparable to the blood of higher forms.

In Allmann’s account of digestion in Hydroids he thus
describes the ‘‘ somatic fluid: ’—‘‘Its basis is a transparent
colourless liquid, and in this solid bodies of various kinds are
suspended. These consist partly of disintegrated elements of
the food; partly of solid coloured matter which has been secreted
by the walls of the somatic cavity ; partly of cells, some of
which have undoubtedly been detached from those walls,
though it is possible that others may have been primarily
developed in the fluid; and partly of minute irregular cor-
puscles, which are possibly some of the effete elements of the
tissues.”

Leaving the Celenterates and turning to the Turbellaria, we
have a striking instance of how the enteric cavity, the gut, may
serve as an organ for the distribution of nutriment. The case
of the Turbellaria also enables us to contrast cases 1 and
2, for in the Rhabdoccels we have animals with a simple gut
surrounded by a tissue, the mesenchyme, with numerous cleft-
like spaces, which may be so far developed as to form a fairly
well-marked space round the gut, as in Mesostoma tetra-
gonum, and to a less extent in Microstomum lineare.
That such a space or system of spaces facilitates the distribu-
tion of material derived from the gut hardly needs stating,
whether we consider the distribution to be brought about by
diffusion or by the agitation of the contained fluid as a result
of the muscular movements of the animal.

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 519

In the Triclads and Polyclads, on the other hand, the same
problem has been solved in a rather different fashion. The
development of spaces round the gut is not the most obvious
fact, but the gut itself ramifies into a network of tubes which
penetrate to every part ofthe body.

The Structure of the Endoderm. — Leaving these
general considerations, we will pass at once to a consideration
of the structure of the endoderm in Myriothela,

The mouth is bounded by a thin membranous lip, or hypo-
stome, which is exceedingly muscular and sensitive, and is
probably of considerable use in seizing the prey. It is figured
by Allmann and Hincks in a condition of extension, In
fig. 20 it is shown as it appears when the animal is retracted.

In the lip the supporting lamella is either reduced to the
thinnest film or is entirely absent. It is always absent from
about half the breadth of the lip from the free edge. The
ectoderm of the external surface is rich in nerve-cells, and
overlies a well-developed layer of radial muscles. At the free
edge there is no loss of histological continuity, the ectoderm
being continued on to its under surface for a certain distance,
and having the same structure. Passing downwards towards
the attachment of the lip, there appears at the base of this
ectodermal epithelium a more and more defined layer of
highly vacuolate cells, which are the first commencement of the
endoderm. In other words, the ectoderm for a short distance
overgrows the endoderm, thus forming a distinct zone of mixed
character.

At the point where the lip merges into the tentacle-bearing
region the ectoderm, as a distinct structure, has finally dis-
appeared, and we have the arrangement shown in fig. 14.
Three kinds of cells may be distinguished: (1) a super-
ficial layer of elongated cells, staining well and uniformly
with picro-carmine, ach possesses a nucleus and nucleolus
in its basal portion, which is tapering and wedged in between
the subjacent cells. In favorable preparations these cells
appear covered with short, fine cilia. These are, however, not
always visible, mainly because the animal in dying usually

520 W. B. HARDY.

retracts this region to such an extent that the epithelium is
thrown into deep folds, and the free surfaces of the cells ap-
posed. This ciliated zone forms only a narrow band. Between
the bases of the ciliated cells are (2) numerous rounded and
deeply-staining cells, which are more numerous the nearer the
lip. Delicate processes may be sometimes seen passing from
them to the surface, and they are probably sense-cells. Below
these, and forming by far the greater portion of the epithe-
lium, are strikingly characteristic palisade-like cells. Each
has a very scanty and ill-staining protoplasm, which surrounds
a large irregular vacuole occupying the bulk of the cell. Two
nuclei, each with a small nucleolus, are usually present, and
may be either close together about the middle of the cell, or
one at either end. There is not the slightest evidence that
the presence of these twin nuclei indicates cell division.

At the lower edge of the ciliated zone conical cells appear
between and rapidly replace the ciliated cells, while at the
same time the deep-staining sense-cells disappear. Each of
these conical cells resembles in its general appearance a goblet-
cell of an ordinary mucous membrane. We can, therefore,
conveniently style the next region the goblet-cell zone.
This embraces a considerable portion of the tentacle-bearing
region. It is, however, impossible to reduce the relative
dimensions of these various zones to numerical exactness
because of the extreme extensibility of this portion of the
animal.

Each goblet-cell is, as its name implies, flask-shaped, and
consists of an expanded part which stains lightly with picro-
carmine (fig. 16) or osmic acid (fig. 15). The contents
of this part are turbid from the presence of ill-defined,
granular masses. ‘The expanded portion of the flask is
continued downwards into a tail, which contains a small
nucleus embedded in deeply-staining granular protoplasm.
The numerous and coarse granules of the basal portion stain
deeply with osmic acid, and less so with picro-carmine. These
cells are undoubtedly of the nature of gland-cells, and the well-
formed basal granules may be regarded as the first stage in

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 521

the elaboration of the secretion product which occupies the
expanded portion of the cell. In the upper part of the goblet-
cell zone the endoderm is composed of goblet-cells lying
wedged between the apices of palisade-like cells exactly
resembling those occurring in the ciliated zone, except in the
fact that they now abut on the free surface. When this
portion is retracted the deep folds present the appearance in
sagittal sections of long tubular glands, which, from the
character of the abundant goblet-cells, closely simulate the
crypts of the large intestine.

In the middle and lower part of this region the endoderm is
thrown up into low conical villi. These structures are char-
acteristic of the whole of the endoderm with the exception of
the part already described, that is in the neighbourhood of the
mouth, and in the foot. They vary very much in length in
the different regions of the animal, but are usually longest in
the lower portion of the tentacle-bearing region, where they
are long filiform structures, sometimes branched, and may
measure ‘3 to ‘5 mm. from base to apex. Generally speaking,
the villi are not muscular, but in the goblet-cell region they
appear to have a distinct muscular axis.

Structure of a Villus from the Goblet-cell Zone.

A section through the axis of one of these is shown in
fig. 16. On the sides of each villus, and between their
bases, we have the same arrangement of palisade-cells and
goblet-cells as that described above. At the apex, however,
are a group of cells presenting many new features. These
I propose to call the apical cells, and they form not only
the apex of the villus, but are also continued downwards as its
muscular axis.

Each apical cell has the following general structure. The
protoplasm is abundant, and stains deeply, thus offering
a marked contrast to the other cells of the villus. It is
also turbid and opaque, but differs very much in this and
in its behaviour with stains in different parts of the cell.
In the muscular stem, however, the protoplasm is always

522 W. B. HARDY.

of a uniform texture, and behaves towards osmic acid and other
stains in a manner closely resembling the muscular elements
of the ectoderm. The protoplasm of the expanded end of the
cell encloses the following structures. One, rarely two, nuclei,
which vary so much in position as to suggest an extreme
mobility of the contents of the cell. A varying amount of
pigment, dark brown in colour, and disposed in scattered
grains near and on the free surface of the cell, but usually in
little heaps of grains in the deeper parts. One or more large
vacuoles. And, lastly, turbid masses of substance, some of
which are certainly the remains of material which has been
ingested by the cell, and all of which are in more or less
obvious relation to the vacuoles.

These factors sum up the constituents of the apical cell as
usually seen; but the extent and character of the vacuoles
and the turbid masses of enclosed matter vary very much at
different periods and in adjacent cells. To this, however, we
will return later.

In the varying position of the nuclei we have some indica-
tion of the mobility of the protoplasm of the apical cells; and
this mobility finds further expression in the fact that from the
free surface of the cells pseudopodial extensions are pushed
out, especially from those cells which are fairly free from
enclosed masses (cf. Allmann’s ‘ Memoir,’ pl. lvi, fig. 2).

In the middle tentacular region the endoderm assumes a
different character. The goblet-cells disappear, and the pali-
sade-cells gradually pass into a shorter and broader type of
cell, mostly with only one nucleus. These cells I will call
vacuolate cells, adopting the term applied by previous observers
to similar cells occurring in Hydra. These cells usually
contain numerous round hyaline corpuscles, which vary in
their characters but stain always with osmic acid and many
aniline dyes (such as methyl blue and green), but typically
take no coloration with hematoxylin and little or none with
carmine stains. These are, without doubt, identical with the
sphere-like masses of reserve nutriment described by Miss
Greenwood as occurring in the vacuolate cells of Hydra

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 523

under the name of “ nutritive spheres.” The vacuolate cells
of Myriothela when free from nutritive spheres may be seen
to possess a large vacuole surrounded by scanty protoplasm.
In this condition they recall the palisade-cells of the oral
region, and the intermediate types are so numerous that I
am disposed to regard these cells as fundamentally the same.
The most constant differences are that the palisade-cells have
almost always twin nuclei, and only rarely contain nutritive
spheres.

Wedged here and there between the vacuolate cells are
other and smaller dark-staining cells (fig. 21), which occupy
the same postion but are not disposed with the same regu-
larity as the goblet-cells. These cells are as variable in
size and appearance as the vacuolate cells, and they correspond
to the “ gland-cells”’ of Nussbaum and Jickeli. Miss Green-
wood has shown reasons for considering that cells similar
to these occurring in the endoderm of Hydra are concerned in
the formation of the digestive enzymes, and I find that her con-
clusions are warranted by the different appearances presented
by these cells under varying conditions in Myriothela To
this point we will return later.

Rarely a gland-cell may be seen apparently bearing a deli-
cate pseudopodium or flagellum, and having the appearance
shown in fig. 184. Nussbaum similarly describes cilia on the
gland-cells of Hydra.

These gland-cells are very widely dispersed throughout the
endoderm. They occur, perhaps, in greatest abundance on the
sides of the villi ; sometimes, however, one or two may occur
at the apex of a villus. Rarely, at the apex of a villus, a group
of two, three, or four small cubical darkly-staining cells is
found (fig. 19). Whether these are or are not stages in the
development or multiplication of gland-cells I was unable to
determine. That they are, however, the antecedents of the
free corpuscles which at certain periods occur in the somatic
fluid I see no reason to doubt.

The gland-cells of Myriothela have a rather wider distri-
bution than those of Hydra, where they are restricted to the

524 W. B. HARDY.

body. In Myriothela they occur in their greatest abundance
in the endoderm of the lower half of the tentacle-bearing
region. But they are also numerous in the middle region of
the body whence the blastostyles spring, and may even occur in
limited numbers in the endoderm of those structures near their
points of attachment.

In the middle and lower regions of the body the endoderm
is to a certain extent different from that already described.
The villi, as a rule, become less muscular, while at the same
time their apical cells change their characters and become more
and more akin to the vacuolate cells. The endoderm of the
body-wall from which the villi spring, and which in the lower
tentacle-bearing region is composed of cells in no wise dis-
tinguishable from those lining the villi, changes its character
in the blastostyle-bearing region. There it is composed of
long columnar cells, each with a single nucleus, and each
composed of dense well-staining protoplasm free from
vacuoles.

The endoderm maintains these characters in the foot; that
is to say, the villi, which here are of the nature of broad flange-
like folds, are covered by vacuolate cells with rarely a gland-
cell, while the endoderm of the body-wall is composed of the
deeply-staining columnar cells. At the extreme end of the
animal, however, the supporting lamella ceases to exist, and to
a certain extent the limits between ectoderm and endoderm
become obscured, and a kind of growing-point to the creeping
stolon is the result.

The endoderm of the blastostyles will receive special mention
later. To make this general account of the whole endoderm
complete, however, I will merely state here that the villi in
the blastostyles are low conical structures almost exclusively
composed of vacuolate cells. A few gland-cells lie scattered
in the proximal third of each blastostyle.

Taking the most general view of the structure of the epithe-
lium lining the enteric space of Myriothela as described in
the preceding pages, we see that it may be divided into different
regions. These are—(1) An oral region characterised by the

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGTA. 525

presence of sense-cells and cilia in its upper part, and of
numerous glandular cells, the goblet-cells, in its lower part.
(2) A middle zone comprising the middle region of the entire
animal, and characterised by the presence of numerous gland-
cells. (3) The blastostyles and the foot region, where the
endoderm is almost exclusively composed of vacuolate cells,
usually loaded to the full with stored nutritive material in the
form of nutritive spheres.

Of the function of the goblet-cells I can say little. From
their position I had supposed that their stored material was
discharged when the food was first received, and that they were
concerned in the elaboration of a digestive ferment or ferments.
But I have found them apparently unaltered in animals which
have just taken in their prey (a crustacean). The glairy,
sticky appearance of the contents of the expanded portion of
the goblet, however, suggests the idea that they form astrongly
adhesive surface to what may be called the prehensile portion
of the endoderm. The gland-cells, on the other hand, present
no especial difficulties. Fig. 18a represents one shrunken and
discharged as seen in an animal at the close of a digestive act,
that is with merely the detritus of a meal in its enteric cavity.
Fig. 18 shows one taken from a fasting animal. It is fully
loaded with granules. Fig. 184 represents an intermediate con-
dition. The granules are large and coarse, and appear to be
formed in the deeper portions of the cell. Their discharge is
characteristic, and may be witnessed in preparations from an
animal which has just ingested its prey. The granules are
extruded, apparently unchanged, into the somatic fluid, there
to be dissolved. That is to say, they do not break down to
form the digestive enzymes until they are free from the cell in
which they were formed (fig. 21).

The Process of Digestion.—For a long time I was
unable to determine the natural food of Myriothela, while at
the same time all endeavours to induce it to ingest pieces of
raw meat or fragments of Molluscs and Crustacea completely
failed. I therefore, in the meantime, turned my attention to
carmine and sodium sulphindigotate, and with a fine pipette,

526 W. B. HARDY.

inserted through the mouth, injected a drop of sea water con-
taining the one in suspension or the other in solution. Later,
however, I was more fortunate, and succeeded in obtaining
specimens with food in the enteric cavity. In one case the
prey was a Crustacean of some considerable size, so that it
produced a very obvious bulging of the animal. I did not
witness the capture and ingestion of the prey, but the specimen
was killed before the digestive fluid had produced any change
in the tissues. Fig. 21 is taken from this specimen. It also
contained the remains of a previous meal in the lower part of
the enteric space. Other specimens furnished other stages,
and in this way, as a result of an examination of a large number
of animals, I was enabled to obtain a series representing, to a
certain extent, the various stages of digestion.

Myriothela is carnivorous, and captures small Crustacea.
In one case the meal consisted of a half-digested egg, either
derived from the gonophores of the individual in question, or
from those of its neighbours.!

Digestion is carried on at first in the lower portion of the
tentacle-bearing region—that is, in that region where the
gland-cells are most abundant, and results in a disintegration
of the prey, brought about by the agency of the digestive
fluid.

The gland-cells which first discharge their contents are
those in the immediate neighbourhood of the meal, but even
there all the cells are not affected at once, those which are
most loaded with granules probably being discharged first.
Some of the gland-cells in the proximal region of the blasto-
styles and in the foot may be found undischarged until nearly

1 The huge yolk-laden eggs, when set free from the gonophores, are taken
by tentacle-like bodies, the ‘“claspers,”” which hold them while development
proceeds and until the actinula larva is fully formed. In March and April, how-
ever, the claspers are not always present, and the eggs formed then when ripe
are shed and sink to the bottom, where they become attached. I think that
further research will show that these early eggs have a more direct develop-
ment than those formed later, and pass at once to the form of the adult without

the intervention of the free-swimming actinula stage. It was one of these
free eggs which apparently had found lodgment in the enteric cavity.

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 527

the close of a digestive act. The disintegration of the prey is
accompanied by a great amount of solution, so that after
digestion has been in process for some time the enteric cavity
contains a number of fragments of the prey floating in a fluid
rich in proteids, which form a deeply-staining granular precipi-
tate after treatment with corrosive sublimate. The disin-
tegrated fragments of the meal find their way into the foot
and blastostyles, the somatic fluid probably being circulated
by the active movements of the animal, which include the ex-
tension and retraction of the blastostyles, and possibly by the
cilia, which appear to be borne here and there by the endo-
derm-cells. The process of digestion is therefore largely
extra-cellular ; but this is not all, intra-cellular digestion takes
place to a marked, but undoubtedly a subordinate extent, and
is mainly, perhaps entirely, confined to the ameboid and
mobile apical cells of the villi in the teutacular region.

If these cells be examined in an animal killed towards the
close of digestion they will often be found to contain the more
or less imperfect capsules of thread-cells or irregular frag-
ments of hyaline material, which recall the cuticle of a Crusta-
cean, and probably are fragmeuts of that structure.

In fig. 17 such a nematocyst is shown embedded in a turbid
mass of darkly-staining material, which occupies a vacuole
in the cell protoplasm. Atm is another nematocyst, now no
longer lying in a vacuole, but embedded in the cell proto-
plasm. The digestion of the food-mass with which this nema-
tocyst was associated having been completed, the vacuole has
filled up.

Other interesting points may be mentioned which point to
this intra-cellular digestion. In some cases an enclosed mass
may be found embedding a more or less perfect nucleus. By
a careful examination of different cells we may see such nuclei
in various stages of disintegration, until they are finally resolved
into chromatin granules which are scattered through a pseudo-
podial-like lobe of protoplasm, which appears to be thrust into
a vacuole as the digestion of its contents becomes complete.

Carmine grains injected into the enteric cavity are eagerly

528 W. B. HARDY.

ingested by the apical cells, both in the body of the animal and
in the proximal region of the blastostyles.

Towards the close of digestion a few free cells appear in
the somatic fluid. Each is rounded and composed of dark-
staining protoplasm embedding a nucleus with contained
nucleolus.

Before turning to the further fate of the food, that is to say
before considering the process of absorption, it will be well to
describe more particularly the contents and characters of the
vacuolate cells.

These when unloaded with nutritive spheres present the
appearance shown in fig. 18, where the cell is seen to consist
of a thin pellicle of protoplasm surrounding a large central
vacuole and embedding a nucleus. The protoplasm stains only
slightly. The outline of the cell is exceedingly sharply defined
by some staining material which has almost the appearance of a
cuticle. The process of loading with nutritive spheres is remark-
able, and essentially similar to that described by Miss Green-
wood as occurring in Hydra. The protoplasm at one point
develops a small vacuole which increases in size, and bulges into
the large vacuole. In this the nutritive sphere is formed from
the turbid semi-fluid material which first fills it. This process
continues until the whole of the cell becomes occupied by small
vacuoles, each containing a nutritive sphere. The size of the
cell, therefore, does not necessarily vary according to the amount
of reserve nutriment it contains. This, however, only holds
good for the vacuolate cells of the body. In the blastostyles
they are slightly different. In the first place the large central
vacuole is not developed, with the consequence that when the
cell discharges its nutritive spheres it frequently shrinks to the
condition of a cell with dense, non-vacuolated protoplasm, which
stains deeply with picro-carmine. In fig. 22, at 1 is a cell
without nutritive spheres, and at 2 one partly filled with
vacuoles containing nutritive spheres.

But the discharge of the nutritive spheres does not always
leave the cells smaller and solid. The vacuoles may persist
(fig. 22, 4), and the result is a remarkable cell with bubbly

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 529

protoplasm. Such cells form a very striking feature of osmic
acid preparations. The fluctuations in the size of the individual
cells in the endoderm of the blastostyles naturally leads to a
corresponding fluctuation in the total bulk of that tissue. In
specimens taken in May or June it sometimes almost fills the
cavity of the blastostyles.

We therefore have in the blastostylar endoderm, in addition
to the scattered gland-cells, the following:—(1) Small dense
cells with nucleus and nucleolus which stain deeply. (2) Cells
whose protoplasm is completely occupied by vacuoles, each of
which, some of which, or none of which contain nutritive
spheres. And there are numerous intermediate stages between
(1) and (2), with either a small or a large portion of the cell sub-
stance occupied by vacuoles. The protoplasm of cells (1) and
(2), or of the intermediate stages, appears in osmic acid prepara-
tions to be remarkably dense and almost glassy, and the vacuo-
lation is limited to the large vacuoles which embed the nutri-
tive spheres. But other vacuolate cells occur, forming a third
class, whose protoplasm is not of this character, but is so
occupied by vacuoles of all sizes as to give the whole cell a
very characteristic appearance (fig. 22). The vacuoles always
differ in size in different parts of the cell, being larger near the
free surface where they may contain young nutritive spheres.
These cells, forming the third type of vacuolate cell to be seen
in sections through the blastostyle, may be regarded as cells
which are actively forming nutritive spheres.

Before considering the processes going on in the vacuolate
cells generally, it will be well to turn to the nutritive spheres
themselves, The endoderm generally contains three kinds of
formed bodies—that is to say, bodies which are not of the
nature of material merely ingested by the cells, but rather are
new formations resulting from the activity of those cells.

Ofthese the most common are small spherical bodies, gene-
rally about 3 m» in diameter. These crowd the endoderm-
cells in great numbers, but are always most numerous in the
foot and blastostyles. They stain uniformly, but not so deeply
as to become opaque, with osmic acid, and when perfect appear

VOL. XXXII, PART IV.——-NEW SER. NN

530 W. B. HARDY.

singularly hyaline and structureless even with the highest
powers ; they do not stain at all with picro-carmine or hema-
toxylin, but take a deep tinge with aniline blue and methyl
green.

They are of a complex character chemically, with probably
a proteid basis. That they are not wholly proteid is, I think,
shown by the fact that the coloration obtained with the xan-
thoproteic reaction, and with acidulated ferrocyanide of potas-
sium and ferric chloride, is never intense though it is sufficiently
distinct. Neither iodine nor iodine and sulphuric acid give
distinctive results. They are also not of a fatty nature, for
exposure for many hours to hot turpentine fails to materially
change them. These are not only the most abundant, but
the most permanent nutritive spheres.

The second class of bodies is markedly distinct from those
just mentioned, and they occur sometimes in considerable
abundance. They are perhaps most noticeable at the close of
a digestive act. They measure as a rule 12 » in diameter,
and are spherical bodies, each embedded in a vacuole of a
vacuolate cell, which may or may not contain at the same time
the small nutritive spheres. Like the latter they are some-
times homogeneous bodies showing no internal structure,
and then they stain very intensely with picro-carmine (fig.
27). But they may be also found composed half of intensely
staining homogeneous material, and half of more turbid
material which stains scarcely at all (fig. 28). In yet other
cases the intensely staining material is reduced to a small
spherical nodule or patch placed excentrically on the surface
of the sphere. There can be no doubt that these are various
stages in the formation or destruction of the same bodies, but
I do not think that the evidence at my disposal justifies me in
deciding which stage is which. Asa general rule, however, the
smallest of these bodies, perhaps but little larger than the first-
mentioned type of nutritive sphere, are homogeneous and deeply
staining, while without exception the largest forms are those
with the deeply-staining material reduced to an excentrically
placed bleb. These bodies resemble in some respects the yolk

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 531

spherules of the ripe egg, and they are also found in some
abundance in the young buds. I can throw no light either on
their special significance or on their relation to the small nutri-
tive spheres. I might finally add that they are rarely widely
distributed throughout the endoderm, but usually occur in
localised patches as though they were related to some localised
and special metabolic process. They are perhaps never com-
pletely absent.

Though the term nutritive sphere can, I think, be applied
with justice to both the preceding bodies, its application to
the third class of endodermal products would be misleading,
since they are merely a specialised product of the endoderm
of the tentacles.

When abundant, each cell of the endoderm of a tentacle
may contain one of these bodies, and then they doubtless give
rise to that opacity which was noted by Allmann. They form,
however, a very variable element, for while one tentacle may
be fully charged with them its neighbour may contain few or
none. Each of these bodies is, when fully formed, 10 mu in
diameter, and consists of a sphere which stains intensely
with picro-carmine (fig. 26), and is embraced by a cup-shaped
capsule with expanded edge.

That these bodies play any great part or are at all concerned
in the general nutrition is extremely improbable. Under
certain circumstances they are discharged from the endoderm-
cells of the tentacles and find their way into the enteric space,
and are ingested and digested by the apical cells of the
adjacent villi.

In what I have to say concerning the formation and fate of
the nutritive spheres in the following pages, I shall refer ex-
clusively under that title to the small nutritive spheres which
are so abundant and numerous. The method of formation of
these bodies has been already described, and it was seen that
they develop in vacuoles formed in the protoplasm of the
vacuolate cells ; and I see no reason to doubt, but rather every
reason for agreeing with Miss Greenwood’s view, that these
bodies are formed from material absorbed in the fluid form

582 W. B. HARDY.

from the results of digestion in the enteric cavity. The fate
of sodium sulphindigotate when introduced into the enteric
cavity is interesting in this connection, for though a consider-
able portion of that pigment is rapidly decolourised, some may
be found after a short period in the vacuolate cells associated
with the nutritive spheres in the various stages of their forma-
tion in such a way that these bodies are tinged with blue in
irregular patches. We may conclude, therefore, that the sub-
stances resulting from the solution of the tissue of the prey
are mainly absorbed by the vacuolate cells, while at the same
time ingestion of disintegrated fragments, possibly of a re-
sistent nature, takes place to a limited extent, and is carried
on by the apical cells of the villi in the middle region of the
body. And the distribution of the products of digestion to
the more remote portions of the endoderm is effected by the
agency of the general somatic fluid.

