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4

CONTENTS

CONTENTS OF No. 213, N.S., SEPTEMBEE, 1909.

MEMOIRS : page

Development and Origin of the Respiratory Organs in Araneae.

By W. F. Purcell, Ph.D., Bergvliet, Diep River, near Cape
Town. (With Plates 1 — 7, and 7 Text-figures) . . 1

Notes on the Nephridia of Dinophilus and of the Larvae of
Polygordius, Echiurus, and Phoronis. By E. S. Goodrich,
F.R.S., Fellow of Merton College, Oxford. (With Plate 8) . Ill
Further Notes on a Trypanosome found in the Alimentary Tract
of Pontobdella muricata. By Muriel Robertson, M.A.,
Carnegie Research Fellow ; Junior Assistant to the Professor of
Protozoology in the University of London. (With Plate 9, and
5 Text-figures) ...... 119

CONTENTS OF No. 214, N.S., OCTOBER, 1909.

MEMOIRS :

Dendrosoma radians, Ehrenberg. By Sydney J. Hickson, D.Sc.,

F. R.S., Beyer Professor of Zoology in the University of Man-
chester, and J. T. Wadsworth. (With Plate 10) . . 141

On the Structure of the Excretory Organs of Amphioxus. Part 2.

— The Nephridium in the Adult. Part 3. — Hatschek’s Ne-
phridium. Part 4. — The Nephridium in the Larva. By Edwin
S. Goodrich, F.R.S., Fellow of Merton College, Oxford. (With
Plates 11 — 16 and 1 Text-figure) .... 185

Intra-cellular and General Digestive Processes in Planariae. By

G. Arnold, from the Cytological Laboratory of the University

of Liverpool. (With Plate 17) . . . . 207

Professor Hubrecht’s Paper on the Early Ontogenetic Phenomena
in Mammals ; an Appreciation and a Criticism. By Richard
Assheton, M.A., Trinity College, Cambridge; Lecturer on
Biology in the Medical School of Guy’s Hospital, in the Univer-
sity of London. (With 5 Text-figures) . . . 221

IV

CONTENTS.

CONTENTS OF No. 215, N.S., DECEMBER, 1909.

MEMOIRS : PAGE

The Formation of the Layers in Amphioxus, and its bearing
on the Intei'pretation of the Early Ontogenetic Processes in
other Vertebrates. By E. W. MacBride, D.Sc., LL.D., F.K.S.,
Strathcona Professor of Zoology in McGill University, Montreal.
(With Plates 18 — 21, and 10 Text-figures) . . . 279

The Structure, Development, and Bionomics of the House-fiy,
Musca domestica, Linn. Part III. — The Bionomics, Allies,
Parasites, and the Relations of M. domestica to Human
Disease. By C. Gordon Hewitt, D.Sc., Late Lecturer in
Economic Zoology, University of Manchester. (With Plate 22) 347

The Develojnuent of the Temnocephalese. Part I. By Professor
W. A. Haswell, M.A., D.Sc., F.R.S. (With Plates 23 — 25) . 415

Experimental Observations on the Organs of Circulation and
the Powers of Locomotion in Pennatulids. By Edith M.
Musgrave, D.Sc. (nee Pratt), Late Honorary Research Fellow
in the University of Manchester. (With Plates 26 and 27) . 443

CONTENTS OF No. 216, N.S., FEBRUARY, 1910.

MEMOIRS :

On Certain Features in the Development of the Alimentary Canal
in Lepidosiren and Protopterus. By J. Graham Kerr,
Professor in the University of Glasgow. (With 13 Text-figures) 483
The Phylogeny of the Tracheie in Araneie. By W. F. Purcell,
Ph.D., Bergvliet, Diep River, near Cape Town. (With Plate

28, and 21 Text-figures) ..... 519

On the Reproduction of Kalpidorhynchus arenicola;
(Cnghm.). By Margaret Robinson, University College,
London. (With Plate 29) .... 565

Studies in the Experimental Analysis of Sex. By Geoffrey
Smith. (With Plate 30) ..... 577
Some Points in the Physiology of Lamellibranch Blood-Corpuscles.

By G. H. Drew, B. A. Cantab. (With Plate 31) . . 605

Note on the Cytology of Calothrix fusca. By Dr. N. H.

SwELLENGREBEL, Amsterdam. (With Plate 32) . . 621

Tropidonotus and the “ Archenteric Knot ” of Ornithorhynchus.

By Richard Assheton, M.A. (With Plate 33) . . 633

Title, Index, and Contents.

RESPIRATORY ORGANS IN ARANE.E.

1

Development and Origin of the Respiratory
^ Organs in Aranese.

By

\V. F. Fiiiooll, Pli.D.,

Bei'^vliet, Diep Rivei’, near Cape Town.

With Plates 1 — 7 ami T Te.xt-fignres.

CoNTKNT.S.

I. Introduction . . . . .

Material . . . . .

Biological Observations . . . .

Treatment ....

II. General Orientation . . . .

Ining-hooks . ... .

Tracliea; . . . . .

III. Historical (Development) . . . .

Development of the Lung-books in Arachnida.
Development of the Trachea; in Aranese

IV. The Provisional A1)dominal Appendages in the Embryo

of Attus floricola

V. The Development of the Lung-hooks .

Stage with two Pulmonary Furrows

Stages with three or more Pulmonary Furrows

Formation of the Spiracle . . . .

Sinking of the Apj^endage

Formation of the Pulmonary Saccules

Comparison with the Gill-hooks of Limnlus

Later Development of the Pulmonary Saccules

The Chitinous Lining of the Pulmonary Saccules

The Moidting of the Lung-hooks

The Operculum of the Lung-hooks .

The Lung-hooks of the Young Spider

Critical Remarks on the Literature

The Fully Developed Lung-hooks of Spiders .

VOL. O-I, PART 1. NEW SERIES. i

PAOE

2

;i

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7

9

9

11

12

12

P)

Hi

17

17

20

02

o.>

2;l

25

28

31

32

34

35
30
41

2

W. F. PUECELL.

PAGE

VI. The Development of the AEclominal Longitudinal Muscles

and their Tendons . . . . 4F

VII. The Entapophyses (Ectodermal Tendons) of the Pul-
monary Segment . . . .47

The Inteii^xilmonary (Epigastric) Fold in the Adult of
Attus . . . . . 4h

The Inteipulnionary Fold in other Spiders . . iiO

VIII. The Development of the Trachea; and the Entapophyses

of the Tracheal Segment . . . 53

The Post-embryonic Development of the Tracheal
Plate . . . . .57

Critical Remarks on the Literature . . .61

Tlie Attus Type and Similar Types of Tracheae in
Other Spidei's . . . . .62

The Agelena Type of Trachea; and its Develoiunent . 63

The Tracheae in the Dysderidae . . .68

Tiie Tracheae in Argyroneta aquatica . . 70

The Tracheae in the Scytodidae. Palpimanida*. and
Filistatidae . . . .71

IX. The Entapophyses of the Third and Fourth Abdominal

Appendages (the S2>inners) . . .74

X. General Conclusions . . . .75

The Origin of the Tendinal or Medial Tracheal Trunks
in Araneae . . . . .76

The Origin of the Lateral Tracheal Trunks in Araneae 78

The Origin of tlie Secondary Tracheal Tubules . 80

The Origin of the Lung-books in Arachnids . . 81

The Homologies of the Pulmonary Segments in

Arachnids . . . . .85

XI. Historical List of Papers concerning the Lung-books of

Arachnids . . .02

List of Literature . . . .06

Exf)lanation of the Plates . . .103

I. Introduction.

It is just one hundred years ago that the first anatomical
account of the lung-books of Arac hnida was published by
Meckel (’09), who, like his immediate successors, looked upon
these organs as gills, and it was not until 1828 that their
pulmonary nature was recognised by Johannes Muller (’28a,
’28b) and Straus-Durckheim (’28j. The latter was also, 1 be-

EESPIRATORY ORGANS IN ARANE^.

3

lieve, the first to point out (p. 315) that the lung-books could
be regarded as a special form of tracheae, a view which was
later on elaborated by Leuckart (’48, p. 119 note, and ’49) and
for a time generally accepted, until the appearance of Ray
Lankester’s paper, “Lim ulus: an Arachnid,” in 1881, opened
up the probability of the branchial origin of these organs.

While working at certain points in the embiyology of a
spider some years ago it occurred to me that a more careful
and detailed investigation of the development of the lung-
books and tracheae than had hitherto been attempted would
probably reveal some points of interest in connection with the
origin of these organs, and indeed it soon appeared that two
important facts had been entii’ely overlooked, viz. (1) the
appearance of the earliest lung-leaves on the free
posterior side of the provisional abdominal ap-
pendages quite outside of the pulmonary invagi-
nation, and (2) the origin of a considerable part of
the trachem from ectodermal tendons (entapophyses)
and not from lung-books. This latter appeared to me a
point of particular interest, as it is the only case, I believe, in
which the origin of a trachea from another organ not re-
spiratory in nature can be clearly demonstrated.

My investigations were carried out in the years 1894 and
1895, in the Zoological Laboratory of the University at Berlin,
and my thanks are due to Geheinn’ath Prof. F. E. Schulze
for the use of his splendidly equipped laboi’atory. About one
third of the text had already been written and most of the
figures drawn when I left Berlin in 1895 for South Africa,
where various circumstances prevented the completion of the
paper for the press until quite recently.

Material. — The material for the development was collected
in the neighbourhood of Berlin, and consisted of the embryos
and young of Sitticus (Attus^) floricola C. K., of which
T had an unlimited supply of all the required stages of de-
velopment. Besides these T examined a small number of

* Tins name has been recently discarded by E. Simon and Sittici;s
snb.stitnted in its stead.

4

W. F. PURGE r.L.

embryos and young of A<relena 1 ab jM’i n t li i ca and of
Tegenaria atrica. but the account of the embryology in
the following pages applies only to Attus floricola, unless
the contrary is expressly stated.

Tlie material required for anatomical purposes consisted of
adult or snbadnlt specimens of foi’t}'-one species mostly ob-
tained in the neighbonrboods of Berlin or Cape ^'own, as
stated in the list given below. The specific determination of
the European specimens (except Tegenaria atrica) were
made from Dahl (’83), but the families and genera are in
agreement with E. Simon ('Hist. Nat. Araign.,’ 2nd ed.).

List of the Species used.

(The twenty-nine species marked with an asterisk [*] were
also examined in sections.)

et r ap n e u m o n o u s Spiders.

Earn . A V i c n 1 a r i i d a3 .

Sub-fani. A vi c ul arii na) .

*Crypsidromns intermedius, Paraquay.
Harpactira atra, Latr., Cape Town.

Sub-fam. Ctenizime.

Stasimopns unispinosns, Pure., Cape Colony.
Hermacha sp., Cape Town.

Dipnenmonons and Apnenmonons (Caponia)
Spiders.

Earn. Ere si dm.

Eresns sp., Cape Town.

Earn. Sic arii dm.

*Scytodes testudo. Pure., Cape Town.

Earn. Dysderidm.

*Dysdera s])., Berlin.

*Harpactes Hombergi, Scop., Berlin.

*Segestria seuoculata, L., Berlin.

EESPIRATOKY ORGANS IN ARANE^’,.

5

Fam. Caponiidje.

*Caponia spii’alif era. Pure., Cape Colony.

Fain. Drassidae.

*Drassodes (Drassus) inf iiscatus, Westr., Berlin.
D. tessellatus, I’urc., Cape Colony.
*Melanopliora (Prostliesiina) Petiveri, Scop.,
Berlin.

Fam. P a 1 p i m a n i d le .

*Palpimanns sp.. Cape 'I'own.

Fain. Theridiidie.

Latrodectus geometricus, C. K., Cape Town.
*Tlieridion linen turn, Cl., Berlin.

Fam. Argiopidm.

Sub-fam. L i n y p li i i n tu.

*Ijinypliia triangularis. Cl., Berlin.

Sub-fam. Te t r agna t li i n a? .

*Pachygnatha Bisteri, Sund., Berlin.

Sub-fam. Nephilime.

Nepliila sji., Senegal.

Sub-fam. Argiopime.

Argiope clatlirata, C. K., Cape Town.

Fam. Tliomisidm.

*Pliil odrom UK (Artaues) f uscomarginatus, De
C., Berlin.

*P. (Artanes) pallidus, Walck., Berlin.

*Tibellus oblongus, Walck.

Fam . C 1 u b i o n i d £6 .

Palystes s]).. Cape Town.

*Clubiona holosericea, De G., Berlin.

*Zora sp., Berlin.

Fam. Agelenidai.

*A rgy roneta aquutica. Cl., Berlin.

*Textrix lycosina, Sund., Berlin.

*Agelena labyrintliica, Cl., Berlin.

*Tegenaria atrica, C. K., Berlin.

'1'. domestica. Cl., Cape Town.

6

W. F. PUKCELL.

Fani. J’isauridas.

*Pisaura (Ocyale) mirabilis, Cl., Berlin.

*Dolomedes sp., Berlin.

Fam. Ly cosi daj.

*Lycosa (Trochosa) sp., Berlin.

*L. (Pi rata) hygropliila, Thor., Berlin.

*L. (Tarantula) aculeata. Cl., Berlin.

1j. Darliiigi, Poc., Cape Town.

*L. sp., Berlin.

Fain. Salticidai (Attidie).

*Sitticus (Attus) floricola, C. K., Berlin.

S. (Attus) sp., Berlin.

*Marpissa (Marpessa) mucosa. Cl., Berlin.

Biological observations. — Attus floricola fastens its co-
coons on dead branches, etc., on the edges of the lakes in the
Grunewald, a forest near Berlin, and I have found as many
as twenty or thirty cocoons closely packed together in a group
at the N.W. corner of Hundekehle See. The number of eggs
in a cocoon varies normally from about thirty-five to fifty,
and eggs may be found in the cocoons throughout June, July,
and the first half of August.

A number laid in captivity on July 12th hatched (i.e. burst
the egg-shell) on July 28th and 29th, i. e. after sixteen to
seventeen days. At the time of hatching the embryos are still
very imperfectly formed and very much resemble Locy’s
fig. 10, except that the legs, which are curved inwards and
ventrall}', are segmented. The pedipaljis are each provided
at the base with a small conical tooth, which is broader than
high and drawn out at its apex into a tiny brown point.
Shortly before hatching the egg-shell becomes stretched and
raised on the tips of the two teeth, which then split it across
in front of the chelicera.^

' I also found the two teeth in Xysticus, Tegenaria, and Age-
lona, the teeth and the part of the cuticnla on rvliich they stand being
Idack in the two latter genera. These teeth do not ajijjear to have been
pi-evionsly observed, and they have been recorded in Korschelt and

EESPIRATOEY ORGANS IN ARANEA^.

7

After hatching the embryos remain motionless for five to
sis days or even a little longer before the first post-embryonic
moult takes place, after which the young spiders acquire the
use of their limbs. They are still, however, in a very im-
perfect condition, especially as regards the eyes. They remain
in the cocoons until after the second moult, which takes place
sixteen to seventeen days after the first. The young spiders
then emerge in a perfect condition, with fully-developed eyes,
and have also acquired the definite shape of the adult. ^

The entire development, therefore, takes from about thirty-
seven to forty days, less than half of which number is spent
within the e"sr-shell.

Treatment. — The preserving reagent upon which I mostly
relied was a hot concentrated alcoholic solution of corrosive
sublimate, which 1 make use of in the following manner :
A quantity of sublimate is placed in a small, loosely corked,
boiling fiask with sotne alcohol of 70 per cent, and heated
over a flame with constant shaking until the alcohol begins
to boil. Home of the concentrated solution is then poured
into a small tube of about 3 c.cm. capacity, which is imme-
diately corked and suspended by a string in a basin of water
heated to 80°C.“ A few eggs are now dropped in and
removed almost immediately afterwards by means of a thin
glass rod, which is flattened at one end and here bent at
right angles to form a scoop large enough to take out one
egg at a tiine.^ The sublimate will probably be precipitated
during the latter operation, but that does not matter. The
eggs are then placed in G3 per cent, alcohol (70 percent, will

Heider {’92, jj. 588). Later oii I found a similar black tooth on the base
of each pedipalp in the embryo of a Tetrapneunionous spider (Harpac-
tira atra) from Cape Town. •

' Similar phases occur in the development of all other Dipneunionous
spiders which I have examined.

^ A temperature of 70° has much the same effect.

^ It sometimes happens that the egg-shell bursts, in which case the
embi-yo is destroyed by the violent action of the reagent. As a rule,
however, it remains intact and only just sufficient reagent penetrates to
preserve the embryo.

8

\V. F. ITHCELl,.

do as well) lor one lionr, tlie alcoliol Leiiig changed several
times, then for one to two hours in 78 per cent, alcohol, and
finally in 93 per cent, and absolute alcohol. Chlorofoim was
used for embedding in paraffin and the sections were stained
on the slide in alum carmin, wdiich I found the most suitable
for them.

. A number of embryos were also preserved in a hot aqueous
solution of corrosive sublimate (concentrated at the ordinary
temperature) for the purpose of control. I did not, however,
require to make much use of these embryos, Avhich could not
compare with the others in point of preservation.

The great disadvantage attached to the use of plain hot
water or hot aqueous solutions, which ai'e the methods
hitherto usually employed in spider embryology, lies in the
cii’cumstance that the embryonic tissues take the appearance
of a syncytium when so preserved. It accordingly becomes
difficult to distinguish between two or more e})ithelia in con-
tact, the several layers of nuclei being then usually the only
feature by which the nature of the tissue can be guessed at
rather than recognised.

I devised the hot alcoholic sublimate method desenbed
above only after repeated experiments. It has the merit of
rendering the contours of the cells much more distinct, so that
not only the boundary line between ttvo epithelia in contact,
but generally also the boundary lines between adjacent cells
within an epithelium becomes recognisable.

In the sections these cell-contours may appear iu the form
of hue dark lines or they may be indicated meiely by a
difference in the staining between the protoplasm of adjoining-
cells or by the presence of paler lii.es between the cells, as if
these had been slightly drawn apart.

As the appearance of the protojilasm was of minor imjiort-
auce, but the contours of the ejiithelia and their cells of the
greatest importance iu my investigations, 1 have for con-
venience represented tlie contours in the figures by dark lines,
by which, of course, I do not intend to imply that regular cell-
walls are present.

KKSriK’ATOKY OKGANS ]N AlfANK/E.

9

I found it quite impossible to obtuin uu accurate idea of
the rudimentary luug-books and trachea} iii the embryos,
except by means of reconstructions, of which extensive use
was made. For the complicated lung-books a large number of
the ordinary reconstructions wiih wax tablets were made
(thickness of sections 5‘82 q, of wax tablets 2 mm.; magnified
343’7 diameters), but for the simpler trachea} the following-
method was employed:

A sheet of transparent paper is placed over another of white
paper ruled with a series of parallel lines 2 mm. apart. 'J'he
width of the organ to be reconstructed maguified the required
number of times (343‘7 times for sections 5-82 /i thick), is
measured with a])air of compasses in each section and marked
off on the parallel lines, each of which represents a section.
When all the sections have been marked in this way on the
transparent paper the outlines of the organ will be obtained
in their coiTect proportions. This method is much quicker
than the other and very suitable for reconstruction in outline
from transverse sections of any bilaterally symmetrical organ
of simple form, such as the embryonic trachete in the later
stages (figs. 28 and 29). By drawing a line down the middle
of the paper at right angles to the parallel lines to re])re-
sent the median line of the body, and marking each transverse
section symmetrically on each side of this line, the symmetiy
of the reconstructed organ will be preserved.

If. (iKNEKAL OltlEXTATiON.

Lung-books. — A typical well-developed lung-book of a
Dipueumonous spider has the following }>arts (figs. 20
and 21) ;

(1) A more or less transverse spiracle (•'■-p.) or stigma
placed laterally at the junction of the ventral and lateral
surfaces of the second abdominal segment along its hind
margin (text-fig. 1).

(2) A short flattened tube leading forwards from the
spiracle into the body in a slightly upward and medial direc-

10

\V. F. PUKCEU,.

tion, forming a stalk or pedicel {iie.d.) to the whole lung-
complex. This opens into —

(3) An elongated-lanceolate hollow hand^ the pulmonary

sac (ante-chamber or vestibule, a.), 'which runs

from just in front of the medial angle of the spiracle at fii’st
in a dorso-lateial direction, but becomes procurved at a
greater or less distance beyond the lateral angle of the
spiracle to form the horn (Schneider, li.) and terminates in a
short, blind, apical pouch [aj).).

(4) A series of long, flattened, hollow pouches (saccules,
6'.), which are triangular in shape, like a flattened butterfly-
net, generally horizontal, and placed one over the other in a

Text-fig. 1.

Attns floricola. Ventral surface of abdomen. Ih. Pulmonary
operculum, iniltn. xp. Pulmonary spiracle, tr. xp. Tracheal
spiracle. Magnified 13.

slightly imbricating manner (each being slightly more lateral
than the one below it), like the leaves of an open book. The
saccules, being invaginations of the anterior wall of the ante-
chamber, communicate with its lumen by their open posterior
ends, which form a series of parallel slits, like an oven-grate
(Bertkau), extending obliquely across the entire anterior
surface of the ante-chamber, including the corresponding
ventral surface of the procurved horn, being absent only from
the small apical pouch of the latter.^

' In some text-books, e. g. Korschelt and Heider ('92. p. 605, fig. 382)

EESPIBATORY ORGANS IN ARANE/E.

11

All these parts, being hollow, contain air in direct com-
munication with the external atmosphere. The partition walls
between the air-spaces of adjacent saccules, I shall term the
‘‘septa.”i The dorsal side of each septum is studded Avith
numerous, simple, blunted spines, which keep the lumens of
the saccules open, while the walls of the ante-chamber
(including its fenestrated anterior wall and the apical pouch
of the horn), are covered with pecnliar hooped spines
(spines with anastomising apical branches). The pedicel is
for the most part unspined.

The two spiracles are generally united by a transverse
fold, the epigastric or interpulmonary fold {interp.
fill.), which also connects the two pedicels and the extreme
medial corners of the two ante-chambers (see text-fig. 1).
The lumens of the latter at the same time communicate by
the interpulmonary canal of communication (can.), or
passage Avitli hooped spines in the upper edge of the fold.

Further remarks on the lung-books of the adnlt are given
on p. 41, and an historical account of the literatnre will be
found at the end of the paper.

Tracheae. — The usual form of tracheae in a Uipneuinonous
spider has the folloAving parts (figs. 21, 25 and 31) :

(1) A median, transverse, ventral spiracle (sj).), placed
on the hind mai-gin of the third abdominal segment usually
just before the spinners (text-fig. 1).

tlie procAirvecl honi is wrongly represented as having no saccules opening
into it.

‘ In order to avoid ambiguity I have substituted the terms "sac-
cules” and "septa” in place of the old terms "leaves” and
"lamellai.” The older writers almost invariably meant to indicate the
saccules when they used the term "leaves” (feuillets, Blatter),
but since about 1881 the term has generally been employed for the septa,
like the term " lamellae.” Neither term, however, has at present tniy
definitely recognised use. Thus "lamelles” signifies the septa with
MacLeod ("84), but only one of the layers of a septum Avith Berteaux
('89), Avhose term for a Avhole septum is “lame,” Avhile "feuillet”
signifies a septum Avith SchiuikeAvitsch ('84) and Plateau ('86), Imt a
saccule Avith Scl)neider ('92).

12

W. F. PUKCEM,,

(2) A short, flattened cdiamber (vestibule, rest.), leading
lorwards and upwards from the spiracle into tlie body and
giving off at its anterior or deepest part —

(3) A pair of medial [vi.tr.) and a pair of lateral
tracheal trunks [l.tr.), which may again give rise to
tracheal branches (tv., fig. 21), the finest of these being the
tracheal tubules [tr.tnh., fig. 31). The trunks and
branches are lined with anastomosing spines (more rarely
with spiral threads oidy), but the fine tubules have only
spiral threads.

'I'he anterior or deepest part of the cavity of the vestibule
is always widened to form a transverse canal of communi-
cation [can.) or passage with hooped spines, connecting the
cavities of the tracheal trunks. The remaining or smooth
portion of the vestibule forms a stalk or pedicel [yed.) to
the whole tracheal system, and is supported cu each side by
chitinous thickening or rod [rd.).

111. lllSTOKICAL (DeVELOPMKNt).

Development of the lung-books in Arachnida. — Metschnikoff
(’71) gives an account of the development of the lung-books
in scorpions, and observes that they arise as ectodermal
invaginations just behind the four posterior pairs of abdo-
minal ajipendages, which latter subsequently atrophy. Towards
the end of the embryonic period the folds in the pulmonary
sacs appear.

Salensky (’71) was the first to study their development in
Aianeas, and believed that the lung-books were formed
by the invagination of the abdominal appendages (teste
Jaworowski, ’94, p. 55).

Bertkau (’72) showed that in the young spider, after the
completion of the embryonic period, the lung-books continue
to develop, new leaves being added at the growing dorso-
lateral end, each new leaflet arising next to the one previously
formed.

Locy (’86) gives a detailed description of the later stages

RESPIRATORY ORGANS IN ARANE.E.

13

in a spider (Agelena ntevia), and lie is the first to give an
account of the transformation of the embryonic epithelial
foldings into the definite pulmonary septa (lamellee) with their
chitinons coverings. According to him the lung-books arise
as a pair of invaginations late in the period of the reversion,
but he makes no mention of their connection witli appeu-
d a.<res.

Bruce (’86a, ’86b, ’87) is of opinion that the pulmonary
folds in spiders are formed on the anterior surface of the first
abdominal appendage, which subsequently becomes involuted,
so that its anterior surface with the folds now faces the pos-
terior end. Probably two abdominal appendages are invagi-
na ted for each lung-book.

Schimkewitsch f’87; also in ’86a and’86b, teste Jaworowski,
’94) states that the lung-book arises as an invagination of the
ectoderm and forms a true trachea, consisting of a main trunk
divided into five branches, in the embryo of Lycosa saccata
just before hatching. Recently, hovvevei', Schimkewitsch
(:06, pp. 45, 46, footnote) has withdrawn this interpretation.

Kowalevsky and Schulgin (’86) merely note that the pul-
monary sacs in the scorpion (.\ndroctonus ornatus) arise
as simple invaginations into a space containing plenty of
blood.

IMorin (’87) found that 'the lung-books in the spider
(Theridion) arise from a pair of ectodermal invaginations
at the base of the first pair of abdominal appendages, which
themselves become the lung-opercula. In his later paper
(’88) he appears to have given more details of the formation
of the lainelhe, of Avhich those nearest the operculum are
furthest developed (teste Jaworowski ’94, pp. 57 and 58).

Laurie (’90) states that in the scorpion (hluscorpius
italicns) the four last pairs of abdominal appendages are
pushed in on their posterior part, so as to form shallow, cup-
like cavities, which later on are divided up by lamellm growing
down from their upper ends (pp. 125 and 127). A later stage
with lamellae is also described (p. 129). In a later paper
(’92) he deals with the development in Scorpio fulvipes.

14

W. F. PURCELL.

Kishinouye (^90') confirms the statement regarding the lung-
books and the opercula contained in Morin’s earlier paper
(’87), and adds that that wall of the invagination which faces
the distal end of the appendage is much thickened, filling the
interior of the appendage, the cells becoming after a while
arranged in parallel rows to form the septa. He examined
Lycosa, Agelena, Theridion, Epeira, Dolomedes and
P h o 1 c u s .

Simmons’ (’94) paper is the most important that has yet
appeared on the development of the spider’s lung-books, and
was based on embryos of Agelena nmvia and Theridion
tepidariorum. He confirms Morin’s and Kishinouye’s state-
ments regarding the formation of the pulmonary invagina-
tion, and was the first to describe and figure an early stage
of the formation of the pulmonary septa (lamellae), which he
states arise as infoldings upon the posterior surface of the
abdominal appendage in the same manner as described by
Kingsley for the gills of Limulus.

Javvorowski (’93 and ’94) describes the presence in a spider
embryo (Trochosa) of embryonic tracheae, which ultimately
becomes rudimentary, excepting the portion adjoining the
spiracle. The wall of this portion is thi-own into folds and
persists as the lung-books. The author thus totally differs
iu some most important points from all his predecessors.
’I’liese tracheie arise from invaginations under the abdominal
appendages, the latter becoming the opercula (’95, p. 43).
Jaworowski also gives a valuable account of the formation of
the definite pulmonai’y septa out of the folded embryonic
epithelia.

According to Laurie (’94) the embryonic abdominal
appendages are not paired in the Pedipalpi, but stretch
right across the abdomen, and in Phrynus the lung-books
evidently arise as foldings of the posterior wall of an appen-
dage.

Brauer (’95) confirms Metschnikoff’s and Laurie’s observa-
tions on the earlier stages of the pulmonary invaginations at
the base of the four posterior pairs of abdominal appendages

EESPLKATOKY ORGANS IN AEANEiE.

15

in scorpions, and giv^es the best account of the early stages
of the pulmonary folds (in Euscorpins car pat hie us).

In my own paper (’95) the appearance of the earliest pul-
monary folds on the free posterior side of the first pair of
abdominal appendages in Aranem is described.

Sophie Pereyaslawzewa (:01) investigated the earliest
appeai-ance of the lung-books in Phrynidmin Phryniscus
b a c i 1 1 i f e r and the later stages in Damon m e d i u s. Accord-
ing to her the lung-books are formed from the third and
fourth abdominal appendages, which belong to the third and
fourth abdominal segments (p. 194). The outer integument,
the cuticula of which is regularly wrinkled (fig. 61), is deeply
infolded into the body behind the third and fourth appen-
dages to form the lung-sacs, the grooves in the invaginated
wrinkled surface deepening to form the saccules, while the
ridges become the septa (pp. 248-252). The embryonic
septa are also described (p. 262) and figured (fig. 69).

Gough (:02) states that the lung-books in an embiwo of a
Pedipalp (Phrynid) belong to the first and second abdominal
appendages. The author gives no further account of the
development of the lung-books, but mei*ely states that it does
not differ from that in other Ai’achnids (p. 616).

Schimkewitsch (:03, ;06) gives a more detailed account of
the development of the lung-books in Thelyphonns
ca lid at us. According to him the lung-books are formed
from pulmonary sacs or invaginations at the base of appen-
dages, which are placed on the hind margins of the second
and third abdominal somites. The lung-leaves arise as folds
in the lower wall of this sac, and later on the leaves, which
were formed in the sac, come to lie outside of it on the
posterior side of the appendages so that the saccules then
open to the outside instead of into the sac (fig. 46). Several
sections of later stages of the lamellae are figured.

Sophie l^ereyaslawzewa (:07), in a posthumous memoir,
describes and figures some interesting stag-es in the develop-
ment of the lung-books of a scorpion (Androc tonus orna-
tus) from the material left by Kowalevsky and Schulgiii.

10

W. F. PURCELL.

According to her tlie invagination -which forms the pulmonary
sac is situated ou the anterior edge of the lateral part of
the base of an abdominal appendage and is apparently uncon-
nected with the latter (p. 177).

Development of the tracheae in Araneae. — Schimkewitsch (’87)
states that the tracheae in Lycosa saccata arise by invasri-
nations of the ectoderm. In his Russian paper (’86a) he
gives a figure of a. developing trachea [ect, fig. 29a), without,
however, recognising it as such.

Kishinouye (’90) observed an ectodermal invagination in
various spiders in the basal part of the second abdominal
appendage on the interior side. This invagination forms a
deep tube at the time of hatching and the author calls it an
“ abortive trachea.”

Simmons (’94) found the same invagination in Agelena
nmvia and Theridion tepidarior um, and in addition
what he considers to be aborted lung-leaves.

Finally, in my abstract (’95) of the present paper the origin
of the greater part of the tracheae in Attus floricola from
ectodermal tendons is stated in outline.

TV. Thtk Provisional Abdominal Appendages in the Embryo
OK Attus Floricola.

The description begins with the stage^ immediately
preceding the apnearance of the pulmonar}’- folds
(stage 1, F)t. 1). 'Phe embryonic band has attained its
o-reatest length, and the process known as the reversion
is about to commence. A sagittal section (PI. 1, fig. 4)
through the abdominal region shows eight abdominal
segments with coelomic sacs. The first abdominal (seventh
post-oral) segment bears no appendages in this species, but
the following four (eighth to eleventh post-oral) are each
provided in their posterior region with a low, flat-topped,

' CoiTespondiny to the stage in Korsclielt and Heider. p. 581. fig.
and to Locy’s PL ii, fig. 8, and Balfour's fig. fi. A list of the v.irious
stages and of the figures referring to each is given in the explanation of
tlie plates.

RESPIRATORY ORGANS IN AUANE.E.

17

provisional appendage {ah. app. 1-4) in successive stages of
growth, that of the eleventh segment being the smallest and
that of the eighth the largest.

The segments are marked off from each other by distinct
transverse grooves, which are shallow, except immediately
behind the appendages, where they are considerably deepened
(f/r.), and where the ectoderm forms a distinct post-appen-
dicular fold, projecting at right angles, or nearly so, to
the general surface into the body. The posterior wall of this
fold is comparatively thin, like the adjacent epithelium of
the following segment, but the anterior wall is much thicker,
being, in fact, a direct continuation and a part of the posterior
wall of an appendage, as I shall presently show.

A similar post-appendicular infolding (as distinct from the
pulmonary .sac to be described later) appears to be also found
in Limulus (Kingsley, ’85). In the older spider-embryo
those of the posterior pairs of appendages serve as places
of attachment for the ventral longitudinal muscles of the
abdomen.

The deep infoldings behind the first pair of abdominal
appendages extend from the medial end of the hind margin
of each a])pendage nearly, but not quite, up to the extreme
lateral end, and, moreover, the lateral part of the infolding
{t/r., fig. 7a) is always .slightl}', but distinctly, deeper than its
medial part hg. 7). I’liese two figures represent the

appendages just before the earliest appearance of the rudi-
ments of the lung-books.

V. The Deveeoi'Siext ok the Lung-Kooks.

Stage with two pulmonary furrows (stage 2, SL 2). — 'I’he
appendages of the pulmonary or eighth post-oral segment
undergo considerable changes in passing from the stage just
described to the next one, which I shall term the “stage with
two pulmonary furrows.” Fig. 1 is a transverse section of
this stage, and shows that the appendages are still near
together, although the reversion has commenced. This stage

VOL. 54, PART 1. — NEW SERIES. 2

18

W. P. PURCELL.

follows so quickly upon the last that it is at first very puzzling
to make out the changes accurately, hut with the aid of
numerous reconstructions in wax 1 have been able to ascer-
tain the more important phases with certainty.

Fig. 14 is a sketch made from such a reconstruction, and
represents the typical appearance of the right appendage
seen somewhat from behind. Its distal surface is flat and
often, although not always, distinctly transverse. Measured
at the base, however, the breadth of the appendage is about
equal to the antero-posterior diameter, and remains in this
relation throughout the later stages. Seen fi’om the distal
surface the appendage appears distinctly four-sided, with its
posterior side placed transversely to the embx’yonic band.

Fig. 8 is another reconstruction made from a series of
longitudinal sections cut parallel to the principal axis {pr. ax.,
fig. 1) of the appendage, and a number of sections from this
series are given in figs. 8a-8h, the positions of the sections
being indicated by the vertical lines in fig. 8.

'Phe first point to be noticed is the subsidence of the epi-
thelium (ejx., figs. 8a-8g) lying immediately behind the first
abdominal appendage and forming the posterior lip of the
post-appendicular groove^ (yr., in stage 1, fig. 7). The two
lips of the latter thus become drawn completely apart along
its whole length, so as almost to obliterate the groove as such
(except at a single place to be mentioned presently) and lay
free the whole posterior side of the appendage. In its median
Imlf the former bottom of the groove is now indicated only
by a shallow furrow (yr., figs. 8a-8d), which at the same time
marks what in the previous stage (fig. 7) was the base of the
posterior side of the appendage. This shallow fui'row behind
which the subsidence was greatest is moi-e or less curved
owing to a shifting backwards of the tissue in which it lies
(yr., fig. 14), so that the posterior side of the appendage
comes to slant in its xnedial part at base (.v/., figs. 8a-8d,

' A corresponding subsidence also takes place anteriorly to the fii-st
ji])pendage, causing the obliteration of the groove between the seventh
and eiglith segments.

RESPIRATORY ORaAXS IX ARAXE.E.

19

and 14) much raora than was the case ia the previous stage,
where the groove was straight and transverse. The angle of
the slanting surface varies, the latter being in some embryos
nearly perpendicular, in others nearly parallel to the adjacent
body surface and in the latter case the curved furrow

may entirely disappear. The above will become clear by com-
paring figs. 8b and 9 of this stage with the corresponding
section (fig. 7) of the previous stage.

In the second place we notice a little pocket-like cavity
(palm. .s\, figs. 8 and 14) extending from the middle of the
base behind in a latei’al direction for about one third of the
breadth of the appendage. This cavity, which we may term
the pulmonary sac, is practically all that remains of the
once extensive post-appendicular groove, and is to be con-
sidered as a portion of the latter which had become especially
deepened and so escaped the obliteration which befel the rest
of the groove — for a subsidence has also taken place in the
tissue immediately behind the pocket. (Compare fig. 8g
with the corresponding section, tig. 7a, of the previous
stage.)

The ])ulmonary sac was fii*st described and figured by
.Metschnikoff (’71) for the scorpion and was found by most
subsequent investigators, but generally in a later stage of
development.

'rhe cells which form the wall of the sac undergo from
now on repeated division (fig. 8o), causing the sac to grow
rapidly, at first in a forward direction in the form of an in-
pushing under the appendage, but later on in a latero-dorsal
or dorsal direction (fig. 10). The anterior wall of the pul-
monary sac yields the cell-material for all the lung saccules,
except the first two, whose appearance forms the third and
most important point to be noticed in this stage.

On the medial half of the posterior side of the appendage
there appear two parallel furrows of varying length (/. 1,/. 2,
figs. 8 and 14). These are the first beginnings of the two
oldest saccules of the lung-book. They are never transverse
but always incline to the longitudinal axis of the appendage

20

W. F. PURCELT,.

fit varying ang-les. The first or medial furrow [ f. 1) is
always much the deeper and extends from near the medio-
posterior angle of the base of the appendage in a latero-
distal direction. As a rule when the posterior face of the
appendage is strongly inclined, the furrow takes a more
transverse direction and does not then reach the distal
surface (as in figs. 8 and 14), but when the posterior face is
less inclined, the furrow takes a direction more nearl}-
parallel to the axis of the appendage, extends right up to
tlie distal surface of the latter, and comes to be situated on
its medio-posterior corner (fig. 10). In such cases, in fact, it
is sometimes more on the medial than on the posterior side
of the appendage. The second furrow (/. 2) appears
almost simultaneously with the first, and is situated between
the latter and the base of the appendag-e, so that its medial
end terminates on the proximal side of the lateral part of the
first furrow. It never extends right to the base nor to the
distal surface of the appendage, and if produced medially
would run proximally to the first furrow.

Compared with the preceding stage the medial half of the
appendage has developed considerably and is sharply set off
from the body surface. Further, in its lateral part (fig. 8g)
the anterior side has become much more inclined than in the
preceding stage (fig. 7a), so as to be parallel to the slanting
anterior wall of the pulmonary sac. In longitudinal sections
through this part (fig. 8g) the appendage has the false
appearance of being directed backwards, and this becomes
still more marked in later stages (as for instance figs. 12 and
IGc). That this appearance is deceptive and merely due to
the pulmonary sac will be readily seen if it be remembered
that fig. 8g is a section lying between the sections fig. 8f and
fig. 8h, and fig. 16c a section between fig. 16a and fig. 16n.
The main axis of the appendage remains in all cases at right
angles to the body.

Stages with three or more pulmonary furrows (stages 8 to 5).
— I’lie third furrow ( /. 8, fig. 16) appears at the middle of
the base of the posterior side of the appendage. It is

liESPIKATOKY ORGANS IN ARANE.®.

21

parallel to the others and lies partly inside and partly outside
of the pulmonary sac. Its medial part lies proximally to the
lateral end of the second furrow, and, iu some cases at least,
is continuous with the lateral end of the curved furrow
mentioned above [gr,, fig. 14), which limits the appendage
posteriorly. The fourth fui-row (/. 4, fig. IG) and all sub-
sequent ones lie wholly within the pulmonary sac and appear
successively as oblique grooves in its anterior wall, all more
or less parallel to those already formed and with the medial
ends of each lying on the proximal side of the lateral part of
the previously formed furrow.

After the appearance of the first two furrows the appen-
da ges rapidly move from a ventral to a lateral position owing
to the reversion of the germinal band, and it is necessary to
bear in mind that we must substitute the terms “dorsal” and
“ ventral” for “lateral” and “medial” after the lateral posi-
tion has been I'eached.^

Tigs. 1-3 will make this clear. Fig. 1 is the position at the
end of the 2-furrow stage; fig. 2 that at the end of the 3-
furrow stage, and fig. 3 re])resents the position from the end
of the 4-furrow stage, and here the appendages remain till
near the close of the embryonic period. 'I'he whole segment
which bears the appendage participates in this wandering,
and the position of the appendage relatively to the adjacent
surface is, of course, not affected by the movement.

It will be observed that the youngest furrow (fig. 3) is the
most dorsal one, and, if produced, would lie on the proximal
side of all the older ones.

'J'he pulmonary sac increases hand in hand with the forma-
tion of new furrows, almost filling out the dorsal part of the
hollow of the appendage. At the 5-furrow stage its blind
end grows as a tube with a considerable lumen iu an upward
or dorsal direction, raising up the outer epithelium as it
pushes its way underneath (see hgs. IG, IGd, IGe).

' For tlie sake of imiforiuity and in order to facilitate comparison
between them, the sections of the earlier and later stages of the appen-
dages have been drawn in the same positions throughout.

22

W. F. I’UHCELL.

Formation of the spiracle. — After the appendage has attained
its greatest elevation (genei’ally late in the 3-furrow stage) the
whole region between the three oldest furrows begins to sink
below the level of the appendicular posterior surface by a for-
ward moveineiitj causing* it to be over-topped by the distal edge
of the appendage (fig. 16a). Tliis sinking movement^ which
must not be confounded with the formation of the pulmonary
folds described further oip commences next to the pulmonary
sac, and the latter thus comes to include first the third furroAv
(fig. 16b), then the second, and finally the first, while the
common opening becomes the spiracle fig- 13^ which com-
pare with figs. 13a and 13b of the same series).

Meanwhile that portion of the body epithelium which lies
immediately in front of each of the four appendages in a row
becomes absorbed into the anterior side of the appendage
(compare figs. 4, 5, and 6), so that the four appendages appear
closer together, while the original opening of the pulmonary
sac comes to lie at the bottom of the groove so formed between
the first and second appendages.

The lateral (afterwards dorsal) end of the spiracle is the
first to be formed, and is already clearly elefined immediately
after the appearance of the pulmonary sac (fig. 14). The
jirogressive development of the lateral part of the spiracle
may be followed in figs. 8g, 12, and 16c. In the latter embryo
the surface posterior (fig. 16c) and dorsal (fig. 16n) to the
lateral end of the spiracle is almost on a level with the distal
surface of the first appendage. The medial (later ventral)
region of the spiracle remains open and undefined for a much
louger time.

Sinking of the appendage. — This is a very simple process
and begins about the 5-6-furrow stage {St. 5). The anterior
and ventral sides become more slanting, so as to pass, like the
dorsal side, more and more gradually over into the adjacent
body surface, while the appendage itself decreases in eleva-
tion and sinks graduall}' into the body, until tinally only a
slight convexit}' in front of the spiracle marks its former
])osition. (Compare fig. 13b, with five pulmonary furrows.

EESPIRATORY ORGANS IN ARANE.p].

23

witli the corresponding section, fig. 17, of a nincli later
stage.)

Formation of the pulmonary saccules. — Next to their position
on the posterior side of the appendage, the precise manner in
which the saccules begin to form is the point of greatest
interest and importance, when considered with regard to their
possible direct origin from gill-lamellm. 1 have been
successful in obtaining a number of excellent sections through
the region in question, showing the cell-boundaries with
perfect distinctness. The position of these and of the nuclei
of each individual cell have been drawn with the aid of a
drawing apparatus in the sections figured in the plates, which
are in this respect exact reproductions of the original sections
(see p. 8).

The three figures 7, 0, and 8c represent longitudinal sections,
cut parallel to the axis of the appendage, and, as nearly as
possible, through the same region of the latter, in each case
indicated by the line marked {fig. 8c) in fig. 8. All these
sections are through the region in which the first fuiTOW
appears, and represent three consecutive phases following
close upon one another.

In the youngest stage (fig. 7) no trace of the furrow is
apparent, and the appendicular epithelium is composed entirely
of elongated cylindrical cells.

In the next stage (fig. 9), however, the distal wall of the
oldest pulmonary saccule has appeared, and is seen still better
developed in fig. 8c. The formation takes place as follows :
A cleft {cl. 1) in the epithelium appears on its internal surface
at the junction of the posterior and distal sides of the
a])pendage, while a similar cleft (the first pulmonary furrow,
/. 1) is formed almost simultaneously on the outer surface.
'I’he cylindrical cells between these two clefts immediately
begin to shorten to about one-half of their former length and
rearrange themselves as a one-layered epithelium, whose basal
and free surfaces are now represented by the internal and
external clefts respectively.

'I’he proximal surface of the first pulmonary' furrow is still

24

W. F. PURCELL.

hounded hy the original cylindrical cells (figs. 8b and 8c).
In these two figures we see, however, the commencement of a
second internal cleft {cl. 2) and a second external furrow,
the latter being the second pulmonary fur row (/. 2). In a
later stage the cells betAveen the second internal cleft and
the two external furrows are seen in the process of shorten-
ing' to one half of their former length in order to re-arrange
themselves to epithelia having this cleft and the two furrows
for their basal and free surfaces respectively (figs. 10 and
11).

d'he walls of the oldest saccule, embracing the first pul-
monary furrow between them, and the distant Avail of the
second saccule, are, therefore, uoav present. In a similar
manner the proximal wall of the second saccule and the Avails
of all subsequent saccules are formed (fig. 15).

The external pulmonary furroAvs are ahvays provided Avith
a distinct lumen, cutting deep into the sides of the appendage
and sac, and they have been thus figured by Simmons, who
Avas, I believe, the first to observe them. The internal clefts,
on the contrary, have no real lumen AvliateA^er, and are
indicated on the visceral surface of the epithelium by slight
grooves only.

The process described above may be summarised as follows :
The cells of the epithelial region Avhich contains the
pulmonary furroAvs shorten and re-arrange them-
selves in the form of a folded epithelium, Avhich has
one half the thickness of the original epithelium
and occupies about the same total A’^oliime.

Ily re])eated cell-division the folded e[)ithelium expands
in such a manner that all those folds Avhich are directed
iiiAvards and Avhich contain a pulmonary furroAv between
their Avails groAv into and ultimately fill out the cavity of the
appendage (figs. 10 and 16a). Each holloAv pouch thus
pn'oduced gives rise, of course, ultimately to a hollow air-
containing saccule, Avhile the outAvardly directed folds com-
prised betAveen tAVO pulmonary furroAvs ultimately form the
septa betAveen the air-compartments of the lung-book. The

RESPIliATORY ORGANS IN ARANE^,

25

oldest saccule is the fii'st to grow into the interior, while the
others follow in turn in the order in which they were formed.

Simmons (^94) is, so far as I am aware, the only author who
has described and figured the earlier stages of the formation
of the saccules in spiders. He gives two figures of sagittal
sections from embryos of the same age, one (fig. 6) showing-
five and the other (fig. 5) two pulmonary furrows. The
position of these two furrows in the latter figure shows that
they are not the two oldest, the others having apparently
been missed by the section, which is probably of about the
same stage as his fig. 6. Simmons’ account is as follows:
The outer wall of the pocket “has its ectoderm thrown into
folds,” the nuclei in this ectoderm “ being rather iri-egnlarly
arranged, the pulmonai-y ingrowths [i.e. the furrows] forc-
ing their way between them.” dlie more distal gill-lamellm
(by which the author means the se])ta) are the oldest, as in
Ijimulus (p. 217). Simmons’ paper is dealt with again
further ou (p. db).

Comparison with the gill-books of Limulus. — It would be
profitable here to institute a comparison with the gill-books
of L i in u 1 u s.

According to the description and the figures of Kingsley
(’85), the gill-leaves of the American Limulus arise as out-
growing folds of the epithelium of the posterior side of the
apjiendages, their formation being accompanied by a slight
in-tucking of the epithelium between them, and taking place
in the same order as the pulmonary saccules in spiders. The
epithelial walls of each outwardly directed fold are, however,
not in contact along their ba.sal surfaces, and have apparently
not been suddenly reduced in thickness, thus differing in
these two points from the rudimentary lung-books.

In the Japanese Limulus the process appears to be some-
what different. According to Kishinouye (’91), the proximal
jiortion of the appendage is much thicker than the distal
jiortion and is ])rovided with many transverse furrows or
invaginations, the tissue between two furrows giving rise to a
gill-lamella. At any rate both forms agree in one main

26

AV. F. I’UKCELL.

point, namely, that the out-growths or out-foldings are
accompanied by an invagination of the ectoderm between
them in the earliest stage.

Now, in the case of the rudimentary lung-books in spiders,
as summarised on pp. 17-20, it is evident that the pulmonary
folds cannot be considered as due to simple ont-foldings or
in-foldings of an epithelium whose thickness was that of the
walls of the folds, as is the case in the American Lira ulus
at least. On the contrary, they arise by a peculiar process,
which results in the transformation of a very thick but even
epithelium into a folded one of one half the thickness, but
occupying the same volume, and unaccompanied, therefore,
by any out-growth or in-growth at first.

1 am of opinion that these two modes of forming a folded
epithelium are not fundamentally different, for the one may
be readily derived from the other. On the contrary, I believe
that the method which obtains in the spider is merely an
abbreviation of some such process as occurs in the American
Limulus, being the most convenient one for rapidly throw-
ing a limited area of a very thick epithelium into folds, for
this could not easily be done by ordiuaiw folding, as the
breadth of the area in question is only equal to the thickness
of the epithelium itself. Which of the two methods was the
original depends, of course, on the thickness of the appen-
dicular epithelium in the common ancestor, and is a question
of but secondary importance. The Japanese form, according
to the description of Kishiuouye, appears to bear some
resemblance to the spider in the origin of the respiratory
lamella}.

'The result of the folding in Limulus and the spider are at
first practically the same in each case, namely, an undulating
folded epithelium, and it is only in the subsequent growth of
the folds that a real difference between the two cases becomes
apparent. For in each the epithelial cells Tiiultiply by
division in such a manner that the walls of the folds expand
and grow, in the case of the spider, into the interior of the
appendage, but outwards and away from the latter in

liESriKATOKY OKGANS IN AUANE/E.

27

Li m ulus. We should have iio difficulty in imagiuing a case
in which the cells divided so as to cause the folds to expand
simultaneously in both directions, and the result would be a
structure intermediate between the gill and the lung-book.

The foregoing paragraphs lead up naturally to the simple
and ingenious hypothesis first put forward by Kingsley (’85)
to explain the derivation of the lung-books from gdl-books
(see Kingsley’s explanatory figs. 18-20). He simply assumes
that simultaneously with the sinking of the whole organ the
inwardly directed folds of the gill-books became exaggerated,
while those directed outwai'ds correspondingly decreased. In
this way an intermediate type of respiratory organ would
first be obtained, representing the condition in tlie animal
when it was leaving the water and seeking a terrestrial life.
Finall}-, the lung-book type would be reached by the com-
plete suppression of any tendency of the folds to grow
outwards.

Now, from a morphological point of view there should be
no difficulty in accepting this hypothesis. The passage from
a gill-lamella with three free outer edges to a lung-septum
with only one such edge is perfectly simple and easy to
imagine. It now really constitutes the only assumption not
directly proved ontogenetically which we have to make in
deriving the Arachnid lung-book from a Limuliue gill-book.
For the two remaining conditions necessary for such an
origin, namely, the appearance of the oldest septa on the free
posterior side of the appendage and the subsequent subsi-
dence of the latter, are observed embryological facts. To
return to the first point, the ontogeny, although it does not
exactly prove it, furnishes us, nevertheless, with some evi-
dence which tends to show that the folds were originally
designed to grow outwards and not inwards. For, so far as
1 could make out, the two walls of the most distal se2)tum or
outwardly directed fold are formed simultaneously, and
followed later by the simultaneous apjiearance of the two
walls of a second fold also dii-ected outwards, and so on (see
fig. 11). It cannot be denied that each such fold, on its first

28

AV. F. rUKCELL.

iippearaiice, creates the impression that it \va^ designed and
is about to grow outwards, aud one is perhaps justified in
asking why, if the saccules were originally derived from
tracliea-like invaginations, the two walls of a saccule do
not appear simultaneously as we should expect from a fold
originally designed to grow inwards ? I do not, however,
wish to attach too much importance to this point, as it is very
difficult to ascertain with certainty, and would not even then
constitute a clear proof either way.

Passing to the physiological side of the question, one
benefit derived from the sinking of the gill-leaves into the
appendage and of the latter into the body would, of course,
as Kingsley says, be protection from the increased wear and
tear incidental to terrestrial motion. I’he delicate gill-leaves
wdth the three unattached edges would be very liable to
injury when deprived of liquid support, while a lung-septum,
having only one unattached edge, is perfectly secure. At
first, no doubt, the gill-leaves would be very sensitive to
evaporation, and the cavities between their basal portions in
the intermediate stage (fig. 11) in Kingsley, '85) may have
formed convenient reservoirs for retaining water to moisten
the respiratory surfaces during terrestrial excursions.

Various other theories have been suggested by different
authors (Milne-Edwards, ’72, p. oG ; Kay Lankester, ’81, ’85a
and ’85b; iMacLeod, ’82 and ’84; Jjaurie, ’92 and ’93) to explain
how gill-books like those of Limulus, may have been con-
verted into lung-books, but none of them correspond exactly
to the embrvological facts, so I shall not consider them
further in this pa]>er.

Later development of the pulmonary saccules. — 1 resume the
descripition at the 0-6-iun’ow stage represented in figs.
18-1 8b and 1G-1Gk. 'I'lie interior of the appendage has
become nearly filled out by the ingi-owing saccules, which
push befoi'e them the intra-appendicular part of the coelom
and ultimately occiqty its place. They continue to grow
till the anterior side of the aj)])endage is reached. The oldest
saccules are still the longest, but are exceeded in breadth by

liESPIKATOKY ORGANS IN ARANE.R.

29

the younger ones (fig. 13b) — so much so, indeed, that in the
dorsal region the latter project for at least half their mass
into the body cavity, while the oldest saccules are still
entirely contained within the appendage (a condition still
apparent at the time of hatching, fig. 17).

The plane of each saccule is still an inclined one, slanting
upwards anteriorly, owing to the presence of the genital duct
iu the now ventral (originally medial) portion of the appen-
dage. When, in later stag’es, the duct has migrated else-
where, the saccules come to lie horizontally and parallel to the
ventral side of the appendage (figs. 17 and 18). A slight
twist in the plane of a saccule may always be noticed in the
o-G-furrow stage, by which each becomes distinctly more
horizontal in its anterior region (fig. 13b) than at the orifice
(fig. 13a). 'I'his twist does not seem to be retained through-
out all subsequent stages.

From the 5-furrow stage until the period when the
cuticula and the lacuna; first appear in the lung-books the
latter present various characteristics, best studied in trans-
verse sections, such as fig. loB. The ventral wall of each
of the saccules (.v. 1, .v. 2, etc.), is distinctly thicker than the
dorsal wall, its cells being more cylindrical and more
nursierons, its nuclei more oblong and situated nearer the
ventral (basal) ends of tlieir cells, which thus come to have
more protoplasm at the free (dorsal) ends than do the corre-
sponding (or ventral) ends of the cells of the dorsal wall.
'I’he saccules are each provided with a considerable cavity,
but between the closely appressed walls of two adjoining-
saccules no lumen whatever is found.

^Vith the appearance of the chitinous structures and the
blood-lacunae at the end of the reversion a great change takes
])lace in the appearance of the walls of the saccules, the
older ones being, as usual, those first affected. There appear
between the walls and cells of two adjoining saccules irregular
spaces figs. 17 and 18), which are at first small, but

rapidly eidarge and communicate with one another and with
the blood-cavities on the medial and lateral sides of the lung-

30

W. F. PURCELri,

book, tlins forming a passage for the blood and blood-
corpuscles {hd.e.) from the one side to the other. All
mitoses definitely cease in such saccules, although they are
common enough in the previous stages, as well as in younger
not yet chitinised saccules of all subsequent stages. The two
adjacent walls do not, however, lose contact with one another,
for each cell of a dorsal wall of a saccule (with a few excep-
tions) remains united with one or two cells of the ventral wall
of the adjacent saccule by means of a column of protoplasm,
in the formation of which both or all three cells (ir., fig. 18)
take part. Owing to the excess of nuclei in the ventral wall
of the saccnle we often find a column provided with two nuclei
at its dorsal and one at its ventral end (//., fig. 18), while
some of the cells of the ventral wall become simple plaster-
cells unattached to a column {z., fig. 18). Similar double
nuclei and plaster-cells ai*e rarely found in the dorsal wall of
a saccnle. This arrangement of the nuclei is retained through
all subsequent stages up to the adult form, and was found in
the adults of all other spiders examined.' I also found it in
embryos of Agelena labyrinthica, and it is evidently
general amongst Dipnenmonous spiders.

The nuclei vary greatly in shape. IMany are more or less
depressed in the plane of the septa, becoming plano-convex
oi- conical, the plane side facing the chitinous cuticula.

'Phe cells of the ventral wall of the oldest saccnle (.v. 1)
require special mention. These also form columns, which
attach themselves to the body hypodermis, but the cells of
the latter do not contribute to these structures. The nuclei
of this saccule are often drawn out in a peculiar way into the
thinnest part of the ventral columns (fig. 17). Locy, who
describes these columns, considers them to be probably of a
muscular nature, but there does not .seem to me to be any
reason for thinking that they are any more muscular than
the colmnns of the septa. Their greater length is simply
explained by the fact that each cell has to form a column, at

' The plaster-cells were first noticed by Berteanx ('89) in fully
developed spider’s lun"s.

RESPIRATORY ORGANS IN ARANEiE.

31

least as long as the two-celled columns of the septa, in order
to allow sufficient space for the blood-corpuscles to pass
between the ventral saccule and the outer hypodermis.

Two authors, Locy and Jaworowski, deal with the formation
of the definite lung-septa from the embryonic epithelia.
According to Locy (’86), whose account differs from mine,
the nuclei, which are in parallel rows, become plano-convex
and arrange themselves in pairs, the convex side of each
nucleus in one row being exactly opposite that of an adjacent
parallel row (i. e., of an adjacent epithelium). Ultimately
the cells of each pair of nuclei, which thus face each other,
come in contact and fuse together to form the columns.
’I’lie cells of such a pair of rows constitute the two walls of a
flat, hollow sac, a respiratory lamella (i. e., a septa). Blood
has a free access to the lamelhn at their anterior attachments.
(Locy’s statement that a septa represents a hollow sac is, of
course, incorrect. He apparently considers them attached at
their anterior ends only.)

Jaworowski’s account (’94, pp. GO-Gl), is more in agree-
ment with mine. According to him the space between the
two layers of nuclei of a septum is filled with protoplasm and
tlie lacuna) appear between the cells, and are at first small
and roundish, and later on large and elongate. Jaworowski
evidently intends to imply that the columns are the remains
of the protoplasm left between the lacunae, and his fig. 12
illustrates this very clearly. Here two, or even three nuclei
may be observed at one or both ends of a column at first, but
later on this is rarely or never the case, only one nucleus
being found at each end of the column (in agreement with
Locy).

The chitinous lining of the pulmonary saccules. — Shortly
before the appearance of the lacuna) the walls of the saccules
appear to collapse, and on the surfaces of contact, where the
cavity was situated, two chitinous membi’anes are secreted.
These pass over into one another at their medial, lateral, and
anterior edges, so as to form a flattened chitinous .saccule
within the epithelial saccule, and are further connected by

32

W. F. I’UHCEU..

iiimniierable tiny cliitinous rods, which are firmly soldered to
each membrane and distributed over their entire inner sur-
faces (.s. 1, figs. 17 and 18). The ante-chamber is also provided
Avith a smooth cnticnla fig. 18), except in the dorsal

growing ]>art {pulm. 'prol.).

The walls of the cliitinous saccules are lined on one(thebasal)
surface with a thin layer of protoplasm, which is, of course,
the matrix, and althoug’h this layer may become very thin (as,
for instance, in Agelena laby rin thica), it is always dis-
tinctly recognisable at this stage. Locy could not trace the
protoplasm on the chitin away from the columns in Ag-elena
naevia, ivhile Jaworowski (’94) describes these columns as
atmeboid in shape, sending out processes over the surface
of the chitin to connect with those of neighbouring cells of
the same epithelium.

The moulting' of the lung-books. — It is well known that at
each monlt of the young spider the entire cliitinous lining of
both the ante-chamber and saccules is cast off (Menge, ’51,
p. 22; W. Wagner, ’88, p. 315), and that the ventral walls of
the latter produce the innumerable free spines on the surface
of the cnticnla (W. Wagner, ’88, p. 314). A’’arious points of
interest still remain to be described in connection wdtli the
growth at monlting-.^

Already at the time of hatching we find the saccules pre-
paring for the first post-embryonic moult, although the latter
does not take place until nearly a week later. The epithelia
of each saccule expands in a medial, as well as in an anterior
direction, considerably beyond the corresponding edges of its
primitive cliitinous lining, while the lateral and posterior
edges I'emain stationary. 'I’he enlarged saccule thus created
then secretes over its interior surface a new cuticula forming
a, second cliitinous saccule (/., fig. 34), Avhich encloses the
one first formed (.v.) and differs from it in structure. For its
ventral membrane bears over that part of its area which is
co-extensive with the primitive cuticnlar saccule (.s.) nume-

' The fullowing' remarks on this suhject apply equally to Attiis
floricola, Agelena lahyrinthica, and Tegenaria atriea.

RESPIEATORY ORGANS IN ARANE.E.

33

rous short cones (c.), not attached to the dorsal membrane,
while in the newly added medial portion (s'.) the rods are
fused with both membranes.

Herein lies the explanation of the greater thickness of, and
the larger number of cells in, the ventral wall of the saccules
in the earlier- stages (fig. 13b) described on p. 29 ; for we
may assume that the ventral wall secretes the numerous
minute rods as well as the ventral cuticula of the primary
chitinous saccules, and that only their dorsal cuticula is con-
tributed by the doi’sal wall of the saccules. Being in contact
at first the rods of the ventral cuticula are able to fuse with
the dorsal cuticula, but at the first moult and all subsequent
moults the two cuticulas are sepai-ated by the previously
formed chitinous saccule except along the newly added medial
and anterior portions. The chitinous saccules first formed are
cast off at the first moult, but they previously become squeezed
very thin and are thus difficult to i-ecognise as such.

At each subsequent moult the saccules are enlarged in the
way described for the first moult, and since in the medial and
in the anterior portion of the chitinous saccules at any period
of life the rods ai’e found soldered to both membranes, I con-
clude, generally, that this soldered region represents the
portion that was added at the previous moult. ^

My account of the primary chitinous saccules differs from
that of both Locy and Jaworowski. The first-named author
(’86) describes and figures the dorsal chitinous membrane of
each saccule as smooth and the ventral membrane as den-
tigei'ous, but not united to the dorsal one in the embryo in
Agelena naevia. In my sections of the embryos of
A g e 1 e n a 1 a b y r i n t h i c a the two membranes of the primiti ve
saccules are undoubtedly fused together, exactly as in Attus
floricola. According to Jaworowski’s description, in the

' Tlie same appears to he the case in many other spiders, although it
has liitherto escaped the notice of investigators: so, Argyroneta,
Drassodes, Lycosa, Philodromus, etc. There is no special reason
wliy the added region should never have free rods, hence the above
statement must not be applied too strictly to all spiders.

VOL. 54, PART 1. — NEW SERIES.

3

34

\V. 1’. rURCELL.

embryos of Trocbosa singoriensis both the membranes
bear granules (i. e. the teeth), and from his figures it is clear
that these membranes are not fused together.

Both these authors’ accounts may very easily be reconciled
with one another and with mine, if we assume that their
figures represent stages in which the preparation for the first
post-embryonic moult had already begun. Locy’s figures then
would represent sections in which the new cuticiila of the
dorsal \vall of the saccule had separated from the primary
chitinous saccule and so appeared smooth, while the ventral
cuticula would still appear dentigerous. It may happen in
Attus floricola that the ventral w^all of the secondary
chitinous saccule (*■'.) becomes pulled apart from the
primary saccule (.s-.), which, adhering to the dorsal wall,
causes it to appear as if both walls of the saccule were pro-
vided with denticles. This, no doubt, is the explanation of
Jaworowslci’s statement.

The operculum of the lung-books. — It is -well known from
the observations of Morin (’87), Kishinouye (’90) and others
that the outer epithelium of the pulmonary appendage forms
the operculum, which covers each lung-book after the appen-
dage has sunk into the body.

It will be observed from a comparison between figs. 13b
and 17, and between figs. IGa or 1Gb and 18, that the sides,
as well as the distal wall, of the abdominal appendage con-
tribute to the formation of the operculum. ’Phus, in fig. 17
the ventral portion, w'.x'., of the operculum, to 'which the
ventral columns of the oldest saccule, .s.l., are attached, cor-
respond to the ventral wall, w' .x'., of the appendage in fig. 13b,
while the distal and dorsal walls, x'.y'. and y'.z'., of the latter
correspond as nearly as pos.sibleto the portions x'.i/ .and y'.z.
of the operculum in fig. 17 (both figures being magnified the
same number of times). Aline {jir. ax.) through the centre of
the area x'.y'., or, say roughly, of the entire operculum, and
perpendicular to its surface would, 1 think, correspond
approximately with the original axis of the appendage.

Since the positions of the septa and the operculum remain

EESPIBATORY ORGANS IN ARANEyE.

35

practically unclianged after the stage represented in fig. 17,
we can disting-uish in the operculum of the adult spider (1) a
nearly horizontal portion to which the ventral saccule is
attached, and which belongs to the ventral surface of the
abdomen, and (2) a strong-ly inclined portion on the lower
part of the lateral surface of the abdomen. The horizontal
part corresponds to the ventral wall of the embryonic appen-
dage in hg. 3 {iv'.x'. in figs. 13c and 1 7), or the median wall of
ail earlier stage (fig. 1), while the inclined portion, which forms
much the greater part, is the distal and dorsal wall of the
appendage, i . e. — the part x'.z'. in figs. 13c and 1 7, or the distal
and lateral wall of an early stage (fig. 1). Anteriorl}’^ the
operculum curves strongljr towards the median line, and this
incurved part corresponds, of course, to the anterior wall of
the embryonic appendage (fig. 18). All the surfaces pass over
gradually into one another and cannot be sharply dis-
tinguished.

The lung-books of the young spider. — Not much remains to
be added on the subsequent development.

At the time of hatching the lung-book has much the appear-
ance of fig. 18, except that the pulmonary .sac (now the ante-
chamber) has much thinner walls, lined with chitin internally,
and the dorsal saccules are long’er. Moreover, that portion of
the epithelium of the pulmonary sac immediately adjoining
the spiracle now forms a thin-walled, narrow, hollow neck or
stalk (pedicel) connecting the ante-chamber proper with
the edge of the spiracle.

'I'his pedicel persists throughout all later stages, and its
chitinous lining acts both as an air-passag’e to the ante-
chamber and as a sort of ligament by means of which the
lung-complex is firmly attached to the outer cuticula of the
body.

3’he dorsal horn of the ante-chamber preserves its charac-
teristic curved form, and, as Bertkau (’72) showed long ago,
continues to provide new lung-septa. According to W.
Wagner (’88), the addition of new septa goes on until the age
of sexual maturity is reached. In Attus floricola at the

36

W. F. PUJiCELL.

time of hatcliiug there are about seven or eight developed
saccules. At the time of the second moult there are perhaps
twelve to fourteen, while in the adult about thirty-four or
thirty-five appear to be present, but I cannot state the exact
numbers with certainty.

Critical remarks on the literature. — Aranefe. — According to
Locy (’86,p.81) the in-foldings for the lung-books in Agelena
ngevia arise late in the period of the reversion. From his
figure (fig. 73) and description of “early stages” (p. 89), in
which the lung-books appear as extensive gi’oups of cells
with the nuclei arranged iu parallel rows, as well as from the
fact that he makes no mention of any connection with the
abdominal appendages, it is clear that Locy was really deal-
ing with late stages after the appendage had already sunk
into the body and long after the earlier saccules had been
formed. Of the formation of these latter he gives no
account. His account of the formation of the definite septa
has already been dealt with on a previous page (p. 31).

Bruce^s (^86a, ’86b, ’87) statements may be dismissed as di.s-
proved by later researches. Both Kishinouye (’90) and
Simmons (’94) are of opinion that Bruce (’87) has misinter-
preted the parts in his figures Ixxix and Ixxix'. Certainly
the fold L' is not a pulmonary fold, and is not on the anterior
surface of the first abdominal appendage, as Bruce supposes
it to be.

Simmons (’94) states that the pulmonary sac arises as an
in-pushing behind and under the abdominal appendage, “ so
that eventually a pit is formed, actually extending into the
general body surface.” The pit is considered as bounded on
its outer side by the appendage itself, its outer wall being-
described as “the morphologically posterior surface of the
appendage” (p. 217), which is represented as lying flat on
the body surface and directed backwards. The opening of
the pit under the posterior or distal end of the appendage
persists as the spiracle. The outer wall of the pit “has its
ectoderm thrown into folds, the rudiments of the leaves of
the lung-book,” and sections of early stages are figured, one

KESFIRATORY ORGANS IN ARANE.E.

37

section (fig. 6) showing five pulmonary furrows and the other
(fig. 5), although of the same age, only two such farrows.

It is plain that the author considers that the earliest lung-
leaves are fonned entirely within the pulmonary pit or sac
and not on any part of the free surface of the appendage
outside of the sac; so that, as far as the position of the
lung-leaves in regard to the appendage at their first appear-
ance is concerned, the author has not advanced beyond what
was known to his predecessoi’s. Nevertheless, in the sum-
mary at the end of the paper we find the following statement,
that ‘Hhe lung-book of the spider (and presumably of all
Ai’achnids which possess one) arises at first as an external
structure upon the posterior surface of the abdominal
appendages” (p. 219).

If we accept the theory that the lung-books are derived
from gill-books as indisputable, then we can say that the
appearance of the lung-leaves on the outer or anterior wall of
the pulmonary sac proves that this wall is morphologically
the posterior side of the abdominal appendag’e, but we cannot
conversely first call this wall the posterior side of the
appendage and then say that the appearance of the hing-
leaves upon it proves that they are formed on the posterior
side of the appendage, as Simmons does. For if we choose
to consider that the lung-h-oks were derived from internal
tracheae and not from external gill-books, the pulmonary sac
would be the trunk of a ti’achea, and no one would then call
its outer wall the posterior wall of the appendage. Thus, if
Simmons’ description of the early development were correct,
then the lung-books would not arise at first as an external
structure, but as an internal one in an invagination.

As a matter of fact Simmons’ representations of the abdo-
minal appendage in his figs. 5 and 6 are very misleading, as
will appear if we refer to his fig. 10, which represents an
entire embryo of the same age as those in figs. 5 and 6.
Here the first abdominal appendage has its usual stumpy,
knob-like form, and is situated on opposite sides of the
abdomen, almost antipodal in fact, just as in Attus f loricola.

38

W. F. PURCELL.

Sagittal sections, like Simmons’ figs. 5 and 6, therefore, cut
the appendage more or less transversely to its main axis,
which in the two figures would be, not in the plane of the
paper, but almost perpendicular to it.

In fact I cannot believe tliat the appearance of the a])peu-
dage in Agel ena n gev ia at this stage differs so essentially
from the corresponding stage in Attus floricola, such as
that represented by my fig. 1(5. A sagittal section in the
case of the appendage represented in this figiu'e would, of
course, be more or less perpendicular to the plane of the
paper and cut the appendage parallel to the line ep.-ep. If
the section were slightly more inclined towards the lower part
of the paper (say along a.-h., fig. IGn) we should get a section
like fig. 15, but if it were inclined more towards the upper
pai-t of the paper (say along c.—d., fig. 16b), we should get
sections exactly resembling Simmons’ figs. 5 and 6, according
as two or five of the furrows were cut. This 1 believe to be
the true explanation of the appearance of Simmons’ figures.
It is extremely difficult, if not impossible, to get an exact idea
of the structure of an appendage without the aid of wax
models, of which Simmons does not say he made any use.

'I’he last paper on the spider’s lung-book to be considered
is that of -Jaworowski (’94), who studied Trochosa singo-
riensis. He discovered in this species an embryonic structui e,
which he de.scribes as an embryonic trachea, consisting of an
ante-chamber, a trunk, and branches. 'I’he ante-chamber is
inverted funnel-shaped, with the apex pointing upwards and
the broad end terminated ventrally by the abdominal appen-
dage or operculum. The sides of the ante-chamber are
closely appressed to one anotlier (j). 56) and extended in a.
sagittal plane (since they are seen broadways in sagittal
sections). The pulmonary lamelhe are formed by parallel
folds of the wall of the ante-chamber, “ the edges of the folds,
which jut into the lumen, being more or less (figs. 1 and 2)
undulate” (p. 62) and parallel to the surface of the operculum,
i. e., transverse to the axis of the ante-chamber and trachea.

According to Jaworowski’s idea, therefore, the free edges

BESPIKATORY OEGANS IN AEANBiB.

39

ot: the septa run parallel to the longitudinal axis of the
abdomen instead of at right angles to it, as they do in Attus
floricola, etc. Now if we compare my figs. 13b and 17
of transverse sections with Jaworowski’s figs. 3 and 5/ which
come just in between mine in point of development, it will be
seen that the lung-books of botli species exactly correspond,
so that the free edges of the septa cannot run longitudinally
to the body axis. In fact, Jaworowski has evidently mistaken
the direction of the folds, which are seen laterally in his
figs. 3 and 5 and not from their free edges ; and, moreover,
the funnel-shaped area which he calls the ante-chamber in
his sagittal sections is not the ante-chamber at all.

The trunk of the embryonic trachea, according' to Jawo-
rowski, extends dorsad from the apex of the ante-chamber
and then divides, the branches reaching to the dorsal blood-
vessel and subdividing into smaller brancblets. These have
sometimes the appearance of a cuticular tube provided with
regular internal thickenings (fig. 6). Ultimately both trunk
and branches degenerate and disappear, only the “ ante-
chamber'’ remaining to form the lung-book.

In the later stages of the spider-embryos which I ex-
amined, I find the yolk-mass continuous along the median
region but divided towards the sides by partial septa, which
are transverse and doubtless of mesodermal origin. The
surfaces of the yolk are lined with very thin Hat cells, and the
intra-septal space between these two layers of cells contains
muscles, blood-corpuscles, and a number of large vitello-
phagous cells resembling those marked i'it. in my figs. 16d, 16e,
17, etc. Ventrally the intra-septal spaces widen out, the
widened part appearing funnel-like in stigittal sections (see
tig. 41, which shows three such septa). The lung-books lie in
the ventral widening of the septum between the eighth and
ninth segments. The space between the lung-books and the

‘ The author calls these " frontal sections,” hut since the abdomen is
inclined ventrally to the longitudinal axis of the cephalothorax, frontal
sections of the hitter would cut the abdomen more transversely than
frontally. (See Locy’s fig. 10 or Korschelt and Heider, p. 585, fig. 372b. J

40

W. F. PUKCELL.

yolk also contains blood- corpuscles, vitellopliagous cells, and
various iiiesoderinal elements, besides fluid.

Jaworowski’s tracheal trunk and ante-chamber undoubtedly
correspond in position to the lower part of the septum and
its funnel-shaped widening, but 1 have found nothing in them
in my sections which could possibl}’ be taken for trachea}.
Jaworowski states that the trunk has a nucleated epithelium,
the nuclei being smaller than those of the pulmonary
lamellje (p. 62). These may well be, 1 think, the nuclei of
the mesodermal septa, but I am at a loss to account for the
tracheal branches and branchlets drawn by Jaworowski in
his tigs. 1 and 2. At any rate the tracheal nature of the
structure cannot possibly be maintained so long as no
embryological evidence at all is advanced to prove that they
are of ectodermal origin and derived from the same mass of
cells which form the lung-books. It will be noticed further
that the lumen of the ante-chamber is closed off from that of
the tracheal trunk by a diaphragm (p. 68).

No other investigator has ever found anything like these
embryonic tracheae, and although Jaworowski (’94, p. 55)
asserted that Schimkewitsch (’86a, ’86b) had previously
observed a similar structure, the latter author has recently
(:06, p. 45) disclaimed any connection between that figured
by him and those found l)y Jaworowski.

Scorpiones. — Metschnikoff ('71), Laurie ('90 and ’92j,
Brauer (’95) and Pereyaslawzew'a (:07) all agree that the lung-
books of scorpions arise as folds in the wall of the pulmonary
sac, which according to the first three authors is formed by
invagination on the posterior sides of the four posterior pairs
of abdominal a})pendages. According to Pereyaslawzewa,
however, this sac arises on the anterior side of the appen-
dages, but it appears to me probable that this author has mis-
taken the intersegmental folds which separate the sternites
for appendages, the true appendages described by previous
authors having evidently already disappeared.

Brauer states that, so far as he could make out, the oldest
pulmonary fold occurs at the innermost part of the sac, the

KESPIKATORY OEGANS IN AllANEiE.

41

following folds occurring on the distal side of this one (i. e.
exactly opposite to what takes place in spiders). The autlior
does not appear to be cjuite certain about this point, and is,
moreover, corrected by Pereyaslawzewa, who maintains that
tlie oldest fold is the one nearest to the outer body wall (1 . e.
as in spiders).

Brauer’s text-hg. 15c (p. 413) very closely resembles my
fig. IbB, so far as the ectoderm is concerned. He thinks
there can be scarcely a doubt that the lung-book is not
formed behind or apart from the appendage, but is the
posterior half of the latter itself, which is invaginated and
on which the pulmonary folds appear (p. 415).

Laurie (’92) makes an interestiug statement regarding the
position of the lung-septa in the older scorpion-embryos,
Here they are placed horizontally, as in the older spider-
embryos, whereas in the adult scorpion they are vertical

(p. 102).

Pedipalpi. — The development of the lung-saccules and
their relation to the abdominal appendages do not appear to
me sufficiently clear, from the existing embryological data, to
make a comparison with the Aranese possible. Apparently
the abdominal appendages are not so obvious in this group
as they are in A ran e a) and Scorpiones, since the parts
described by Laurie (’94j under this name are not identical
with those to which Schimkewitsch (:06) applies the term.

A remarkable point in the development, as described by
ISchimkewitsch, is that the oldest saccules are said to be
formed within the pulmonary sac and to subsequently migrate
out of it ou to the posterior side of the appendage. In such
a case their development would be exactly the opposite to
tliat in A ran etc, as well as to what we should expect from
phylogenetic considerations.

Pereyaslawzewa’s (:01) descrijjtion of the formation of the
lung-septa out of the cuticular wrinkles of the body-surface
is altogether fanciful.

The fully-developed lung-books of spiders. — A. Schneider (’92)
has given an excellent account of the coarser anatomy of the

42

W. F. PUKCELF;.

lung-books in spidei-s, the descriptions of MacLeod (’84) aud
Berteaux (’89) being unsatisfactory in this respect. Berteaux’s
account of the chitinous structures (spines, etc.) of the lung-
books is, however, very detailed and the best we possess, but
his description of the bi-nucleated cell-columns in tlie septa
as nnicellnlar structures is misleading and not in accordance
with the embryological facts, since these columns are formed
by the fusion of opposed cells in two separate epithelia.

Both MacLeod and Berteaux made a curious error regard-
ing the free edges of the septa. The edges of the septa they
describe as being free, not only along the posterior border
but along tlie posterior part of the lateral side as well. As
a matter of fact tlie lateral sides of the septa are never free,
but may have the appearance of being so in horizontal
sections through the dorsal procurved portion of the ante-
chamber. The apparently free edge is merely that of a
septum with its lateral part cut off by the razor, hence the
irregularities in its occurrence observed by these authors.
'I’hat these lateral edges are not free can easily be demon-
strated by examining the lung-books under a hand-lens after
treatment with caustic jiotash, and I can strongly recommend
this old-fashioned method to anyone who wishes to obtain a
clear idea of the coarser structure of the lung-books in a short
time (see fig’. 20). It will be found much more satisfactory
than if one were to rely on sections only.

Bbrner (:04) has recently stated that the se})ta are placed
more or less vertically in the majorit}^ of the Aranete, and
has thrown doubt on MacLeod’s well-known diagrams, in
which the septa are i-epresented as lying horizontally.

I have examined one or two species of most of the larger
families and I found the septa as nearly horizontal as they
could well be in the following Lipneumonous spiders :
Attidm (Attus floricola), Lycosidm (Lycosa Uar-
lingi), Agelenidte ('I’egenaria domestica, Textrix
lycosina), Clubionida) (Clubiona holosericea, Paly-
stes sp.), Thomisida) (Philodromus f uscomargina-
tus), Theridiid^ (Theridion lineatum), Drassidte

KESPlliATOKY OKGANS IN AHANEiE.

48

(Drassodes tesselatus), Sicariidie (Scytodes tes-
tudo); also iu the following Tetrapneumonous spiders
(Aviculariidse) : subfam. Avicu lar i inse (Harpactira
atra)j subfam. Cteuizinae (Sbasimopus unispiiiosus
and Her mac ha sp.).

In the following forms the septa were inclined at an angle
of 45° or less to the horizontal, sloping downwards from the
higher medial edges to the lower lateral edges : Argiopidae,
subfam. Argiopiuae (Argiope clathrata), Theridiidm
(Latrodectus geometricus), and Eresidie (Eresus
sp.).

If the above examples are any indication of the usual posi-
tion in the families to which they belong, then Horner’s state-
ment niust be wrong, and cannot hold good for the great
majority of Araneie. Even in the three cases where the
septa were incliued they were nearer the horizontal than the
vertical (except, perhaps, in Latrodectus geometricus,
where they formed an angle of about 45°).

Moreover, the operculum iu the spiders with horizontal
septa is similar to that of Attus floricola described on p.
49 (see hg. 17), and since this type of operculum represents
that of any Dipneumonous or Tetrapneumonous spider in
which the abdomen is not greatly developed anteriorly, we
may faiidy assume that the septa must be horizontal, or very
nearly so in the great majority of s})iders.

^Vllen, however, the anterior upper region of the abdomen
is abnormally distended above the opercula, it may happen
that the lateral region of the latter becomes pushed down-
wards into a more horizontal position than is the case in
fig. 17, and at the same time the septa become tilted upwards
on the medial side, that is to say, they take a more or less
incliued position, such as one finds in Argiope, Latro-
dectus, and Eresus, and no doubt in many other genera of
the same families, d'he inclined position of the septa cannot,
therefore, be a primitive condition in these families, but, I
think, merely due to the abnormal distension of the abdo-
men, for iu closely allied forms, in which the abdomen is not

44

W, F. PUECELL.

unusually distended and the operculum more upright, I find
the septa placed almost horizontally (e. g. in a specimen of
Nephila from Senegal).

VI. The Development op the Abdominal Longitudinal
Muscles and theik Tendons.

In their earliest stage the coelomic sacs of the eighth to
eleventh segments each protrude an evaginated portion of
their somatic wall into a provisional appendage, completely
lining the cavity of the latter (fig\ 4). At this stage {St. I)
the cells of the somatic wall of the sac are cylindrical, and
much higher than those of the splanchnic wall.

At the time when the first pulmonary furrows begin to appear
(stage 2) the intra-appendicular portion becomes partially
cut off from the main coelomic sac by an infolding of its wall
along the medial basal edge of the appendage (fig. 1). The
infolded layer grows in a lateral direction halfway across the
cavity of the appendage, converting the medial half of the
intra-appendicular coelom into a short tube. Each of these
segmental tubes lies in a transverse plane, is blind at the
medial end, and opens laterally into the coelomic sac.^

In a longitudinal section through the lateral region of the
appendages {ah. ai)p. 1-3, fig. 5a) the main coelomic cavity is
seen to be continuous with the intra-appendicular portion,
whereas the latter portion in a more medial section of the
same series (fig. 5) appears separated from the main coelom,
the segmental tubes {seg. t. 3-10), of course, being seen in
cross section. If this latter section be compared with a
corresponding section of a later stage (fig. b, stage with five
pulmonary furrows), it will be noticed that the portion of the
somatopleura which formed the inner layer of the above-

‘ I have sliown elsewhere ('95) that the segmental tubes of the eighth
or pulmonary segment become the genital ducts. In fig. 23b (adult
male) the mesodermal part of the genital duct [mes. fj. d.), derived from
the segmental tubes, is seen sharj^ly differentiated from the ectodermal
portion {ec. <j. d.).

RESPIKATORY OROAXS IX ARAXE.E.

45

mentioned fold, and which overlies the segmental tube, has
increased considei*ably in thickness in each segment. Its
cells (»i., fig. 6) are no longer cubical and one-layered, as they
are in fig\ 5, bnt have become spindle-shaped, elongated
longitudinally to the body, and arranged in several layers.
Each such group of elongated cells forms a bundle whose
ends are in contact with the anterior and posterior basal
edges of the appendages, thus completely bridging over the
.segmental tubes, and ultimately gives rise to a corresponding
segment of one of the two great ventral longitudinal muscles
of the abdomen.

The ectodermal areas to which these muscular
segments are attached are of primary importance
in enabling us to determine the homologies of the
tracheae. In their eailier stages it will be seen that these
areas (nr. 7-11, figs. 6 and 27) are precisely similar to each
other in position and arrangement with regard to each
appendage. They occupy the visceral surface of the medial
half of each post-appendicular (intersegmental) in-folding,^
and their transverse extension is at first nearly the same as
that of the segmental tubes with which the areas alternate.

Each contact area soon becomes marked by the appearance
of the intermuscular tendons or en toch on drites.
These organs, whose mesodermal origin has already been
demonstrated by Schimkewitsch (’94) are formed in various
parts of the body by the fusion and metamorphosis of the
ends of the muscular cells at those places where the ends of
two or more muscle-bundles come in contact with each other
and with the hypodermis." In the embryo they form a non-

' For the area (ar. 7) Ijetween segments 7 ami 8 there is, of course,
no post-appendicular in-folding, hut its ijosition is otherwise precisely
similar to that of the othei-s.

- Schimkewitsch, to whom we mainly owe our knowledge of the
development of the entochondrites in Arachnida, has a somewhat
different view of the conditions necessary for the formation of these
organs. He says ('94, j). 20(5) : “ Mann kann hehaupten, dass in jenen
Fiillen, wenn zwei Muskelanlagen einander entgegen wachsen, sie mit
einander verwachsen ; wenn sie aher unter einem Winkel zusammen-

46

W. F. rUKCELI-.

nucleated homogeneous mass, which stains very deeply with
carmine, and is, histologically, readily distinguishable from
all adjacent tissues. All the cells of the ventral longitudinal
muscles become subsequently attached by each end to these
entochondrites only, which in turn firmly adhere to the bases
of the ectodermal cells of the deepest part of the contact-
area described above, and provide us with a certain means of
locating- these latter cells long after the subsidence of the
appendages. It should be noted that the entochondrites are
much less extensive at first than the ectodermal areas (ar. 7,
etc., figs. 6 and 27) Avith Avhich the ends of the forming
muscles Avere originally in contact.

From Avhat has just been said, there can be no doubt that
the places of attachment of the entochondrites inserted in
each ventral longitudinal muscle are serially homologous, a
fact Avhich may also be readily inferred from a glance at
fig. 41, Avhich represents the stage shortly before hatching.
From this figure it may also be observed that scA^eral other
muscles are attached to the entochondrites {f. 7-11), in addi-
tion to the segments (r. 1. m. 7-11) of the great longitudinal
muscles as, for instance, (1) a long dorso-A’entral muscle
{<1. V. m. 7-10); (2) a short, oblique muscle (p. oh. vi. 8-11)
running- obliquely foi-Avards and attached near the middle of
its segment to the hypodermis Avithout the interposition of
an entochondrite ; and (8) a corresponding oblique muscle^
{a. oh. r/i. 8 and 10), running obliquely backAA-ards, present in
segments 8 and 10. In Agelena lab3"rinthica I also

treffeu, so liildet sich an der Stelle ihrer Beriilu-uiig eiiie Seliiie meso-
dei-iualen Ui-sjn-ungs.” To my mind it does not matter whether the
muscles are in a line or meet at an angle, the essential conditions being
only that the ends of tivo or more muscles should meet at one
spot and that that spot should be on tlie basal surface of
the hypodermis. When muscles are attached singly to the hypo-
dermis they do so directly AA-ithout the intervention of an entochondrite.

‘ I have called this muscle-series and that pi-evioAisly mentioned the
anterior {a. oh. m.) and posterior oblique muscles (p. ub. w.), because
they lie in the anterior and posterior regions resjAectively of their
soinites.

HESPIRATOKV ORGANS IN ARANEA;.

47

found an oblique muscle on the medial side of the enta-
pophysis of the pulmonary segment, which, perhaps, corre-
sponds to a muscle {a. oh. m. 9) not observed in Attus
f loricola.

Ill the embryo and the growing spider the ectodermal
areas to which the eutochondrites of the longitudinal muscles
are attached become drawn out in the form of hollow pro-
cesses {ec. f. 8-11, fig. 41), lined with chitin, and projecting
into the interior of the body. These are the ectodermal
tendons or entapophyses ' (apod ernes) of the ventral
longitudinal muscles, and it was the purpose of the foregoing-
paragraphs to establish their serial homology.

Since the segments of the longitudinal muscles are each
formed out of the entire length of the somatic wall of a
coelomic sac, it follows that their points of attachment (or the
entapophyses) must be intersegmental, and this is also the
case with the points of attachment of the dorso-ventral
muscles where they coincide with those of the longitudinal
muscles (as, for instance, the ventral end of the muscle
(l.r.'ia. 8, and both ends of d.r.iu. 9 and 10 in fig. 41). But
it frequently happens in the Arachnida that the dorso-
ventral muscles disassociate themselves at one or both ends
from the insertions of the longitudinal muscles, and may then
form separate external depressions, which are not necessarily
intersegmental, but, in fact, very frequently placed more
forwards on the face of the segment. Thus in fig-. 41 the
muscle d. v. m. 8, although actually in contact at the place of
crossing with the dor.sal longitudinal muscle {d. I. m. 8) of the
eighth somite, is inserted dorsally at some distance in front
of the intersegmental entochondrite which lies between the
muscles d. 1. m. 8 and 9.

VII. The Entapophyses (Ectodermae Tendons) of the

PUDMONAHV EeGMEN'I'.

When the entochondrites beg-iu to form at about the stage
with five pulmonary furrows, the first pair of provisional appen-
‘ A term introduced l>y Ray Lankester.

4S

W. P. PURCBLL.

dages has reached its most lateral position (fig. 3). The
anterior wall of appendage 2 has approached nearer to
appendage 1, so that the ectodermal area («r. 8, figs. 6
and 16), to which the entochondrite becomes attached, now
forms the bottom of a rather wide groove lying between the
two appendages towards their medial side. Shortly after-
wards the three posterior provisional appendages commence
to move towards the posterior venti-al part of the abdomen
away from appendage 1.

Now, as the latter retains its lateral position for the pi’esent
and is very close to appendage 2, we find the area {ar. 8)
between them, to which the tendon is attached, also shifting
slightly ventrad. In fig. 27 the appendages are still in a row,
blit the ventrad movement is commencing; in fig. 16 fa
slightly later stage) appendage 2 has moved ventrad to the
region comprised between the sections nos. 1-18, and the
area of attachment {nr. 8) is no longer just behind the
region of the two oldest furrows (/. 1 and/. 2), as it appears
to be in fig. 27, but lies ventrally to them. When this area
has reached the extreme postero- ventral corner of the base of
the appendage, it remains there, while appendage 2 con-
tinues its ventrad movement.

The subsequent development up till after the second moult,
is a simple one. After the completion of the reversion and
shortly before hatching (stage 6) we find the entochondrite
{t. 8, fig. 41) situated alongside the posterior median corner
of the lung-complex {Ih.), just next to the medial end of the
spiracle. The ectodermal area {ec. f. 8) to which the ento-
chondrite is attached, is composed of elongated cells and is
somewhat sunken-in, forming a shallow, groove-like con-
tinuation of the spiracle (sp., fig. 18). At the second moult,
when the young spider attains its definite form, this groove
becomes obliterated, so that the hypodermis of the attach-
ment area comes to lie on a level with the adjacent body
surface and completely outside of the spiracle.^

’ In A^^elena liibyrinthica the groove is present in all stages
after the time of hatching.

RESPIKATORY ORGANS JN ARANEA5.

49

As tlie youug spider, however, a.pproaches maturity, the
attachment area aorain nndero-oes a considerable alteration in
its form and. position, resulting mainly from two processes.
These consist in (1) the formation of a transverse in-folding
of the hypodermis (the interpulmonary fold) between the
two spiracles, which thus become connected by a deep
ventral groove ; and (2) the drawing-out in the fonn of a
blind tube (entapophysis) of the dorsal edge of this fold at
the two spots to which the pair of mesodermal entochondrites
are attached.

The interpulmonary (epigastric) fold in the adult of Attus. —
Fig. 20 represents a posterior view of the abdominal chitinous
skeleton of an adult of a species of Attus, after the removal
of everything posterior to the transverse plane which passes
through the spiracles. It will serve to illustrate the form of
the interpulmonary fold in the adult.

In Attus floricola the interpulmonary fold in the male
differs greatly from that of the female.

Figs. 23-2dB are three sagittal sections through this fold
in the adult male in the regions indicated in fig. 20. The
much crumpled, anterior and posterior surfaces of the groove
contained in the fold are normally closely applied to each
other, so as to leave very little space between them. Along
the dorsal edge, however, this is not the case, for here the
groove suddenly widens to a nearly cylindrical canal {can.),
which opens on each side into the pulmonary ante-chambers
at their medio-ventral corners, thus forming a permanently
open communication between the two lungs canal of com-
munication” observed b}' Berteaux (’89) in Agelena and
Kpeira]. The chitinous wall of the canal is thick and
covered with branched anastomosing spines, quite similar
to and directly continuous with those of the ante-chambers.

The two entapophyses (ec. t. 8) have each the form of a
short, strongly compressed pouch, whose blind end is
directed upwards, backwards and laterad. The cavity in the
ventral part of each branches off from the interpulmonai’y
canal, and is provided, like the latter, with anastomosing

VOL. o4, PART 1. — NEW SERIES. 4

50

W. E. PURCELL.

spines {-spi., fig. 23a), while near its blind dorsal end it is
vvitlionb any spines (ec. t. 8, fig. 23a).

The hypodermis of the blind end is of special interest.
Its cells assume a fibrous structure fig. 23) and are not

pigmented like the adjacent hypodermis, and the large ento-
chondrite {t. 8) is firmly attached to their basal ends.

The numerons and powerful muscles which are attached to
this entochondrite have been described by Schimkewitsch
(’84), Vog’t (’89), A. Schneider (’92), and others. It cor-
responds to the anterior of the three pairs of abdominal ento-
chondrites described by these authors.

In females of Attus floricola, at any rate in matured or
nearly matured specimens, the interpul rnonary fold differs in
a very remarkable manner from that just described. Instead
of the cylindi-ical spinous canal of communication one finds a
broad, thin-walled, much wrinkled, band-shaped canal, with-
out spines inteimally, and strongly compressed from before
and behind {can., fig. 22). Moreover, the two walls of the
fold itself, apart from the canal, are much more strongly
wrinkled than is the case in the male.

In the adults of both sexes the opening of the genital
organs is found on the anterior wall of the medial reg’ion of
the fold between the pair of entapophyses {g. o., figs. 20 and
23b).

The interpulmonary fold in other spiders. — The interpnl-
monary fold ^ was found in all Dipneumonous spiders
examined by me, and in the majority of the genera resembled
the conditions occurring in either the male or the female
of Attns floricola (Marpissa $, Clubiona $, Age-
lena ?, Pisaura ?, Dolomedes 2, Melanophorad' ? ,
Drassodes ?, Zora 2, Linyphia 2, etc.).

’Phe pair of entapophyses are very variable in shape, even
in the same species ; thus out of five specimens of Tegenaria
domestica (fig. 21) examined, no two had the entapophyses
shaped exactly alike. In some forms these tendons may be

‘ This fold was known to Treviranns (T2), and is descriljed Ly
MacLeod (’84). Berteanx (’89) and others.

KESPIRATORY ORGANS LN ARANE®.

51

far apart (Attus sp., fig. 20), in others, again, close
together (Tegenaria domestica, fig. 21), or even fused to
a single, rounded, median lobe (Lycosa Darling!) . The
free end or ends are either sub-entire or else drawn out
into short finger-like processes (fig. 21). Except in the
Dysde ridse the entapophyses are more or less inclined
backwards.

The interpulmonary canal varies considerably, and is fre-
quently spined and even cylindrical in the female as well. In
the majority of cases it forms a canal of direct communication
between the ante-chambers of the pair of lung-books, as in
Attus floricola, but in the Lycosidm and in Philo-
dromus this is not the case, the interpulmonary fold
being rudimentary in the lateral part between the enta-
pophysis and the lung-book. In many Lycosidge this por-
tion of the fold is represented merely by a slight internal
thickening of the cuticula [interp. jld,, fig. 19a), but in Philo-
dromus there is a slight in-folding of the outer surface as
well (fig. 19b) without an actual lumen being formed. The
presence of these rudiments indicates that there was once a
well-developed fold connecting the pulmonary sacs with the
entapophyses, and that the pi’esent condition is a secondary
one.

In Argyroneta (J') the fold is well developed throughout,
but there is no canal of communication, the two surfaces of
the fold being closely apposed and without a lumen between
them (fig. 19c).

In the Dysderidae, too, there is no spinous canal of com-
munication, although in the median part the lumen may be
widened {interp.jid., fig. 40). In Dysdera and Segestria
the fold is deep and well developed between the pair of enta-
pophyses, but on the lateral side of these it is rudimentary and
not continuous with the spiracle. In Harpactes the fold is
much less deep than in the two other genera, and the entapo-
physes are hardly specially distinguishable at all, being merely
slightly deeper portions of the fold, to which the entochon-
drites are attached. The lateral portion of the fold is, how-

52

\V. F. PUECKLL.

ever, here directly coutinuous with the spiracles. The male
(but not the female) of Harpactes is also remarkable in that
the opening of the genital organs lies immediately in front of,
but separate from, the interpulmonary fold (p. o., fig. 40),
whereas in all other Dipneumonous spiders examined the
genital opening lies in the anterior wall of the fold.

In the Tetrapneumonous spider ^ examined I found no inter-

Text-fig. 2.

Crypsidromus intermedins. Ventral surface of abdomen.

//>.. opercula of lung-books ; .sp.. spiracles of lung-books ;
ec. t. 8 and 9., muscle insertions; fj. o., genital opening.
Magnified fi.

pulmonary fold connecting the .spiracles, but on the medial
side of the latter (but separate from them and from the
genital cleft) shallow depressions resembling stigmata in the
cuticula were observed (er.i. 8 and 9, text-fig. 2), which proved
in sections to be the places to which the entochondrites of the
ventral longitudinal muscles are attached. These rudimentary

‘ Specimens labelled “Crypsidromus intermedins, Paraguay,”
obtained from the Berlin Zoological Lal>oratory.

KESPIBATORY OEGANS IN AEANEiE.

53

eiitapopliyses (ec. t. 8, fig. 36) were similar in both pulmonary
segments.

The only other order possessing* the interpulmonary folds
is the Pedi palpi, in which these folds are very well deve-
loped in both pulmonary segments and much resembles
that of Dipneumonous spiders (see Tarnani, ’89 and :04, and
Piirner, ;04).

VIII. ’1’he Development of the Tkachea: and the Entapo-

PHYSES OF THE TkaCHEAL SEGMENT.

'I'he tracheal appendages are, as nearly as possible, the
exact counterparts of those of the pulmonary segment in the
earliest stages, up to, say, the period when the pulmonary
furi-ows begin to appear (compare ah. aj>p. 1 and 2 in fig. 4).
'I'lie post-appendicular groove {(jr.) extends along the whole
posterior side of the appendage (except, perhaps, as in
ajjpendage 1, at the extreme lateral part), but it does not
appear to be deeper laterally than medially.

In the stage with two pulmonary furrows (figs. 1, 5 and
5a), however, after the simultaneous subsidence of the epithe-
lium lying between consecutive abdominal appendages we
find that the post-appendicular groove is not almost obliterated
in its medial half {tr. *■., fig. 5), differing in this respect from
the corresponding groove of the pulmonary segment [yr., fig.
5). On the contrary the infolding containing the groove has
increased in depth along its whole extent, and continues to
deepen in the following stages in such a way that its blind
bottom is directed slantingly forwards {fr. .v., fig. 6a). This
in-folding is the tracheal sac.

If we examine a reconstruction of the appendage from the
inner surface (fig. 27) at this stage (when about five pulmonary
furrows are present and the mesodermal entochondrites
begin to be formed), we find a broad transverse ridge [tr. s.)
projecting into the body and nearly co-extensive with the
posterior side of the base of the appendage. This ridge is
the ectodermal in-folding which forms the tracheal sac. The

54

W. F. FUKCELF.

space (ar. 9) occupying the medial region of its visceral
surface and enclosed by the dotted lines in the figure is the
area with which the ends of the longitudinal muscles are in
contact, and to the deepest part of which the entochondrite
becomes attached. The medial area (ar. 9) of the tracheal
sac is, therefore, serially homologous with the corresponding-
area (nr. 8) behind the pulmonary appendage (see p. 20),
and has consequently nothing to do with the region in which
the earlier pulmonary furrows appear, nor with any portion
of the lung-books. It will be observed that owing to the
presence of the lung-leaves the area (ar. 8) in the pulmonary
segment is more widely separated from the segmental tube
{seg. t. 8) than is the case in the tracheal segment.

The lateral region of the ti’acheal in-folding is of especial
interest, as it is the only part which is serially homologous
with the pulmonary sac. It will be remembered that the
pulmonary sac proliferates in a lateral direction (position as
in fig. 1), later in a dorsal direction (position as in fig. 3), in
the form of a hollow tuber-like process creeping along the
inner surface of the outer epithelium; and that this sac and
its proliferations yield the cell-material for the formation of
the fourth and following pulmonary saccules.

Now the tracheal post-appendicular in-folding begins to
proliferate laterally simultaneously with the pulmonary sac
in precisely the same manner and direction. But the walls
of the tracheal sac have not to furnish cell material for lung-
saccules, of which no traces are present at any time, and, no
doubt, on this account the pulmonary sac rapidly outgrows
the corresponding tracheal sac, and in the stage of fig. 27
already greatly exceeds it in size. In this figure the groove
{tr. 1.) behind the tracheal appendage extends dorsally up to
section No. 16, while the proliferation extends through five
more sections; and the groove [imlm. 1.) behind the pulmonary
appendage reaches to section No. 24, while the corresponding
proliferation extends further likewise through five more
sections.

Figs. 35-35a and lOn-lbE represent longitudinal sections

RESFIBATOKY OECUNS IN AKANBJ].

55

through the tracheal and pulmonary proliferations respectively
of one and the same embryo from a series of sections similar
to those from which fig. 27 has been reconstructed and re-
presenting the same stage. It is to be noticed that the tracheal
proliferation {tr. proZ. ) is solid throughout, while that of
the pulmonary sac (pulm. *•.) is provided with a considerable
cavity, though this is, of course, not a fundamental difference
but is to be considered rather as due to a mere difference in
the rapidity of growth. In other respects both proliferations
closely resemble each other: in each the incision between it
and the outer epithelium is deepest on the anterior side and
dorsally at the apex. The opening of the tracheal sac does
not extend dorsally beyond section Ko. 10 (fig. 6a) of fig. 27,
the following five sections (compare figs. 35 and 35a) showing
no trace of a post-appendicular groove, exactly as in the case
of the corresponding five sections (Nos. 25-29) of the pul-
monary appendage. In both cases the dorsal ends of the
openings represent the latero-dorsal ends of the permanent
spiracles, the medial ends of which are still unformed.

Shortly after the stage I have just described the migration
of the three posterior pairs of abdominal appendages, already
alluded to on a previous page, commences. This process, which
may be considered as characteristic of all Dipneumonous
spiders with the tracheal spiracle near the hind end of the
body, consists of a double movement, namely, a medio-ventrad
movement of each of the three pairs of appendages and a
caudad one caused by the enormous elongation of the ninth
somite. Near the end of the reversion, as a result of this
process, these appendages come together in pairs in the median
line in the posterior half of the abdomen {tr. pL, figs. 41 and
43). At the same time the tracheal appendages gradually
sink to the level of the body surface.

During this period the formation of the tracheal spiracles
is completed, the lateral ends of the spiracles having already
been formed at an earlier stage. The unformed median ends
become approximated by the migration of the appendages
towards the median line, and subsequently the region of the

56

W. F. PURCELL.

body-epitlielium Iviiig in between {inf., fig. 28) folds into
the body, and the two spii’acles become united to a single one
{sp., fig. 28).

Meanwhile, important changes hare taken place in the post-
appendicular sac of the tracheal appendage. Fig. 28 repre-
sents a reconstruction seen from above of the ectoblast of the
two tracheal sacs (together with four muscles and two ento-
chondrites) at the end of the reversion. Fig. 41 is a sagittal,
and fig. 48 a transverse section of the same stage. The right
lialf of fig. 28 is equivalent to the ti'acheal in-folding [fr. s.)
in fig. 27, dorsal in the latter figure corresponding, of
course, to lateral in fig. 28.

We observe in the first place (fig. 41) the great longitudinal
elongation of segment 9, bringing the tracheal spiracle
and sac nearer to the posterior end of the abdomen. As if to
compensate for this backward migration the medial region of
each tracheal in-folding, that is, the region corresponding to
the area ar. 9, fig. 27, to wdiich the entochondrite is attached,
becomes drawn out in the form of an elongated plate (enta-
])ophysis, ec. t. 9, figs. 41 and 28), which is directed forwards
and slantingly upwards and is much compressed dorso-
ventrally.

8'he entochondrite (f.9) is attached to the narrowed anterior
end of the plate, and at the extreme posterior lateral corner
of the latter a second entochondrite (/., fig. 28) is found
attached to those cells which bound the lateral angles of the
spiracle. Between these two entochondrites the anterior
oblique muscle («. oh. m. 10) is stretched.

'I’lie bulging lateral jjortion {tr. prol.) of each plate cor-
responds to the dorsal proliferation of a previous stage
[tr. 2n’ol., fig. 27). In the line of the transverse section,
fig. 43 (see fig. 28), the lateral edge of the plate {tr. pil.) is
some little distance from the outer hypodermis ( hy.) but
more posteriorly, near the entochondrite t., fig. 28, the
edge of the plate comes nearly or actually into contact, with
the outer hypodermis.

The pair of plates are connected like the spiracle by the

KESriKATOKY OKGAN<>; IN AUANlvE.

57

in-foldiug {inf,, fig. 28) of the epithelium between them, and,
therefore, possess a common lumen, which, however, is con-
fined to the basal region only and denoted in the figures by
the area, tr. I, within the dotted lines. The greater part of
each plate is, therefore, solid. Fig. 28 gives the correct out-
line of the pair of plates, as they appear near the end of the
embryonic period, and I have tested the accuracy of the
reconstruction by comparisons with sections cut parallel to
the tracheal plates, so as to contain the whole width of the
])air of plates in one section. I could not detect a distinct
chitinous lining within the lumen of the plate at this stage,
for the cuticula which previously covered the body surface
always appeared quite loose and outside of the cavity, as if
the embryo were undergoing a moult such as Locy describes
for Ageleua.

If the tracheal and pulmonaiy appendages be now com-
pared, the diffei'ence in the relative development of the two
main organs connected with them becomes apparent. The
entapophysis is small in the pulmonai-y segment but large in
tlie tracheal segment, where it forms the greater part of the
tracheal plate, while the large mass of cells composing the
lung-book is represented by the comparatively small, lateral,
bulging portion of the traclieal plate.

The post-embryonic development of the tracheal plate. — After
the hatching of the embryo very important changes take
place in the shape of the tracheal plates. In the first place
the medial tendinal portion of each becomes drawn out in a
forward and upward direction to form an elongate, spathulate,
hollow process, which is strongly flattened dorso-ventrally
and much broader anteriorly than in the middle. Its shape
may be seen in fig. 29, wliich gives an accurate representation
of the pair of plates after the first post-embryonic moult
(stage 8). 'I'he tracheal lumen {tr. 1.) now extends to near
the anterior end, where it is also broader, but since the dorsal
and ventral surfaces are practically in contact (fig. 29a) this
portion can scarcely function as a respiratory organ at this
stage.

58

W. F. PURCELL.

In the secoud place^ the lateral tracheal proliferations
{tr. prol.) have also considerably elongated, but in a lateral
direction to form a broad Hattened lobe on each side. A
portion of the lumen of the trachea is continued into the
basal part of this lobe, and I have indicated the lumen by
the dotted lines (tr. L), as far as 1 could trace it with certainty,
but there are indications in the sections that the lumen
penetrates even further. It is extremely difficult to ascertain
the exact shape of the lateral ends of the lobes, as they are
wedged in between several other tissues, and it is just possible
that they are bilobed and not rounded as I have drawn them.
'I’hat portion which could be followed with cei’taintyis drawn
with plain lines, and the uncertain parts are indicated by the
dotted outline in tig. 29.

In the third place, a short basal portion has been added,
forming a hollow stalk or pedicel {pad.) connecting the whole
apparatus with the outer epithelium. This pedicel is
supported on each side by a chitiiious rod-like thickening {rd.)
in the form of a fold springing into the lumen from the lateral
edges of the chitinous lining and corresponding to the “ pro-
longeiiient chitineux” described by Schimkewitsch (’84, p. (36,
PI. ii, hg. 6) in the adult of hipeira.

'riie small entochondrite (b), which in tig. 28 is attached
to the hypodermis near the extreme lateral ends of the
spiracle, is now found a long way off from the spiracle. By
comparing the two figures it will be seen that the ento-
chondrite has not actually changed its position but that the
spiracle (.sy>.) itself has greatly contracted, being now, in fact,
less than half its former width, and thus the tissue bounding
its lateral ends now comes to lie some distance away from
the entochondrite. In shape this entochondrite {t., tig. 29)
has greatly elongated. It is broader towards the ends and
sle)iderer just behind the middle and is attached at its
posterior end (at x) directly to the hypodermis. To the
larger anterior portion three muscles are attached, viz. the
anterior oblique muscle (u. oh. 'in. 10) and two other muscles,
the medial and lateral spinner muscles {m. ,sp. m. 10 and

RESPIRATORY ORUANS IN ARANE^.

59

1. sp. m. 10), which pass posteriorly and attach themselves to
the medial and lateral parts respectively of the base of the
left anterior spinner. The same muscles are seen in fig. 28.
The smaller posterior portion of the tendon is further con-
nected with the lateral edge of the tracheal pedicel by a
small transverve column of cells [tr. vi.), apparently of a mus-
cular nature and plainly corresponding to the little tracheal
muscle found by Schiinkewitsch (’84, p. 66, PI. ii, fig. 6)^
and subsequently also by Lamy (:02, p. 160, PI. viii, figs. 4,
5) in the adult of Epeira. Schiinkewitsch considers these
muscles to serve the purpose of closing the lumen of the
tracheal pedicel, which in the adult, as well as in the young,
is strongly compressed dorso-ventrally. The lateral part of
the tracheal proliferation lies under the two spinner muscles
and the entochondrite, t., and the posterior edge of the
proliferation is apparently wedged in between the spinner
muscles and the transverse tracheal muscle.

'I'he lumen of the whole trachea at this stage is lined with
a smooth but strong cuticular membrane {cn., fig. 29a). The
great ventral longitudinal muscles {v. 1. m. 10) of the tenth
somite are stretched some distance above the trachea between
the entochondrites t. 9 and t. 10. 'I’he former of these
entochondrites is attached as before to the anterior end of
the tendinal portion of the trachea (er. t. 9), while the latter
lies above the spiracle and is attached to a long hollow
entapophysis from the posterior side of the anterior pair of
spinners.

After the second post-embryonic moult (stage 9) the trachete
appear for the first time as a fully functional respiratory
organ. In shape they are not much changed, except that
the lateral proliferations now branch at the ends into two
smaller trachejB, but beyond these I could not find any other
brauchlets at this stage. 'The chitinous lining is now covered
(except in the pedicel) with the palisades of hooped (anasto-
mosing) spines, ahso found in the adult spider, which keep

' In the figure the muscle is marked ep., hut in the text (p. 88) these
letters stand for the chitinous tliickening.

(K)

W. !■'. r UK CELL.

the liimeu permanently open and allow the air to circulate
freely through it.

The anterior (ventral) and posterior (dorsal) walls of the
pedicel are close together and lined with a smooth, stout,
chitinous membrane, but the two main tracheal trunks are
now connected by a spined intertracheal canal of communi-
cation, e.xactly resembling the similar canal already described
for the lungs.

I have no other stages between this and the adult form,
the chitinous skeleton of which is drawn in fig. 31, and may
be readily derived from the post-embryonic stages just
described. In fact the only essential difference between the
adult form and that after the second moult consists in the
])resence in the former of a large number of fine tracheal
tubules or secondary branchlets, which spring from the main
trunks either singly or in clusters, particularly from the ends
of the tendinal trunks and of the two branches of the lateral
trunks.

The entochoudrite {f. 9) of the earlier stages is now found
attached to the apex of a main tendinal trunk (w. tr.), which
is not continued beyond the entochondrite in this s])ecies
except in the form of a bunch of fine tubules.

In Attus, therefore, the two main tracheal trunks
[m. tr., fig. 31) are serially homologous with the pair
of entapophyses or ectodei'inal tendons (ec. f. 8, fig.
20), to which the entochoudri tes of the ventral
longitudinal muscles of the pulmonary and tracheal
somites are attached, and are actually homologous
w i t h t h e c o r r e s p o n d i n g e n t a p o p h y s e s of the second
]) ulmonary segment of Tet ra p n e n m onous spiders.

The lateral basal lobes (/. fr., fig. 31) of the ti achem
are directly derived from the lateral proliferation
of the earlier stages, and are serially homologous
with the pulmonary sacs of the previous segment,
and are to be considered as actually homologous
with the pulmonary sacs of the second pair of lung-
books o f T e t r a ]■) n e u m o n o u s spiders.

RESPIRATOKY ORGANS IN ARANE.E.

61

Critical remarke on the literature. — Schimkewitscli was the
lirst to figure a stage in the development of the trachea of a.
spider, for in his Russian paper (’86a) he gives a sketch (fig.
29a) of what is evidently the tendinal portion of the trachea
{ect.) and the entochondrite {L. 2) attached to it. I am
unable at j^resent to consult his principal paper on the
development of Spiders (’87), but apparently Schimkewitscli
failed to recogni.se the tracheal nature of the ectodermal
tendon, ect., which he considered to be a provisional struc-
ture, as is evident from the following remark in a later
paper (’94, p. 210) : “ Bei den Araueinen, wo das Endoskelet
im Abdomen fehlt, enstehen beim Embryo unter den hintern
Sehnen provisorische Ectodermfalten, die von mir auf fig. 11,
tab. 22 [’87] abgebildet sind.” By ‘^Sehnen” the author
refers to the entochondrites of the ventral longitudinal
muscles.

Simmons (’94) gives two figures of the developing trachea.
His earliest stage (fig. 8) is a sagittal section cut at a period
when the tracheal appendages are on opposite sides of the
embryo (my stage 5). It, therefore, represents a section
through the dorsal proliferation of the tracheal sac, and is, as
Simmons correctly claims, homologous with the pulmonarv
sac. On the other hand, his second figure (fig. 9), cut after
the reversion, evidently represents the tendinal portion of
the trachea, and cannot be the same structure as that repre-
•sented in fig. 8, ns Simmons claims it to be.

Simmons also claims to have found rudiments of the pul-
monary folds, and interprets certain undulations on the surface
of the embryonic trachea and two iu-pushings at its ends
(fig. 8) as such, but without, 1 think, sufficient justification
for doing so. Similar undulations may be found in Attus
floricola (e.g. on the posterior surface of appendage 2 in
fig. 5), which certainly bear a superficial resemblance to the
pulmonary folds in appendage 1, but these undulations are
produced by the mitoses of nuclei lying quite near the surface,
and may occur on any part of the body. They have certainly
nothing to do with pulmonary folds. Also, the two in-pushings

62

W. F. PUKCELL.

figured by Simmons do not resemble pulmonary folds; being
on the opposite sides of the tracheal tube, and they can
hardly be “ tracheal twigs ” as Simmons suggests, since the
lateral trachea? are, I believe, unbranched in the two forms
examined.

The Attus-type and similar types of tracheae in other spiders. —
The Attus-type of tracheae has been found in various other
genera of Attidm (Bertkau, Lamy), and is possibly the pre-
vailing type in this family.

A very similar type, not sharply separable from the
Attus-type, has been described by Bertkau and Lamy under
the name arborescent type of tracheae, on account of
the moi’e frequent branching of the twigs given off by the
imiin trunks. Other differences, according to Lamy, are
the presence of a spiral thread in the main trunks, and the
prolongation of these trunks into the cephalothorax. Such
trachea? have been found in the Uloboridse (Uloborus,
Lamy, :02, fig. 3, Miagram mopes, fig. 5), Prodidomidae
(fig. 26), Zodariidae (Zodarion, fig. 31), Clubionidae
(Anyphaeua, fig. 51) and Attidae (Ballus, fig. 67).^

In all these forms the main trunks probably represent the
entapophyses, while the small, branched, lateral lobe at the
base of each trunk is, no doubt, the i-udimentary homologon
of a pulmonary sac, exactly as in Attus floricola.

Lamy failed to recognise the homologon of the pulmonary
sacs in these lateral lobes in the arborescent and Attus-
types of trachea?, and supposed that here the ectodermal
tendons and the lateral trunks (representing the pulmonary
sacs) were completely fused together and no longer distin-
guishable. He also strangely misunderstood my statement
on the subject, for he quotes (:02, pp. 257 and 260) me as
having said that in the Attida? the homologon of the pul-
monary sac takes no part in the formation of the trachem,
which are entirely formed of the entapophyses, and he then
proceeds to dissent from this view.- My actual statement

' The figirres referred to are all in Lamy (:02).

■i Thus on p. 2fi0 he says: “ En tout cas, I'opinion de Purcell sur les

ItESriBATOBY ORGANS IN ARANE^.

63

was (’95, pp. 398, 399) : “ The homologon of the lung is
represented in the latter groups [Agelenidm, etc.] by the
lateral pair of tracheal trunks, but in the Attidse by a mere
rudiment in the form of a short lateral process on each side
at the base of the two large trunks.”^ There was, therefore,
no need to have differed from me as to the presence of the
homologa of the lungs.

The Agelena-type of tracheae and its development. — It was
shown long ago, first by v. Siebold (’48) and later by
Bertkau (’72, ’78) and Lamy (:02), that many families of
Dipneumonous spiders (about half of the genera examined,
according to Lamy, p. 227) possess a much simpler tracheal
system than that which occurs in the Attidae. This simpli-
fied system^ consists of four long trunks united behind at the
base, as in fig. 21, but without any of the fine secondary
tubules found in the Attidae.

Such tracheae occur in the Agelenidae, Clubionidee,
Drassidae, Argiopidae, Lycosidae, Theridiidae, etc., and
their relation to the Attus-type of trachea may be at once
seen by comparing fig. 21 (Tegenaria) with fig. 29 (young
Attus). Here the tendinal trunks [m. tr.) in the latter are
obviously equivalent to the medial pair of trunks in Tege-
naria, while the lateral branch {l.tr.) on each side in Attus
is represented by the pair of lateral trunks, which, therefore,
are serially homologous Avith the dorsal proliferation of the
embryonal pulmonary sac.

That this is really the case may also be easily shown from
the embryological material of Agelena labyrinthica in
my possession. Shortly before the hatching of the embryo
and after the completion of the reversion in this species, the
pair of tracheal plates have very much the same form as in

tracliees des Attidae, aux(juelles il donne une origine entiHement
entapophysaire, ne me semble pas acceptable.”

‘ On p. 248 Lamy cAirioiisly enough correctly quotes this statement.

® Literature : v. Siebold (’48, jj. 535), Leydig ('55, p. 460), Bertkau
('72. '78), Schimkevvitsch ('84), W. AVagner ('88, figs. 26, 67, and 68),
Vogt ('89, p. 226), Lamy (:00, :01, :02).

64

W. K. PUKCEU,.

Attus floricola (Hg. 28) at tlie same stage (stage 6).
They are, however, mucli further apart, and, therefore, with a
wider intertracheal infolding connecting them, the latei’al
proliferations being also more pronounced. Further, each
plate is much tliinner in the middle and lateral region than
at tlie base and along the anterior and medial mai’gins.

In embryos one to two days after hatching (stage 7) the
tendinal portion of each plate has considerably increased in
length and is, like the rest of the plate, very thin, except at
the apex, where it rather suddenly swells out and ends in a,
thick knob to which the eutochondrite is attached.

After the first post-embryonic moult (stage 8) the tendinal
portion of the trachea has much the same appearance as in
the previous stage, except that it has increased in length,
but the lateral proliferations have grown for some distance
in a lateral direction close to the hypodermal covering of the
body and are now provided with a distinct lumen in the form
of a very fine canal lined with chitin and communicating
with the spiracle. The chitinous lining both of this and of
the tendinal portion is smooth at this stage.

At the second moult (stage 9) the trachea assumes its
permanent shape. The chitinous lining, except in the pedicel,
becomes provided with the usual anastomosing spines and
the lateral proliferations increase considerably in length,
still growing in a lateral direction. The pedicel and the
canal of communication also appear. In fig. 30 (just before
the second moult) the hooped spines (ft'pt.) of stage 9 have
already appeared in I’eadiness for the moult.

Both in this stage and in the previous one the lumen of
the lateral proliferation {tr. prol., fig. 30), in its basal region
at least (i. e. near the pedicel), is eccentric, lying posteriorly
to the axis, the posterior wall of the trachea being much
thinner than the anterior wall, which contains nearly all the
nuclei. Towards the apex this wall becomes much thinner
and the lumen lies practically in the middle. This eccen-
tricity of the lumen is significant of the origin of the lateral
trachea, and may be at once understood if we remember that

EBSPIRATOKr ORGANS IN ARANE^.

65

the thicker anterior wall of the trachea is equivalent to the
anterior wall of the pulmonary sac together with the lung-
saccules produced by the latter.

Three forms of this type of trachea are mentioned by
Bertkau (’72), namely, those having : (1) the two median
trunks united at base to a short common tube, as in the
Theridiidse and some Argiopidae (Bertkau and Lamy) ;
(2) a medial and lateral trunk united at base in pairs on
each side to form two short common trunks, as in Tegeuaria
(fig. 21), this being the usual form, according to Bertkau
and Lamy; and (3) the four trunks springing separately
from the pedicel, as in some Argiopidae, e.g. Linyphia
(fig. 25).

The resemblance between this third form of
tracheal system (fig. 25), in which the lateral trunks
at first take a lateral course before running for-
wards, and tlie pulmonary system of an Attus (fig. 20)
is very striking, and clearly shows the homology
of the medial trunks {vi. tr.) with the eut apophyses
(ec. t. 8), and of the lateral trunks (/. tr.) with the
pulmonary sacs {pulm. ».). The parallel between the
spinous intertracheal canal of communication (connecting
the median trunks with one another and with the lateral
trunks at their base) and the interpulmonary canal of com-
munication (connecting the entapophyses with one another
and with the pulmonary sacs) is complete, as may be seen by
comparing fig. 24 with fig. 23 (sagittal sections through the
lateral region of the canal [can.'] along the lines indicated in
figs. 25 and 20), and fig. 26 with fig. 23b (median sections
along the lines indicated in figs. 25 and 20). From these
figures it will also appear perfectly clear that the
medial trunks of the tracheal system cannot be con-
sidered as branches of the lateral trunks any more
than the entapophyses of the pulmonary segment
are branches of the pulmonary sac.

In the Agelena-type of tracheie the medial trunks
generally take a fairly straight course as far as the euto-

VOL. .54, PART 1. NEW SERIES.

o

66

W. P. PURCELL.

cliondrite, where they may either terminate and tlius remain
comparatively short, as in Araneus (Epeira), according to
Lamy, or they may become longer and be continued beyond
the point of attachment on the lateral side of the entochon-
drite, often winding about for a short or even a considerable
distance further before coming to an end (Agelena, Te gen-
aria, Melanophora, Pachygnatha, Clubiona, etc.).
These long trunks are frequently bifid for some distance from
the apex, a character first observed by W. Wagmer (’88) in
Lycosa, and subsequently by Lamy (:02) in several other
forms (Agelena, Zora, Tib ell us, etc.).