But, as I said before, there are reasons for believing that
the somatic fluid is more than a vehicle for the distribution of
the immediate results of digestion. Let us now turn to a
consideration of those reasons.

In the blastostyles the extensive accumulation of nutritive
spheres is in obvious relation to the active and important pro-
cesses carried on in those structures. But the nutritive
spheres are stored in equal abundance in the endoderm of the
foot, where, during the summer and autumn, there appears to
be no great call for this enormous reserve of nutritive mate-
rial. On the other hand, throughout the whole tentacular
region and usually in the endoderm of the tentacles themselves
no abundant reserve of food-stuff is present. It is, as I
pointed out above, highly doubtful whether the peculiar bodies
sometimes found in the endoderm of the tentacles are of the
nature of simple reserve nutriment. On the other hand, it is
equally certain that the actively motile and sensitive tissues
of the tentacular and oral regions are not supplied directly
and solely from the immediate products of digestion. We are,
therefore, compelled to conclude that the nutritive material
stored in one region of the body may be conveyed as occasion

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 533

demands to regions quite remote. In other words, we must
suppose that the endoderm is not merely a collection of cells
each fighting for a share of the nutritive material to be
ultimately used by itself or by the ectoderm with which it
is anatomically in immediate relation, but rather that the
metabolic activities of the units of the endoderm throughout
its whole length are linked together into one consistent and
interdependent whole.

What is the nature of that link, and how is the interchange
of material which it implies between widely separated regions
brought about? That it is effected entirely by the laborious
passage of material from cell to cell throughout, perhaps, a
considerable length of the animal is, I think, an impossible
suggestion. Yet this process undoubtedly takes place to a
certain extent, and I think we may see in the palisade-cells
of the oral region, from the contents of whose enormous
vacuoles amorphous masses are precipitated by the action of
corrosive sublimate, a mechanism for facilitating such a process,
and whereby nutritive material may find its way even to the
lip, a region which during the early stages of the digestion of
prey of any considerable size must be more or less cut off
from the general somatic fluid. This same method of distri-
bution of nutritive material must also obtain between the
endoderm and the ectoderm, the interchange being facilitated
by the numerous pores which may be seen to penetrate the
supporting lamella when horizontal sections of that structure
are examined with the highest powers. But the stored nutri-
tive material of the foot can only be rendered available for the
body generally through the agency of the somatic fluid, and to
a certain extent histological facts support this conclusion.

If we attempt to follow the further fate of the nutritive
spheres we find that two things may happen to them. They
may either undergo a gradual disintegration, their substance
becoming at the same time studded with pigment grains which,
after the nutritive sphere as such has ceased to exist, remain
as a little heap of dark granules, bound together by a scanty
amount of unstaining substance (fig. 25), while, as the dis-

534. W. B. HARDY.

integration of the sphere approaches completion, the vacuole
in which it lay becomes obliterated; or they may be rapidly
and entirely dissolved and discharged from the cells, the
vacuoles meanwhile persisting and leaving the striking honey-
combed appearance described above (fig. 22). Whatever may
happen to the constituents of the sphere in the first case,
which agrees with the fate of the nutritive spheres of Hydra
as described by Kleinenberg and Greenwood, we can only con-
clude that in the second case they have been dissolved and
discharged into the enteric cavity. It is even possible that we
may divide the endoderm-cells, other than the gland-cells, into
two sets: (1) Those which are concerned in the elimination
of waste matter from the nutritive spheres or from the somatic
fluid directly. These are the apical cells of the villi and the
vacuolate cells in their immediate neighbourhood. And (2)
Those which discharge their stored material, leaving, so far as
can be detected, no residue. These lie towards and between
the bases of the villi in the blastostyles and middle regions of
the body and foot.

This conclusion, which may be accepted as a provisional
hypothesis until the processes taking place in the endoderm
shall have been worked out more fully, is based upon two facts ;
namely, that the pigment, as was noted by Allmann, is located
only in the cells near the free ends of the villi, and that the
bubbly cells in all their various conditions of incomplete or
complete discharge always lie between or towards the bases of
the villi.

Another point of evidence in favour of the view that the
somatic fluid conveys stored nutriment from one part of the
body to another is derived from a study of the histology of the
spadix of the gonophores.

Structure of the Spadix of a Gonophore.—This,
in the completely formed female gonophore, is composed of
a considerable number of tongue-shaped villi, which have their
apices turned towards the axis of the spadix, and project
a considerable distance downwards towards the centre of
the blastostyle. The cells between their bases, and therefore

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 535

forming the apex of the spadix, are long, narrow, and columnar
in character ; and their protoplasm is filled with numerous
small vacuoles, the contents of which become precipitated by
preserving reagents, thus conferring a characteristically turbid
character on the entire cell. That end of the cell which abuts
on the basement membrane separating the endoderm of the
spadix from the developing generative elements is excavated
to form a vacuole. If we examine the cells which form the
villi we find that they have fundamentally the same structure,
except that the basal vacuole is now so much elongated that
we may almost speak of that portion of the cell as being
canalised (fig. 23). In other words, the whole spadix is a
specialised structure for the absorption of nutriment from the
somatic fluid of the blastostyle, which nutriment is doubtless
largely derived from the stored material of the vacuolate cells,
through the help of the somatic fluid.

The absorbed nutriment, probably after it has undergone
important changes at the hands of the cells of the spadix, is
discharged into the vacuoles at their base which abut on the
supporting lamella, whence it passes to supply the remarkably
abundant fluid present in the entocodon of the female gono-
phore.

These different points—(1) the general distribution of the
nutritive spheres, (2) the method of the discharge of those
bodies, and (3) the fact that the gonophore possesses an organ,
the spadix, the histological characters of which lead us to
suppose that itis designed to absorb nutriment from the
somatic fluid (which, especially in the autumn, when Myrio-
thela appears to exist largely at the expense of its stored
material, and probably throughout the year, must be largely
recruited from the reserve material of the vacuolate endoderm-
cells)—although when taken singly they are of slight value,
yet when considered together and as mutually supporting
one another they justify the statement that the metabolic
activities of the different parts of the endoderm are brought
into relation with one another through the agency of the
somatic fluid.

536 WwW. B. HARDY.

If no such link existed, and if, therefore, the animal were
unable to direct its entire resources towards the accomplish-
ment of any metabolic act, we should expect to find evidence of
the fact in the more marked exhaustion of the endoderm during
starvation in the immediate neighbourhood of a gonophore as
compared with the other parts of the blastostyle or of the
body generally. But such evidence appears to be wanting,

EXPLANATION OF PLATES XXXVI & XXXVII,

Illustrating Mr. Hardy’s memoir “On some Points in the
Histology and Development of Myriothela phrygia.”

Fic. 1.—Section through the ectoderm of the distal portion of a blasto-
style. [Animal killed in May.]

Fic. 2.—Section of the ectoderm of the gonophore-bearing region. [Ani-
mal killed with osmic acid in May.] th ob.

Fic. 3.—Teased preparation from the same specimen. The primitive germ-
cells have dropped out. - th ob.

Fic. 4.—Section through the generative region of an exhausted animal
killed in autumn.

Fies. 5, 5a, 56.—Ectoderm elements isolated by teasing. Osmic acid. In
5 and 5a are represented parts of the nerve network. -

Fic. 6.—Isolated primitive germ-cells. Osmic acid. ,th ob.

Fie. 7.—Section showing the process of absorption of the supporting
lamella.

Fic. 8.—First stage in the formation of a gonophore. Hctoderm thickened
and containing a cluster of primitive germ-cells in its lowest part.

Fic. 9.—The second stage in the formation of a gonophore.
Fic. 10.—The third or blastema stage.
Fig. 11.—A completely formed young gonophore.

Fic. 12.—Piece of the cuticle which covers the ectoderm stripped off.
zsth ob
165 .

HISTOLOGY AND DEVELOPMENT OF MYRIOTHELA PHRYGIA. 537

Fie. 13.—A Myriothela bearing buds in various stages.
Fic. 14.—Section through ciliated cell zone of the endoderm.
Fie. 15.—Goblet-cell. Osmic vapour.

Fie. 16.—Villus of goblet-cell zone.

Fic. 17.—Two apical cells.

Fic. 18.—Loaded gland-cell and empty vacuolate cell.

Fic. 18¢.—Discharged gland-cell.

Fic. 184.—Intermediate condition of gland-cells.

Fic. 19.—Group of small cells from apex of villus.

Fic. 20.—Oral end of Myriothela with lip retracted.

Fic. 21.—Villus from middle region of the body, with gland-cells just com-
mencing to discharge.

Fic. 22.—Group of cells from the endoderm of a blastostyle. Osmic acid.

Fie. 23.—Villus from the spadix of a gonophore, showing canal-like
vacuoles in the lower portion of the cells. At a@ and a’ these are cut across.

Fic. 24.—Two cells from the endoderm of a blastostyle. Osmic acid.

Fic. 25.—Nutritive spheres containing pigment granules. Osmic acid.
zsth ob.

Fic. 26.—Body from the endoderm of a tentacle.

Fic. 27.—Endoderm-cell containing the large type of nutritive sphere.

Fic. 28.—Large nutritive sphere.

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STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 5389

On the Structure of an Earthworm allied to
Nemertodrilus, Mich., with Observations
on the Post-embryonic Development of
Certain Organs.

By

Frank E, Beddard, M.A.,
Prosector of the Zoological Society of London, Lecturer on Biology at
Guy’s Hospital.

With Plates XXXVIII and XXXIX.

TABLE OF CONTENTS.

I. Introductory, p. 589. VI. Homologies of the Reproduc-
II. List of Memoirs consulted, p. tive Organs in Hudrilide, p.
BA]. 579.

VII. Definition of Genus Libyo-
III. External Characters, p. 542. drilus and Species L. vio-

IV. Anatomy and Histology, p.544. laceus, p. 588.

V. Description of some Young | VIII. Summary, p. 583.
Stages, p. 572. IX. Explanation of Plates, p. 585.

I. Introductory.

Tue investigations of Rosa (5),! Michaelsen (8, 4), and more
recently of myself (1), have shown that the earthworm fauna of
tropical Africa is characterised by an extraordinary abundance
and diversity of types of the family Eudrilide. This family
had been known for a long time by the genus Eudrilus alone,
which seemed to occupy—principally on account of the struc-
ture of the reproductive system—a somewhat isolated position
among the Oligocheta.

* The numbers in brackets refer to the list of papers on p. 541.

540 FRANK E. BEDDARD.

Eudrilus itself has been only lately received from the
African continent; but no less than eight new genera of this
family have been within the last year recorded from thence.
I have myself on two separate occasions received worms from
Lagos, which proved to be the types of three new genera;
this shows the great abundance of forms which must remain
to be discovered.

In a recent number of this Journal (1) I described two
genera—Hyperiodrilus and Heliodrilus—which I ob-
tained from the Royal Gardens at Kew; they were found in
earth which had come from Lagos. In all probability the
genus Siphonogaster, of which I have given a brief notice
in the last published number of the ‘ Proceedings of the Zoo-
logical Society,’ will prove to be a Eudrilid; this worm was
discovered by Mr. Alvan Millson, Assistant Colonial Secretary
at Lagos, who has contributed to the ‘Kew Bulletin’ of
October, 1890, a very interesting account of its habits. The
same gentleman was so good as to bring over with him a large
box full of living earthworms from Lagos, which included
some specimens of the ‘‘ Yoruba worm ;” unfortunately these
latter disappeared during the voyage, but the other species
survived ; it proves to be the type of a perfectly distinct and
new genus, which presents some remarkable peculiarities of
structure. In the box were also some examples of asmall worm,
which appears to be an Allolobophora, and some very young
examples of the new Eudrilid. The opportunity thus afforded
me of contributing something towards the development of the
reproductive system and nephridia of Libyodrilus violaceus
causes me to feel less regret that it was impossible to keep the
species alive in order that they might breed.

I am greatly indebted to Mr. Alvan Millson for his
kindness.

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 541

II. List of Memoirs consulted.

(1) Bepparp, F. E.—“ On the Structure of Two New Genera of Eudrilide,
with Remarks on Nemertodrilus, Mich.,” ‘Quart. Journ. Micr.
Sci.,’ March, 1891.

(2) Bepparp, F. E.—‘*On the Occurrence of Numerous Nephridia in the
same Segment in Certain Earthworms, and on the Relationship between
the Excretory System in the Annelides and in the Platyhelminths,”
‘Quart. Journ. Mier. Sci.,’ vol. xxviii, N. 8., p. 397.

(8) Micnuartsen, W.—“ Beschreibung der von Herrn Dr. Franz Stuhlmann
im Miindungsgebiet des Sambesi gesammelten Terricolen,” ‘Jahrb.
Hamb. Wiss. Anat.,’ Bd. vii.

(4) Micuartsen, W.—<Oligochaeten des Naturhistorischen Museums in
Hamburg,” iv; ibid., Bd. viii.

(5) Rosa, D.— Lombrichi dello Scioa,’ ‘ Ann. Mus. Civ. Genova,’ 1888.

(6) Horst, R.—“ Note on a New Earthworm,” ‘ Zool. Anz.,’ 1891.

(7) Horst, R.— Sur quelques Lombriciens exotiques appartenant au genre
Eudrilus,” ‘ Mém. Soc. Zool.,’ 1890, p. 223.

(8) CexrontaIne, P.—“ Recherches sur le Systéme Cutané et sur le Systéme
Musculaire du Lombric Terrestre (Lumbricus agricola, Hoffmeis-
ter), ‘Mém. cour. et Mém. sav. étr. Acad. Roy. Belg.,’ 1890. (This
paper is reprinted in ‘ Arch. de Biol.’)

(9) Spencer, W. B.—“ The Anatomy of Megascolides australis (the
Giant Earthworm of Gippsland),” ‘Trans. Roy. Soc. Victoria,’ vol. i,
part 1.

(10) Perrier, E.—‘‘ Mémoires pour servir a V/histoire des Lombriciens
terrestres,” ‘Nouv. Arch. Mus.,’ t. viii.

(11) Bennam, W. B.—“ Atrium or Prostate?” ‘ Zool. Anz.,’ 1890.

(12) p’Uprxem, J.—“ Histoire Naturelle du Tubifex des Ruisseaux,”
‘Mém. cour. et Mém. des sav. étr.,’ 1853.

(18) Horst, R.— Descriptions of Earthworms,” I, ‘Notes Leyden Mus.,’

vol, ix, p. 97.
(14) Erste, H.— Die Capitelliden,” im ‘Fauna and Flora des Golfes von
Neapel.’

(15) Bepparp, F. E.—* Contributions to the Anatomy of Earthworms,”
No. I, ‘ Proc. Zool. Soc.,’ 1887.

542 FRANK E,. BEDDARD.

III. External Characters.

The worms were brought over in a large wooden box nearly
filled with a sandy loam; they preferred the bottom of the
box, and were in nearly every case found in close contact with
the wood; this may be perhaps owing to the greater warmth.
The worms were very inactive in their movements; but this is
possibly not so much a characteristic of the species as due to
the coldness of the weather. Nevertheless I have not found that
tropical Pericheta, which are the most agile of earthworms,
are in any way influenced by such causes.

The colour of the species is a dull purplish brown with a
distinct pinkish tinge; the clitellum is only to be distinguished
by its darker colour. The colouring substance was largely
dissolved out by alcohol staining the fluid of a greenish yellow.
There remained, however, after treatment with alcohol, a dark
coloration on the dorsal surface.

The worms, when placed in spirit, protruded the buccal
cavity, which appeared of a bright red colour; they do not
protrude it in locomotion, as Pericheta does.

Prostomium.—The prostomium is short, and not continued
by a groove on to the buccal segment.

Setze.—The setz are strictly paired, and lie upon the ventral
surface of the animal. There is nothing unusual in their
form. The ventral sete of Segment 17 are absent, as shown
in the illustration (fig. 2). The lateral and ventral setz of each
side of the body are connected by a muscular slip shown in
fig. 19, M.

Dorsal pores appear to be, as in other Eudrilids, entirely
absent.

Nephridiopores, usually conspicuous enough in this
family, could not be detected in the present species (see, how-
ever, p. 556) even with a lens.

The clitellum is continuously developed all round the
body : in most specimens it occupied only two segments, viz
Nos. 15 and 16; in some, however, it extended over the 14th

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 543

also. The extent of the clitellum is more limited than in
other Eudrilids, where four to six segments appear to be the
rule.

Spermathecal Pore.—This is situated in the middle
ventral line (see fig. 2, sp) of Segment 13; it lies between
the ventral pairs of sete; the aperture itself is small, but
its position is rendered somewhat conspicuous by being
placed upon the summit of a slight swelling of a yellowish
colour.

Oviducal Pores.—These pores occupy what is, at present,
an unique position among earthworms. With the exception
of certain species of Moniligaster, and with the possible
exception of Eisenia (= Tetragonurus of Hisen), the ovi-
ducts of earthworms appear to open invariably upon the 14th
segment. In the present species they are situated on Seg-
ment 15. I took particular pains to assure myself that this
was really the case; in half a dozen or so of individuals which
were marked by having these orifices very conspicuous they
were certainly situated on that segment. The oviducal pores
in this worm are not difficult to see ; indeed, these apertures
are for the most part particularly plain in the Eudrilide. This
is due to the fact that the end of the oviduct is often slightly
protruded, and forms a round whitish elevation contrasting in
colour with the surrounding dark integument. I mention
these facts more particularly for the purpose of showing that
I am not likely to have fallen into an error in assigning this
abnormal position to the oviduct pore. The pore in Libyo-
drilus has the same appearance as I have figured and
described in Heliodrilus (1); that is to say, it is a whitish
knob upon the segment: it lies to the outside of and a
little in front of the lateral pair of setz, but well behind the
groove which separates its segment (the 15th) from the 14th.
These are the only apertures belonging to the genital system
which are paired.

The male genital pore is unpaired and median; it lies on
the border line of Segments 17,18. The aperture lies upon
the summit of a conspicuous elevation of the integument; the

544, FRANK E. BEDDARD.

two penial sete may be often seen protruding from the
orifice.
I found no genital papille of any kind.

IV. Anatomy and Histology.
§ Body-wall.

Epidermis.—The chief point to be noted about the epi-
dermis is the absence of the peculiar sense organs so generally
found among the Eudrilide. I have described them in
Eudrilus, Hyperiodrilus, and Heliodrilus. Dr. Horst
has recently described them in an African species of Eudrilus,
and was the first to suggest that they were possibly sense
organs ; he compares them to the Pacinian bodies of Verte-
brata. I had originally considered these bodies as repre-
senting rudimentary sete, and as corresponding to those
epidermal structures in Urocheta which Perrier first de-
scribed, and considered as rudimentary sete; but a more
detailed examination of their structure led me to express in
my paper upon Hyperiodrilus and Heliodrilus an
opinion that they were of a sensory nature, and to make the
identical comparison that Dr. Horst made, previously in point
of time of publication. Rosa has found these structures in
Teleudrilus; but they do not exist in Nemertodrilus or
in Libyodrilus. The absence of these epidermal sense
organs unites these two genera, which have other points in
common.

The cells of the epidermis are of the usual kind; the large
glandular cells appear to be perhaps unusually abundant. I
have not made a special study of the histology of the epi-
dermis, which has been recently so elaborately worked out
and so beautifully illustrated in Lumbricus by M. Cerfon-
taine (8).

Muscular Layers.—The usual circular and longitudinal
muscles are present; the relative thickness of the two coats
may be judged from figs. 4 and 13. In both layers the fibres

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 545

are embedded in a gelatinous-looking matrix, which is more
abundant in proportion to the muscular tissue in the case of
the longitudinal coat. As to the fibres themselves, I find that,
as M. Cerfontaine has described in Lumbricus (8), both
those of the longitudinal and circular coat present a radiately
striate appearance, the striz converging towards a central
clear space. I have not taken any special means to investi-
gate thoroughly the histological structure of the muscles, I
only describe what I have seen in transverse and longitudinal
sections of the entire worm; but I have little doubt from what
I have been able to see that the results of M. Cerfontaine’s
investigations into Lumbricus will hold for Libyodrilus
also. The matrix in which the fibres are embedded can hardly
be termed granular in Libyodrilus; it has a gelatinous
homogeneous appearance, and is but slightly affected by borax
carmine. It forms a specially thick layer (fig. 14) between
the end of the longitudinal muscular fibres and the perito-
neum. The fibres—both those of the circular and the longi-
tudinal coat—tend to be grouped into definite areas, which
are only separated from each other by the thickness of the
matrix lying between them. I quite agree{with M. Cerfon-
taine that there is nothing comparable to the septa described
by Claparéde. No doubt such an appearance might be pro-
duced by great shrinking, caused by preservation in strong
alcohol without previously fixing the tissues.

The arrangement of the fibres of both muscular coats is
regular, but the regularity is not so great as in certain species
of Lumbricus and Allolobophora; each “ compartment ”
is made up of fibres, which are three, four, or five deep—even
more sometimes.

It is an interesting fact that in the young worm the fibres
are disposed much more nearly in the bipinnate fashion origi-
nally made known by the descriptions of Claparéde.

§ Celom and Intersegmental Septa.
The first septum separates Segments 4—5; in front of
this the pharynx is attached to the parietes by numerous muscu-
VOL, XXXII, PART IV.—NEW SER. 00

546 FRANK E. BEDDARD.

lar bands which occupy a good deal of the coelomic space: it
is probable that the reduction of the nephridia in this region
of the body is brought about by the development of this exten-
sive meshwork of muscles. The septa which lie behind the
first are thickened, and formed often of distinct layers of fibres.
The last of the thick septa separates Segments 11—12;
the next septum, however (see fig. 1), although not so
thick as those which precede it, is thicker than those which
follow.

A noteworthy point is the non-coincidence of the insertion
of the septa with the intersegmental furrows. This frequently
occurs inearthworms, but according to Rosa it is possible to
trace the fibres belonging to the septum from their apparent
attachment to the middle of the segment to their real insertion
into the intersegmental furrow. I could not trace any such
arrangement in Libyodrilus. The first septum which has an
abnormal insertion is 12—13; this and the following six or
seven are attached near to the middle of the segment just in
front of the sete. It is only dorsally and laterally that the
septa are so attached ; ventrally their insertion is quite regular.
I have mentioned on page 571 the effect produced upon the
position of the oviducal pore by the shifting of the interseg-
mental septum 14—15.

The principal point of interest in connection with the celom
is the separation of spaces which surround some of the organs.
In my description of the vascular system I have referred to the
perihemal space which encloses the subcesophageal blood-
vessels; besides this space, the areas surrounding both of the
lateral pair of setze are enclosed by a membrane shutting them
off from the general body-cavity of their segments. A series
of transverse sections, such as the one illustrated in fig. 19,
shows that this membrane is perfectly continuous everywhere,
forming a distinct chamber, which is prolonged round the seta
sac and nearly reaches the circular muscular coat. In the
Capitellidee Eisig has described somewhat similar lateral
chambers, into the floors of which the setz are inserted ; but I
am not aware that anything of the kind has until now been

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS, 547

described in the Oligocheta. As will be seen from fig. 19,
the muscle connecting the two pairs of sete lies within the
chamber. The membrane which forms the walls of the
chamber is nearly homogeneous in appearance, with a covering
and lining of epithelium only recognisable by the nuclei.

§ Alimentary Canal.

The alimentary canal of Libyodrilus presents certain im-
portant differences from that of other Eudrilide. There is in
the first place no trace that I could discover of csophageal
glands, or of those impaired ventral pouches termed “ chylus-
taschen” by Michaelsen, which occur in so many Eudrilids.

The position of the three gizzards is another point of interest.
In Heliodrilus and Hyperiodrilus there are a larger
number of gizzards, which are situated as in Lumbricus at
the junction of the cesophagus and intestine ; the three gizzards
of Libyodrilus occupy a similar position.

In his memoir upon the classification of earthworms Perrier
remarked upon the occurrence in all the Anticlitellians of the
gizzard behind the organs of generation and the contractile
hearts of the vascular system ; from this group he distinguished
both the Post- and Intra-clitellian worms by the fact that the
gizzard was placed in front of the essential organs of repro-
duction.

This distinction held good until my own discovery of
the position of the gizzards in the two Eudrilids above
referred to. In some forms—for instance, a Teleudrilus—the
gizzard has the forward position which characterises the
majority of worms belonging to Perrier’s two groups of Intra-
clitellians and Postclitellians; but I am disposed to think
that the arrangement of these organs found in Heliodrilus,
Hyperiodrilus, and Libyodrilus will prove to be more
characteristic of the group. There are no particular reasons
that I can see for placing the Eudrilide in the neighbourhood
of the Lumbricide, and thus one of the characters of the latter
group can no longer be regarded as distinctive. Dr. Horst

548 FRANK E. BEDDARD.

has already shown in his brief account (6) of an evidently very
interesting form, Gly phidrilus Weberi, that the position of
the clitellum hitherto found only among the Lumbricide is
also to be found in a worm clearly referable to Perrier’s
Intraclitellians. The varying position of the gizzard is less
remarkable now that there is some probability of its epithe-
lium being always derived from the hypoblast; if the older
view that the gizzard belonged to the stomodzal invagination
had been confirmed by more recent researches, it would have
been necessary to regard the calciferous pouches as being
epiblastic when in front of the gizzard and hypoblastic when
lying behindit. The relations of the same diverticula show that
when the gizzard is followed immediately by the “ large in-
testine ” it does not mean that there has been a suppression of
the small intestine immediately following the gizzard. The
absence of any relation between the position of the gizzard (or
gizzards) and the position of other organs seems to show clearly
that this modification of the walls of the anterior section of
the gut may take place anywhere. There is no need to assume
that the gizzard of one earthworm is the exact homologue of
that of another.

The pharynx occupies the first five segments.

The esophagus is of great length, and divided into two
distinct sections, neither of which, as already mentioned, is
furnished with glandular diverticula. The first section of the
tube is of narrower calibre, and extends as far as the end of
Segment 19. After this the tube becomes suddenly dilated and
forms akind of crop, which immediately precedes the gizzards.
The “ crop” occupies Segments 20—22 inclusive.

As to histological structure, the cesophagus does not differ
from that of other earthworms; its walls are exceedingly
vascular, and the epithelium is in many parts rather thin.
Even in the 12th segment there are a few chloragogen. cells
upon the cesophagus. These become largely developed in Seg-
ment 15, where the cesophagus is as closely enveloped by them
as is the intestine; the transition, however, appears to be
gradual. The latter part of the cesophagus from about Segment

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 549

14 onwards is ciliated ; its epithelium is thrown into regular
longitudinal folds ; the cells are tall and columnar, and the
muscular coat is of an appreciable thickness.

The cesophagus is extremely vascular ; in longitudinal and
transverse sections its walls have from this cause a moniliform
appearance (fig. 16). So far as I could make out, the plexus of
blood-vessels which lies immediately below the epidermis was
nowhere a sinus, although the individual vessels were so close
together as to give this appearance; still a more careful
examination showed the longitudinally running connecting
branches between the much wider circular vessels.

The three gizzards are in Segments 23, 24, and 25—one
gizzard to each segment; they are not all three directly con-
tinuous, but are separated by a certain amount of soft-walled in-
testine. The soft-walled portion occupies the anterior part of
each gizzard segment ; posteriorly, the gizzard comes into con-
tact with the septum.

The width of the crop gives the gizzards the appearance of
being strung, as it were, upon the intestine; but that the
crop belongs to the cesophagus and not to the intestine is shown
by the fact that it has no typhlosole.

The intestine is furnished with a typhlosole which is of a
somewhat unusual form. From its very commencement to as
far back as Segment 37, three folds can be seen on a dissection—
one median, and one on each side. The median fold corre-
sponds, of course, to the dorsal vessel, and is of about thrice the
vertical diameter of each of the two lateral folds. All three
folds were quite conspicuous in dissections from their red
colour. After the 37th segment the two lateral folds disap-
peared, but the median typhlosole continued on to near the
end of the body.

In dissected worms, both fresh and after preservation, the
intestine was of a rich mahogany brown colour, quite different
from the colour of our British Lumbrici; Nemertodrilus
and some of the other Eudrilids agree with the present species in
this particular, which is due to the abundance and darkness of
the granules in the chloragogen cells.

550 FRANK E. BEDDARD.

§ Circulatory System and Perihemal Spaces.

The principal account of the circulatory system in this family
of Oligocheta—and that is very incomplete—is contained in
a paper upon Eudrilus by myself (15). The blood-vessels
of Libyodrilus are very similar, but I did not find a sub-
nervian trunk. Very frequently in transverse sections a blood-
vessel could be seen lying beneath the peritoneal membrane,
and vertically beneath the nerve-cord ; but by following out
the vessel through a few sections it was invariably found to be
merely a part of the integumental blood plexus running for a
short distance in a longitudinal direction. The dorsal vessel
is united with the ventral by a series of paired “hearts.”
These increase in size posteriorly. The dorsal vessel is
ensheathed by a layer of “‘ chloragogen ” cells, beneath which
is a thin layer of very delicate muscular fibrils; the aper-
tures of the hearts into the dorsal vessel are guarded by
valves consisting of a mass of granular columnar cells with
small nuclei; there are similar valves where the hearts
debouch into the ventral vessel. The supra-intestinal
trunk is recognisable only in the cesophageal region of the
worm; it is very easily seen in dissections, and its calibre
is quite equal to that of the dorsal vessel. The supra-intestinal
differs from the dorsal blood-vessel in having only a very thin
peritoneal layer ; there are no large chloragogen cells like the
pear-shaped cells which cover the dorsal vessel, only a cover-
ing of much flattened cells: the supra-intestinal trunk is
united with the dorsal by a vertical sheet of mesentery. The
supra-intestinal vessel is directly concerned with the blood
supply of the esophagus. As already mentioned, the cesophageal
walls contain an extremely rich meshwork of blood capillaries ;
in specimens that happen to have been killed with this part
of the vascular system naturally injected, the walls are rendered
turgid by the contained capillaries, which have almost the
appearance of forming a continuous sinus; here and there the
supra-intestinal vessel is connected with this network by a
short tube.

The presence of infra-csophageal, vessels has been

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 551

described by myself in Eudrilus; as they occur in Libyo-
drilus they will probably prove to be characteristic of the
family. Similar vessels appear to occur in other earthworms,
but their distribution has not been much studied. In Libyo-
drilus as in Eudrilus they are paired trunks, but
they do not unite together at intervals as in the latter genus.
These vessels are, like the supra-cesophageal (supra-intestinal)
trunk, concerned with the blood supply of the ceesophagus ; they
are probably efferent trunks, taking the blood away from the
cesophagus, which has reached it by way of the supra-cesopha-
geal vessel.

Two mesenteries are connected with these vessels
aud with the ventral esophageal wall; these enclose
a space which appears to be completely shut off from
the celom of the segments. The arrangement of these
mesenteries differs in different parts of each segment. Towards
the middle of the segment, as is shown in fig. 17, the two
mesenteries originate from the lateral regions of the cesophagus
(the histological details of the cesophagus are not shown in the
figure) ; they are thrown into folds by the contraction caused
by the preserving fluid. These mesenteries unite considerably
above the ventrai blood-vessel, and thus enclose a space which
is roughly crescentic in outline but of considerable vertical
depth. The membrane consists of a faintly staining nearly
homogeneous core, covered on both sides by peritoneal cells,
of which the nuclei were alone visible; here and there small
groups of longitudinally running bands of muscular fibres, not
shown in the figure cited, are embedded in this homogeneous
layer. The figure, however, does show two longitudinally
running bands of muscle (m); these are special muscles
attaching the ventral surface of the cesophagus either to the
septa or to the ventral parietes. Above these, i. e. nearer to the
esophageal wall, are the paired sub-cesophageal trunks (4/. ves.) ;
in the section represented in the figure the two blood-vessels
are seen to be attached to the mesenteries, but in other sec-
tions they lie freely within the mesenteries, and further back
they may come to lie outside. These facts appear to show that

552 FRANK E, BEDDARD.

we are dealing here with a perihemal ccelomic space not found
in connection with supporting mesenteries of the blood-vessels ;
further forward in the segment the two lateral membranes
come to be attached to the intersegmental septum where this
forms the floor of the perihemal chamber. The supra-ceso-
phageal vessel is for the most part not enveloped in a peri-
hzmal space, but it is connected in parts by the membranes
with the walls of the esophagus; these gradually shift their
areas of attachment to the csophagus until they are nearly
continuous with the two membranes which enclose the sub-
oesophageal vessels.

The formation of perihezmal spaces is an interesting, but
not a perfectly new fact in the anatomy of Oligocheta,

I believe that I was the first to call attention to the fact
that, in an earthworm (Deinodrilus) belonging to quite a
different family, the dorsal vessel is for the greater part of its
course enclosed in a “pericardium.”” An almost identical
structure has been figured by Professor Spencer in Mega-
scolides australis. It is noteworthy that in both these
cases the “ pericardium” contains abundant perivisceral cor-
puscles which stain deeply, and appear to be merely the
younger forms among the corpuscles of the perivisceral fluid.
I suggested the possibility of these corpuscles being principally
formed in that space.

Similar corpuscles occur in the space which surrounds the
supra-cesophageal vessel in Hyperiodrilus and Helio-
drilus, and I have found them in Libyodrilus. I did not,
however, observe any proliferation of these cells from the lining
membrane of the perihemal space in Libyodrilus; there
were simply masses of these cells (fig. 17, corp.) lying here and
there within the space.

Fig. 21 represents the course of the sub-cesophageal vessels
in Segments 9—11.

They generally lie at some distance from the cesophageal
wall, and are connected at frequent intervals by branches with
the peri-cesophageal plexus. Towards the anterior part of
Segment 9 the two trunks are connected by a branch which

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 553

was prolonged backwards and downwards to the neighbour-
hood of the ventral vessel ; this branch (a) does not, however,
open into that vessel, but runs into one of the muscular bands
which unite the cesophagus with the parietes. Farther back
still each vessel gives off a branch which supplies the inter-
segmental septum; here its capillaries no doubt unite with
those of branches of the ventral blood-vessel. In the 10th
segment there was no connection between the two sub-ceso-
phageal trunks; each of them gives off a branch (a) which
supplies the muscle (m.) running from the cesophagus to the
parietes.

These branches pass along the walls of the perihzemal space
and reach the interior of the muscle, along which they pass to
its point of insertion; these two branches, of course, cor-
respond to the single cutaneous branch which I have described
in Segment 9. At the septum dividing this segment from the
11th a branch is given off from each vessel, which supplies not
only the septum but also the sperm-sac. In the next segment
the branches of the sub-cesophageal vessels are the same, and
therefore need no description. I imagine that these sub-
cesophageal vessels represent, wholly or in part, the lateral
trunks! of other earthworms, which appear at least frequently,
if not always,-to take their origin from the peri-cesophageal
blood network.

§ Nephridia.

In dissecting the worm the nephridia were quite obvious in
most of the segments of the body; and where visible, pre-
sented a paired arrangement which characterises all the
Eudrilide. In the posterior region of the body the nephridia
were particularly conspicuous, both in fresh and alcoholic
specimens, by reason of their opaque white coloration. In
the anterior segments of the body the nephridia were largely
adherent to the posterior septum of their segments, and did
not show the white opaque appearance of the posterior
nephridia. We have evidently here another instance of what

1 “ Tntestino-tegumentary.”

554 FRANK E. BEDDARD.

is very common among the terrestrial Oligocheta, viz. a sepa-
ration of the nephridia into two series, an anterior and a
posterior; this is seen, among other forms, in Pontodrilus
and in Microcheta. In the segments occupied by the
spermatheca, viz. in Segments 13—17,! no trace of nephridia
could be detected in a dissection. I am disposed to refer
this partial obliteration of the nephridia in this region of
the body (we shall see immediately that they have not abso-
lutely disappeared) to the enormous development of the
spermathecal sac, which cccupies most of the available space ;
the case is, in fact, analogous to that of many aquatic forms
in which the nephridia disappear with the appearance of the
genital organs.

Each nephridium (fig. 11, f.) has a funnel lying, as usual,
in the segment anterior to that which bears the external
pore. The nephridia of the posterior segments are enve-
loped by a mass of peritoneal cells; such at least appears
to be the nature of the very peculiar tissue illustrated in
fig. 10,c. I may remark here that both Heliodrilus and
Hyperiodrilus have a similar investment to the nephridia.
A covering of this kind which is simply an exaggeration of the
ordinary peritoneal layer is very characteristic of the aquatic
Oligocheta, but it has before now been recorded in earth-
worms; it was first described in an earthworm by Perrier in
Pontodrilus.

In sections of Libyodrilus which have been stained with
borax carmine this mass of cells is very deeply coloured; the
staining fluid has, however, chiefly affected not the protoplasm
of the cells, but innumerable spherical particles with which
they are crowded. Among these deeply-stained spherules are
masses of others, rather larger, which are not stained at all
(cf. fig. 6, c’.). The cells were not, however, always found
to be loaded with the products of their activity; occasionally
the nephridial tube is embedded in a mass of deeply-staining
cells, of which the nuclei were very conspicuous; the out-

1 There was some variation, probably due to different stages of maturity,
in the number of segments from which nephridia were apparently absent.

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 555

lines of the cells were not in every case so clear. Compara-
tively few of the cells contained agglomerations of secreted
spherules. The worm which furnished the material for the
section described here was immature ; but it must not be inferred
from this that the activity of the glandular cells surrounding
the nephridia does not commence until the animal is fully
mature ; in the youngest individual which I have been able
to study the cells in question were crowded with spherules.
Their activity is evidently intermittent.

In fig. 11 are represented three successive nephridia from
the post-clitellar region, belonging to as many segments.
The nephridia themselves are opaque white bodies, broader
towards the middle line of the body as there represented ; on
the upper surface it is quite easy to see, even without using a
lens, a single loop which appears darker than the surrounding
tissue on account of the thinness of its walls. This part of
the nephridium will be referred to in connection with the
minute structure of the glands. A narrow tube leading from
the broad end of the nephridium and perforating the septum
traced in the funnel, situated in the segment in front, is quite
easy to make out; so is another tube which passes direct to
the body-wall in front of the ventral pair of set; this tube is
the external duct of the nephridium. Anyone who was con-
tented, as were some of the earlier investigators, with dissec-
tion only, would be satisfied that in such a drawing as fig. 11]
the nephridium was sufficiently displayed. The nephridium
itself is shown, the funnel and the duct leading to the exterior.
But there is apparently no external orifice corresponding to the
place where the duct appears to perforate the body-wall on
its way to the exterior.

In examining the body of the worm with a lens I did not
succeed, as already mentioned, in discovering the nephridio-
pores, which are generally quite easily visible, even without a
lens.

This failure to find the nephridiopores was at once accounted
for when I examined a fragment of the cuticle stripped off and
mounted in a drop of water.

556 FRANK E. BEDDARD.

Besides the minute orifices of the unicellular glands of the
epidermis (fig. 8), which here, as in other species, show a
cross-like form illustrated in many memoirs upon the anatomy
of earthworms, there were other larger orifices of a circular or
perhaps slightly elliptical form. In the figure cited the number
of these apertures relatively to those of the unicellular glands is
shown ; they are extremely numerous in each segment, and have
no particular regularity of arrangement.

These apertures obviously suggest the external pores of the
nephridia, which are so numerous in Pericheta and other
forms with diffuse nephridia. In Perichezta the apertures
in question are of moderate size, though considerably smaller
than the apertures through which the sete protrude. They
are continuous with a little cylinder of chitinous membrane,
which is, of course, the lining of the nephridial tube.

In Libyodrilus the apertures that I have referred to were
smaller than those of Pericheta, and I could not detect the
involution of the chitinous membrane. Otherwise I should
have thought myself justified in stating, if necessary without
any further investigation, that Libyodrilus belonged to that
group characterised by a diffuse, irregular nephridial system.

This suggestion, however, appears to be somewhat at variance
with the statement that Libyodrilus, like other Eudrilids,
possesses paired nephridia, one pair to each segment. I have
found, however, by a series of sections, that the body-wall is
permeated by asystem of branching and anastomosing
canals, opening on to the exterior by numerous aper-
tures, and connected on the other hand with the
paired nephridia. I took these canals at first for blood-
vessels, to which they bear not a little resemblance ; indeed, I
am still unable to find any very marked histological difference
between these tubes and the blood-vessels. That they belonged
to a different system was shown by the fact that they were
frequently found to be accompanied by blood-vessels. It is
hardly perhaps necessary to say that there was no connection
between these tubes and the blood capillaries; the former never
contained blood-clots, while it was impossible to follow out a

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 557

blood-vessel through more than two or three consecutive
sections without finding coagulated blood in its interior; and
it is not difficult in well-preserved earthworms to detect the
blood-clots, nor is it easy to confuse them with any other
structures as a general rule.

The tubes that I have referred to consist of larger trunks

with a definite arrangement, and smaller vessels which form a
plexus ; the longitudinal and circular muscular coats were per-
meated by them. It is, perhaps, necessary to explain here
that the tubes in question have nothing to do with the
** Lymphspaltraume ” which Dr. Kiikenthal has tigured and
described in certain Oligocheta, particularly in Tubifex; I
have met with spaces similar to those, and crowded with lymph-
corpuscles, in various genera of earthworms. I found them to
be particularly abundant in the two genera Heliodrilus and
Hyperiodrilus described in a recent number of this Journal
(1). They consisted in those genera, as Dr. Kikenthal has
pointed out for other forms, of spaces, with no special walls,
between the muscular fibres. I quite agree with Dr. Kiiken-
thal in regarding these as the first beginning of the lymphatic
system of the Vertebrata, in which group the finest branches
of the lymphatic system are without intrinsic walls. The
tubes which I describe here have definite though thin walls,
darkly stained by borax carmine ; the substance of which they
are composed is granular in appearance, and there are evident
nuclei attached here and there to the outside of the walls. The
principal trunks run in a longitudinal and a transverse direc-
tion; there are four main longitudinal trunks, which
run on a level with the four pairs of sete; they are
continuous from segment to segment.
. These tubes are of considerable calibre, almost equal in
diameter to one of the setz, or even greater; it is not particu-
larly useful to give accurate measurements of them, for the
reason that they varied considerably in size from place to
place, contracting here and expanding there. Their walls are
thin, but quite as thick as those of blood-vessels of about the
same size, ~

558 FRANK E. BEDDARD.

The longitudinal muscular coat consists, like that of other
earthworms, of fibres embedded in a matrix of connective
tissue. As I have already mentioned (supra, p. 545), this con-
nective tissue forms a thickish layer below the level where the
muscular fibres end; it is in this layer that the longitudinal
vessels run. At the implantation of the couples of sete the
longitudinal vessels come to lie within the arch formed by the
two sete—which converge at their deep ends, and diverge super-
ficially. Just behind the setz the longitudinal vessel gives offa
wide branch, which gradually gets nearer and nearer to the thin
peritoneal epithelium, Ultimately (fig. 7) it comes to lie within
the body-cavity, surrounded at first by a prolongation of the
almost hyaline connective tissue of the longitudinal muscular
layer. This covering finally disappears, and the tube comes to
an end; in a few cases I have traced it into connection with a
very rudimentary coil of nephridial tubules. I have already
mentioned that in the clitellar segments the nephridia are not
visible with a lens. The tube lying within the celom was
always accompanied by a blood-vessel of about the same dia-
meter, and I have observed the two to be suspended by a
common mesentery. The longitudinal vessels are also pro-
vided with numerous branches, which pass among the longi-
tudinal muscles; some of these branches are smaller, some
larger ; a few of the larger ones pass obliquely upwards in the
direction of the circular muscular layer ; the tract which these
branches traverse is free from muscular fibres; traced up-
wards, these branches are seen to open into a circular
vessel, which runs right round the body; these circular
vessels, which are repeated metamerically, lie just between the
circular and longitudinal muscular coats, close to an important
nerve, which also passes right round the body (see fig. 20).
The circular vessels are of considerable width, and can be easily
recognised in sections examined by quite low powers; their
outline is somewhat crenated, which probably means that they
can be expanded or retracted in accordance with the move-
ments of the body. The fact that the vessel lies between the
two muscular coats, instead of within the substance of one of

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS, 559

them, probably ensures less injury through pressure. These
tubes, like the rest, appeared to be entirely empty, or, at any
rate, to be filled with an absolutely clear fluid; they give off
numerous branches on all sides; the commencement of the
branches, which run along the segments from end toend—that is,
in a longitudinal direction—could be seen in transverse sections
as elliptical foramina in the walls of the tube (see fig. 14). The
branches which pass downwards ramify among the longitudinal
muscles, and even on the dorsal side of the body of the worm,
Where there are no conspicuous longitudinally-running trunks
the branches reach the thick layer of connective tissue which
lies to the inside of the longitudinal muscles ; arrived there,
they pass in different directions, longitudinal as well as trans-
verse. Besides the main transverse trunks running between
the two muscular coats I observed vessels of less calibre, run-
ning, as shown in the diagram (fig. 16, a), just below the peri-
toneum, and apparently continuous right round the body.

The branches which pass upwards could be traced as far as
the epidermis; they were constantly, like the other branches,
accompanied by blood-capillaries. I could not detect the
actual orifices, but at frequent intervals the non-glandular cells
of the epidermis were closely crowded together. At these
points I imagine lie the external orifices which are so evident
upon the cuticle when it is stripped off and examined.

In no case were cilia observable along the course of the
tube. It is clear that this system of tubes forming a plexus
within the muscular coats or the body-walls is a new form of
excretory organ, and unlike any that has been hitherto described
in any Oligocheete.

A plexus of nephridial tubes, not interrupted by the septa,
occurs in a good many forms; but it has been hitherto found
to lie in the celom, or is at most partly retro-peritoneal ;
besides, in the genera (Pericheta, Megascolides, &c.)
which have excretory organs of that pattern the tubes are to
some extent ciliated; in those forms there are numerous
external pores, and so the integument is perforated by numerous
nephridial tubes, These may even branch on their way to the

560 FRANK E, BEDDARD,

exterior, as I have figured in Acanthodrilus (2, pl. xxx,
fig. 1), and as Prof. Spencer has figured in Megascolides
australis (9, pl. vi, fig. 21); but there is no branching
and anastomosis combined, such as is described above in
Libyodrilus; in Pericheta, &c., the network is confined to
the coelom, and is formed out of the proximal part of the
nephridia; in Libyodrilus the network is in the thickness of
the body-wall, and is formed out of the distal part of the
nephridia. It has been already explained that the paired
nephridia of the clitellar segments become rudimentary,
though they do not entirely disappear; if the ccelomic part of
the nephridia were to disappear—-and they have all but done
so in the clitellar region—the nephridial system would
consist merely of a rich network of smaller and larger
tubes in the thickness of the body-wall.

The figure (fig. 14) which illustrates the arrangement of this
portion of the excretory system represents a combination of
several sections. It should be stated, however, that it is not
meant as a diagram ; such an arrangement as is there depicted
might well occur and be visible in the thickness of any one
section. To thoroughly demonstrate the network in the longitu-
dinal muscular layer probably needs some process of injection
which I have not been able to apply. Had I been aware of
this interesting network of fine tubes in the skin I should have
certainly attempted some injection while I had living speci-
mens at my disposal. As it is, Iam inclined to think that I
have not in my drawing done full justice to the complexity of
the meshwork ; the scheme (fig. 16), which is purely diagram-
matical, expresses my idea as to this network. In fig. 14 two
longitudinal ducts (d@) are shown, and the connection of each
with the circular vessel: the latter (c) is represented as cut at
different levels; the thickness of the walls of the tubes ramifying
among the longitudinal muscles is, perhaps, rather exag-
gerated. At aa branch of the longitudinal trunk d is seen
to leave the muscular layer, and to project into the body-cavity,
still surrounded by a connective-tissue sheath; this branch
joins one of the paired nephridia,

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 561

Fig. 7 represents one of the longitudinal trunks seen in
longitudinal section ; it runs for the greater part of its course
in the thick layer of gelatinous tissue which bounds the longi-
tudinal muscles below; at the implantation of the sete it
passes up into the muscular layer itself. These vessels, as
will be seen from the figure, have a somewhat undulating
course; they are occasionally diverted by a large blood-vessel.
I have not shown in this figure many branches passing
from the longitudinal trunk, because these were not visible,
except at a, in the section of which it is a drawing. At
N the duct of a nephridium is seen to join the longitudinal
vessel. In the anterior segments of the body the ducts of the
nephridia pass down in close contact with the hinder septum
of their segment to join the integumental network; in the
posterior segments, as is shown in fig. 7, they are not in
such close relation to the septum.

A connection of the nephridia with the pharynx
occurs in this earthworm. The ventral wall of the pharynx,
as in other Oligocheta, is formed of but little more than the
lining epithelium ; the dorsal] wall, on the contrary, is extremely
thick, owing to the mass of muscles which overlie the epi-
thelium. Here and there the epithelium of the ventral
pharyngeal wall gives off very short tubular diverticula
which are connected with nephridial tubules; these same
nephridial tubules can be traced in the other direction into the
longitudinal vessels of the body-wall and so to the exterior
(fig. 15).

An opening of nephridia into the stomodeal region of the
alimentary tract in earthworms was first discovered by myself
in Acanthodrilus multiporus; subsequently Spencer
found numerous nephridial apertures into the pharynx in
Megascolides.

In fig. 15, the opening of a tubule into the ventral side of
the pharynx is shown; these apertures are not numerous, and
appear to be regularly paired ; the epithelium of the pharynx
dips down, its cells gradually getting shorter ; the nephridium,
into which this short diverticulum opens, forms a coil within a

VOL. XXXII, PART IV.—NEW SER. PP

562 FRANK E. BEDDARD.

layer of connective tissue and muscles belonging to the
pharyngeal wall.