The form with short medial trunks has been care-
fully studied by Lamy (:02), wlio was the first to describe
the method by which these trunks are attached to tlie ento-
chondrite. The tendinal trunks, according to Lamy, are
produced at their ends into a chitinous fibrous piece which
adheres to the entochondrite and is not furnished with a
spinons cavity and therefore presents “ absolument uu aspect
entapophysaire ou tendineux ” (:01b, p. 178). This fibrous
termination was observed by Lamy in most Theridiida;
aiid various Argiopidm (Liuyphia, Araneus, etc.).

The form of trachea with long medial trunks is very
widely distributed, but its mode of attachment to the ento-
chondrite has evidently eluded the observation of Lamy, for
he nowhere makes any definite statement nor gives any figure
regarding this point, except in the case of Tegenaria. In
this genus the medial trunks are said to terminate at the
entochoirdi'ite in the same fibrous process which was observed
in the Theridiidae, etc. (Lamy, :01b, p. 178), and one of the
trunks is figured as ending in such a process (:02, p. 213,
fig. 58).

I have examined five adult specimens of Tegenaria
domestica after treatment with caustic potash, and always
found the medial trunks evenly rounded off at the apex and
spined internally to the very tip, but without any trace of
terminal fibres. At a distance from the apex equal to about
two fifths of the whole length I found one or more short.

RBSPIKATOKY OKGANS IN ARANE/E.

67

fibrous, chitinous processes ijiy' fig. 42) attached to the medial
side of the trunk, which undoubtedly represent the terminal
fibres found by Lamy in Araneus, etc., and which connect
the trachea with the entochondrite. The part of the trachea
which is produced beyond the point of attachment is thinner
than the part posterior to the entochondrite, and may be
eitlier a single tube as in B, fig. 42, or it may consist of two
equal (left side, fig. 21) or unequal tubes {hr., fig. 42) produced
by the branching of the main trunk at the insertion of the
fibres. In one case each of the branches was again divided
so that the tracheal trunk then ended in four separate points.
There is no symmetry about this branching, for one side may
be branclied and the other not, but in all cases the branches
are lined internally with hooped spines right up to their ti])s,
differing in this respect from the ordinary secondary tubules
of the Attidae, etc., from which such spines are absent.

The only instance which Lamy mentions of a similar
tendinous fibre being attached to one side of a medial trunk
is the genus Chorizomma (;02, p. 219), in which, however,
the trachea) belong to a different type from the one we are
now discussing.

It is probable that the various forms of trachea) with
long medial trunks, whether branched or not, described by
Lamy in a number of families (Drassida), Argiopidm,
Thom i si d 86, Clubionida), Agelenida), Lycosidas, etc.),
all resemble one or'other of the variations of Tegenaria in
their mode of attachment to the entochondrite.

In Nephila, which Lamy reckons with the forms with
short medial trunks, I observed the tendinous fibres both at
the apex and also on the medial side at some distance from
the apex. This form may have, therefore, two places of
attachment.

In all cases these medial fibres, and a good part of the
terminal ones, are certainly nothing else but the intercelluhir
fibres usually produced by the hypodermal cells of an euta-
pophysis to connect the cuticula with the attached entochon-
drite or muscle (e. g. hi/., figs. .42, .‘36, etc.). They do not

68

W. F. PURCELL.

themselves constitute the eutapophysis, which is, of course,
formed by the entire ectodermal invagination — that is to say,
in this case the medial tracheal trunks.

The tracheae in the Dysderidse. — These tracheae, which have
been frequently described,^ were the first found in spiders
(by Leon Dnfour® in 1834, teste Bertkau, ’72, and Lamy,
:02), and ai’e of considerable interest from a comparative ana-
tomical point of view. I have myself examined sections of
Segestria, Harpactes, and Dysdera.

The tracheal spiracles of the Dysderidm are widely
separated, lying in the anterior region of the body a little
behind the pair of pulmonary spiracles (text-fig. 3, p. 69),
and entirely unconnected with one another. Each leads into
a large tracheal trunk, which rises upwards from the spiracle
and then runs forwards and bi'eaks up at its anterior end,
either in the pedicel of the abdomen or a little behind it, into
a larg-e bunch of fine secondaiy tubules. At the base a
shorter posterior trunk projects backwards, and also gives
off a number of fine tubules. The chitinous lining of the
trunks is provided Avith spines, which support a spiral thread
(Dysdera) or an inner perforated tube (Segestria).^ To
these well-known facts I have to add the following observa-
tions :

The segments of the venti’al longitudinal muscles belonging
to the tracheal somite are very short in this family, like the
somite itself, and the entosternite is attached on the medial

' Literature : Duges ['36, '49 ; v. Siebold ('48. p. 535) also cites the
following of the year 1835 : ‘ Feuill. Acad, des Sci. Seance du 9.
Fevr.,’ also Froriep's ‘ Notizen,' xliii, p. 231. also ‘ Ann. Sc. Nat.,’ at, p.
183], BertkaAi {'72), MacLeod ('80). Lamy (:02).

- 1 am unalde to find the reference to this pajAei-, unless it he ‘ le
Temps,’ No. 1942, cited l>y Menge ('51. jj. 22), Avhich, howcA-er, v. Siebold
('48. p. 5.35) accredits to A. Duges, both authors giving tlie year 1835,
and not 1834.

Bertkau (’72) states that in Segestria these spines do not anas-
tomose, but Lamy (:02, j). 183, fig. 23) has since shoAvn (and 1 can
coi’roborate his statements) that they certainly anastomose at their
ends, forming an inner, fenestrated, chitinous tul)C. In Harpactes
the anastomosing branches of the spines form a sinq)le network only.

KESPIRATOKY ORGANS IN ARANE^.

69

side of the base of each tracheal pedicel. In Segestria the
ectodermal area of attachment is drawn out in a mediad
direction in the form of a short, flat, unspined pouch (ec. t. 9,
figs. 32 and 33), whicli opens into the short, smooth, flexible
pedicel {ped.) connecting the rigid outer cuticula (cu.) with
the spinous main trunk {tr.) of the trachea. This enta-
pophysis is not respiratory, and the entire trachea is to be
regarded as homologous only with the embryonic pulmonary

Text-fig. S. Text-fig. 4.

Segestriii senocnlata, ?. Argyroneta aquatica, $.

Ventral surface of abdomen. — jmlm. sp., pulmonary spiracle; tr. sp.,
tracheal siiii-acle ; Ih, operculum of lung-books ; (j. o., genital opening ;
Magn. 12.

sac and its proliferation, as I have already stated in a previous
communication (’95). That this must be the case may fairly
be concluded from the position of the pair of spiracles (text-
fig. 3) corresponding to the second pair in Tetrapneumonous
spiders (text-fig. 2, p. ~)2) and from the position of the enta-
pophysis on its medial side. Lamy (:02) has also expressed
himself in agreement with this view, which differs entirely
from that of Bertkau (’72), who considered the short posterior

70

AV. ¥. rUL’CE],L.

truuks only to be equivalent to the lateral trunks in other
spiders.

The tracheae in Ai’gyroneta aquatica. — The highly-developed
tracheae ^ of this water spider are very peculiar. The two
main branches, as Bertkau (’78) showed, have a common
sjeiracle, which opens far forwards (text-fig. 4), close to the
pulmonary spiracles. The remarkable point about these
trachete is the circumstance that they lie entirely on the
medial side of the longitudinal ventral muscles. The seg-
ment of this muscle belonging to the tracheal somite is very
short, corresponding to the anterior position of the spiracle,
and it is stretched between two large entochondrites, the pos-
terior of which is attached to the upper surface of a short
basal process on the lateral side of the tracheal trunks. This
process, which is figured by Lamy (:02, p. 212, fig. 56), is
provided in its cavity with spines, like the main trunks, and
gives off at its apex a number of fine tracheal tubules (the
lateral bunch of trachem described and figured by Menge
[’51, dd, PI. i, fig. 7] and Lamy).

The process is flattened dorso-ventrally and corresponds to
the tendinal trunk of other Uipneumonous spiders. The two
large main trunks appear to be outgrowths from the medial
side of each very short tendinal trunk. They are joined at
their bases by an intertracheal fold provided with the usual
spinous canal of communication. On the posterior side of
each main trunk, near its base, is a transverse out-folding of
the tracheal wall forming a deep spinous groove ou the inside
of the trachea, connecting the canal of communication with
the lateral or tendinal branch. This transverse folding, Avhich
was described by Bamy (:02) as an abdominal trunk resembling
that in Segestria, also gives off numerous tubules, which,
together with another group just below, springing directly
from the main trunk, form the posterior bunch of tracheaj
figured by Menge (’51, ee., PI. i, fig. 7).

In Argyroneta, therefore, the entire tracheal

' Literature : Grube (’42), Menge (’51), Bertkau'.('78, p. 384). MacLeod
('80. "84), Lamy (.02).

KESPIRATOKY ORGANS IN ARANE^E.

71

system appears to be derived from the teiidinal
portion of the trachea, and there is no distinguish-
able trace left of the lateral trunks, ■which may be
homologised with the pulmonary sacd This leads us
to the conclusion that the ti'acheal systems of Argyroneta
aud the Dysderidse, although superficially closely resem-
bling one another, are yet apparently not homologous struc-
tures.

The tracheae in the Scytodidae, Palpimanidae and Filistatidae. —
The trachefe of these three small families possess a peculiar
interest, inasmuch as Lamy has shown that their medial
trunks are uon-respiratory and serve solely as entapophyses
for the attachment of the entochondrites. Bertkau (’78)
observed that the medial trunks were reduced to an unpaired
median rudiment in Scytodes, only the lateral ones being
developed, but our knowledge of the trachefe in the other
forms is due to Lamy (;00, ;01b, :02).

The most interesting is the tracheal system of Filistata,
of which I reproduce Lamy’s figure (:02, p. 173, fig. 12), as I
have no material of this family at my disposal. Here, accord-
ing to Lamy’s description, the spiracle is very broad and
placed about midway between the interpulmonary fold and the
spinners. The two short lateral trunks (Z. tr.) are pointed
sac-like and of the simplest form, exactly as a pulmonary sac
would appear if it lost its saccules. The four trunks are con-
nected at base by an intertracheal fold with spines in its
deepest part (which no doubt forms a canal of communica-
tion). The two entapophyses (ec. t. 9), too, have some internal
spines in their basal part, but are otherwise unspined, while
their free ends are jagged and tendon-like. If we compare
this text-figure Avith the figure of the pulmonary
system of Attus (fig. 20) and leave the saccules
out of account, the parallel between the two

' MacLeod’s (’82, p. 785, and ’84, p. 29) view that the trachea of
Argyroneta is nothing else than the dorsal chamber of the second
pair of lung-books of a My gale, enormously developed, is certainly
incorrect.

72

W. P. PUECELL.

respiratory systems appears complete and their
homologies almost self-evident.

In the Palpimanidm, according to Lamy, the medial
trunks are separate at least at their apex, while in the
Scytodidae they are confluent to the apex and form a single
median trunk.

1 have examined preparations in caustic potash and sections

Text-fig. 5.

Filistatii caiiitata Hentz. Tracheal apparatus (after Lamy). ec. <. 0.
entapophysis ; L fr., latei’al tracheal trunk. Magnified 100.

from both families. The long, lateral trunks are connected
by a canal of communication {can., fig. 38), lined internally
with hooped spines {spi.), which also spread into the basal part
of the median trunk. This latter is unpaired, and although
in my examples of Palpimanus there are indications of a
bifurcation at the apex, it is not nearly so prominent as in the
species figured by Lamy (;02, p. 187, fig. 29).

The median trunk is flattened dorso-ventrally and hollow

RESHRATOKY ORGANS IN ARANEA<].

78

internally nearly to the apex, but its chitinous lining is
plainly much too thick to allow it to be used for respiratory
purposes. In Scytodes by far the greater portion of this
chitinous lining is smooth internally (figs. 38 and 39), only a
small part quite at the base being spined {sjji., fig. 38), but iu
Palpimanus nearly one half is lined with hooped spines
(fig. 37a). In both genera the greater part of the unspined
portion of the entapophysis is in contact with the entochon-
drites {t. 9, fig. 37).

The histological structure of these tracheal eutapophyses
ajid of those of the pulmonary segment of Attus is quite
similar. The section through the basal half of the tracheal
entapophysis of Palpimanus (fig. 37a) should be compared
with the spinous part {spi.) of the pulmonary entapophysis
given in fig. 23a (in the latter the juatrix is not drawn iu),
while fig. 37 of Palpimanus is comparable with fig. 23 of
Attus, both passing through the places of attachment to the
entochondrites, t. 9 and t. 8. The same fibrous hypodermis
{hy.') and flattened smooth cuticula (c?t.) is observable in both
figures.

In his description of Palpimanus gibbulus Lamy says
there are two short medial apophyses without a spinous
lining (:02, p. 188), but his figiu’e clearly shows that the two
trunks are confluent for the greater part of their length and
separate only towards the apex. Lamy evidently considers
the confluent portion to be part of the vestibule. In other
places, too (:01b, p. 178; :02, p. 174), he states that in all
these forms the medial trunks ai’e reduced to the unspined,
terminal, tendinous (i.e. fibrous) part found at the end of the
medial trunks in Epeira, etc., by means of which the attach-
ment to the entochondrite is effected, while the whole portion
of the trachea in Epeira between the entochondrite and the
vestibule are said to be absent in Palpimanus. 1 cannot
consider this view to be quite correct, for the entire median
process in Pal piman u s and Scytodes, including the un-
paired part in the former and the spinous portion in both,
constitutes the entapophyses, and the spinous portion lying

74

W. F. rURCELL.

between the entoclioudrite and the vestibule is homologous
with the much longer but corresponding portion of the
medial tracheal trunks in Araneus, Tegenaria, etc.

Thus, the medial trunks in Filistata, Palpimanus
and Scytodes are homologous with the entire medial
trunks in Araneus, etc., and not merely with their
unspined, fibrous, apical portion, as Lamy suggests.

The unspined portion of the medial trunks in Filistata,
Palpimanus and Scytodes may well be compared to tlie
trachem of a young spider previous to the second moult
(staged), while this organ is still in its primitive spineless
condition. (Compare the transverse section, fig. 29a, of
the medial tracheal trunk of a young Attus with that of the
cuticular lining of the entapophysis of Scytodes given in
fig. 39.)

In the pulmonary segment the unpaired median entapo-
phvsis of a Scytodes has its exact parallel in the unpaired
median entapophysis of such forms as Lycosa Darliugi,
described on p. 51. We thus see all the variations of the
pulmonary entapophyses repeated in the tracheal segment.

IX. The Entapophyses of the Third and Fourth Abdo-
minal Appendages (The Spinners).

These tendons are unconnected with the respiratory org-ans
and need only be briefly described. They arise at a very
early stage, being formed out of the post-appendicular grooves
{(jr., fig. 4), which appear behind the third and fourth
pair of abdominal appendages shortly before the beginning
of the reversion (stage 1). At the time of the appearance of
the first pulmonary furrows (stage 2) these grooves have
deepened and become more pronounced (figs. 5 and 5a), and
they may be easily followed through all the later stages (fig.
6).^ At the end of the reversion they form invaginations {ec.

' I may point out that no trace of a lateral proliferation corresponding
to that of the pulmonary and tracheal sacs is ever found in connection
with these grooves.

KESIMRATOKY (ORGANS IN ARANE/E,

75

t. 10 and 11, fig. 41), which may exceed that of the pi’octo-
daeiim {proc.) in size. After moulting they form internal
cones or processes with a chitinous axis situated at the
posterior inner angle of the anterior and posterior spinners
respectively. At the stage of fig. 29 (stage 8, after the first
moult) the entochoudrite {t. 10), to which the anterior of
these entapophyses is attached, is placed just over the tracheal
spiracle, but is, of course, not attached to it.

The chitinous skeletons of the entapophyses of an adult
Tegenaria and their relation to the anterior and posterior
spinners are shown in fig. 21. These spinners are, of course,
the third and fourth abdominal appendages, but the middle
pair of spinners (m. spin.) do not, according to Jaworowski
(’95), correspond to a pair of appendages and have con-
sequently no entapophyses. An entochoudrite of the longi-
tudinal muscles is attached to the anterior part of each of
these entapophyses, the posterior of the three well-known
pairs of large abdominal entochondrites^ described by
Schimkewitsch (’84, p. 38) and others being that {t. 10)
which is attached to the entaj)ophyses of the anterior pair of
spinners.

The four pairs of serially homologous entapophyses {ec. t.
8-11) may all be seen in fig. 21. 'I’hey are, of course,
connected on each side by a longitudinal muscle, and the
positions of the four intermuscular tendons {f. 8-1 1) are
indicated in brackets. This figure may, therefore, serve to
give a general idea of the inter-relationship of all
these tendons of ectodermal (ec. f. 8-11) and meso-
dermal (f. 8-11) origin.

X. Gknekal Conclusions.

The embryological data furnished in the preceding pages
will, 1 believe, enable us to arrive at definite conclusions with
regard to certain questions concerning the phylogenetic

‘ These tliree entochondrites, marked I'l, /I, and ?3 in Schimke-
witsch's PI. vii, fig. 1, correspond to t. 8, t. 9, and t. lO respectively in my
fig. 41.

76

\V. F. FUKCELL.

origin of the trachea?, as well as of the lung-books in
A r a n e a?.

The origin of the tendinal or medial tracheal trunks in Ai’aneae.
— As the pair of ventral longitudinal muscles is a very primi-
tive structure, and must originally have been attached to the
outer hypodermis, it follows that the tracheal nature of the
tendinal or medial tracheal trunks must be a secondary
character, for if this were not the case we should have to
assume that all the ectodermal areas of attachment of the
ventral longitudinal muscles were originally derived from
trachese, since they are all seihally homologous, but this
would be an absurd supposition and quite contrary to the
facts of embryology and comparative anatomy.

I have also already pointed out that these medial trunks
cannot be considered as branches of the lateral ones, nor does
the embryological evidence show that they are otherwise
than independent metamorphosed entapophyses united at
their base with the lateral trunks by an intertracheal fold
and canal of communication, exactly in the same way as the
entapophyses of the pulmonary segment are united with the
pulmonary sacs by an iuterpulmonary fold and canal of com-
munication. The independent nature of the tendinal trunks
is obscured in the adults of such forms as the Attida? (fig.
dl), owing to the partial fusion of the rudiments of the lateral
trunks with the base of the medial ones, but it is clear
enough in most other forms. Even in such forms as
Segestria, Scytodes, and Palpimanus, where the enta-
pophyses have not been converted into trachem, they remain
attached to the smooth pedicel at the base of the lateral
trachem (Sege stria, fig. 32) or to the spinous canal of com-
munication uniting the two lateral trunks (Scytodes [fig.
38], Palpimanus), and do not shift their jiosition on to the
spinous part of these trunks. These forms, therefore, do not
provide us with any grounds for supposing that the spinous
parts of the medial trunks in other spiders have originated
as outgrowths from the spiuous part of a lateral trunk.
In fact, we have no other alternative, in view of

EESPlKATOliY ORGANS IN AKANE^.

77

both the embryology and comparative anatomy, but
to consider the medial trunks of the trachem as
equivalent in their entirety to metamorphosed enta-
pophy ses.

It is, moreover, a common feature in the Arachnida for
the ectodermal areas of attachment of various muscles to be
invaginated into the body in the form of pouches or tubes for
the purpose of serving as tendons, as, for instance, theentapo-
physes (ec. t. 10 and 11, fig. 21) of the two following- abdo-
minal segments already described.

In order that an ectodermal tendon may become converted
into a trachea it is only necessary that it should be hollow and
sufficiently thin-walled, with free access of air to its interior,
and that it should lie in blood or tissues requiring aeration.
It is also evident that a tendinal trachea must have existed
first as a simple entapophysis, since it could not possibly
function as a trachea until after it had attained a tubular
form. The entapophyses could not, therefore, have been
originally produced for respiratory purposes.

In the case of Araneae I have already sought to explain
the elongated tracheal entapophyses by the great elongation
of the ninth somite, and since the tubular entapophyses so
produced are hollow and lie in the large ventral blood sinus
(r. sin., figs. 41 and 43) we have here all the conditions
necessary for their conversion into a trachea. For it is well
known that the blood passes from this sinus to the lung-books
and thence to the heart, and that the sinus, therefore, contains
venous blood requiring aeration (Blanchard A9, ’50, Claparede
’63, Schneider ’92, etc.).

In the Tetrapneumouous spiders and in some Dysderidm
(Segestria) we find the rudiments of the entapophyses of the
ninth segment in the form of shallow depressions {ec. t. 9,
text-fig. 2, p. o2) or pouch-like invaginations (ec. f. 9, fig. 32),
already described on previous pages. These rudiments have
no respiratory function, and if they were to approach near to
the median line and be united at base by an iutertracheal in-
folding we should obtain the conditions found in Filistata,

78

\V. V. PURCELL.

etc. (text-fig. o, p. 72), and we have only to further imagine
these entapophyses lengthened and to become thin-walled and
provided internally with spines throughout in order to con-
vert them into the tendinal tracheae of other Dipneumonous
spiders. It is evident that the condition in the Tetrapueu-
monous spiders, at any rate, is a primitive one, on account of
the other primitive characters of this group, but the possibility
of a reversion from an elongated tracheal tendon back to a
very short one must be borne in mind, and may, perhaps,
occur in some Dipneumonous spiders in which the ninth
somite has secondarily become shortened again. I do not
think that this has been the case in the Dysderidm, how-
ever, on account of the primitive position of the tracheal
spiracles (text-fig. 3, p. 69) and other primitive characters in
this family, but in Argyroneta (text-fig. 4, p. 69) I believe
there is evei*y probability that the spiracle was once more
posterior and has subsequently shifted forward again to suit
a newly acquired, aquatic habit. This would account for the
fact that, although the actual tracheal entapophyses are
extremely short, they are lined with the usual anastomosing
spines and provided with a large medial outgrowth. This
outgrowth may originally have been merely a medial pro-
longation of the tracheal entapophysis beyond the ento-
chondrite, and when the spiracle moved forwai-ds and the
entapophysis shortened, its medial prolongation may have
increased in inverse proportion, so as to maintain the effective-
ness of the entire trachea as a respiratory organ.

It would certainly appear that the tendinal trunks are more
effective breathing organs than the lateral trunks are, pro-
bably on account of the position of the former in the great
ventral sinus of venous blood. For we frequently find the
tendinal trunks v'ery strongly developed, and the lateral ones
correspondingly reduced to a mere rudiment (Attidae) and
sometimes apparently to vanish altogether (Argyroneta).

The origin of the lateral tracheal trunks in Araneae. — I’he
second question to be considered is whether the pair of lateral
trachem of Dipneumonous spiders was derived from the second

EESPIKATORY ORGANS IN ARANE.E.

79

pair of lung-books of Tetrapneumouous forms or whether the
reverse was the case.

That the lateral trache® are serially homologous with the
pulmonary sacs of the preceding somite and, therefore, homo-
logous with the same part of the lung-books of the ninth somite
in Tetrapneumonous spiders, cannot, I think, be disputed,
although the embryology of the latter group is not yet
known.

Ill deriving the lung-books from tracheae the simplest theory
and the one that has been usually adopted by those who
favoured this view, is to consider the pulmonary sac or ante-
chamber to represent the main trunk of a trachea and the
saccules merely modified lateral branches arranged in a single
row and flattened by mutual pressure.

A very serious objection to this view lies in the appearance
of the two oldest pulmonary saccules on the embryonic ap-
pendages q uite outside of the pulmonary sac. These
two saccules cannot be branches of the main trunk, and in
order to account for their presence we should have to assume
that they themselves at one time each represented a separate
tracheal trunk. This, however, could hardly have been the
case, since all the saccules are formed in the embryo in exactly
the same manner (apart from their position out of or within
the sac) and should, therefore, have exactly the same phy-
logenetic origin.

Another view based by Jaworowski (^94) on embryological
grounds and adopted by Bernard (’96, p. 375) on theoretical
ones is to the effect that the lung-books arose by horizontal
folds in the basal part of a vertical tracheal trunk. Hei’e
also the appearance of the two oldest saccules, entirely out-
side of the pulmonary sac, is too strong an argument against
our acceptance of this theory, which, moreover, Jaworowski
lias failed to prove embryologically, as I have already pointed
out on a previous page (p. 33).

In fact the only way we can derive the saccules of lung-
books from tracheal tubes which appears to me at all feasible
is to assume that an ancestral form of the Araneae possessed

80

W. F. PURCELL.

abdominal appendages, on the posterior side of which were a
number of separate tracheje arranged in a row, and that these
appendages were sunk into the body in later forms. The
tracheated appendages of such an ancestral form would, in
fact, be very similar to one of the transitional stages which
Kingsley assumes for his theory of the origin of lung-books
from gills (p. 27), but it would be totally different from
anything actually found in the tracheal system of existing
spiders.

F rom purely embryo logical considerations, there-
fore, and quite apart from the branchial theory of
the origin of the lung-books, we have to assume that
the pair of lateral branches of the tracheae of the
ninth somite in Dipneumonous spiders must have
been derived from the pulmonary sac and not the
reverse. This conclusion is, moreover, strongly confirmed
by the fact that the Tetrapneumonous spiders, and particularly
the remarkable genus Liphistius, are more primitive in their
other characters than are the Tracheate spiders.

The origin of the secondary tracheal tubules. — The third
question is the nature of the tracheal branchlets, those fine
tubules {tr. tuh., fig. 31) given off by the main trunks in
certain forms (Attidte, Dysderidoe, Argyroneta, etc.).
It is usual to consider these as homologous with the saccules
of the lung-books, whatever view ^ may be taken of the origin
of the latter. I think, however, that this homology is, for the
most part, erroneous.

Since the pulmonary saccules occur only on the anterior
side of the pulmonary sac, we should expect to find the
tracheal tubules on the corresponding surface of the lateral
tracheal trunks, but this is by no means the case.” Thus, in

' Except, however, Jaworowski and Beniard.

- In the remarkable anterior paii‘ of trachea; of the Apneumonons
Family Caponiidse, described and figured in Simon (‘Hist. Nat.
Araign.,' 2e cd., i. pt. ii, pp. 326, 327, figs. 294 and 295, 1893) after
Bertkaii, the tubtiles are nearly all placed, however, on the anterior side
of an oval ante-chamber, and here, no doubt, do correspond to pulmo-

EESPIKATORY ORGANS IN ARANE/E.

81

the Dysderid®, in which the entire tracheal system is
probably derived from lung-books, we find the tubules arising
in a dense cluster from the apex of the elongate trunks
and from a small basal branch on the posterior side, but
none from the anterior or under surface of the trunks.
Further, in all tracheae of the Age! ena-type, which is that of
the majority of the Dipneumonous families, the lateral trunks
have no secondary branchlets at all.

On the other hand we find these tubules at various places
on the tendinal trunks in the Attidas amd other groups (fig.
31), which shows that the tubules may arise anywhere on a
tracheal trunk, when required, and quite independently of
the pulmonary saccules, since in this case they could not have
been derived from the latter. In Attus floricola there is
no embryological evidence that the tubules of the lateral
tracheal branches have anything to do with pulmonary
saccules, for whereas these latter commence to form in the
pulmonary segment at an early embryonic stage the tubules
do not appear until long after the young spider has been
hatched. It is, however, conceivable that the earlier lung-
saccules may have been entirely suppressed in the tracheal
.segment, so that only the post-embryonic lung-saccules
reappear as secondary tracheal tubules in certain cases, and
the possibility of the anterior terminal tubules of the Dysde-
ridte and those of the lateral lobes of the Attidas being of
this nature must be borne in mind.

Bertkau (’72, ’78) attempted to utilise the presence or
absence of secondary tubules as the basis of a system of
classification, but Lamy (:02) has shown that this character
has little value for this purpose, since within the same
family some forms may be provided with tracheal tubules,
while closely related forms are entirely Avithout them.

The origin of the lung-books in Arachnids. — A fourth question
in connection with this subject is Avhether the lung-books of

nary saccules, since the ante-clianiber is doubtless that of the pair of
lung-books which the tracheae have replaced. I examined sections of
C. spiralifera.

VOT.. o4, PART 1.

•NEAV SERIES.

6

82

'W. F. rUECELL.

tlie Araclinids were derived in the first instance from
tracheal books or from gill-books.

I have endeavoured to demonstrate in a preceding para-
graph that, since all the lung-saccules within the pulmonary-
sac precisely resemble in their formation and structure the
two oldest which appear outside of this sac, all the saccules
must have had the same phylogenetic origin and must
consequently all have originally been upon the posterior
surface of the abdominal appendage. The question, there-
fore, is whether the saccules of this primitive appendage in
the ancestral Arachnid were tracheae or whether they were
produced from sunken-in gill-lamellm. Whei'eas the appear-
ance of a number of tracheae in such a position seems most
improbable, the arguments in favour of tbe branchial origin
appear overwhelming. Most important amongst these, next
to the embryological evidence, is the undoubted general
agreement and affinity between Limulns and Arachnida,
first pointed out by Straus-Durckheim arid v. Beneden, and
afterwards so ably demonstrated by Ray Lankester. The
embryological side of the cpiestion and the probable manner
in which the transition from gill-books to lung-books may
have taken place has already been fully discussed (pp. 17-44)
and need not be considered again.

I shall only introduce here two figures of the abdominal
appendages of Limulus for comparison with the pulmonary
segment of a spider drawn in fig. 20. The appendages of the
genital segment (text-fig. 6), which are homologous with
those of the pulmonary segment, have no gill-books, but
possess the pair of genital openings [cj. o.), tvliich would lie
between the gill-books, if the latter were present. Text-fig 7
represents a branchiate appendage, and it will be seen that if
the gill-leaves were sunk into the appendage and the latter
into the abdomen, we should have exactly the condition found
in a spider (fig. 20). The large entapophyses {ec. t.) shown in
the text-figures are not, however, homologous with those of
the pulmonary segment {ec. t. 8, fig. 20).

fi’he endeavour to derive all trachem in Arthropods from a

KESPIKATOEY ORGANS IN ARANEiE.

83

common origin has no doubt weighed considerably against the
acceptance of the branchial origin of lung-books, but this
should not be the case in Anew of the undoubted diphyletic
origin of the trachem in Aranese, which, I think, I have suffi-
ciently demonstrated. Further, in one and the same spider
both parts of the tracheae, although of different origins, have
exactly the same histological structure, hence similarity
of structure in the fully developed tracheae does
not mean similarity of origin. I mention this here
expressly, since this similarity of structure has been used as

Text-fig. 6. Text-pig. 7.

Text-figs (! and 7. — Appendages of the genital segment and a
pair of abdominal branchiate appendages of Limiilns, seen
from behind (after Ray Lankester). ;/. o., genital openings;
hr., gill-book ; ec. t.. external opening of an ectodermal tendon.

an argument in favour of the monophyletic origin of all
tracheie.

The appearance in spiders of trachea) as newly acquired
organs derived from two separate and distinct sources simul-
taneously with the occurrence of other Avell-developed organs
of respiration clearly shows how readily trachea) may be
acquired.^ Why, then, should they not have originated

' Pocock ('93), who was of opinion that tracheal tubes replaced lung-
books at least twice in the gi-oup Aracbnida, viz. in the Dipneu-
mones and in the Pseudoscorpiones, remarks (p. 17); “ The fact
that these tubes have been developed tivice in the same group bears
veiy strong evidence as to their efficacy as breathing organs. They

84

AV. F. rUECELL.

equally readily over and over again in the Arachnida^ and
particularly in so large and diversified an assemblage as
the Tracheata? Thus in the Solifugfe the thoracic
trachem, which open at the base of the third pair of legs and
have always been an unexplained anomaly in view of the
branchial theory,^ may easily have originated from the entapo-
physes of some muscle. The same remark applies to the
occurrence of the remarkable pair of tracheal spiracles dis-
covered by Hansen (’93, p. 198), and subsequently confirmed
by Loman (’96) on the tibiae of the four pairs of legs in the
Phalangii dae."

I do not mean to imply that these abnormal tracheae were

must, in fact, be Ijetter adapted for their purpose than the hmg-book
tracheae.” This remark of Pocock's may possiidy explain why such
highly segmented forms as the Solifugmhave highly developed trachea;
only, since the extraordinary activity of the members of this group
would require the iJi’esence of the most effective breathing organs.
Bernard ('90, p. 374) mentions that these are the only Arachnids in
which the primitive tracheal tubes anastomose (as in the Insecta),
and to this I may add an observation which I have often made on living
Solpngidse, which is that regiilar and pronounced respiratory
movements are observable in the middle part of the body, especi-
ally after the animal has been running. Similar movements have not
hithei’to, so far as I am aware, been recorded for any air-breathing
Arachnids (see Plateau, '86).

' Bernard ('92, p. 521), for instance, remarks that the presence of
trachea; on the cephalothoi’ax in Arachnida is “one of the principle
difficulties in the way of those who would deduce the Arachnidan abdo-
minal trachea; from embedded gills. ... It compels us, for instance,
to assume that the cephalothoracic ti-achea; have had an entirely dif-
ferent origin, so that . . . it is necessary to assiune that the same

strncBires, tubular ti-achea;, have had two independent origins in the
same animal! . . . there is absolutely no difference between the

trachea; which open through the large stigmata of the thorax and those
opening tlu’ongh the more insignificant stigmata in the al)domen [in
the Solifuga;]. It is difficult to believe that they had a separate
origin. The embedded gill theory must, I think, definitely give way
before some simpler theoi'y, such as that here put forward.” So also
Weissenborn ('87, p. II4).

- It is interesting to note that Loman foiind these spiracles absent in
very young Phalangii da;.

EESPIRATOEY ORGANS IN ARANEJl,

85

probably muscular tendons. They may have bad any other
origin. Thus J. Wagmer (’94, p. 126), who has investigated
the embryology of Acari and admits the branchial origin of
hing-books, explains the cephalothoracic tracheae of Acari
and Solpuga by deriving them from unicellular hypodermal
glands, such as are found in water mites; while Borner
(:02, pp. 455, 461, and 463) considers it very probable that
the spiracle on the prosoma is that of the genital seg’ment
displaced forwards. Further, Ray Laiikester’s suggestion
that tracheae may have arisen by the tubefaction of meso-
dermal strands may apply.

The primitive nature of the lung-books in comparison with
the trachem within the class Arachnida is in full agreement
with the teachings of the comparative anatomy of other
org’ans. Thus we 6nd only lung’-books in the highly seg-
mented orders Scorpiones and Pedipalpi, tracheae in
the orders with more concentrated bodies. Op i Hones and
Acari, while in the Araneae the more primitive Tetra-
pneumones have lung - books . only, the more highly
specialised Dipneumones tracheae as well. The Solifugae,
however, which are highly segmented, have tracheae only
(see footnote on p. 84).

Bernard’s theory that trachem have arisen from bristle-sacs
of Chaetopod Annelids cannot be maintained for a moment as
an explanation of the lung-books or trachem of Araneae in
spite of the resemblance which the ectodermal tendons of the
tracheal segment in my fig. 41 bears to the bristle-sac with
its two oblique muscles figured by Bernard (’92, text-fig. 1,
p. 512). Indeed, Bernard does not attempt to derive enta-
pophyses from bristle-sacs, and the rudimentary spiracles
(vestigial stigmata) which he claims to have found in Pseudo-
scorpiones (’93a, p. 422, and ’93b, p. 26) and Pedipalpi
(’94, p. 151) are always placed by him on the lateral side of
the depressions caused by the dorso-ventral muscles of the
abdomen.

The homologies of the pulmonary segments in Arachnids. —
On the accompanying page I have given a table representing

lotnes of the Abdominal Appendages and their Derivatives.

80

W. V. PURCI-'Lt.

Argy ronetii not at all.

KESPIRATOEY ORGANS IN AEANEJ].

87

the homologies of the abdominal appendages of the eighth
to thirteenth somites in Limulus and the three pulmonate
orders of A.rachnida, based upon the most recent embryo-
logicad researches. The segmental homologies given in this
table agree with that of the same six somites given by Borner
( 02, pp. 456, 457), and may be taken, so far as the pulmonate
Arachnida are concerned, as sufficiently established, whereas
the homologies of these segments in most of the Tracheate
orders of Arachnida cannot be considered as satisfactorily
established, since the necessary embryological evidence is
wanting and that afforded by comparative anatomical research
insufficient.

The most important point in connection with this question
is the position of the genital opening.

(1) Aranem. — I have shown in a previous paper (’95)
that the genital ducts in Attusfloricola are formed out of
part of the coelom of the pulmonary somite and open extern-
ally into the interpulmonary (epigastric) in-folding, which lies
between the eighth and ninth somites. The genital segment
in Dipneumonous spiders is, therefore, identical with the first
pulmonary segment, which has been shown to be the eighth
post-oral by all the most recent investigators (Kishinouye,
’90; Simmons, ’94; Jaworowski, ’94; and myself, ’95).

(2) Scorjiiones. — Brauer (’95) has clearly shown that
the seventh somite in the scorpions had been overlooked by
previous authors, and that the genital operculum belongs to
the eighth somite, the pectines to the ninth, and the four
pairs of lungs to the tenth to thirteenth. Pereyaslawzewa
(;07) also places the four pairs of lung-books in the tenth to
thirteenth somites (pp. 174-176).

The homologies of the abdominal appendages in scorpions
and spiders given in the table on p. 86 may, therefore, be
considered as fully established by embryological evidence.

We have thus the remarkable fact, which I pointed out
before (’95), that none of the lung-books in scorpions
are actually homologous with the tw'O pairs in
spiders, and further, the two pairs of lung-books in spiders

88

W. F. rOL’CELL.

are represented by external appendages in the adult scor-
pion, and the two anterior pairs of lung-books in the latter by
external appendages in the adult spider. Now I cannot
imagine that the pectines of scorpions could have been
derived from appendages which had already sunk into the
abdomen and been converted into lung-books, and the con-
verse, that these external organs, after having lost their
branchial nature and acquired new functions could ever have
been converted in lung-books, is equally improbable. I con-
sider, therefore, that the lung-books of the scorpions
and those of the spiders must have been derived
from branchiate appendages quite independently
of each other, a ndt hat the terrestrial Arachnids are
not monophyletic but must have bad at least a
diphyletic origin from primitive aquatic Arachnids
with six pairs of abdominal branchiate appendages
on the eighth to thirteenth somites.^

Laurie (’93) has expressed a similar opinion but based on
palaeontological grounds, that the lung-books in scorpions
arose independently of those in other Arachnids.

(3) Pedipalpi. — It has been recently shown by Schimke-
witsch (:06) that in the embryo of Thelyphonus the lung-
books belong to the second and third abdominal somites
(p. 43), while the genital opening is found between these two
segments (pp. G3, 64), that is to say, exactly as in the
A r a n e ae .-

’ It is interesting to note in this connection that Schinikewitsch
('94, p. 207) discovered in the embryos of a scorpion on each side on the
genital opercnlum three to four teeth (Warzen, Km., fig. 12) which were
formed on tlie same plan as those of the j^ectines b\it vanished again
before birth. Schimkewitsch thinks it veiy probable that the genital
oijercnlum was once a sense organ like the pectines, and asks whether
both were not once gills f

2 Hansen ('83, p. 105) had previously pointed out that in Thely-
2)honus the first abdominal sternite should l)e sought for in the small
sclerite at the anterior end of the abdomen, so that the large anterior
sternite, which covers the genital ojjening and the first 2>air of lung-
books, would, according to Hansen, belong to the second abdominal

EESriKATOKY OEGANS IN AEANE^.

89

Lam-ie and Gough, who examined embryos of Phrynids, are
not quite clear as to the segmental position of the lung-books.
Laurie (’94, p. 34) states that the first pair belongs either to
the first or second abdominal somite, while the second pair
belongs to the third somite. According to Gough (:02,
p. G16) the lung-books belong to the first and second ab-
dominal appendages, but the author does not say to which
somites they belong. Pereyaslawzewa (:01), on the other
hand, describes distinct paired appendages on each of the
first five abdominal somites, the lung-books being formed
from the third and fourth pairs (p. 194).

In view of the definite statements made by iSchimkewitsch,
as well as of the anatomical evidence afforded by the adult
(see footnote on preceding page), and of the close relationship
which the Pedipalpi bear to the Aran etc, we may accept as
certain that the lung-books in the former group belong to
the second and third abdominal segments, i. e. the eighth and
ninth post-oral somites, and that the genital segment is the
second and not the first of the abdomen, as stated by Laurie
(’94). This would make the lung-books in the Pedipalpi
directly homologous with the corresponding ones of the
Araneaj, as represented in the table on p. 86.

I consider that the pulmonate Arachnids comprise two
distinct groups, which have separately originated from
branchiate ancestors, namely, (1) the Scorpiones, and
(2) the Aranete and the Pedipalpi. To the latter phylum
some, if not all, of the remaining orders of tracheate
Arachnida may perhaps be added, but 1 shall not at present
enter further into the relationships of these other orders.

Pocock (’93) has already expressed the opinion that the
Scorpiones, although the most primitive of all terrestrial
Arachnida, could not have been the ancestors of any other
orders of Arachnida, because the useful tail would not be
likely to be lost. Pocock, who based this opinion on grounds

segment. Recently Taruani (:04, text-figs, on pp. 51, 52, and 121) and
Burner (:02, :04) have also adopted this view.

90

W. F. PUECELL.

which are totally different from mine, accordingly divides
the Arachnida into two snb-classes, viz. (1) Ctenophora^
for the scorpions, and (2) Lipoctena for the remaining
terrestrial orders. Burner (:02, p. 459) in his paper on the
segmientation and general classification of the Arachnida,
accepts this subdivision, bnt on other grounds, viz., on account
of the difference in the number of the segments of the meso-
and metasoma which appear to exist between the Scorpiones
and the Lipoctena. Bdrner, however, considers tha.t both
Scorpiones and Lipoctena must have been derived from
a common ancestral group provided witii at least five pairs
of lung-books (pp. 459 and 463), but the difficulty (in my
opinion almost an impossibility) of deriving a lung-book from
a pectine, or vice versa, does not seem to have occurred to
him.

The question of the conversion of a sunken-in lung-book
into the external spinners of the Aranem would also present
difficulties, but these do not appear to me nearly so great as
in the case of the pectines, because the reconversion of the
lung-septa (lamellae) into external gill-like organs is not
involved. 1 have, however, already pointed out that no
trace of a lateral proliferation, corresponding to that of
the pulmonary and tracheal sacs, is found in the embryo of
Attus floricola, the entire post-appendicular invagination
becoming the entapophysis in these two segments. Moreover,
the spinning glands appear at quite an early stage (stage 5,
■sp. g., fig. 6) at the apex of the appendages, which always
remain recognisable as such to the end of the development.
In fact, they have every appearance of having been directly
develo))ed into spinning organs from external appendages
which were not sunken into the body, and, therefore, not
lung-books.

So far as our knowledge goes, therefore, we may
say that there is no evidence of any sort to indicate
that the spinners of the Aranem were derived from

* For which word the term Cteidophora lias been substituted by
Bonier (:02, p. 4b5).

EESriRATORY OliGAXS IX ARAXEAi].

91

sunken - in lung-books, or that the spinner - segments
ever possessed such organs in any ancestral form of
this o r d e r.

When abdominal segments bearing spiracles in other
Tracheate orders (Solitugae, Pseudoscorpiones, Opi-
Hones, and Acari) are homologised with those bearing
spinners in Araneae, as is done by Bbrner (;02, p. 457), the
difficulty of deriving spinners from lung-books should be
taken into account. For if the Lipoctena represent a
natural group and the tracheae leading from these spiracles
are derived from lung-books, as is often assumed to be the
case, it follows that the spinners in Araneae must also have
been derived from lung-books. But if we cannot admit the
latter derivation, then either some or all of these tracheae are
not homologous with lung-books (i . e . they are new forma-
tions), or else the segments bearing them are homologous
with the pulmonary segments in Araneae (and not with
those bearing spinners), or, finally, some or all of these
oi’ders maiy have originated independently of the Fed i palpi
and Araneae from branchiate ancestors (whether in connec-
tion with the Scorpiones or not is another question).

In the fSolifugm two (or at least one) of the three
tracheate segments of the abdomen must be homologous with
segments bearing spinners in Araneae, and a knowledge of
the development of the tracheae would be necessary before
one could determine the relaitionships of this order.

(4) Li m ulus. — According to Kingsley (’85) the genital
segment in the American species of Li m ulus is the seventh
post-onil segment, but Kishinouye (’91) has since discovered
an additional somite between the last thoracic segment and
the genital segment in the Japanese specie.s, thus making the
latter segment the eighth post-oral one. It is possible that
this aidditional seventh somite was overlooked in tlie American
Limulus, just as it has fre(^uently been overlooked in the
spiders and scorpions, for its presence would bring the
segmentation of the abdomen of Limulus into line with
that of spiders and scorpions.

92

W. F. PUrx’CFI.L.

Thus the homologies of the abdominal appendages in
Limulus may with the greatest probability be represented
as in the table on p. 86.

XL Historical List of Papers concerning the Lung-
books OF Arachnids (Exclusive of those Dealing
ONLY WITH Enervation, Embryology or Theoretical
Considerations.^)

Meckel (’09 and ’10) gives the earliest anatomical descrip-
tion of the lung-books of a scorpion and spider. He observed
the leaflets (saccules) attached to a stalk (pedicel and ante-
chamber) leading to the spiracle, and thought the stalk might
be hollow. He looked upon the organ as a real gill-book.

Gr. K. Treviranus (’12, ’16) describes the lung-books of
scorpions and spiders (as true gills) and the interpulmouary
fold and its muscles in spiders. He thought the “gills” may
be mainly organs for absorbing moisture from the air, while
the respiratory functions are carried out principally by four
jiairs of stigmata on the back of the abdomen (muscle
impressions) and four pairs on the sides of the cephalo-
thorax.

H. jM. Gaede (’23) describes the four lung-books of a
My gale (as gills) and observed the “granulation” on the
leaves. He thought tlie respiration took place, not in the
“gill-leaves,” but on the fine membrane behind them (i . e.
on the posterior wall of the ante-chamber, which is smooth
in some Tetrapneumonous spiders, e. g. in Cry])sidromus
inter m e d i u s) .

Johannes Muller (’28a, ’28b) discovered that the stalk
(pedicel), ante-chamber and leaves (saccules) in scorpions
and spiders are hollow by blowing air into the spiracle, and
so proved the pulmonary function of the lung-books. He

‘ The coiuiiarative anatomy of the lung-books is outside of the scope
of this paper, so I give this list in the form of an appendix for the use
of future workers on the subject, as it is more complete than any list
yet given.

EESPIEATOEY OEGAXS IN AEAXE^.

93

correctly surmises the passage of the blood between the
saccules and denies the presence of blood-vessels. This is
the most important description np to Leuckart’s time.

H. Straus-Durckheim (’28) describes the luug-books of
spiders, and says one may consider the saccules of these
organs in Arachnids as non-ramified tracheal trunks, repre-
senting merely a continuation of the external integument
in-folded into the interior of the spiracles (pp. 315-318).

J. F. Brandt (’33, p. 89) gives a poor description of the
lung-books of Epeira diadema (as gills), apparently
without knowledge of the work of the two previous authors.

A. Duges (’36, p. 181) injected spiders’ lung-books with
carmine. There is also a note on the lung-books in Duges,
’38, p. 5G8, teste Duvenoy (’40, p. 465).

G. L. Duvenoy (’40) describes the lung-books of spiders.

•1. van der Hoeven (’42) describes the lung-books of
Phyrnus niedius,^ calling them gills.

G. Newport (’43) describes the appearance of the lamellae in
scorpions and the circulation of the blood through the lung-
books (“ branchim ”) (pp. 295-297).

Pappenheim (’48) has a note on the lung-books of spiders.

A. Duges (’49) gives figures of the lung-books of Mygale
(PI. ii, fig. 8, and PI. iv, fig. 6), Segestria (PI. iv, fig. 5),
Pholcus (PI. iv, fig. 7), and Scorpio (PI. xviii, fig. 1/).

R. Leuckart (’49) describes the lung-books of scorpions
and spiders. He discovered the spines of the ante-chamber
in spiders and recognised the network on the leaves in
scorpions as a chitinons thread on the surface of the mem-
brane. He insists that lung-books are merely modifications
of tracheje (also ’48, p. 119 note), and his paper is the most
important that appeared before MacLeod’s.

E. Blanchard (’49, ’50) proved by injection that the blood
passes through the septa and thence to the heart.

A. Menge (’51) describes the lung-books of Argyroneta
(water spider). He failed to find any I'espiratory movements

' According to Kraepelin ('95, p. 41) v. d. Hoeveu’s species was in
reality Charon Grayi, Gerv.

94

W. F. I’URCELI-.

eitliei- in the lung-books or in the enclosed air, and observed
that the entire cnticula of these organs is shed at moulting.
He doubted their respiratoiy function.

F. Leydig (’55) found that the “ granulations ” observed by
previous authors in the lung-leaves of spiders are really
internal processes, like those in the posterior trachem.

L. Dufoiir (’56) describes the scorpion’s lung-books (pig-
ment and reticulation of the leaves, etc.).

E. Claparede (’63) describes the circulation of the blood,
with some notes on the lung-leaves, in the spider.

P. Bertkau (’72) gives a g’ood description of the lung-books
in spiders, and the earliest account of their growth in young
spiders.

C.Chun (’76), from a brief i-emark (p. 42), evidently implies
that he has found an epithelium with regular cell-boundaries
on the lung-leaves of Arachnids, but reserves the proof for a
later occasion.

H. Lebert (’77, p. 25) makes some very curious observa-
tions, such as his discovery of a second pair of smaller lung-
books (Xebenfachertracheen) in other spiders besides Tetra-
pneumones (e. g. in some Argiopidas); also bifurcate
saccules.

J. MacLeod (’80, ’82, ’84) advanced our knowledge greatly
beyond the works of his predecessors by the use of sections.
In his first paper (’80) he describes the lung-books as “ un
faisceau de trachees aplaties, foliiformes” (p. 48), but
influenced later by the branchial theory he re-casts his method
of Heating the subject (’82, ’84). His principal paper (’84)
is, perhaps, the best known of all works on the lung-books.

B. Bay Lankester (’81, ’85a, ’85b) in his first paper com-
pares the Inng-books of scorpions with the gill-books of
liimnlus from actual preparations, and derives lung-books
from gill-books by a theory. This paper (’81) affected most
subsequent studies of the subject, and made the development
and comparative anatomy of the respiratory organs a subject
of paramount interest in the Arachnida. His later papers
(’85a, ’85b) describe the circulation of the blood through the

IIESPIKATORY ORGANS IN ARANEAR

95

lung-bookS; and the histology of the lamellae in Scorpio, aTid
a new theory of the origin of the lung-books.

P. J, Mitrofanof (’81) makes some remarks on the lamellte
in Argyroneta (teste Schimkewitsch ’84, p. 64).

W. Schimkewitsch (’84) describes the lung-books of E peira,
without knowledge of MacLeod’s principal paper (’84).

F. Plateau (’86) searched for respiratory movements in
living Scorpiones, Araneae, and Opi Hones, with nega-
tive results.

W. Wagner (’88) describes the moulting and growth of the
lung-books in immature spiders.

L. Berteaus (’89) describes minutely the lung-books of
spiders and scorpions. This paper is the most complete on
the histological structure, particularly that of the cuticular
formations, and various errors made by MacLeod (’84) in this
respect are corrected. (His description of the form of the
lung-leaves and the ante-chamber is, however, unsatisfactory,
and is improved upon by Schneider [’92].)

J. Tarnani (’89) figures the topography of the two pairs of
lung-books in Thelyp bonus, and describes the interpul-
monary folds. In his later work (:04) these lung-books are
also figured and described (p. 121).

C. Vogt (’89) gives an original description of the lung-books
of E peira diadem a.

A. Schneider (’92) describes the circulation of the blood
through the lung-books, and gives an account of the general
structure of these latter organs in spiders. This very excellent
I>aper is indispensable as a supplement to Berteaux’s important
histological work.

M. Laurie gives an account of the structure and histology
of the lung-books in I^edipalpi (’94)^and of the difference in
the chitinous armature of the septa in different groups of
scorpions (’96a, 96b).

Sophie Pereyaslawzewa (:01) figures some sections of the
luiig-books of Bhrynidm (figs. 59, 62 and 64), and gives a
number of descriptive notes, especially on the histology of
the septa and on the pulmonary muscles (pp. 251-262).

06

AV. P. PURCELL.

A. Borner (:04) gives an account of the lung-books in
Pedipalpi, and some diagrams to illustrate their structure
in Arachnids generally.

List of Literature.

(Tlie titles of papers wliicli I have not seen are enclosed in hrackets.)

'92. Bernard, H. M. — “An Endeavour to show that the Tracheae of the
Arthropoda arose from Setipai’ous Sacs,” ‘ Zool. Jahrb., Ahth.
Anat.,’ V, pp. .511-524, 3 text-figs., 1892.

'93a. “Notes on the Chernitidae, with Special Reference to

the Vestigial Stigmata and to a New Form of Trachea.” ‘ Jonrn.
Linn. Soc. London,’ Zool., xxiv, pp. 410-430, Pis. xxxi and xxxii,
189.3.

'93b. “Additional Notes on the Origin of the Tracheae from

Setiparous Glands,” ‘Ann. Mag. N. H.,’ (0), xi, pp. 24-28, 189.3.

'94. “Vestigial Stigmata in the Arachnida.” ‘Ann. Mag.

N. H..’ (6). xiv. pp. 149-1.5.3. .3 text-figs., 1894.

'96. “The Comparative Moiphology of the Galeodidae.”

‘ Trans. Linn. Soc. London.' (2). Zool., vi.pt. iv. pp. 305-417, Pis.
xxvii-xxxiv. 1896.

"89. Berteanx, L. — “ Le Poumon des Arachnides,” ‘ La Cellule.’ v, pp.
‘2.5.3-317. 3 Pis.. 1889.

'72. Bei-tkau. P. — “ Uher die Respirationsorgane der Araneen,” ‘Arch,
f. Naturg.,' xxxviii, Bd. i. pp. 208-2.33, 1 PL. 1872 ; also ‘ Inaug.
Dissei’tation,’ -32 iip. (without plate), Bonn, 1872.

'78. “ Versuch einer natiiriichen Anordnung der Spinnen neh.st

Bemerkungen zu einzelnen Gattungen,” ‘Arch. f. Naturg..’ xliv,
Bd. i. pp. .351-410, PI. xii, 1878.

'49. Blanchard. li. — “ De I'aniiareil Cii’culatoire et des Organes de la
Respiration dans les Arachnides,” ‘ Ann. Sci. Nat. Zool.’ (3), xii,
pp. .316-.351. Pis. vi-viii, December, 1849.

'50. “ De I’appareil Circulatoire et des Organes de la Respiration

dans les Arachnides,” ‘ Compt. Rend. Acad. Sc., Paris.’ xxx, pp.
(i0_64, 18.50.

:02. Biirner, C. — “ Arachnologische Studien.” ii and iii, ‘ Zool. Anz.,’
XXV, pp. 43.3-466, 14 text-figs., 1902.

“ Beitriige zur Moi’phologie der Arthropoden. I : Ein

Beitrag zur Kenntnis der Pedipalpen,” ‘ Zoologica.’ Heft, xlii,
2 pts., 174 pp., 114 text-figs., 7 Pis., 1904.

04.

KESPIRATORY ORGANS IN ARANBAR

97

'33. Brandt, J. F. — “ Mediziii. Zoologie, od. getreue Darstellung u.
Besclireib. der Tliiere, die in der Arzeneiinittellelire in Betracht
koninien, etc.,” Von J. F. Brandt n. J. T. C. Ratzebni-g, ii, 44
Pis., 4to., Berlin, 1830-34 (Anat. of Lnngs of Epeira, by
Brandt, p. 89, 1833).

'95. Brauer, A. — “ Beitrage znr Kenntnis der Entwicklnngsgescliichte
des Skorpions, II,” ‘ Zeit. f. wss. Zool.,’ Kx, pp. 351—435, 20 figs..
Pis. xxi-xxv, 1895.

'86a. Bruce, A. T. — “ Observations on the Embryology of Insects and
Arachnids,” ‘ J. Hopkins Univ. Circ.,’ v, p. 85 ; also in ‘ Ann. Mag.
N. H.’ (5), xviii, pp. 74-76, 1886.

'86b. •* Observations on the Emluyology of Spiders,” ‘ Anier.

Natural,’ xx, p. 825, 1886.

'87. “ Observations on the Embryology of Insects and Arach-

nids ” (a memoi’ial volume), 31 pp., 6 Pis., 4to., Baltimore,
1887.

'76. Clmn, C. — “Uel)er den Ban, die Entwicklung und physiologische
Bedeutung der Rectaldriisen bei den Insekten.” ‘ Aldiandl. d.
Senckenb. Naturf. Ges.,’ x, pp. 27-55, Pis. i-iv, 1876.

'63. Claparcde, E. — ‘ Ktudes sur la Circulation du Sang chez les
Arances du geni-e Lycose,” ‘Mem. Soc. Phys. et Hist. Nat.,.
Geneve,' xvii, pp. 1-22, 1 PI., 1863; also ‘Ann. Sc. Nat. Zool.'
(5), ii, pp. 259-274, 1 PL, 1864.

'83. Dahl, Fr. — ” Analytische Bearbeitung der Spinnen Norddeutsch-
lands mit einer anatomisch-biologischen Einleitung,” ‘ Schr.
naturvv. Ver. Schleswig-Holstein,' v, pp. 13-86, Pis. i-ii, 1883.

'56. Dufour, L. — ‘‘ Histoire Anatomique et Physiologique des Scor-
pions,” “ Mem. presentes a I'Acad. Sci. Paris, Sc. Math, et Phys.,'
xiv, 4 Pis., i^p. 561-657, 1856.

'36. Dugcs, A. — “Observations sur les Araneides.” ‘Ann. Sc. Nat.
Zool.’ (2), vi, pp. 159-218, 358-360, 1836.

['38. “ Traite de Physiologic Comparee de I’liomme et des

Animaux,” ii, Montpellier, 1838.]

'49. ‘ Le Regne Animal,’ etc., par G. Cuvier, noxiv. (4®) cd..

Paris. ‘ Les. Arachnides,’ by A. Duges and Milne Edwards, 106

pp., 31 PL, 1849.

'40. Duvenoy, G. L. — “ Lemons d’ Anatomic Comparee,” par G. Cuvier,
2e cd., vii, Paris, 1840.

‘23. Gaede, H. M. — “ Beitrage zur Anatomic der Insecten,” ‘ Nova acta
phys.-med. acad. Leop. Carol, nat. cixr.,’ xi, pt. i, p2x. 323-340.
PL xliv, Bonn, 1823.

VOL. 54, PART 1. NEW SERIES.

7

98

W. F. FUKCELL.

:02. Gough, L. H. — “The Development of Adnietus pumilio : A
Contrilnitiou to the Embryology of the Pedijjalijs,'’ ‘ Quart.
Journ. Micr. Sci.,’ (2), xlv, pp. 5115-630, Pis. xxxii-xxxiii, 1902.

'42. Grube, E. — “ Einige Resultate aus Uutersuchimgen iiber die Ana-
tomie der Araneideii,” ‘ Midler's Arch. f. Auat. Phys. and Wiss.
Med.,’ Jg. 1842, pp. 296-302, 1842.

'93. Hansen, H. J. — “ Organs and Characters in Different Ordei’s of
Arachnids,” ‘ Entomolog. Meddalelser Kjbbenhavn,’ iv, pp. 137-
144, Pis. ii and iii, 1893 ; pp. 145-249, Pis. ii-v, 1894; also separate,
Copenhagen, 1893.

’42. Hoeven, J. van der. — “ Bijdragen tot de kennis van het geslacht
Phrynus Oliv,” ‘ Tijdschr. voor natuur. Geschied. en Physiol.,’
ix, pp. 68-93, Pis. i-ii, 1842.

['93. Jaworowski, A. — “Some Remarks on the Development of the so-
called Lung Trachete in Spiders,” Polish, 1893 (cited from
Jaworowski, '94).]

'94. “ Die Entwicklung der sogenannten Lungen bei den Arach-

niden und speciell bei Ti-ochosa singoriensis Laxm., nebst
Anhang iiber die Crustaceenkiemen,” ‘ Zeit. f. Wiss. Zool.,’ Iviii,
l)t. i, pp. 54-78, PI. iii, 1894.

'95. “Die Entwicklung des Spinnapparats bei Trochosa

singoriensis Laxm., mit. Beriicksichtigung der Abdominalan-
liiinge und der Fliigel bei den Insekten,” ‘ Jena. Zeit. Naturwiss,’
XXX, pp. 39-74, Pis. iii, iv, 1895.

’85. Kingsley, J. S. — “Notes on the Embryology of Limulus,”
‘ Quart. Journ. Micr. Sci.’ (n. ser.), xxv, jjp. 521-576, Pis. xxxvii-
xxxix, 1885.

'90. Kishinouye, K. — “On the Development of Araneina,” ‘Journ.
Coll. Sci. Univ. Japan,’ iv, pp. 55-88, Pis. xi-xvi, 1890.

'91. “ On the Develoiiinent of Limulus longispiiia," ‘Journ.

Coll. Sci. Univ. Japan,’ v, pp. 53-lUU, Pis. v-xi, 1891.

'92. Korschelt, E., and K. Heider. — “ Lehrbuch der vergleichenden
Entwicklungsgeschichte der wirbellosen Thiere,” ‘ Si^eciel. Theil,’
Hft. ii, i>p. 309-908 [Arthrojjoden], Jena, 1892.

"86. Kowalevsky, A., and M. Schulgin. — “ Zur Entwicklungsgeschichte
des Skorpions (Androctonus ornatus),” ‘Biol. Centralbl.,’ vi.
No. 17, pp. 525-532, Noveml.>er, 1886. [Also in ‘ Mem. (Sapiski)
New Russ. Natural Soc., Odessa ’ (Russian), xi, pt. i, 19 jjp.]

'95. Kraepelin, K. — “Revision der Tarantxdiden Fabr. ( = Phryniden
Latr.),” ‘ Abhand Ver. Naturwiss., Hamburg,’ xiii, ^xp. 1-53, 1895.

;00. Lamy, E. — “ Note sur I’appareil respiratoire trachcen des Ara-
neides,” ‘Bull. Soc. Ent. France,' No. 13, pj). 267-270. 1900.

RESPIRATOEY ORGANS IN ARANE^.

99

Ola. *• Sm- les diiferentes formes de I’appareil tracheen dans ime

meine famille d’Araneides,” ‘ Bull. Soc. Ent. France,’ No. 2, pp.
25, 26, 1901.

;01b. “ Siir la terminaison des trachees chez les Araneides,”

“Bull. Soc. Ent. France,’ No. 9, pp. 178, 179, 1901.

:02. ■* Recherches anatomicpies sur les trachees des Araignces,”

‘ Ann. Sci. Nat. Zool.,’ (8) xv, pp. 149-280, 71 figs.. Pis. v-viii, 1902.