The development of these nephridia, so far as I have been
able to trace it (see on p. 574), seems to indicate that the
network in the body-wall is a secondary arrangement. It is
undoubtedly necessary to be very careful in arguing from on-
togeny to phylogeny, but so far as the facts go they seem to
point in this direction. In this case it will be hardly permis-
sible to compare the network to that of flat-worms, particularly
of Cestodes ; in the Cestodes the nephridial network appears
to exist not only in the mesoderm, but also immediately
beneath the cuticle in the layers of the body-wall.

On the other hand, it may be permissible to compare these
structures with something of the same kind in the Nematoids.
These worms usually possess at least remnants of a celom
which is well developed, and with a limiting epithelium in
Gordius;! in Ascaris, Joseph has stated (‘ Zool. Anz.,’ Bd.
v, p- 603) that it is possible to inject from the excretory pore
a fine system of canaliculi lying between the muscle-cells, and
in fact pervading the body generally, while Schneider (Mono-
graph Nematoden) has figured the branches in Ascaris. In
any case the canals running in the lateral lines are not a little
suggestive of the longitudinal canals which I have described in
Libyodrilus. In the Acanthocephala also, where there is a
coelom, the lemnisci, which have been stated to occur in certain
Nematoids also by Hamann, appear to be processes of the
body-walls depending into that ceelom ; they are permeated by
tubes which are connected with a system of vessels in the
body-walls. These tubes may be excretory in nature, though
they have been described as vascular. In any case I am not
aware that a similar network of tubes has been described in
the integument of any Oligocheta ; this worm, therefore, has

1 According to Vejdovsky’s recent work upon the structure of Gordius
(‘ Zeitschr. f. wiss, Zool.,’ Bd. xliii) these Nematoids show many points of
affinity with segmented worms; so much so that they should be removed
altogether from Nemathalminthes. The excretory organs appear to be repre-
sented by coelomic canals,

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 568

an excretory system which is at least of interest in being
formed on quite a novel plan.

§ Male Reproductive System.

The testes are paired structures lying in Segments 10 and
11; they are not enclosed in any sacs, and are quite indepen-
dent of the sperm-sacs. The gonads are remarkable on account
of their great length and thinness; the extremities are frayed
out, as is usually the case among earthworms, into a number of
jagged processes.

S perm-sacs.—There are two pairs of sperm-sacs attached
to the anterior walls of Segments 11 and 12; each sac of each
pair is quite independent of its fellow. In the living worm
the sperm-sacs have a whitish appearance with the exception
of the basal part, which is greyish brown. In this part of the
sac the Gregarines, which appear to be parasitic in all earth-
worms, are chiefly lodged, and the colour is due to brown pig-
ment deposited round their cysts. As to the histological
structure of these sacs, they agree with those of other earth-
worms in having their cavity greatly broken up; the sperm-
sacs on their first appearance are solid, except for a very small
cavity which communicates with the interior of the segment in
front of that in which they lie. There are no seminal reser-
voirs, and thus the testes, and funnels of the vasa deferentia,
depend freely into the ccelom.

Sperm Ducts and Funnels.—In all the members of the
family Eudrilide (excluding from this family Eudriloides
and Pygmeodrilus) with the exception of Nemertodrilus
griseus, and perhaps Preussia siphonocheta, the sperm
ducts are dilated before their connection with the funnel.
In Libyodrilus there is no such widening of the sperm ducts
either in the neighbourhood of the funnels or anywhere else.
The funnels themselves occupy the usual position in Segments
10 and 11; those of Segment 11 are more nearly opposite to
the sperm-sacs than to the testes ; the sperm ducts are narrow
tubes of the usual appearance, but are unusual in passing
through the segments which they traverse at some distance

564 FRANK E. BEDDARD.

from the body-wall; their course is straight without any
windings; the two sperm ducts of one side appear to unite
just behind the intersegmental septum 12—13; asa matter of
fact they do not unite here or anywhere else until at their
actual orifice; they lie, however, in close contact. In trans-
verse sections the sperm ducts are seen to have quite the
ordinary structure ; there is no development of muscular fibres
round them such as occurs in Hudrilus. The funnels are
large and their walls are much folded.

The atria have the tubular form which is characteristic of
the family ; but they are short, and usually contained entirely
within the 18th segment. Towards the external aperture they
become narrower. These organs have the “ nacreous”’ appear-
ance of the corresponding organs in EKudrilus, which is due
in both cases to the great development of the muscular coat.
This reaches an extraordinary thickness in Libyodrilus,
which is thus remarkable even among the Eudrilide.

The external orifice is single, and is situated upon the summit
of a rounded elevation occupying the middle of the ventral
surface of the body and extending over a portion of Segments
17 and 18. The epithelium which covers this elevation is fur-
nished with numerous glandular cells, conspicuous on account
of their dark staining. With each atrium is connected a sac
in which lies a single short penial seta. The sac communicates
with the distal part of the atrium that is embedded in the
thickness of the body-wall.

The penial seta is illustrated in fig. 8; it is remarkable
on account of its shortness and its blunt rounded free extremity,
which is not ornamented.

The atrium is lined throughout with epithelium, which is
separable into two strata.

The cells, however, had not in any case (I examined the
atria of three individuals) the extremely glandular appearance
of the corresponding epithelium in Eudrilus. This may, no
doubt, be owing simply to a cessation in the secretory activity
in the cells.

Tn nearly all the Eudrilide the vasa deferentia open into the

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 865

atrium. Perrier, who was the first to make known the struc-
ture of Eudrilus, described the vasa deferentia as opening
into a pouch, into which also opened the atrium, regarded by
him as the equivalent of the prostate of other earthworms.
It is so figured in his memoir (10, pl. ii, fig. 26). I myself
showed that, unless we were dealing with different forms, this
description was incorrect ; in specimens of Eudrilus coming
from several distinct localities I found that the vasa deferentia
opened into the structure that had been termed prostate. This
glandular tube, I pointed out, was itself divided into. two
separate tubes by a longitudinal septum not visible-or hardly
visible externally. Into the longer of the two tubes, at a point
corresponding with the apex of the shorter compartment, open
the vasa deferentia, each by its own orifice. The structure of
Eudrilus has been recently reinvestigated by Dr. Horst (7),
who gives a diagram (7, p. 6) agreeing with my figure (15, pl.
xxxi, fig. 15), and remarks that “V’appareil génital male est
exactement conforme a la description détaillée qui M. Beddard
a publiée de cet organe.”

In Teleudrilus Rosa describes the opening of the vasa
deferentia into the atrium not far from the anterior extremity
of the latter, i. e. not far from their point of opening on to
the exterior. The same thing occurs in Heliodrilus
and Hyperiodrilus; in Nemertodrilus the communica-
tion between atrium and vasa deferentia takes place, as it
apparently does in Teleudrilus, not far from the external
aperture of the former.

In Libyodrilus the arrangement is very interesting. On
a dissection of the worm I could not follow the course of the
vasa deferentia beyond the point of opening of the atrium;
this was due to the fact that the two tubes, which retain their
distinctness though they are in very close juxtaposition, at
that point perforate the muscular tissue of the atrium; they
pass up, at first within the thickness of the muscular tissue, and
later within the epithelium, to nearly the summit of the atrium.
At this point they open into its lumen by a common orifice ;
the two vasa deferentia unite just at the orifice ; the cilia are

566 FRANK E. BEDDARD.

not continued on to the epithelium which lines the atrium.
At the orifice of the vasa deferentia they are particularly long,
and project for some way into the interior of the atrium. In
Libyodrilus, therefore, the atrium has come to be, as it is in
such worms as Nais, a kind of continuation of the vasa
deferentia. Among the Eudrilide the point of opening of the
vasa deferentia gradually moves down the atrium until, in
Nemertodrilus, it comes to be placed near to the external
orifice.

This series of facts lends further support to the original
suggestion of Vejdovsky, which was strengthened by my own
observations, as to the homology between the atrium of the
aquatic genera and the prostates of the terrestrial genera.

If we are to follow Dr. Benham (11) in objecting to this
identification we shall be plunged into obvious difficulties, for
Dr. Benham considers “that a portion of the prostate in
Pericheta, Eudrilus, and other genera, in which the sperm
duct and the prostate join, is probably the homologue of the
atrium of Tubifex.”

I should mention that the interior of the atrium in Libyo-
drilus is not divided by a septum, and that the lining epithe-
lium is greatly folded ; this is carried to such an extent that in
longitudinal sections the lumen appears to be furnished with
tubular diverticula; transverse sections are required to show
that these are merely foldings of the epithelium.

§ Female Reproductive System.

The extraordinary development of ccelomic pouches in con-
nection with the different parts of the female reproductive
system is the most distinguishing character of the Eudrilide,
and also occurs in the present species.

Spermathecal Sac.—In dissecting the species which
forms the subject of the present memoir, the most conspicuous
organ of the body is a large sac which lies upon the dorsal
surface of the esophagus. This structure is illustrated in
fig. 1, which represents a general view of such a dissection, and

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 567

more diagrammatically in fig. 9; the latter figure shows the
relations of the sac to neighbouring organs. The sac is of a
brownish-yellow colour, and-of an irregular elongated form ;
it is marked here and there by furrows, which indicate that it
is capable of being extended to a much greater size; it com-
pletely covers the dorsal vessel and cesophagus, to which it is
attached by a series of thread-like ligaments. The sac extends
when fully developed from the anterior boundary of Segment
13 to the posterior extremity of Segment 18; it gives off on
either side three diverticula; from the first pair of those
diverticula arise the oviducts which pass outwards so far as
already stated upon the 15th segment. Anteriorly the sac
divides so as to embrace the cesophagus, round which it forms
a ring; immediately beneath the cesophagus the two halves
become reunited, and pass forwards and downwards in close
contiguity to the septum separating Segments 12 and 13. In
this region the sac is very much narrower; at the nerve-cord
the sac again divides so as to surround the cord, and the un-
paired duct which arises from this perineural ring opens on to
the exterior in the 13th segment. The sac was found to be
filled with bundles of spermatozoa of irregular shapes and
sizes; a fragment of this sac taken from the living ovum and
teased up upon a slide showed that it was lined with peculiar
granular cells. The cells are so easily detached that it appeared
as if they were free in the interior of the sac; an examination
of hardened material showed that this was not so, and that the
cells form a lining to the sac. The fresh cells are of an irre-
gular, generally somewhat elongated form; occasionally they
are branched; the protoplasm of the cell is very clear and
transparent ; in the protoplasm are a quantity of spheroidal
granules, usually massed together in the widest part of the
cell; the cells resemble very closely the peritoneal cells which
cover the alimentary tract, but the granules in their interior
are greyish instead of brownish green.

In sections of the mature spermathecal sac (see fig. 12)
its walls are seen to be muscular, with a coating and lining of
epithelium.

568 FRANK E. BEDDARD.

Embedded in the thickness of the muscular coat is a dense
layer of tissue which stains darkly ; this layer is of some thick-
ness, but apparently homogeneous throughout ; here and there,
for instance, in that part of it which covers the egg-sac as
shown in fig. 12, favorable sections demonstrate that it is
a membrane, e/; it is usually much bent, showing the zigzag
contour illustrated in the figure; in all probability it is
of the nature of elastic tissue, and permits of the easy recovery
of the original dimensions of the sac after it has been unduly
distended with sperm.

The lining epithelium of the sac is, as a rule, more than one
cell thick ; there appears usually to be an innermost layer of
fairly regular smallish cells; here and there an extensive pro-
liferation of these has taken place. The whole character of
the cellular lining of the sac indicates that it is not formed as an
invagination of the epidermis; nor, as will be seen later, is
it. It has been said that the anterior wall of the sac passes in
close relation with intersegmental septum 12—13. In
longitudinal sections—which are, I may remark, much more
useful for studying the structure of worms than trans-
verse sections—the anterior wall is seen to be not merely
in continuity with the septum; it is formed by the
septum itself.

Ovaries.—I could not find the least trace of these organs
in fully mature worms, either by dissection or in sections.
That the ovaries should be evanescent structures in an earth-
worm is rather unexpected. They are present in young stages,
where they exhibit the usual position and structure, though
like the testes they are much elongated and very narrow.

Egg-sacs.—In a dissection the egg-sacs can hardly be dis-
tinguished from the great unpaired spermathecal sac in which
they lie; a careful inspection shows that at the point where
the oviduct arises there is a slight bulging outwards of
the sac; this has a browner coloration than the rest of
the walls of the sac. This, however, is no more conspicuous
than is represented in fig. 9, where the spermathecal sac
and the oviducts are displayed in aslightly diagrammatic form.

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 569

In transverse sections the egg-sac of each side is seen to be a
spherical body lying within the spermathecal sac, and supported
at one side by the wall of this sac; its cavity, however, is
nowhere continuous with the cavity of the spermathecal sac;
the two structures are so far perfectly independent, and appear
to be as it were accidentally associated. I shall show later
(on p. 574) that there are also embryological reasons for
regarding the egg-sac as entirely unconnected with the great
unpaired sac in which it hes. The egg-sac of all earthworms
in which it has.been found, with the exception of the Eudrilide,
lies in the 14th segment, attached to the anterior wall of this
segment. In Libyodrilus there is a septum separating the
13th from the 14th segment; but this septum does not traverse
the spermathecal sac ; hence the egg-sacs do not depend from
any intersegmental septum in the fully mature worm.

In its microscopical structure the egg-sac presents no pecu-
liarities of any particular interest. Its cavity is, as is usual
with this organ, subdivided into numerous chambers, packed
with ova and germinal cells in various stages of development.
The walls of the sac are formed largely of muscular tissue with
abundant nuclei; the compartments which are formed by in-
growth of this layer are also muscular, but seem to be irregularly
lined with granular peritoneal cells. The relations of the egg-
sac to the spermathecal sac are shown in fig. 12; it will be
noticed that it projects into the interior of that sac, and in other
sections, taken at a different level from the one figured, it
appears to lie freely in the interior of the sac. It is, however,
easy to see from the histology of the parts concerned that the
egg-sac is not really within the spermathecal sac ; the walls of
the latter are not perforated ; they are merely pushed in by
the egg-sac: these points will be understood by a reference to
fig. 12.

The figure shows the egg-sac just where the oviduct opens
into it; two parts of the oviduct are shown; the lettering od
is placed in the oviduct where it commences to widen out into
the funnel; to the right of the figure lies a section of the
narrower part of the tube. Running round the walls of the egg-

570 FRANK E. BEDDARD.

sac where it projects into the spermathecal sac isa band of elas-
tic tissue (e/.), which I have already spoken of in connection with
the spermathecal sac; it will be noticed that this band of tissue
does not belong to the egg-sac but lies outside it, thus showing
that the egg-sac does not really project into the spermathecal
sac through its walls; it is, therefore, unnecessary to state that
there is not any communication between the internal cavities
of these organs. They are perfectly independent of each other
though in very close contact. The egg-sacs have, in fact, the
same relation to the spermathecal sac that the sperm-sacs have
in certain earthworms to the “seminal reservoir.” For
example, in Dichogaster the sperm-sacs are enclosed in large
thin-walled seminal reservoirs. Bergh distinguishes these sacs
in Lumbricus as “ Samenkapseln”’ (= seminal reservoirs),
and ‘“‘ Samenblasen”’ (= sperm-sacs). The former may, and
generally do, enclose other organs, e.g. the ventral blood-
vessels, and, in Dichogaster, the sperm-sacs themselves.

The mature ova have a thick, darkly-staining membrane,
in which I could find no striz such as occurin Hyperiodrilus
and Heliodrilus.

Oviduct.—lIn describing the external characters of this
worm, I have already remarked upon the abnormal position
of the oviducal pores. Their position ‘is seen in longitudinal
sections to be rather less abnormal. This is due to the fact
that the septum which separates Segments 14—15 is attached
to the middle of the latter segment, just in front of the
sete ; accordingly the oviducal pores are really situated
within the fourteenth segment,reckoning by the
septa and not by the external furrows; they are, how-
ever, as in Teleudrilus, just in the boundary line between
the two segments.

In dissections of the worm (see fig. 1) the oviduct is
seen to pass in a straight line without any windings from the
egg-sac to the exterior; its length is considerable when com-
pared with the oviduct of Lumbricus, for example; in dissec-
tions the egg-sac cannot be seen, as it is entirely enclosed by
the spermathecal sac. In trahsverse and longitudinal sections

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 571

of the genital region of the worm the oviduct is seen to be
ensheathed in a tolerably thick muscular coat, as is the case
with all other Eudrilide. In the young the oviduct is con-
nected both with the interior of Segment 13 and with the egg-
sac in the way that is generally found in earthworms. In the
adult worm the large spermathecal sac nearly fills the 13th
segment and encloses the egg-sac, though, as has been already
said, there is no communication between thetwo. Near to its
opening into the egg-sac the oviduct is invested with a specially
thick muscular coat ; it opens into the interior of the egg-sac
by a not very extensive funnel ; just before its opening into the
egg-sac it gives off a narrow branch which passes into the
wall of the spermathecal sac, becoming narrower as it proceeds,
and ultimately opens into the interior of the sac by a very
small aperture. The aperture into the spermathecal sac can-
not be of any functional importance, since the ovary is an
evanescent organ, and there are no ova to be found in the sac
of the adult and nearly adult worm. At the point where the
branch of the oviduct opens into the spermathecal sac the
walls of the latter are specially thickened, and project into its
interior in the form of a rounded pad. The formation of two
openings from the originally single aperture is, of course, due to
the growth of this spermathecal pouch, which cuts off the open-
ing of the egg-sac into the body-cavity, and at the same time
that part of the oviducal funnel which opened in the younger
stages into the egg-sac.

V. Description of some Young Stages.

The youngest individual which I examined was rather more
than half an inch in length. I imagine that it could not have
long escaped from the cocoon.

It is quite evident from an examination of this embryo that
the present species is hatched in a much more imperfect con-
dition than either Lumbricus or Acanthodrilus, the ouly
two genera of whose development we have at present any
knowledge. In the embryo of Lumbricus, according to
Bergh, the testes and ovaries are present while it is still within

572 FRANK E. BEDDARD.

the cocoon; in the New Zealand Acanthodrilus multiporus,
whose development I propose to treat of in another paper, not
only the gonads but the funnels of the sperm ducts and ovi-
ducts are fully recognisable in advanced embryos extracted
from the cocoon.

I could not find any trace of the gonads in the young Libyo-
drilus. This may not be enough to prove that they are not
present in the stage which I examined; but it seems unlikely
that they would have been overlooked had they reached any-
thing like the development that these organs have reached in
the corresponding stages of Lumbricus and Acanthodrilus.

The gonads being absent, or at most very slightly developed,
it is not surprising that there is no trace whatever to be found
of this duct or of the terminal apparatus with which the ducts
are connected. All these structures then appear to be later
developments than in Acanthodrilus or Lumbricus,

One of the most striking features in sections of the embryos
of Acanthodrilus is the enormous quantity of perivisceral
corpuscles present; they are so numerous as to occupy the
greater part of the celom. The amount of the perivisceral
corpuscles in Libyodrilus is not at all large. M. d’Udekem
(12, p. 31) has commented upon the great abundance of peri-
visceral corpuscles in the young Tubifex just escaped from the
cocoon.

The immature condition of this stage was also shown by the
minute structure of the alimentary canal and by its contents.
I thought it possible that I might detect some trace of the
pouches connected with the esophagus, which are so important
and characteristic a feature of most Eudrilide; but there was
not the remotest trace visible; the three gizzards were just
commencing to be formed, but they had not acquired anything
like the proportions which they ultimately assume. In describ-
ing the structure of these organs in the adult worm I have
mentioned that they lie in three consecutive segments, and are
separated from each other by a short, soft-walled segment of
gut. These tracts (cf. fig. 1) are of very much less extent
longitudinally than the gizzards themselves; in the youngest

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 573

worm (fig. 18) the reverse is the case ; the interspaces between
the gizzard are very much longer than, about three times as
long as, the gizzards.

The intestine is lined by a columnar epithelium, the cells of
which are still loaded with spherical granules ; they resemble,
in fact, the cells of the intestine in the advanced feetus of
Acanthodrilus. The interior of the intestine seems toindicate
that the young worm had not yet commenced to swallow the
soil in which it was found ; the intestine contained a granular
substance, which presented the appearance of a coagulated
albuminous fluid; in this were embedded a large number of
sete of small size; the structure of the sete showed that they
probably belonged to the same species; at least there was
nothing to indicate that they did not. So far as my experience
at present allows me to say, it is not possible to distinguish the
setee of the Eudrilide from those of the Lumbricide, and, in-
deed, of most other groups of earthworms. The question is,
How did these setz get into the intestine of the young worm?
I noticed in the body-cavity of the same individual a few
setze embedded among the peri-intestinal ccelomic cells; these
were evidently set of the same individual that had become
freed into the body-cavity ; if they can work their way through
the intestinal epithelium we have an explanation of how they
come to be found in the cavity of the intestine. It seems
more probable, however, that they were originally swallowed,
and in this case it appears likely that the cocoon con-
tains up to a comparatively late period more than
one developed embryo; the individual or individuals
which succeed in reaching the full term of their development
must feed upon the others when alive or upon their dead
bodies.

The structure of the gizzard in longitudinal section is shown
in fig. 18. The circular muscles appear to be derived from
columnar cells, the nuclei of which are shown; whether these
cells are produced by the division of the hypoblastic cells, or
from the splanchnopleure, which is most probable, I am unable
to say. The muscular fibres themselves are arranged in a

574 FRANK E. BEDDARD.

more or less regular fashion along both sides of those cells.
The ty phlosole is a simple fold.

The integument in the youngest specimens at my disposal
was chiefly interesting on account of the structure of the
epidermis and of the longitudinal muscular coat. In the epider-
mis the gland-cells are limited to an area in the middle of each
segment ; it isevidently here that they first appear. In the
longitudinal muscular coat the fibres are embedded in the same
gelatinous material recognisable in mature worms, but this is
proportionately less in amount, and there is no thick layer of
it separating the longitudinal muscles from the peritoneum.
The fibres themselves approximate in their arrangement very
closely to Lumbricus; that is to say, they are disposed in
double columns, which are not, however, perfectly regular.

The nephridia in these embryos presented features of con-
siderable interest. The paired nephridia were recognisable in
all the segments after, and including, the fourth ; the first pairs
are not covered with the coating of peritoneal cells containing
numerous spherules, as are the succeeding pairs. This invest-
ment of the nephridia commences gradually, inasmuch as those
of the fifth pair had only a very slightly developed peritoneal
layer, in which, however, the granules of secretion, which stain
deeply, were perfectly plain.

The anterior pairs of nephridia are, as also in the adult worm,
closely attached to the posterior wall of their segment. They
are not bound thereto by any mesentery, but lie in actual
contact with the septum. The duct leading to the exterior
passes down, in the case of the anterior nephridia at any rate,
also in close contact with the septum; it is much coiled, and
so in longitudinal sections appeared, in every section, cut at
right angles to its course. Arrived near to the insertion of the
septum on to the body-wall the duct joined a continuous
longitudinal duct, passing along the first few seg-
ments of the body and putting the nephridia of these
segments into communication with each other. On
each side of the nerve-cord this duct was found having similar
relations to the nephridia of the segments through which

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 575

it passes. The duct is not embedded in the longitudinal
muscular coat or in the peritoneum, but appears to lie in the
coelom.

If this duct had turned up two or three years earlier than it
has, I should have been able to claim it as a discovery of some
importance. The account given by Balfour of Hatchek’s state-
ments about the developing nephridia of Criodrilus gave
additional strength to them, although Balfour considered them
to need confirmation, and the apparent confirmation which is
given by a young, only just out, foetal earthworm would have
almost clinched the matter. The discovery even of Meyer and
Cunningham that the adult Terebella conchilega is fur-
nished with such a longitudinal duct was regarded of the
greatest importance in that connection. As it is, the new
fact recorded here has lost a good deal of importance by appear-
ing too late. Since Hatchek’s paper the aspect of the nephri-
dial question has changed. Nevertheless it is of some little
importance. In Perichxta there does not appear to be, in
the adult at any rate, a pair of distinct longitudinal ducts
passing from segment to segment ; there is simply the irregular
network of each segment which is connected here and there
with that of adjoining segments. Spencer, however, has
described (9) in Megascolides australis a pair of longitu-
dinal ducts which run from segment to segment, putting the
complicated nephridial network and the large paired nephridia
of successive segments into direct communication. It is not yet
known how early these ducts appear in Megascolides, but
it is interesting to notice that they appear at least compara-
tively early in the development of Libyodrilus. In theadult
worm they persist (fig. 7), but their importance is lost in the
complications of integumental network. For all that I know
the paired longitudinal ducts may be the first of the nephridial
system to appear. I very much regret that my material does
not enable me to settle this question. At any rate, it is certain
that in Libyodrilus the integumental network con-
nected with the paired nephridia is preceded by
paired longitudinal ducts uniting those nephridia.

576 FRANK E. BEDDARD.

In the stage that I am describing here, there was no dis-
coverable trace of the integumental network ; the external
pores of the nephridia were apparently paired, but as they
were very minute, I may possibly have overlooked other orifices.
It should be mentioned that the nephridial funnels exist at
this stage; they lie ina plane considerably above that in which
the longitudinal ducts lie, and there is, therefore, no chance of
my having confounded the longitudinal duct with the tube
leading to the funnel.

Generative Organs.—lIn a specimen without a clitellum,
but as large or nearly so as some specimens with a clitellum,
the spermatheca was not visible from the exterior; that is to
say, there were no traces that could be seen, even with a lens,
of the external pore, which, though small, is quite evident in
mature or nearly mature specimens.

Longitudinal sections revealed a very remarkable condition
of the female reproductive apparatus.

The ovaries, receptacula ovorum, and oviducts present the
same arrangement as in more typical genera of earthworms
(Lumbricus, for example).

The epidermis in the middle of Segment 13 for a small
space is different from the rest; this difference, however,
merely consists in the entire absence of glandular cells.
This modification of the epidermis, which is illustrated in
fig. 18, first appears in this stage; it is not directly in-
herited from the epidermis of the embryo, as yet undif-
ferentiated into gland-cells and packing-cells. In the earlier
stages which I have already described there is no means of
distinguishing by the characters of the epidermis the median
part of Segment 13, which is so marked in the present stage.
A complete series of sections through the 13th and neighbour-
ing segments showed a small sac (fig. 13) lying immediately
beneath the nerve-cord and in contact with the body-walls.
The walls of this sac are chiefly muscular; it is lined witha
layer of cells, of which the nuclei alone were darkly stained ;
the outlines of the cells themselves were rather hard to make
out, but the epithelium was evidently composed of very low

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 577

cells; these cells had the appearance rather of peritoneal cells
than of invaginated epidermic cells.

The sac, where it lay beneath the nerve-cord, had a flattened
appearance, as if it had been squeezed between the nerve-cord
and the body-wall; its cavity was chiefly developed in a hori-
zontal, not in a vertical, direction. It could be traced round
the nerve-cord for a short way on each side as a narrowish
tube running forwards and backwards in close contact to the
thinnish septum which divides the 13th from the 12th seg-
ment; coming into close relation with the base of the ovary,
it ended in the neighbourhood of this organ.

Traced in the other direction the sac penetrated the longi-
tudinal muscular coat for a short distance, and gave off a
narrow tube, ending blindly some way below the circular mus-
cular layer: the space between the ends of this tube and the
epidermis was crowded with nuclei, but there was no break
that I could find in the circular layer of muscles, and no in-
vagination of the epiblast. ‘The sac is thus entirely closed ;
it has no communication with the exterior, which might have
been expected.

The opposite extremity of the sac does not form a closed
sac, as indicated in the figure; it appears to do so in some
sections, such as the one illustrated in my drawing ; a complete
series of sections shows that this sac communicates freely
with the body-cavity. One wall is formed by the
septum which separates Segments 13/14, lettered spt;
the opposite wall is thinner, and consists of a mem-
brane, which connects that septum with the one
behind; the membrane in question is, however, not attached
to the intersegmental Septum 13/14 for the whole of its extent ;
it thus forms an aperture which leads into the subneural
pouch.

In specimens rather more immature than the one just
described, the relations of the subneural pouch will be under-
stood by a reference to figs. 4—6, which represent three sec-
tions of a continuous series; those selected for illustration
represent the critical points in the series ; the first one (fig. 4)

VOL. XXXII, PART IV.—NEW SER, QQ

578 FRANK E. BEDDARD.

shows the continuity between the subneural pouch and the
body-cavity ; the walls are seen to be formed from the interseg-
mental Septum 13/14, and form amembrane (S), which, traced
backwards, comes into continuity with intersegmental Septum
14/15. In the second section (fig. 5) the two membranes join
above the subneural pouch ; the third section (fig. 6) is through
the ventral median line, and the nerve-cord is, therefore, indi-
cated. The subneural pouch is seen to lie below the nerve-
cord, and in contact with the longitudinal muscular coat of the
integument ; the intersegmental Septum 13/14 does not reach
the nerve-cord, and to its free extremity is attached the mem-
brane uniting it with the following intersegmental septum.
In this specimen the epidermis lying immediately below the
subneural pouch is exactly like the epidermis elsewhere, i. e.
the glandular cells are not absent. It will, I hope, be clear
from the above description and from the figures that the sper-
mathecal sac is first of all formed by certain membranes in the
celom, which become approximated and attached in parts to
form asac. Later on the sac burrows its way towards the
exterior, where it appears to be met by a very limited invagi-
nation of the epidermis. This mode of formation has a remark-
able analogy to the formation of the oviducts in certain ganoid
and teleostean fishes. It is very probable that, as I have
already suggested, the second oviduct in Eudrilus is formed
in a similar way. It will be observed that in early stages the
ovary comes to lie within the spermathecal sac. If, for some
reason or other, the backward extension of the sac were
checked, it would form simply a duct for the ova.

There is also an interesting analogy to the formation of the
‘nephridia and genital ducts in Peripatus, which have been
recently so successfully investigated by Mr. Sedgwick.

In a species of Moniligaster—M. Houteni—Dr. Horst
has described the oviduct in a way which suggests that
it resembles the early stage of the spermathecal sac of Libyo-
drilus. His description (13, p. 99) is as follows :—“ As
stated before, the 12th and 13th septa are placed close
against each other ;: these two septa seem to form together on

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 579

each side a sort of funnel, the inferior part of which communi-
cates with one of the pores on the 14th ring. Although the
ovaries could not be found, I suppose that this funnel may
function as an oviduct.”

In Nemertodrilus the spermathecal sac is in a condition
of arrested development; it resembles to a certain extent the
early stages in the development of Libyodrilus, but secondary
modifications—in the complicated fringe which surrounds the
pores on Segment 13—appear to prevent their performing the
function of an oviduct. It would be very interesting to have
some information about the development of the large sacs in
Nemertodrilus, particularly as to whether they originally
communicate with the egg-sacs or not.

VI. Homologies of the Reproductive Organs in Eudrilide.

The homologies of the different parts of the reproductive
system in Eudrilide are by no means easy to decipher.

It is possible, however, to attack the problems with greater
confidence in view of the developmental facts that I have been
able to bring forward. Hitherto nothing but the adult struc-
ture was known, and that in certain cases evidently imperfectly.
It is clear that in many points the Eudrilide differ greatly from
all other earthworms; and I think that this paper rather
accentuates the differences than removes them.

On the other hand, I have been able to show that at a certain
stage of development the female reproductive system closely
resembles that of more normal genera; the special peculiarities
of the Eudrilide are a later formation; this argues for the
isolated position of the group, which I have elsewhere urged,
but not-for its primitive characters ; the reproductive system
must be looked upon as much modified from that of other
earthworms.

It seems fairly clear that the large unpaired sacs which exist
in the genera Heliodrilus, Hyperiodrilus, Stuhlmannia,
Polytoreutus, Paradrilus, Preussia, Nemertodrilus,
and Libyodrilus correspond in every case. But they are in

580 FRANK E. BEDDARD.

this event not spermathece, except in function. A true sper-
matheca, as I have shown, exists in Heliodrilus and Hy-
periodrilus, partly or entirely enclosed by the median sac.
In Paradrilus, too, judging from Michaelsen’s figure, there
is a spermatheca of this kind, but apparently not in some of
the other genera; nor is this structure represented in Libyo-
drilus. In Libyodrilus the large unpaired sac is
obviously a spermatheca in function, for I found
sperm in it, but its development shows that it cannot
possibly be compared to the spermatheca of, e.g.,
Lumbricus. It is a mesoblastic pouch which has
acquired an external opening; whereas the spermatheca
of Lumbricus are known, through the investigation of Bergh,
to be purely epidermic involutions. The homologue of the
spermatheca of Heliodrilus and Hyperiodrilus in Libyo-
drilus appears to be the orifice only of the mesoblastic sac.
In this connection the structure of Nemertodrilus as
described by Michaelsen is of great interest. He has described
(3), and I have been able (1) to entirely confirm his descerip-
tion, a pair of apertures in Segment 13 which lead into the
interior of that segment ; they have no connections with the
sacs extending backwards from Septum 18—14, but Michaelsen
has suggested that they represent the orifices of those sacs; I
am now of opinion that this suggestion is perfectly correct.
So far Nemertodrilus appears to be a degenerate form, and
other structural facts confirm that view. It is a little difficult
to understand the arrangement of the various organs in
Preussia, since Dr. Michaelsen was unable, through being
obliged to respect museum specimens, to make out the relation
of the ovaries.

The following is Michaelsen’s description of the parts in
question: I think it worth while to quote it in full, as the
journal in which it is contained may not be accessible every-
where.

“Der weibliche Geschlechtsapparat zeigt wieder eine neue
Modifikation der eigenartigen Verwachsung, wie sie fiir die
Teleudrilen charakteristisch ist. Die ventral median Offnung

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 581

im 15 Segment fiihrt in eine lange, gestreckt birnformige
Tasche, die sich bei dem untersuchten Exemplar bis in das 19
Segment nach hinten erstreckt. Nach vorne schien der Stiel
dieser Tasche in zwei grosse, ovale, dinnhautige Blasen
ueberzugehen, die fast bis an den Anfang des 13 Segments
reichen. In dem nach hinten gerichteten Pole dieser Blasen
miunden die Hileiter ein. Diese gehen von ihrer Ausmiindung
seitlich am 14 Segment, zuerst in grader, senkrechter Richtung
auf die Medianebene zu. Weit bevor sie dieselbe erreichen
biegen sie nach hinten um. Zugleich verdicken sie sich
bedeutend. Das Dissepiment 14/15 durchbrechend, verlaufen
sie nach hinten bis vor das Dissepiment 15/16, biegen sie dann
wieder nach vorne um und miinden schliesslich in die erwahnten
Blasenemeni. 22.) . jeder Hileiter tragt an der ersten Knick-
ung, an der Ubergangsstelle von dem engen distalen Teil zu der
erweiterten, nach hinten gerichteten Schleife, ein verhiltniss-
missig lang gestieltes Receptaculum Ovorum.”

It appears to me that the “ thin-walled vessels’”’ and the tube
leading to the receptaculum correspond to the first of the
diverticula of the median sac in Libyodrilus, and that the
distal extremity of the “ pear-shaped pouch ” is the true sper-
matheca. It is rather harder to understand Eudrilus and
Teleudrilus. The former genus has been redescribed by
Horst (7), his description of figures being confirmatory of my
own. There seems, therefore, to be little doubt, now that so
careful a worker as Dr. Horst has looked into the matter, that
the adult structure of the female genitalia of Eudrilus is
known.

I must, in the first place, admit that the structure of other
Eudrilids is not confirmatory of my view that in Eudrilus
there are two pairs of ovaries and two pairs of oviducts. I do
not, however, admit that that view is yet entirely disproved.
In order to show that the tube, which I have regarded as the
oviduct of Segment 13, is not an oviduct, it must be shown
that the organ regarded by Perrier, Rosa, Horst, and myself as
a spermatheca is a coelomic sac comparable to that of Hyperio-
drilus, &c. Iam inclined, however, to believe that this will

582 FRANK E. BEDDARD.

be proved ; but then it will have to be farther proved that the
duct of that sac up to a point beyond the “‘ oviducal” orifice
is also derived from the coelomic sac. Now, Horst says (7, p.
232) that the muscular layer of the spermatheca becomes very
thick near to the point of opening of the ovarian duct (= my
“ oviduct of Segment 13”). This looks as if the true-sperma-
thecal invagination began, or rather ended, at this point. If
so, it would be reasonable to speak of the tube in question as
an oviduct, whereas if it were merely a portion of the ccelomic
sac it could hardly be termed an oviduct, but rather an exten-
sion of the said sac forwards.

It is clear that two pairs of ovaries and oviducts formerly
existed in earthworms ; not only do they occasionally occur as
a sport (see my remarks upon varieties of Perionyx), but they
occur normally in Phreoryctes. Considerable traces of the
missing parts are met with in embryos of Lumbricus and
Acanthodrilus. Dr. Horst has overlooked these facts, some
of which were known when he wrote (7, p. 239), “ Chez
tous les genres d’Oligochétes, on n’a trouvé jusqwici qu’une
paire d’ovaires.” To return to the immediate subject of
discussion, it seems probable that the large sacs termed sper-
mathece in Eudrilus and Teleudrilus correspond to the
ceelomic pouches of Libyodrilus and the other forms. The
question is how much of them corresponds ; it may probably be
safely assumed that up to the point of opening of the ovi-
ducts there has been an epidermal invagination ; but beyond
that inference I do not consider it safe to go at present.

VII. Definition of Genus and Species.
Genus Libyodrilus, F. E. B.!

Nephridia paired, but connected with a network of tubes
ramifying in the integument; those of some of genital seg-
ments disappear in mature worms, but network remains. A
large unpaired sac opening on Segment 13 extends through
‘ T have briefly characterised the species in ‘ Proc. Zool. Soc.,’ 1891, p. 172.

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 583

five segments and lodges receptacula ovorum; oviducts pass
from these to apertures on 15th segment. Atria two, with
thick muscular walls opening by a common orifice on middle
line between Segments 17, 18. Each is furnished with a
single penial seta. Vasa deferentia without dilatations or
muscular coat, open near to summit of atria. Césophagus
without calciferous glands or ventral pouches; three gizzards
present at end of cesophagus in Segments 23, 24, and 25.
Integument without sense bodies.

Libyodrilus violaceus, F. E. B.

Sete strictly paired and ventral in position. Penial sete
short with a rounded free extremity. Clitellum occupying
three segments (14—16). Dorsal vessel single. Colour dark
violet with a tinge of pink.

Habitat—Lagos, West Africa.

The above definitions must in the present state of our
knowledge be regarded as only tentative.

VIII. Summary.

The principal new facts in this paper may be briefly sum-
marised as follows :

(1) The nephridial system consists of paired nephridia
which do not open immediately on to the exterior, but are
connected with an extensively ramifying system of tubes
embedded in the circular and longitudinal muscular coats ;
these tubes consist of four principal longitudinal trunks con-
tinuous from segment to segment, and of a single large cir-
cular vessel in each segment passing right round the worm at
the junction of the circular and longitudinal muscles; these
are connected by a plexus of vessels, and numerous tubules,
leading to the exterior, are given off from the circular trunk.
In some of the genital segments the paired nephridia have
almost disappeared, leaving only the integumental network.
Nothing of the kind has been yet described in any Oligochete.
In the young worm, just escaped from the cocoon, there is no

584. FRANK E. BEDDARD.

integumental network, which must, therefore, be regarded as
secondary, but the anterior nephridia at any rate are con-
nected on each side by a continuous longitudinal duct lying
within the celom.

(2) In the young worm the reproductive organs agree with
these organs in other earthworms; in the adult, a large
unpaired sac lying over the gut is developed ; this sac encloses
the receptacula ovorum, and opens by a median pore on Seg-
ment 13. It is developed from mesoblastic tissues, and is not
therefore the morphological equivalent of the spermathece in
Lumbricus, &c., but it performs the same function ; the sac
is formed internally and then grows out towards the epi-
dermis ; it is at first in open communication with the celom ;
its front wall is formed out of the intersegmental septum
between Segments 12, 13; the ovaries are enclosed by it, but
disappear early, before the sac is completed; otherwise the
ova would be probably unable to enter the egg-sac which
becomes nearly completely shut off from the sac; the two are
in communication only by the oviducal funnel, which has
become divided by the growth of the spermathecal sac into
two separate tubes, one opening into the spermathecal sac, the
other into the closed egg-sac; they unite, of course, to form
the oviduct itself, which opens on to the 15th segment,
reckoning by the external furrow, but on to the border line
between Segments 14, 15, reckoning by the septa.

(3) The testes and the vas deferens funnels are quite typical
in their structure and position; so, too, are the (two) pairs of
sperm-sacs (in Segments 11,12). The sperm ducts are not,
as they are in other Eudrilidz, dilated to form sperm reser-
voirs ; they open into tubular atria, with thick muscular walls
and glandular lining, near to their blind extremities; the two
atria open by a common pore upon the border line between
Segments 17,18; each is furnished with a short penial seta
not ornamented.

(4) The alimentary tract has no calciferous glands or
ventral cesophageal pouches such as are found in other Eudri-
lidee ; at the end of the cesophagus are three gizzards, one to a

STRUCTURE OF EARTHWORM ALLIED TO NEMERTODRILUS. 585

segment ; the intestine which immediately follows has at first
three typhlosolar folds ; later on the two lateral and shorter
folds disappear. The ventral wall of the pharynx is con-
nected with the nephridial tubes of its segments ; they open
into the interior of the pharynx.

(5) The area surrounding the sete of each side of the body
is shut off from the general body-cavity, forming a paired
series of chambers ; in the oesophageal region is developed a
perihemal celomic space surrounding the subcesophageal
vessels.

EXPLANATION OF PLATES XXXVIII & XXXIX,

Illustrating Mr. Frank E. Beddard’s paper ‘“‘ On the Structure
of an Earthworm allied to Nemertodrilus, Mich., with
Observations on the Post-embryonic Development of
Certain Organs.”

ANATOMY OF LIBYODRILUS VIOLACEUS, Nov. Gen., N. Sp.

Fic. 1—Dissection to show the principal organs. ¢. Testes. Sp.s. Sperm-
sacs. od. Oviduct. ».d. Vasa deferentia. a. Atrium. SS. Spermathecal
sac. g. Gizzard.

Fic. 2.—Ventral surface of Segments 13—18. sp. Spermathecal pore.
9. Oviducal pores. ¢. Male pore.

Fic. 3.—Fragment of cubicle, showing nephridial pores (z.).

Fies. 4, 5, and 6.—Three figures drawn from a continuous series of sec-
tions, aud illustrating the development of the spermathecal sac. Sp¢. Septum
separating Segments 12,13. i. Hinder wall of spermathecal sac, the cavity
of which is lettered Sp. sac. ov. Ovary. WV. Nerve-cord.

Fic. 7.—Longitudinal section through a portion of one of longitudinal
nephridial ducts. «@. Branches given off from duct. 4. Junction with one
of paired nephridia. The thick gelatinous connective tissue in which the duct
is shown; z. Nuclei of this tissue.

Fic. 8.—Penial sete.

Fig. 9.—Spermathecal sac and neighbouring organs. 4. Sac. a. Diver-
ticula of the same. 7.0. Receptaculum ovorum. od. Oviduct. qs, (isopha-
gus. #. Nerve-cord,

Fic. 10.—Section through a portion of one of pairednephridia. a, 6. Ne-
phridial tubes. ~. Nucleus of same, ¢, c’, Peritoneal investment.

586 FRANK E. BEDDARD.

Fre. 11.—Three nephridia of posterior segments on one side of body, as
seen on a dissection. xz. Nerve-cord. Spt. Intersegmental septum. zph. Ne-
phridium. f Funnel. §. Sete of ventral pair.

Fic. 12.—Transverse section through receptaculum ovorum and spermathe-
cal sac. Sp. sac. Spermathecal sac. ov. Ova. od. Oviduct. el. Elastic
membrane surrounding sac.

Fic. 13.—Spermathiecal sac at a later stage of development than Figs. 4—6,
illustrated by a transverse section through body-wall. Zp. Epidermis. wm.
Circular muscles. m’, Longitudinal muscles, with lymph spaces (/.). IV. Ne-
phridial tube. Sp. sac., Sp. sac?. Two portions of spermathecal sac. Spé.
Intersegmental septum 12—13. ov. Ovary.

Fic. 14.—Transverse section through body-wall, to show the ramifications
of the excretory tubes. ep. Epidermis. m. Transverse muscular layer.
m'. Longitudinal muscular layer. Bl. Blood-vessels. dd. Longitudinal
nephridial vessels. d’. Smaller tubes, forming network among the muscles.
a. Junction with one of paired nephridia. c. Circular nephridial vessel. 7.
Tube running to external orifice.

Fic. 15.—Section through ventral wall of pharynx, to show opening of
nephridium. o. Orifice of nephridium, ev. Nerve in pharyngeal epithelium.
m. Muscles. mph. Nephridial tubes.

Fic. 16.—Diagrammatic transverse section of body, to show integumental
nephridial network. D.v. Dorsal blood-vessel. S. Z. Supra-intestinal blood-
vessel. J. J. Infra-intestinal vessels. v. Ventral blood-vessel. Nps. Nephri-
dium. 7, Longitudinal trunks (four in number) of integumental network.
ec. Circular trunk lying between circular and longitudinal muscles. a. Cir-
cular vessel lying beneath longitudinal muscular layer of body-wall.

Fie. 17.—Transverse section to show perihemal space surrounding infra-
cesophageal blood-vessels (BZ. vess.). @s. Gsophagus. corp. Perivisceral
corpuscles. mes. Walls of periheemal space. m. Muscular bands.

Fic. 18.—Longitudinal section through one of the three gizzards in a very
young specimen. giz. Epithelium of gizzard. Per. Peritoneum. Spt.
Septum.

Fie. 19.—Transverse section through body-wall, more diagrammatic and
less highly magnified than Fig. 14. §&. Sete sac. B/. Blood-vessel. MM.
Muscle, connecting sete of lateral and ventral couples. Cal. Coelomic space
cut off from general cavity of segment by membrane Sp. JV. Nephridial
tube. JZ. Longitudinal vessel of integumental nephridial network; the net-
work is also indicated in white.

Fie, 20.—Highly magnified transverse section to show circular nephridia]
vessel (z.). m. Nerve accompanying it. JZ. m. Longitudinal muscles.
¢. m. Transverse muscles,

Fic. 21.—Infra-cesophageal blood-vessels in Segments 9, 10,11. a. Branches
of the same. Sp. sac. Sperm-sacs.

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 587

Some Points in the Development of Scorpio
fulvipes.

By

Malcolm Laurie, B.8Sc., F.L.S.
With Plate XL.

In the introduction to my paper on the development of
Euscorpius italicus (‘ Quart. Journ. Micr. Sci. vol.
xxxi) I mentioned that a species of scorpion (Scorpio
fulvipes), which I was permitted to examine by the kind-
ness of Professor Lankester, showed a considerable difference
in its mode of development from that of Euscorpius. This
difference is, as there stated, fundamentally due to the absence
of yolk from the egg of Scorpio fulvipes, in which particular
it contrasts strongly with that of Euscorpius, in which the
proportion of yolk is enormous. Further examination of more
abundant material, which Professor Bourne of Madras collected
and preserved with great care and forwarded to Professor Lan-
kester, shows that the specialisation of the embryo in relation
to its mode of nutrition has reached a very high pitch.

A few notes with regard to this particular form of Scorpion
development will, it is hoped, be of some service towards an

understanding of the embryology of Scorpions, and Arthro-
poda generally.

Ovary and Ovarian Egg.

The species of Scorpion of which the female reproductive
organs are described by Duvernoy appears to agree in the pecu-

588 MALCOLM LAURIE.

liarities of those organs with 8. fulvipes. Duvernoy, how-
ever, deals only with the external appearance of ovaries in
which the development of the embryo had already advanced
some way.

In Scorpio fulvipes the ovary agrees with that of
Euscorpius in its anatomy, in so far as it consists of a network
of tubes bearing a number of eggs which cause the tubes
to bulge at intervals into the body space. Beyond this, how-
ever, there are many important differences. The ovarian tube
in place of the large oval sessile ova of Euscorpius bears a
number of long diverticula, each of which ends in a solid
coiled appendix (fig. 1). These diverticula were described by
Duvernoy! as the eggs, but the comparatively small egg occu-
pies only one tenth of the length of the diverticulum, and lies
at its distal end at the root of the appendix (fig. 1, ov.).

The formation of these diverticula can be easily understood
from fig. 2, which shows a section of a young egg such as ov’
in fig. 1. The egg, which is formed as in Euscorpius by the
growth of one of the cells of the inner layer of the two-layered
ovarian tube, is carried out at the end of an outgrowth of the
ovarian tube, and is at first completely surrounded by a mass
of cells of the inner layer. At a stage somewhat later than
that in fig. 2 an opening is formed through these cells, so that
the ovum is only separated from the cavity of the diverticulum
by the vitelline membrane which surrounds it. There is no
specialisation of any of the cells for the purpose of forming
yolk, and in fact the egg is, when ripe, entirely without food
yolk.

The whole diverticulum, as is seen in fig. 1, consists of four
distinct regions :—A long stalk (s¢), a thickened collar (c), a
somewhat conical portion in which the ovum lies (ov), and a
long appendix. The stalk is peculiar in that the continuation
of the inner layer of the ovarian tube is divided into two
distinct portions. The inner one (fig. 3, 7./.) next the lumen is
formed of a single layer of long cylindrical cells with their

1 Duvernoy, “ Fragments sur les organes de la Génération de divers ani-
maux,” ‘Mém. de |’Acad. Sci. de l'Institut,’ t. xxiii.

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 589

nuclei at various levels, giving at first sight the appearance of
a folding of the walls. The outer portion (fig. 3, 7./’.) is com-
posed of a closely packed mass of small cells of which only the
nuclei can be made out. Outside this outer portion is the con-
tinuation of the outer layer of the ovarian tube (0./.), which con-
sists of flattened cells.