’81. Lankester, E. Ray. — “Limnlus: an Araclinid,” ‘Quart. Jonrn.
Micr. Sci.’ (n. ser.), xxi, pp. 504-547, 609-649, 1881.

’85a. “ A New Hypothesis as to the Relationsliip of the Lnng-

hook of Scorpio to tlie Gill-book of LiinnliTs,” ‘ Quart. Joimi.
Micr. Sci.’ (n. ser.), xxv, pp. 339-342, 1885.

'85b. W. B. S. Benhain and E. J. Beck, “ On the Muscular and

Endoskeletal Systems of Limnlus and Scorj^io, with some
Notes on the Anatomy and Generic Characters of Scori)ions,”
‘Trans. Z. Soc. London,’ xi, pp. 311-384, Pis. Ixxii-lxxxiii, 1885.

'90. Laurie, M. — “The Embryology of a Scoi'pion (Eusc orpins
italicus),” ‘ Qua id. Jouru. Micr. Sci.’ (2), xxxi, jip. 105-141,
Pis. xiii-xviii, 1890.

■02. " On the Development of the Lung-books in Scorpio ful-

vipes,” ‘ Zool. Anz.,’ xv, pp. 102-105, 4 figs., 1892.

’93. “The Anatomy and Relations of the E urypterida;,”

■ Trans. Roy. Soc. Edinburgh,’ xxxvii, pt. ii, pp. 509-528, 2 Pis.,
1893.

'94. “On the Morphology of the Pedipalpi,” ‘ J. Linn. Soc.

London,’ xxv, pp. 20-48, Pis. iii-v, 1894.

'96a. “Notes on the Anatomy of some Scorjiions, and its

Bearing on the Classification of the Order,” ‘ Ann. Mag. N. H.'
(6), xvii, pp. 185-194, PI. ix, 1896.

'96b. “ Further Notes on the Anatomy and Development of

Scorpions, and their Bearing on the Classification of the Order,”
• Ann. Mag. N. H.’ (6), xviii, pp. 121-133, PI. ix, 1896.

'77. Lebert, H. — “ Die Spinnen der Schweiz, ihr Ban. ihr Leben, ihre
systematische Uebersicht,” ‘ Neue Denkschr. Schweizer. Gesell.
gesammt. Naturw.,’ xxvii, pt. ii, 1877.

'48. Leuckart, R. — ‘Ueber die Morphologie und die Verwandschafts-
verhiiltnisse der wirbellosen Thiere,’ 180 pp., Bi-annschweig, 1848.

’49. “ Ueber den Ban und die Bedeutiing der sog. Limgen bei

den Arachniden,” ‘ Zeit. f. wiss. Zool.,’ i, pp. 246-254, 1849.

'55. Leydig, F. — “ Zum feineren Ban der Arthropoden,” ‘Miillei''s Ai’ch.
f. Anat.,’ 1885, pp. 376—480 ; 4 plates.

100

W. F. PURCELL.

'86. Locy, W. A. — “Observations on the Development of Agelena
nmvia,” ‘Bull. Mns. Harvard Coll.,’ xii, pp. 63-lU3, Pis. i-xii,
Jannai-y, 1886.

'96. Loman, J. C. C. — “ On the Secondary Spii'acles on the Legs of
Opilionida),” ‘ Zool. Anz.,’ xix, pp. 221, 222, 1836.

'80. MacLeod, J. — ‘ La structure des trachces et la circulation pcri-
trachcenne ; Mem. couronne an concours universitaire de 1878-
1879,’ 72 pp., 4 Pis., Bruxelles, 1880.

'82. “ Recherches sur la strixcture et la signification de I’appareil

respiratoire des Arachnides (Comm. ])relim.),’’ ‘ Bidl. Acad. roy.
Belg.’ (3), iii, 11 text-figs., pp. 779-792, 1882.

'84. “ Recherches sur la structure et la signification de l'ai)pareil

respiratoii’e des Arachnides,” ‘ Arch. Biol.,’ v, pp. 1-34, Pis. i-ii,
1884.

'09. Meckel, J. Fr. — “ Bruchstiicke aus der Insecten-anatomie,'’
‘ Meckel’s Beitriige zur vergleichenden Anatomie,’ i, Hft. ii, pp.
105-131, Leipzig, 1809.

'10. ‘Cuvier, O. L., Vorlesungen iilier vergleichende Anatomie,’

iv ; iibersetzt ;i. mit. Anmerk. u. Zusiitzen vermehrt von J. F.
Meckel, Leipzig, 1810.

"51. Menge, A. — “Uber die Leliensweise der Arachniden,” ‘ Neueste
Schriften Naturf. Ges. Danzig.,’ iv, pp. l-(i4. Pis. i-iii, 1851.

'71. Metschnikoff, E. — “ Emljryologie des Scorpions,” ‘ Zeit. f. wiss.
Zool.,’ xxi, Hft. ii, pp. 204-232, Pis. xiv-xvii, 1871 ; also separate,
W. Engelmann, Leipzig, 1870.

'72. Milne-Edwards, A. — “Recherches sur rAnatomie des Limules,"
‘Ann. Sc. Nat. Zool.’ (5), xvii. Art. No. 4, 67 pp.. Pis. v-xvi,
November, 1872.

['81. Mitrofanof, P. J. — “ Sur I’Anatomie de I'Argyronete aquatique,”
‘ Mem. Soc. Amis Sci. natur.,’ xxxvii, 1881 (Russian, cited from
Schimkewitsch, '84).]

'87. Morin, J. — “ Zur Entwicklungsgeschichte der Spinnen,'’ ‘ Biol.
Centralbl.,’ vi. No. 21, pp. 658-663, January, 1887.

['88. “ Contrilmtions to the Embryology of the Spiders,” ‘Mem.

(Sapiski) New Russ. Natural. Soc. Odessa' (Russian), xiii, with
Plate, 1888.]

'28a. Midler, J. — “Beitriige zur Anatomie des Scorpions," ‘Meckel's
Arch. f. Anat. u. Phys.,’ pp. 29-71, 1828.

'28b. “ Ueber die Atheniorgane der Spinne,” ‘ Isis,' xxi, pp. 707-711,
Leipzig, 1828.

RESPIRATORY ORUAXS IX ARAXE^,

101

’43. Newport, G. — “ On the Stractare, Relations, and Development of
the Nervous and Circulatory Systems, and on the Existence of a
Complete Circulation of the Blood in Vessels, in Myriapoda
and Macronrons Arachnida — First Series,” ‘Phil. Trans.,’
1843, pt. ii, pp. 243-302, Pis. xi-xiv.

’48. Pappenheim. — “Note snr les ponmons des Araignees,” ‘Revue
Zool.,’ 1848, p. 250.

:01. Pereyaslawzewa, Sophie. — “ Developpement emhryonnaire des
Phrynes,” ‘ Ann. Sc. Nat. Zool. ’ (8), xiii, pp. 117-304, Pis. ii-ix,
1901.

■01 . “ Contributions a I’histoire du developpement du Scorpion

(Androctonus ornatus),” ‘ Ann. Sc. Nat. Zool.’ (9), vi, pp. 151-
214, Pis. iv-xvi, 1907.

'86. Plateau, F. — “De I’abscnce de mouvements respiratoires percep-
tibles chez les Arachnides,” ‘Arch. Biol.,’ vii, pp. 331-348, 1886.

'93. Pocock, R. I. — “ On some Points in the Morphology of the
Arachnida (s.s.), with Notes on the Classification of the
Croup," ‘ Ann. Mag. N. H.’ (6), xi, pp. 1-19, Pis. i-ii, 1893.

'95. Purcell. W. P. — “ Note on the Development of the Lungs, Entapo-
l)hyses. Trachea-, and Cenital Ducts in Spiders,'’ ‘ Zool. Anz.,’
xviii, pp. 396-400, 2 figs., 1895.

['71. Salensky, W. — ‘•Embryology of the Aranese,'’ ‘Mem. (Sapiski)
Kieff Soc. of Naturalists,’ ii, pt. i, jjp. 1-72, Pis. i-iii, 1871
(Ru.ssian). Abstract in ‘ Hofman u. Schwalbe’s Jahresbei’., fiber
Anat. u. Physiol.,’ ii, pp. 323-325, 1875.]

'84. Schimkewitsch, W. — “ Ktude sur 1' Anatom ie de I’Epeire,” ‘ Ann-
Sc. Nat. Zool.' xvii,’ pp. 1-94, Pis. i-viii, 1884.

'86a. “ Material towards the Knowledge of the Embryonic

Develo])inent of the Araneina,” (Russian), St. Petersburg, 1886.
[Cited from Jaworowski, '94.]

['86b. “ Les Arachnides et leur affinitcs,” ‘ Archives Slaves de

Biologie,' 1886. (Cited from Jaworowski, '94.)]

'87. “ Ktude sur le Developpement des Araignees,” ‘ Arch. Biol.,’

vi, pp. 515-584, Pis. xviii-xxiii, 1887.

'94. “ TJeber Ban uud Entwicklung des Endostei-nits der Arach-

niden," ‘Zool. Jahrb. Abth. Anat. u. Ontog.,’ viii, pp. 191-216,
Pis. x, xi, 1894.

03. “Ueber die Entwicklung von Telyphonus caudatus

(L.), ‘Zool. Anz.,’ xxvi, pp. 665-685, 6 figs, 1903. The same with
slight alterations, without figures, in Russian and Cerman under
the title “ Zur Embryologie der Thelyphonidje,’ ‘ Travaiix
Soc. Natural. St. Petersbourg Zool. et Physiol.,’ xxxiii. nn.
57-110, 190-i.

102

\V. F. I’UKCEI.L,

06. "UeEerdie Entwickluiig von Telyplioniis caudatiis (L.),

verglichen init derjenigen eiiiiger anderer Araclmiden,” ‘Zeit.
f. Wiss. Zool.,’ Ixxxi, pp. 1-95, Pis. i-viii, 1906.

'92. Schneider, A. — “ Melanges Araclinologiqnes," ‘ Tablettes Zool.

Poitiers,' ii, pp. 135-198, 16 Pis., 1892.

'48. V. Siel)old, C. Th. — “ Lehrbnch der vergleiclienden Anatoniie," von
V. Siebold und Stannins, i. ‘ Wirbellose Thiei’e,' von v. Siebold, 679
pp., Berlin, 1848.

'94. Simmons, O. L. — “ Develoi^ment of the Lungs of Spiders,” ‘ Amer.
Journ. Sci.’ (3) xl. pp. 119-128, 1 PL, 1894. Reprinted in ‘Ann.
Mag. N. H.’ (6), xiv, pp. 210-221, PI. vi, and • Tuft's CoU. Stud.,’
No. ii, jjp. 49-62, 1 PL, 1894.

'28. Straus-Durckheim. H. — ‘ Considerations gencrales sur I'anatomie
comparee des animaux articules, etc.,’ 435 + 36 pp., 10 Pis.. 4to.,
Strasbourg, 1828.

'89. Tarnani, J. K. — “Die Genitalorgane der Tbelyiihonus.’' ‘Biol.
Centrall)!.,' ix. No. 12, pjj. 376-382, w., 5 text-figs., 1889.

:04. “ Anatoniie de Thelyphonus caudatus (L.),'' ‘Meni.de

rinst. Agrononiique et Forestier il Novo- Alexandria.' xvi
Supplement, pp. 1-288, Pis. i-vi, 1904 (Russian).

12. Treviranus, G. R. — “ Ueber den innern Ban der Araclmiden,”
48 pp., 5 Pis., 4to, Nurnberg, 1812.

'16. Alihandlungen fiber den innern Ban der ungefliigelten

Insekten.” ‘ Verm. Schrift., Anat. u. Phys. Inbalts., von G. R. u.
L. Clir. Treviranus,’ i, pp. 1-84, 12 Pis., 1816.

'89. Vogt, C. — Article " Classe der Araclmiden," in ‘ Lebrbuch der
praktischen vergleiclienden Anatoniie, von C. Vogt and E.
Yung,’ ii. pp. 193-263, with 22 original text-figs., 1889.

'94. Wagner, J. — •“ Beitriige ziir Phylogenie der Araclmiden," ‘Jena.

Zeit. Naturw.,' xxix, pp. 123-156, 1894.

'88. Wagner, W. — ^“La mue des Araigiiees,” ‘ Ann. Sc. Nat. Zool.' (7),
vi, pp. 281-393, Pis. xv-xviii, 1888.

'87. Weissenborn, B. — “Beitriige ziir Phylogenie der Araclmiden,”
‘ Jena. Zeit. Naturw.,’ xx, jip. 33-119, 1887 ; also sejiarate as
‘Inaug. Diss.,’ 71 pp., Jena, 1886.

RKSI’IRATOHY ()R(iANS IN ARANKA'].

103

EXPLANATION OF PLATES 1—7,

Illustrating Mr. W. F. Purcell’s paper on '•'Development and
Origin of the Respiratory Organs in Araneae.”

Stages in the Development.

Stage 1 {St. 1), just befoi-e the appearance of the pulmonary furrows :
figs. 4, 7, and 7a (all from same emhi-yo).

Stage 2 {St. 2), with two pulmonary furrows: figs. 1, 5, 5a, 8, and
8a-8h (5, 5a, 8, 8a-8h all from one embryo) ; figs. 9, 10, 14.

Stage 3 {St. 3), with three pulmonary fun-ows ; figs. 2, 11.

Stage 4 {St. 4), with 4-5 pulmonary furrows : figs. 12, 15.

Stage 5 {St. 5), with 5-G pulmonary furrows : figs. 3, 6, Ga, and 27
(G, Ga, and 27 from one embryo) ; figs. 13, 13a, and 13b (all from one
embryo) ; figs. IG, 1Ga-1Ge, 35, and 35a (all from one embryo).

Stage G {St. G), after end of reversion and shortly before hatching :
figs. 17 and 43 (from same embryo) ; figs. 18, 28, 41.

Stage 7 {St. 7), after hatching : fig. 34.

Stage 8 {St. 8), after first post-embryonic moult : figs. 29 and 29a
(from same embryo) ; fig. 30.

Abbreviations for all the Plates.

The yolk is coloured yellow in the figures representing sections, all
of which have been drawn with the aid of a drawing apparatus. The
letters {St. 1). {St. 2), etc., alongside the numbers of the figures denote
the stage of the embryo from which the section has been made.

a. ob. m. 8 and 10. Anterior oblique muscles of somites 8 and 10.
«. spin. Inner openings of anterior pair of spinners, ah. app. 1-4. Ab-
dominal appendages 1-4. ant. Anterior side. ap. Apical pouch of
honi of pulmonary sac in developed lung-book. app. G. Sixth prosomatic
apiDcndage. ar. 7-11. Areas in contact with the ends of the segments
of the ventral longitudinal muscles of somites 7-11 at the time of the
formation of the entochondrites. h.f. Basal fold of tracheal trunk.
bd. c. Blood-coiqmscles. br. Branches of tracheal trunk, c. Cones of
a cJiitinous saccule, can. Canal of communication, cent. Centre of
section, cl. 1, cl. 2. Clefts on the distal side of first and second pul-
monary saccules. coel. G-14. Ccelomic sacs of 6th-14th post-oral
somites, cu. Cuticula. cu.', cu." Cuticula formed at first and second
post-emljryonic moults, d. 1. m. 8-15. Segments of the dorsal longi-
tudinal muscles in somites 8-15. d. v. 1. m. Longitudinal muscle along-
side of the proctoda;um uniting the last segment of the dorsal with that

104

W. F. I’UIIOKM,.

of the ventral longitnclinal muscles. d. v. m. 7-10. Dorso-ventral
miiscles hehind somites 7-10. dist. Distal side. dors. Dorsal side.
dors. (led.). Dorsal (originally lateral) side. ec. (j.d. ectodermal j^oi-tion
of genital duct. ec. t. 8-11. Ectodermal tendons (entai^ophyses,
apodemes) of the appendages of the 8th-llth somites, end. Endoderm.
ep. epithelium, f. \,f. 2, etc. First, second pulmonary fuiTOws, etc., in
the oi'dei' of their formation, (j. Genital cord. (j. o. Genital opening
into interpulmonary fold (to the outside in fig. 40). (jr. Groove behind
abdominal apj^endages. h. Horn (procurved end) of pulmonary sac.
horiz. pi. Horizontal plane of body. hy. Hypoderrnis. luj'. Fibrous
parts of the hypoderrnis of the ectodermal tendons, inf. Infolding of
the hypodermis. irdorp. fid. Interpulmonary (eiDigastric) fold or its
rudiment. 1. Lumen. 1. S2nn. m. 10. Lateral muscle to anterior side
of first spinner (in lOtli somite). /. tr. Lateral trunks of tracheal
system. lac. Lacuna. led. Lateral side. lb. Lung-book or tissue
forming it. m. Muscles or tissue forming them. m. sp>in. Inner
openings of middle pair of sj^inners. m. spin. m. 10. Medial muscle to
anterior side of first spinner (in 10th somite), m. tr. Medial trunks of
tracheal system, ma. Matrix cells, ined. Medial side, wicf/.p/. Median
plane, mes. y.d. Mesodermal part of genital duct. nv. <j. Nervous
ganglion of the pulmonary somite, op Opening at anterior end of
abdomen, p. oh. m. 8-11. Posterior oblique muscles of .somites 8-11.
2>. S2un. Inner ojienings of ijosterior ijaii’ of spinners, pcd. Pedicel.
2iost. Posterior side. pr. ax. Principal axis of appendage, proc.
Proctodaeuni. pidm. 1. Lumen of pulmonary sac. pidm. 2»'ol. Pul-
monary proliferation or gi'owing end of pulmonary sac. pulm. s. Pul-
monary sac. rd. Chitinous thickening on lateral side of tracheal i>edicel.
•s., s. 1, s. 2, etc. Pulmonary saccule ; first, second saccules, etc., in the
order of their formation, s'. New chitinous saccule forming before
the first post-embryonic moult, scy./. 8-11. Segmental tul)es of 8th-
11th somites, si. Slanting medial part at base of posterior wall of first
appendage, sp. Spiracle, sp. y. Spinning gland, spi. Anastomosing
spines, spin.m. lO. Muscles to the anterior side of first pair of spinners.
sp)z. Si>erma. st.p). Stercoral jmeket. t. Mesodermal tendon (ento-
chondrite). t. 7-11. Entochondrites at hind ends of the segments of the
ventral longitudinal muscles of somites 7-11. (t. 8-11.) Indicates the

position of these tendons, where not drawn in. tr. Tracheal trunk.
tr. 1. Lumen of tracheal plate or sac. tr. m. Ti-ansverse muscle on
lateral side of tracheal pedicel, tr. pi. Tracheal plate. tr.2Jrol. Tracheal
proliferation. tr. s. Tracheal sac. tr. tub. Tracheal tubules or finest
branchlets. v. 1. m. 7-11. Segments of the ventral longitudinal muscles
in somites 7-11. v. sin. Ventral sinus of abdomen, vc. Vacuole.
vent. Ventral side. vent. (nied.). Ventral (originally medial) side.
vest. Vestibule, vit. Vitellophagous cell. u\ Two-celled column, x.

TiESPIKATOKY ORGANS TN ARAXEiE.

105

Point at wliich the entochonclrite is attaclied to the liy]^)odennis.
y. Thvee-celled column, z. Pavement cell. w'. x' x' . if and y'. z' .
Anterior, distal, and jjosterior sides of apjDcndage.

PLATE 1.

Embryology of Attns f lor i col a.

[Longitudinal sections are cnt piirallel to the principal axis (pr. ax. in
figs. 1-3) of the appendage and at i-ight angles to the posterior margin
of the latter. Transverse sections are parallel to the principal axis and
to the posterior margin.]

Figs. 1-3. — (Zeiss oh]. C, oc. I, hot ale. snbl.) Transverse sections
showing the change of position of the first pair of abdominal appendages
during the reversion of the embryo. Fig. 1 I'e^n’esents the stage with
two pnlmonary furrows, fig. 2 witli three, and fig. 3 with four, five, or
more furrows.

Fig. 1. — (Zeiss C, 111, hot ale. snbl.) Longitudinal section tlirough
tlie lateral region of the four abdominal appendages, just ju'evious to
the appearance of the pulmonary furrows and the commencement of
the reversion.

Figs. 5 and oa. — (Zeiss C. III. Iiot ale. snbl.) Longitudinal sections
through the abdominal api^endages at tlie stage with two indmonary
furrows (/. 1,/. 2), con-esponding to fig. 1. Fig. o glasses tlu-ough the
medial, fig. 5a through the lateral region of the anterior appendages.

Figs. () and ()A. — (Zeiss C, III, hot ale. snbl.) Longitudinal sections
through the abdominal appendages at the stage with five or six pul-
monary fuiTows, corresponding to fig. 3. Fig. (i passes through the
medial, fig. (>A through the lateral region of the appendages.

Figs. 7 and Ta. — (Zeiss oil im.. II. hot ale. subl.) Longitudinal

sections through the medial (fig. 7) and lateral paid (fig. 7a) of the first
abdominal appendage (enlarged from the same embryo as fig. I) just
before the formation of the pulmonary furrows; cp., epitlielium (belong-
ing to somite 9) behind first appendage.

PLATE 2.

Embryology of Attns floricola.

[Longitudinal sections are cut parallel to the principal axis (pr. ax.
in fig. 1) of the appendage.]

Fig. 8. — Diagrammatic view of the posterior side of the first abdo-
minal appendage at the end of the stage (con-esponding to fig. I with
two pulmonary fuiTOws,/. !,/• -) (from a wax reconstruction). The
parallel lines represent the planes of sections; ep., the ejiitheliuni

106

W. !■'. rURCELL.

1‘epresented as cut along the groove {gr.) in figs. 8a-8d, and along the
deepest part of the pulmonary sae (pulm. s.) in figs. 8f and 8g.

Figs. 8a-8h. — (Zeiss oil iin., II, hot ale. suhl.) Longitudinal
sections through the first al>doniinal appendage, of which fig. 8 is a
reconstruction. Figs. 8a-8e pass through the medial and 8p-8h
through the lateral halves of the appendage, and their i^ositions are
indicated in fig. 8 ; ep., ejpithelium (belonging to somite 9) behind the
first al)dominal appendage.

PLATE 3.

Embryology of Attus floricola.

[Longitudinal sections are cut parallel to the principal axis (pr. ax.
in figs. 1-3) of the appendage.]

Figs. 9 and 10. — (Zeiss oil im., II, hot. ale. siahl.) Longitudinal
sections (from different embryos) through the medial region of the first
aI)dominal appendage at the commencement (fig. 9) and the end (fig. 10)
of the stage with two pulmonaiy furrows ; ep., epithelium (belonging to
somite 9) behind first appendage.

Fig. II. — (Zeiss oil. im., II, hot ale. suhl.) Section through the

medial region of the first abdominal appendage at about the com-
mencement of the 3-furrow stage, cut at a slight inclination to the
longitudinal axis of the appendage.

Fig. 12. — (Zeiss ^ oil im., II, hot ale. suhl.) Longitudinal section
through tlie lateral region of the first al>dominal appendage at the stage
with four to five pulmonary furrows.

Figs. 13-13b. — (Zeiss oil im., II, hot ale. suhl.) Transvei'se

sections cut parallel to the anterior side of the first abdominal appen-
dage at a stage with at least five well-develojoed pulmonary furrows,
fig. 13 being the second, 13a the fifth, and 13b the eighth section from
the posterior side of the ajjpendage (13 and 13a ai’e in outline and show
ectodermal tissue only), sp. The primitive sj^iracle ; s. 1-s. 5, the five
oldest saccules.

Fig. 14. — Sketch of first alKlominal appendage of the right side at
tlie stage with two pulmonary furrows, from a wax reconstruction, seen
from behind and distally.

Fig. 15. — (Zeiss oil im., II, hot ale. suhl.) Sagittal section (cut

parallel to the median plane of the embryo, c.f. fig. 2) thi’ough the first
abdominal appendage at the commencement of the stage with four
pulmonary furrows (slightly later than fig. 2).

RBSriilATOKY ()K(;ANS IX ARANEA'l.

107

PLATE 4.

Embryology of Attus floricola.

[Longitudinal sections are cut parallel to tbe principal axis (pr. ax.
in fig. 3) of the appendage.]

Fig. 16. — Diagrammatic view of the posterior side of first abdominal
appendage at the stage with five well- developed pulmonary fim-ows
(corresponding to fig. 3) from a wax reconstruction. The parallel lines
represent the planes of section ; ep., the epithelium represented as cut
along the line marked (ep., fig. 16) in figs. 16a-16d.

Figs. 16a-16e. — (Zeiss oil ii>i-> H- bot ale. subl.) Longitudinal

sections through the first abdominal appendage, of which fig. 16 is a
reconstruction.

Fig. 17. — (Zeiss oil im., 11, liot ale. subl.) Transverse section

tlirough first alidominal appendage shortly before tlie hatching of the
emlnyo, cut along the line indicated in fig. 18.

Fig. 18.— (Zeiss oil im., II, hot ale. sul)l.) Sagittal section of tlie
same stage as fig. 17 and cut along the line indicated in the latter
figure.

PLATE 5.

Fig. 19. — (Zeiss ^ oil im., I.) Sagittal sections through the cuticula
of the lateial part of the inteipulmonary fold (between the entapo-
pliysis and the lung-book), or its rudimentary remains in A, Lycosa
sj). ; B, Philodromus fuscomarginatus (subadult) ; and C, Argy-
roneta aquatica (adult ). The sections are all arranged in the same
positions.

Fig. 29. — (Caustic potash.) Attus sp. (adult). Chitinous skele-
ton of abdomen anterior to the pulmonary spiracles, drawn from
behind. The posterior wall of the ante-chamber has been removed on
the right side, and on the left the grate-like openings are visible through
this wall.

Fig. 21. — (Caustic potash.) Tegenaria domestica (ad. $ ).
Chitinous skeleton of abdomen from above.

Fig. 22. — (Zeiss oil im., I, picro-sulphuric acid.) Attus floricola
(mature, or nearly mature $ ). Sagittal section through the inter-
p\ilmonary fold between the lung-book and the entapophysis, showing
the cuticula only.

Fig. 23. — (Zeiss oil im., I, Flemming's sol.) Attus floricola

(ad. (5*). Sagittal section through the interpulmonary fold and the
entapophysis in the region indicated in fig. 20.

108

W. V. PURCELL.

Figs. 23a and 23b. — Similar sections of the same in the regions
indicated in fig. 20. Fig. 22a shows the cuticnla only.

Fig. 24. — (Zeiss oil im., IV.) Linyphia triangnlaris (ad. (J).

Sagittal section through the vestibule of the trachea in the region indi-
cated in fig. 25, showing the cuticnla only.

Fig. 25. — (Zeiss C, IV, caixstic potash.) Linyphia triangiilaris
(ad. ? ). Cuticnlar skeleton of basal region of tracheal system.

Fig. 26. — The same as fig. 24, hut in the median plane along the line
indicated in fig. 25.

PLATE 6.

Adult or snh-adult spiders.

Fig. 27. — (Zeiss F, I, hot ale. suhl.) Embryo of Attns floricola at
the stage with 5-6 pulmonary furrows. Reconstruction (made from
tlie same series of sections as figs. 6 and 6a) of the right pulmonary and
tracheal appendages seen from their inner side. The only mesodermal
elements shown are the two segmental tubes, sej/. f. 8 and 9. The
l)asal outlines of appendages 1-3 are indicated by dotted lines, as are
also the lumens of the pulmonary and tracheal sacs.

Fig. 28. — (Zeiss F, I, hot aqueous subl.) Embryo of Attus
floricola after the reversion. Reconstruction from transverse sec-
tions of the rudimentary trachea), together with the muscles and ento-
chondrites attached to the right half. The sketch is imagined as taken
directly from above, the anterior part being therefore higher in the
figure than the posterior part (cf. also figs. 41 and 43 of the same
stage). The rudimentary lumen is outlined by the dotted line.

Fig. 29. — (Zeiss F, I, hot ale. subl.) Young Attus floricola,
after the first post-embryonic moult. Reconstruction of the tracheal
system (imagined as taken directly from above) together with the
muscles and entochondrites connected with the right half and some of
those on the left. 29a (Zeiss oil im., IV). Transverse section
through a right medial trunk at the line indicated in fig. 29.

Fig. 30. — (Zeiss y^ oil im., II, hot ale. snbl.) Young Agelena
labyrinthica just l>efore the second post-embryonic moidt. Sagittal
section through a lateral tracheal trunk and the hypodei-mis below it ;
the cuticnla of the second moult is already formed.

Fig. 31. — (Zeiss C, III, caustic potash.) Attus floricola (ad. ? ).
Tracheal system. (The terminal portions of the secondary tubules are
not drawn in.)

Fig. 32. — (Zeiss C, III, equal parts of Flemming’s sol. and abs. ale.)

KESriEATORY ORGANS IN ARANE^.

109

Segestria senociilata (ad. $ ). Obliqiiely transverse section cut at
an angle of 38° to the horizontal (of. fig. 33) though the hasal part of a
traclieal trunk {combined from a couple of sections).

Fig. 33. — Similar to the last but a sagittal section, showing the
entapophysis cut across the line indicated in fig. 32.

Fig. 34. — (Zeiss oil iui., IV, Flemmings sol.) Embryo of Tege-

naria atrica, just after hatching. Transverse section through two
upper pulmonary saccules.

PLATE 7.

Figs. 35 and 35a. — (Zeiss oil im., II, hot ale. subl.) Embryo of

Attus floricola at the stage with five pulmonary furrows (from the
same series of sections as figs. I6-1()E). Longitudinal sections through
the tracheal appendage along the lines indicated in fig. 27.

Fig. 36. — (Zeiss jV oil im., I.) Crypsidromus intermedins.
Sagittal section through the rudimentary entapophysis or muscular
stigma of the first pulmonary sternite, showing the distal j^ai’t of some
of tlie long hypodermal filn-es (hij.') to which the entochondrite of the
pulmonary segment is attached.

Figs. 37 and 37a. — (Zeiss j'.j oil im., I, ale.) Palpimanus sp.
Transverse sections through the anterior (fig. 37) and tlie pf)sterior
(fig. 37a) regions of the median entapophysis of the tracheal system.

Fig. 38. — (Zeiss C, IV, caustic potash.) Scytodes testudo.
Basal part of chitiuous skeleton of tlie tracheal system.

Fig. 39. — (Zeiss oil im., IV, ale.) Scytodes testudo. Trans-

verse section through the ciiticular lining of the median entapophysis
of the tracheal system along the line indicated in fig. 38.

Fig. 40. — (Zeiss oil im., I.) Harpactes Hombergi (ad. (^).

Median sagittal section through the cuticula of the iuterpulmonary fold
and the genital opening.

Fig. 41.— (Zeiss 0, II, hot ale. subl.) Embryo of Attus floricola
after the reversion in sagittal section, showing the principal muscles,
ectodermal and mesodermal tendons and segmentation of the alidomen
(combined from several sections). The lungs, genital cords, and the
stercoral pocket (imagined as seen from the medial side) are di-awn in
to show their toi)ography, the last being represented in median section.
The muscle (p. ob. m. 8) is the only one lying between the genital duct
(fj.) and the lung-books (lb.).

Fig. 42. — (Zeiss A, II, caustic potash.) Tegenaria domestica
(adult). Portions of the pair of median tracheal trunks, showing the

110 W. F. PURCELL.

place of attachment to the entochondrite (seen from the dorsal side).
A, part of left trunk ; b, anterior part of right trunk (see fig. 21).

Fig. 43. — (Zeiss '^'1 im., II, hot ale. suhl.) Embryo of Attus
floricola after the reversion. Transverse section through the ventral
sinus of the al^domen in the region of the tracheal plate (along the line
indicated in fig. 28; same stage as figs. 28 and 41).

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DINOPHILUS, POLYGOEDIUS, ECHIUEUS, AND PHOEONIS. Ill

Notes on the Nephridia of Dinophilus and of the
Larvae of Polygordius, Echiurus, and Phoronis.

By

E. Ciioodricli, F.K.S.,

Fellow of Mei'ton College, Oxford.

With Plate 8.

In this paper are recorded some observations made on four
different types of true nephridia provided witli closed internal
ends bearing some form of solenocyte. Although all these
nephridia have been more or less completely described at
some time or other by various authors^ I am able to add
some details, not without interest, which help to complete our
knowledge of these organs.

Dinophilus.

The nephridia of this hee-swimming Annelid have been
described by Schmidt, Korschelt (7), Meyer (9), Mariner (5),
Schimkewitsch (11), and Shearer (14). It is stated by
Korschelt that in D. apatris, and by Meyer that in D.
gyrocil iatus, the internal extremity is blind, and ends in a
flame cell. In Manner’s excellent account of D. taeniatus
the end is said to lie in a cavity near the gut, and to bear a
ciliated appendage or knob forming the base of attachment
for the flame-like bunch of cilia which beat down the lumen
of the canal. Marmer is not positive as to the absence of an
opening.

In 1906 Shearer made the interesting discovery that the
“ ciliated appendage ” of Marmer is really formed of a number
of solenocyte tubes, comparable to those I have described in

112

E. S. GOODRICH.

many Polyclia3tes. These tubes had already been indistinctly
seen and described by Hariner as “ elongated, pear-shaped
bodies ” which “ vibrated individually.”

The blind internal end of the nephridium of D. tEeniatus
ends in a bundle of blind, slender tubular extensions, at the
extremity of each of which is attached a long flagellum. The
flagellum woi'ks down the tubule into the lumen of the nephri-
dial canal, which is not otherwise ciliated.

While able fully to confirm these important observations
made by Shearer on the living worm, I am also in a position
to complete his description from sections of well-preserved
material.

Fig. 1 is a slightly diagrammatic reconstruction, from
sections, of the end of the nephridium. These structures are
very small, and can only be made out with the highest
powers. The nephridial canal is relatively thick-walled ; the
cells of which it is composed are loaded with excretory
globules and pierced by an intra-cellular lumen. Nuclei can
be seen at r:u-e intervals along its course. One conspicuous
large nucleus is always found at the extreme end, which pro-
jects into a free space, supported by strands of mesenchyma-
tous tissue. Here the nephridial lumen expands into a
chamber bounded on one side by a thin wall from which
arise the “ solenocyte ” tubes. They may be considered as
hollow outgrowths of the wall. In sections they appear
rather shorter and stouter than in the living worm ; this is
probably due to contraction of the preserved material. Each
tube bears a little lump of })rotoplasm at its free extremity,
from which starts the flag’ellum. No nuclei are found on or
neai‘ ihe tubes themselves. The whole apparatus thus con-
sists of a single cell bearing some twenty or thirty tubes and
flagella, all controlled by the one large nucleus mentioned
above.

It is clear that in Dinophilus we have a new type of soleno-
cyte formation representing perhaps an intermediate step
between the Platyhelminth “ flame-cell ” and the more
typical Polychmte solenocyte, in which each tube, with its

DINOPHILUS, rOLTCORDIUS, ECHIUBUS, AND I'HORONIS. 113

flagellum, lias its own nucleus. This latter state might be
reached by a multiplication of the nuclei until they came to
correspond in number with the tubes. That Dinophilus is
related to Polygordius has long been suspected; it is there-
foi’e interesting to note that in the larval nephridium of
Polygordius there is a similar multiplicity of tubes, as I
showed some years ago (2).

The Larva op Echidrus.

It is to Hatschek that we owe the first detailed description
of the larval nepliridia of Echiurns (6). According to his
account there are a pair of larval nepliridia withont internal
opening. In the late larva there is a membrane separating
the coelom near the gut from a space below the body-wall
traversed by mesenchyniatous strands. Hatschek describes
the nephridium as passing inwards from its external pore
through tins space, to spread out in a system of fine branches
on the outer surface of the membrane. The nephridium ends
in “ein Hiishel von Ihidorganen,” and die Endknopfehen
der fei listen Caniilchen je einen Zellkern enthalten.”

'I'his description and the accompanying figures had long
led me to suspect that the “ Endkndpfchen ’’ were in reality
“ solenocytes.” It was not, however, till this spring that I
had an opportunity of confirming my suspicions by the exami-
nation of living larva3 at Naples.

Fig. 3 is a careful drawing from life of the nephridium of
a larva of about the a^e shown in Hatschek’s fisf. 4. A
glance at my figure shows that the nephridium is, in fact,
provided with typical solenocytes. The canal leading from
the external pore has granular, rather thick walls; when it
reaches the membrane outside the cceloniic epithelium it
divides into a number of thin-walled branches, which spread
over the membrane. These terminate in delicate, almost
cylindrical tubes, at the end of which are the nucleated cell-
bodies. 'I'he nuclei can be seen even in the living state.

As figured by Hatschek, fine protoplasmic threads extend
VOL. 54, PART 1. — NEW SERIES. 8

E. S. flOODlv'IOll.

1 14

from the cells, by means of which they are attached to the
surrounding structures. From each of the cell-bodies a long
flngellum passes down the tube into the canal of the nephri-
dium almost to the external pore. A few similar but shorter
flagella spring also from the wall of the canal itself. Sections
show that the lumen is inti-a-cellular, that nuclei are present
in the wall of the duct leading to the pore, but absent in the
fine branches. It is intei-esting to find that the solenocytes
project freely both into the mesenchymatous spaces on the
outside of the membrane, and into the coelom, passing through
the coelomic epithelium.

We may now add the Echiuroidea to the already long list
of groups in which are found typical solenocytes.

What may be the origin of the anterior kidney tubes of the
adult Echiurus is still quite unknown. Obviously they are
not derived from the larval organs, which open much farther
forwards in front of the large paired setee. Hatschek (6) and
Salensky (10) have given some acconnt of the development of
the posterior so-called anal kidneys, and these appear to be
not nephridia at all, but coelomoducts derived from the
coelomic epithelium. The more anterior kidneys of the adult
are probably of the same nature. I may add that I have
found no trace of the supposed opening of the duct of the
larval nephridium into the coelom mentioned by Salensky.

fi’HK Actinotkooha Lakva of Phouonis.

In a paper published in 1903 (3) I was able to show that
the larval nej)hridia of this larva project into the hmmocoele,
where they end blindly in bunches of solenocytes. It is
unnecessary for me to repeat here this account, or to again
refer to the older literature on the subject. But attention
may be drawn to several papers since published. Cowles (1)
and De Selys Longchamps (12) have confirmed most of my
statements with regard to the structure of the nephridium.
Moreover, De Sel3^s and Cowles agree with Ikeda (8) as to
the origin of the nepliridia from an ectodermal pit. But it

DINOPHILUS, POLYGOKDIUS, EOHIUKUS, AND PHORONIS. 115

is to Shearer (13) that we owe the first clear and definite
account of the development of the solenocytes themselves.
They arise from the wall of the ectodermal pit, as is con-
vincingly shown in his figui’es.

During a visit to Helgoland last year I was able again to
study the nephridium in a living Actinotrocha larva (A.
branchiata). In this form it is a large organ with abi’anched
internal end lying in the preseptal haemocoele. As shown in
fig. 4, multitudes of solenocytes project from the wall of the
nephridial canal. The nuclei are placed, not at the extreme
end of each tube, but rather at the side. Now it is well known
that an accumulation of blood-corpuscles is generally found
in the immediate neighbourhood of the nephx’idium in these
larvse, and the interesting new point I have to bring forward
is this — that very long, stiff cilia are set among the soleno-
cytes, attached by their base to the wall of the nephridium
between the solenocyte tubes.

Although I did not observe these cilia move very actively
in the larva compressed under a cover-slip, yet I have little
doubt that the}”^ agitate the blood-fluid, and so lead to the
gathering of the corpuscles just mentioned. I have described
almost exactly similar cilia on the nephridia of the Alciopids
(2), but in these Polychaetes they lie, of course, in the coelom.
Actinotrocha is the only form, so far as I know, in which
they occur on a nephridium projecting into a haemocoelic
space.

The Larva of Polygordius.

Since the figure of the nephridium of a larva from Ceylon
published in 1900 (2) w'as based on somewhat doubtful observa-
tions made on scanty material, I have here given a new and
more complete representation of the whole organ (fig. 2) as it
occurs in the Neapolitan species. The drawing was carefully
made from the living larvae of about the stage shown in fig.
8, PI. 11, of Fraipont’s well-known monograph (4). It will
be seen at once that the new figure confirms in every I’espect
the account I previously gave, in which it was first shown

116

E. S. GOODRICH.

that tlie blind iutevnal extremities are provided with soleno-
cyte tubes.

The stiff refringent solenocyte-tubes run free from the
protoplasm of the umbrella-like web, and are attached to it
only at one end, which may be called the outer end. The
opposite end of the tube plunges though the wall of the
nephridial canal, into which it opens. The flagellum passing
down the tube into the nephridial lumen is of remarkable
length. Thickened protoplasmic ridges pass radially along
the web from the central mass to the periphery, where they
embrace the outer extremities of the tubes. The apparatus
may be either expanded or contracted, as can be seen in the
figure. In the expanded condition the web is flattened out with
the tubes widely diverging; the central mass and contained
single nucleus then lies somewhat flattened in the middle.
When the web closes up, the tubes are on the contrary drawn
together until they become almost parallel, and the central mass
with its nucleus is made to bulge outwards as a convex knob.

There is no doubt, then, that in P. neapolitanus a single
nucleus, at the tip of each branch of the nephridial canal, con-
trols a set of from six to seven solenocyte tubes. If Woltereck’s
recent description of the nephridium of a North-sea larva is
correct, it would appear to differ considerably in structiu’e ; for
he states that there is a nucleus to each tube, that the tubes are
covered by the cytoplasm, and figures no web between them
(16). In an important paper on the development of Poly-
gordius. Shearer (15) has given a most careful account of the
origin of the larval nephridia ; they arise from two cells
differentiated quite early, lying on the inner surface of the
ectoderm, and probably derived from it. Each of these cells
multiplies to form a chain which develops into the whole
nephridium. T'he solenocytes arise from the extremity of
the nephridium itself. These observations entirely confirm
the view that the canal and solenocytes of the Annelid
nephridium form a whole, a single organ derived from one
rudiment, strictly comparable to the canal and flame-cells of
the platyhelminth excretory organ.

May 12th, IhOT

DINOPHILUS, POLYGOEDIUS, BCHIUETIS, AND PIIOEONIS. 117

Literaturk.

1. Cowles, R. P. — “ The Body-cavities and the Nephridia of the

Actinotrocha.” ‘ J. Hopkins Univ. Circ.,’ 1904; ‘Ann. Mag. Nat.
Hist.’ (7), vol. xiv, 1904; and Brooks and Cowles, “Phoronis
Architecta,” ‘Mem. Nat. Ac. Sci., Washington,' vol. x, 1905 (not
seen).

2. Goodrich, E. S. — “ On the Nephridia of the Polycha;ta,” part 3,

‘ Quart. Journ. Micr. Sci.,’ vol. 43, 1900.

3. “ On the Body-cavities and Nephridia of the Actinotrocha

Larva,” ibid., vol. 40, 1903.

4. Fraipont, J. — “ Le genre Polygordius,” ‘ Fauna u. Flora des Golfes

von Neapel,’ Monograph 14, 1887.

5. Hariuer, S. F. — “The Anatomy of Dinophilus,” ‘ Jouni. Mar. Biol.

Ass., Plymouth,’ vol. i, 1880.

6. Hatschek, B. — “ Ueber Entwick. Echiurus,” ‘ ArV). Zool. Inst.,

Wien,’ vol. iii, 1880.

7. Korschelt, E. — “ Ueber Ban u. Entwick. des Dinophilus Apatris,”

‘ Zeit. f. wiss. Zool.,’ vol. xxxvii, 1882.

8. Ikeda, 1. — “ Observations on . . . Actinotrocha,” ‘ Journ. Coll.

Sc., Japan,’ vol. xiii, 1901.

8. Meyer, E. — “ Studien ii. d. Korperbau der Anneliden," ‘ Mitth.
Zool. Sta. Neapel,’ vol. vii, 1887.

10. Salensky, W. — “ Ban den Echiurus Larva,” ‘ Mcmoires Ac. St.

Pctersbourg,’ vol. xvi, 1905.

11. Schimkewitsch, AV. — “ Z. Keniit. d. Banes u. d. Entw. des Diiio-

philus,” ‘Zeit. f. wiss. Zool.,’ vol. lix, 1895.

12. Selys Longchamps, M. de. — “ Devel. des Phoronis,” ‘ Mem. Ac. Sc..

Belgicpie,’ vol. i, 1904; and “Phoronis,” ‘Fauna u. Flora des
Golfes von Neapel,’ Monograph 30, 1907.

13. Shearer, C. — “ Dcvel. of Larval Nephridia,” part i : “ Phoronis,”

‘ Mitth. Zool. Sta. Neapel,’ vol. xvii, 1900.

14. “The Nephridia of Dinoiihilus,” ‘Quart. Journ. Micr. Sci.,’

vol. 50, 1900.

15. “Devel. of Larval Nephridia.” part 2: “Polygordius,”

‘ Phil. Trans.,’ vol. cxcix, 1907.

16. Woltereck, R. — “ Trochophora-Studien ; 1, Polygordius-Arten der

Nordsee,” ‘ Zoologica,’ vol. xiii, part 34, 1902.

E. S. GOODRICH.

1 18

EXPLANATION OF PLATE 8,

Illustrating’ Mr. E. S. Goodrich’s “Notes on the Nephridia
ot‘ Dinophilns and the Larvas of Polygordins, Echiurus,
and Phoronis.”

Reference Letters.

h. c. Blood-corpuscles, c. Cilium. c. m. Centi’al mass of cytoplasm
containing a single nucleus, c. t. Connective tissue, fe. Flagellum
inside the solenocyte tube. n. Nucleus, n. c. Neijhridial canal.
Nephridiopore. pr. Cytoplasmic process, Mass of cytoplasm

at end of tube. s. Septum separating the co3lom from the hajinocade
in which lie the solenocytes. t. Tube of solenocyte. w. Thin cyto-
plasmic web.

Fig. 1. — Restoration from sections of the inner end of the nephridium
of Dinophilns tseniatus. The organ is represented as if cut longi-
tudinally.

Fig. 2. — Drawing from life of the whole nephridium of the larva of
Polygordius neapolitanus.

Fig. 3. — Drawing from life of the whole nephridium of Echiurus sp.

Fig. 4. — Drawing from life of the inner end of the nephridium of
Actinotrocha branch iata.

. uo0.o‘t,iv.o.£Ko.a.

I

FURTHER NOTES ON A TRYPANOSOME.

119

Further Notes on a Trypanosome found in the
Alimentary Tract of Pontobdella
muricata.

By

yiiii'iel Kohertsoii, IVl.A.,

Ciirnct'ie Reseai’ch Fellow; Junior Assistant to the Professor of
Protozoology in the University of London.

With Plate h and 5 Text-figures.

Introduction.

In a previous paper published in the ' Proceedings of tlie
Royal Phys. Soc. of Edinburgh/ vol. xvii, No. 3, 1907, an
account was given of the main features in the life cycle of T.
raise as it appeared in the leech Pontobdella muricata.

I could not, however, at that time get clear knowledge of
the early stages of the infection in the leech, nor could I get
conclusive evidence that the form appearing in Ponto-
bdella was really Trypanosoma raite. The following
observations till in these important gaps, and appear to me
to make it clear that the flagellates found in such abundance
in this leech are derived from the skate and form part of the
life cycle of T. raiae.

I have also taken the opportunity afforded me by a fresh
set of infected leeches of again working through many of the
stages in that host. On this occasion I have abandoned the
Giemsa method and the dry fixation of the films, and have
used instead such fixatives as Schaudinu’s fluid, Hermann

120

JIUKIEL KOBEKTSOX.

and Flemming’s fluid. The stains used were iron liaema-
toxylin, Delafield’s lisematoxylin, Twort’s combination of
licht-griin and neutral red, and acid fuchsin. Tliese are all
useful stains. The Gienisa dry film method has been so much
criticised that one felt that cytological work carried out with
this stain was on an insecure basis until confirmed or criti-
cised by other methods.

The corrections in this paper apply only to cytological
detail, and my view of the life cycle, as a whole, has only
received further amplification from the new facts at my
disposal.

In the spring of 1907 I received a number of Pontobdella
cocoons from the Marine Station at Plymouth ; they had been
deposited on clam shells. I put them into a glass jar filled
with clean sea water, covered it with a loosely-fitting glass
lid, and simply put it on to a shelf in the laboratory. I went
to Ceylon in the course of the summer, and was later informed
that the leeches hatched out about the beginning of October,
1907, although the exact date was not noted. On my return
in the autumn of 1908 they were still alive in the original jar
and the same water. In November I took them to the
Marine Station at IMillport,^ on the Clyde, and fed them on
infected skate to obtain the first stages of the parasite in the
leech.

I may say in passing that the Trypanosome infection is
vei’y common in the skate caught in the Clyde area; more
than 50 per cent, of these fishes are infected, a circumstance
which accounts for the great frequency of the parasite in the
leech, only occasional specimens being uninfected.

The leeches, it was observed, showed the greatest excitement
— waving about actively in all directions — when a shadow
was made to fall upon the jar containing them. This reaction
to the appearance of a shadow is very marked indeed, and
must be of service to the creature in its natural state. The

' I wisli to expre.SK my thanks to the director of the station for
affording me all iiossihle facilities in carrying out these experiments.

FUETHEE NOTES ON A TEYPANOSOME.

121

skate, coming over the ground upon whicli a leech is attached,
will, of course, cast a shadow, and this will be one of the
ways the leech becomes aware of the presence of its host.

The young leeches attached themselves readily enough to
the skate, except in one or two cases where they required a
good deal of coaxing; ultimately, however, they all fed.

The Pontobdella appai'ently does not feed at once, possibly
on account of the difficulty in piercing the skin. The attitude
of the leech when attaching itself with the intention of
feeding is very characteristic. It takes firm hold with its
posterior sucker, and then bends its anterior sucker in so
that it is touching its own body. It then slides the anterior
sucker down along the body, finally attaching it to the skate
at a spot close beside the posterior sucker. These leeches
made quite a large wound for such small creatures.

In some cases, after feeding for a time, the leech detaches
the anterior sucker, but does not, as a rule, remove the pos-
terior one. Finally, after a greater or less interval of time
has elapsed, the feeding is resumed. On one occasion I was
able to observe the pimcess very clearly. A leech was put on
to a skate at 2.30 p.m., on the 14th November, 1908; at
10 a.m. on November 15th it was attached by the posterior
sucker only. 'I'he leech had fed, but had not taken very
much (the leech is, of course, rather transparent when young,
and it is possible to tell at a glance if it lias fed or not).
Three quarters of an hour later it was still in the same posi-
tion, with the anterior sucker free. I then held the skate
under water in the tank in such an attitude that 1 could see
the leech, which had begun to make searching movements
with the anterior sucker. It presently slid the anterior
sucker down its body and on to the skate, but moved it
about until it passed over the old wound, when it buried its
sucker in it. Tlie leech seems to force its proboscis in pretty
deep, judging from the wound and its attitude while sucking.
At 7 ]).m. on the 15th the leech was again only attached by
its posterior sucker, but it was much swollen, and had blood
right up to the anterior end. It was left attached to the

122

>rUKIEL ROBEFrrSUN.

skate till the moniing of the IGth, but did not seem to feed
again.

Anyone who has reviewed a large number of a given
species of leech will have observed that there is a good deal
of individual variation in the processes of digestion. The
broad lines are of course the same, but there is always a
certain amount of individual idiosyncrasy. This in Ponto-
b del la is chiefly expressed in the greater or less fluidity of
the blood in the crop and the nature of the bacterial flora
moulds, schizomycetes, etc., present. These complex circum-
stances no doubt react upon the Trypanosomes and may
explain a certain variability in the detail and also in the time
co-efticient of some of the developments.

The blood in the crop of Pontobdella has a tendency to
coagulate. It forms a rather dry mass with fluid in the
interstices. The time factor in this stiffening of the blood is
rather variable. It always occurs, but the time at which it
happens and the length to which it goes differs a good deal
in individual specimens. Late in digestion the mass in the
crop tends to become fluid again.

In Pontobdella the crop is a single rather thin-walled
sac passing back from the oesophagus right to the posterior
end of the body. The opening from the crop into the intes-
tine is placed at a point about two thirds of the way from the
anterior end and it passes back as a narrow tube lying on
top of the crop.

The young Pontobdella which had been hatched in cap-
tivity were opened at different intervals after feeding on
infected skate.

The condition of the parasites was carefully observed and
the following course of development was found to take place.
It must be observed in ])assing that skaters blood contains an
immense number of leucocytes, which present very con-
fusing appearances; also the parasites are rather scarce until
the multiplication period sets in. ksome searching is there-
fore required to find the Trypanosomes in the earliest stages.

FURTHEil NOTES ON A TEYl’ANOSOJIE.

123

A leech (5a) was put on to an infected skate at 9.30 p.m. one
evening; 12 hours later it was feeding, and had already
ingested a good deal of blood ; 2-T hours later it had ceased
feeding. The leech was opened I?! hours after it had been
first ])iit on to the skate, aud 3^ after it had finished feeding.
So that the earliest ingested Trypanosomes had been in the
leech about 16 — 17 hours, and the last ingested ones about
3 — 4 hours.

The blood was very fresh-looking, and no obvious changes

Text-fig. 1.

Drawing of live Trypanosome from the crop of Pontobclella.

'J'he animal, which has been recently ingested with the blood,
is in process of rounding off.

had taken place in the blood corpuscles. The Trypanosomes
showed very variable appearances. A good number still
showed the flagellum, but were no longer in the typical Try-
paniform condition. For the most part, they were somewhat
pyriform (text-fig. 1), with an immensely long, thick flagellum
protruding from one end. Some very fantastic appearances
were seen where the body of the Trypanosome had assumed
an irregular shape with curious rounded bulges, and where
the flagellum had broken loose from the membrane, and had
become tangled round the body, the end was usually free

124

MURIEL ROBERTSON.

and still motile. In othei' cases free flagella still actively
motile were seen ; this has often enough been observed with
Trypanosomes, but in this case the flagellum does not take
the kinetonucleus with it. Uninucleate stages of this parasite
have never been seen. One of these free motile flagella was
seen to become secondarily attached to a resting individual.
This animal was watched for many hours in case the process
might prove to be of more than merely casual significance,
but no development took place. Besides these flagellate
creatures, others were present which had already discarded
the flagellum (text-fig. 4). These were rounded egg- or
pear-shaped individuals with a characteristic clearly visible
nucleus. It is composed of a softly refractile circular body
surrounded by a bright halo. I'he nucleus lies towards
the broader end of the body. These animals pi’esent a very
characteristic appearance, but, nevertheless, are easily over-
looked in the mass of leucocytes and blood-corpuscles.

These uou-flagellate organisms were already in a few in-
stances undergoing division but no sign of the new flagellum
was as yet forthcoming.

Another leech (7a) opened forty-eight hours after it began
to feed showed only resting forms. The blood in the crop
had coagulated into a rather dry mass but the corpuscles
showed no signs of degeneration.

'I’hese resting stages of the Trypanosome are identical in
appearance with those so frequently seen in Pontobdella
found infected in nature. A resting pear-shaped individual
was chosen as a subject for observation at 4.30 p.m. When
it was first observed ^ the trophonucleus was clearly visible
and had its usual appearance of a sphere suri’ounded by a
halo. Half an hour later the animal was more rounded, the
nucleus was less distinct, and a slight groove had appeared at
the broad end. By 5.30 the nucleus, as such, had disap-
peared, but a large clear oval space had appeared in its stead.
At about 5.50 the two nuclei began to reappear, joined by a

' I am inclel)ted to Mr. C. H. Mai-tiii for kind assistance in carrying
out some of these continuous observations uiion the live specimens.

FUETHEH NOTES ON A TRYPANOSOME.

125

clear area. At 6.15 the two nuclei were once more quite
clearly defined but the clear area joining them remained
visible till about 9 o’clock after which it was no longer to be
detected. This clear band is the remains of the division
spindle : it is a very characteristic feature in the stained
specimens. During the division of the nucleus the body had
gradually become flattened in an antero-posterior direction
and correspondingly widened laterally. Grooves also began
to appear in the antero-posterior direction. It is to be noted
that these arose both at the anterior and the posterior end —
the anterior end being the broad end at which the nucleus
lies when the animal is in the pear-shaped condition and at
which the flagellum is later developed.

These gTooves altered a good deal in appearance during the
next few hours and towards 4 a.m. had deepened till the
animal presented the picture of two pears stuck together in
the middle, with however, the two broad ends and the two
pointed ends free. This is a point of some slight importance.
When in the trypaniforiu state division of the protoplasm of
this parasite usually begins from the anterior end and pro-
ceeds to the posterior. This is likewise the rule in the crithi-
dial state. This question of the grooves arising at both the
ends of the parasite is not in itself deserving of much remark
but it explains some curious stages where division of the
protoplasm goes from the posterior to the anterior end, to be
described in a later part of the paper.

This individual was watched for another two hours and one
of the two daughter-individuals developed a stiff and very
short flagellum which, however, showed no signs of move-
ment. The other individual appeared to have a little projec-
tion suggesting a flagellar rudiment. At 0.30 a.m. the animal
was finally abandoned, although the complete separation of
the pi'otoplasm had not yet occurred.

The flagellum seems to appear for the first time somewhere
between the second and third day after feeding. It is a very
characteristic feature that it generally arises at a division
stage. The flagellum is at first a stiff and relatively thick

126

:\ruRrEL rorektson.

little rod which sticks straight out from the anterior end of
the organism. A very considerable time seems to elapse
before it becomes motile, I cannot say exactly how long, but
it seems to be moi’e than twelve hours.

A leech (6n) opened six days after feeding showed a typical
infection of the varied type so characteristic of Ponto-
b del la.

True Trypanosomes had already appeared, some of these

Trypanosomes from the blood of the skate rounding off on a sealed
slide. The drawings are from live si>ecimens.

were broad individuals and others were much more slender,
but not of the elongated type which appears at a much later
stage of digestion. 'I’he broad and slender types were joined
u}) by innumerable intermediate forms. Besides these crithi-
dial forms were also present. I call crithidial forms those
where an undulating membrane is developed, but which have
not the typical Trypanosome arrangement of the kineto- and
tropho-nucleus. Herpetomonad forms with the flagellum
sticking straight out from the broad end of the body and
with as yet no membrane were also to be seen. And finally

Text-fig. 2.

Text-fig. 3.

FURTHER NOTES ON A TRYPANOSOME.

127

many rounded forms, some dividing and some developing tlie
flagellum, were likewise present. Some of the trypanifonn
individuals were already dividing.

Conjugation was very carefully searched for as it seemed
pi’obahle that it might occur at this stage of the life cycle,
but no signs of such a process were detected. Two individuals
were found joined by their posterior ends, one slightly broader
than the other. They were watched continuously from 6 p.m.
till 3.15 a.m., and the protoplasmic junction between the two
was seen to become much more slender and pulled out,
showing that the individuals were dividing.

Text-fig. 4.

Text-fig. 5.

Text-fio. 4. — Rounded off Trypanosome.

Text-fio. o. — Division of resting Trypanosome.

Drawings made from live specimens in a sealed slide of skate’s

blood.

An interesting corroboration of the stages above described
was obtained from blood drawn from a skate and sealed up
between a coverslip and slide (text-figs. 2 — 5). A Try-
panosome was continuously watched from 2.45 p.m. when the
slide was made. At 4.30 the animal had come to rest. The
flagellum which, when it breaks free from the membrane, is
seen to be of a relatively immense length was tangled up
round the animal. The slide was watched for some hours
longer, but as the Trypanosome was no longer motile it was
left. Xext morning it was found to have divided into two.

The behaviour of the Trypanosomes on a sealed slide is
interesting — a number do not alter at all, others very soon
after the slide is made begin to react to the altered condi-

128

MURIEL ROBERTSON.

tions. They adopt a dumpy spiral shape, or the posterior end
may become much thickened at the expense of the rest of the
body (text-figs. 2 and 3). Sometimes the most fantastic shapes
are seen, finally the flagellum breaks free but may remain
attached to the Trypanosome by its posterior end. It may
then become tangled round the body and stick out iu stiff
loops. The Trypanosomes in these phases on the sealed slide
made from the skate’s blood are absolutely identical in appear-
ance with the motile forms described in leech 5a (cf. text-
fig. 1). The time at which the Trypanosomes come to rest
varies much.

On another occasion a rounded non-motile Trypanosome
which had discarded its flagellum was chosen for continuous
observation on a slide of skate’s blood which had been
mounted for fourteen hours. The animal was seen to divide
into two about 11 o’clock in the forenoon. During the after-
noon, at about 4 o’clock, these two individuals each divided
thus forming four little rounded animals lying more or less in
contact. By 7 the same evening they had become more oval
and were identical in appearance with the resting phases in
the leech. By 9.30 p.m. short pi'ojectious were seen at
the broad end of two out of the four creatures under obser-
vation. The slide was left about 10 o’clock that night
as the animals wei’e not motile. Next morning at 9.30
— thirty-six hours after mounting the slide, observation was
a'min resumed and it was found that the creatures had each
divided. The eight resulting individuals wei'e still closely
apposed but not connected with each other. Unfortunately
these creatures were lost owing to a careless movement. The
slide was, however, iu the following condition. Unaltered
'rrypanosomes, still actively moving, were to be seen, non-
motile groups of four and also a few groups of six and eight
individuals Avere present.

I had often noticed that the Trypanosomes on a sealed slide
of skate’s blood altered their shape, but, thinking this was
merely a pathological manifestation, had not persevered with
the observation. The process is easily enough passed over

PUETHEE NOTES ON A TEYPANOSOME.

129

unless the observation is continuous, as the infections are
generally rather slight, and, once the Trypanosome has come
to rest, it is not quite a simple matter to see it among the
large number of leucocytes and red corpuscles. Moreover,
the curious fact that all the Trypanosomes on a slide do not
round off leads one to imagine that no development has
taken place.

A low temperature seems to favour the process. This
work on the live skate’s blood Avas carried out at the Millport
Marine Station, and the most successful set of observations
Avas obtained in very cold Aveather, Avhen the temperature of
the laboratory Avas much loAver than usual.

It appears to me that these experiments Avith the young
leeches, which confirm Brumpt’s short sketch of the life cycle
of T. raiae, are good evidence that the Trypanosome in
I’ontobdella is T. raiae. Of course it may be urged by
those who do not consider that Trypanosomes have a cycle of
evolution, however simple, outside the vertebrate host, that
the young leeches Avere hereditarily infected Avith a flagellate
of their oAvn. Against this I can only advance that the long
fast of more than a year, to Avhich these neAvly hatched leeches
Avere subjected, is not in favour of the survi\'al of such para-
sites; and that the close correspondence betAveen the early
stages in the leech, and those obtained upon the sealed slide
from the skate, is strongly in favour of the identity of the
parasites. Further, even if, in addition to the direct infection
from the skate, a hereditary infection also exists in Ponto-
bdella, it has still to be proved that the form inherited is
not T. raim.