In the collar (fig. 4) the two portions of the inner layer are
no longer distinguishable. The whole layer, which is enor-
mously thickened, consists of clear, highly refracting cells with
oval nuclei and well-marked outlines. As development pro-
ceeds the collar gradually moves down towards the ovarian
tube, and as it passes the cells of the diverticulum change
their form and come to resemble those of the collar. This
change is preparatory to the active excretion of nutritious
matter for the nourishment of the embryo in its earlier stages.
The greater thickness of the collar is simply the sign of the
activity of the cells within, which require more space in which
to undergo their change in form. When this change is
accomplished the diverticulum returns to its former dimen-
sions.

The conical portion above the collar (fig. 1, ov) consists of
the egg, which when ripe is less than *2 mm. in length (that
of Euscorpius measuring 1°5 mm.), and of the inner layer
cells surrounding it. This follicle is several cells thick, the
cells being the same in appearance as those already described
in the collar. Between the egg and the lumen of the diverti-
culum is a small passage, the cells surrounding which are long,
and have their nuclei at their outer ends. These three
proximal portions of the diverticulum together reach a length
of about 2°5 mm.

The coiled appendix, which is longer than all the rest of
the diverticulum, consists of a solid rod of cells (fig. 5, 2./.)
forming a continuation of the inner layer, and is surrounded
by the flattened cells which represent the outer layer of the
ovarian tube (0.l.). The appendix plays an important part in
the nutrition of the embryo, being eeainalty absorbed as
development proceeds,

590 MALCOLM LAURIE.

The embyro undergoes the whole of its development in the
diverticulum, and does not pass into the ovarian tube until it is
ready to be born.

The development extends over more than six months, my
earliest stages having been preserved towards the end of
October and the latest in May. How much longer it may take
I cannot say, but the oldest embryos which I have examined
are still some way from being fully developed.

Development of the Embryo.

In the earliest stage examined by me the ovum is com-
pletely segmented. The external appearance of the diver-
ticulum and appendix is shown in fig. 1, and has been
described above. A transverse section (fig. 6) shows the
ovum, which is about ‘12 mm. in diameter, to consist of an
irregular mass of cells with faintly marked outlines and large
oval nuclei. There appear to be spaces between the cells, but
this may very likely be due to shrinkage. The nuclei for the
most part stain very faintly with carmine, and show a slightly
granular structure. Here and there one of the nuclei stains
very darkly, and such nuclei appear to be undergoing divi-
sion. The nuclei are scattered about irregularly, and show no
tendency to the formation of layers. There is no trace of
yolk either in the cells or in the spaces between them.

In the next stage, between which and the one described
above there is a considerable gap, various changes have taken
place. The ovum has increased in size to about ‘(15 mm.
diameter, though there is considerable individual variation in
this respect, and shows the beginnings of several structures.
The cells are chiefly aggregated along one side—the ventral—
and towards the posterior end, leaving a space in the middle
which is full of finely granular substance. This space is well
seen in longitudinal section (fig. 8), and a section in this
direction also shows very clearly the stomodeum (figs. 7
and 8, st.). This structure is a tube, the walls of which are
at first one cell thick (fig. 7, st.), and the lumen of which
opens to the exterior of the embryo at the anterior end (i. e.

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 591

the end furthest from the ovarian tube), and at the posterior
end into the space in the middle of the embryo described
above. It is certainly extraordinary that the stomodeum
should be the first structure developed, but the reason will be
found in the peculiar mode of nutrition of the embryo in its
later stages.

The outermost layer of the embryo (fig. 7, am.) is
separated from the rest by a distinct space extending all
round. It represents the so-called amnion, which is so well
developed in Euscorpius, but in this form appears to be
confined to the earlier stages, as I have been unable to find
any trace of it in the later ones. A protective envelope of
this sort is not so necessary in this form, where the embryo
remains in situ till it has attained its full growth, as it is in
a form like Euscorpius, where the eggs pass into the ovarian
tube at an early stage, and its slight development and early
disappearance are probably due to this change of habit in the
embryo. Its presence seems to point to the condition in
Euscorpius being the more primitive, a conclusion which is
supported by many other facts in the development.

In the next stage, in which the embryo has increased
very considerably in length, the body-wall (fig. 9, ep) is
very thin, and composed of flattened cells on the dorsal
and lateral surfaces; while on the ventral surface, where
the mesoblast and nervous system will afterwards make their
appearance, it is formed of two or three layers of cells with
large round nuclei.

The gut is not yet formed, but its position is shown by a
large cylindrical mass of yolk, the central portion of which
has a curiously honeycombed appearance. ‘The rest of the
body space is filled with yolk spheres, the material for the
formation of which must have been derived from the cells of
the diverticulum.

The cavity of the diverticulum beyond the point to which
the embryo extends is full of a finely granular substance
secreted by the cells which line it. The granular structure
may be due to coagulation of a fluid by alcohol. Here and

592 MALCOLM LAURIE.

there, in the midst of this granular substance, may be seen the
nuclei of free cells. These, probably, play some part in the
preparation of the nutritious material for absorption by the
embryo, but whether they are derived from the embryo or
from the walls of the diverticulum, I have not been able to
ascertain, though I think the latter most probable.

At the posterior end of the body there is a considerable
mass of cells, the growth and multiplication of which provide
for the rapid increase in length of the embryo. The head
end (fig. 10) consists of an almost solid mass of cells, in the
middle of which is seen the laterally-compressed lumen of the
stomodeum, which has a highly refractive, probably chiti-
nous lining.

At the sides of the stomodzum the cells have become elon-
gated (m), and form masses of muscular fibres, extending from
the stomodeum to the lateral walls of the embryo. A pair
of solid outgrowths on the ventral surface represent the
chelicere (fig. 10, I).

No traces of the other appendages are present, and it is
noteworthy that this pair of appendages, which in Euscor-
pius appears later than the five succeeding pairs, should here
be the first to be formed.

In the next stage only a few points need be noted. Among
the most curious of these is the formation of aseries of dorsal
outgrowths of the body, one to each segment, which give it,
when viewed from the dorsal surface, the form shown in fig.
11, while in section it has the shape seen in fig. 12. The reason
for this curious change in the shape of the body is not clear, as
the spaces thus formed are unoccupied by any structures except
a few thin bands of mesoblast. A section (fig. 12) shows the
gut completely formed as a tube of large cells surrounding a
granular mass of food material. The rest of the body-cavity
is occupied only by thin trabeculze of mesoblast except on the
dorsal surface, where the mesoblast is present in considerable
amount and is hollowed out to form the heart. The yolk
spheres, which in an earlier stage occupied the body-cavity, have
been completely absorbed, and, except for a slight thickening

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 593

of the epiblast where the ganglia will be formed, there is no
trace of the nervous system in the body. In the cephalic
region the brain is beginning to form as in Euscorpius by
the proliferation of the cells of a pair of cerebro-optic in-
vaginations, which are more lateral in position than in the latter
species, but otherwise completely correspond to them.

At a considerably later stage the gut begins to be constricted
by both circular and longitudinal bands of mesoblast (fig. 13),
which divide it into a central tubular portion with a series of
large diverticula which form the so-called liver. Both the gut
and the liver remain full of granular matter, from which the
body-cavity is quite free. The ventral nervous system is
by this time completely separated from the epiblast, but as its
formation presents no special points of interest I have not de-
scribed it indetail. In the head the stomodeum becomes very
chitinous, and is furnished with powerful lateral muscles (fig.
14, m). Just opposite the aperture of the stomodzum is the
lower end of the cord of cells which forms the coiled appendix
described above (p. 589), and it is by the destruction of this cord
of cells that the embryo nourishes itself through all the later
stages of embryonic life. This mode of nutrition, which was
imperfectly described by Duvernoy,' is very exceptional, and,
indeed, more like the nutrition of a young marsupial than any-
thing else. The cord is held in position by the chelicere
(fig. 14, I), and this is the object of the phenomenally early de-
velopment of these appendages as well as of the stomodzum.
The comparatively early formation of the gut is due to the
same cause. As might be expected, the chelicere are con-
siderably modified to serve their peculiar function. This is
best seen in the oldest stage in my possession (figs. 15—17),
where the chelicere, apart from their great proportional size,
are seen to have the third joint specially developed (fig. 17, I, 3).
This third joint is very much larger than the other half of the
pincer, and is further provided with a strong band of chitin
which runs in a somewhat sinuous manner from the base of
the joint up to the tip. This band of chitin is grooved, and the

‘ Loe. cit.
VOL. XXXII, PART 1V.—NEW SER. RR

594 MALCOLM LAURIE.

two chelicere are twisted so that the chitinous plates come into
contact with each other, leaving a small hole (fig. 16, co)
through which the cord of cells passes to the mouth. I think
it is probable that the chelicerze do not merely hold the cord
in position, but serve also to crush it. The lower end of the
cord consists of horny-looking cell-walls from which all the
protoplasm seems to have disappeared. Whether the embryo
nourishes itself on the protoplasm of the cells or whether the
cell contents are some special substance I have not been able
to find out. It is perhaps worthy of remark that the chitinous
plates on the chelicerz are covered in places with a scale-like
marking very similar to that which is so characteristic of
the fossil Merostomata. The epistomial lobe (figs. 16 and
17, est) is very well developed, and also furnished with chitinous
plates.

The five succeeding pairs of appendages closely resemble
their adult state, and need no description here.

The genital opercula are in my latest stage not yet visible
in a surface view, contrasting in this respect with the pectines
(fig. 15, vzz).

The six metasomatic or caudal segments present one point of
interest in Sc. fulvipes, namely, that the tail is bent up over
the back, contrasting in this respect with Euscorpius, in which
the tail lies along the ventral surface.

From what I was able to make out as to the development
of the median eye, I think that the pigment-cells are epi-
blastic. The retina of the eye is formed, as in Euscorpius, from
a thickening of the dorsal wall of the cerebro-optic invagina-
tion. The thickened portion consists at first of a mass of
large spherical nuclei, which are distributed through the whole
thickness of the retina. In the latest stage which I examined,
in which there is already a considerable quantity of pigment,
the nuclei are confined to the inner half of the retina, leaving
the outer portion quite clear (fig. 18), The nuclei are, how-
ever, no longer all alike; but, while the majority of them retain
their spherical form, a certain number, forming a band in the
middle of the retina, have assumed an elongated form, These

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 595

latter are the nuclei of the retinal cells, and as from the dis-
tribution of the pigment special pigment-cells seem to be
present, the large spherical nuclei probably belong to these
latter. Absolute confirmation of this view by dissociation of
the elements of the retina has unfortunately proved impossible,
owing to their state of preservation.

Conclusions.

The development of this form adds another to the numerous
types of development in the Arachnida. It is, as is shown
by its mode of nutrition, a highly specialised form. There is
no doubt that the type of development represented by Eu-
scorpius is the more primitive of the two. The chief argu-
ments in favour of this view are the formation in Scorpio
fulvipes of (1) a rudimentary amnion, and (2) the formation
of yolk spheres in the earlier stages, and a mass of yolk round
which the gut is formed.

Further, so far as can be made out from the description by
Kowalewsky and Schulgin,! the development of Androctonus
follows much the same lines as that of Euscorpius. It is
curious that Euscorpius should resemble the very distantly
related Androctonus so closely, while differing so markedly
from a comparatively near relation like Scorpio; and further
study of the mode of development in other forms would, pro-
bably, throw an interesting light on the value of the present
system of classification.

The mode of nutrition explains many of the peculiar points
in the growth of the embryo, everything being sacrificed to
the rapid development of the organs—chelicerz, stomodzum,
and gut—necessary for nutrition, and the other appendages,
together with the mesoblast and nervous system, being formed
at leisure after nutrition is provided for,

I have, unfortunately, not been able to find the remarkable
sense organs described by Patten.?

! «Biol. Centribl.,’ vol. vi, p. 525.
? Quart. Journ, Micr, Sci.,’ vol, xxxi,

596 MALCOLM LAURIE,

EXPLANATION OF PLATE XL,

Illustrating Mr. Malcolm Laurie’s paper on “ Some Points in
the Development of Scorpio fulvipes.”

Abbreviations.

am. Amnion. ap. Appendix of diverticulum. 4. §. Air-space. Bl. 8.
Blood-sinus. c. Collar of diverticulum. co. Cord of cells on which embryo
feeds. ep. Epiblast. est. Epistome. 4&¢. Heart. Ay. Hypoblast. 7.7. Inner
layer of ovarian tube. m. Muscles of stomodeum. mes. Mesoblast. 2. ¢.
Nerve-cord. o. Central eyes. o'. Lateral eyes. o. 7. Outer layer of ovarian
tube. o.d. Ovarian tube. 0.x. Optic nerve. ov. Ovum. ~7. Nuclei of
retinal cells. s. Stalk of diverticulum. s¢. Stomodeum. sfg. Stigma. v.
Vitreous layer. y&. Yolk. The appendages are numbered 1, 1, U1, &c.
For the sake of simplicity the diverticulum round the embryo has not been
figured, except in Fig. 14.

Fic. 1.—Part of ovary of Scorpio fulvipes, to show the diverticula in
which the eggs are formed. x 4.

Fic. 2.—Longitudinal section of a diverticulum and ovum at an early
stage. X 82.

Fic. 3.—Transverse section of the lower part of a diverticulum. x +35.

¥ic. 4.—Transverse section of the swollen “collar” of a diverticulum.
x 48.

Fic. 5.—Transverse section of the solid coiled appendix in which the
diverticulum ends. xX 4%.

Fic. 6.—Transverse section of a fully segmented egg. x 259.

Fic. 7.—Transverse section of front end of a very young embryo, showing
stomodeeum, amnion, &. x 259.

Fic. 8.—Longitudinal section of an embryo slightly older than Fig. 7.
x 148,

Fic. 9.—Transverse section. Body of embryo considerably older than
Fig. 8. x +858,

Fic. 10.—Transverse section through head of same embryo, showing
stomodeeum, chelicere, &e. Xx +5,

Fic. 11.—View of the dorsal surface of an embryo, older than Fig. 9,
x 4,

SOME POINTS IN DEVELOPMENT OF SCORPIO FULVIPES. 597

Fic. 12.—Transverse section of body of same embryo. x 8°.

Fic. 13.—Transverse section of body of an older embryo. x 82.

Fic. 14.—Transverse section through head of same embryo, showing mode
of nutrition. x 82.

Fic. 15.—View of ventral surface of advanced embryo. The iii to vi
appendages have been removed on the left side. x &.

Fre. 16.—View of mouth parts of same embryo, from the front. x 8,

Fie. 17.—View of mouth and surrounding appendages of same embryo.
The left chelicera has been unfolded to show its structure, and the left chela
has been removed. x 143.

Fic, 18.—Section through retina of central eye. x 292,

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MAUPAS S RESEARCHES ON CILIATH INFUSORIANS. 599

Abstract of Maupas’s Researches on Multipli-
cation and Fertilisation in Ciliate Infusorians.

By

Marcus M. Hartog, D.Sc., M.A., F.L.S8.,
Professor of Natural History at Queen’s College, Cork.

THE interest so widely felt in all questions of heredity and
reproduction removes all need for excuse in presenting an
abstract of two of the most important contributions to this
question.! I drew this up originally for my own guidance,”
and have now enlarged it with a few modifications in what I
may term “notation,” to make the nuclear relations more
easy to master.

The methods used are given at length. Cultures are made
from single progenitors in a drop of water placed on a slide
and covered, with bristles, &c., placed under the thin glass to
prevent pressure, the total amount of water being about
10 cgms. These slides are kept in moist chambers, and the
slight loss of water from evaporation is made up as needed with
distilled water. The Infusoria under these circumstances live
and thrive, assembling in a circle just within the cover, and
not moving about much when well supplied with food, so that
counting them is no difficult task. The Ciliata may be divided
into herbivorous and carnivorous. The former are fed by

1 «Sur la multiplication des Infusoires Ciliés,” in ‘ Archives de Zoologie
Expérimentale,’ sér. 2, vol. vi, pp. 165—2738, t. ix—xii; “‘ Le Rajeunisse-
ment Karyogamique chez les Ciliés,” op. cit., vol. vil, pp. 149—517,
t, ix-—Xxxill.

2 Tn the autumn of 1889; the abstract in its present form was completed
in November, 1890.

600 MARCUS M. HARTOG.

cautiously supplying very dilute boiled paste to the colony.
The latter require to be supplied with smaller Infusoria, for
which purpose Cryptochilus nigrescens is raised in hay-
decoction; the drop of the liquid with this form is always
examined under the microscope to ascertain its freedom from
other forms, and only after this control drawn off with the
pipette to be given as food. In the research on fission observa-
tions were made daily, and entered with the temperature in
the journal. Twenty species were examined, and their rate
of bipartition carefully studied. For any given species this
rate varies with (1) the temperature, (2) with the quantity,
and (3) the quality of the food; while hght or darkness is
absolutely without influence.

It is found with each species that for temperature there is a
minimum below which no bipartition takes place, an optimum
at which it proceeds most rapidly, and a maximum beyond
which, again, there is no occurrence of reproduction. Ex-
haustion of the food-supply determines encystment if the
culture is of close relations; conjugation when the indi-
viduals are of mixed origin, and their age is suitable.
This is a discovery on which the second research was largely
based, as Maupas could now induce conjugation at will.

The greatest frequency of bipartition at temperatures of
15°—18° was observed in Glaucoma scintillans, namely,
five times in the twenty-four hours, and the least in
Spirostomum teres, viz. once in forty-eight hours, or one
tenth the rate of the former species.

Maupas started the cultures with an individual that had just
separated from its partner in conjugation, and which he terms
an “exconjugate.” In this way it has been possible to ascer-
tain whether the rate of bipartition increases or diminishes
as we recede from the ancestor; and he finds that neither
change takes place at first, but that the rate is fairly constant
for all the individuals of a single cycle for a long period, after
which it diminishes: there are, however, fairly marked dif-
ferences between the descendants of different ancestors of
the same species.

MAUPAS’S RESEARCHES ON CILIATE INFUSORIANS. 601

Considering Weismann’s assertion, that Protozoa are im-
mortal, it is of extreme interest to note that the later individuals
of a cycle differ from the preceding ones in a way that fully
justifies Maupas’s use of the term “senescence.” The later
individuals are reduced in size—sometimes to one fourth of
their normal length; the buccal wreath is more and more
reduced as the stock ages; the nuclear apparatus undergoes
degeneration in various ways, rendering fertile conjugation
impossible. In this stage, however, it is remarkable to find
that sometimes a sexual hyperesthesia is observed, inducing
conjugation between close relations, though this is of course
sterile, and accelerates the death and disappearance of the
stock. This ultimate death by senescence took place in
Stylonychia pustulata in 316 generations from the excon-
jugate progenitor; while in a stock of Leucophrys patula,
of which the progenitor was possibly not an exconjugate, it
occurred only after 660 generations.

In a section of the second paper these results are completed
by the proof that for many species, at least, the cycle shows
from its origin in conjugation to senescence first of all a stage
of “immaturity,” in which conjugation cannot be induced ;
then, after a certain number of bipartitions, “ puberty” is
reached, and during the period of “eugamy” that ensues fer-
tile conjugation can be induced at any time by suitable treat-
ment : this second stage merges into the last—of “ senescence.”
Puberty is reached in Stylonychia pustulata at the 130th
bipartition, in Onychodromus grandis at the 140th, and
in Leucophrys patula only at the 300th; and senescence
commences in the first about the 170th (though some senile
individuals showed as early as the 100th), in the second at the
230th, and in the third at the 450th. Since bipartition pro-
ceeds in some cases in senile individuals that have lost
their micronucleus, it is obvious that this organ plays no
necessary part in the vegetative reproduction of the Ciliata.

602 MAROUS M. HARTOG.

In the second paper we come to the study of conjugation,
and the first full account of it yet given, illustrated by ex-
quisite figures as well as by numerous diagrams, some of which
we shall reproduce in a modified form. The materials were
obtained by the methods described above, the starvation of
mixed colonies of a species at a suitable stage always bringing
about conjugation, In these experiments it was found that
the method of starvation and mixing colonies of different
stocks failed if tried too soon after conjugation, and this was
the starting-point of the discovery of the puberty of the cycle
mentioned above.

In some cases (e. g. Leucophrys patula) the first result
of starvation is to induce the production of dwarf forms by
repeated fission, and it is only these dwarf forms that con-
jugate; if, however, a food-supply be introduced at a suffi-
ciently early stage, the gametes quit company, and resume the
ordinary process of growth and bipartition. The formation of
similar dwarf forms occurs only in certain members of the
stock of Vorticellines, and eventuates in the production of the
microgametes or “males,” which swim to and conjugate
with the undivided attached megagametes.

By careful observation the time for each stage of the process
was determined ; and by killing and fixing couples at successive
intervals, preparations of every stage were easily obtained for
research. The fixative was a 1 per cent. solution of corrosive
sublimate, and the animals were stained on the slide with
picro-carmine or a solution of methyl green in 2 per cent.
acetic acid, and then cleared in situ with glycerine or balsam,
The best homogeneous immersion objectives are absolutely
needed for a complete study of these processes.

I may recall the fact that the nuclear apparatus of Ciliata
is double, comprising the large ‘‘meganucleus’’ (macronucleus,
endoplast, nucleus of authors), and the ‘ micronucleus ” (endo-
plastule, paranucleus, or nucleolus), which latter is multiple
in some species. In bipartition the meganucleus divides by
mere constriction, while the micronucleus shows all the stages
of karyokinesis save the disappearance of its membrane. In

MAUPAS’S RESEARCHES ON OILIATE INFUSORIANS. 603

conjugation a pair of Ciliata approach and adhere, fusing more
or less completely by their oral surfaces, and then separate
after a time: it has been long known that in this process
the nuclear apparatus undergoes a complete reconstruction.
What the nature of this reconstruction is, Maupas has now
revealed.

The process of conjugation is divided by the author into
the eight following stages. In A the micronucleus enlarges
greatly. In B, C, D it undergoes successive bipartitions. Two
sister nuclei of the third bipartition (stage D) are specialised
as “pronucléei:” the one of these migrates into the other
gamete to fuse with the stationary pronucleus there; and this
process constitutes stage E. The “ copulation-nucleus” so
formed undergoés two fresh bipartitions in stages F and G.
In stage H the nuclear apparatus is reconstituted and the first
bipartition takes place.

In the five stages B, C, D, F, G, involving nuclear division,
we can distinguish the following substages or phases: (1)
Spirema, the nucleus shows a reticulation; (2) aster, the
chromatin forms distinct thickish rods, extending the greater
part of the fusiform nucleus, its poles being occupied by fine
achromatic filaments; (3) equatorial plate; (4) diaster, the
chromatin rods are in two groups united by achromatin fila-
ments; (5) dispirema, the two chromatin groups are more
distant, and contorted, and united by achromatin filaments
which are strangulated in the middle, or contained in a dilated
connective tube (boyau), which is soon absorbed into the
ambient cytoplasm. Thus G, would indicate the aster stage
of the second bipartition of the copulation-nucleus,

In stage A we may also distinguish four substages: (1) the
micronucleus unchanged ; (2) swelling without much change
of form; (3) further swelling with changes of form charac-
teristic of the species; (4) the micronucleus condenses and
shrinks by way of preparation for its first bipartition in
stage B,

Stage E is also divided into phases; E,, the male pronucleus
approaches the point of exchange; E,, it passes into the fellow-

604 MARCUS M. HARTOG.

gamete; E,, it reaches and touches the female pronucleus ;
E,, the male and female pronuclei fuse completely to form the
copulation-nucleus.

Stage H is divided into the following phases: H,, the off-
spring of the copulation-nucleus are all similar; H,, they are
differentiated into enlarging meganuclei and small micro-
nuclei; H,, the meganuclei still enlarge, but refuse nearly or
altogether to take up stains; H,, completion of the growth of
the meganuclei, which now stain well; the exconjugate is now
ready to undergo bipartition.

The separation of the exconjugates usually.takes place in
stages F' and H. I have tabulated the cases given by Maupas,
and find that in one species they separate in E, four in F, three
in G, eight in H ; while in one species, Spirostomum teres,
they may separate in any stage from F, to Hy.

For the sake of additional clearness, I have used the follow-
ing notation, u or vy = the micronucleus of a gamete; Z, the
copulation-nucleus formed by the fusion of the two pronuclei.
M, the mega-, and m, the micro-nuclei of the exconjugate. To
denote the daughter nuclei after any given number of mitoses
I use this same number as an index to the letter of the original
nucleus ; thus Z! (abbreviated for Z~+ 2‘) represents one of the
two nuclei formed by the first mitosis of the copulation-
nucleus; > is one of the nuclei resulting from the third
mitosis of the original micronucleus of a gamete.

We may now describe fully the process in one of the sim-
plest forms, Colpidium Colpoda, which has a single micro-
nucleus. This enlarges and divides twice in stages B and C; of
the four nuclei so formed, three abort and are finally absorbed
by the cytoplasm, as ‘“‘corpuscules de rebut ;”’ the fourth under-
goes a new bipartition (stage D) to form the two pronuclei
of the formula p?. The point that determines which of the
four nuclei, 4”, should be preserved for bipartition seems to be
the accident of position; it is always the one at the
frontal end of the animal.

So, too, position determines the respective fates of the
two pronuclei; that which is next the point of union of the

MAUPAS’S RESEARCHES ON CILIATE INFUSORIANS. 605

two gametes is the male or migratory pronucleus, the other
the female or stationary pronucleus. In stage E each
male pronucleus crosses over the other at the point of contact
of the gametes, and fuses with the female pronucleus of the
other gamete: the copulation in this species takes place in
the resting stage of the pronuclei. Hach conjugate has now
a “ copulation-nucleus,” which we letter Z.

3 3 K
pes em
ary Bea oi 2
: H
Epa Gy heey C
eee aed aocstcnttnanntrnntnenntnnnannninnnmennennnnnnennnntn
Cieraes Mone ae B
B A

Fic. 1.—Schema of nuclear processes in a conjugating Ciliate, from the first

division of the micronucleus (#) to the formation of the conjugating
pronuclei (u*s, p’m). The arrow indicates the path of the migratory
pronucleus ; a bar is placed above the rejection nuclei. The dotted lines
separate the successive stages, marked by capital letters at the right
hand.

The first mitosis, stage F, takes place in the front of the
body, but the nuclei Z! pass to the hinder half to divide again
in stage G. By the elongation of the connective tubes in the
dispirema stage, G;, two of the nuclei Z* pass into the
anterior end and become meganuclei; and their respective
sisters pass to the hinder end and become the micronuclei of
the offspring of the first bipartition.

Separation takes places in the stage H,; and young excon-
jugates seem to have neither mouth, gullet, nor frontal pro-
minence: from three to eight days elapsing before the mouth
forms afresh and feeding recommences; and from eighteen to
twenty-four hours more until the first bipartition. Before
this takes place the micronuclei move up and place themselves
beside the meganuclei; one set goes to the fore, the other to
the hinder half of the body; while a second contractile vacuole
appears in the front half. Transverse fission then takes place.

606 MARCUS M. HARTOG.

“It is obvious,” says Maupas, “ that this division, unprovoked
by any nuclear division, must be due to the forces of the pro-
toplasm, the nuclear elements only playing a passive part.”

tt :
M:M m (lst fission).
errerriiiiririis tr Poem ascanseseneeees . . H
Tt ia p shee :
PR felt SE Gk See ee AOR
Gesdowst A i ctty F
SE Pett SS Ee rc

Fic. 2.—Reconstitution of nuclear apparatus from conjugation-nucleus in an
exconjugate Colpidium Colpoda, and reversion by a single fission to the
normal type with single mega- and micro-nucleus (M,m). The dotted
vertical line indicates the plane of fission.

We may express this by equations thus :
Z=477 (Stages F, G).

=2M+2m (Stage H).
= {M+m} + {M+m} (Ist fission).

The original meganucleus is passive throughout, and retains
its granular structure till H,. At H, it becomes homogeneous,
and still stains well; while in H, it passes to the hinder end
of the body, becomes smaller and irregular, loses its staining
power, and finally disappears completely. The above dia-
grams (Figs. 1, 2) are modified from Maupas.

We may sum up matters thus :—In each gamete the micro-
nucleus undergoes three bipartitions ; two sister nuclei of the
third bipartition are differentiated into stationary or female
pronucleus, and migratory or male pronucleus respectively.
By fusion of a male and female pronucleus a copulation-nucleus
is produced; by two successive mitoses are formed four
nuclei of the formula Z’; the sister nuclei of either pair are
differentiated in their development to form the one a mega-,

MAUPAS’S RESEARCHES ON CILIATE INFUSORIANS. 607

the other a micro-nucleus; and on fission each set so formed
constitutes the nuclear apparatus of the first offspring of the
exconjugate. During these processes the original meganuclei
of the gametes have undergone disorganisation and ab-
sorption.

We may express the above still more briefly thus:—Two
Ciliata of the same species approach by their oral surfaces ;
the original micronuclei by repeated bipartition form a pair of
pronuclei; one pronucleus in either gamete migrates into the
other to fuse with the stationary pronucleus therein; then
a new nuclear apparatus is constituted by the bipartitions of
the copulation-nucleus ; the gametes now separate to found
each a new life-cycle of the species,

Several important variations occur on the schema given
above; thus, in Coleps hirtus, &c., with a single micro-

nucleus, stage G is doubled (G G), so that we have eight

nuclei of the form Z, instead of four of the form Z?; two of
these become micronuclei, as in Colpidium; and it seems
probable that the other six are transformed into meganuclei,
which, by interfusion, have their number reduced to 2.

As a more complicated variation I may cite Paramecium
caudatum, where the stages are identical with Colpidium
as far as E, and stage G is doubled as in Coleps.

We may represent the process of reconstruction in (a single
exconjugate) Paramecium caudatum by the following
equations, using the symbol for multiplication by zero (x 0)
to designate the elimination of certain of the nuclei descended
from the conjugation-nucleus :

@) 782" (Stages F, G, G).
(6) 822 = 4M+m+3Zx0 (Stage H).
(c) 4M+m = 4M+2m!' = {2M+m'} + {2M+m'} (lst fission),
(dZ) 2M+m' = 2M+2m’?= {M+m*} + {M+m*} (2nd fission),

Here one of each of the four pairs of nuclei Z> becomes a
meganucleus, their sister (micro-) nuclei all aborting save one.
This divides during the first bipartition, of which the offspring

608 MARCUS M. HARTOG.

have hence two meganuclei; and it is only at the second
bipartition that the nuclear apparatus is reduced to one mega-
and one micro-nucleus as characteristic of the species. The
primitive meganucleus of the gametes undergoes fragmen-
tation into as many as sixty parts, which persist in the ex-
conjugates and their offspring for some time; they usually
undergo absorption, or are expelled as feces; but if the
culture is starved, some may fuse with the new mega-
nucleus.

Paramecium aurelia, with which P. caudatum is
usually confounded, has two micronuclei, so that at stage C
eight nuclei of the formula ,? are present ; only one of these
undergoes the next bipartition to form the pronuclei, all the
others being “‘ corpuscules de rebut.” Stages FandG are as in
Colpidium ; but at the first bipartition the micronuclei divide,
one daughter nucleus of each going to either of the offspring.
The following equations represent Stage H and the first
fissions of Paramecium aurelia:

47? = 2m+2M (Stage H).
2m+2M = 4m'+2M.

= {2m'+M} + 2m'+ M} (1st fission).

Still more complicated is the process in Vorticellines where
the gametes are of unequal size, the smaller ones being formed
by vertical fissions of an individual into four or eight, and
swimming freely. One of these microgametes conjugates
with a megagamete and fuses completely with it, so as to
form a single zygote—a process first fully described by Stein.
Either gamete here only forms a single functional pronucleus,
their sister pronuclei aborting. As the two pronuclei advance
to meet they might, perhaps, be regarded both as equivalent
to the male pronuclei of other species; but it is impossible
to regard the pronuclei as really differentiated into distinct
sexes. Stage G is double, so that eight nuclei are formed,
Of these, seven evolve into meganuclei; one into the micro-
nucleus of the zygote. Successive fissions of the zygote take

MAUPAS’S RESEARCHES ON CILIATE INFUSORIANS. 609

place, in which the meganuclei are distributed between the
offspring, while the micronucleus undergoes bipartition: the
normal state of the species is reached in one offspring of the
second, and in six of the third fission.

The processes of nuclear reconstitution in Vorticellines, with
the fissions necessary to bring back the type form, possessing
a single mega- and micro-nucleus, are represented in the

following equations. In each case a vinculum is put
above the normal form.
825 = 7M+m (Stage H).

7M+m =7M+2m'= {8M+m'}+ {4M+m'} (Ist fission).
3M+m' = 3M+2m?= {M+m%}+{2M+m} (2nd fission).
4M+m! = 4M+2m?= {2M+nm?}+{2M+m?} _,, ys

2M+m?= 2M+2m* = {M+m3}+{M+m5} (8rd fission).

The relations are further complicated by the reduplication
of the micronucleus of the microgamete in the beginning of
stage A; both these micronuclei undergo the bipartitions of
stages B and C; but only one of the eight nuclei (2v?) so
formed divides in stage D, which is thus identical with that
of the megagamete. A similar reduplication of the micronu-
cleus occurs in both the isogamous gametes of Euplotes.

The fusion of the two cytoplasms takes place at the end of
stage E. The old meganucleus undergoes fragmentation, and
its pieces may be all absorbed, or some may finally fuse with
the new meganuclei.

I have cited a number of distinct cases from Maupas to
show that, with wide differences of detail, the phenomena are
everywhere essentially the same in the Holotricha, Hetero-
tricha, Hypotricha, and Peritricha: his observations on the
Suctoria or Acinetina show that the law may be extended to
this group—a sure proof, if one were needed, of its true
affinities.

A series of elaborate discussions follows the systematic review
of the phenomena of conjugation in the Ciliata. The double
nature of the nuclear apparatus in this group is explained by a

VOL, XXXII, PART 1V.—NEW SER. 88

610 MARCUS M. HARTOG.

physiological division of labour. The mEGANUCLEUs presides
over the nutrition and growth of the individual, and holds the
power of repairing injuries ; it divides by simple constriction.
The MIcRONUCLEUS presides over the preservation of the species,
and is the seat of that power of rejuvenescence which allows
continued reproduction, and the substratum of inherited
qualities ; it always divides by mitosis.

In the mitosis of the micronucleus the connecting tube of
the two daughter nuclei always seems to be lost by absorption
into the cytoplasm, and as this contains the greater part of the
nucleo-hyaloplasm, we may regard it as an additional proof
that the chromatin is the essential element of the nucleus, in
which reside its special functions, those of transmitting in-
herited properties ; while the hyaloplasm has a purely mechani-
cal function. The greater or lesser compensation for loss of
substance in mitosis is effected by osmosis only, as the nuclear
wall always persists.

It is shown that fertilisation is essentially a nuclear pheno-
menon: the union of the chromatic elements of two nuclei of
different origin so as to constitute a new “ nucleus of rejuvenes-
cence.” As to its physiological purport, it is very obvious that
the common view is wrong, and that fertilisation is not neces-
sarily linked with multiplication. In Ciliata fertilisation
interrupts the ordinary rapid multiplication of the race for
hours and days together. Thus Onychodromus grandis
is six days in conjugation; in which time by fission each
gamete could have produced in the thirteen bipartitions some
8000 offspring ; while in Vorticellines the one of the gametes
is practically suppressed and its power of fission annihilated.

Rolph’s theory of isophagy is only mentioned to be waived
as not corresponding with the facts.

Let us now examine Weismann’s view: that sexual or amphi-
gonic reproduction has the chief and sole function of mixing
inherited tendencies, and of creating individual differences,
through which selection forms new species. This view excludes
fecundation from all share in the sustenance and preservation
of the original species, on the ground that living matter pos-

MAUPAS’S RESEARCHES ON OILIATE INFUSORIANS. 611

sessed as a primordial attribute an unlimited power of growth
and reproduction, as inherent as energy is in matter, and
quite independent of the actual mode of reproduction, sexual
or asexual.

This latter part of Weismann’s theory is completely over-
turned, so far as the Ciliata are concerned, by Maupas’s proof
of the existence of senescence and death as the normal ter-
mination of every cycle, when the possibility of normal conju-
gation is excluded. Thus the hypothesis of the essential
immortality of the Protozoa has lost its foundation in fact.!

Maupas’s own conclusion as to the real explanation is, that
fertilisation determines rejuvenescence, a view enunciated
by Biitschli and Engelmann, and adopted by Hensen and E,.
van Beneden.

Fertilisation is, in its essence, independent of sexual differ-
entiation. This is clearly shown by the above studies on the
Ciliata, where both gametes and pronuclei are clearly identical.
Henceforward we can only look on sex as a secondary adapta-
tion to facilitate the act of fecundation.

Besides rejuvenescence, fecundation does determine the
transmission of inherited qualities; and we must admit this
part of Weismann’s view: that it plays an important part
in variability.

What is the essence of this act of fecundation considered
as a general phenomenon of organic life? Maupas answers
this question in the following theses :

1. Morphologically it is essentially a nuclear phenomenon.

2. The nuclei retain their existence throughout; Haeckel
was mistaken in assuming their disappearance in a ‘‘ moneran
stage.”

8. A preliminary reduction takes place in the nuclear sub-
stance, which by successive normal mitoses is reduced by
three fourths; the parts so eliminated disappear by absorption
as “ noyaux de rebut,” or else are expelled as polar bodies,

1 My own standpoint is somewhat different, for I admit the existence of
primitive forms, like the Monadina of Cienkowski, that are completely
agamous and “ immortal.”

612 MARCUS M. HARTOG.

The reduction so effected is purely quantitative, and appears
to involve male as well as female cells. (The grounds on
which this thesis is founded are referred to below.)

4. “ Noyaux de rebut” and fertilisation-nuclei are absolutely
equivalent; their respective fates are determined by the ac-
cident of position.

5. By the last mitosis both “ noyaux de rebut” and fertilisa-
tion nuclei cease to be true nuclei, capable of indefinite
normal evolution, and are simple pronuclei, only possessing
restricted powers of development.

6. The peculiar characters of the pronuclei are due then to
the preliminary double mitosis; they can only accomplish
their functions by union (copulation) with a nucleus from a
different germinative cell.

7. Normal copulation can take place between two pronuclei,
and two only.!

8. The copulating nuclei, though of distinct origin, are
equivalent to one another, and either plays the same part to
the other. In the nuclear union, the supreme act of fecunda-
tion, there is neither male nor female ; the sexual modifications,
which are so obvious to our eyes, are mere accessory adaptations
to facilitate and ensure the approximation of the pronuclei,
which themselves are of nosex atall. Fecundation, reduced to
its lowest terms, is distinct from and independent of sexuality.

9. The high evolution of these accessory sexual adaptations
is only a proof of the extreme physiological importangaty of the
nuclear copulation in fecundation.

10. The chromatin of the nuclei represents their permanent
personality ; the other nuclear structures are subject to con-
tinual changes, destructive and recuperative, and only play an
accessory part.

11. The act of fecundation is completed by the union of the
chromatic elements of the two pronuclei into one single
nucleus, and this union may be effected at different stages
of nuclear evolution.

1 Maupas seems ignorant of the many cases of “ multiple isogamy” in
Protophytes: see ‘Some Problems of Reproduction,” in this Journal,

MAUPAS’S RESEARCHES ON CILIATE INFUSORIANS. 613

12. Probably the chromatic elements of the two pro-
nuclei retain their individuality in the copulation nucleus,
while only the nuclear accessories, their hyaloplasmas and
juices, undergo complete fusion. If this be so, fecundation
might be stated as the approximation of chromatic elements of
two distinct origins, and their incorporation into a single
nucleus.

The above statements, No. 3 especially, require fuller explana-
tion. In the first place, while two mitoses are usually required
for the formation of the female pronucleus in Animals, three, at
least, are always requisite in the Ciliata; a point which Maupas
dismisses as ‘‘a special adaptation of the organism of these
Protozoa.” His attempt to carry on this rule to the vegetable
world is very forced: it can, indeed, only be adopted for the
Archegoniata, where the successive formation of neck- and
belly-canal-cells compares very fairly with the formation of
polar globules in Animal ova; the first of which is known to
divide again in some Animals, like the neck-canal-cell of many
Archegoniates. But the identification with the processes in
the embryo-sac of Flowering Plants is only carried out by the
suggestion that all the authors who have described the evolution
of this structure are wrong in their description! Maupas
admits that there is difficulty in finding similar mitoses, and
anything corresponding universally to ‘corpuscules de
rebut ” in the evolution of spermatozoa, though some cases
present marked homologies. He also interprets the “ vege-
tative nuclei” of the pollen as corpuscules de rebut, which
may be physiologically correct, but finds, at least in part,
another, morphological, explanation. He points out that all
authors are agreed in regarding the production of polar bodies
as involving the expulsion and elimination of a quantity of
nuclear substance useless for the normal evolution of the germ
cells ; but that Weismann’s view as to the significance of this
elimination is disproved by the absolute identity of polar and
germ-nuclei in Metazoa; and of the sister offspring of the
micronuclei in the Infusorian gametes. For the present
Maupas abstains in the chapter on a “General Theory of

614 MARCUS M. HARTOG.

Fertilisation” from giving any explanation of his own; but in
the preceding one he suggests that ‘‘ we may always suppose
that a reduction in the quantity of material elements may act
with effect on molecular forces, attenuating their strength and
giving more delicacy to their play.”” Now, what we know of
molecular forces would be rather in favour of the view that
reduction of the masses would give more energy and violence
to the action of the nuclei. He is clearly right in rejecting
all explanations of “ polar bodies” which are inapplicable to
the three preliminary mitoses of the micronucleus of the
infusorian gamete.!_ Probably the phenomena are essentially
morphological, a question which will be discussed in my paper
on “ Some Problems of Reproduction.”

In the above abstract many interesting points have been
left untouched; much has been treated far too summarily.
But I trust that I have given an idea of the value and wealth
of the papers, and brought out their two cardinal points: the
limits of reproduction by continuous fission alone; and the
nature and results of conjugation.

1 A singular omission from this discussion is that Maupas nowhere brings
in his discovery that certain brood-nuclei of the rejuvenated conjugation-

nucleus itself abort as “ noyaux de rebut,” instead of becoming mega- or micro-
nuclei.

PSEUDOPODIA OF DIATOMS. 615

On the Occurrence of Pseudopodia in the Diato-
maceous Genera, Melosira and Cyclotella.

By

J.G. Grenfell, B.A., F.G.S., F.R.M.S.
With Plate XLI.

Tue diatoms on which these pseudopodia have been found
do not belong, as might have been expected, to the motile, but
to the non-motile forms, to the genera Melosira and Cyclotella.
The Cyclotella is certainly C. Kiitzingiana. The Melosiras
belong to one or two small species which have not yet been
satisfactorily determined.

They were first met with in April, in the large pond in the
gardens of the Royal Botanical Society of London, in the
Regent’s Park. This gathering consisted almost entirely of
small isolated frustules of Melosira, with a few Cyclotellas and
some Archerina Boltoni.! Later on filaments of frustules
of Melosira became commoner, while Archerina increased
enormously in number, Finally Cyclotellas replaced the
Melosiras, and the Archerinas vanished.

I next met with pseudopodia on Cyclotellas at Stanstead, in
Hertfordshire, whither I was directed by a friend.

Later I went to stay at Heytesbury, in Wiltshire, where I
found the river Wiley and the brooks full of a Melosira in
small isolated frustules, with long, delicate pseudopodia. Re-
cently I have found a good set of Cyclotellas with pseudopodia
in Kew Gardens, and others at Eastbourne. I infer that these

1 See Professor Lankester’s description of this organism, ‘ Quart. Journ.
Micro. Sci.,’ vol. xxiv, 1884.

616 J. G. GRENFELL.

small isolated Melosiras and Cyclotella Kiitzingiana have
these pseudopodia normally. At Stanstead, Heytesbury, Kew,
and Eastbourne there was no trace of Archerina. The reason
for mentioning this is that it was suggested that the external
protoplasm of Archerina migrated on to the diatoms.

The pseudopodia of the first gathering and of the Stan-
stead ones were easily seen for a part of their length with
a 4 object-glass, magnifying nearly 400 diameters; but in the
case of the Heytesbury Melosiras and most of the Cyclotellas
from the Botanical Gardens and Kew they are generally in-
visible, even when specially looked for. I think this helps to
explain why they have not been found before.

To study the whole length of the pseudopodia I found it a
good plan to dry the material on a cover-glass, which could
then either be mounted dry or stained or roasted. All the
figures on Pl]. XLI are from specimens thus treated. Quite
lately I have stained and mounted some of the Kew gathering
without drying. The results I hope to give in a future paper.
So far they have simply confirmed the main conclusions at
which I had already arrived.

The principal points to notice in the structure of the pseu-
dopodia are these: they are fairly stiff, and are non-retractile
to ordinary observation.

The length varies in Cyclotella from two and a half to six
times the width of the valves. In the Heytesbury Melosiras
this reaches fully nine times the width. They are very per-
manent; a slide prepared in April by simply sealing the
diatoms in the water in which they were found showed the
pseudopodia apparently unchanged after five months. The
dried slides show that the great majority of the pseudopodia
are arranged fairly symmetrically round the margin of the
valves. This is best seen in side view (figs. 6 and 7).

In slides of the Kew gathering prepared in the wet way a
great many Cyclotellas show a series of small tooth-like pro-
jections of protoplasm round the margins of the two valves,
arranged as regularly as the teeth of a circular saw. These
projections are the thicker bases of the delicate pseudopodia. On

PSEUDOPODIA OF DIATOMS. 617

a typical specimen I counted about forty-six of these projections,
while a roasted cover of the same gathering gave about forty-
six as the number of radiating ribs on the valve of the Cyclo-
tella. Hence there would appear to be a close connection be-
tween the number of the pseudopodia and the structure of the
diatom—a point of very great importance. Sometimes pseudo-
podia spring from the surface of the cingulum as well as from
the valves.

The pseudopodia are generally fairly straight ; occasionally
they branch at some distance from the valve. This was
especially the case in the earliest gathering (figs. 1 and 2),
where they branched repeatedly.

On dried cover-glasses it is common to find two or three
pseudopodia springing from a short thickened base. In water
unstained these bases are extremely hard to see.

The number of the pseudopodia is, on the whole, dndienpe
regular. On dried covers seventeen to twenty is the ordinary
number seen round a valve in side view (fig. 6); sometimes
twice that number (fig. 7).

The pseudopodia vary a good deal in thickness in different
gatherings. On dried unstained cover-glasses they often show
short portions of their length more opaque and solid-looking
than the rest, sometimes getting a beaded look. In this they
agree with the pseudopodia of Archerina (figs. 5, 6).

Rarely one comes across quite thick pseudopodia. Fig. 5
represents a striking form, with a few very stout ones.
It is noteworthy here that these four spring from the coarser
markings on the valve. Occasionally one of the pseudopodia
is thickened in the centre, as in fig. 6. Lastly, the pseudo-
podia of two diatoms seem to be able to fuse into each other,
and increase greatly in width, so as to form a broad band con-
necting the two diatoms. Whole chains are thus formed,
though two frustules only are more common (figs. 2 and 8).

Next as to the use of these pseudopodia, and the question
why other diatoms do not have them. The chief point to be
remembered is that these little Melosiras and Cyclotellas occur
mainly as isolated frustules, and are without the power of loco-

618 J. G. GRENFELL.

motion. Under these circumstances the pseudopodia serve three
purposes: 1. Protection. 2. Means of attachment. 3. Floats.

1. Protection.—The pseudopodia act in the ordinary way
as defensive spines. I have often seen large predatory In-
fusoria knocking these about, and absolutely unable to touch
them. The ordinary isolated diatoms can creep into mud or
débris out of harm’s way. To these the stiff pseudopodia
would be quite useless.

2. Means of Attachment.—At Heytesbury I found the
diatoms in running water, especially amongst filamentous
weeds. Here the use of the pseudopodia is quite obvious.

3. Floats.—In the Botanical Gardens they are all over the
still waters, and no doubt the large extra surface given by
these pseudopodia helps to keep them floating. The remark-
able pelagic diatoms Chetoceros also have long processes, but
of a different kind: that is to say much coarser, and obviously
forming part of the siliceous skeleton.

The next question is the substance of which these pseudopodia
are made. I think the facts point conclusively to that sub-
stance being protoplasm. ‘They are destroyed by nitric acid,
while those of Cheetoceros are not. All the finer parts are at
once destroyed by roasting at the lowest red heat. The thick
connecting bands and the thickened bases of the pseudopodia
will stand a low red heat occasionally, while they, too, are
entirely destroyed by a strong heat.

They stain readily with Kleinenberg’s hematoxylin. With
Schultze’s solution they give no cellulose reaction, nor with
iodine and sulphuric acid. Boracic carmine, which does not
stain cellulose, stains the bases of the pseudopodia strongly,
and just the same colour as the cell-membrane—the fine part
slightly. It is quite probable that some kind of cuticle is
secreted by the protoplasm in contact with the water, and that
this gives to the bases and connecting bands their resisting
power at a low red heat. The bases and cell-membrane always
behave in nearly the same way to stains. Two other proteid
stains, said not to stain cellulose, were tried. Picro-nigrosine
stained the diatom bodies well, the bases fairly, and also the

PSEUDOPODIA OF DIATOMS. 619

finer parts. Alcoholic safranin just stained the pseudopodia ;
but all alcoholic stains are apt to fail with them.

The evidence of the stains, therefore, joins with the other
tests in pointing to the presence of protoplasm with or without
a very fine cuticle of uncertain nature. The thick pseudopodia,
with their variation in density (fig. 5), also point to a very fine
cuticle with protoplasmic contents. There is no evidence in
any of my slides of a layer of protoplasm normally present out-
side the diatom shell. The pseudopodia or their bases spring
straight from the shell. Most probably there is a fine layer of
protoplasm outside the shell, for Imhoff has shown that the in-
ternal protoplasm reaches the surface, but it is not thick enough
to be visible in optical section.