The only Avay in Avhich quite conclusive proof of the ques-
tion could be obtained is by means of experiments Avith both
leeches and skate hatched in captivity. I hope ultimately to
be able to carry these out, but the difficulties iu the Avay are
obvious, as the obtaining of Pontob della cocoons is a matter
of chance, and the breeding of skate takes time and is often
uncertain.

VOL. 54, PART 1. NEW SERIES.

9

130

MURIEL ROBERTSON.

I cannot agree with Capt. Patton and Mr. C. Strickland,
who, on the basis of tuy previous description, consider this
parasite to be an independent Crithidia having no connection
with T. raiae. In a recent paper ^ I observe that they place
it in this group with the name of C. robertsoni. I would
like to point out that, quite apart from the question as to
whether this parasite belongs to the cycle of T. raim or not,
it certainly is a Trypanosome. The Crithidial, like the Her-
petomouad stage which precedes it, is transitory; the animal
ultimately adopting the Trypanosome state. I admit at once
that the distinction between a Crithidia and a Trypanosome
is not a very important one, but, such as it is, I do not see
that there is any scientific point in neglecting it.

I now wish to give a brief account of the parasite as seen
in stained films, fixed, for the most part, in Schaudinu’s
(alcohol-acetic corrosive sublimate) fluid. The stains used
were Delafield’s hmmatoxylin, Heidenhain’s iron liEematoxylin,
T wort’s licht-griin, and neutral red combination and fuchsin.

Delafield gives an excellent result, staining the nuclear
structures with clearness and precision; the flagellar appa-
ratus stains, but does so a little faintly.

Iron haematoxylin gives a very clear picture, staining the
nuclear parts coal black, and bringing into good relief flagellar
and cytoplasmic detail. Great cai'e, however, must be taken
in the interpretation of this stain, as it leads one into much
the same errors as Giemsa’s method, in so far as it stains
chromatic and acromatic structures alike. Tberefore, while
lleidenhain’s method gives a really splendid picture, it is
necessary to check the results by Delafield’s limmatoxylin,
which is a much safer stain.

Twort’s combination of neutral red and licht-griin was also
used. This is a clear, transparent stain, giving a red reaction

' “A Critical Review of the Relation of Blood-sucking Invertebrates
to the Life Cycles of the Trypanosomes of Vertebiates, etc.," by
Captain Patton and C. Strickland. ‘ Parasitology,’ vol. i, No. 4, Dec.,

iyu8.

FURTHER NOTES ON A TRYPANOSOME,

131

for chromatin, and a green reaction for cytoplasmic and
achromatic structures. The drawbacks to this stain are the
uncertainty of action which seems to attach to all delicate
double stains, and the fact that there is another loophole for
uncertainty in the process of washing* out the stain; so that
it remains doubtful in some cases whether the nuclear colour
is absent from a structure, owing to the absence of chromatin
or through the staiu having been washed out.

Fuchsin gave quite good nuclear pictures, but did not
bring up the flagellar apparatus sufficiently well.

The drawings in the plate are made from two well infected
leeches found infected in nature. They were at the early
part of what I have called the middle period of digestion.

For convenience sake the periods of infection may be
divided into three, corresponding to the stage of digestion:

(1) The early stage, when the blood is just coagulating,
and when the first signs of the dai*k greeu-browu fluid is
visible in the upper part of the intestine. The parasite at
this time is in the condition of throwing ofi^ the original
flagellum and adopting the resting state, during which
division begins to take place. The parasites are for the
most part still in the crop.

(2) Middle period of digestion, when the intestine is full of
the green-brown fluid, where breaking down of the blood is
going on actively. The parasite is now in the intestine in
large numbers. It shows the whole range of forms, from the
spherical non-motile creature to the typical Trypanosome,
(xreat variation in size and thickness of the pai’asites is to be
observed. Very slender, long forms are only occasionally to
be seen. This middle period is of very long duration,

(3) Final period of digestion, when the crop is empty (or
almost so) of blood, and the intestine nearly, or completely,
free from the characteristic green-brown fluid. The Try-
panosomes are now long, slender forms, with the kineto-
iiucleus in the typical Trypanosome position. The forms now
begin to remount the crop, and are also to be found in a still
more slender condition in the proboscis.

132

IMTTRIEL ROBERTSON.

The drawings, being made from leeches in the earlier
phase of period (2), do not show the long, slender Trypano-
somes developed during period (3). This final stage of the
Trypanosome is most stinking ; but it is, of course, a matter
of chance to get a leech in this condition in nature. This
year I only got one ; it was only slightly infected, and I ivas
unfortunate in not getting fixed films.

It will be convenient first to give an account of the Try-
panosome phase as found in the intestine of the leech (figs.
8 — 11), and then to give any points of interest in its develop-
ment from the resting form.

The protoplasm is finely and evenly granular without
vacuoles; indications of alveolar structure can be detected
in some specimens, but are not very conspicuous. Proto-
plasmic inclusions are only occasionally present.

The trophonucleus is composed of a large central karyo-
some surrounded by a wide halo, which is in turn surrounded
by a membi’ane.” Fine, but perfectly distinct, rays pass
from the karyosome to the outer membrane. The karyosome
is quite obviously made up of two substances, namely, the
chromatin and an achromatic substance, in which the chro-
matin lies embedded. This achromatic substance frequently
receives the name of plastin, and, while this does not convey
any very clear idea, it is nevertheless a convenient and useful
term. In Delafield’s preparations the plastin stains a pale
greyish-blue, in iron hfematoxylin it is brownish, and it takes
the green colour in the Twort’s combination. The nature of
the rays is a little obscure : they stain, as a rule, rather
faintly with Delafield, but in some cases take the colour more
deeply; Heidenhain shows them up black, but they wash out
easily. I am iucliued to regard them as more of the nature
of plastin, but they seem at times to carry chromatin. The
membrane shows very often little condensations of chromatin-
staining material at the points where the rays meet it. The
membrane stains well with Delafield and also with fuchsin,
likewise with Heidenhain, but washes out long before the
karyosome. The condensations on the membrane appear to

FURTHER NOTES ON A TRYPANOSOJEE.

133

me to be chromatin, but in Twort’s stain they do not take up
the red colour. I do not lay very much stress on this point,
as it is just in a question of this kind that I think such a stain
as Twort^s is rather unreliable. There seems to be in the
membrane, as in other parts of the nucleus, an underlying
substance of an achromatic nature, in or on which the chi’o-
matin is deposited.

This nucleus is exceedingly constant in all the stages of
the parasite as found in the leech, the only variation lying in
slight differences in the condensation of the chromatin in the
karyosome and the membrane.

The pictures presented in the dried Giemsa preparations
differ greatly from this account. The most curious feature
about this is that some of the Giemsa appearances give a very
tolerably accurate representation, while others depart com-
pletely from the type shown by the wet method. The eight
chromosomes so often seen in the Giemsa nuclei are not to be
detected in the hmmatoxylin films. The rays and the con-
densations on the membrane are, I have no doubt, the
manner in which these appear in the wet films. The number
of the rays can, liowever, not be made out, as they are
excessively fine, nor do the condensations on the membrane
stand out sufficiently separately to be considered as individual
structures. The Giemsa stain, of course, always increases
the apparent size of any nuclear element into which it
penetrates.

The kinetonucleus takes all the stains mentioned with srreat
intensity; it is relatively large and rod-shaped. In close
proximity, and apparently attached to it, lies the blepharo-
plast (Minchin, ‘ Quart. Journ. Micr. 8ci.,’ May, 1908, vol. 52),
(figs. 9-10, etc.).

This structure will be more fully considered when the
development of the flagellum is discussed. The blepharo-
plast stains with iron luematoxylin, but the stain is washed
out more readily than from the nuclear structm*es ; it
appears grey-blue and rather faint with Delafield, and is
difficult to detect at all in Twort preparations. Tlie flagellum

134

MURIEL ROBERTSON.

runs forward from the blepharoplast, ending, as usual, in a
free whip. It stains green with Twort’s combination and
faintly with Delafield — more deeply with iron hsematoxylin.
The undulating membrane is developed to a varying degree,
but is never much frilled.

Two other structures remain to be described. The first is
a small granule only staining in Heidenhaiu preparations. It
lies posterior to the kinetonucleus and often near the pos-
terior end of the body. Sometimes it appeal’s to be connected
to it by a delicate strand. This granule is also found in the
Giemsa preparations and is clearest in the Trypanosome
phase. It has possibly something to do with the anchoring
of the kinetonucleus (figs. 9 and 10).

The second is an element which I have never seen in the
dried preparations at all, but which is a pretty constant feature
in the wet films and appears equally so with all the four
stains used. Just anterior to the trophonucleus a small con-
densation is to be observed in the protoplasm surrounded by
quite a definite little halo. In rather dark Heidenhaiu pre-
parations it stains almost black (figs. 9 and 10), with Delafield
it looks grey-blue with soft outlines, and is usually only very
slightly darker than the surrounding protoplasm (figs. 8, 11
and 19). It stands out clearly in these films more by reason
of the halo than on account of its greater depth of colour.
In Fuchsine films it is very clear : it is also visible in Twort’s
preparations but does not take on the red colour. It some-
times appears to be double. The nature and function of this
structure is quite obscure ; it is most clearly visible in the
Trypanosome stage and its position is quite constant, but it is
also present in the earlier phases. The position of the kineto-
nucleus often obscures it in these and makes it difficult to
see.

'I’lie Trypanosome just described arises, as has been said,
from a rounded resting form which develops a flagellum (figs.
1 — 11). The body gradually elongates and the kinetonucleus
migrates backwards until it is well posterior to the ti’opho-
nucleus. An undulating membrane develops during this pro-

FURTHER NOTES ON A TRYPANOSOME.

135

cess and the ci’eature takes on the typical Trypanosome
facies. The only point about this that calls for special atten-
tion is the development of the flagellum.

It may be noted in passing that, owing to the flattening of
the Trypanosomes prepared by the dry method, certain details
may be more distinctly visible in such specimens than in those
prepared by the wet method.

The earliest stage of the development of the flagellum, of
which one can be quite certain, is shown in fig. 2. Here it
will be seen that two little projections have grown out from
the kinetonucleus which is itself in process of division. These
little structures sometimes take the Heidenhain rather deeply ;
they are not, however, very easy to make out as any obliquity
in the position of the kinetonucleus is apt to obscure them.
Later stages are shown in figs. 3 — 5. Here the flagellum
appears as a thick strand arising from a granule with not
very definite contours, which is in turn attached to the kineto-
nucleus. This granule at the origin of the flagellum is the
blepharoplast. Tlie minute detail is not very clear in the wet
preparations, but as far as I can make out the blepharoblast
seems to be attached to the kinetonucleus by a double thread.
This may be seen in much later stages (figs. 5 and 6). The
blepharoplast seems like the kinetonucleus to be ultimately a
thin rod-shaped body. It seems to arise from the kineto-
nucleus, but I do not think in the light of its behaviour
with the various stains that it is a chromatic body. It is true
it often stains a deep sharp black with iron-hsematoxylin, but
that is no test for chromatiu. It takes on a grey-blue pale
colour with Helafield and shows up only dimly when at all with
Fuchsin. In Twort’s combination it stains green like the
flagellum and is not very clear or precise. I am therefore
inclined to regard the blepharoplast and flagellar apparatus
which grows out from it as achromatic. Our knowledge of
achromatic nuclear elements is so limited at present that it is
impossible to say whether they can be regarded as an expres-
sion of the achromatic elements of the kinetonucleus or not.

Oiemsa’s stain presents a greatly exaggerated picture of

136

MURIEL ROBERTSON.

the above development. The drying pulls the blepharoplast
away from the kinetonucleus and makes the thread joining
them stand out very clearly; it also greatly enlarges the
apparent size of the blepliaroplast, and very markedly
increases the flagellar rudiments, which are always thick at
this stage. In addition to all this these flagellar structures
stain a deep red corresponding to the chromatic colour.
That such a stain should be a fruitful source of error is
obvious. It must, however, in justice be said of this staiu
that it has done brilliant if unequal service to the advance-
ment of our knowledge of blood parasites, and has, therefore,
amply served its purpose.

The longitudinal division of the kinetonucleus which I
held to be the first stage in the development of the flagellum
has been seen again iu resting forms (fig. 1), but iu the light
of the above observations I do not feel confident as to how
this appearance should be interpreted. It is iu these, as iu
the other specimens of Pontobdella previously examined, a
rai'c appearance, and the evidence for considering this as the
initial step iu the formation of the flagellum is very much
less clear in the wet films than iu Giemsa preparations.
Moreover, I have recently found in T. vittatse that the
longitudinal division of the kinetonucleus is the ordinary
method instead of the transverse, as in T. raim; it is, there-
fore, possible that the point may be open to some variation.

Division. — The figs. 7, 12 — 17 give pictures of various
division stages, and it is perhaps in this point that Giemsa’s
method has, generally speaking, been the least misleading.
The division starts as a rule by the transverse division of the
elongated kinetonucleus, but this point is open to slight
variation.

The trophonucleus shows first an arranging of the chroma-
tin in two masses within the karyosome (figs. 7 and 12). A
well-developed spindle subsequently arises, but the chromatin
is divided without the formation of an equatorial plate. The
wet method brings up the spindle very well (figs. 13 — 17).
Ceutrosomal functions of some kind seem exercised by the

FURTHEB NOTES UN A TRYPANOSOME.

137

condensations at the extreme points of the spindle, and the
chromatin seems to pass along the “fibres”^ from the two
main central masses in such a stage, as fig. 13, to either poles
of the spindle. There is at a slightly later stage (figs. 14 and
15), often a curious double appearance in the nuclei. Figs.
IG and 17 give the final stages. The spindle persists for
some time after the nuclei are reformed, and can be clearly
seen as a bright line in the live specimens.

Division occurs at all the different stages of development.
There is an occasional not very well defined tendency to
multiple division. Ecpial division is the rule, but unequal
division is sometimes met with. Considerable variation in
the division of the protoplasm is seen, as shown in figs.
18 — 24. It will be observed that in figs. 20 — 22 the division
of the protoplasm has started at the posterior end instead of
the anterior, as is more usual. Specimens of this type were
watched for many hours in the live state in the hope that
they might be individuals conjugating, but no evidence in
favour of this was forthcoming, and I was led to believe that
they are in all probability mei’ely cases of division.

Some very curious appearances were observed where the
protoplasm had split into several rod-like processes. This is
shown in fig- 24, and, while not a common appearance, is
still far too frequently seen to be dismissed as a casual
abnormality. The figures, I may say, hardly do justice to
all the varying sizes and shapes to be seen at this stage in
the development.

As I have already said in a recent paper ^ on T. vittatae
from the soft tortoise Emyda vittata, we seem to have in
these forms a type of life-history which is probably very
wide.spread. The rounding off process and the subsequent
development of a flagellate stage occurs, as has been recently
shown in many of the Trypanosomes from cold-blooded hosts.

' I use the word fil>re quite without prejudice. The word con-
veniently expresses the optical effect, and I have no means of knowing
what they actually represent.

- ‘ Quai-t. .lourn. Micr. Sci.,’ vol. .5.3, part 4.

VOL. 54, PART 1 . — NEW SERIES. 1 0

138

MURIEL ROBERTSON.

It seems to occur in the Trypanosome from Hyla
arborea (Franya, ‘Bull. Soc. Portug. Sc. Nat./ 1907), also
in T. granulosum (Franya), also in T. loricatum (Dutton,
Todd, and Toby, ‘Ann. of Trop. Med. and Parasit.,’ No. 3,
1907).

The similarity between the life-history of T. vittatse and
T. raim hardly needs emphasising. T. raiae, of course,
shows clearly the adaptation to the peculiar habits of the
Pontobdella, its relatively slow development and the long
persistence of the resting phase being correlated with the
stiffening of the blood in the crop and the very long period
which the Trypanosome must pass in the leech owing to the
extreme slowness of the digestion.

The work here recorded was carried out partly at the
Millport Marine Station, partly in the Zoological Laboratories
at Glasgow University and the Lister Institute, Chelsea.

Lister Institute ;

April. 11*09.

EXPLANATION OF PLATE 9,

Illustrating Miss Muriel Robertson’s paper on “ Further Notes
on a Trypanosome found in the Alimentary Tract of
Pontobdella muricata.”

Fi^s. 1 — 24 are drawn with 2 mm. apochr. immersion lens by Ziess,
I'fO N.A. long tube and oc. No. 12, with the assistance of the camera
liuada. The magnification is .approximately 4500 diameters.

Fig. 25 is drawn with the No. 2 eyepiece ; the magnification is
approximately IfiOO diameters.

Figs. 1, 2, 3, 4, S, 9, 10, 13, 15, 17, 18, 19, 21 and 23 are from Heiden-
bain’s bamiatox. preiiarations. The remaining figures are from Delafield
preparations.

All fbe figures, with the exception of 25, are from the leech Ponto-
bdella.

FURTHKE NOTES ON A TRYPANOSOME.

139

Fig. 1. — Resting phase, showing longitudinal division of the kineto-
nucleus.

Fig. 2. — Early stage in develoinnent of flagellum.

Figs. 3, 4 and 5. — Stages showing newly-formed flagellum.

Figs. 6 and 7. — Crithidial stages.

Fig. 8. — Trypanosome phase.

Fig. 9. — Trypanosome showing lilepharoplast, granule at posterior
end, and the structure just anterior to the trophonucleus.

Figs. 10 and 11. — Trypanosome phase.

Fig. 12. — Early division phase. Note condition of trophonucleus,
kinetonucleus and hlepharoplast.

Fig. 13. — Division stage showing spindle.

Fig. 14. — Later division stage.

Fig. 15. — Division stage showing trophonucleus spindle and also
second division of the kinetonucleus. Tliis is a rather unusual appear-
ance.

Figs. 1(5 and 17. — Later division stages.

Fig. 18. — Division of Trypanosome.

Fig. 19. — Division of Trypanosome.

Figs. 20-22. — Division stages where the protoplasm divides from the
posterior end.

Fig. 23. — Rather unusual appearance where the posterior part of a
dividing Trypanosome has formed a large I’ounded mass.

Fig. 24. — Division stage showing irregular splitting of the proto-
plasm.

Fig. 25. — T. raiae from the skate's hlood. Note this is drawn at
a much smaller magnification (x IGOO) than remaining figures.

Oumi. Juiini. Micr. Sci. Vol. 54, N.S., PI. 9

25.

!

DENDROSOMA RADIANS, EHRENBERG.

141

Dendrosoma Radians, Ehrenberg.

By

Sydney T. Ilickisoii, D.Sc., F.K.8.,

Beyer Professor of Zoology in the University of Manchester,

and

J. T. Wadsu'orlli.

With Plate 10.

Contents.

PAGE

Previous Observations .... 141

On closely related Genera .... 143

Material ...... 14o

Food ...... 149

Structure of tlie Cytoplasm . .1.51

The Meganncleus ..... 152

The Micronnclei ..... 157

The Gemmulaj ..... lOl

The Free-swim!uing Gemuiulaj and their Development . 164

On Urnnla epistylidis .... 170

On the Systematic Position of T r i c h o p h r y a, L e r n ae o -

phrya, and Dendrosoma . . 174

Summary of Results . .178

Literature . .178

Descrijition of the Plates .... 180

Dendrosoma radians was first described by Ehrenberg
in 1837 (9), but the figures he drew to illustrate its structure
were not published until 1862 (11). In 1840 (10) he gave
the following diagnosis of the species ‘^Dendrosoma
radians: D. corpusculis conicis, ci’assis, mollibus lasvi-
busque, alterne ramosis, rame apice incrassatis et tenta-
culatis ”

VOL. 54, PART 2. — NEW SERIES.

11

142 SYDNEY .1. HICKSON AND J. T. WADSWORTH.

Ehvenberg apparently saw the meganucleus, but in his
first paper expressed the opinion that it was the male genital
gland. In the figure (11, Plate III a) which he published
later he represents a long axial vacuole, probably the mega-
nucleus, and in the description of it apparently abandons the
view that this structure is a s'enital nland.

Claparede aud Lachmann (5) gave a good figure of Dendro-
soma in 1861, but made a grave eri’or in describing: the mega-
nucleus as an elongated contractile vacuole. There is no
reason whatever for believing that there is in Dendrosoraa
any system of elongated canals in communication with the
ordinary spherical contractile vacuoles.

The next important contribution to our knowledge of
Dendrosoma is that of Levick (23), who in 1880 described
and gave a good figure of the gemmule. Levick also gave a
description of the interesting streaming movements of the
protoplasm, which we have confirmed. There can be little
doubt, however, that the germ and sperm elements” of this
author were not correctly interpreted. Notwithstanding the
statement that he actually saw living spermatozoa comparable
to those of Hydra discharged from the Dendrosoma, it is, in the
light of our knowledge of the reproductive processes of other
Acinetaria, impossible to accept the view that Dendrosoma
possesses at any time definite male and female sexnal
glands.

In 1881 Saville Kent (19) published a more elaborate
account of the species and gave a figure of a large specimen
one tenth of an inch in height, which has been copied wdth
some modifications in nearly all the text-books of zoology.

The principal new point of interest in Kent’s accouut of
the species is the description of “ certain exogenousl_y pro-
duced buds similar to those of Acineta mystacina des-
cribed by Stein.” If Kent’s interpretation of these bodies
were correct Dendrosoma would present the very remarkable
peculiarity of producing two different kinds of gemmulte
at the same time. Subsequent authors have referred to
these bodies with caution: Butschli (2) calls them ^'angebliche

DEXDROSOMA RADIANS, EHRENBERG,

143

freie aiissere Knospen ” ; Lankester (21), in his description of
the figure, refers to them as “more minute reproductive (?)
bodies.”

Tliere can be no doubt, however, that these bodies are not
“ buds ” at all, but epizoic Acinetaria belonging to the
genus U r n ula.

The megauucleus was described by Kent as follows : “Endo-
plast ribbon-like, ramifying and much contorted in the
stolon and basal portions of the main stem, continued as a
simple band into the distal region and secondary branchlets.”

He gives a figure of the meganucleus in a small portion of
the basal part of an arm, which is probably correct so far as
it goes, but in the figure of a distal portion of an arm the
nucleus is shown to extend much nearer to the extremity
than we have found it in any of our preparations. In the
figure of the whole specimen the ineganucleus is not shown
at all. This figure has been copied by the writers of many
text-book.s, and in some, as, for example, those of Calkins
(3) and Lang (20), the meganucleus has been added to the
figure and erroneously shown to extend to the extremity of
all the branches.

The other genera of Acinetaria that are apparently closely
related to Dendrosoraa are Trichophrya (C. and L.), Ler-
lueophrya (Perez), Astrophrya (Awerinzew), and Den-
drosomides (Collin). Concerning Astrophrya we have
at present only a short note and figure by Awerinzew (1).
The single specimen obtained was found free in the plankton
of the Volga, and, like some of our specimens of Deudrosoma,
it had a massive test of adherent sand grains and plant
detritus. As it is figured with long arms and rather short
tentacles its affinities are apparently with Dendrosoma rather
than with Lernmophrya.

The genus Trichophrya seems also to be closely allied to
Deudrosoma. It is found on the stalk of Epistylis and on
the appendages of Astacus and various Entomostraca. We
have found specimens which we attribute to the genus
Trichophrya in our material obtained from the Bridge-

144 SYDNEY ,T. HICKSON AND J. T. AYADSWOETH.

water canal. The genus may be distinguished from Dendro-
soma by its small size, by the shortness and unbranched
character of the processes that bear the suckers and by
other chai’acters which are described later (p. 174) (fig. 46).

The genus Lernaeophrya was found by Perez (25)
attached to Cordylophora in the docks at Bordeaux. We
have also found it associated with Cordylophora in the
material from the Bridgewater canal. It varies in size, but
is usually larger than Trichophrya. It has short, un-
branched arms and remarkably long suckers (fig. 48). In
other respects also it appears to be distinct from Dendrosoma
and other genera (see p. 176).

The genus Bendrosomides of Collin (17) was found on
the thoracic appendages of Eupagurus cuanensis. It
appears to have some affinities with Dendrosoma, but on the
whole it seems to be more closely related to the Ophryo-
dendrime.

The genus S taurophry a,^ Zacharias, is a free fresh-
water form, with short arms, long non-capitate tentacles.
Its affinities are obscure.

There is very little information at present as to the
geographical distribution of Dendrosoma. All that can be
said is that it has been found in Europe and in the United
States of America. According to Ehrenberg it is found on
Ceratophy Hum, Callitriche, and on dead leaves. Kent
found it on Anacharis, Myriophyll u m , and other aquatic
plants. Leidy (22) states that it is found in America on
Ceratophyllum, and attached to the rotifer Limnias
socialis. Stein (27) briefly refers to a species, Deudro-
soma astaci, attached to the appendages of the fresh-water
crayfish. As suggested by Kent it is very probable that this
is the same species as D. radians.

' The only account of this genus we luive found is in Deluge’s and
Herouard's ‘ Protozoaires,’ p. 514.

DENDROSOMA RADIANS, EHRENBERG.

145

Material.

The first material we examined was supplied to us some
years ago by Mr. Bolton, of Birmingham. Since that time
he has forwarded to us living specimens on several occasions.
The Birmingham material is usually attached to water
plants.

A few years ago the Kev. T. Robinson, of Hale, informed
us tliat Dendrosoma could be obtained in the Bridgewater
canal, in the neighbourliood of Altrincham, attached to
specimens of Cordylophora, and we have found it there
and obtained an abundant supply at all times of the year.
For many mouths it is principally attached to the hydrocauli
of living Cordylophora, but we have also found it on the
stalks of freshwater Polyzoa and on weeds. In the winter
we have found specimens on the perisarc of the hydrocauli of
Cordylophoras that have died down. It is noteworthy that we
found these specimens a few days after the severe frost of Decem-
ber 25th to doth, 1908. They were very pleutiful, healthy,
and weie giving rise to a great many gemmulae. They were
also provided with a considerable number of epizoic Urnulas.
These facts seem to {irove that there is no special advantage
gained by the Dendrosoma in being associated with the
Cordylophora other than that of position. It seems probable
also that Cordylophora gains no advantage by the presence
of Dendrosoma on its hydrocauli.

A difficulty in the investigation of the Dendrosoma found
in the Bridgewater canal is, that the test becomes encrusted
with a thick coat of Avhat we can only call black dirt (text-
figs. A and B). This renders the observation of the structure
of the stolon and the proximal regions of the arms in anything
but sections extremely difficult. The material obtained from
Birmingham is much cleaner. Associated with the Cordylo-
phora and Dendrosoma in the Bridgewater Canal there are
several other genera of Aciuetaria, Vorticella, Car-
chesium, Epistylis, and Stentor, besides worms,
Rotifers et hoc genus omne.

146 SYDNEY J. HICKSON AND J. T. WADSWORTH.

Of the other genera of these Aciuetaria, two ai’e of con-
siderable importance in relation to Dendrosoma. These are
the genera Trichophrya (Clap, and Lach.) and Leruaeo-
phrya (Perez).

At one period in our investigations we weie inclined to
believe that both these genera represent young phases in the
life-history of, or varieties of, Dendrosoma radians, butwe
have since arrived at the definite conclusion that they are
certainly distinct species. The relation between the three
forms will be discussed later on, but we may state here that
we are in ag'reement with Perez in considering Lernceo-
phrya an intermediate form connecting Trichophrya with
Dendrosoma and justifying the inclusion of the tlu'ee genera
in the distinct family the Dendrosomina (Biitschli). It is a
matter of considerable interest that the three closely allied
genera should be found associated together in this manner in
the same locality. The struggle for existence between the
different Acinetaria and Ciliata must be very keen, and it is
surprising that the three genera should have suiwived side by
side. In the material from Birmingham we found no examples
of either Trichophrya or Lernseophry a.

The specimens from the Bridgewater canal consist of an
irregular base or stolon attached to the perisarc of the host
from which a number of free branches or ‘‘arms” project at
various angles into the water (text-hg. A). It is difficult to
form any vei-y definite conception as to the size to which a
single individual may attain, as the stolon is usually so thickly
encrusted with dirt of vailous descriptions, and bends so
frequently from one side of the C'ordylophora to the other in
its sinuous course, that the limits of the individual are often
impossible to determine. We have measured single un-
branched arms of this form that are ’5 mm. in length (text-
fig. B). The tentacles are very extensile, and may be 75-
90 fi in length. The arms arise from the stolon at irregular
intervals, sometimes in clusters of three or four, sometimes at
intervals of one or two millimetres. There appears to be no
geometric law governing the origin of the arms ; but an arm

DENDROSOMA RADIANS, EHRENBERG.

147

seems to be pushed out wherev'er an opportunity occurs for
one to reach a food supply.

Text-fig. A.

A specimen of Dendrosoma radians, attached to Cordylo-
phora, from the Bridgewater canal, near Manchester, c.v.
Contractile vacnoles. (jem. The “internal buds” or gemmula;,
me. the long strap-shaped meganucleus, me', the meganucleus
in the liase of attachment, showing at me", an irregular dis-
tribution of its chromatin granules in the cytoplasm ; t. the
test covered with dirt and foreign particles of various kinds.

V. The so-called “external buds.” now shown to be a species
of Urnula, ai'e seen at the extremity of some of the branches.

The figure is semi-diagrammatic. The outline was drawui with
the camera lucida from a very large, branched, living specimen,
and the structural details filled in.

The specimens from Birmingham differ in some respects
from those found in the Bridgewater canal. The former

148 SYDNEY J, DICKSON AND J, T. WADSWORTH.

attain to a greater size, their arms are longer and branch
more frequently, and they seem to possess more raicronuclei.
It is difficult to express the differences between them in exact
terras. Some specimens from Birmingham are very similar
to some specimens from the Bridgewater canal, and there is

Text-fig. B.

This figure represents the more usual form in which the variety
of Dendrosoma radians from the Bridgewater canal is
found. It is attached to the h y drocaulus of a Cordylo-
phora. The longest of the three arms was 0'5 mm. in length
and unhranched. The longest suckers are 7.5-90 /t in length,
c. Cordylophora hydrocaulus. c.v. Conti-actile vacuole.

undoubtedly a great range of possible variation in specimens
from both localities. The specimens from Birmingham are
undoubtedly more closely related to the original type speci-
mens of Ehrenberg and to those described by Kent than are

DENDEOSOilA EADIANS, EHEENBEEG.

149

the specimens from the Bridgewater canal, and we have no
hesitation in naming them Dendrosoma radians Ehren-
berg. It is a matter of opinion whether the specimens from
the Bridgewater canal are sufficiently distinct to justify the
constitution of a new species. After careful consideration
we have come to the conclusion that they are not. All the
pi'incipal differences, except that of the number of the micro-
nuclei, may very probably be due to the direct influence of
the environment.

Fooju

For some time we were unable to record any observations
on the food of Dendrosoma, the animal appearing to be
very refractory in this respect as compared with Dendroco-
m e tes.

We have frequently observed small Ciliata and Flagellata
swimming freely among the suckers, and even bumping up
against them, without harm to themselves or producing any
reaction in the suckers. In September of last year, however,
we obtained in the plankton of some ponds near Manchester
specimens of Buplotes and of a green Paramecinm
bursaria; and, on passing some of these under the cover-
slip of a preparation of living Dendrosoma, several were
caught and held fast by the suckers, and the process of
feeding began. The astonishing feature of the pheuomenon
is that Dendrosoma seems to prefer such large prey. A
Euplotes lOO/j in length will be held and devoured by a
Dendrosoma arm that is not more than lo/u in diameter
(fig. :I0).

When an Infusorian is captured by a Dendrosoma it is not
paralysed, but continues to lash its cilia with as great, or
even greater, activity than before. It is almost painful to
watch a Euplotes struggling to escape its doom. It will
make a violent effort to move forward and then fall back
exhausted for a few seconds, or it will endeavour to spin

150

SYDNEY J. HICKSON AND .1. T. WADSWORTH,

round on its axis. But all the time particles of its protoplasm
can he seen streaming down the suckers of the Dendrosoma
into the arm. The Dendrosoma never seems to use the whole
bunch of suckers at the end of an arm for feeding at the
same time. From one to six or seven, according to the size
and strength of the prey, may be used, the others being
stretched out unconcerned with what is going on in their
neighbourhood as if in search of other victims (fig. 30).

The usual statement made in the text-books is that the
Acinetaria paralyse their prey before feeding on them, and
this is used as an argument that a fluid of some kind passes
down the tentacle towards the food before the return current
of the food substance sets in. We are cjuite certain that in
Dendrocometes such a paralysis of the food animals does take
})lace. We have seen on several occasions small Paramecia
caught and paralysed by the arms of this remarkable form.
Biitschli (2) gives several examples of the pai-alysing pro-
})erties of the Acinetarian suckers, and quotes cases observed
by 8tein in j\I etac i neta., Maupas in Sphmrophrya, Plate
in Hypocoma.

Claparede and Lachmann (vol. ii, p. 50), however, record
that a Stylonychia caught by “an Acinetau ” struggled for a
long time and then underwent fission. One of the daughter
individuals escaped from the Acinetau, leaving her sister to
be devoured.

Levick (23) also does not mention that the Infusoria upon
which Dendrosoma feeds are paralysed, but says that the
tentacles are capable of “ resisting the struggles of the
captive.”

Since we made our first observations on the feeding of
Dendrosoma we have occasionally found a smaller infusorian
caught by the tentacles, but time after time we have failed
to induce them to feed upon the common Paramecium
a u roll a from our cultures.

But although we have succeeded in inducing the Dendro-
soma to feed we have failed to keep them alive in the labora-
tory for more than a few days. The material soon becomes

DEXDROSOMA RADIANS, EHRENBERG.

151

putrid owing to the death of the Cordylophora and the decay
of the roots to which it is attached, and we have not succeeded
so far in getting the specimens to become fixed to any other
support.

The Sti:ucture of Denurosoma.

We have very little to add to the knowledge of the suckers
and the general cytoplasm of Dendrosoma. The suckers are
of considerable lengdh, terminating in a small knob or cup.
They are usually quite rigid when extended, like the suckers
of most of the other Acinetaria, and they can be slowly
shortened when circumstances become unfavourable. Being*
very slender and transparent we have not been able to
demonstrate tlie pi’esence of a definite lumen, nor have we
seen even in the shortest suckers any evidence of a spiral
ridge such as can be seen on the retracted suckers of some other
Acinetaria (fig. 45). In several of our series of sections we
have been able to trace very delicate lines i-uuning down
some distance into the arm from the bases of the suckers.

The cytoplasm is usually clear and transparent. In the
living arm a streaming movement of minute granules can be
clearly seen, as originally described by Levick. In well-
stained preparations the cytoplasm appears to consist of a
delicate network of fibrils enclosing a number of minute
granules which stain faintly with acid dyes, but there are
veiy few elements in the cytoplasm that give a deep stain with
basic dyes. We have found no evidence in our prepai ations
of bodies corresponding to the ‘‘Tinctinkorper” of other
Acinetaria in the arms or stolon where the meganucleus is
clearly delimited. In some cases, however, when the mega-
nucleus is scattered (see p. 15G) the cytoplasm is filled with
numerous chromatin bodies which may be derived from the
meganucleus.

IMartin (24) suggests in a recent paper that the “ Tinctin-
kdrper ” of other Acinetaria represent in some cases the
chromatin of the nucleus of ingested prey. The difficulty we

152 SYDNEY J. HICKSON AND J. T. WADS^YOKTH.

have found in getting Dendrosoma to feed may to some
extent account for the absence of Tinctinkbrper.

The contractile vacuoles occur at irregular intervals on the
arms and may also occur in the stolons, but they are not easily
seen except in those parts which are relatively free from the
encrusting dirt (text-fig. A, cv.). The description of these
structures given by Levick and Kent is correct ; the elongated
contractile vacuole figured by Claparede and Lachmann is
obviously incorrect.

The Meganucleus.

In a stained preparation of Dendrosoma the meganucleus
usually appears as a dark band running along the axis of thp
arms and stolon. It is rarely exactly in the centre, but
usually bends first to one side and then to the other, or runs a
course for the whole length of an arm somewhat to one side of
the exact axial line. In the younger parts of a stolon it has
very much the same appearance as it has in an arm, but in the
older parts it is frequently contorted, sometimes broken or dis-
continuous, branched, or knobbed, and not infrequently
dissijjated (tigs. 11, 12, 13, and text-fig. A).

There is so much variation, however, in the general
character of the meganucleus that it is impossible to describe
it adequately in a single sentence.

Although it is usually lound to some extent in every arm it
never extends to the distal extremity as it is drawn in the
figures given by Lang and Calkins, and in the younger and
shorter branches there may be no part of the meganucleus at
all (text-fig. A). We have observed a difference, which
appears to be constant in this respect, between the specimens
fi’om the Bridgewater canal and those from Birmingham.
In the former the meganucleus extends only a short distance
into the arms, in the latter it extends very much further ; but
in this respect, as in others, the meganucleus is subject to
considerable variation. In many specimens the limits of the
meganucleus are very clearly defined, in others the boundary

DEXDEOSOMA RADEANS, EHRENBERO.

153

lines are ill-defined and the difference between the nieo'a-
nncleus proper and scattered chroraidia impossible to dis-
tinguish ffigs. 12, 13, 14).

If the form and distribution of the meganucleus of Dendro-
soma are remarkable, still more so is its minute structure.
The difference between the meganucleus of Den drocometes
and that of Dendrosoma in this respect is very striking, and
the first cause we had to doubt the current view that the so-
called ‘‘ external buds ” (Urnula) are really produced by the
Dendrosoma was the difference we observed in histological
detail between the meganucleus of these bodies and the
mesranucleus of the Dendrosoma.

We described in a former paper (16) the meganucleus of
Dendrocometes as consisting of a distinct meshwork of
darkly staining lines which appears to support a series
of minute, rounded, chromatin granules.'’^ In the meshes
of the darkly staining chromatin there is a homogeneous sub-
stance which stains faintly yellow with brazilin.”

In the meganucleus of Dendrosoma we have found no trace
of a true limiting membrane or of a meshwork. It consists
simply of a number of chromatin granules floating indepen-
dently in a fluid matrix (fig. 52). In this respect, therefore,
the structure of the meganucleus appears to be very ex-
ceptional.

We have not arrived at these conclusions without very
careful study and numerous experiments. We expected to
find some kind of network, whether of plastin or of chro-
matin, such as we found in De n d roco metes , Ur n ula, and
some other Acinetaria, but the various methods we have em-
ployed have given us no positive results. Staining with
hmmatoxylin and congo-red we have obtained very shai-p
differentiation of the histological structures, and in some
sections we have seen a thin line bounding the meganucleus
which might well be mistaken for a membrana limitans, but
after prolonged research we are convinced that this line does
not represent a continuous membrane, and that it belongs, not
to the nucleus, but to the surrounding cytoplasm. Whether

154

SYDNEY .T. HICKSON AND J. 'P. WADSWORTH.

our interpretation of this line is correct or not, the fact
remains that there is a very marked difference between the
ineganncleus of Dendrosoma and that of other Acinetaria
except Lernajophrya we have examined in this respect.

In referring to the meganucleus of Acineta papillifera,
Martin (24) writes : “ Generally in whole stained preparations
numerous spherical dark areas are to be seen resembling the
so-called “ Binnen-korper ” of the Infusoria. In section,
these structures, as in the case of some Infusoria and Dendro-
cometes, are found to consist merely of local thickenings in
the mesh of the nuclear network, and therefore resemble
karyosomes rather than true nucleoli.”

In Lermeophrya, so far as our observations go, the
meeranucleus resembles that of Dendrosoma.

In Acineta tripharetrata, according to Entz (13), the
substance of the meganucleus appears to resemble that of
Dendrosoma and contains a number of sharply defined bodies,
but there is a distinct nuclear membraue.

Collin (7) describes the meganucleus of Bphelota gemmi-
para as consisting of “grains chrom'atiques de forme variee
sur un substance achromatique,” but he finds also a distinct
isolable nuclear membrane.

In the iron-brazilin preparations a faint yellow colour can
be seen in the thicker sections between the granules, and iu
iron-luematoxylin and congo-red preparations a faint pink
colour may be seen in the same place, indicating perhaps that
the matrix in which the chromatin granules float is to some
extent capable of taking a faint oxychromatic stain, but no
structure is seen in it even with the highest powers (Zeiss
2 mm.) of the microscope we have used in the best light.

There can be little doubt that the granules are mainly
composed of chromatin. They give the characteristic stains
with iron-lnematoxylin, iron-brazilin, safranin, and thionin.
They are usually spherical in shape, but occasionally irregular-
shaped granules and large lumps (fig. 51) are found in the
course of the band.

'I'he variability in size may be seen by a comparison of figs.

DENDROSOMA RADIANS, EHRENBERG.

155

50, 51, 52, which are drawn to the same scale. Fi". 50
represents the terminal extremity of the megannclens in an
arm, and the granules are of an average size. The diameter
of the megannclens in this specimen was 2 ju, and the largest
granules were approximately 0'85 /.t in diameter. Fig. 51
represents a longitudinal section near the base of an arm in
which there were two meganuclear bands nearly parallel with
each other. The larger of these bands was about 3‘4 p in
diameter. The granules in the.se bands are extremely small,
but at intervals large lumps of chromatin, in some cases over
4 fi in length, occur. Fig’. 52 represents a section through a
remarkably swollen part of a megannclens (25’5/.t by 20’4 /<), in
which many of the granules are very large; the largest are
about 2 5 fi in diameter. The study of the variation in size
and form of the granules in the meganucleus suggests a com-
parison with the descriptions given of the megannclens in
Carchesium and other Ciliate Infusoria. In Carchesium,
accoi’ding to iMrs. Bidder (Greenwood, 14), there is a nuclear
membrane which in certain stages is “ denionsti-able with
difficulty”; the contents consist of granules of two kinds
floating freely in the nucleochyme, and there is no indication
of permanent linin threads connecting these granules togetlier.
fl'he granules are of two kinds — the proto-macrosomes and
the proto-mici’osomes, and of these the proto-macrosomes
certainly and the proto-microsomes probably undergo
changes in association with nutrition. The larger granules
of the meganucleus of Dendrosoma seem to agree with the
macro.somes of Carchesium, although we have not observed
the vacuolated condition which the latter sometimes exhibit.
A distinction between macrosomes and microsomes has not
been demonstrated in Dendrosoma-, but it is possible that
further investigation might yield similar results to those
obtained in Carchesium. 'Ihis comparison suggests that
the great difference we have recorded in the size of the
granules of the meganucleus may be in some way associated
with nutrition.

Division of the mega nude us. — The only method of

156

SYDNEY .1. H[CFvSON AND .T. T. WADS^YOln'H.

division we have been able to follow is the formation of
the me^anucleus or megannclei of the geramalge. Before
the outline of a young geinmula is seen in the cytoplasm, one
or more knob-like outgrowths are formed from the ribbon-like
meganucleus of an arm or of a part of the stolon. These grow,
and as the outline of the geramula is developed become con-
stricted at the base, and ultimately the knob or knobs are
separated off. The granules in the constricted part of the
outgrowth become elongated, spindle-, or rod-shaped (fig. 20)
dnring the constriction, but regain their spherical form as
soon as the Tueganuclei of the gemmnlge are free. No distinc-
tion whatever can be drawn between the minute structure of
the meganncleus where it is g-iving off these branches and that
of other regions — in other words this division is purely
amitotic. But it is not only amitotic, it is obviously unequal.
In the amitotic division of the meganucleus of many Ciliata
and of some Acinetaria (e. g. Dendrocometes) the division
may be an equal division, the two products of fission in the
former case, and the parent and geminula in the latter may
receive an equal portion of the parent meganuclear substance.
In the case of Dendrosoma it is obvions that the gemmula
receives only a small part of the total meganuclear substance
of the parent. The meganucleus not only divides in gemmula
formation but occasionally in ordinary vegetative growth,
as is shown by the fact that the meganuclear bands are
sometimes discontinuons. We have no reason to believe
that this division is brought about by any other process than
simple constriction.

The fragmented or disintegrated condition of the mega-
nucleus may be seen in many preparations. It is by no
means confined to the parts of the meganucleus in the stolon,
but mav occur in the basal portions of the arms. In some
cases scattered chromatin granules (fig. 12) may be seen in
the cytoplasm in regions where a distinct but not well-
defined meganuclear band occurs, but in other regions the
whole of the meganuclear structure appears to be scattered
in the cytoplasm (fig. 13). If it is a fact that the mega-

DEXDROSO.A[A RADIAXS, EHRENBERG.

157

nncletis under certain circumstances breaks up and is dissi-
pated in the cytoplasm, we are reminded of the condition,
long ago described by Grnber, of the meganuclear fragmen-
tation in the ciliata., Oxytricha and Lacrymaria. An
alternative hypothesis, however, might be put forward to the
effect that the condition we have observed is not disinte-
gration but construction of meganuclens; that the scattei’ed
ofrannles we have observed were not cast out from the meg'a-
nnclens, but are in process of formation in the cytoplasm, and
will be added to the nuclear bands.

This alternative hypothesis will be considered more fully
in a subsequent paper, bnt whatever hypothesis we adopt the
study of the sections exhibiting this fragmented condition
confirms most definitely the statements we have made above
— that (1) there is no true nuclear membrane, and (2) that the
granules of chromatin are not connected by a plastin net-
work.

Micronuclei.

The micronuclei of Dendrosoma have not been previously
described. Their distribution in the arms and stolon varies
a good deal, and it seems very probable that they are not
constant in position but move about in the arms and stolon,
driven hither and thither by the protoplasmic currents. On
this point, however, we cannot speak with certainty as we
have never been able to recognise the micronnclei in the
adnlt living specimens. We have been able to distinguish
them in some of the living gemmulae.

'I’liey are not difficult to see in some of the ordinary stained
whole mount preparations, although the crust of foreign
bodies adherent to the pellicle sometimes interferes with the
clear definition of the smaller internal structures.

'I'he micronuclei may occur both in the arms and in the
stolon. We have not observed them at any time very close
to the distal extremity of the arms, although in one or two
cases we have seen one or two micronuclei lying just beyond
or close to the distal termination of the meganucleus (fig. 50, m.,

VOL. 54, PART 2. NEW SERIES. 12

158

SYDNEY J. HICKSON AND .1. T. WADSWOETH.

and text-fig. A, mi.). The arrangement of the niicronuclei in
the arms and stolon is very irregular. In some specimens they
are found at fairly regular intervals of 30/t or 40 fi in the arms,
in others they seem to be in pairs (figs. 9, 10), indicating,
perhaps, a recent epidemic of division, but in several speci-
mens we have found clusters of three, four, five, or six micro-
nuclei irregularly scattered in tlie arms and stolon. In the
specimens from Birmingham we have noticed that, as a rule,
more rnici’onuclei are present than in the specimens from the
Bridgewater canal. In the clusters there are sometimes eight
or nine micronuclei together, and in one specimen there
Avere as many as nineteen (fig-. 15). This irregularity in the
arrangement of the micronuclei has sugg-ested the view that
they may move about in the living organism.

In Dendroconietes and in other Acinetaria the micronuclei
are usually surrounded by a clear area, the “hyaloplasm ” of
Maupas (fig. 14). Such a clear area is also seen round many
of the micronuclei of Dendrosoma, but it does not appear
to be so constant in character as it is in some of the other
genera. In a former jiaper (16) it Avas suggested that this
clear area may be due to a shrinkage of the micronucleus in
preservation, and therefore of the nature of an artefact.

This vieAv is confirmed by a cai-eful examination Avitli high
poAvers of the area round the micro.nucleus of Dendrosoma,
as it shoAvs not the faintest trace of protoplasmic structure
eA-eu in the most successfully stained preparations. Occa-
sionally a feAv very delicate strands may be seen stretching
across the area (fig. 14), indicating, perhaps, a former
organic connection betAveen the nucleoplasm and the sur-
rounding cytoplasm. But in addition to the perfectly clear
and unstainable area Ave can also distinguish a more irregular
area of cytoplasm around each micronucleus, Avhich is better
defined in some cases than in others (fig-. 13), that differs from
the rest of the cytoplasm in being relatiA'ely free from granules
and stains rather less deeply. It is this outer area Avhich
serves as a guide to the micronuclei and serA-es to distinguish
them from other chromatin granules.

DENDEOSOMA EADIANS, EHEENBERCt.

159

The minute structure of the resting micronuclei is very
difficult to determine as they stain so deeply; but we have
satisfied ourselves that there is a definite nuclear membrane
and a network of chromatin (figs. 22, 23).

The resting micronuclei are always spherical in form.
When seen in pairs and clusters they are about 3'3,u in
diameter; when isolated they are sometimes as large as 4‘5/tt
in diameter. Various stages in the division of the micro-
nucleus have been observed, but others are missing. Our
account of the process is not, therefore, quite complete.
What we have seen may be regarded as a modified form of
mitosis. It cannot be positively asserted that a more direct
mode of division of the micronuclei does not take place, but
as the different stages of division have been seen in areas
where gemmule foiunation has begun, and also in areas where
there is no evidence that gemmule formation is about to take
place, and as, moreover, we have never seen any signs of
direct division in the hundreds of micronuclei we have
examined, it seems probable that amitotic division of the
micronuclei never occurs.

The history of the mitosis so far as we can judge at present
is as follows : The micronucleus swells and then becomes
slightly oval in outline (fig. 24). The smallest specimen we
have seen at this stage is 9%3 p by 5T ,u. It is difficult to
determine whether the chromatin network has broken down
at this stage or not, but definite lines can be distinguished
running in the direction of the longer diameter. In the
next stage the oval shape is changed to a spindle shape and
the size is 15‘3/i by 6'8p (fig. 2(1). 'Phe chromatin seems to
be withdrawn from the points of the spindle, which are
usually quite clear at this stage. The chromatin is in the
form of a large number of minute granules connected by a
network of fibrils. It is possible that Ave have missed a
stage here, as we have not found a spindle form yet with the
chromatin collected together more definitely into an equa-
torial band.

We have found three or four examples of a stage in the

160

SYDNEY HICKSON AND ,T. T. WADSWORTH.

mitosis shown in fig. 2o. In this stage the chromatin
grannies are arranged in lines running from the poles towards
the equator, but there is a clear zone free from chromatin
running through the equatorial belt. The three examples of
micronuclei in this stage we have found measured 6’8 /u
by 6'3 ft, 8'o fi by 5'6 and 6'8ju by 5'7 /u respectively.
They are oval or nearly spherical in shape. Being small and
less pointed than the examples we have seen of the stage
shown in fig. 26 it might be supposed that they come eai’lier
in the mitosis. On the other hand, the separation of the
cln-omosomes leaving a clear zone in the equator suggests
that the metaphase has begun. If this is the correct inter-
pretation of them then it seems probable that the onset of
the metaphase is accompanied by a contraction of the
figure.

In the next stage the chromatin gi’anules are collected in
two groups at the poles of the figure, which is 14'5 ju hy 4 /u in
size (fig. 27). In the broad band connecting these poles
definite plastin lines can be distinctly seen. In the next
stage we have observed the poles are further apart, the total
length of the figure being 20‘4 /t (fig. 28). The poles ai’e
.5'1 fi in diameter and contain an immense number of evenly
scattered minute chromatin granules, but we have not been
able to discern any plastin network connecting them. The
band connecting the two poles has shrunk in the middle to
3‘5 fi in diameter. The same plastin lines may be seen in
this band as in the last stage. In the final stage (fig. 29)
the chromatin grannies are more concentrated towards the
inner hemisphere of each pole and there is an appearance of
a “ pole-plate” similar to that of Paramecium. The spindle
is much narrower. Our examples of this stage are smaller
than those of the last named, the measurements of the example
figured being — total length 15'6ju, diameter of the poles
3'4/j, diameter of the spindle P7 /i.

The history of the division of the micronuclei of Dendro-
soma that we have just described is different in many res-
pects from that we described in Dendrocometes in 1902 ; but

UENDEOSOMA RADIANS, EHRENBERG.

161

it must be remembered that in the case of Dendrocometes we
described only the division of the micronuclei in conjugation.
The mode of division of the micronuclei in gemmule-forma-
tion in Dendrometes differs in some respects from that seen
in conjugation. On this subject we hope to write a further
paper at a later date. We have not yet been fortunate
enough to observe any phase in the conjugation of Dendro-
soma.

It is possible that the minute granules of chromatin seen
in various stages of the division of the micronucleus in
Oendrosoma represent the chromosomes. With this view in
mind we have carefully compared them in the earlier and
later stages of the process to determine whether they are
double in character before their separation in the two poles.
In a recent paper Calkins and Cull (4) have shown that in
the earlier maturation divisions of the micronuclei of Para-
mecium aurelia the chromosomes divide longitudinally.
W e can only state that we have not been able to find any
evidence that the chromatin granules in these karyokinetic
figures of Dendrosoma divide at all. Evidence of this,
however, may be forthcoming from the stages we have
missed.

The Gemmula;.

The gemmulm were first discovered by Levick, but it is
difficult to understand from his figures v/hat is their exact
shape. If his tig. 4 is drawn accurately to scale theg’emmula
it represents was about 45 n in diameter. Kent describes the
gemmula? as “ hypotrichously ciliated embryos of relatively
large size.” Unfortunately he gives no statement of the size,
but according to his figure they are about 36‘3 fx in diameter.

Sand repeats KenPs statement that these bodies are hypo-
trichous, but adds that they are “ en forme de lentille bicon-
vexe aplatie,” and have three contractile vacuoles (in corre-
spondence with Kent^s figure).

In the living material we have examined from the Bridge-

162

SYDNEY HICKSON AND .T. T. WADSWORTH.

water canal two kinds of free-swimming gemmulm were
observed : a large kind which was plano-convex in side
view (fig. 1), provided with a band of several rows of cilia and
from six to ten contractile vacuoles ; and a smaller kind
(fig. 31), biconvex or loaf-shaped in side view, with an
equatorial band of three or four rows of cilia and pi-ovided
with only three or four contractile vacuoles.

lu this material, however, we have found the Acinetaria
attributed to the genera Trichophrya and Lern^ophrya
associated with Dendrosoma, and until recently we had no
definite evidence as to wliich of the three forms the gemmulas
belonged.

In the material obtained from Birmingham no Tricho-
phrya nor Lernseophrya forms were found, and the
gemmulm we found were all of the larger plano-convex type
(fig. 1). This observation suggested that the larger type
is the gemmula of Dendrosoma; but we obtained a definite
proof of the truth of this suggestion by our observations on
the development of two of these gemmulte into the definite
Dendrosoma forms as described below (p. 167).

AVe then examined carefully all our whole mount prepara-
tions for Lernmophrya forms showing gemmulm in the
brood-pouches, and found that the smallest of them was 24 /u
in diameter, and the largest about 37 fx (the outline of this
gemmula is indistinct and the measurement probably not quite
accurate). Tlie average diameter of eleven gemmulEe works
out at about 29' 6 n. The measurement of a lai-ger number of
gemmulae of Dendrosoma, in their brood-pouches, gave us
considerably greater diameters, but there are one or two
complications that must be mentioned before giving the actual
figures obtained. Whereas all the Lernaiophrya gemmulae
were approximately circular in outline, many of the gemmuhe
of Dendrosoma, in the aspect presented to us in the fixed
preparations, were definitely oval in outline.

The smallest of the fully formed gemmulae was 29'6 /x
(circular in outline), the largest 40'7 ju by 55‘5 fx (oval in out-
line). Many of the gemmulae we measured were not fully

PENDROSOMA KADIANS, EHKEXBEKG.

163

formed, and it may be tliat the size was not as great as it
would be when the outline was completed. There is no reason
to believe, however, that there is any considerable growth in
the gemmula after the first faint Hues marking out its contour
ai’e noticeable.

The average of the shorter diameters of sixteen gemmulaj
was about 35’5 /x and the average of the longer diameters
about 41 ju. We may i-oughly express, therefore, the diffe-
rence in size between the two kinds of gemmulae as a difference
of 29'6 p by 29 6 n in Lernaeophrya and 35'5 /x by 41 ^ in
Dendrosoma. There is not much difference in size, probably,
between the lai-gest gemmulae of Lernmophrya and the
smallest gemmulae of Dendrosoma, and there is clearly con-
siderable variation in the size of the gemmula3 of both
genera.

But still it may be considered a fact of some systematic
value that the gemmulm of Dendrosoma are larger than those
of Leriueoph ry a. AVe found, unfortunately, no Tricho-
phrya forms showing formed gemmulm, and we can therefore
give no near details of them. They have been described by
Biitschli, Sand, and others, but we have not found any
measurements given of them in the literature of the genus.

The gemmulae of Dendrosoma are usually found in well-
developed individuals, and are frequently situated at the
base of the arms (text-fig. A), but they also occur in the
course of the arms and moi’e rarely in the stolons.

'Pile gemmulae at the base of the arms is a very character-
istic feature of the specimens from the Bridgewater canal. In
the specimens from Birmingham the gemmula3 are more
frequently found some little distance above the base.

In the earliest stage of gemmule formation there is a slight
swelling in the arm, and two or three curved lines (fig. 19, o.)
appear in the cytoplasm and mark the external boundary
of the future • gemmula. A narrow crescentic space often
appears between these lines aud the cytoplasm. The mega-
nucleus sends off a short branch which ends in a knob-like
swelling towards the concavity of these lines (fig. 20). The

164 SYDNEY .1. HICKSON AND .T. T. '\YADS^Y01^TH.

base of this arm of the megauucleus gradually becomes con-
strictedj and at a later stage divides, leaving the knob-like
extremity as the meganucleus of the gemmula (conf. p. 156).

In several cases, particularly in specimens from Eiimiug-
ham, we have observed two of these processes entering into
the gemmula area, and we have several preparations of young
gemmuhie with two distinct megauuclei (figs. 16 and 21)'
When there are two meg’anuclei in one gemmula there seems
to be a great deal more megannclear substance in proportion
to the cytoplasmic substance than when there is only one
meganncleus. A comparison of the different gemmulse with
only one meganncleus renders it difficult to believe that there
is any definite relation between the cytoplasm and nucleoplasm
in gemmule formation.

One or two, or possibly in some cases more than two
micronuclei in the vicinity of this swelling increase in size
until they are about 11 p in diameter. One, or possibly more
than one of them, becomes spindle-shaped, attaining to a
size of nearly 14 p in length by 11 p in greatest diameter
(fig. 19). This microuucleus then divides by mitosis (see p.
159).

A\4iile the micronucleus is thus dividing the boundary lines
of the gemmula are in the process of completing the enclosure
of the gemmule area. The division of the micronucleus is,
however, completed, and the daughter-nuclei have consider-
ably shrunk some time before the area of the gemmule is
entirely delimited from the pi-otoplasm of the arm.

It is not possible for us to state definitely that there is any
constancy in the number of micronuclei taking part in the
bud formation.

In the bud that is still in the brood-pouch, shown in fig. 17,
there is only one micronucleus. In the specimen shown in
hg. 19 two micronuclei are taking part in the formation of
the gemmula; in the young form shown in fig. 18 there are
three micronuclei (only two are drawn, the third being hidden
by the meganncleus) ; in that shown in fig. 21 there are four,
and in that shown in fig. 16 there are seven. e have never

DKNDKOSUWA HADIAKS, EKKENBERU.

165

seen any evidence of division of the niicronucleus during' tlie
free-swimming gemmula stage nor in younger dxed stages.
It seems probable, tlierefore that the number of micronuclei
takiug pare in the formation of the bud may vary from one
to seven. In JJendrocom etes we also found that the
number is not constant, but varies from two to five.

In JJendrosoma then, as m Dendrocometes, the mega-
nucleus of the gemmula is formed by amitotic division of tlie
megauucleus of the parent, and the micronuclei of the gemmula
are formed by mitotic division of the micronuclei of the
parent.

The Fkee-Swiiiming Gemmei.a.

We have called attention to the fact that there is consider-
able variability m the size of the gemmulte of Deudrosoma.
The gemmula shown m fig. 1 was plano-convex in form, with
a broad girdle of several rows of cilia extending from the
middle almost to the edge of the plane surface, it was 6Up
ill diameter and 4U p in height, it was difficult to count the
number of contractile vacuoles or to be certain they were
constant in number, but there were certainly more than
tliree, and probably from eight to ten. Their rhythmic con-
tractions were not synchronous, and frequently, but not con-
stantly, two or three vacuoles in close contact made their
appearance when previously only one was seen.

The megauucleus in this form could be clearly seen in the
centre of the protoplasm when the gemmula Avas viewed from
above or below, it was sometimes spherical, but varied in
sliape, and frequently showed one or two lobate processes
like tlie pseudopodia of an amoeba.

This particular gemmula was kept under observation lor
two days, and gave rise to a young suctoriau, which was
clearly a Deudrosoma radians.

The Iree-swimming gemmula; of the second kind (p. iOl),
found ill the material from the iiridgew'uter canal, are pro-
bably the gemmula of the Leruteophrya forms. They have

166 SYDNEY .1. HICKSON AND J, T. WADSWORTH.

some resemblauce to the figures and description of the
gemmula of Trichophrya epistylidis given by Butschli
(2, PI. 78, 6h).

According to Sand, however, the reproduction of Triclio-
ph rya epistylidis is “par embryons internes multiples
(tons situes dans la meme cavite), en forme de leiitille bicon-
vexe, munis de 3 couronnes de cils et de 3-8 vacuoles con-
tractiles.”

According to Stein the four to eight gemmulae that may be
found in the brood-pouch of this species are produced by the
fission of a single gemmula.

W'^e have not observed more than a single gemmula in any
brood-chamber of either Lernseophrya or Dendrosoma,
but we have not seen enough specimens of Trichophrya
yet either to affirm or deny Sand’s statement that the gem-
mulm are multiple in that genus.

The gemmule formation iu the specimens of Bern aeo p h rya
resembles that described by Perez (25) for the specimens
obtained by him at Bordeaux in the curious fact that it some-
times occurs very early. In our figure (32), for example, we
have drawn an example of a very young Lernaeophrya (or
Trichoph rya f) whicli has only just settled down, but already
shows a fully formed gemmula in the brood-pouch.

^Ve cannot give a decided opinion as to whether all the
free-swimming geminulm of this biconvex type belong to
Lernmophrya or to Trichophrya, as we have not at
present been able to trace out their history after they have
settled down, but it is almost certain that they belong either
to the one genus or the other, and not to Dendrosoma or any
of the other Acinetaria associated with it.

3'hese gemmulae ai-e usually biconvex, with a slight con-
striction in the middle, an equatorial band of four rows of
cilia, and three contractile vacuoles (fig. 31). There is pro-
bably a considerable range of variation (see p. 162) in size
and shape. One of the conv'ex sides is sometimes rather more
flattened than the other ; there may be only three, or possibly
more than four, rows of cilia in the equatorial band (the exact

DENDROSOMA RADIANS, EHRENBERG.

1G7

Dumber of rows being difficult to determine unless the gem-
mula is seen sideways and the cilia are moving slowly). Tlie
number of contractile vacuoles may in some cases be two or
tour, but is usually three.