In the Kew gathering I have met with two or three speci-
mens snrrounded by a thick layer of what looks like a gelatinous
substance, not granular. The pseudopodia, however, are inde-
pendent of this, and are clearly traced through it up to the
margin of the shell of the diatom.

I have also seen granular or fluffy substance round Cyclo-
tellas, which might be protoplasm; but this is not at all
common. I have seen it occasionally round other forms which
have no pseudopodia.

Next we come to what has been said or suggested as to the
meaning of the pseudopodia.

The first suggestion was that these were not diatoms at all,
but the identification of a well-known species, Cyclotella
Kiitzingiana, with pseudopodia has settled that point. Next
it was suggested they might be extensions of the gelatinous
layer which Professor H. L. Smith has shown to surround
many diatoms.

But there is no evidence here of any kind of normal gela-
tinous envelope, and the stiffness of the pseudopodia and the
permanence of the bases tell against this theory, which is
further negatived by the fact already stated, that the pseudo-
podia penetrate the gelatinous layer when this is present.

A third hypothesis, based on the remarkable slides where
the diatoms and Archerina are mixed in countless profusion, is

620 J. G. GRENFELL.

that the pseudopodia are a vegetable growth covering every-
thing in the field. But here the Heytesbury set are conclu-
sive : on a single slide you may have some hundreds of diatoms
of various kinds ; about 200 may be the small Melosira, nearly
every one of which will have pseudopodia, while nothing else
in the field has any. Besides, botanists will, I think, agree
that there is nothing plant-like in these forms. Another sug-
gestion was that they are like the filaments of Polysiphonia.
But these latter never branch and are not granular, and could
not form connecting bands.

_ Another class of suggestions was that these pseudopodia do
not belong to the diatoms, but to an investing animal like
Vamppyrella, which devours Gomphonema. But if they belong
to an investing animal, where is the animal which invests?
Vampyrella, any way, is visible both when wandering in search
of diatoms and when investing. Besides, Vampyrella projects
its pseudopodia from any part of the diatom, while here
they are mainly confined to a definite tract. And Vampy-
rella and other predatory animals do not have pseudopodia
so regularly symmetrical, nor so constant in number. I have
seen what appeared to be Leidy’s Biomyxa vagans, which is
something like Vampyrella, investing diatoms. And here also
the pseudopodia had no relation whatever to the structure of
the diatom, while the animal itself was clearly visible outside
of the diatom. But, strongest proof of all, Vampyrella devours
the diatom and kills it in a couple of hours; while these
diatoms lived for five weeks in one bottle, quite healthy all the
time, and dead ones were not found.

I do not think that any theory based on Vampyrella or vam-
pyrelloid animals (as suggested at the British Association) will
explain the facts. If the pseudopodia are foreign to the
diatom, it would have to be a case of symbiosis between the
diatoms and unknown invisible animals. I know of no similar
case,

All the phenomena seem to me to point to these pseudo-
podia being filamentous extensions of the cell protoplasm, pro-
bably strengthened by cuticular deposit.

PSEUDOPODIA OF DIATOMS. 621

The fact that they do not move or quickly retract under
stimulus would not necessarily distinguish them altogether
from true pseudopodia ; at any rate, if the processes of Arche-
rina are to be called by that name these must also, for it is
impossible to separate them. It might, however, be an advan-
tage to have some distinctive name for such stiff, slowly
changing or unchanging pseudopodia.

It remains to treat briefly of the morphological resemblances
between these diatoms and other forms. In doing this I wish
simply to point out resemblances or differences, not to draw
any large conclusions.

The first point that strikes one about these pseudopodia is
their extreme unlikeness to any plant or part of a plant.
Secondly, it is remarkable that when pseudopodia are dis-
covered on diatoms these resemble a type which, so far as I
know, is absolutely confined to the Heliozoa, forms with which
the diatoms are already connected by the presence of a more
or less siliceous test. All the Heliozoa are characterised by the
relative stiffness of their pseudopodia ; but in Archerina this
relative stiffness becomes practically absolute, as in these dia-
toms. I do not know of any described form except Archerina
which has these non-retractile pseudopodia.

But the similarity in type of the pseudopodia is not confined
to rigidity ; it extends to general form. As far as I know,
every single detail of form in the diatoms can be matched
amongst the Heliozoa with the exception of the repeated
branching of those of Melosira. ‘The resemblance of the diatom’s
processes to those of Archerina is still closer: nevertheless in
99 cases out of 100 it is quite easy to distinguish on a dry slide
the pseudopodia of Archerina from those of the diatoms.

They are distinctly wider and taper regularly. But in the
100th case you get an Archerina colony the pseudopodia of
which are absolutely undistinguishable from those of the
diatoms. Their size and shape are the same as those of
Cyclotella, and on a dry cover they show the thickened bases
and the denser portions of the filament which I have described
as characteristic of the diatoms.

622 J. G. GRENFELL.

If one bears in mind that the little Melosiras have very little
silica, possibly none at all at times, and that both they and
the Archerinas contain chlorophyll bodies, the resemblance
becomes still more striking. It is also a striking fact that the
connection of the diatoms by bands of protoplasm finds its
counterpart amongst Heliozooid Protozoa, as in Monobia
confluens. Another point of agreement is the fact that
Archerina at times has very few but very large pseudopodia,
just like the Cyclotella.

A fact about Archerina as yet unpublished is worth notice.
In staining with Schultze’s fluid and with iodine and sul-
phuric acid I obtained the clearest evidence that the cuticle
of the chlorophyll bodies is made of cellulose. In this it is
more plaut-like than the diatoms themselves, which do not
give the cellulose reaction. Cellulose has been found in a
number of animals, but still it is interesting that while the
pseudopodia draw the diatoms nearer to the animals, this cel-
lulose draws Archerina nearer to the plants. This fact is
likely to be used as an argument in favour of Archerina being
a case of symbiosis, and such a view may be extended to
these diatoms. I will not discuss that view now. I do not
think it is conclusive in the case of Archerina, much less in
the case of the diatoms, where all the facts seem to point to
the pseudopodia being integral portions of the diatoms.

PSEUDOPODIA OF DIATOMS, 623

EXPLANATION OF PLATE XLI,

Illustrating Mr. J. G. Grenfell’s note “‘ On the Occurrence of
Pseudopodia in the Diatomaceous Genera, Melosira and
Cyclotella.”

Fic. 1.—Melosira of the first gathering in the Botanical Gardens, much
branched. The second frustule shrivelled in drying. Stained.

Fic, 2.—Melosira of the same gathering. Stained.
Fie. 3.—Melosira filament from Botanical Gardens. Stained.
Fic. 4.—From Heytesbury, in Wiltshire. Unstained.

Fie. 5.—Cyclotella from Botanical Gardens. Four thick granular pseudo-
podia. Unstained.

Fie. 6.—Cyclotella Kiitzingiana from Botanical Gardens. Unstained.
Fie. 7.—Ditto, with more pseudopodia,

Fic. 8.—Two forms from Botanical Gardens, roasted at low red heat,
showing projections and connecting band,

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REVIEW.

A Monograph of the Victorian Sponges, by ArtHUR DENpDy,
D.Sc., F.L.S., Fellow of Queen’s College, and Demonstra-
tor and Assistant Lecturer in Biology in the University of
Melbourne. Part I.—The Organisation and Classification
of the Calcarea Homocela, with Descriptions of the
Victorian Species (with plates i—xi). Melbourne, July,
1891.

UnpeEr the above title a most interesting paper on Sponges
has appeared, giving, it is not too much to say, the first
attempt at an accurate description of the histology of the lower
Ca.carea since Metschnikoff’s paper in 1879. I have been
working at the same group myself for about half of the last
five vears. As it is likely to be still some months before my
results appear, I think that it may facilitate discussion, while
noticing some of the principal features in Mr. Dendy’s paper,
to indicate at the same time the extent to which my work has
led me to similar or different conclusions. And I may com-
mence generally by saying that in the plates of this paper I
have had for the first time the pleasure of seeing drawings
which represent accurately the structures which have come
under my notice.

The introductory criticism on Haeckel (p. 3) appears some-
what to underrate the power of variation in calcareous sponges,
particularly their plasticity to environment. Thus Sycandra
raphanus has formed a special variety in the Naples Aquarium,

VOL. XXXII, PART 1V.—NEW SER. sia

626 REVIEW.

unknown in the bay and hitherto undescribed. It outwardly
resembles 8S. capillosa. Vosmaer’s views on Leucandra
aspera (‘ Mitt. Zool. Stat. Neapel.,’ vols. ili and iv) may require
some modification, but its variation is certainly enormous. It
is true that the Homoccela of Naples seem fairly constant in
outward form and canal-system (sensu Dendy), but evidence
points to their being identical with species of very different
aspect in other localities. I disagree with Mr. Dendy that
‘there is no doubt that a species has no existence in nature.”
This, however, is an academic question of general zoology,
which should be treated by more competent hands than mine.
In his rejection of von Lendenfeld’s Homodermide and
Leucopside he has taken the only course open. It is to
general advantage that it should be stated plainly that the
histology figured in Homoderma is alone sufficient to con-
vince any student of Calcarea that the structures described
were never seen.

The attempt to group the Homoceela according to structure
is valuable and suggestive. It does not claim to be sufficiently
developed to be considered as a natural classification. There
are three sections of the Homocela: I. Simp licia, solitary, or
with individuals of easily recognised individuality. II. Reti-
cula, anastomosing tubes, in which individuals are unrecog-
nisable. ILI. Radiata (one species), a (large) central
Ascon-tube bearing secondary radial tubes. The last group
or species (Leucosolenia tripodifera) is perhaps the most
important discovery recorded in the paper. The radial tubes
branch and branch again, until they are set thickly together
like a wall round the wide central osculate sac into which they
open, so as to simulate precisely the canal-system of a Sycon.
But the central tube retains its collar-cells and pores, and
its true wall differs in no structural respect from that of its
tributaries.

This observation and its treatment by the author appear to
me most suggestive. I think that an examination of inter-
mediate forms will convince him that there are many other
Homocela to be included in the Radiata. Ascandra

REVIEW. 627

Lieberkuhnii! without doubt, as it is found at Naples,
comes under his definition of the group, and in its fir-tree-like
form, branching at right angles into smaller and smaller tubes,
shows a stage antecedent to L. tripodifera. Its main oscular
tubes are as much (in the specimen before me) as 1°5 cm. by
‘4 cm. ; its smallest branches about ‘01 cm. ; between these all
intermediate sizes. A most suggestive feature, on which this
is not the place to dwell, is that the unpaired rays of the
spicules in the branches are mostly distally directed. In
Ascaltis cerebrum the oscula (not pseudoscula ; Haeckel
was probably misled by the outward appearance when he
figured what Dendy would term an “ inverted canal-system ”’)
open from spaces 0°3 cm. wide, lined with collared cells con-
tinuously up to the granular lip; its wide tributaries are not
superficial as in the oscula of Ascandra reticulum, but
deep, and covered with secondary ramifications, ranging down
to about ‘01 cm. diameter. This sponge might seem almost
to lead us to Dendy’s Leucosolenia stipitata, placed by
him in Section II. But without going further I would point
out that the three previous forms all possess triradiates and
quadriradiates more or less slender and pointed ; that while
those of A. Lieberkuhnii are of nearly the same dimensions as
L. tripodifera, and the apical ray similarly curved, A. cere-
brum possesses on its external surface the very tripods which
give L. tripodifera its name. Will Mr. Dendy consider the
possibility that his Radiata may already advantageously step
forth as a genus? e.g. Homocela. Genus 1. Leucosolenia.
The growth of the tubes is mainly confined to new branches ;
type L. clathrus, L. dubia. Genus 2. Nardoa. The
growth of the tubes is continuous, the newest branches have
consequently smallest diameter; type N. tripodifera, N.
Lieberkuhnii (?).

The Reticulata are again divided according as the gastral
cavity is or is not traversed by ingrowths of mesoderm. I
must state my strong suspicion that the “ingrowths of meso-

* For my own observations I use Haeckel’s names to avoid the coining of
new combinations,

628 REVIEW.

derm ” are the ameeboid cells observed long since in the gastral
cavity of certain sponges after digestion, which, as in Ascetta
clathrus, form such traversing processes, and which I
believe, with the older workers, to be collar-cells; to the
amceboid metamorphosis of which Dendy makes no allusion.
In these Australian sponges there appears to occur none with
a many-layered endoderm. This structure, observed by
Haeckel and since universally discredited, certainly appears
in Ascetta clathrus, and—I hope I am not wrong in say-
ing—was observed some years ago by Mr. Hardy, of Caius
College, Cambridge, in a Leucosolenia found by him at
Plymouth.

Turning to histology, Dendy finds “the ectoderm of the
Homoceela agrees precisely with what Schulze has described
for Sycandra raphanus.” Although this form occurs in
the Homoceela, it is in my experience rare. The typical
ectoderm (e.g. Ascetta clathrus) I find composed of onion-
shaped gland-cells containing a nucleus and granules, and
provided with a usually fine duct, the expanded end of which
forms the hexagonal area whose boundaries are, in the case of
most sponges, all that has been observed. In Ascetta
clathrus and blanca almost the whole ectoderm is of this
type, and at least a large part of it in Ascaltis cerebrum,
Ascandra reticulum, and Ascetta primordialis; on
the external surface in Sycandra raphanus, Leucandra
aspera (sensu Vosmaer), and a new sponge which I pro-
visionally name Sycaltis leuconides (having a Sycaltis
skeleton and a Leucon-like canal-system, and thereby neces-
sitating a change of classification among the Heteroceela).
Making such a statement without details or figures, 1 will
add that in 1887 Dr. Vosmaer very kindly volunteered to me
permission to quote him as being convinced with respect to
the ectoderm of Leucandra aspera. This structure of
ectoderm was described and figured by Merejkovsky for
Halisarca in 1878 (‘Mém. Acad. Petersburg’), by Metsch-
nikoff for Ascetta blanca in 1879 (“Spong. Studien,”
‘Z. f. w. Z.,’ 32). Though occurring in one of the latter’s

REVIEW. 629

best known papers I have never seen this description alluded
to, and the entire paper has curiously escaped Dendy’s notice.
Metschnikoft’s pl. xxi, fig. 1, gives evidence of the flask-
shaped epithelium in the young Halisarca, corroborating
Merejkovsky’s description, with which, indeed, the figures
of Schulze and similar ones of Metschnikoff’s are at variance
only in interpretation. I have found this glandular ectoderm
in a horny sponge (Aplysina ?), an Axinellid, and a Renierid.
Von Lendenfeld’s “mesodermal gland-cells” are certainly
_ nothing else in Calcarea, and as the descriptions are identical
they are probably the same in horny sponges; many “ eestho-
cytes ” and similar structures in all likelihood have the same
foundation in fact. I therefore personally believe that the
typical ectoderm, not only of Calcarea, but of sponges, is a
glandular epithelium of flask-shaped cells with dilated mouths,
and that on external surfaces this is probably the usual form.’
Mr. Denby has shared with most others the disadvantage of
working on specimens more or less shrunk by preservation in
alcohol ; to this shrinking, to generalisation from the epithe-
lium of canals (more easy to observe than the defended
exterior), and to deduction of the existence of a flat epithelium
from mere hexagonal silver lines, I attribute the overlooking
of this structure by all but the two Russian authors.

Dendy has not found cilia on the ectoderm of Homoceela,
and throws much doubt on the figures of Lendenfeld, where
they are invariably introduced. After long comparison of
Dr. von Lendenfeld’s descriptions of Calcarea with the original
structures I have no hesitation in saying that his “low flat
plates filled only partially with protoplasm ; from this plate
threads extend which pervade the cell-cavity,” are com-
pletely imaginary. His flagella certainly do not exist in the
Calcarea I have examined ; they are probably a generalisation
from some structures he has seen in horny sponges. In
Aplysina (?) I have found threads standing vertically from the
ectoderm and precisely simulating flagella; they are undis-

1 T think that it may prove the primitive Metazoon ectoderm, and will
probably be found in various larve.

630 REVIEW.

solved in 30 per cent. caustic ammonia, and are probably
vegetable.

In the case of the sheath on the apical ray of gastral
spicules, Dendy most justly says that there is no evidence to
prove their mesodermal origin, and I differ from him in that I
am thoroughly prepared to accept them as endoderm. The
time has come to free the study of the Porifera from the fetish
of mesoderm, and to render to her grasp only that to which
embryology can prove her entitled. The presumption lies in
favour of the old layers, the ectoderm and endoderm; the
onus probandi is on the new-comer.

My own work has led me to regard the endoderm not only
as multiform, but as most proteic. Dendy recognises it as
** polymorphic,” but this appears only to refer to the relative
state of “ retraction”’ of the collars and flagella. I agree with
all his description and figures of these, both in this and preced-
ing communications, and I have now myself observed, in the
living Sycandra raphanus, the coincidence of flagella and
Sollas’s membrane which (in Halichondria p anicea) he was
the first to meet with. But I have come to the conclusion that
this coincidence is only transitory ; and while he most generously
yields priority for the theory of filtration, I have been brought
to relegate it to the disunited collars, and to believe that the
membrane of Sollas is a valvular adaptation to prevent the
reflux of water past satiate and therefore inactive cells. Where
he writes “retraction ” I would write ‘‘ disappearance,” and I be-
lieve that in the old “ Verwandelung der Geissel bewegung ”’ in
«© Amoeboide Bewegung ” lies the key to many of the anomalies
of the intimate structure of sponges. He draws and describes
in Leucosolenia cavata “ yellow granules,’ which he more
than suggests are symbiotic alge. I have worked at then—
besides in former years—during the last nine months in Ascetta
clathrus (where, besides the description he quotes from Bower-
bank, they were described and figured by Metschnikoff, loc. cit.)
and A. primordialis. I find Dendy’s drawings and descrip-
tions of their behaviour and relations most accurate; in Asc.
clathrus there is an additional point of interest that the

REVIEW. 631

granules in the (glandular) ectoderm-cells differ from these only
in being of smaller size. I have been very slowly and gradually
led to the conclusion that the bodies in question, which I propose
to call “ Metschnikoff cells,” are metamorphosed collar-cells; that
by their reaching to the exterior and becoming perforated, pores
are formed; and that the granules of these and of the ectoderm,
and of the glandular ectoderm in general (and possibly the
granular cells so frequently described beneath it in Silicea),
are excretory.

In the nucleus of the ovum Dendy finds in L. pelliculata
nucleoli and circumferential granules; in L. depressa, in
addition, a faint reticulum. In Asc. clathrus (nitric acid
and borax carmine) I find a distinct and typical reticulum
with small granules at the nodes. I have found also a large
nucleolus with vacuoles, possibly artificial. In the matrix
capsule of sponge embryos Dendy has almost established a pro-
prietary interest; in Leucosolenia Wilsoni he finds they
have no connection with ectodermal cells. In A. clathrus
ova occur which appear to have such a connection, but when full
of yolk they lie in sacs dependent in the gastral cavity, clothed
with collared cells, of which some are always metamorphosed,
and which are in continuity by the neck of the sac with the
general endoderm. It may be worth adding to his instances of
specially robust external spicules, besides Asc. cerebrum,
whose “tripods ” are confined to the outer surface, Ascandra
reticulum, in some varieties of which the acerate (*‘ orceote ”)
spicules are so confined, while in others they disappear. The rod-
like bodies he describes on the gastral surface of Sollas’s mem-
brane inL. tripodifera Ido not believe to belong to the sponge ;
he himself accepts them doubtfully. But it is curious that
in the allied A. cerebrum and the variety of A. primordialis
which simulates its form (variety of A. cerebrum simulating
the spicules of A. primordialis? nova species ?) the collars and
flagella most frequently appear to be replaced by a network of
threads. Nothing but its “ constancy and peculiar and regular
arrangement,” to quote Dendy’s words, could, however, give any
doubt that these are vegetable, which on the whole is at present

632 REVIEW.

my belief. With reference to this membrane, the descrip-
tions of von Lendenfeld, in contradiction to the statements of
Sollas and Dendy, are, I have convinced myself, as worthless as
those of the unique and quite different form of collar-cell till
very recently described and figured by him in all groups of
sponges.

Dendy’s observations of afferent canals lined with ectoderm
perforating the walls of L. stolonifera are most interesting,
and in my judgment probably of generic importance. Though
A. clathrus is much more thickly walled than most of the
group, the communication is established by a single perforated
granular cell as in other Homoceela, and strictly homologous
with the granular ring round a prosopyle in the Heterocela
(cf. Poléjaeff, Grantia tuberosa). This description does not
refer to other large pores which occur in this lipostomous
sponge, whose structure and morphology I have not worked
out.

In conclusion, if in this review points of difference rather
than of agreement are accented, it is because the former take
more words to express. This paper lies throughout in lines
parallel to those on which I have been long labouring; and it
may so not be impertinent to give it the cordial welcome of
a fellow-worker who finds a great stride made forward towards
the knowledge of a group that appeared almost insoluble.

GrorGE BrippeEr.

Napues; August, 1891.

NID XS OW Olli ees Tain

NEW SERIES.

Allopora, gonophores of, by Hickson,
375

Amphioxus, the later larval develop-
ment of, by Willey, 183

Archenteron of Rana, 451

Assheton and Robinson on the primi-
tive streak, archenteron, and ger-
minal layers of the frog, 451

Beddard on an earthworm allied to
Nemertodrilus, 539
»» on two new genera of earth-
worms, and remarks on Nemerto-
drilus, 235
Benham on the nephridium of Lum-
bricus, 293
Bidder, G., review of Dendy’s mono-
graph of the Victorian sponges,
625
Bourne, A. G., on Megascolex, 49
3 on Pelomyxa viri-
dis, and on the vesicular nature of
protoplasm, 357
i on the naidiform Oligo-
cheta, and on Pristina and Ptero-
stylarides, 335

Crisia, the British species of, by
Harmer, 127

Decapod Crustacea, renal organs of,
by Weldon, 279

Dendy, sponges of Victoria (review),
625

VOL. XXXII, PART IV.—NEW SER.

Dendy, studies on the comparative
anatomy of sponges, No. III, 1
re a No. IV, 41
Diatoms, occurrence of pseudopodia
in, by J. G. Grenfell, 615
Distichopora, gonophores of, by Hick-
son, 3875

Earthworm allied to Nemertodrilus,
539
Earthworms, course of the blood in,
by A. G. Bourne, 49
Re two new genera of, by
Beddard, 235

Frog, primitive streak, archenteron,
and germinal layers of, 451

Germinal layers of Rana, 451

Grantia, Dendy on, 1

Grenfell, J. G., on the occurrence of
pseudopodia in diatoms, 615

Halichondria panicea, Dendy on,
41

Hardy, W. B., on the histology and
development of Myriothela, 505

Harmer on the British species of
Crisia, 127

Hartog, abstract of Maupas’s re-
searches on ciliate Infusorians, 599

Hickson on the medusz of Millepora,
and on the gonophores of Allopora
and Distichopora, 375

UU

634

Infusorians, ciliate, researches on; by
Maupas, 599

Laurie on the development of Scor-
pio fulvipes, 587

Lee on a little-known sense-organ in
Salpa, 89

Lumbricus, nephridium of, by Ben-
ham, 293

Maupas’s researches on multiplication
and fertilisation of ciliate Infusoria
—abstract by Hartog, 599

Medusz of Millepora, by Hickson,
375

Megascolex ceruleus, by A. G.
Bourne, 49

Microbes, immunity against, by Ruffer,
Part I, 99

y as Part II, 417

Millepora, meduse of, by Hickson,
375

Mpriothela, histology and develop-
ment of, 505

Naidiform Oligocheta, by Bourne, 335
Nemertodrilus, earthworm allied to,
539
%y remarks on, by Bed-
dard, 235
Nephridium of Lumbricus, by Ben-
ham, 293

Oligocheta, naidiform, by Bourne, 335

Pelomyxa viridis, by Bourne, 357

INDEX.

Phymosoma, a new species of, by
Shipley, 111

Primitive streak of Rana, 451

Pristina, by Bourne, 335

Protoplasm, vesicular nature of, by
Bourne, 357

Pterostylarides, by Bourne, 335

Renal organs of certain decapod Crus-
tacea, by Weldon, 279

Robinson and Assheton on the primi-
tive streak, archenteron, and ger-
minal layers of the frog, 451

Ruffer on immunity against microbes,
Part I, 99

33 eB)

Part II, 417

Salpa, sense-organ in, by Lee, 89

Scorpio fulvipes, development of
587

Sense-organ in Salpa, 89

Shipley on a new species of Phymo-
soma, 11]

Sponges of Victoria, by Dendy (re-
view), 625

», Studies on the comparative

anatomy of, by Arthur Dendy, No.
Til al

No. IV, 41

Teichonide, Dendy on, 1

39 33

Weldon on the renal organs of certain
decapod Crustacea, 279

Willey on the later larval development
of Amphioxus, 183

PRINTED BY ADLARD AND SON, BARTHOLOMEW CLOSE.

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