A remarkable point in the structure of these gemmulse is
that at present we have no definite evidence of the presence
of micronuclei. Neither in the free-swimming’ gemmul®
themselves nor in the preparations (both whole mounts or
sections) of the gemmulaj in the brood-chambers, nor in the
body of the Lernteophry as and Trichophryas have we
been able to find the micronuclei in any form.

Certain deeply stained granules are sometimes found
scattered in the protoplasm, and some of these may be, and
probably are, micronuclei. It is very improbable that these
or any other Aciuetaria are not heterokaryote, but we imagine
that in Leriunophrya, and possibly in Trichophry a too,
the micronuclei are very small or involved with the mega-
nuclei.

Further investigation is needful before the mystery of the
micronuclei of these forms is solved.

Develoi'jient of the Gemjiula.

We have observed the development of the gemmula in the
Birmingham material twice. The history of one of these,
September loth, 1908, was as follows:

A free-swimming gemmula settled down on the cover-glass
between 11.45 and 12 noon. The cilia disappeared entirely
in forty minutes, and as they disappeared short suckers
were developed all over the body, except, of course, the part
attached to the cover-slip. As to the method of disappearance
of the cilia we have nothing but the negative evidence to offer
that we have not seen them break off. The probability is that
they are withdrawn. We feel certain that they are not con-
verted into suckers. At 12.30 the embryo was in a similar
stage to that represented by fig. 3, and closely resembles the
stage figured by Biitschli for Trichophrya (2, PI. 78, 6 c.).

168

SYDNEY .1. HICKSON AND J. T. WADSWORTH.

It was slightly oval iu outline, the two diameters being
55’5 /ii by 518 /i. A preserved and stained gemmula of this
stage is shown in tig. 18. At 1.30 the tentacles had con-
siderably increased in number and in length on one quadrant
of the gemmula, but had diminished in number over the rest
of its surface. At 2.10 the main body of the gemmula had
increased to 62 p in diameter, but on one side a short arm
process (7'2 ,u), supporting a great many tentacles, had been
protruded. There was only one tentacle left on the main body.
Six contractile vacuoles were observed at this stage, and the
meganucleus was visible and had an amoeboid form with two
short, thick, pseudopodia-like processes. At 3.10 the arm
was 25'9 /i in length and the tentacles confined to its distal
extremity. For the first time a single contractile vacuole
w’as seen at the base of the arm. At 3.55 the arm had
increased to 33'3 in length, and a new arm at right angles
to it was beginning to be formed. At 4.40 the main body
was losing its circular outline. The new arm, about 14‘8 ju iu
length, exhibited tentacles, and the first formed arm was
48T in length. Three inicronuclei were clearly visible at
this stage in the clear protoplasm. The meganucleus had four
branches, one directed towards the longer arm, but not extend-
ing into it, one towards the shorter arm. Another specimen of
a Fendrosoma, which was probably of the same age as this
one, was found in a whole mount preparation and is shown in
hg. 21. The meganucleus is in two parts, but the general
form of it, apart from this peculiarity, is very characteristic
of young Deudrosomas of this stage. In this particular speci-
men there were four well-developed micronuclei.

The other case of the development of a young Fendrosoma
from a free-swimming gemmula is illustrated by the figures
2-7. The ciliated gemmule, at the time it settled down, had
from eight to twelve contractile vacuoles. The meganucleus
could be clearly seen, as it was quite colourless as compared
with the pale yellow tint of the surrounding cytoplasm. It
was distinctly amceboid, constantly, but slowly, changing
its shape (fig. 2). Before the cilia disappeared numerous

DENDROSOMA RADIANS, EHRENBERG.

169

short suckei’s were produced, scattered irregularly but
principally near the margin of the body. The body of this
specimen did not retain its circular form, but became irregu-
larly quadrilateral. After a period of two hours (fig. 4) the
cilia had disappeared and the suckers were mainly collected
at the two ends, but two or three odd suckers were observed
in the middle. In another hour there were three distinct tufts
of suckers (fig. 5), but two hours later still all the suckers
had disappeared except one isolated one, and a dense tuft at
one end (fig. 6). The next morning’, i.e. eighteen hours
later, the end supporting the tuft of suckers had grown
considerably to form a definite Dendrosoma arm (fig. 7).

The stage just described corresponds vvith that figured by
Savile Kent (in his Pi. 47, fig. 21), but differs from it in the
absence of suckers on the general surface of the body. We
have seen a good many specimens of this stage but never one
which, possessing a well-defined arm, had suckers scattered
over the rest of the body, as shown in KenPs figure. In
Kent’s figure of this stage only three contractile vacuoles
are shown. Levick, however, gave another figure in which
six contractile vacuoles were shown. The latter is in this
respect, as well as in the actual size, more in accordance
with our observations than the former.

'r h e s o - c a 1 1 e d “ e X t e r n a 1 buds .” — In the literature of
Dendrosoma reference is made by nearly all authors to
another method of reproduction than that by the gemmulge
previously described. Kent observed at or near the distal
extremity of the arms of many specimens a number of
spherical or oval bodies, which he believed to be “ exo-
genously produced germs similar to those of Acineta
mystacina of Stein.” It is probable that these are the
same bodies as those previously described by Levick as
“ovaries.” Biitschli describes them as “ angebliche freie
iiussere Knospen,” and Sand as “ gemmes externes ciliees
quelqiiefois tentaculees, produites a I’extremite des rameaux.”
We have found bodies similar in position, form, and size to
these so-called external buds in our specimens from the

170

SYDNEY J. UICKSOX AND J. 'J'. WAUSWOETH.

Hi-idgewater canal and from Birmingliam, and we have dis-
discovered that they are epizoic Acinetarians belonging to the
species U r n n 1 a e p i s t y 1 i d i s .

The possibility that the bodies described and figured by
Kent are different from those we have observed has of course
occuri’ed to us. It is, however, very improbable that external
buds could be formed in the position of these bodies for the
following- reasons :

In the formation of a bud it would be necessary for the
meganucleus to take part. The meganucleus of Deudrosoma,
however, does not extend as far as ttie extremity of the
branches, and could not possibly take part in the formation of
buds in the position assigned to the “ external buds ” by
8aville Kent.

"NVe have examined a great number of specimens ofpendro-
soma from the two localities, obtained at different seasons of
the year and in varying phases of activity, but we have never
seen any reproductive bodies of the form and in the position
assigned to the “ external buds”; butUrnula epistylidis
does occur in this position in a large majority of the specimens
examined, and frequently in considerable numbers.

On Uknula Epistylidis.

This interesting species was first described by Claparede
and Lachmann (5). It is mentioned in their first volume
(1858-9), but the reproduction is more fully described in the
second volume (1860-61). It was found on the stalk of an
Epistylis. Owing to the appearance of a branching
tentacle in one of their specimens these authors regarded
Urnula as a Khizopod and placed it next to the genus
Eugl}'pha in the family Actinophryina.

Engelmann (12) and subsequent authors have, howev-er,
agreed that it is an Acinetarian, and Butschli places it in the
family Urnulina with the genera Bhynceta and Acine-
topsis — an arrangement that is followed by Sand.

Only one species has been described — U rn ula epistylidis

DENDROSOJIA RADIANS, ElIKENBERG.

171

— and the measurements given bj Sand of tliis species are :
Length of test, 20/i-120n ; diameter of test and body, 15g-80//;
diameter of the nucleus, The genus is characterised

by the definite but very thin test, which is usually conical in
shape and curved proximally towards the disc of attachment.
There are one, or two (rarely more than two) tentacles. The
reproduction is by oblique unequal fission, and the smaller of
the two products of fission escapes as a ciliate gemmula.

The question whether the species of Urn u la we have found
on Dendrosoma should be referred to the species U. episty-
lidis or placed in a new species may be open to discussion;
but we can find no satisfactory reasons at pre.sent for adopt-
ting the latter course.

The specimens attributed to U. epistylidis by Sand and
other authors are very variable in size (15/<-80/t in body
diameter), so that the fact that the largest of the specimens
we have measured is not more than 30 /< in this diameter and
the average is about 25 /x does not signify more than that the
Urnula on Dendrosoma belongs to a small race.

In the original figure given by Claparede and Lachmann
the two tentacles are shown to arise from the side of the
body turned towards the host. In most of our specimens the
tentacles ari.se from the distal surface or apex, as shown in
figs. 35-39. The oinginal figure with the tentacles arising
from the sides has been copied in the subsequent papers and
books, and it seems to be b}" no means certain that this
origin is normal in the species. The body may rotate more or
less in the test and the appearance shown may be only
temporary, but we have observed only one or two cases in
which the origin of the tentacles may have been lateral.

3’he position of the contractile vacuole was not constant in
our specimens. It is usually situated, as shown in figs. 35-39,
near the centre of the distal convexity, but in several speci-
mens we have seen it more deep-seated. In the figures of the
species given by other authors it is shown by the outer side of
the nucleus. The structure of the test, which is extremely
delicate, appears to be the same as that previously described.

172

SYDNEY .T. HICKSON AND .T. T. ^YADS^YORT^.

ex:cept that we have observed a slight disc-like swelling at the
point of attachnient.

After full consideration of all these points of apparent
differences, and bearing in mind the possibility of considerable
variation in adaptation to the conditions, we have come to the
conclusion that the Urnnla found on Dendrosoma should be
referred to the species U. epistylidis (ClaparedeandLach-
mann).

It is not necessary to give a full description of the species,
but we will be content with a few remarks on some special
character’s. The tentacles are the most remai-kable featnr’es.
In the first place they are compai-atively rai’ely seen, the body
of the Urnnla being usually rounded off within the test and
at rest. In the majority of specimens which do exhibit
tentacles at all only one is seen. When there ar*e two they
usually cross one another as shown in fig. .39.

It is extremely probable that a specimen that at one tinre
exhibits only one tentacle may at another time exhibit two, or
even three, tentacles. The tentacles of Urnnla differ from
those of the typical Acineta.ria in two respects. They are
relatively ver*y long and flexible, moving actively with cnrions
sei’pentine curves as if in search of food. They do not
terminate in a sucker.

When fully extended they are very delicate and attenuate
at the distal extremity to a very fine point (fig. 35). When
partially retracted or not fully extended they are much thicker,
show a spiral marking (fig. 45), and terminate in a spindle-
shaped, or sometimes bluntly club-shaped, extremity. There
can be no doubt that thev are protruded and withdrawn into
the body with considerable rapidity.

We have not been able to satisfy ourselves as to the food
of Urnnla. We have frequently observed a tentacle bent
over towards the head of the Dendrosoma, with its pointed
end buried among the bases of the Dendrosoma suckers.
This attitude suggested that the Urnnla is parasitic on the
Dendrosoma, and this suggestion is confirmed by the fact
that the heads of Dendrosoma affected by U rnnla do not look

DENDKOSOMA RADIANS, EHRENBERG.

173

so healthy as those that are free from them. On the other
hand, healthy Urnnlas are frequently found on Dendrosoma
itself in positions that would not permit them to penetrate
the delicate unpi'otected ectosarc of the head region, and also
on other bodies, such as weeds, stalks of Epistylis, etc. We
have, moreover, never observed a stream of food particles
passing from the Dendrosoma to the Urnula body in the
tentacle that is apparently attached to the former, as we
should certainly find if the latter were feeding parasitically
upon it. We are inclined to the opinion, therefore, that the
Urnula is epizoic and not strictly pai-asitic.

The meganucleus is usually spherical in shape and central
in position. In one specimen (fig. 40) in which the diameter
of the body as preserved was 19 q, the diameter of the mega-
nucleus was 8’5 /n. The chromatin of the meganucleus is usually
in the form of a network. It never shows the granular character
that is such a marked feature of the meganucleus of Dendi’O-
sotna. It i.s sometimes difficult to determine whether a micro-
nucleus is present or not, but a small deeply staining granule
about T7 /.i in diameter may frequently be observed in sections
which we believe to be the micronucleus (fig. 40). No stages
in its enlargement or division have been observed.

We have observed the same method of reproduction in
our specimens as that previously described for the species
by Claparede and Lachmann. The individual divides by
oblique fission into two parts, one usually larger than the
other (figs. 41, 43). Of these the smaller becomes holo-
trichously ciliated and escapes. The larger may remain in
the undivided lorica and increase in size until it is again full
grown. Of this, however, we have no positive evidence. It
is possible, however, that the escape of the smaller product of
fission entails the death of the larger product, but if this
were the case we should expect to find attached to the Den-
drosoma a certain number of empty loricse. We have, how-
ever, never found an empty lorica attached to the Dendrosoma
nor any signs of degenerating protoplasm in a lorica.

The Urnula was found on specimens of Dendrosoma from

VOL. 54, PART 2. — NEW SERIES. 13

174

SYDXKY J. HICKSON AND .1. T. WADSWORTH.

both localities. Sometimes a few specimens may be found
that are quite free from these epizoites, but it is very rai-el}''
the case that a single collection of Dendrosoma is brought
in that does not show some infected specimens. The number
varies a good deal, but there is no reason to believe that
they are more numerous at one season of the year than at
another.

The settlement of the gemmiila and the development of
the lorica have been observed by one of us (W.) on two or
three occasions. The free-swimming holotrichously ciliated
geininnla is about 20 by 15 ju in size. The cilia stop moving
and begin to disappear about ten minutes after settlement
on the Dendrosoma is effected. The lorica must be formed
very rapidly as the protoplasm is contracted into an oval
form near its free end, as in fig. 35, about five minutes after
the settlement. The cilia ai’e still plainly visible, but in
another five or ten minutes they disappear. A single tentacle
begins to grow out a few minutes after the cilia have dis-
appeared.

A curious feature that was observed on both occasions was
the presence of two or three minute capitate tentacles at the
time the cilia are disappearing-. They are, however, only
present for a few minutes, and cannot be recognised at all
when the characteristic Urnula tentacle is developed.

In the figure given by Saville Kent one of the supposed
“external buds” is drawn with six short capitate tentacles.
It is possible that Kent may have observed an Urnula that
had just settled down and still retained the temporary
capitate tentacles we have described.

Further Remarks on the Systematic Position of Dendro-
soma, Lern.eophrya and Trichophra'a.

The relation of these three genera has already been briefiy
referred to in the introduction, but a further summary of the
characters that distinguish them may be useful in the light
of the observations we have recorded in this paper.

DEXDROSO^[A RADIAXS, ERREXBERG.

175

The genus Trichophrya was described by Claparede and
Lachmanii, 1858-61. The original type-species is T.
epistvlidis, a common species usually attached to the stalk
of Epistylis. We have found it frequently in the Bridgewater
canal collections, and the specimen we have drawn in fig.
46 Tr. was about 129‘5/_t by 111 ju in size.

'Paking this as a type we may say that it differs from a
full-grown Dendrosoma in its small size and the relative
shortness of its anus. It might be thought to be a young
Dendrosoma, but it differs from the young Dendrosoma in
having several shoi’t arms instead of only one or two. The
voung Dendrosomas shown in figs. 47 and 48 are smaller than
the Trichophrya shown in fig. 46, but nevertheless exhibit
the characteristic Dendrosoma form.

There inust also be some important difference between them
in the character of the micro-nucleus, but the nature of this
difference we cannot describe. It is perfectly easy, as our
figures show, to demonstrate the presence of micro-nuclei in
young Dendrosoma, but we have not yet been able to find
definitely the micronucleus in any specimen of Trichophrya
we have examined.

In Trichophrya epistylidis, according to Stein,
Biitschli, Sand, and other writers, the single gemmula that is
formed in the brood-chamber may divide into four or eight
gmnmuhe before liberation. AVe have not observed a similar
mode of repi’oduction either in Lernfeophrya or in Dendro-
soma.

The figure given by Biitschli of the free-swimming
gemmula of T. epistylidis shows that it must be very
similar in shape to the gemmula we have ascribed to
Lernajophrya (fig. 31).

Several other species of the genus have been described by
Sand and others, but of these we have very little detailed infor-
mation. Some of them, such as T. salparum, T. amoe-
boides, T. odontophora and T. mirabilis are marine.
One of these at least, T. mirabilis, found attached to hydroids
at Banyuls, may possibly be more closely related to the genus

176

SYDNEY .T. inCKSON AND ,T. T. AYADS^YORTH.

Leriifeophi’ya as it is characterised by its very long
suckers.

The genus Lerupoophrya was described in 1903 bv
Perez (25). We have found it in the Bridgewater canal, and,
like the Bordeaux type-specimens, attached to Cordylo-
ph ora.

Lernaeophrya is a larger form than Trichophrya.
According to Perez it may attain to a size of 400 ju.
Our specimens are not as large as this, bnt we have fonnd
them over 200 /x in length (fig. 49).

They difPer from both Trichophrya and Dendrosoma
in the extraordinary length of the suckers. Perez says he
has measured suckers 400 /« in length. Our specimens
were smaller than his, but we have found some of the
suckers to be over 275 n in length. Perez states that the gem-
mulpB are frequently formed at a very early stage, befoi’e the
arms are formed. We have fonnd the same peculiarity in some
young forms which we attribute to Lern asophry a. In our
fig. 32 we have drawn a young Acinetarian, which is probably
a young Lernaeopli rv a, although we have no conclusive evi-
dence to prove that it is so, in which there are no arms and
only four suckers, but it nevertheless contains a full-gi’own
gemmula in its brood pouch.

As in 3'richophrya so in Lerna'ophrya the micronnclei
have at present escaped onr observation, and as Perez does not
mention these structures in the description of his specimens
it is possible that some peculiarity of the micronuclei, Avhich
renders them obscure in the resting stage, is a character
which Lernaeophrya and Trichophrya have in common.

Only the one species, L. capitata Perez, has at pre.sent
been described. Onr specimens do not appear to differ from
the type except in .size, and we are inclined, therefore, to
regard them as a small race of the type-species.

The gemmulae described by Perez were 50 fi in diameter,
whereas the largest we have measured were only 37 /i in
diameter, but in other respects they seem to agree.

The important characters in which the genus Dendro-

J)ENDROSOMA EAJJIANS, EHKENEEKG.

177

soma differs from Trichophrya and Lerumoplirya are
its greater size, the greater length of its arms, and the
characters of the gemmula. The suckers of Dendrosoma
vary a great deal in length, according to circumstances, but
they never attain to the same actual or relative length as the
suckers of Lernasophry a. The free-swimming gemmulm
of Dendrosoma differ from the n'emmulm which we attribute

O

to Lernmophrya in size, in shape, in having several instead
of only three or four bands of cilia, and in having several
contractile vacuoles instead of only three.

A prolonged study of the specimens of Dendrosoma from
the Bridgewater canal and from Birmingham give some
grounds for the view that they belong to different species.

These differences have been jn-eviously mentioned (p. 147) ;
it is only necessary in this place to refer again to the difference
in the number of the micronuclei. In looking through a
number of preparations of specimens from the tw'o localities,
the large number of the micronuclei iu the Birmingham speci-
mens is often a very .striking feature. To take two extreme
ca.ses, the piece of an arm that is drawn iu fig. 15 showing
nineteen micronuclei in a clu.ster round the meganucleus, and
a gemmula showing seven micronuclei, such as that drawn in
fig. U), we should recognise at once as belonging almost
certainly to the Birmingham variety. On the other hand,
when the micronuclei are isolated or in pairs at considerable
distances apart, as shown in text-fig. 1 and in figs. 9 and lU,
there would be a strong probability that they were taken from
specimens of the Bridgewater canal variety.

But nevertheless specimens of the canal variety are some-
times found iu which several microuuclei are aggregated
together, as shown, tor instance, in fig. 8, where six micro-
nuclei form a cluster, and in many specimens from Birming-
ham the microuuclei are scattered iu very much the same way
as in the canal variety.

To endeavour, therefore, to make a specific character of the
number of the micronuclei, a character which is obviously
subject to great variation, would be a task of great difficulty

178

SYDNEY .1. HICKSON AND .1. T. WADSWORTH.

and very little practical value. Joseph (18) has shown
that there is a great range of variation in the number of
inicronuclei in the Ciliate Loxodes, and it is clear from this
and from other evidence that it is not safe to base specific
differences on the number of the micronuclei.

SuMMAKY OF KeSULTS.

Specimens of Uendrosoma Avere found in the Bridgewater
canal attached to C ordy lophora, Polyzoa, and Aveeds. They
differ in some respects from the type of Dendrosoma
radians, and may be regarded as constituting a distinct
race.

The meganncleus does not extend to the extremity of the
arms as pre\dously described and figured. It has no true
nuclear membrane and no linin or plastin supporting netAvork,
but consists of numerous chromatin granules, “ chromidia,”
floating freely in a nuclear sap.

There are numerous micronuclei, usually about 4 g in
diameter, Avhich divide by mitosis. Beproduction is effected
by the formation of internal buds, “ gemmulm.” They are
plano-convex in form, 85'5 x 41 g in diameter, have several
contractile A'acuoles and a broad band of cilia. The descri])-
tion of these gemnmlm given in this paper differs in some
respects from that of previous authors.

The gemmula3 of Lernmophrya are also described.

The development of the gemmuhc of Dendrosoma is
described.

ddie “external buds’^ of Saville Kent are proved to be
epizoic Acinetaria belonging to the species Urnula epi-
s t }■ 1 i d i s .

Litekatl'ke.

1. AAverinzeAv, S. — “ Astrophrya areiiaria,” ‘ Zool. Aiiz.,’ xxvii, ia04.

p. 42.5.

2. Biitsclili, (). — “Protozoa. Abt. iii. Infusoria," in Bronn’s • Klassen

u. Ordnnngen.’ Leipzig, 1887-89.

DENDROSOMA RADIANS, EHRENBERG. 179

3. Calkins, G. N. — “ Protozoa,” Columbia University, ‘ Biol. Series,’

1901.

4. and Cull, S. W. — “The Conjugation of Paramsecium

aurelia,” ‘Arch. Protist.,’ x, 1907.

5. Claparcde, E., and Lachmann, J. — ‘ Etudes sur les Infusoires,’

Geneva, 1857-1861.

6. Collin, B. — “Note preliminaire sur nn Acinetien nonveaii,

Dendrosoiuides paguri,” ‘Arch. Zool. Exp.,’ (4) v, 1906,
p. 64.

7. “Note preliminaire snr qiielcpies Acinetiens,” ibid., (4), vii,

1907. Notes et Revue.

8. Delage, Y., and Herouard, E. — ‘ Zoologie Concrete Protozoa,' 1898.

9. Ehrenberg, C. G. — “ liber eine neue Thiergattung,” ‘ Monats.

Ak. Berlin,’ 1837, dee. 11, p. 152.

10. Polygastrica exclusis Bacillaries,” ibid. , 1840, p. 199.

11. “ “ Uber die seit 27 Jahren noch wohl erhaltenen Organisa-

tions Praparate,” ‘ Abh. Ak. Berlin,’ 1862, plate iii.

12. Engelmann, W. — “ Zur Naturgeschichte der Infusionsthiei-e,”

* Zeitschr. wiss. Zool.,’ xi, 1862, p. 370.

13. Entz, G. (sen.) — “ Ueber einige Patagonische Protozoen,” ‘ Math.

naturw. Ber. Ungarn,' xxi, 1903, published 1907.

14. Greenwood, M. — “ The Macronucleus of Carchesium,” ‘ Journ.

Physiol.,’ XX, 1896, ]). 427.

15. Hickson, S. J. — *• Heterokaryota ” in Lankester's ‘A Treatise on

Zoology,’ part i, fasc. 2, 1903, p. 421.

16. and AV^adsworth, J. T. — " Dendrocometes paradoxus,”

Part 1, ■ Quart. Jouni. Micr. Sci.,’ 45, 1902, p. 325.

17. “ Note on the Structure of Dendrosoma radians,”

■ Rep. Brit. Assoc.,’ 1908, Dublin.

18. Joseph, H. — " Beobachtungen iiber die Kernverhaltnisse von

Loxodes,” ‘Arch. Protist.,’ viii, 1907, p. 344.

19. Saville Kent. — ‘ Manual of the Infusoria,’ 1881-2.

20. Lang, A. — " Lehrbuch der Vergleichenden, Anatomie,” ‘Protozoa,’

1901.

21. Lankester, E. R. — Article “ Protozoa,” ‘ Zoological Articles,’ 1891,

p. 36.

22. Leidy, J. — “Notice of some Fresh-water Infusoria (Cothurnia

and Dendrosoma),” ‘ P. Ac. Philad.,’ 1874, p. 140.

23. Levick, J. — "On Dendrosoma radians,’’ ‘ Midland Natiu’al.,’ iii,

1880, p. 29.

180 SYDNEY J. HICKSON AND J. T. WADSWOETH.

24. Miirtin, C. H. — “Some Observations on Acinetaria,” ‘ Quart. Jonrn.

Micr. Sci.,’ 53, 1909, p. 353.

25. Perez, Cli. — "Sur un Acinetien Noiiveau,’ ‘ C.R. Soc. Biol.,’ 1903,

p. 98.

26. Sand, R. — ‘ Etude nionograpliique des Infusoires tentaciiliferes,’

Brussels, 1901.

27. Stein, Fr. — ‘ Der Organismus der Infusionstliiere,’ Leipzig. 1868.

EXPLANATION OF PLATE X,

Illustrating the memoir by Messrs. Hickson and Wadsworth
on Dendrosoma radians.

Lettering.

A. Arms of Dendrosoma. chr. Chromatin granules, c.v. Contractile
vacuoles. D. Swollen end of the arm of Dendrosoma in tig. 30.
E. Eujilotes. Rp. Epistylis. y. Gemmula in lirood-pouch. 31. Mega-
nucleus. 31.y. Meganucleus of gemmula. m. Micronucleus. 0. Out-
line of gemmula in fig. 19. s. Sucker, sf. Stream of food-particles.
f. Tentacle of Urnula. Tr. Trichophrya in fig. 45.

Figs. 1-7. — Illustrating the free-swimming gemmula of Dendrosoma
and its development after fixation.

Fig. 1. — Side view of the free-swimming gemmula, showing the
band of several rows of cilia and four of the peripheral
contractile vacuoles. X 250.

Fig. 2. — Surface view of a gemmula immediately after fixation.
Tlie meganucleus (31.) has an amoeboid form.

Fig. 3. — Gemmula as seen about thirty minutes after fixation,
showing the suckers (s.) that have begun to sprout out
from the general surface. Cilia are still present but
comparatively few in number.

Fig. 4. — Young Dendi’osoma two hours after fixation.

Fig. 5 . — Young Dendrosoma three hours after fixation.

Fig. 6 . — Young Dendrosoma five hours after fixation.

Fig. 7. — Young Dendrosoma one day after fixation. All the
suckers are now confined to the extremity of the single arm.

Fig. 8. — Section through a part of the stolon of a Dendrosoma showing
a cluster of six micronuclei (»i.).

J3ENDK0S0MA lUDlANS, EHEENBERG.

181

Fig. 9. — Section through an arm on the same slide showing the
micronuclei in pairs. Each of these micronuclei are aljout 3'.5 /t in
diameter.

Fig. 10. — Section through another arm in the same preparation
showing another pair of micronuclei and a contractile vacuole.

Fig. 11. — Section through a portion of a stolon showing a double
and contorted meganucleus.

Fig. I'i.. — Section through a portion of an arm showing chromatin
grains scattered in the cytoplasm. M., the main meganucleus.

Fig. 13. — Section through another part of an arm showing a single
micronucleus (»i.) about 4 // in diameter and the fragmented meganucleus.

Fig. 14. — Section through another arm more highly magnitied to
show tlie structure of the cytoplasm and nuclei. The micronucleus is
3 3 fi in diameter, the largest chromatin granules in the meganucleus
about l o jx ill diameter.

Fig. 1.5. — Drawing of a part of the arm of a whole-mount preparation
of a sjiecimen of Dendrosoma from Birmingham showing a cluster of
nineteen micronuclei.

Fig. lb. — Section through a gemmula of Dendrosoma from
Birmingham showing two distinct meganuclei (My.) and seven micro-
nuclei (tn.). The diameters of this gemmula were 5.5 /i X 48 /i.

Fig. 17. — Transverse section through an arm showing a newly
formed gemmula in position. The meganucleus of both the gemmula
and of the arm have discharged some chromatin grains (chr.) into the
general cytoplasm. The diameter of the bud is 22 ji, of the micro-
nuclei about 5‘5 y.

Fig. 18. — A gemmula soon after it has become fi.xed from a stained
preparation, showing the liand of cilia (c.) and two suckers (s.). The
diameters of the gemmula are 38 x 37 fi and of the micronuclei 5 /i
and 4'8 y,.

Fig. 19. — An oblkpie section through an arm of Dendrosoma showing
an early stage in the formation of a gemmula. At o. are shown the
curved lines that mark the Vioundai'y of the gemmula. Two micro-
nuclei of the arm have enlarged previous to division. The sizes of the
micronuclei (m.) in this preparation were 15‘3 fi x b'8 n and 9'3 /t X 5'1 y
respectively.

Fig. ‘2U. — Transverse section through an arm showing the outline of
a gemmula and the method by which a part of the meganucleus of the
arm is pinched off to form the meganucleus of the gemmula. In this
preparation some of the contractile vacuoles could be seen. No micro-
nuclei were obseiwed in this section. The diameter of the gemmula
from u-h was 37 '4 y.

182 SYDNEY J. HICKSON AND J. T. ^YADS\YOKTH.

Fig. 21. — Young Dendrosoma observed alive from free-swimming
gemmula stage (September, 1908) and then fixed. It is 85 /j x 70 /i in
size, the arms about 20 /j. in length, and the four micronuclei each aljout
4 n in diameter.

Figs. 22-29. — Series of stages seen in the mitotic division of the
micronuclei. X 1000.

Fig. 22.— A full-sized micronucleus in a resting condition.

Fig. 23. — Enlarged mici’onucleus previous to mitosis.

Fig. 24. — Micronucleus in stage of spindle-formation.

Fig. 25 . — Stage of division in which there is an equatorial baud
free from chromatin.

Fig. 20. — A stage in mitosis occasionally seen in which the
2)oles are j^ointed and free fi-oni chromatin. The relation
of this stage to tlie other stage in mitosis is not clear
(see jj. 159).

Figs. 27-29.— The chromatin is, in these stages, collected into
the two 2>oles wdiich are connected by an achromatic
S2>indle.

Fig. 30. — Dendrosoma feeding u^jon a EujDlotes (E.). The swollen end
of the aian of the Dendrosoma (D.) was 15 fi in diameter, the length of
the Exqjlotes body lUO /x. A stream of jxarticles (St.) could be seen
2>assing down into the arm through the attached suckers ; the other
suckers were (piite indifferent.

Fig. 31. — Side view of a gemmula of Lernajoixhrya (!■') showiug two
of the three contractile vacuoles. None of these gemmuhe exhilxited a
micronucleus.

Fig. 32. — A very young, ^u'obably (juite recently fixed s^xecimen of
Lerna'ojxhrya with only four suckei's, showing a com^xletely formed
gemmula in position. There are no micronuclei to be seen.

Figs. 33 and 34. — Two sketches of young Dendrosomas showing the
method of arm formation in Dendrosoma.

Figs. 35-39. — Series of studies of Urnula e^xistylidis e2>izoic on
Dendrosoma showing the different forms assumed by the tentacles. In
figs. 35-38 the S2)ecinieus have only one tentacle, in fig. 39 it has two.

Fig. 40. — Section through an Uniula from a stained 2U'eparation. In
this S2iecimen the diameter of the laxly is 19 fx, of the meganxxcleus (M.)
8'5 fx, and of the micronucleus (m.) 1'7 /x.

Fig. 41. — Transvei’se section through an Urnula after the formation
by fission of a gemmula (y). Drawn to the same scale as fig. 4(t

Fig. 42.— A S2)ecimen of Urnula e2Jistylidis, showiug the body
retracted below the mouth cf the test. Co2xied from Engelmann (5) (PI.
30, fig. 13).

DENDKOSOMA KADIANS, EHRENBEEG.

183

Fig. 43. — Urnula epistylidis, showing the formation of the
gemmnla. Copied from Claparede and Lachmann (5) (PI. 10, fig. 3).

Fig. 44. — Free-swimming gemmnla of Urnula. Copied from Clapai-ede
and Lachmann (5) (PI. 10, fig. 3).

Fig. 45. — A portion of the body of an Urnula very mnch enlarged to
show the spiral marking of the tentacle (C).

Fig. 46. — A specimen of Trichophrya epistylidis (s^j. .‘') found in
the Bridgewater canal attached to the stalk of an Epistylis. (From a
stained preparation.) No microuucleus could be seen. Size 129 5 /i
X 111

Fig. 47. — A very young Deiidrosoma with one arm and one micro-
nucleus, also attached to an Epistylis stalk. Tlie size of this sj)ecimen
is 6P3 n in gi’eatest length, including the arm. From a stained pre-
paration.

Fig. 48. — Another rather older Deiidrosoma with three micronuclei.
Size 60 IX, -f the arm 60 /i = 12(f fi. From a stained preparation.

Fig. 49. — Lernaeophrya (sj). ?) from the Bridgewater canal. Drawn
from a living specimen January, 1909.

Figs. 50-52. — Three figures drawn to the same scale ( x 1000) to
show the varying structure of the meganucleus of Dendrosoma.

Fig. 50. — Section through a part of an arm (Birmingham
material) in the region where the meganucleus terminates.
The terminal extremity was in the direction of the upper
side of the figure, but was not included in the actual
section. Two micronuclei are seen beyond the mega-
nucleus. The size of the largest granules was only 0'85 /x.

Fig. 51. — Section through an arm showing two ineganuclear
bands. The chromatin granules are smaller than in fig.
50, but the meganucleus contains peculiar, large, iiregular
bodies which give the chromatin reaction.

Fig. 52. — Section through a meganucleus (Bridgewater canal
material). The largest chromatin granules seen in this
section are 2'5 p in diameter.

Snu/r^jKijc/T^Sct,. Vol.SJf-.NS :0^.1O

STRUCTURE OE THE EXCRETORY ORGANS OF AMl’KIOXUS. 185

On the Structure of the Excretory Organs of
Amphioxus.

Part 2. — The Nephridium in the Adult. Part 3. — Hatschek’s
Nephridium. Part 4. — The Nephridium in the Larva.

By

Edwin S. Ciioo€li‘i<-li, F.IC.K.,

Fellow of Merton College, Oxford.

With Plates 11 — 16, and 1 Text-figure.

Part 2. — The Nephridium in the Adult.

In Part 1 of this contribution, which appeared some years
ago (7), it was shown that the nephridia of Amphioxus bear
a startling resemblance to the nephridia of certain Polychaete
worms, such as Phyllodice ; they are segmental, they are
formed of an excretory canal opening to the exterior (atrium),
but ending internally in blind branches; these blind ends are
provided with typical solenocytes. The last fact was the only
striking novelty then contributed to the descriptions of Weiss
(13) and Boveri (1). Although I can now add but little of
real importance to my previous account, it is necessary to
return to the subject again owing to the publication by
various authors of certain statements as to the presence of
internal openings. These statements, if not soon disproved,
will spread confusion and error in the literature of the subject
which it may take years to eradicate. Had not other pressing
work prevented me, I should myself long ago have attempted

186

EDWIN S. GOODRICH.

to dispose of tlieiu. Sucli convincing evidence can now be
brought forward against these views, tliat it may be hoped
the question will soon be definitely settled. Moreover, I take
this opportunity of adding certain details which serve to
complete our knowledge of these interesting organs.

In 1904 Boveri published a note (la) in which, while
accepting my description of solenocytes, he still maintained
that the lumen of the nephridial canal opens into the dorsal,
or hyper-branchial, coelom by one or more funnels.

Felix, in his excellent acconnt of the development of the
excretory organs of the Vertebrata (2), fully adopted my
view as to the structure of the nephridia of Amphioxus, and
gave some figures, derived from Boveri’s original paper (1),
but “corrected after Goodrich.” In these figures the funnels
wei-e closed up.

Shortly afterwards Felix changed his opinion, having exa-
mined Boveri’s sections, and republished the latter’s figures
in their original condition (with open funnels) in a second
work on the excretory organs of the Vertebrata (3). K. C.
Schneider likewise accepts Boveri’s description, but gives no
new figures to support his opinion (12).

Xow, it may at once be stated that I am firmly convinced
that such internal funnels do not exist. Indeed, I am
prepared not only to affirm that they do not occur in any
Amphioxus I have examined, but also to prove the correctness
of my description to any competent person who is willing to
look at my preparations. My affirmation is based on a long
and patient study of numberless specimens, both living and
preserved. It is naturally to sections that one turns for the
final verdict, and I may say that, although I have examined
hundreds of sections of specimens of different sizes and ages,
preserved according to a variety of methods, cut in all
directions, stained in various ways, never once have I been
able to discover such an opening. Occasionally, if the section
is broken, the preservation defective, or the staining im-
perfect, one may meet with what at first sight appears to be
a communication between the coelom and the nephridial

, STRUCTURE OF THE EXCRETORY ORC4ANS OF AMPHIOXUS. 187

canal; but this deceptive appearance is soon exposed on a
more critical examination of the preparation. Thick sections
are especially misleading. ISTo observation made on a section
more than 5 thick is in the least conclusive. The technical
difficulties are very great in the study of Amphio.Kus;
the tissues are brittle, the cells very small and difficult
to stain satisfactorily. Formol and Flemming’’s fluid, cor-
rosive-acetic, and picro-sulphuric-formol are all good pi-e-
servatives. Great care must, however, be taken to avoid
shrinkage, and for this purpose the method of double
embedding in celloidin and paraffin is most useful. By far
the best sections are obtained from pieces of the pharynx
removed from the fresh animal, and preserved separately.
One may use either carmine or hmmatoxylin for staining the
nuclei ; but it is quite essential to add some suitable cyto-
plasmic stain such as acid fuchsin. For the particular
])urpose Ave are now concerned with, perhaps some strong
staining reagent like Mann’s methyl-blue eosin is the best
for working out minute details under high powers, though
picro-nigrosin also yields valuable results.

Turning now to the structure of the nephridium, we find
the external pore opening at the very top of the atrial cavity,
on the anterior outer surface of the secondary or tongue bar
{op., figs. 1, 2, 7, and text-figure). The pore leads into
a canal which gives off a short posterior limb, and a much
longer anterior limb. The latter passes forwards to the next
primary bar, and downwards into the triangular coelomic
cavity delimited by the ligamentum denticulatnm. In a
fully developed nephridium both the anterior and posterior
limbs give off diverticula of varying length, which may
sometimes branch. These are shown in fig. 1 of Part 1 (7),
and are seen again in the reconstructions given in this paper
(figs. 1, 2, 3).

Let us pass to the conclusive evidence which can only be
obtained from sections. The wall of the nephridial canal
contains many nuclei (figs. 7, 13). In some places they are
so closely packed that they seem to press against each other.

188

EDWIN S. GOODlilCH.

In other regions of the canal they may be more sparsely
distributed. Cell outlines are rarely visible. The cytoplasm

Diat^nun of a, section tlirough the nepliridimn, passing along a
plane at right angles to the long axis of the animal, and
parallel to the gill bar.

a. e. Atrial epithelium, h. Base of secondary gill-bar. hv.
Blood-vessel, c. ep. Coelomic epithelium, cli. Chamber con-
taining solenocytes. v. Wall of nephridial canal, op. Ne-
phridiopore. koI. Solenocyte cell, and t. its tube containing
a flagellum.

usually contains numerous granules of an excretory nature.
As we approach the tip of a diverticulum, we find that the

STEUCTUKE OE THE EXCEETOEY OEGANS OF AMriIIOXDS. 189

nuclei do not gradually decrease in number, but suddenly
stop in the immediate neighbourhood of the solenocyte tubes
(figs. 6, 20). Here, where these tubes spring out of the
canal, there are no nuclei; but the wall itself is continued as
a sheet of more or less granular cytoplasm completely closing
off the lumen of the canal (figs. 6, 9, 20, 21). This canal wall
may be thick or thin, the variation in thickness depending,
I believe, chiefly on the state of tension of the fluid inside
the canal. In good thin sections the wall is always visible.
Indeed, the better the section, and the more perfect the
stain, the clearer becomes the limiting wall, whatever may
bo the direction in which it is cut.

Figs. 19 and 20 represent two sections taken parallel to
the surface of the nephridium, sagittal sections of the animal.
The first just shaves through the outer wall of the canal, and
shows many solenocytes lying on the blood-vessel. The
second, which ouly corresponds to the left hand portion of
the first figure, cuts deeper into the canal through the
extremity of one of the branches, where may be seen the
solenocyte tubes piercing the closing wall. In the next section
the nuclei of the opposite side begin to appear, the whole
thickness of the small solenocyte-beariug offshoot having
been nearly cut through, 'fhe following section would show
only a slice of the wall. There is no opening. Fig. 21 gives
a similar view of another nephridium in the same animal.

Two consecutive sections through the lowermost tip of the
anterior limb of the nephridium are drawn in figs. 13, 14.
Here again are seen the tubes piercing the wall, in which
there is no trace of an opening.

Figs. 5 and G represent sections from a series nearly
transverse to the animal and parallel to the bar. That in
fig. 5 passes through the external- pore, and shaves off the
wall of a diverticulum. The next section (fig. 6) cuts through
the extremity of this diverticulum. It is seen that the lumen
is closed off from the coelom by a distinct cytoplasmic wall,
through which pass solenocyte tubes. In fig. 7 is drawn a
portion of the same section when the microscope has been

VOL. 54, I'AKT 2. — NEW SEKIES. 14

190

EDWIN S. GOODKICH.

focussed to the lower surface ; the nuclei of the wall are again
visible. There is no opening.

Innumerable figures could be given of series of sections all
telling the same story. But the critic will say : it the diver-
ticula are really closed, sections taken at right angles through
their tip should show the tubes cut across embedded in the
thickness of the wall. Such sections are not difficult to find,
and I figure several on Plates 12 and 13.

Figs. 10 and 12 represent two consecutive sections across
the tip of a brancli. In the first are seen the tubes entering
the wall, while the next (fig. 12) strikes the lumen. A small
})art of this figure is shown slightly diagrammatised (fig. 11)
on a larger scale. Again three consecutive sections are
di'awn in figs. 15, IG, and 17. Here two sections cut through
the solid wall before the lumen is reached. Lastly, fig. 18
represents a section through two adjacent processes, one of
which has been cut so as to expose the lumen, while the other
shows very clearly the soleuocyte tubes piercing the wall and
embedded in its cytoplasm.

The evidence of all these sections is quite unequivocal ; it
would serve no good purpose to multiply instances ; there is
no opening, the wall is continuous, and is traversed by the
tubes of the solenocytes.

But there is other evidence of a different nature leading to
the same conclusion. I have observed in a living nephridium
the fluid inside the nepliridial canal so compressed, perhaps
by the overlying cover-glass, that it dilated the tip of the
diverticulum so as to give rise to a bulging vesicle at its
extremity. Now, such a swelling could obviously not be
formed if the tip were open.

We may now turn to injections to corroborate our view. 1
have recently injected the dorsal hypei’branchial coelom with
Indian ink. The minute black particles were held in sus-
pension in sea-water. Such a fluid, if introduced with a
hypodermic syringe, can be made to fill the coelom. It is
clear that if the nephridium communicated with the coelom
the ink would jienetrate into the canal ; this would happen

STRUCTUEE OF THE EXCRETORY ORGANS OE AMPHIOXUS. 191

all the more easily, since a powerful ciliary current works
towards the external pore. Sections of such injected speci-
mens show conclusively that not a single particle of ink has
entered the nephridial canal, although the ink has penetrated
into every chink of the coelom.

But, it may be asked, if the facts are so plain and conclu-
sive, hoAV is it that so keen-sighted and accm-ate an observer
as Boveri has been deceived? Well, if it will not be con-
sidered presumptuous on my part, I will attempt to explain
how the mistake arose. ^ To begin with, the sections he
examined were not appropriate!}'’ stained. The nuclei are
clear, but the cytoplasm scarcely stained at all. In the
majority of the sections which I had the opportunity of
seeing the wall which closes the tips of the diverticula was
very difficult to make out, though I could detect it on close
e.xamination in a suitable light. I naturally turned with
great interest to the section given on PI. 33, fig. 17, of the
original memoir (1), and of which a ])hotograph is published
in the ' Anatomischen Anzeiger’ (la). Anyone on first
looking at this section might be led to believe in the exist-
ence of a funnel. The appearance is extraordinarily decep-
tive. But it is deceptive, and the deception is due to two
things. First of all the nuclei are deeply stained, but the
cytoplasm practically colourless and transparent; in the
second place the section is thick. The figure given by Boveri
is really an optical section of the preparation. The closing
wall can, indeed, be seen, but only with the greatest diffi-
culty. The misleading appearance of a funnel is due to the
sudden cessation of the nuclei round the base of the soleno-
cyte tubes; an appearance which is further heightened by
the limit of the coclomic epithelium at the same spot (see
p. 193, and text-figure).

Jjet it not be thought that in insisting on the absence of an
opening I am unduly influenced by a priori considerations

* Soon after the publieatiou of liis paper (la) I wrote to Professor
Boveri, who then very kindly sent me his preparations, and I gladly
take this opportunity of thanking him for his courtesy.

192

EDWIX 8. (JUODKICH.

clue to theoretical bias. It is true that I hold that the renal
organ of Aiuphioxus is a nephridiuin homologous with the
nephridia of Annelids and Platyhelmiuths, and not homo-
logous with the kidney tubules of the Craniata (5, 7) ; but it
is now well known that the true nephridia of Annelids may
open into the coelom. There is no a priori reason why
they should not do so in Amphioxus. However, no nephri-
dium has yet been found possessing both solenocytes and an
internal opening, though such intermediate stages must pre-
sumably have existed.

The Relation of the Nephridium to the Blood-
supply. — The general blood-supply has been well described
and figured by Boveri (1). But according to my observa-
tions the vessels occur not so much as narrow capillaries, as
in the form of a large expanded vessel spreading over the
area occupied by the excretory organ. This is shown in
sections (figs. 7, 23), and also in the reconstructions given on
Plate 11. It will, moreover, be noticed that, although the
greater part of the bloodvessel lies on the inner or atrial
surface of tin; nephridium, yet several loops pass round to
the outer or coelomic surface. Thus a considerable part of
the nephridial canal is entirely surrounded by the blood-
vessels. The solenocytes radiate out from the canal, and
always lie on the wall of a bloodvessel, being attached to it
by a })rotoplasmic process (tigs. 4, 15). 'I’he way in which
these cells are distributed is shown iu figs. 14, 19, and
diagrams 2 and 3, and the text-figure. It will there be seen
that the longer tubes, which are of course those belonging to
cells furthest away from the canal, pass over the shorter
tubes to reach their destination. Never do the solenocytes
project freely into the ccclom ; when they appear to do so in
sections this is, I believe, due to the cell having become
detached accidentally, either during the process of preserva-
tion or of cutting. The tubes are therefore fixed at both
cuds.

In the text-figure may also be seen the peculiar disposition
of the solenocytes at the top of the secondary gill-bar. Here

STRt'CTURE OF THE EXCRETORY OR(T\XS OF AMFHIOXUS. 193

tlie canal of tlie nephridium gives off two or three short
diverticula, which are turned away from the coelom towards
the middle line. The numerous solenocytes projecting from
these diverticula lie in a sort of pocket or chamber (figs. 1, 2,
3, 23), which only communicates with the coelom by means of
a dorsal opening, over which pass a large number of soleno-
cyte tubes. In one region the inner wall of this chamber is
fortued by the skeletal rod of the gill-bar (figs. 3, 23). Some-
what similar pockets are found occasionally in connection
with other parts of the nephridium, as, for instance, the
anterior limb of the canal. The cavity in the chambers is,
I believe, rather of the nature of a lymph space than of a
true ccclomic cavity.

The Relation of the Nejihridium to the Ccclomic
Epithelium. — It is important to determine exactly what is
the disposition of the coolomic epithelium in the neighbour-
hood of the nephridium. Boveri (1) and Weiss (13) have
already shown that the canal is covered by the ccclomic
epithelium ; but this epithelium only clothes the outer or
ccclomic surface (text-figure). It passes on to the nephri-
dium from the atrial wall, covering the canal and its blind
branches to their extremity. Here it is not reflected so as to
pass over to the inner or atrial surface of the organ, but ends
abruptly near the base of the solenocyte tubes (figs. 2, G,
8, 9).

Thus the nephridium and the bloodvessels which accom-
pany it maybe said to lie ‘‘morphologically” eiitirely outside
the coelom; between the ccclomic epithelium and the atrial
epithelium. The nephridium is, in fact, retroperitoneal.
'I’liis is true, I believe, of the solenocytes themselves, though
less easy to prove. For the ccclomic epithelium stops short
where the solenocytes begin (figs. 6, 8, 2), passing neither on
the inner side over the bloodvessel, nor outside them over
their ccclomic surface. For a long time I was under the
impression that a very delicate membranous extension of the
epithelium covered over the coolomic surface of the soleno-
cytes; but I am now satisfied that this is not the case,

194

EDWIN S. OnODRICH.

altliougli sometimes the epithelium seems to stretch over the
base of the solenocyte tubes for a considerable wny. The
coelomic epithelium is not continuous with the wall of the
canal at the tip of the diverticula, but often can be seen in
sections to end with a free and jagged edge. Over the region
where the solenocytes occur there is a gap in the coelomic
epithelium, so that coelomic fluid freely bathes the solenocyte
tubes (figs. 2, 3). That the space in which lie these tubes,
and even the deep pockets described above (p. 193), commu-
nicate with the coelom is evident in specimens injected with
Indian ink.

Since no epithelium covers the solenocytes their true rela-
tion to the coelom cannot be made out for certain in the
adult. Without going into the question of their develop-
ment in this paper, I may say that a careful examination of
M. Legros’s excellent preparations has convinced me that in
the very earliest stages of its development the whole rudi-
ment of the nephridium and solenocytes lies enclosed betw'een
the coelomic epithelium and the atrial wall. There is nothing
unusual in the solenocytes coming into secondary contact
with the coelomic fluid. We know that in the Actinotrocha
larva the nephridium pierces the wall of the preseptal
hmmocoel, and the solenocytes project freely in the blood
(8). In many Polychmtes also the nephridium passes tlu’ough
the coelomic epithelium and the solenocytes lie naked in the
coelomic fluid (6) .

To sum up the chief points in this contribution : — The
careful examination of the nephridia in sections and in the
living state shows that they have no internal opening. The
tubes of the solenocytes pierce the wall of the nephridia!
canal, and open into its lumen. The flagellum passes down
the tube into the lumen. The solenocytes are attached to
the wall of the bloodvessels, which expand in this region, and
may surround the canal. Both the bloodvessels and the
nephridial canal are covered by the coelomic epithelium,
being situated between it and the atrial wall. Over that
region which is occupied by the solenocytes there is a gap in

STRUCTIIKE OF THK EXCRETORY OR(4AN.S OF A.MPHIOXrS. 195

the cocloinic epithelium, allowing the fluid to bathe the tubes.
Nevertheless the whole excretory organ is to be considered
as retroperitoneal.

Part 3. — The Nephridium of Hatschek.

'I’his organ was first described by Hatschek in a paper
without illustrations (9), wherein he states that the first
somite of the left side divides into two halves, of which the
first acquires an opening to the exterior, and becomes the
“ Riiderorgan” or ciliated glandular joit in the buccal cavity,
while the second and inner half becomes the nephridium.
“ Es entwickelt sich in der Larve als rnesodermaler wim-
pernder Trichter und canal, und zwar nnr linkerzeits vor der
Munddffnung, in der Region des ersten Metamers ; es wiichst
spilter weiter nach hinten aus. Bei dem ansgebildeten Thiere
erstreckt sich das Organ an der linken kdrperseite lungs des
ventralen Randes der Chorda von nahe dem vorderen Mun-
drande bis dicht hinter das Velum. Hier scheiut es in den
Kiemendarm zu rniinden (die Ausmiindung muss ich noch-
mals priifen). Es liegt in einem engen Fortsatz der Leibe.s-
holile iiberlagert von der linken Carotis ” (9).

Lankester and Willey incidentally refer to the organ in
their important memoir on the larva of Amphioxus (10), and
there state that “ in the condition in which we have observed
this structure (viz. in larvic ranging from the stage with
three gill-slits up to closure of the atrial cavity) there does
not seem to be any special reason for regarding it as a
nephridium. We should prefer to call it the subchordal
tube. It appears to end blindly anteriorly, and to open into
the buccal cavity near the recurved extremity of the gland-
ular tract which accompanies the club-shaped gland.”

Willey in his later contribution (15) incompletely repre-
sents the nephridium of Hatschek in his figures, but actually
draws the solenocytes without, hoAvever, realising their
significance. He says, “1 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

19G

EDWIN S. (iOODRICll.

of Annelid larva) (ti'ochospliores), but I can form no opinion
as to the reality of any such resemblance.”

The next author to mention the organ is MacBride, who
briefly describes its development, believing that it arises
from the communication between the gut and the second
myotome (11).^

Van Wijhe (14) describes the canal of Hatschek’s nephri-
dium in the adult, applying to it the name Schlundforsatz :
^^eine enge Rohre, welche dem linken Seiteurande der linken
Aorta angeschmigt ist. Das enge lumen wird von einem
einschichtigen Cylinderepithel begrenzt und bildet strecken-
weise seitliche Ausbuchtungen. Wo eine solche ange-
schnitten wird, kbnnen zwei Lumina im Schnittbilde auf-
treten. Unmittelbar hinter dem Velum miindet die Ebhre
mit einer feinen Offnung in den Schlund aus.” He denies,
however, the presence of the coelomic cavity described by
Hatschek, and does not accept the latter’s theory as to the
organ’s function. “ Nach meiner Meinung,” says van Wijhe,
“ ist das organ nicht anderes als ein Rudiment des vorderen
Darmendes, welches beim Embryo in das Plimmersiickchen
(linke Entodermsackchen) ausmiindete.”

It is to Goldschnidt that we are indebted for the first
description of solenocytes in the nephridium of Hatschek (4),
placing its homology with the posterior nephridia beyond
doubt. His account seems, however, to be based on imper-
fect material, and he falls into the error of ascribing to the
canal an internal opening such as Boveri had described in
the paired nephridia.

I have recently had the opportunity of studying this
interesting organ in adult and larval specimens in Helgo-
land,^ and am thus able to give a more complete description
of it.

' I am unal)le to agree witli tlie view of either Hatschek or MacBride
as to the origin of this nepliridium.

- I gladly seize this opportunity of thanking Prof. Heincke, Prof.
Hartlauh, and the staff of the Kiinigl. Biologische Anstalt for the kind
way in which they received me in Helgoland.

STIUHITURE OK THE EXCHETORY OROANS OF AMI’HIOXUS. 197

The nephi’idium of Hatschek reaches its maximum develop-
ment in the adult, where it is indeed the largest nephridium
in the body, some 2 mm. in length. Lying on the left side,
below and parallel to the notochord, it opens just behind the
velum into the pharynx,^ and runs forward a long distance
to a point just in front of the ciliated groove (Riiderorgan).
Here it ends blindly, and along its course are given off short
blind diverticula (figs. 27, 42, 43, 44). Solenocytes are set
on the dorsal and lateral surfaces of the organ along almost
its whole length, being especially numerous on the diverti-
cula (fig. 28). Altogether an enormous number of soleno-
cytes are present on this nephridium in the adult Am-
phioxus.

The canal runs along the floor of a narrow cavity beside
the aorta (figs. 42 — 44). It is to the wall of this cavity that
the solenocytes are attached, and it appears to be of coclomic
nature; at all events it is in open communication with the
myococle of the first myotome in larval stages (figs. 25, 20,
33). In the adult, however, it is closed off, and the lining
epithelium seems to be very irregularly developed, forming
Jio distinct layer of cells (fig. 28).

In the larva of about 13 gill-.slits, of the left series only
(fig. 33), the nephridium cau be well seen by transparency
as a short tube opening behind into the phai’ynx (fig. 24).
Its dorsal surface is entirely beset with solenocytes in several
closely packed rows (fig. 29). An optical section of the
organ at this stage is represented in fig. 38, showing clearly
the way in which the tubes of the solenocytes pierce the thin
dorsal wall.

We may summarise as follows the observations recorded
above : — The nephridium of Hatschek is a true nephridium,
similar in structure to the posterior paired nephridia. In
the adult, where it reaches its maximum development, it
extends along the left aorta from in front of the ciliated

‘ On one occasion only I have found an opening from the canal into
the hinder region of the huccal cavity itself, as well as the posterior
opening into the pharynx.

19S

EDWIN S. (iOODKICH.

groove backwards to the pharynx into which it opens.
Very numerous solenocytes are set chiefly on short blind
diverticula. It has no internal opening, and lies in a cavity,
Avhich is in communication with the myocoele of the first
myotome in the larva.

That this nephridium is in every way similar to and homo-
logous with the paired posterior nephridia there can be no
doubt. Van Wijhe’s suggestion, mentioned above, must
therefore be abandoned. Two peculiarities, however, still
remain to be explained ; its unpaired character and its open-
ing into the alimentary canal. No one, so far as I am aware,
has yet worked out the exact relation of the gill-slits to the
somites in the larva of Amphioxus, and my own observations
on this point are very incomplete. But judging from the
course of the dorsal spinal nerves (fig. 30), the first gill-slit,
of the left (on the right side) series, which is the first to
appear in the larva, corresponds to the third myotome.
Probably its true morphological position is between the
second and third myotome. Presumably Hatschek’s nephri-
dium would correspond to the next gill-slit in front, between
the second and first myotonies, did such a slit exist. As for
its unpaired character, I can for the present offer no better
explanation than this, that it is the left of an original
anterior pair of nephridia, the one-sided development of
which is no doubt correlated with the general asymmetry of
the anterior region so conspicuous in the larva. But this
question can only be profitably discussed after an exhaustive
study of the development, and must therefore be put aside
for the pi’esent. In the same way a detailed knowledge of
the development of this organ, and of the posterior nephridia,
is necessary before one can discuss the significance of the
anomalous position of the opening.

STEUCTURE OP THE EXCRETORY OWOANS OF AMPHFOXUS. 199

Part 4. — The Development of the Left Series of Nephridia in

the Larva.

For many years I have been trying to trace the develop-
ment of the nephridia in Amphioxus. In 1902 I collected a
large amount of material from the Pantano at Faro ; but ill-
health prevented my working out the development on the
living larva, and I failed to do so on the preserved speci-
mens. It was not till last year that I was again able, in
Helgoland, to study the living larva, and succeeded in
tracing some stages of the development of the excretory
organs. In the meantime Legros had been studying the
same subject in Naples, and published anonymously a pre-
liminary notice of his results a short time ago (16).^

In the present paper I shall not discuss in detail the first
origin of the nephridia, but restrict myself to a description of
the stages found in the larva with from ten to fifteen gill-slits
of the left hand series, and no trace of the right hand series.
These are the only stages which I have been able to study
sufficiently in the living state.

Fig. 30 gives a left side view of a young larva with eleven
slits. The anterior gill-slits are still well on the right side,
but the hinder slits are in or near the middle line. The
future dorsal edge of each slit may, of course, at this stage
be more ventral than the future ventral edge. The nephridia
are seen as small rounded sacs near the posterior ventral
corner of each slit. Every slit from the first to the last has
such a nephridium. At this early stage there is no atrium,
the slits have an internal margin of thick branchial epi-
thelium, which is thrown into characteristic folds when the
branchial muscles contract, while the external margin of the
slit is formed by a thin fold of the body wall, acting as a sort

' Tlirougli the kindness of M. Legros I have had the oi^portunity of
examining Ins sections, and I cannot agree witli liis conclusions as to
the origin of the nephridia from the ccelomic epithelium, nor as to the
presence of inteimal openings. But I believe he has modified his views
considerably on these points since the publication of the note.

200

EDWIN S. OOnDEICH.

of sphincter (fig’s. 33 and 36). A shallow branchial chamber
lined with epidermis is thus formed, leading from the external
to the internal opening. It is in this chamber that the
nephridium opens, at a place corresponding apparently to the
point of junction of the ectoderm with the endoderm (fig. 41).
The position of the nephridiopore can be seen in figs. 36 and
30. When the atrinm becomes formed by the closing off of
tlie space between the metapleural folds, with which the
branchial cavities become merged, the pores open into the
atrium. A ventral view of a stage where the atrium has just
begun to be formed posteriorly shows one or two nephridia
behind the last open gill-slit (fig. 35). Probably these
nephridia belong to the posterior gill-slits, which have closed
up (Willey, 15) ; they open now directly on the surface
(fig. 30).

The young’ nephridium is a flattened sac, without internal
opening (figs. 36 and 39). From its inner end spring a large
number of solenocytes; their tubes pierce its wall, and their
flagella pass into the lumen of the sac. The majority of the
solenocytes spread over the blood-vessel which runs along
the future dorsal edge of the slits. The solenocytes of the
first few slits scarcely extend beyond this limit; but, passing
backwards to more posterior nephridia, we find that the
solenocytes spread farther and farther up towards the dorsal
aorta, the tubes lengthening out as the cells lie farther from
the nephridial sac. At about the fifth or sixth nephridium
some of the solenocytes actually reach the aorta (fig. 40).
fl'he tubes in this case ina,y attain a really astonishing length,
stretching right across the field of a yVth oil-immersion
objective with oc. 8.

Fig. 34 represents the posterior gill region of a living
larva, in which the remarkable development of the .soleno-
cytes is well shown. Here a group of the longest solenocytes,
some twelve to eighteen in number, spread out over the aorta
in a most beautifully regular fan-like arrangement in each
segment. A section of this region is shown in fig. 31 ; the
fan-like disposition is found in each segment to the hindmost

STRUCTURE OF THE EXCRETORY ORGANS OP AMI'HIOXUS. 201

limit of the series of nephridia. Presumably the dorsal
solenocytes degenerate later^ since they are not known to
exist in the adult.

The observations on the larval nephridia recorded in this
part may be summarised as follows : — To every gill-slit
corresponds a uephridinng consisting of a sac closed in-
ternally, but opening to the exterior apparently at the point
where the ectoderm joins the endoderm in the shallow bran-
chial chamber. From the internal blind end of ihe nephridial
sac spring numerous solenocytes, some of which reach and
spread over the aorta at every segment in a fan-like arrange-
ment. This structure is only fully developed from about the
eighth segment backwards to the last nephridium.

July 3rd, 1909.

List of Kffkkfncks.

1. Hoveri, Tli. — “Die Niereiikauiilcheii des Aiiiphioxiis," ’Zool. Jahrl).

Aiiat. Abt.,’ Bd. 5, 1892.

la. “ Bemerk. iiber d. Ban. d. Niereiikaiialcheii des Ampliioxus,"

‘ Aiiat. Aiiz.,’ Bd. 25, 1904.

2. Felix, W. — “Entw. des Hainapparates," ‘ Hertwig’s Handbucdi d.

Eiitw. d. Wirl^eltiere,’ Bd. 3, 1904.

3. “ Entw. des Exkretions-system,” ‘Aiiat. Hefte. Ergeb. Aiiat.

11. Entw.,’ Bd. 13, 1904.

4. Goldschmidt, R.— “Ampliiuxides,” ‘ Wiss. Ergel). d. deutscli. Tiefsee-

Expedition “Valdivia,”’ Bd. 12, 1905.

5. Goodrich, E. S.— “On the Cadom, etc,” ‘Quart. Journ. Micr. Sci.,’

vol. 37, 1894.

6 “ On the Nephridia of the Polycha-ta,” Parts I, II, and III,

ibid., vols. 40, 1897, 41, 1898, and 43, 1900.

7. “ On the Structure of the Excretory Organs of Ainphioxus,”

Part I, ibid., vol. 45, 1902.

8. “ On the Body-cavities and Nephridia of the Actinotrocha

larva,” ibid., vol. 47, 1903.

9. Hatschek, B. — “ Mitth. iiber Ainphioxus,” ‘ Zool. Anz.,’ Bd. 7, 1884.

202

EDWIN S. DOODEICII.

10. Laiikcster, E. R., and Willey, A.— “ The Development of the Atrial

Chamber of Amphioxus,” ‘ Quart. Journ. Micr. Sci.,’ vol. 31, 1890.

11. MaeBride, E. W. — “The Early Development of Amphioxus," ibid.,

vol. 40, 1898.

12. Schneider, K. C. — ‘Lehrhuch des Vergl. Histologie,' Jena, 1902.

13. Weiss, F. E. — “Excretory Tubules in Amphioxus,” ‘Quart. Journ.

Micr. Sci.,’ vol. 31, 1890.

14. Wljhe, J. W. van. — “ Beitr. z. Anat. des Kopfregion des Am-

phioxus,” ‘Petrus Camper,’ vol. 1, 1901.

15. Willey, A. — “ The Later Larval Development of Amphioxus,”

“Quart. Journ. Micr. Sci.,’ vol. 32, 1891.

16. Legros, R. — “ Sur le dcvel. des fentes hranchiales et des canalicules

de Weiss-Boveri cliez I’Amphioxus,” ‘Anat. Anz.,’ Bd. 34, 1909.
Published anonymously.

List of Refkkence Letters.

a. c. and «. ep. Atrial epithelium, ao. Aorta, h. Secondary gill-har.
b. c. Buccal cavity, b. ep. Epithelium of buccal cavity, hr. Brain.
br. c. Branchial eiiithelium. bv. Blood-vessel. cii. Cavity in whicli
runs Hatschek's nephridium. c. ep. Coelomic epithelium, c. gl. Club-
shaped gland. ch. Solenocyte chamber. cr. Cirrhus. c. w. Cut
wall of nephridial canal, d. n. Dorsal nerve, e. <j. s. Edge of gill-
slit. e. m. Margin of mouth. end. Endostyle. c. op. External oijen-
ing of club-shaped gland, cp. Ejjidermis. fl. Flagellum, g. Glandular
area. g. m. Branchial muscle, g. s. i— Gill-slits. gt. Wall of
gut. h. c. Pre-oral head-cavity. H. ncph. Hatschek’s nephridium.
ibw. Inner blood-vessel, i. op. Internal opening of club-shaped gland.

I. Lumen of nephridial canal. 1. p. Lower margin of mouth, 'lucl.
Myocojle. mt. i— -. First and second myotome. vit.f. Metapleural
fold. my. Myotome, n. Wall of neidiridial canal, negdi. Nephridium.

II. c. Nerve cord. ne. Nerve from left side. id. Notochord, oji.
External opening of nephridium. pi). Peripharyngeal ciliated tract.
p. III. Pre-oral muscle, p. o. m. Anterior oral muscle, p. t. m. Pos-
terior oral muscle, sol. Solenocyte. s. s. Pre-oral sense-organ, t. and tb-
Tul)e of solenocyte. up. Upper margin of mouth, v. Velum, v. b. v.
Ventral l)lood- vessel now on right side. vt. Velar tentacle.

STEUCTUEE OF THE EXCUETOEY OEUANS OF AMPHIOXUS. 203

EXPLANATION OF PLATES 11—16,

Illustrating Mr. Edwin S. Goodrich’s paper “ On tlie Struc-
ture of the Excretory Organs of Ainphioxiis.”

PLATE 11.

Fig. 1. — Diagnunniatic reconstruction of a left nephridiuui and the
neighbouring lilood-vessels, from a series of sections taken parallel to
the gill bar. The solenocytes are not represented. Side view from the
outside. Op. indicates the position of the nephridiopore on the opposite
side.

Fig. 2. — Similar reconstruction of a right nephridiuui, fiom a series
of sections transverse to the gill-bar. In two places the coilomic
epithelium and the solenocytes are shown.

Fig. 3. — Reconstruction of a portion of a left nephridium, the blood-
vessels, and the toji of the secondary gill-l>ar, seen from behind.

Fig. 4. — Small portion of a section shaving off the wall of a
nephridial canal, and showing the bases of the solenocyte tubes
embedded in the cytoplasm. Cam. Z. 2 mm. ap. oil imm., oc. 8.

PLATE 12.

Figs. 5 and 6. — Two consecutive sections, parallel to the gill-bar,
through a nephildium, showing the solenocyte tubes xiassing through
the thickness of the wall of the canal. Cam. Z. 2 mm. oc. 4.

Fig. 7. — Drawing of the lower surface of the section of which the
upper surface is represented in fig. (1. Cam. Z 2 mm. ap oil-imm.,
oc. 4.

Fig. 8. — Section across the anterior limb of a nephridium, showing
the ccelomic epithelium passing over the outer surface of the canal.

Fig. 9. — Similar section showing solenocyte tubes piercing the wall
of the canal. Cam. Z. 2 mm. ap. oil-imm., oc. 4.

Fig. 10. — Section parallel to a gill-bar, cutting the wall of a diver-
ticulum of the neiihridial canal (at th.). Cam. Z. 2 mm. ap. oil-imm.,
oc. 4.

Fig. 11. — Diagi'amniatic view of a small portion of the wall of the
diverticulum in the same section, showing the bases of the solenocyte
tubes.

204

EDAVIN S. (lOODRICII.

Fig. 12. — Next section to that drawn in fig 10.

Figs. 13 and 11. — Two consecutive sections through the ventral end
of the anterior limb of a nephridial canal. Cam. L. jb oil-imni., oc. 3.

PLATE 13.

Figs. 15, 1(>, and 17. — Three consecutive sections, parallel to a gill-
bar. through the extremity of a diverticulum of the nephridial canal.
Cam. Z. 2 mm. ap. oil-imm., oc. 12. In figs 15 and 10 the solenocyte
tubes are cut in the thickness of the wall of the canal.

Fig. 18. — Section across the ends of two adjacent nephridial diver-
ticula. The bases of solenocyte tubes are clearly seen embedded in the
cytoplasmic wall. Cam. Z. 2 mm. ap, oil-imm., oc. 18.

Fig. 19. — Longitudinal section cutting the surface of a nephridium.
Cam. L. oil-imm., oc. 3.

Fig. 20. — View of the portion of the next section corresponding to
the left-hand region of fig. 19.

Fig. 21.— Similar section of another nephridium.

Fig. 22. — Diagram to illustrate the direction of the sections drawn in
figs. 5, 9, 10, 13, 15, and 19.

PLATE 14.

Fig. 23. — Section across the toj) of one primary and two secondary
gill-bars, showing the position of the solenocyte chambers (ch.), and of
the blood-vessels. The position of the external pore at a lower level is
indicated by a cross X.

Fig. 24. —Transverse section of a larva, passing through the mouth,
and oiiening of Hatschek's nephridium. Cam. Z. D., oc. 3.

Fig. 25. — Transverse section farther forward passing just beyond the
anterior end of Hatschek's nephridium, where the cavity in which it lies
opens into the first myocade.

Fig. 26. — More enlarged view of a portion of the next section,
sliowing the solenocyte tubes in a cavity continuous with the first
myocccle.

Fig. 27. — Anterior end of an adult Amphioxus, ventral view. The
buccal cavity has been opened up by cutting along the mid-ventral line
H;itschek's nephridium is seen on the left side of the notochord.

STRUCTURE OP THE EXCRETORY ORCxANS OF AMPHIOXUS. 205

Fxg. 28. — Small poi-tion of a transverse section of the head, showing
Hatschek's nephridium. Cam. L. oil-imm., oc. 3.

Fig. 29. — Similar view of a larva (the same as that in fig. 2t, from
Helgoland, with about thirteen gill-slits). Cam. L oil-imm., oc. 3.

Fig. 30. — Portion of a longitudinal section of a larva, showing a
nephridium opening behind the last open gill-slit. Cam. Z. 2 mm. ap.
oil-imm., oc. 4.

Fig. 31. — Portion of a longitudinal section of a larva, showing the
fan-like group of solenocytes on the aorta. Cam. Z. 2 mm. ap. oil-imm.,
oc. 4.

PLATE 15.

Fig. 32. — Left side view of a larva, drawn fi-om living and preserved
specimens.

Fig. 33. — Left side view of the anterior region of a slightly older
larva on a larger scale, from living and preserved specimens. The cilia
are not indicated.

Fig. 34. — Left side view of the posterior branchial region of a larva,
showing the disposition of the solenocytes. From the living.

Fig. ,35. — Ventral view of a region of a larva, showing the last open
gill-slit, and two more jio.sterior nephridia. From the living.

Fig. 36. — Ventral view of two posterior gill-slits of a living laiwa.

Fig. 37 —Solenocytes from Hatschek's nephridium in the larva.

Fig. 38. — Optical section of Hatschek’s nephridium in the larva.
From the living.

Fig. 39. — Ventral view of a nephridium showing its opening just
within the margin of a posterior gill-slit in a larva. Solenocytes cut
short.

PLATE 16.

Fig. 40. — Left side view of a single nephridium in a larva. From
the living.

Fig. 41. — Portion of a transverse section of a laiwa, passing through
the nephridiopore. Cam. Z. 2 mm. ap. oil-imm., oc. 4.

Figs. 42, 43, and 44. — Portions of three transverse sections of the
head of the adult, showing Hatschek’s nephridium. In front of the
ciliated pit (fig. 42), at the level of the ciliated pit (fig. 43), and behind
it (fig. 44).

VOL. 54, PART 2. — NEW SERIES.

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DIGESTIVE PROCESSES IN PLANARF7E.

207

Intra-cellular and General Digestive Processes
in Planariae.

By

O. Arnold,

From the Cytological Laboratory of the University of Liverpool.
With Plate 17.

In 1878 Metsclinikotf drew attention to the plieuoinena
of intra-cellnlar disfestion occnrrino' in Tnrbellarian worms.
Since that time bnt little lias been published dealing with
this very interesting subject.

Metschnikoff’s short notice was followed by a paper by
Lankester dealing with intra-cellnlar digestion in the endo-
derm cells in the medusa of Limnocodinm, and two years
later, 1883, Metschnikolf published further observations on
intra-cellnlar digestion in the mesoderm cells of Synapta and
l^hyllirhoe.

Intra-cellular digestion has been observed in Ccelenterates
generally, sponges. Protozoa, and in the leucocytes of the
blood.

Within recent years several observers have dealt with the
digestion in the Protozoa, but apparently no work has been
published dealing with the cytological details of intra-cellnlar
digestion in any of the Enterocoela.

Mouton in 1902 and Nerinstein in 1905, following on the
earlier work of Greenwood and Saunders, have given long
and detailed accounts of the process of digestion in Amoeba,
Paramoecium, etc. These authors, however, have limited
their attention almost entirely to the intimate history and
staining reactions of the food vacuoles of those animals, and
their conclusions afford few data which shed any light on

208

0. ARNOLD.

the digestion in more highly organised animals such as
Planaria. Moreover, the methods of research are necessarily
different. In unicellular animals a considerable number of
facts may be ascertained by the observation of the effects of
intra-vitam staining. In animals such as the Planaria this
is impossible on account of their lai’ge size and opacity. The
observations here described have therefore been made upon
carefully preserved specimens, and the staining reactions ai-e
therefore post-mortem.

The methods used were as follows :

A number of Planaria lactea, which had been deprived
of all food of any sort for fifteen days (after which period of
time the cells of the intestine are entirely devoid of all food
remains, see fig. 11) were fed with fresh clotted pig’s blood,
and fixed in Flemming’s strong solution at various intervals
after feeding.

These intervals after feeding were as follows: j, i, IJ, 3i,
27, 48, 52, 70, 76, 96, 118. When a Planarian has just fed,
the fixation is attended with difficulty owing to the fact that
immediately the animal is immersed in the fixing fluid it
contracts and ejects the recently ingested food Avith consider-
able violence, not through the pharynx, but anyAvhere through
the skin. If, however, the animal is cut into several pieces
at the same time that the fixative is poured upon it, this
difficulty is partially obviated, the whole procedure being too
rapid to permit of any violent contraction. Forty-eight hours
after feeding the lumen of the intestine is almost empty, most
of the blood having been ingested, and the Planaria fixed
after that interval did not eject any of the remaining contents.

The stains used were: (1) A triple stain — Basic fuchsin,
methylene blue and orange G.,^ and (2) iron-alum-hmma-
toxylin, acid fnchsin and orange G.

All the figures, except fig. 5, are drawn from preparations
stained by the former process.

' I have given an account of tlie inethocl of using this stain in a
paper on the “Ovi- ami Spermatogenesis of Planaria lactea," ‘ Arch,
f. Zellforschung,’ Bel. iii, Heft 3, 1909.

DIGESTIVE PROCESSES IN PLANARI^.

209

Some cells from the intestine of a planarian which has been
without food for some seven or eight days are shown in
PI. 17, fig. 13.

The cells of the intestine are of two sorts :

(1) Long, irregularly columnar cells. The cytoplasm of
the cell (fig. 13) consists of a clear protoplasmic network^
enclosing several large vacuoles at its distal end, the vacuoles
towards the middle of the cell being smaller and fewer. The
proximal part of the cell consists of very much denser cyto-
plasm, in which the reticulum is very fine and close, showing
an almost fibrillar structure at its extreme end. The spaces
between the network take the acid stain, but the network
itself is stained by the basic colours, so that the proximal end
of the cell where the reticulum is very dense is much darker
than the rest.

In this part lies the nucleus, which is small in proportion
to the cytoplasm. The nucleus is round or ovoid, with a
deeply staining membrane, and a nucleolus which is stained
bright blue by the methylene blue.

In an animal which has been starved for fifteen days the
vacuoles in the cytoplasm are more numerous and larger
(fig. 11). The cytoplasm around each vacuole is denser and
more granular than elsewhere, but a definite membi’ane
cannot be made out.

(2) (xoblet-shaped gland-cells, only half as large, or less,
than the former, invariably with a small nucleus extremely
irregular in outline, and taking the basic fuchsin stain very
markedly. The cytoplasm is very granular, and is peculiar
in having a greater affinity for basic than acid stains,
staining as deeply as the nuclear material. It is full of large
vacuoles, in which now and again is to be seen a residue also
stained by the methylene blue (tigs. 11 and 13.)

Absoki'tion of Pat.

We will deal with the functions and history of these gland
cells first. There is generally one of them to every ten of the

210

(1. ARNOLD.

others. When the intestine is empty they are large and the
vacuoles are full (fig. 11). Very soon after food has been
taken into tlie intestine the whole cell diminishes in size, till
at about the twenty-seven hour stage it is shrunken to a
fifth of its original size and quite flaccid (fig. 16). In this
condition it lies squeezed in between the columnar cells, so
much so that sometimes these cells appear to lie quite outside
the intestine, between the latter and the sui-rounding
parenchyma.

There can be little doubt that the gland cells secrete a
digestive ferment, which is probably used entirely for the
digestion of fat.

AVithin a quarter of an hour after feeding it will be seen
that the columnar cells are full of fat-globules, stained black
by the osmic acid of the fixative (fig. 1). Even when the
lumen of the intestine is full of blood (red corpuscles, leuco-
cytes, etc.) no fat-globules are to be seen lying free in the
lumen, nor can any pseudopodial extensions of the cytoplasm
containing fat-globules of the columnar cells be seen, suggest-
ing that the fat has been ingested in an amoeboid fashion.

The gland-cells do not begin to return to their normal size
till after about the forty-eighth hour, when almost all the
columnar cells are devoid of unaltered fat, and reach their usual
size again at about the seventieth hour. It is very noticeable
that no ingestion of solid particles (i.e. true intra-cellular
digestion) takes jtlace until the absorption of fat is over, and
the latter has undergone marked changes in the columnar
cells. A large part of the fat absorbed by the columnar cells
is digested in the cytoplasm of these cells, but some of it
is again passed out at their bases unaltered lying in the
parenchyma. The fate of these extruded globules will be
dealt with later ou.

The fat first appears in the cytoplasm of the columnar cells
in very small globules, which by fusing together form much
larger ones, so that some cells within an hour after feeding
seem to be one mass of fat.

The researches of Munk, Aloore and Rockwood, and others

DIGESTIVE PEOCESSES IN PLANAEIiE.

211

have shown that in the higher animals, especially in mammals,
the absorption of fat by the epithelial cells of the intestine is
brought about by the fat of the food being converted into
fatty acids and glycerine by the action of lipolytic enzymes.
Only in that form can the fat be taken up by the epithelial
cells, which then again synthesise the fatty acids into fat,
and the latter is seen in the cytoplasm of the cells in the form
of globules, being passed on by them to the lymphatic cells
and the lymphatic capillaries.

The process appears to be very similar in the Planarim,
and judging by the facts stated above, there is reason to
believe that the goblet-cells of the Planarim function as
organs secreting a lipolytic enzyme. Possibly they may
elaborate other secretions as well, but their ability to secrete
a fiit-digesting fluid can hardly be doubted.

It has been pointed out that when the columnar cells are
full of fat-globules stained by the osniic acid, no such fat is
to be seen in the lutnen. It was therefore necessary to see
whether there was any fat in the lumen in a form not acted
on by osmic acid. It is well known that the staining with
osmic acid is due to the presence of unsaturated compounds.
In view of the work of Lorrain Smith (’07) it was thought
desirable to test the action of Nile-blue sulphate, which
stains not only the neutral fat, but differentiates the fatty
acids. For this purpose some Planaria were fixed a quarter
of an hour after feeding in a weak solution of foruiol and
cut with a freezing microtome. By this means all fat solvents,
such as xylol, etc., were avoided. The sections were then
stained for fifteen minutes in a strong aqueous solution of
the dye. In spite of the fact that the colour was slightly
masked by the blue colour taken by all the tissue, charac-
teristic globules of fat in the columnar cells were seen, red to
reddish-yellow in colour. Care has to be taken not to
confuse loose red blood-corpuscles which have been shifted
from the lumen on to the cells with these globules. The
colour is, however, entirely different, the fat-globules being
definitely red under a high power lens, whereas the corpuscles

212

G. AUNOLD.

are yellow. Apart from this, fat-globules can be seen in the
cytoplasm of the endotlerm cells, far too large to be mistaken
for any corpuscle lying over or under one of those cells. It
must be remembered that no ingestion of the corpuscles takes
place until some considerable time after the one hour stage,
at which these Planaria were killed.

This fact indicates, at least, that the fat which appears in
the endoderm cells is a neutral fat, but whether the secretion
of the gland cells breaks down the fat of the blood into fatty
acids could not be ascertained, for the colour of the blood-
corpuscles completely masks any blueness which might be
present in the food magma.

However, sections of the one and a half hour stage, cut
in paraffin and stained with Nile-blue sulphate, showed a
definite bluish tinge in the magma, but not a trace of red.

The significance of this fact is important, for it shows that
the digestive process in Planaria is not, as has been stated by
Metschnikoff (‘ L’lmmunite,’ 1902), entirely intra-cellular, and
at the same time indicates the first step in the formation of
the highly complex digestive apparatus found in the higher
animals.

This first step is, we have seen, the production of a secre-
tion by certain cells which enables fat to be absorbed. Such
cells are unicellular glands. If during the course of evolu-
tion these unicellular glands, instead of being difl^used
throughout the intestine, become aggregated in certain areas,
we are enabled to picture the formation of any of the multi-
cellular glands which line the intestinal tract by the
subsequent invagination and enlargement.

Metschnikoff (’02) and in his work ‘ L’lmmunite dans les
(Maladies Infecteuses,’ says that in Planaria digestion is
entirely intra-cellular, and this seems hitherto to have been
widely accepted.

Mesnil (’01) comes to the same conclusion in regard to the
Actinia, but this disagi-ees with the results of several other
workers.

Pratt’s (’05) observations on the digestive organs of the

DIGESTIVE PROCESSES IN PLANAKI^.

213

Alcyonaria, lead her to conclude that large food bodies are
rapidly broken up into small particles, and in some cases
apparently acted on by some digestive ferment in the coelen-
teron of the zooids before being ingested by the cells of the
ventral mesenterial filaments, and that “ we have evidence
in the Alcyonarige as in the Madreporaria of an intercellular
digestion by the secretion of a digestive fluid in the coelenteron
of the zooids, as well as an intra-cellular digestion which
occurs throughout the coelenterates.”

Jordan (’07) has come to similar conclusions on the diges-
tion in Actinia, and says that in them digestion is both inter-
and intra-cellular. He put little paper bags containing fibrin
in the gastric cavity of some Actinia, and found that the
contents were digested although the bags remained intact.
His results are in agreement with those of Krukenberg.

Even in Hydra, according to Hadzi (’00), an appreciable
amount of extra-cellular digestion takes place, the food being
slightly predigested in the lumen before being ingested by
the pseudopodia of theendoderm cells.

We need not be surprised then tliat in the more highly
organised Planarian, digestion is not entirely intra-cellular.

The alteration which the fat-globules undergo in the
columnar cells is characterised by very marked alterations in
their staining reaction. At first they are deep black owing
to the action of the osmic acid in the fixing fluid in which the
animals were preserved (figs. 1 and 2).

Each globule is enclosed in a vacuole. Within half an
hour after feeding, some of the globules at the free end of
the cell become paler, changing from black to grey, and then
brown. Within two hours after feeding (figs. 2, 4, and 5) the
change had proceeded a great deal further. The black
reaction to osmic acid is no longer present, and the fat takes
the less basic of the two basic stains, the fuchsin, till eventu-
ally it is only stained by the acid cytoplasmic stain, the
orange G. (figs. 4, 0, and 7). A vacuole is no longer visible,
and eventually the fat-globules are incoi’porated in the sub-
stance of the cytoplasm.

214

(1. AKX(3J,D.

Intka-cellular T)[GEST10N.

After all the fat hits been absorbed, and when all the
gland cells are empty (fig. 10), true intra-cell ular digestion
(phagocytosis) commences.

The columnar cells push out at their free ends long pseudo-
podial extensions into the lumen of the intestine, and shortly
afterwards large vacuoles appear in which masses of red
blood-corpuscles are seen (fig. 3).

At this stage, one and a quarter hours after feeding, the
selective action of the columnar cells is very noticeable, for
only the red-corpuscles are ingested, but none of the leuco-
cytes. The latter are ingested last of all, some forty-eight
hours after feeding (fig. 6).

The digestion of the red corpuscles takes place very slowly.
Kven ninety-six hours after feeding (fig. 8) ihey may be seen
intact in some vacuoles.

As digestion proceeds, the corpuscles lose their shape (fig.
b, h.e.), till at last the vacuoles contain an amorphous mass of
particles, consisting chiefly of the envelopes of the corpuscles,
which are stained by the methylene blue.

The leucocytes are ingested singly, and a vacuolar space
soon appears round them.

The leucocytes which lie in the lumen of the intestine do
not appear to undergo any change at all. Even after forty-
eight or fifty-two hours they can be seen scattered about in
the lumen, their cytoplasm stained orange by the acid stain,
and the nuclear inembi-ane and the chromatin in the nucleus
quite intact. But immediately a vacuole has formed round
them after ingestion (figs. 6 and 7, L.) their staining reaction
changes. The cytoplasm then takes a pink colour, due to
the basic fuchsin, their nuclear contents become diffused,
and shortly afterwards the separate chromatin masses are no
longer distinguishable (fig. 9).

This marked and rapid change in the staining reaction is
undoubtedly due to the fluid in the vacuole secreted by the
surrounding cytoplasm.

DIGESTIVE PROCESSES IN PLANAlllJi.

215

It is now a generally recognised fact that intra-cellular
digestion in Protozoa is accompanied by a secretion of acid
in the vacuoles,, but witli regard to the part played by this
acid in the process of digestion there is a large difference of
opinion.

(rreenwood and Saunders (’94) sliow that proteolysis com-
mences when the acid reaction is over, and is replaced by a
neutral reaction. That the vacuole fluid also contains a
proteolytic enzyme there can be no donbt.

IMouton (’02) succeeded in extracting from cultui-es of
AmoebEe a diastase, chiefly of ei proteolytic action and
approaching trypsin in its nature. This diastase he identified
with the fluid in the interior of the digestive vacuoles.

Nirenstein (’05j does not think that the acid in the
vacuoles has anything to do with digestion, and Mouton has
shown by a most carefid series of experiments that the
ama'bo-diastase which he extracted from AnioebEe has a
digestive action in an alkaline, neutral or faintly acid medium.

On the other hand, Metschnikoff (‘ L’lmniunite ’), by feeding
Planaria with blood with which had been mixed some grains
of blue litmus, came to the conclusion that digestion in those
animals takes phice in an sxcid medium.

“ L’etude des planaires nous montre que la nourriture des
ces animaux subit exclusiveinent la digestion intra-cellulaire,
dans un milieu fsiibleinent acide et avec I’aide d’un ferment
soluble. Pile nous fournit deja nne preuve de ce que la
digestion intra-cellulaire typique est un processus chimiqne,
du si I’intervention d’enzymes.”

I have shown in connection with the absorption of fat
that digestion in Planaria is not entirely intra-cellular, but
the sudden change in the staining reaction of the ingested
leucocytes is strong evidence in support of the view that the
intra-celluhir digestion in these animals tsikes place in an
acid medium. The chsinge in the staining reaction of the
cytoplasm of the ingested leucocytes from the normal acid to
the basic stains would seem to indicsite that the ingested
material becomes impregnated by an acid fluid.

216

(!. ARNOLD.

Occasionally, in even the earlier stages, some ingested
bacteria are seen, but they are not numerous. But in the
cells of two Planaria killed 118 hours after feeding they were
extremely numerous (fig. 10, h. and c.), and also in the lumen
of the intestine.

At this stage the intestine is practically empty, except a
few masses of blood-corpuscles and leucocytes, with numerous
bacteria. That they appear in greater numbers only when
the food, or what is left of it, has been in the intestine for a
long period of time would suggest that the remainder of the
free food is undergoing putrefaction. No great importance
is to be attached to this isolated observation, but perhaps we
have here the indication of the formation of a definite
intestinal bacterial flora.

Changes in the Nucleus.

In all the columnar cells of the starved examples the
nucleus contains only one nucleolus (figs. 11 and 13). At
the most active state of digestion (fig’s. 7 and 10) there are
two nucleoli, and sometimes even three. It is a question
whether this multiplication of the nucleoli is to be inter-
preted as an absorption of material from the cytoplasm to the
nucleus, or as an expression of increased activity of the
nucleus during digestion, with the consequent formation of
waste products.

Bxcketoey and I’igment- Gkanules.

In all the columnar cells of the intestine certain granular
masses are seen. They are highly refractive and preserve a
yellow colour independently of the staining (fig. 15). Most
of them are excretory products, but some can not be distin-
guished from the pigment granules which form the greater
part of the eyes of these animals. As digestion proceeds
they increase in number, but always occur in groups, and are
not evenly distributed through the cell.

DIGESTIVE PROCESSES IN PLANAEIJi:.

217

Passage of Fat and Excretory Gteanules into the
Parenchyma.

Some of the fat absorbed by the columnar cells is not
digested but passed out in globules at their bases into the
parenchyma (see fig. 2 ; on the right a fat-globule is being
extruded). These globules are taken up by some amoeboid
wandei’ing cells (fig. 14), and also by the yolk-cells (fig. 12)
and the large parenchyma cells (fig. 17). How these globules
reach the interior of the yolk-cells I have not been able to
ascertain. Any digestive power of an amoeboid nature in the
yolk-cells or even in the parenchyma-cells is extremely
unlikely. Nevertheless it is very striking that after feeding,
the yolk-cells which lie in proximity to the intestine are
crowded with fat-globules, whereas in unfed specimens the
yolk-cells contain scai’cely anything but yolk-globules. After
feeding, fat-globules are numerous at the bases of the
columnar cells, and lying free in the meshwork of the paren-
chyma (fig. 14). The parenchyma-cells also contain numerous
excretory granules, massed together in vacuoles (fig. 17).

It would be expected that in an animal like Planaria devoid
of an anus, the excretory products would be shed into the
intestine to make their way out to the exterior by the
pharynx.

An examination of a very large amount of material, con-
sisting of some hundreds of slides, has afforded no evidence
in support of this view. Not only have I been unable to see
any extrusion of waste matter into the intestine, but a careful
search through numerous sections has failed to show any
trace of extruded excreta in the shape of the cliaracteristic
yellow concretions in the lumen of the intestine. Are they
so soluble that they are all removed when lying free in the
intestine by the process of preparing the material for
sectioning ? If not, it is difficult to explain how they are
removed from the body of the Planarian to the exterior.

I wish to express my thanks to Dr. Roaf, of the Department

218

G. ARNOLD.

of Physiology in tliis University, for valuable advice on
staining for fat with Nile blue sulphate.

Conclusions.

Digestion in Planaria lactea, and probably in all
Triclads, is both inter- a,nd intra-cellular.

The intercellular digestion is limited to fat. Tlie fat is
broken down in the lumen of the intestine by the secretion
of tiie goblet-cells into fatty acids, which are then absorbed
by the columnar cells and synthesised again into neutral
fat.

Most of the fat is digested in the cytoplasm of the columnar
cells, but some of it is extruded into the parenchyma at their
base, and appears in the yolk-cells and in the wandering
cells.

The digestion in the vacuoles takes place in an acid medium,
as evidenced by the change in the staining reaction of
ingested leucocytes.

Bibliography.

'78. Metschnikotf, E. — “ tlbev die Verdiimmgsorgane einiger susswasser-
turbellarien,” ‘ Zool. Anz.’

'81. Lankester, Ray. — “On the Intra-cellular Digestion and Endoderin
Cells of Liinnocodinni,” ‘ Quart. .lourn. Micr. Sci.,’ vol. 21.

'83. Metsclinikoff. E. — “ Untersucli ungen iiber die lutracelluliire
Verdauung bei Wirl.)ellosen Tieren." ‘ Arb. Z. Inst., 'Wien.' Bd. 5.
'8fi. Greenwood, M. — “ On the Digestive Process in some Rhizoiiods."
‘ .Journ. of Physiol.,' vol. vii.

'8(>. Krukenberg. — ‘ Grnndziige einer ‘Vergleichenden Physiologie der
V erdanung.'

'89. Metsclinikoff, E. — " Recherches snr la Digestion Intracellulaire."
‘ Ann. de I'Inst. Pasteur,’ tome iii.

'01. Metsclinikoff, E. — ‘ L'lmmunitc dans les Maladies Infectieuses.’

'01 . Mesnil, M. — “ Digestion chez les Actinies," ‘ Ann. de I’Inst. Pasteur.'
’02. Mouton, H. — “ Recherches sur la Digestion chez les Amibes,'

‘ Ann. de I’Inst. Pasteur.’

DIGESTIVE PROCESSES IN PLANARIA:.

219

'05. Nii’enstein, E. — “Beitra<^e ziir Erniilirungspliysiologie dev Pro-
tisteii,” ‘ Zeits. Allg. Pliysiol.,’ Bd. 5.

'05. Pratt, E. M. — “ The Digestive Organs of the Alcyonaria and their
Relation to the Mesogloeal Cell-plexus,” ‘Quart. Journ. Micr-
Sci.,’ vol. 49.

’00. Hadzi, J. — “Vorversuche zur Biologie von Hydra," ‘ Arch. Entw.
Mechanik.,’ Bd. 2’2.

'07. Jordan, H.— “Die Verdaiu\ng bei den Actinien," ‘ Arch. Gesanimte
Phys.,' Bd. 110.

'07. LoiTain Smith. — “ On the Simultaneous Staining of Neutral Eat
and Fatty Acids by Oxazine Dyes,” ‘ Journ. Phys. and Bact.,’
vol. xii.

EXPLANATION OF PLATE 17,

Illustratiiio- Mr. G. Aruoltl’s paper on “ Intra-cellnlar and
General Digestive I’rocesses in I^lanarise.”

All the hgiu’es, except IJ, are drawn dii’ect, using a 2 mm. oil-
immersion Zeiss and 8 compens. -ocular. Figs. 9 and 15 with 18 compens.-
ocular. Fig. 13 i in. Swift and 0 ocular.

All the figiu’es except 5, which is stained with iron-alum ha;matoxylin-
acid fuchsin and orange G., are stained with the triple stain mentioned
in the paper.

Fig. 1. — A columnar cell from material fi.xed 1

Fig. 2.— .. .. .. A

Fig. 3.- .. .. .. li

Fig. 4.— .. .. .. 3A

Fig. 5.— .. .. .. Ij

Fig. 0. — .. .. . 48

Fig. 7. — ., ., ,. 52

hour aftei
hours

feediiu

Fig. 8.— Portion of a columnar cell fixed 90 hours after feeding,
showing pseudopodial ingestion of a leucocyte.

Fig. 9. — Portion of another cell, same stage as 8 (18 ocular).

Fig. 10. — A columnar cell from material fixed 118 hours after feeding.
Fig. 11. — A columnar cell and a gland-cell from an animal starved
for fifteen days.

220

G. ARNOLD.

Fig. 12. — A yolk-cell containing fat- and yolk-globnles. Yolk
coloured blue.

Fig. 13. — Several columnar cells and one gland-cell from a Planarian
starved for five days. Normal appearance.

Fig. 14. — Mesh work of the parenchyma, showing a free fat-globule
and two amoeboid wandering cells, also containing fat. some of which is
undergoing alteration.

Fig. l.'i. — Excretory and pigment granules (18 compens. -ocular).

Fig. Ifi. — An empty gland-cell lying at the base of two columnar
cells, cf. fig. 11.

Fig. 17. — A parenchyma cell containing excretory granules massed
together in vacuoles.

/. Osmicated fat. nf. Fat very much altered and partially
absorbed. (. Ingested leucocyte. he. Ingested blood-corpuscles.
hac. Bacteria, g. Goblet-shaped gland-cell.

». P. PRESS SC. ET IMP.

EARf;Y ONTOGENETIC PHENOMENA IN MAMMALS. 221

Professor Hubrecht’s Paper on the Early Onto-
genetic Phenomena in Mammals : An Appre-
ciation and a Criticism.

By

Kicliard As^lieton, 31.A.,

Trinity College, Cambridge ; Lecturer on Biology in the Medical
Scliool of Guy’s Hospital, in the University of London.

With 5 Text-figs.

Pkbface.

Professor Hubrecht’s paper in a recent number of the
‘ Quarterly Journal of Microscopical Science ’ brings together
“ tlie I’esults of new investigations and recent reflections with
such as had already been published on earlier occasions.”

Even if “ tlie new investigations and recent reflections”
do not contain a great deal that is new, nevertheless the
whole is an invaluable expression of the Professor’s present
opinion on a subject which he is doing so much to advance
and to cause others to devote attention to. At the same time
it is not possible to ignore the feeling that this new paper
would have been of still greater interest had the author, in
addition to the resume of his results, thought fit to discuss
more fully the difficulties which have arisen in the minds of
some who are unable to accept his theoretical conceptions.

Hubrecht has no doubt deliberately chosen to leave for the
moment unanswered the objections urged against his views,
possibly with the hope that objections — if such there are —
may be formulated more precisely than heretofore, in which
case we may hope for a chapter making good this omission
at a no very distant date.

VOL. 54, PART 2. — NEW SERIES.

16

222

KICHAKD ASSHETOX.

In the hopeful expectancy of sucli a chapter I venture, as
one who has taken a practical part, though but a small part,
in the attack upon the problems in question, and as one who
appreciates to the utmost the magnitude and inspiring in-
fluence of the Professor’s work, to urge the force of certain
objections which appear to me as formidable obstacles to the
acceptance of some of Hubrecht’s vieAvs.

Chaptek I.

'J’he Ei'thekian Blastocyst.

Beginning with the question of the segmentation of the
ovum of Eutherian mammals, and passing rapidly over
the manner in which the morula stage is produced, Hubrecht
describes the embryo of this stage as consisting of an inner
mass of cells which he calls the embryonic knob, and an
outer layer called the trophoblast, and gives three figures
(2, 3, 6, p. 7) in which the inner cells (the embryonic knob)
are shown to exhibit “a different reaction against staining
reagents than the peripheral” (p. 6), the inner cells being
lighter in colour than those of the outer layer.

Perhaps the gist of the whole paper is foreshadowed in
the second paragraph of Chapter I, p. 3, where the author
speaks of “the erroneous conclusion that the mammalian
blastocyst was derived from the Sain-opsidan by a process
consisting in the gradual disappearance of the yolk, with
retention of the other developmental characters.”

It is upon the establishment of the erroneous nature of
this conclusion that the greater part of the rest of Hubrecht’s
conclusions must be based.

It seems to me, therefore, to be of the greatest importance
to weigh with care the evidence of the manner in which this
morula stage of the mammalian segmented egg is attained, and
to consider other views which have been advanced, whether
from actual observation or as the outcome of reflection.

This part of the development of the mammal Hubrecht
treats very cursorily.

EARLY ONTOGEXETIC PHEXOMEXA IX MA5IMALS. 223

[By the way, on p. 4 Hubrechfc Avrites: ‘'There seems to
be hardly any doubt that both in Ainphioxus and in man —
the tAvo opposite exti'emes in the phylum of the Chordata —
the two first cleavage cells, if separated from each other, may
under favourable conditions each of them develop into a
perfect, full-grown individual.” Wilson, Morgan, and others
have shoAvn that this may be true of Amphioxus, but Avhat
evidence is there that in man the division that gives rise to
homologous tAvins occurs at this early stage of development?
Some years ago I found a case of tAvinning in the sheep (’98)
Avhich I believe is the earliest case known among mamniids,
and the evidence from that specimen tends to shoAv that the
division Avhich results in tAvinning may occur at a later period,
namely, during the formation of blastocyst cavity. I may
mention that I recent!}" found in the ferret a condition Avhich
at first sight I took to be a similar case; but investigation
by sections shows that it is probably not a case of twinning
though it may be derived from a bi-ovular follicle after the
manner of the pluri-o\’ular follicles of some Edentates.]

On Hubrecht’s hypothesis that the trophoblast is derived
“from a larval layer, an Embryonalhiille ” comparable to
those of Desor’s laiwa, the Pilidium, or the Sipuncnlid larva
(p. 17), it is clearly convenient to shoAV that the trophoblast
originates by delaminatiou as suggested by the figs. 2, 3, G,
mentioned above, producing a typical Embryonalhulle like
the hypothetical figure of Hubrecht (’95, fig. li, Taf. Ill), of
the originating trophoblast. This also is the AA"ay in Avhich
arise those superficial layers of anarania, Avhich Hubrecht sub-
sequently— though, as I hope to show, in some cases quite
erroneously — claims as homologous to the mammalian tropho-
blast.

On the other hand such cases of segmenting mammalian
ova as those Avhich supply evidence of the origin of the outer
layer by epibole are inconvenient, and this is a point Avhich
surely should have been considered A"ery carefully, because it
is opposed to the method of formation of the supposed homo-
logous layer of the anamnia, and because it suggests an

224

ErCHARD ASSHETON.

entirely different, and, in many respects, more consistent
explanation of the origin of the mammalian trophoblast, as
lias already been urged by other Avorkers on the embryology
of mammals.

But what does Hubrecht say with reference to these cases?
“ Tlie so-called metagastrula stage of mammals, first described
by van Beneden (’80), has since been abandoned by that
author (though taken up again by Duval [’99, p. 64]).” It
is quite true that van Beneden has abandoned his explanation
offered in 1880 of the epibole, which he described as occurring
in the rabbit, but he has not I’enounced his faith in the fact,
but has reiterated it (’99) and described a similar pheno-
menon in the bat, V. murinus, as Duval also has done
(Duval, ’99). But although DuA'al supports the metagastrula
theory (one, however, Avhich is almost certainly untenable),
van Beneden gives a new, and, to my mind, a much more
plausible explanation. And at the same time, discussing
Hubrecht’s theory, van Beneden says of it that “ I’hypothese
de Hubrecht heurte a des difficultes morphologiques et
physiologiques insurmontables ; elle laisse inexpliquee I’exist-
ence, chez les Mammiferes placentaires, d’une vesicule
ombilicule et d’une foule de caracteres communs ii tous les
Amniotes et distinctifs de ces animaux” (p. 333).

Hubrecht attempts later to meet some of the objections
referred to by van Beneden (also in his former paper, 1902),
but he nowhere now discusses the question and significance
of" the epibole. Does he deny that it may occur?

To me it seems that the evidence in its favour is too strong
for the possibility to be ignored. Evidence for its occurrence
rests on the observations of van Beneden on the rabbit, van
Beneden and Duval on the bat, and myself on the sheep, but
it may as Avell be admitted at once that it is an extremely
difficult matter upon which to come to an unhesitating
opinion, because in many cases, as, for example, the pig,
phenomena Avhich are so strikingly apparent in the bat and
sheep are not to be seen at all.

There can be no doubt, however, that in the bat and sheep

EAliLY ONTOGENETIC PHENOMENA IN MAMMALS. 225

and rabbit many specimens show what apparently are stages
in epibole with such diagrammatic plainness that the proba-
bility of an epibole cannot be ignored, and although such
plainness is absent from many, e.g. pig, mouse, dog, etc.,
there is nothing in these cases which prevents a similar inter-
pretation being placed on them. A difference in staining
reaction does not become established until a later stage. In
such cases one can neither affirm nor deny epibole, but my
point is, there is no evidence against it even in them. In
Tupaja, according to Hubrecht, the staining differentiation
does not arise until after epibole has occurred. If Tupaja
were typical, if the cases of Lepus, Ovis, Vespertilio, were
only like Tupaja, then Hubrecht’s theory of formation of
the trophoblast by delamination might be regarded as
established.

I know that in 1894 I myself doubted van Beneden’s con-
tention that epibole occurs iu the rabbit, but the specimens
of segmenting ova of the sheep which I obtained and
described iu 1898, being extremely well preserved and in
excellent condition histologically, were to my mind so con-
vincing that I was quite converted to the view of van Beneden,
at least as regards the fact of epibole, although I differed
from him in the interpretation of the facts. And since that
time we have had the further evidence of Duval and van
Beneden derived from their study of Cheiropterau develop-
ment. If this epibole occurs, that is to say, if thei’e really is
a growth of one set of segments rouud another set during
the early stages of the segmentation of the Eutherian
mammabs egg, it seems to me possible to hold one of three
quite plausible views. Either it is :

(1) An early separation of trophoblast and a growth of
trophoblast cells round the embryonic knob;

(2) A growth of the epiblast over the yolk mass like the
sliding of the “extra embryonic” epiblastic edge of the
blastoderm over the yolk in a bird’s egg, as van Beneden
suggests (though he does not use the terms “epiblast” and
“ hypoblast ”) ; or —

226

ItICHAED ASSHETON.

(3) A growtli over the temporarily lethargic epiblastic
mass by the yolk or hypoblast cells (Minotj myself).

'I’he last two interpretations pi’e-snppose a derivation of
the Eutherian mammal from Sauropsidan-like ancestors with
large-yolked eggs.

All these are plausible theories, and it would have been
very interesting to have had Hubrecht’s opinion upon them,
especially as the last two are completely opposed to his own
views. Incidentally Hubrecht, in connection with the forma-
tion of the cavity of the blastocyst, says (p. 6), “ E. van
Beneden has ascribed the origin of the free .space between
the epithelial outer layer and the inner mass to the extension
of intra-cellular vacuoles (’9d). His interpretation has found
no support in the results obtained by Keibel and myself, nor
in those of Selenka for the opossum.” 1 should like to say
thfit as far as my experience goes the cavity of the blastocyst
appears to arise, as van Beneden says, as the extension of
inti’a-cellular vacuoles in the pig and ferret, less clearly so in
the sheep (and from general appearance of later stages still
less in the goat), while in the rabbit it would seem as
distinctly to be intercellular.

Perhaps there is not very much in it, but so far as it goes,
if the origin of the cavity is intra-cellular rather than inter-
cellular, it tends towards the probability of the cavity in
question being a vacuolation in a yolk bearing syncytium
like the germinal wall of the Sauropsidan egg rather than a
space between “embryonic” cells and an “ Embryonalhiille ” ;
that is to say, it supports the last theory of the three suggested
better than either of the other two. There can be no doubt
tliat there are in the sheep, pig, ferret, goat (Assheton,
’08, fig. 5), strands of protoplasm which connect the inner
lining of the inner mass to the wall of the blastocyst, and
this tends to sliow that the inner lining of the inner mass is
of common origin with the wall of the blastocyst; that is to
say, the hypoblast and trophoblast are one.

With reference to the three diagrams on pages 229, 231,
233 of my paper referred to above (’08), 1 fear I have not

EAELY ONTOGENETIC PHEN05IENA IN MAMMALS, 227

made it sufficiently clear that they do not represent any
particular animal, hut that they are to be regarded as
generalised diagrams representing three plausible interpre-
tations of the observed facts of Eutherian early develop-
ment. The first was suggested by van Beneden’s papers on
the rabbit and bat. In this I ought to have shown the
epiblast thickened at the embryonic pole from the first,
because van Beneden lays stress on the fact that the inner
mass contains from the first the embryonic epiblast. As
drawn it is a compromise between van Beneden’s and Duval’s
account of the bat.

But to return for a moment to the three alternative
suggested explanations of the epibole. The first alternative
would satisfy Hubrecht’s hypothesis so far as the trophoblast
of Eutherian mammals is concerned, but how can he accept
the explanation if, as he desires to do, he regards this
mammalian trophoblast as the homologue of the epidermic
layer of epiblast in the Amphibian, or the outer coat and
periblast of Teleostean, or certain superficial layers in
Sauropsida ? In all those cases the layer in question arises
later and by delamiuation, as an investing sheath. There is
no hint of a growth round an inner mass. If homologous, is
it not sti’ange that the mammalian trophoblast should be
formed by epibole ? That is to say, Hubrecht cannot well
accept this explanation of the epibole, as it would be
inconsistent with the rest of his theory.

Other objections to Hubrecht’s view are that it does not
give a satisfactory explanation of the phenomena known as
entypie, nor for the rejection of the trophoblast cells by the
epiblast of the embryonal area (pig, rabbit, mole, etc.),
whereas if, as the tliiid alternative requires, the trophoblast
has had a yolk -mass or hypoblast origin in evolution the
rejection is natural enough. Van Beneden’s view of the
epibole is plausible, but this again does not satisfactorily
ex2)lain the rejection of the trophoblast cells by the embryonal
area, nor does it really explain entypie; and it is not
su|-)ported by the nature of the epibole suggested by the

228

KlCHAKl) ASSHETON.

segmenting egg of tlie sheep. Moreover, the growth round
would seem to be in the opposite direction to that required
by the hypothesis. As I have said on a former occasion
(^08), “van Benedeu is also a little inconsistent, for in his
former papers on the rabbit he shows that the epibole is in
the opposite direction to that required by his newer hypo-
thesis. In 1880, in his description of the rabbit, he describes
the epibole as occurring in such a way as to place the inner
mass at the point where the enveloping rim coalesces (vide
van lieneden, ’80, fig. 7, fig. 5^^'), and marks the spot where
the embryonal area will eventually be.”

Tlie third alternative I still believe to be the most com-
pletely consistent explanation of the early stages of the
development of Eutherian mammals, and so strongly do I
believe in it that I would urge the development of the sheep
and bat as strong evidence against Hubrecht’s attempt to
destroy the old group of Amniota.

This third alternative (p. 22G) accounts (1) for the epibole
by regarding it as a feature peciiliar to Eutherian mammals
and due to an overflow, as it were, of the yolk or hypoblast
cells over the epiblastic rudiment, which, as the centre
whence the great bulk of the animal will be formed (for it
includes the whole of the secondary growth centre), is bound
to l emain inactive.

(2) F or this lethargic state of the epiblastic mass continuing-
through many days, until the space necessary for development
of the embryo, on the old Sauropsidan egg type, has been
provided.

(3) For the sharply marked off character of the epiblastic
mass.

(4) For the frequent protoplasmic connections between the
inner layer of the inner cell mass and the blastocyst wall.

(5) For the fact that cells of the inner mass pass into the
outer cell wall (Assheton, 1908).

(6) For the rejection by the embryonal area of cells of the
trophoblast layer.

Lastly, it may be said that the theory demands no change

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 229

of function, the hypoblast or yolk-cells having- retained their
function of providing- nourishment for the developing embryo
throughout the entii-e period of transition from meroblastic
to holoblastic conditions.

But of course it involves descent of mammals from large
yolked eggs of the Sauropsidan type, and is, therefore, so
diametrically opposed to Hubrecht’s hypothesis that he would
seem to consider it unworthy of consideration.

2. Thk Mktathekian and Pkotothekian Blastocyst.

Hubrecht terminates tlie above discussion with the con-
clusion that “all the Didelphia and Monodelphia hitherto
investigated show at a very early moment the didermic stage
out of which the embryo will be built up enclosed in a cellular
vesicle (the trophoblast), of which no particle ever enters into
the embryonic organisation.” Leave out the words “ Didel-
})hia and,” and agreement with the conclusion will be easy
enough.

The only cases we have recorded of the earliest stages of
the Didelphia are that of tlie opossum by Selenka (’87), and
those of Dasyuriis and Perameles by Hill (’U8). Hubrecht’s
interpretation of Selenka’s figures is well known from his
paper on Tarsius (’02), in which (p. 55 et seq.) he argues
that the outer layer of the vesicle, Pig. 10, Taf. xvii of
Selenka’s j)aper, is trophoblast, and that the large inner
ceil, cu., gives rise to the whole embryonic ectoderm and
endoderm.

In the accompanying text-figure, fig. C represents Selenka’s
description of the opossum blastocyst.

Hubrecht’s interpretation would be legitimate enough if
we regard the character of the cells as drawn by Selenka
as only diagrammatic. If, however, Selenka’s sections are
accurately represented, it is very hard after studying
Selenka’s figures 2, 3, 4, 8 on Taf. xviii to believe that the
thickened outer part of the outer layer labelled ex in fig. 4
has really been derived from the group labelled en in fig. 2

Text-fig. 1.

Diagrams to sliow the formation of the hlastocyst in : A, Proto-
theria after Caldwell, Semon, and Hill and Wilson's description ;
B, Metatheria after Hill's description of Dasynrus ; C. Meta-
theria, after Selenka's description of the opossum. White =
epihlast ; black = hypoblast and yolk ; dotted area = tropho-
blast ; shaded area = embryonic knob.

EARLY ONTOGENETIC THENOMENA IN MAMMALS. 231

in fig. 10, Taf. xvii. When we turn to Dasyurus there is
still greater difficulty. According to Hill’s (’08) interpreta-
tion of his specimens the descendant cells of the lower of his
two pi'imary rings which result from the segmentation of the
ovum are to be regarded as “ trophoblast,” the descendants
of the upper ring as epiblast and hypoblast as indicated in
my diagram, text-fig. 1 B.

If this is the correct interpretation clearly there is no time
when the conditions are as all agree them to be in Eutherian
mammals.

There are certain fundamental differences between the
conditions of the segmenting Metatherian and Eutherian
eggs— if the only two cases so far known of the former are
typical of the group. Thus, in the iVIetatherian egg in the
four-segment stage the segments are like those of Amphioxus
or frog, in one plane, whilst in the Eutherian egg they are
always eventually in pairs lying across one another, as 11.
Hertwig points out in his article in O. Hertwig’s ‘ Handbuch,’
vol. i. With this is probably correlated another great differ-
ence, namely, that the result of further segmentation is in
the Metatheria a hollow blastula, in Eutheria a solid morula.
Whether Hill is right in calling the lower ring ectoblastic
trophoblast is perhaps doubtful, but clearly there is no hint
in his description (‘ Nature,’ ’08, p. 049) of any Embryo-
nalhiille, any layer lying over the definitive epiblast, at any
time of segmentation. Hill has not yet published any
detailed account of the formation of the hypoblast ; but
there is nothing in what he has published to pievent one
regarding the trophoblast as of yolk-cell or hypoblastic
origin. The difference, then, between the Metatheria and
the Eutheria would in that case be that while in the latter
the hypoblast and yolk-cell mass overflow the embryonal
area and give rise to the “ Rauber layer,” to use an old term,
in the Metatheria it never overflows. So in the Metatheria
there is neither Rauber layer nor entypie. The condition of
the opossum is less ea.sy to bring into line with my view unless
one takes it to have been brought about by a diminution of

232

KICHARD ASSHETON.

the whole yolk and hypoblast mass, so that the large inner
cell, called by Selenka hypoblast, corresponds to the lower
ring of Hill’s description and has slipped inside the upper
ring, resulting in a condition not unlike the Monotremes
(vide diagram A in the above text-figure).

I should like to take this opportunity of collecting an error
in a drawing on p. 681 of O. Hertwig’s ' Handbuch,’ vol. i.
Fig, 244 is said, by Professor Richard Hertwig, to be
“ Furchungsstadien des Schafes nach Assheton,” and in it
two figures are given of what appears to be the four-cell
stage of the sheep. The drawings are really those of the
rabbit, not sheep, and are taken from my ’94 paper on the
“ Re-investigation into the Early Stages of the Development
of the Rabbit.”

The second figure shows the four-celled stage with the
segments arranged in the way I have just taken to be typical
for Metatheria. I have no recollection of the specimen now,
so cannot say whether the manipulation can have in any way
accounted for the abnormal displacement — if abnormal it is,
as I believe.

Anyhow, it is the only figure I have been able to find after
searching Rischoff, Coste, van Beneden, Duval, Selenka,
Sobotta, Heape, Melissinos, etc., who have observed specimens
of Eutherian four-segment stages in which the segments
are arranged thus.

A possible — and indeed probable — explanation is that this
specimen was obtained immediately after division, at which
time it is possible that they may lie in one plane, the twisting
being due to some cytotropic influence which places the two
pairs almost immediately in the crossways position, where
they remain for the next two or three hours till the third
cleavage plane arises. It is clear that whereas it may very
rarely happen that the stage immediately succeeding the
second division is obtained if it last but a few minutes, the
foui’-celled stage in the crossways position lasting two to three
hours will turn up much more frequently. Of recent years
1 have seen four-segment stages of rabbit, pig, dog, ferret.

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 233

and hedgehog, and I have always found the pairs cross-
ways.

On the other hand, the fact that a three-segment stage is
often met with suggests that one segment as a rule divides
before the other segment, and so brings about the cross-ways
position.

Discussing the arching of the epiblast plate while bursting
its way through the trophoblast as it does in Tupaja, Talpa,
Ovis, Sus, etc., Hubrecht sa,ys : “ The causes of the folded
condition of the embi’yonic shield can hardly be so simply
mechanical as Selenka supposed. They remain obscui’e for
the present, and will come anew under consideration when the
origin of the amnion will be discussed ” (p. 10). It is diffi-
cult to believe that the arching does not aid in the rupture ot‘
the trophoblast, though the fact that it brings about the
rupture — if it is a fact — is not necessarily the reason why the
arching occurs, because the rupture is brouglit about by other
means in cases like Lepus or Sorex.

The causes are less obscure on the hypothesis that Eutheria
derive their peculiar couditions from Sauropsidan ancestry.

With reference to the Monotremes Hubrecht speculates as
follows. He admits that “ our knowledge is as yet very
scanty,” but brings what is known about the segmentation
into line with the higher mammals by homologising the
outer layer (which Wilson and Hill regard as epiblast) with
the trophoblast of the Eutherian, and Wilson and Hill’s (also
Semon’s and Caldwell’s) entoderm Avith the embi-yonic knob
of Eutheria, and regards the yolk as an accumulation on an
ancestral type peculiar to Prototheria and not derived from
Sauropsida.

For this view I can see no reason derivable from actual
specimens described and figured by those four authors.
Hubrecht’s fig. 07, p. 23, might be that of a Sauropsidan, e.g.
sparroAv ; nor do 08, 07, GO, really differ from the Saurop-
sidan blastoderm. Possibly the segments in 60 look rather
less part of the yolk than is often the case in Sauropsidan
eggs, but it is certainly remarkable that there should be the

234

lilCHARD ASSHETOX.

usual plug of fine yolk (characteristic of the Avine egg as the
nucleus of Pander) under the segmented area if the Monotreine
egg is to he regarded as a trophoblastic vesicle, including
“ besides an embi’jonic knob a very considerable amount of
food yolk, the development of which will have gone parallel
with the change in the ancestral line from viviparity to ovi-
piirity.”

There is no trace of a breaking through the trophoblast by
the inner cells, which, on the contrary, seem to spread out
under the outer layer into a thin membrane. When at a later
period the primitive streak proliferating area seems to project
through (Wilson and Hill, PI. 3, fig. 26) after the manner of
Selenka’s figure of the mesoblast pushing through the epi-
bhist in fig. 9, Taf. xviii, of the opossum, this condition could
not be taken as the pushing of epiblast through a trophoblast,
as the neural plate is undoubtedly in front of this area, and
has been formed from the originally outer layer.

So that neither in the Prototheria or the Metatheria is
there any really tangible evidence of a trophoblast occurring
as a covering layer over the definitive epiblast as in Eutheria.

Summarising up to the present stage I submit that
Hubrecht, while ignoring alternative interpretations, has not
Tuade good his own case either for the presence of the tropho-
blast layer in Prototheria and Metatheria, or for the origin of
the trophoblast in Eutheria, as a special Embryonalhulle
formed by delamination from the epiblast, and has not
attempted to meet any of the objections presented to his
theory by the study of the segmenting- stage of such mamma-
lian eggs as those of Lepus, Ovis, and Vespertilio.

Phylogenetic Origin of Trophoblast.

Confirmed in an opinion which as regards Prototheria and
l\Ietatheria is based on very doubtful evidence, that all classes
of mammals have a larval envelope, the trophoblast, Hubrecht
proceeds to speculate upon its origin outside the group, and
starting with a ccfilenterate ancestor, says: "A tendency to

EARLY ONTOGEXETIC PHENOMENA IN J1AMMAL8. 235

exchange the radial for a bilateral symmetry and to separate
the coelom from the enteron must at one time have character-
ised certain coelenterate ancestral forms, as has already been
advocated b}^ Sedgwick (’84) and by myself (’05) on earlier
occasions. It is not straining the imagination to assume that
in this line of descent closely related forms may have deve-
loped, some with, others without, a larval envelope, tem-
porarily ensheathing the cellular elements that will build up
the embryo itself, and thus foreshadowing the separation
among their later vertebrate descendants of such with and
such others without a trophoblast.” lie then shows how this
sporadic appearance of larval envelopes occurs in Nemerteans
and Annelids. AVhy, therefore, not also in Yertebrata ?

On the assumption of a terrestrial life an animal ‘Avould
doubtlessly score certain advantages if at the same time it
became viviparous. . . . And towards the efficiencyof this
viviparous condition the larval envelope could immediately
contribute by the mere change of its protective or locomotor
significance into an adhesive one” (p. 18), and thus we are
led on to the conception of the origin of the mammalian
placenta.

' Hubrecht quotes Mehnert as showing the existence of an
outer or trophoblastic layer in Sauropsida, e.g. tortoise,
lizard, and snakes and many birds. But Hubrecht himself
finds that Mehnert is proving too much, and rejects those
cases which are inconvenient, but retains one case, that of
Clemmys, described by Mitsokiiri, and also Sphenodon and
Cliaimeleo, on the authority of Schauinsland, as being truly
trophoblastic, and dismisses the rejected ones as cases
“ distantly comparable to a mammalian epitrichial layer.”
Of course on the theory I advocate, the trophoblast is of
Eutherian mammalian origin only, and is not homologous to
any form of envelope outside the group of Eutherian mammals.

Turning to the Icthyopsida Hubrecht finds the Embryonal-
hiille present as the Deckschicht in Amphibia and Ganoids
and Dipnoi, and “ more unquestionably ” in the Teleosts.

As regards tlie Teleosts I have myself (’08), in my paper on

236

EICHAl^D ASSHETON.

the “ Teleostean Eggs and Larvfe from the Grambia River,”
shown how like the conditions are, that is to say physiologi-
cally, to the mammalian egg, for the Deckschicht is continuous
with the yolk mass, which is one piece with the hypoblast ;
but I see no reason to think this is anything but analogy.

The Deckschicht of the Amphibia is, however, a verv
different thing. It was to the Deckschicht of Amphibia that
Hubrecht in his paper (’95) drew attention as being the
homologue of the trophoblast and amnion of mammals. But
as he says now, “ I would never look upon the Deckschicht of
the Amphibia as having been the first starting-point of what
afterwards becomes amnion and chorion of the higher
mammals. We may safely say that Deckschicht and tropho-
blast are homologous and of similar descent, but we cannot at
present fully picture to ourselves what has been the arrange-
ment of the larval envelope in the common parent form from
which both have derived ” (p. 81).

Again, ‘^the cells of the Deckschicht proclaim their transi-
tory and larval significance yet further by the fact that they
disappear in later developmental stages, and that it is only
with the constitution of peculiar larval organs that they play
any part ” (p. 80).

“It should, however, be observed that if we are willing to
admit the homology of the Amphibian Deckschicht with the
Mammalian trophoblast, we must then unhesitatingly go one
step further” (p. 81). Thus, Hubrecht clearly relies chiefly
upon the assumed homology of the Amphibian Deckschicht
and Mammalian trophoblast.

I think I can convince Hubrecht that the Amphibian
Deckschicht must, like the “ epitrichial layers” of the
Sanropsids, be rejected as something different from a tropho-
blast or purely larval envelope.

If Hubrecht will examine sections of the segmented egg of
Ran a temporaria, Bufo vulgaris and probably other
Anura and of larval stages up to 5 or 6 mm., he will find that the
Deckschicht layer takes a very important part in the forma-
tion of the tissues of the brain. They become the neuroglia

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 237

cells of the brain and spinal cord, while the “ GrnndSchicht ”
becomes the nervons tissue. It is, in fact, a separation into
what His termed “ spongioblastic ” and “ neuroblastic ” tissue.

The early stages of this process are described in a paper
by myself with figures in vol. 37 of this Journal, pp. 166-169
and PI. 18.

To summarise the present section I would urge that
Hiibrecht has not established the homology of the envelopes
he mentions either among themselves or to the trophoblast
of Eutherian mammals, because :

(1) The epidermic layer of the Anura, upon which great
reliance is laid for the support of the theory (p. 81), is neither
morphologically nor physiologically similar to the trophoblast
of Eutherian mammals, but is in fact the result of an early
separation of spongioblastic from neuroblastic elements of the
ectoderm.

(2) The vestiges of an outer envelope which Hnbrecht
retains as instances of larval envelopes among the Sauropsida
differ from those he rejects as epitrichial only in the time of
their development. The former appear before amnion forma-
tion, the latter after. Quite similar vestiges may be found
among the Mammalia which render the comparison uncon-
vincing, because the real trophoblast in the Mammalia is
obviously present elsewhere.

It is curious that Dendy (’99) should make no mention of
the occurrence of this layer in Sphenodon. But if we take
Schauinsland’s figures as being correct and regard the
presence of this layer as a well-established fact, we cannot
compare it with the deckschicht of Anura because it takes no
part in the formation of the embryo, nor with the Teleostean
layer because it is quite Hee from the yolk-mass.

A comparison with the Eutherian trophoblast is hardly less
difficult. Hnbrecht attempts to do so by comparing the
whole extra-embryonic epiblast of Sauropsida with the
trophoblast of Eutherian mammals by supposing the double-
layered condition of, for instance, Chameleo (Schauinsland,
’03, figs. 182-219) to be a separation into cyto-trophoblast
VOL. 54, PART 2. — NEW SERIES. 17

238

RICHAED ASSHETON.

and plasmodi-troplioblast. But where, even in Erinaceus or
Vespertilio which he cites, can a plasmodiblast layer be found
as a continuous thin layer of squamous cells ? The character
of the layer as drawn by Schaninsland is that of an ordinary
siiperficial layer such as may be found on the embryo them-
selves of Sauropsida and Mammalia alike.

Nevertheless pages 19-26 ai*e by no means the least inter-
esting of Hubrecht’s stimulating work.

(3) The comparison of the outer layer of the blastocyst of
Prototheria and Metatheria (which is admittedly developed in
a very diffei*ent manner) to the trophoblast of Eutheria seems
to me, with all respect to his great authority, to be based on
the slenderest of foundations.

This leaves ns with the traces of larval envelope found in
Dipnoi and Ganoids and Teleosteans. There is certainly in
Dipnoi a separation into the layers of epiblast, though not so
distinct a one as in the Anura, and its subsequent history is
not like that of the Anura with reference to the central
nervous system.

Graham Kerr (1903) (PI. 4, fig. 18, ‘ Quart. Journ. Micr. Sci.,’
vol. 43), shows a neural groove into which the outer layer of
epiblast passes, but the groove is very shallow, and although
a chink is present for a short time (fig. 19) in which some of
the outer layer cells might be imbedded, it is not at all pro-
bable that the outer layer takes any part in the formation of
the central nervous system. Graham Kerr says : ‘^The whole
thickening of the keel is confined to the deep layer of the
ectoderm — the outer layer passing unaffected over the floor
of the groove.”

But neither in Ceratodus nor Lepidosiren does the distinc-
tion seem to be of the same nature as in Teleostei.

In the Teleostean the condition is far more like that of the
trophoblast of the Eutherian mammals than is any other of
the supposed larval layers mentioned by Hubrecht. It clearly
is not concerned structurally in the embryo fonnation, and
being continuous with the periblast — indeed part of the peri-
blast— it may be said to be trophic. Also it separates off

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 239

at an extremely early age (Kopscli, ’01, Assheton, ’08), thus
resembling the Eutherian trophoblast.

The Teleostean Deckschicht differs from those of the
Amphibia and Dipnoi in being quite free from any connec-
tion with the lips of the blastopore, over which it passes as a
continuous layer, e. g. Gymnarchus, Salmo.

In view of the uncertainty of its presence in Sauropsida,
of its widely diffei'ing character and relations among Amphibia,
Teleostomi and Dipnoi, of its unique character and function
in Eutherian mammals, of the dubious nature of its existence
in Monotremes and Marsupials, is it not rash to regard all
these outer layers as homologous, and as constituting a
feature of such importance as to justify the abandonment of
the old group Amniota and the formation of a totally new
association, having this feature as the chief diagnostic
character? Anyhow, the points to which I have drawn
attention seem to me to deserve further consideration.

Another point I might make here. On p. 106 Hubrecht
writes, with reference to his very remarkable observations
published in 1899 on the blood formation in the Tarsius
placenta: “The production of blood-corpuscles by the cells

of a larval envelope is surely an unexpected histological
phenomenon. Still, the details of differential segregation
during the successive stages of cell lineage are not yet well
enough known to justify any apodictic negation. The
possibility is not excluded that at the first cleavage (suppose
this to separate trophoblast from embryonic knob) certain
potentialities of haematogenesis may be passed on to this
trophoblast mother cell.”

On the hypothesis that the Eutherian trophoblast is really
of yolk-cell or hypoblast origin, the formation of blood-
corpuscles from it is much less extraordinary, for it is from
this layer that the first formed blood-cells arise in Sauropsidan
and other vertebrate embryos.

240

RICHARD ASSHETOy.

Feotogenesis or Kephalogenesis ?

Deuterogenesis or Notogenesis ?

I hope I have a desire no greater than is legitimate to
support iny own view of processes etnbryological.

I should, however, like to state clearly why I persist in
using the terms “ protogenesis and “deuterogenesis” in
preference to the terms coined by Professor Hubrecht,
namely, “Kephalogenesis” and “Notogenesis,” which
obviously refer to the same phenomena.

I do so because Hubrecht’s words express conceptions,
which, although having reference to the same phenomena
which I wish to express by protogenesis and deuterogenesis,
signify a different interpretation, which, in my opinion, does
not represent the actual facts. And I would even claim some
consideration because I believe that I was the first to recognise
that protogenesis is in essence the production of a radial
symmetry due to growth from one centre involving gastrula-
tion, and that deuterogenesis is growth in length, bringing
about bilateral symmetry and has nothing to do with gastrula-
tion (though it may be an inevitable consequence of it), which
conceptions I expressed at an earlier date under the terms
primary and secondary growth centres (1894). Protogenesis
and deuterogenesis form a, convenient paraphrase of those
terms.

On p. 63 Hubrecht claims to have been Avith Keibel the
godfather of the unwarranted hypothesis that gastrulatiou
occurs in two phases (Keibel, ’89, Hubrecht, ’88).

Who claims actual parentage I do not know. But that it
Avas a most mischievous and aAvkAvard child I can Avell
believe.

Fortunately both godfathers have fianlly disclaimed, b}'
their papers in the year 1905 (^ Anat. Anz.,’ ‘ Quart. Journ.
^licr. Sci.’), any further responsibility in their adopted off-
spring, and they noAV admit that it is a matter of great
importance in vertebrate embryology to distinguish betAveen

EA.RLY ONTOGENETIC PHENOMENA IN MAMMALS. 241

the true gastruhi stage and the post gastrula stage, whicli
latter is the growth in length of the embryo, and it is because
I also believe so strongly in the importance of the distinction
that I wish to establish the more accurate terms above. I
may also claim to have arrived at my conclusion by actual
experimental observation, having spent mnch time in so
doing’, and, as evidence of this, I may mention my papers of
the years 1894, 1896, 1905.

Hnbrecht hardly does me justice in ignoring my experi-
inental evidence on the subject. Moreover, this theme was
the gist of my tliree papers in 1894, “Re-investigation of
the Early Stages of the Development of the Rabbit,” “ The
I’l’imitive Streak of the Rabbit,” and “On the Growth in
Length of the Erog Embryo,” from the last of which I may
quote one paragraph from p. 288 : “ In other words, I

believe that as in the rabbit, so in the frog, there is evidence
to show that the embryo is derived from two definite centres
of growth, the first, and phylogenetically the older, being a
protoplasmic activity which gives rise to the anterior end of
the embryo (= gastrula stage) ; the second, which gives rise
to the growth in length of the embryo; which centres of
growth occupy the same relative positions in location and in
sequence of time, and probably to each are due the same
parts of the embryo.”

In my subsequent papers on the chick, 1896, and gi-owth
centres, 1905, I have described some of a good deal of
experimental work which I have done in confirmation of this
opinion.

Rut although Hubrecht now tpiite agrees that notogenesis
has nothing whatsoever to do with gastrulation, it is quite
evident that his conception of notogenesis is not the same as
mine of deuterogenesis, and since I believe that my conception
is nearer the truth than his, I must explain where, as it seems
to me, his error lies.

In the first place I should like to refer to a footnote which
appears in Hubrecht’s English edition of his paper, “The
Gastrulation of Vertebrate.s,” in the ‘Quart. Journ. Micr.

242

EICHAED ASSHETON.

Sci./ vol. 49, in which he says that I quite misunderstood
his German version, in so far as I believed him to “hold the
vertebrate mouth to be in any way derived from the stomo-
daeum of an Actinia-like animal.” I am sorry I made the
mistake, which I made at least by implication, confounding
in my mind Hubrecht’s with Sedgwick^s very similar theory,
published in 1884, to which, by the w^ay, Hubrecht made no
reference when he put his forward in 1902.

The accompanying text-fig. 2 shows clearly enough what is
the difference between Hubrecht’s conception and mine, and
why I prefer protogenesis and deuterogenesis to kephalo-
genesis and notogenesis.

The aboral surface of the coelenterate, according to
Hubrecht, becomes the ventral surface of a vertebrate ;
according to my interpretation the aboral surface becomes
the anterior. According to Hubrecht the oral surface of the
coelenterate becomes the dorsal surface of the vertebrate ;
according to me the oral surface of the coelenterate is the
posterior surface of the vertebrate, and so on, as shown in
the figure.

I claim that my interpretation is founded upon actual
experiment on the living embryo, which can be tested by
an}'one.

Where can Hubrecht find experimental evidence in support
of the elongation of an actinian or other mouth in, for instance^
the frog, with concrescence or coalescence of its walls ? Or
how can a theory of concrescence be reconciled with experi-
ments such as those of Kopsch on the trout?

I claim that the experiments which I described in my
paper of 1905 prove that in the frog at any rate the embryo
does grow in the way illustrated by my figures, and that this
is absolutely opposed to tlie method of growth required by
Hubrecht’s theory.

Again, with reference to notogenesis, Hubrecht seems
to have no very clear conception as to tlie extent of its
influence. In his paper on Gastrulation ” (’06), he defines
kephalogenesis and notogenesis thus : “ The distinction

Text-fig. 2.

a.

CL

CL.

Diagium to show the difference in the conception of kephalo-
genesis and notogenesis on the one hand and protogenesis and
denterogenesis on the other. The middle figure, (3, represents
the gastraea or early coelenterate stage, with blastopore on the
lower surface. The upper figiu’e, a, represents the vertebrate
according to Hubi-echt, derived by elongation of the gastrula
in the direction v d, producing an elongated actinian stomo-
damm, followed by concrescence of the walls of the stomodseuni
to foiTu the notochord. The lower figure, y, represents the
vertebrate according to the author's conception by elongation
of the gastrula in the direction a p by activity of cells forming
the blastopore lip. Subsequently, the activity of the ventral
part dies out, and the more dorsal paid continues active and
fonns the tail.

244

RICHARD ASSHETON.

here intended between ‘ kephali ’ and ‘ notes ’ is not iden-
tical with that between head and trunk (trunk segments
having been ascertained to enter into the composition
of the head), but that on one side should be ranged the
very foi’emost portion of the head to which the ophthalmic
[olfactory?] and optic nerves belong, whereas on the other
we place the further subdivisions of the brain with their
cephalic nerves, as also the basis of the skull with the remains
of the notochord it contains, the visceral arches and the whole
of the trunk.”

I am inclined to think that a very considerable part of the
gut, including the pharynx, and certainly the heart, are proto-

Text-fig. 3.

Diagram of a vertebrate to show approximately the parts clue to
protogenesis and denterogenesis respecdively. a, anterior ;
d, doi-sal ; p, posterior ; v, ventral suii'aces.

genetic. But the details of this form a subject for further
experimental research.

Then in his paper of last autumn Hubrecht says (p. 43) :
“ I think we may safely say that by the rapid extension back-
wards of the differentiation process, . . . the dorsal region

of the trunk is laid down in outlines (hence the word noto-
genesis), whereas the derivates of the ventral mesoblast find
employment in the construction of the posterior and postero-
ventral portion of the embryo.”

The ventral mesoblast, by which Hubrecht means the
mesoblast proliferated from the posterior or ventral lip of the
blastopore, is, so far as the mesoblast is concerned, the evidence
of the extent of the effect of deiiterogenesis on the ventral

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 245

wall of the embryo, and as long as it is distinguishable from
the protogenetic inesoblast (Hubrecht’s protochordal plate
and annular zone mesenchyme) its anterior margin forms a
landmark between the protogenetic and deuterogenetic areas.

I have on a later page referred to a difficulty with
reference to the notochord. But I will here ask Hubrecht
two questions :

Is any part of the notochord formed — to use his own
terminology — by kephalogenesis ? That is to say, is there
any notochord anterior to his supposed vermactinian stomo-
daeum coalescence ? If he answers No, then I will ask how
he explains his earlier observation, e. g. on Sorex, where the
protochordal plate is shown to give rise to the anterior-
portion of the notochord. If he airswei-s Yes, then I will ask
how can it be due to coalescence of the vermactinian
stomodmum ?

Chapter II.

So far as my own personal observations go I can support
Hubrecht’s contention as to the origin of mesoblast (mesen-
chyme) from hypoblast in front of the primitive streak, both
as regards that in connection with the protochordal plate and
that from the annular zone in the sheep and rabbit.

Hubrecht describes the peripheral mesenchyme-producing
region in these words: “ As an elongated ring-shaped zone of
entoderm which is situated under and somewhat outside the
border of the ectodermal shield and which, standing back-
wards from the protochordal plate both right and left, meets
under the hinder j3art of the shield in the region where the
mesoblast has acquired that median thickening which is
known as the primitive streak, continued in the Primates into
the connective stalk (Uaftstiel).”

From my point of view this protochordal mesenchyme and
annular zone mesenchyme which reaches round posterior
and ventral to the primitive streak mesoblast is the mesen-
chyme of protogenetic origin, while the primitive streak

246

laCHAED ASSHETOX.

mesoblast, including Hubrecbt’s ventral inesoblast, indicates
the extension of deuterogenetic mesoblast. This is approxi-
mately shown by the dotted line of the figures in my paper on
the primitive streak of the rabbit, 1894, though the dotted line
should have been considerably nearer the “ embryo ”
(vide Assheton, 1898, p. 246, on sheep).

Hubrecht, on pages 35-37, disputes the view that there is
any forward extension of material from the primitive streak
area to form a Kopffortsatz, and holds that the protochordal
wedge becomes lengthened, “ not, however, by its sending out
any ^Fortsatz,^ but by its being, so to say, ‘spun out ^ in
consequence of the backward growth of the tissue that is
going to be the notochord,” with which description I am in
sympathy, for it is the view put forward by me and illus-
trated by the diagrams on Plate 22 of my 1894 paper on
the primitive streak.

At the same time I think this view must be slightly modi-
fied in accordance with the results obtained by actual experi-
ment on tlie growth of tlie embryo in the chick (Assheton,
1896), Avhich show that the “primitive streak” in the bird
actually becomes converted into the embryo. It may seem a
rather subtle difference, but really the elongated part of a
primitive streak such as one sees in bird or rabbit is the
stretched-out anterior part of the product of the deutero-
genetic centre of activity rather than the anterior part of the
deuterogenetic proliferative area itself, this product becoming
subsequently differentiated directly into mesoblastic somites
(which are always deuterogenetic), the deuterogenetic part of
the neural plate, and of the notochord, etc.

Hubrecht then goes on to describe another growth centre,
namely that which gives rise to the ventral mesoblast, which he
says has preceded the formation of the protochordal wedge,
and refers to four of his figures, 47-50, which, however, do
not to my mind indicate any marked distinction either iu
space or time. Surely the so-called third centre of growth is
nothing more than the equivalent of the ventral lip of the
blastopore of Amphioxus and other Anamnia, and is the

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 247

ventral portion of the naturally circular deuterogenetic pro-
liferation area.

This area is essentially circular in itself, but it assumes a
different shape by reason of varying- conditions and is a
circular ring in Anamnia, which have blastopores like Rana,
of an elongated ring- or bottle-shape (v. Sedgwick, ‘ Quart.
Journ. Micr. Sci.,' vol. 33, p. 564) in Elasmobranchs, and
streak-like in many mammals, e.g. rabbit, Carnivora-
Tarsius and birds, truncate and almost disc-like in other
mammals, e.g. Mus, Cavia, and reptiles. But in all it must
have anterior, lateral and ventral margins from which pro-
liferation of cell material takes place. I cannot conceive on
what grounds Hubrecht can separate the anterior part (his
protochordal wedge) and lateral wings from the ventral part.
The figures he refers to do not support it, and besides, it is
well known that the mesoblast sheet formed by deutero-
genesis is a continuous sheet passing posteriorly completely
round the primitive streak, broken only in front by the
separation of the notochord.

The so-called extra-embryonic coelom which develops so
early in Tarsius and man is probably protogenetic coelom, as
it is well ventral to the posterior sheet of mesoblast (Hubrecht’s
ventral mesoblast), figs. 49, 50; and indeed, in the former
figure, 49, it appears to be distinctly marked off from it. More-
over, the extra-embryonic coelom develops long before any
trace of the primitive streak, whether protochordal wedge or
ventral mesoblast is present.

Hubrecht seems to have abandoned the position he took up
in 1890 when writing on the embryology of Sorex. At that
time he was sure that the middle region of the protochordal
plate gave rise to the notochord :

“A most remarkable fact, to which I must now call atten-
tion, is this, that it is not in the posterior region of the epi-
blastic shield that the formation of the middle layer and its
earliest representatives — notochord and lateral mesoblast
plates — is first inaugurated. It is in the hypoblast that the
first differentiation occurs, which ultimately leads to the

248

KICHARD ASSHETOX.

formation of the above-mentioned structures” (‘Quart. Journ.
Micr. Sci.,’ vol. 31, p. 508). Again : “ This patch of modified
hypoblast-cells has at the beginning an oval shape, with
the long axis perpendicular to that of the embryonic shield.
Part of this patch will develop into the anterior portion of
the notochord ; for this reason I will call it the protochordal
plate.”

How does Hubrecht i-econcile the formation of part of the
notochord (what I should call the protogenetic portion of the
notochord) from the gut lining with his theory of the origin
of the notochord from the supposed fused Actiniau stomodaeum
according to his present theory? (p. 109). Does Hubrecht
abandon his earlier conclusions, or does he allow that some of
the notochord is not formed from the stomod^al protochordal
wedge ?

In my humble opinion the Professor was on much sounder
ground in 1890 than now, althongli, as related elsewhere, I
cannot follow him in the idea of two gastrulation periods, an
hypothesis which he has, as we have seen, abandoned since.

Hut surely the difficulties of coenogenetic and palingenetic
tissues, of mesoderms of various origin, of notochord of
epiblastic or hypoblastic origin, are to a large extent
smoothed away and the problem vastly simplified by an
appeal to the way the things actually grow in the embryo as
evidenced by exjierimental observation.

For we find that there is a part of the organism formed, so
to speak, on the egg — in situ — roughly radially symme-
trical, which is what I call the protogenetic ai’ea, and this
alone represents a gastrula or coelenterate stage.

To this subsequently is added tissue from a growing point
(the deuterogenetic area of proliferation), which area itself
can be called neither epiblast, hypoblast, nor mesoblast; but
the material produced by active proliferation of this growing
point becomes epiblast, hypoblast, or inesoblast, according to
the nature of the layer with which it is in direct con-
tinuity.

The protochordal plate and the annular zone inesoblast,

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 249

with, of course, the hypoblast from which these parts are
derived, together with the epiblast which overlies them, as
well as everything ventral to the annular zone, constitute the
protogenetic tissues, which are of more ancient origin than
the primitive streak tissues, and everything this secondary
area of proliferation gives rise to (deutei’Ogenesis) are of a
more recent origin, and include the protochordal wedge of
Hubrecht (Kopffortsatz of others), the so-called ventral
mesoblast and the lateral plates of mesoblast which are
really one and the same thing.

I entirely agree with Hubrecht’s comparison of these areas
in question with corresponding ones in Amphibia, pp. 46-54,
though I claim to be not included among' those who were so
^‘naturally biassed” as to fail to see these points in Amphibia
(vide H^uart. Journ. Micr. Sci.,’ vol. 37, PI. 24, figs. 8-11,
13, 14).

I have also indicated in the figure herewith reproduced
from a paper in the ^ Guy’s Hospital Keports,’ 1907, the
respective regions formed by the protogenetic and deutero-
genetic centres in various groups of vertebrates, though in
these I have omitted details such as the annular zone of
mesenchyme producing hypoblast, which is all included in
the general white area marked with black dots, but the
essential features are unmistakably indicated.

Hubrecht (p. 55) refers to birds and reptiles, and has no
difficulty in finding evidence in them as in the Amphibia of
the truth of the presence of two great embryonic growth
centres in a vertebrate (protogenesis and deuterogenesis), and
finds the same parts, protochordal plate and annular zone,
protochordal wedge and ventral mesoblast.

I am inclined to doubt, from my own observations on the
sparrow, whether the well-marked mass of cells which occurs
in the sparrow at an early stage between hypoblast and epi-
blast, shown so clearly in Schauinsland’s figures and called
by Hubrecht “ protochordal plate,” is really the material from
which any of the notochord is formed. If followed it is found
to gradually take up a more and more forward position, and

250

inCHAUD ASSHETON.

may possibly form the extreme anterior part of the heart and
pericardium, but in any case I am inclined to think it is

Archenteron Metenteron

Diagrams representing sagittal sections of vertebrates at corre-
sponding ages to show the parts of the embryo derived from the
])rimaiy growth centre (protogenesis), and that from the secon-
dary growth centre or blastopore lii^s (denterogenesis). The
latter, the deuterogenetic tissues, are shaded. The dots repre-
sent nuclei within the yolk mass and endodermal tissues of
protogenetic origin. (From ‘ Guy's Hospital Reports,’ 1907.)

altogether anterior to the notochord, and is connected with
the vascular system rather than that organ.

Text-fig. 4.

Archenteron’ Metenteron

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 251

In reference to the relation between the protochordal plate
and the secondary growth centre, Hubrecht (p. 58) writes :
“ The confluence between the earliest ectodermal downgrowth
with the protochordal plate has up to now not been specially
examined in reptiles. Still, one may conclude from the figures
here given, which I copy from other authors, that it comes
about in exactly the same way as we noticed it in I’arsius for
Mammals, and in Hypogeophis for Amphibians.”

Certainly the condition of Hypogeophis is more like the
reptilian condition than is the condition such as occurs in
the Annra or Urodela, in which there is from the first an
opening from the exterior into tlie future archenteron, namely,
a true blastopore.

Therefore, in comparing reptilian gut formations with
others, we ought to compare it more closely with mammals
and birds, and contrast it with Amphibians such as Anura
and Urodela, and with fishes and cyclostomes.

Might it not be well to restrict the term ‘‘archenteron” to
the part of the gnt which is due solely to protogenetic in-
fluence, and call the part of the gut (including neurenteric
canal) which is due to deuterogenetic influence by some such
term as “ metentei’on ” (vide diagram). If this is a true dis-
tinction, then we see that there is a very marked difference
between the Amniotes on the one hand and all the other
vertebrates.

In the Amniotes there is an archenteron formed by infil-
tration of fluid between an upper and anterior wall of cells,
usually a thin membrane, and a lower or ventral mass of
either cells or yolk mass, which, when formed, is not in open
communication with the exterior — that is to say, there is no
real blastopore. This cavity is, of course, that known as
blastocyst cavity in mammals, subgerminal cavity in birds
and reptiles. It is only at a later stage after the growth in
length has started by the oingin of the deuterogenetic centre
that a passage is formed which varies very much in its degree
of development in different types. Thus in birds it is only
recognisable as a narrow and evanescent canal, the neurenteric

252

RIOHARD ASSHETOX.

canal ; in mammals it is usually not more developed, but in a
few cases it is for a while recognisable as a distinct perfora-
tion or neurenteric canal, as, for instance, in the human
embryo (Spee), the hedgehog (Hubrecht), Ornithodelphia
(Wilson and Hill). But in the reptiles it is so evident as a
passage leadiug at first into a blind pouch (but later com-
municating with the subgerminal cavity) that it has been
mistaken for a true archenteron and true blastopore.

The true archenteron or subgerminal cavity is not so well
marked in some (Tropidonotus) as it is in others (Lacerta,
Platydactylus) ; but in all the “ metenteron ” is obvious, and
subsequently a perforation occurs, and communication
between the protogenetic cavity (the subgenuinal cavity
or archenteron) and the deuterogenetic cavity (neurenteric
canal or metenteron) is established.

And I do not think this is a subtle distinction only, but
one Avhich is essential to the understanding of the true way
in which the vertebrate embryo grows, and to my mind it
tends towards simplification.

If it were not for the hypothetical vermactinian ancestor I
see no reason why Hubrecht’s and my own views should not
absolutely coincide. For Hubrecht quite agrees with the
view that this so-called invagination cavity of reptiles, this
cavity slit or porus,” is not archenteron, and I believe that
if he would himself perform operations on developing
Amphibian, Avine, or Teleosteau eggs, he would convince
himself that the relations of the protogenetic and deutero-
genetic centres are morphologically and pln’siologically as I
have several times indicated them to be.

Gastrulation in the Ornithodelphia.

With reference to the description given by Wilson and
Hill (’07) of the corresponding stages in Ornithorhynchns, I
should like to make one suggestion.

Hubrecht accepts their interpretations, and regards the
curious mass situated some distance in front of the primitive

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 253

streak, and called by Wilson and Hill the “primitive or
archenteric knot,” as tlie equivalent of the protochordal
Avedge of his nomenclature. Nevertheless I think Hubrecht
is not very happy about it, as he says: “However, the data
concerning the earliest appearance of this protochordal plate
in Oruithorhynchus are too scanty than that I have ventured
to mention it when in the preceding pages we discussed the
protochordal plate. And it seems advisable on this point to
await yet further researches on these rare mammals, of which
it is so very difficult to obtain the required developmental
stages.”

In spite of this no doubt admirable caution, I venture to
suggest that the object called primitive knot by Wilson
and Hill (pr. K., text-fig. 7, Wilson and Hill) has nothing at
all to do with either gastrulatiou or primitive streak, but is
really the morphological vegetative pole of the egg.

I have tried to make my meaning clear by the accompany-
ing diagram (text-fig. 5).

A. represents a meroblastic egg, such as that of a sparrow.
In this the “ liquefaction ” of the yolk is seen very obviously
to result in an accumulation of fluid between the blastoderm
and the yolk mass, which forms a mass at the lower pole of
the whole egg. The diagram is, however, not an exact
representation. The epiblast, ep., is shown to have nearly
surrounded the yolk, a condition which is in reality not
attained until a much later stage of development of the
embryo itself than is indicated by the upper pole of the egg
in my diagram, which represents a sagittal section of an
early stage, perhaps eighteen hours or so of incubation.

Let us imagine, however, this growth of the epiblast, ep.,
to take place very early (so that the yolk is completely
enveloped by it) and the thickened edges to meet and fuse
early. Let us imagine that the yolk is much reduced, and its
place taken by fluid, so that the nucleated margins {gw.), the
so-called germinal walls, have met and fused also ; we should
then have a condition not unlike the fig. B, except that in
fig. B the part representing the coalesced blastoderm

VOL. .54, PART 2. — NEW SERIES. 18

Text-pig. 5.

ps.

A represents a coniiiosition of two stages in the development of
1 lie egg of a sparrow, that is to say, the “liquefaction of the
yolk ’’ and the enclosure of the yolk liy the epiblast are supposed
to have taken place several hours earlier than is the case. B
represents the blastocyst of Ornithorhynchus after Wilson and
Hill’s desci'iption. It is suggested that the “ primitive knot ’’
of those authors represents the final stage in the “liquefaction ”
of the meroblastic egg. ep., epiblast ; <jw„ junction of hypo-
blast with yolk-mass — germinal wall of Balfour; hy., hypoblast;
kt., “ primitive knot ” in B, and ventral pole of yolk-mass in A;
up., neural plate ; ps., primitive streak.

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 255

margins (epiblast and germinal walls) are eccentric. Now,
is this a possible explanation of tlie primitive knot of Orni-
tliorhynclius ?

Fig. B represents a sagittal section of the monotreme egg,
drawn from Wilson and Hill’s description, except that the
layers are shown too thick, and the primitive knot, M., is
larger than it should be.

On this interpretation the section illustrated diagrammatic-
ally by Whlson and Hill, p. 51 of their paper, should be lettered
thus : “ c.f. cell plug ” = fused edges of epiblast ; “ ent. yolk
entoderm” = hypoblast; “ vi.z. marginal or cortical zone”
= germinal wall or only remains of general yolk mass not
yet liquefied ; “ c.z. central or more cellular zone ” is probably
also fused edges of epiblast, the edge more or less inter-
mingled with the germinal wall yolk- cel Is.

The reason why it is not diametrically opposite the centre
of the embryonal area is to be sought for chiefly in the greater
expansion of the posterior arc or segment of the sphere.
Such a condition is to be found in the rabbit’s blastocyst, but
in a very much less marked degree. If a blastocyst of a
rabbit is taken out of the uterus at about seven days sixteen
hours, i.e. just before it becomes attached by its lower pole
to the walls of the uterus, it will be found to be of an oval
form, with a smaller and acuter anterior and a larger and
obtuser posterior end, and an area more or less ventral which
will be devoid of hypoblast. The centre of this area is not
diametrically opposite the centre of the developing embryonal
area, but more towards the anterior end (vide Assheton, 1894,
PI. 17, fig. 47).

My suggestion is that the position of the primitive knot is
due to a still more marked effect of a similar cause, the cause
being the activity of the primitive streak or deuterogenetic
centre, which allows of the more rapid expansion of the
posterior segment of the blastocyst.

On this interpretation the primitive knot is of little import-
ance, and the Ornithorhynchus egg appeal’s as a condition
intermediate between that of a reptilian and that of an avine

256

RICHAKl) ASSHETON.

egg ; reptilian because the neurenteric canal (metenteron)
is so much more marked in the later stages than in any bird
or even any other mammal, and avine because of the far
greater rapidity with which the subgerminal cavity forms
and swells up by infiltration of fluid and the great lengthening
of the primitive streak, which is so characteristic of all birds
and is so little marked in reptiles, though fairly well mai-ked
in some Eutherian mammals, e. g. rabbit, carnivora, ungulates.

My reasons for suggesting the above are that I cannot
quite follow Wilson and Hill in their identification of the
“ embryonic region ” of their fig. 5, Plate 2, with the structure
in question, called “primitive knot,” fig. 11, Plate 5, or of
this primitive knot with the neurenteric canal or protochordal
wedge development of the later stages, their archenteric
knot of their text-fig. 10 on p. 73.

Very considerable gaps in their material occur between
these three stages.

The earliest of the three stages, a, showing the primitive
plate, is a blastocyst measuring 6 x 6’5 mm.

The middle stage, Q, was found in four eggs which measure
10 ram. X 9‘5 or 9 mm.

The last of the three, P, measures 17’5 mm.

Comparing the three stages with corresponding ones of a
rabbit :

1st, a = 120 hours.

2nd, Q = 168 hours.

3rd, P = 188 hours.

As regards the first the comparison is difficult, because I
cannot see that there is any real ground for considering that
any layer present in the platypus is the homologue of the
troplioblast, unless — which is not at all improbable — the mass
of yolk and cells forming part of the so-called “primitive or
archenteric knot” (= germinal wall ?) is the mass of yolk-
cells into which the epiblast in the Eutherian mammal’s egg
has sunk, and by which it becomes temporarily covei’ed.

But the second stage, i.e. Specimen Q,., fig. 7, is very
much like the Eutherian mammalian embryo of the type which

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 257

resembles more closely the Avine embryo, such as a rabbit at
168-178 hours, except that the mesoblast, especially of the
anterior region, is much more abundantly developed.

Wilson and Hill’s figs. 16 and 15, PI. 5, on this interpreta-
tion pass through what Hubrecht would call the protochordal
plate, showing mesoblast or mesenchyme, which shows no
sign of having been derived from primitive streak. Thei’e is
not a hint of any proliferation of cells from the thickened
epiblast above. In respect of the mesoblast or mesenchyme
development the condition is far more Avine (cf. sparrow)
than Eutherian.

So fig. 14 is perfectly typical of a transverse section
through the front end of the primitive streak of bird or
mammal at this early stage, i.e. when the primitive streak
has attained its full length and before the counteracting-
effects of the primary and secondary growth centres have
reached their highest development.

In fact there are only slight details which distinguish it
from an embryonal area typical of those that develop upon a
vesicle tense under hydrostatic pressure.

4’here is nothing at this stage to suggest any connection
between this embryonal area and the curious mass of cells
and yolk, pr. K. (Wilson and Hill’s “ primitive knot”).

Why should not this embryonal area develop according to
the plan upon which, say, the sparrow embryonal area
develops in which a neurenteric canal is produced though
not quite in so marked a manner (vide Schauinsland’s figure,
Hubrecht, fig. 1 10) ?

In text-fig. 8 the next stage is illustrated, by which time,
according to Wilson and Hill, the embryonal area of text-fig.
7 has expanded on the whole symmetrically so that the
general outlines of mesoblast, thickened epiblast, and area
opaca retain the same general relations to one another, the
primitive streak, however, becoming slightly shorter, and
occupying a i-elatively more posterior position.

'I’his is exactly what occurs in similar types of embryonal
areas in birds or mammals like rabbits. Compare, for instance.

258

JUClIAiiD ASSllETOX.

Duval’s 'Atlas/ figs. 161 and 182, and Asslieton, ‘ Quart. Journ.
Micr. Sei./ vol. 37, PL 20, figs. 4a or 5 and 7 or 8.

Why is it not an expansion within the area itself in
Platypus just as it is in the rabbit or bird ?

What reason is there to suppose that there is in Platypus an
extension forwards in some quite original manner so as to
include a spot some distance anterior to it, except for the
difficulty the authors felt in explaining this spot?

I would therefoi'e submit to them the above suggestion
as a defensible alternative explanation. And again, the
archenteric knot is in no wise histologically like the part of
other vertebrates to which on Wilson and Hill’s hypothesis it
is supposed to be homologous.

They had four eggs showing this spot, namely specimens
D, DD (twins), Q and Y.

They give photographs of sections through Y and DD, a
semi-diagrammatic figure in their previous paper of one (of
the others ?), and also a schematic mesial plane reconstruction
of Y.

Now it must be obvious to everyone that the character of
the dorsal lip of the supposed blastopore has nothing in
common with the character of the dorsal lip of the blastopore
of Amphioxus, or Triton, where it is concerned with gastrula-
tion by invagination, or Raua or Ceratodus or Scyllium, where
it is also concerned with gastrulation but less obviously with
invagination, nor even with the anterior margin of the
" mesodermsackchen ” in any reptile, bird or mammal (where
it has nothing to do with gastrulation) either.

In all these it is a solid lip of more or less columnar actively
dividing cells, making a compact thickened epithelium of
very even and regular character, e.g. figs. 417, 418, 419,
Hertwig’s ' Handbuch,’ vol. i.

In this Platypus ‘'primitive knot” we see from fig. 2,
Wilson and Hill, 1903, quite a different nature of epithelium.
The so-called " dorsal lip ” is an inactive layer of thin
squamous cells passing into an uneven sac utterly unlike any
other dorsal lip hitherto described.

j:akly ontogenetic phenomena in mammals. 259

Also the two structures are unlike in other ways. The
primitive knot appears not very different from a yolk-laden
reticulum with few nuclei^ while the histological characters
of what is alleged to be the same spot of the later stage
present the crowding of nuclei usual to the front end of the
primitive streak.

It is hardly likely that so fundamental a process as the
formation of the neural plate — an organ whose formation in
any other vertebrate is the earliest of all organs — should be
represented by a squamous epithelial layer at a time when the
primitive streak and mesoblast is fully established; especially
unlikely is it when at this very time a plate of epiblast com-
parable in all respects, form, position, and time of origin, to a
normal neural plate, already exists in its correct position, as
shown by figs. 16 and 15 in PI. 5, and text-fig\ 7 of Wilson
and Hill’s ' Phil. Trans. ’ paper.

The next stage, called by Wilson and Hill post-gastrular,
is represented by four eggs, E, F, P, and PP. The diameter
of the first-named was 12’5 mm.

This stage is represented by their text-fig. 8.

In this it is supposed that the embryonic area has expanded
in such a way as to include now the primitive knot.

In text-fig. 8 primitive knot, henceforth called by the
authoi’S the “archenteric knot,” is shown as abutting upon
the front end of the primitive streak. But the primitive
streak itself has not increased in length, in fact it has slightly
diminished.

In fig. 7 the archenteric knot is 22 mm. in front of the
primitive streak.

In fig. 8 it touches it.

lu text-fig. 7 the primitive streak is 39 mm. in length, in
text-fig. 8 it measures 35 mm.

In text-fig. 8 the posterior end of the primitive streak is
8 mm. further off the posterior limit of the mesoderm than in
text-fig. 7.

How can the junction of the archeuteric knot and anterior
end of primitive streak have been brought about ? Has the

260

EICHAKD ASSHETON.

former ploughed its way back so as to join up with the
anterior end of the primitive streak ? This is incredible.

Has the primitive streak actually migrated forward to
meet the knot ?

Unless it has been accompanied by the mesoblast there is
no evidence of a possible advance beyond 8 mm. and the
distance to be traversed is 22 mm. (in the figures).

Has the primitive streak stretched or differentiated
forwards ?

No, it is actually shorter by 4 mm. than it was. Surely it
is simpler to conclude that there has been no junction, but
that the embryonic area has developed in an ordinary fashion
like other mammalian embryonic areas, and that the so-called
archenteric knot was really the remnant of an almost liquefied
yolk-mass, and has now become so much further reduced as
to be unrecognisable. And its position is, as before, well in
advance of the anterior border of the embryonic ai-ea.

When we come to look at the photographs of the sections
of the archenteric knot in the post-gastrula stage and compare
them with the gastrula stage we notice that the histology is
very different, for, instead of a mass of yolk with few small
nuclei we have a quite different tissue, a compact cellular
tissue crowded with nuclei.

To sum up, my suggestion is as follows :

(1) The primitive knot is the remains of the reticulum
which contained the yolk mass.

(2) The “primitive knot,” therefore, forms no part of the
so-called embryonal area.

(3) Gastrulatiou occurs as in other Amniotes by infiltration
of fluid forming a subgerminal cavity.

(4) The embryonic area of the authors, text-fig. 7, expands
into that of text-fig. 8, as corresponding ones do in mammals
like pig or rabbit, or in birds.

(5) The mass of mesoblast lying in front of the primitive
streak of text-fig. 7 is the protochordal plate which differen-
tiates into the anterior part of the vascular system, notochord
(Wilson and Hill, archeuteric plate), and mesoblast.

EAELY ONTOGENETIC PHENOMENA IN MAMMALS. 261

(6) The front end of the primitive streak becomes heaped
up into a mass of tissue, the protochordal wedge, which, fusing
with the tissues in front, adds, as the primitive streak recedes,
deuterogenetic tissues to the notochord, nerve plate and other
tissues affected by the growth in length.

(7) The boundary between tlie protogenetic part and
deuterogenetic part of the notochord may be between the
part of the notochoi’d (archenteric plate) which is described
by the authors as having a smooth under-surface and the part
to which pieces of the broken-down “ Mesoderrasackchen ” are
attached.

(8) A neurenteric canal is here formed as in some other
mammals and birds, by the interaction of the two great
growth centres, but on a much more marked scale which is
exceedingly reptilian in its characters.

If we accept the author^s nomenclature and regard the
Mesodermsackchen as archenteron, we are brought back to
the old difficulty of being unable to account for the sub-
germinal cavity. If, once for all, we abandon all idea of a
true blastopore in any Amniote, and regard the subgerminal
cavity of birds, reptiles, and blastocyst cavity of mammals as
homologous and all as archenteron, the result of protogenetic
activity, and if we regard all subsequent pores which are
invariably connected with growth in length as metenteric (to
use the term suggested before) and deuterogenetic in origin,
the matter seems to me to be greatly simplified, and is in
complete accord with the results of experimental inquiiy as
to liow things actually do grow in the living embryo.

ChAPTEKS III AND IV.

Placentation.

With the general trend of view in Chapter IV, which is
admirable in all respects, few will wish to quarrel. It
contains a clear account of the character of the trophoblastic
changes in most of the mammalian orders. It is a matter of

262

KICllAKD ASSIIETON.

satisfaction to myself to realise that the first paragraph on
p. 101 confirms the conclusion to which I came a year or
two ago, that we have, on the one hand, in the Ungulates
and some other groups great lateral expansion of the tropho-
blast, leading to the type of placenta I called '^plicate,” and, on
the other hand, cell proliferation of the trophoblast pro-
ducing thickenings thereof leading to the type I called
“ cumulate/’

But there is one point which Hubrecht alludes to only
casually, namely, the part played by the uterine glands in
the function of placentatiou. Maybe he does this purposely,
because he proposes to deny the existence of a placenta to
many forms of that type in which the glands play an all
important part. Tims only forms like man, apes, insectivores,
rodents, have placentas, while Equus, “ Sus, Nycticebus,
Galago, and others” have none!

Attempts at systematic arrangements based on placental
characters having never been very successful up to now,
there is no objection to this somewhat radical change in our
conception.” But surely this makes confusion worse con-
founded ! Therefore the sheep is a placental, the cow is a
non-placental mammal 1

We may not be able to agree over terms like “deciduous”
and “ non-deciduous,” “ voll placenta” and “ halb placenta,”
“conjoined” and “apposite,” “cumulate” and “plicate,”
but do not let us quarrel over whether a certain organ of
nutrition or respiration formed in various animals from
identically the same morphological units, appearing at the
same time in all, having the same general function in all,
disappearing at the same period in all, is to be called by
different names according to the mode in which it takes in
its nourishment.

Such a suggestion reminds one of Gideon’s method of
selecting men by the way in which they drank from the
Spring of Harod. An excellent method of determining the
particular merits of the men for certain purposes, but those
individuals did not cease to be men who lapped like a dog.

EARLY OXTOGEXETIO RHEXUMEXA IX MAMMALS. 263

I appeal to Professor Hiibreclit to let us retain the word
“ placenta ” for all types of foetal connection between mother
and offspring for tbe purpose of nutrition^ at any rate within
the class Mammalia.

The size and shape of the uterus^ the character of the sub-
mucosa, the nature of the uterine glands, are, of course, all
features which have exercised profound effects upon the
character of the attachment between mother and offspring,
but I cannot see that they could be compared in morpho-
logical importance to the organ Avhich alone makes them of
any use at all. To say that the laterally expanded tropho-
blast and the distended allantois of the Ungulates^ “ placenta'’
is not an organ equivalent in all respects morphologically and
physiologically with the more compact and confined mass of
trophoblast and allantois of a hedgehog or rat, because the
former obtains its nourishment largely from the glandular
secretions, seems to me to be scarcely tenable.

No doubt the time is not yet i-ipe for using the placenta
even as a broad means of classification, but liiibrecht’s own
work, especially that on Tarsius, has indicated that the
placenta will be found eventually to have most important
diagnostic value.

And I venture to predict that when that time does come the
characters on which the main classification will rest will be
characters of the allantois and trophoblast rather than
maternal features, important though the latter may prove to
be for the smaller subdivisions. As Hubi’echt allows, the
young blastocyst at any rate is parasitic in its relation to the
mother ; and is it not customary to classify parasites more by
their own morphological and physiological features than by
the effects which they have upon their hosts?

Although I did not suggest the terms “ cumulate ” and
“ plicate” as terms for a classification of mammalia on these
lines, still for the present these terms seem to me to represent
the essential character of two extreme forms of mammalian
placenta. The characters of the two types may be tabulated
thus :

264

i; I CHARD ASSIIETOX.

Cumulate placenta.

Great radial proliferation of tro-
phol^last, leading to thickening
of the membrane.

Greater destruction of maternal
tissue.

Much bleeding of mother.

Lacunisation of trophoblast.

Secretion of uterine glands of less
use or in some cases of no use
to foetus.

Degeneration of uterine epithe-
lium and glands severe.

Embryo or embryos seldom fill
the whole cavity of the uterus.

Plicate placenta.

Great tangential proliferation of
trophoblast leading to folding
of the membrane.

Lesser destruction of maternal
tissue.

Little bleeding of mother.

No lacunisation of trophoblast.

Secretion of uterine glands of
prime importance.

Little or no degeneration of
uterine epithelium and glands.

Embryo or embryos usually fill
the whole cavity of the uterus.

The reason for the occurrence of the latter character being
no doubt that fixation is brought about by imbedding or other
iutirnate connection in the cumulate type, while fixation in the
plicate type depends more upon the internal hydrostatic
pressure of the blastocyst up against the uterus — much as a
pneumatic tyre retains its position in the iron rim of a bicycle
wheel.

All those are essential characters and are nearly all due to
the nature of the trophoblast and allantois, i. e. of embryo
rather than mother.

Modifications, especially within an order, maybe due to the
maternal influences. Thus, whether the embryo becomes
wholly imbedded as in the rat, with typical cumulate character,
or approaches the plicate type as in the rabbit (though the
trophoblast undoubtedly shows its intrinsic cumulate charac-
ter in the beginning) depends upon absence of the albumen
layer in the former case allowing the embryo to be caught
ill the narrow chinks of the uterine wall and to become
quickly parasitic, or presence of the albumen layer in the latter
which compels the embryo to remain free for a much longer
time before it can become parasitic.

Or as another instance may be mentioned the presence of
certain areas — Hubrecht’s trophospongia as in Tupaja, or the
cotyledonary burrs of Ruminants which determine the spots

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 265

where either actual phagocytic attack may be made — Tupaja,
Ovis, or only a mutual interdigitation may take place — Bos,
Cervus, the intervening' areas remaining free.

It has occurred to me to wonder whether the copious flow
of uterine gland solution in, for instance, the iutercotyle-
donary areas of Ruminants or the general surface of pigs,
horses, may not obviate the necessity, so to speak, of the
trophoblast eating into the maternal tissues by giving it an
abundant supply of nutritive material.

I entirely agree with Hubrecht in regarding the Carnivora
as a central group' with respect to placentation from which
either cumulate or plicate type could be evolved, or as con-
necting the two extreme types. Whether the carnivora, or
the extreme cumulative or extreme plicate is the most primi-
tive, it is very difficult to say. Hubrecht no doubt makes
out a strong case for the cumulate type, except that it is
based upon the non-sauropsidau origin of mammals, which
seems to me untenable and unnecessary.

As strong a case might be made out by those who believe
in the meroblastic egg and the sauropsidan origin of mammals
for the other view. But, granted that the trophoblast is of
yolk-cell or hypoblastic origin, I can see little difficulty in
deriving either the cumulate or plicate type from the saurop-
sidan meroblastic egg.

Amnion.

One part of special interest is the discussion as to the
origin of the amnion in general and in mammals in particular ;
and the question as to whether the present condition of the
Eutherian developm.ent is to be derived from conditions which
had their origin in a large meroblastic egg, or in a small
holoblastic type, without the intercalation of a mei’oblastic
stage. Hubrecht states his conclusions with great perspi-
cacity in the second footnote to p. 79.

It seems to me that there is no necessity to give up the
idea of mammals having been descended from animals with a

266

RrCIIAlfl) ASSHETON.

meroblastic egg, because certain things point to the proba-
bility of the type of Eutherian development such as we see
in Insectivora, Cheiroptera and Primates (Anthropoidea),
with cumulate placentation, being primitive as regards
Euther ia.

If we accept Hubrecht’s hypothesis that the types just
mentioned are primitive as regards the Eutheria, Ave may
nevertheless derive this from the large-yolked egg on the
supposition that the trophoblast represents the reduced yolk-
mass which has temporarily overflowed the inert epiblast, and
yet retain the meroblastic type of egg for its pre-mammalian
ancestry.

The only difficulty to which this leads one is that the
amnion, such as we find it in Eutherian types like rabbit or
sheep or pig, is not strictly homologous to the amnion of the
Sauropsida. That an amnion so like as that of a sheep and
that of a bird should be not absolutely homologous may seem
to some improbable. Jenkinson (’00) objected to this view
on a previous occasion. To whom I Avould answer thus —

In one sense all the amnions of the Amniota are homo-
logous. There can have been no discontinuity in the presence
of an amnion. The one kind cannot have ceased to exist
until the other had come into existence. It is not a case of
homoplasy only. But I submit that the amnion of the rabbit
is not more homologous to the amnion of the sauropsidan
than the horny teeth of Ornithorhynchus are homologous to
the true teeth of the mammal or reptile, Avhich they have
supplanted.

A similar difficulty is with us already, within the small
group of Eutherian mammals, and must be faced ; for the
amnion of the Eutherians, like the sheep, can hardly be
strictly homologous to the amnion of Cavia, because whereas
the former is formed partly of cells of the epiblast plate of
the embryonal area and partly of trophoblast, the latter is
formed wholly of cells of the embryonal epiblast.

In view of the fact of the curving of the epiblastic plate
noticed in so many cases — Sus, Tupaja, Talpa, Vespertilio,

EART.y OXTOOtEXETLO PJIEXOMEXA IX MAMMALS. 267

Cervus, OviSj etc. — and particularly so in the case of the bat,
to which Hubrecht draws especial attention, the idea that the
formation of the amnion and the incurving of the epiblast of
the embryonal area in Eutherian mammals may have had
their origin in the overgrowth of the yolk by the epiblast
after the manner of Sauropsidans as suggested by me (1908,
p. 252 ; ref. Hubi'echt, p. 23, line 19) seems to me still to have
some value, although no doubt at the present time the reten-
tion of the curvature and the particular form of amnion
formation in some mammals and not in others depends on
various differences in present-time conditions.

This means that the rabbit, sheep, or dog type of amnion
formation has been derived from a type like that seen in
Cavia, whose ancestors had the sauropsidan mode of amnion
formation.

The trace of such a history is perhaps to bo seen in the
blastocyst of Capra (Asshetou, 1908).

Allantois.

I take it that the object of Hubrecht’s suggestions as
regards the Allantois is the removal of the difficulty of
imagining how the Allantois could have acquired its respira-
tory or nutrient functions in the first place, and how it can
have become attached to the somatopleure during early
embryonic life in the second place, in other words, how to
bridge over the gap between Anamnia and Amniota.
Probably all agree that the Allantois as an organ of respira-
tion, or respiration and nutrition, can only have come into
being in connection with the presence of an amnion.

Hubi’echt derives the amnion and trophoblast from an epi-
blastic larval envelope present before and when the ancestral
Amniote became viviparous, and as yet having small, yolk-
less eggs. He discards the idea that the uriuary bladder
of the Amphibian could grow out like the allantois of an
Amniote, and suggests that the allantois is the resulting
effect of an original connection between the splanchnic meso-

268

EICHART) ASSHETON.

blast o£ the g'ut and the larval envelope. I ara not quite sure
whether I understand his theory, but it seems that it must
imply a connection having been made with the larval envelope
by way of the lips ' of the blastopore, and effected at an early
stage of the growth in length, so that the allantois and its
vessels represent a very much more anterior part of the gut
than the urinary bladder of Amphibia, and that the hind gut
arises as a dorsal diverticulum of the allantois. Or are we to
believe that the hind gut of Amniotes is something new and
that the anus of Amniotes is not homologous to the anus of
Anamnia?

Chapter V.

The Gnal chapter on the placenta is full of interest and
less controversial than the rest of this comprehensive and
remarkable treatise. Probably, however, most zoologists
w'ill be sorry that Hubrecht has not attempted to deal more
thoroughly with the question of the value of tbe placenta as
a guide to classification, though no one who has studied
mammalian placentation wdll be surprised at his decision to
postpone this most difficult subject. I feel, however, quite
sure that Hubrecht must believe that the testimony is there
to a large extent, and one will look forward to his exposition
of it at some not distant date.

I entirely agree with Hubrecht’s opinion that the diffuse
placentation of the Lemurs is different from that of the true
plicate forms of Ungulates, Cetacea, some Edentates, etc.

In my paper of 1906“ I did,indeed, mention the Prosimiawith
much hesitation (vide PI. 13) in connection with the plicate
forms, but I felt then — and since having examined a speci-
men kindly sent me from the Zoological Gardens by Mr. F. E.
Beddard, F.R.S., I have felt more strongly — that the condition
of Nycticebus may possibly have been derived from that of
‘ I do not know, liowever, wliat in Hubrecht's opinion the relation
betweeir the larval envelope and the blastopore lips may have been.

^ ‘ Phil. Trans. Roy. Soc. Lond.,’ vol. clxviii.

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 269

a strictly cumulate type by way of such couditions as Hylo-
bates, Semnopithecus, Cercocebus, by the gradual supersession
of the glandular activity of the maternal uterus over the
phagocytic activity of the foetal ti’ophoblast, and the filling
of the blood-spaces, into which the foetal villi originally hung,
with uterine secretions instead of extravasated maternal
blood.

The character of the foetal villi and maternal crypts ai-e
somewhat different in Nycticebus from those of plicate forms.
In the plicate forms each villus fits into its own special crypt
very exactly ; the correspondence is, indeed, extremely inti-
mate in some cases, e. g. the pig, where the processes of the
trophoblast cells even penetrate between the cells of the
uterine epithelium.

In my specimen of Nycticebus the foetal villi appear to
hang in grape-like bunches into the mouths of much wider
depressions, but this character may be artificially exaggerated
in my specimen, as neither Hubrecht for Nycticebus nor
Strahl for Galago indicate this character to the same extent.

Nevertheless, the difference between the true plicate form
and Nycticebus is very well marked. In such plicate forms
as Equus, Bos, Tragulus, Orca, Halicore, the foetal villi are
filose, and fit into distinct crypts in the uterine mucosa, while
in Nycticebus the foetal villi are more lobose and racemose,
and can be described better as partially separated from one
another by lamellate folds of the maternal mucosa, being
elsewhere separated only by uterine gland secretion. The
peculiar chorionic recesses described by Hubrecht (’94) are
recesses where foetal villi project quite freely into a space
filled presumably with uterine milk, as the recesses are open
to the inter-foetal uterine space and certainly contain no
projection of the uterine epithelium itself. Hubrecht says
(p. 145) : “ Nycticebus has foetal investments which, in the
latter half of the period of pregnancy, can, together with the
enclosed foetus, be easily washed out of the maternal crypts,
the trophoblastic villi not being in any way confluent with
maternal tissue.”

VOL. 54, PAKT 2. NEW SERIES.

19

270

RICHAED ASSHETON.

There is, however, no necessity to believe that the real
plicate type has been derived from the extreme cumulate
type, as is suggested for Nycticebus, nor even of the extreme
cumulate from the extreme plicate. A middle view may be
safer, namely, to regard the condition of the carnivora as
that from which the two extremes have evolved, and that the
Lemurs by some such process as that suggested above have
given rise to a secondary condition from the more cumulate
type, and simulate roughly tlie plicate type. The earliest
stages of a Lemur have not yet been described. If there is
anything in the suggestion above, the earliest formed
trophoblast of tlie Lemurs like Nycticebus ought to show
“ cumulations.”

I am entirely at one with the Professor when he recalls the
great value of his discovery of the nature of the placenta of
Tarsius, and not only the placenta, but the character of the
blastocyst, the coelom, mesoderm, and allantois, which so
closely resemble those of juan that they simply cannot be
ignored if comparative anatomy is of any value at all as
evidence of racial affinity. But I think it is not necessary to
take so strong a view as Hubrecht does of the difference
between Tarsius and the other Lemurs in these respects, and
I have indicated a way in which the Lemurine may have
been derived from the Tarsian condition. The condition
seems to me to emphasise the affinity between Lemurs and
Monkeys rather than to necessitate the complete separation
of Tarsius from the Lemurs. So far as it goes the com-
parison supports the view of Dr. Elliot Smith expressed in
his letter in ‘Nature,’ vol. Ixxx, No. 2054. It is, however, a
condition (juite sufficient to justify a grouping of the Lemurs
into those with the pseudo-plicate “ Lemurine ” placenta and
free allantois on the one hand and Tarsius with the cumulate
placenta and early allantoic attachment on the other hand,
the latter group being closer to the Monke}’s than the former,
as, indeed, has been shown by other characters.

EARLY ONTOGENETR! PHENOMENA IN MAMMALS. 271

Chapter VI.

Lastly, Hubrecht wishes to revise the classification of
vertebrates in the light of this supposed homology of all
outer trophic or protective layers within the phylum.

He says (p. 81) : “And it then, of course, strikes us, sup-
posing all these different outer covering layers during early
larval life to be the remnants of an early larval envelope, of
which we find no trace in Amphioxus, the cyclostomes, and
the Elasmobranchs, that the deep significance hitherto
attached to the foetal membranes as a means of sub-dividing-
the Vertebrates into the primary groups of Amniota and
Anamnia runs great risk of losing much of its significance.”

So Hubrecht, leaving out of consideration the Hemi-
chordata and Urochordata, divides the higher chordates into
four “ super-classes,” namely Cephalochordata (for Amphi-
oxus), Cyclostomata (for the Marsipobranchs), Chondrophora
(for the Elasmobranchs), and Osteophora (for the Amphi-
bians, Dipnoans, Ganoids, Teleosteans, Reptiles, Birds, and
Mammals).

Seeing that the bond of union for these last seven groups
is the supposed presence of a larval envelope, one wonders
that Hubrecht has not coined a word suitable to this pheno-
menon instead of falling back upon the presence and absence
of bone. Does the neglect suggest a lack of faith ?

On p. 17 Hubi-echt writes: “A tendency to exchange the
radial for a bilateral symmetry and to separate the coelom
from the enteron must at one time have characterised certain
coclenterate ancestral forms, as has already been advocated
by Sedgwick (’84) and by myself (’05) on eai-lier occasions.
It is not straining the imagination to assume that in this line
of descent closely-related forms may have developed, some
with, others without a larval envelope, temporarily ensheath-
ing the cellular elements that will build up the embryo itself,
and thus foreshadowing the separation among their later
vertebrate descendants of such with and such othei-s without
a trophoblast.”

272

BICHAHD ASSHETON.

I think it is clear from this paragraph that Hiibrecht
suggests that the separation into the great groups Osteophora
with larval envelope on the one hand, and Chondrophora,
Cyclostomata, and Cephalochordata without larval envelope
on the other hand, dates back to coelenterate days, or at any
rate to his vermactinian ancestral phase.

How, then, are we to regard such similarities as the pineal
eye of Sphenodon and the pineal eye of Geotria ; or the
first ten pairs of cranial nerves aiad their distribution in an
Amniote with the ten pairs of cranial nerves of Scyllium ; or,
in fact, any of the hitherto supposed homologous parts which
are to be regarded as comparatively speaking recent acquire-
ments within the phylum Chordata ? Are we not forced to ask
ourselves. Which of three alternatives are we to accept
henceforth ?

(1) That the anatomical resemblances such as those referred
to above are not true homologies, but are accidental resem-
blances.

(2) That tlie coelenterate ancestor “ with a tendency to
become bilaterally symmetrical,” but not at that time having
any orgaus at all visibly like the brain, sense organs, kidneys,
skeleton, etc., of the Lamprey, Elasmobranch or Amniote,
nevertheless was so constituted internally that its descendants,
although separating along different lines of descent, as shown
by the presence and absence of “ trophoblast,” attained
closely similar results in spite of the divers effects that
natural selection is known to have had in other divergent
lines, the Ascidians and Enteropneusts.

(3) That Hubrecht’s attempt to homologise all such super-
ficial layers as the trophoblast of Eutheria, 'Geloderm ” of
Sauropsida, the Deckschicht of Amphibia, of Teleostomes,
etc., fails.

Nor does Hubrecht seem to me to meet the objections
urged by van Beneden, Gadow and others, namely, the many
characters which unite the three groups forming the Amniota
and marking them off from the rest of the Vertebrates. I
may here mention a few and mostly embryological points

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 273

Amniota.

Amnion.

Allantois as respiratory organ.

Umbilical reside or yolk-sac pos-
terior to and qnite separate from
liver.

No true blastopore. If any open-
ing occurs to which this term
has been applied it never occurs
until after the first formation
of the future gut cavity, i . e . it
is a neurenteric canal.

Spiiticle always present and open
as long as any other cleft.

Peculiar epidermic skeleton of ho-
mologous parts (scales, feathers,
hairs).

Vertebral column gastrocentrous
form of Arcocentrous tyjDe.

Ribs well developed and reaching
sternum (except speciahsed
types, e.g. Chelonia, Ophidia).

Hypoglossal comes off within the
skuU.

Neck.

Metanephros in connection with
a special outgi’owth from the
mesonephric duct to form the
ureter.

Mesonephric duct never conveys
urine in adult.

Pronephros of embryo rudimen-

No amnion.

No respiratory allantois.

Yolk-sac either anterior or in con-
nection with liver.

Blastopore ; always correlated
with the fii’st formation of the
gut cavity.

(Gyninophiona may be inter-
mediate.)

Spiracle often rudimentary or
absent.

Nothing comparable.

V ertebral column notocentrous
and pseudocentrous forms of
Arcocentrous type.

No ribs, or if present they do not
reach the sternum.

Hypoglossal always outside the
skull.

No neck.

No metanephros nor special out-
growth from the mesonephric
duct.

Mesonephric duct itself is ureter.

Pronephros of embryo weU de-
veloped.

taiy.

The only other point to which I wnll allude is Hubrecht’s
question, “ Whether many of our Dipnoi, Ganoids, and
Teleosts may not have had terrestrial ancestors,’’ p. 153.

Naturally, I feel proud that I should have been bold
enough to make a similar suggestion for the Teleosteaus in

which are probably of prime importance (though some of
these are already given in other authors’ lists, e.g. Gadow, H.,
‘ Zeits. f. Morph, u. Phys.,’ Bd. 4).

Anamnia.

274

RICHAHD ASSHETON.

my Grymnarchus paper in the ‘ Budgett Memorial Volume ’ ;
and although it is doubtless a “ wild and hypothetical specu-
lation,” yet it seems to me to be au idea which deserves
consideration by both embryologists and palaeontologists. It
is curious, however, that the reasons which led me to the
idea are quite different from those which brought Professor
Ilubrecht to the same conclusion (' Budgett Mem. Vol.,’ p.
407).

And although I was well aware of, and alluded to, the
close resemblance between the trophobhist of Eutherian
mammals and the Deckschicht of Teleosteans (“Report of
Eggs and Larvae from the Glambia River,” ‘ Budgett Mem.
Vol.,' pp. 434-6), it never struck me to regard this as
evidence of affinity between the groups ; and I cannot yet
get over the difficulties of regarding these and the other
outer layers discussed in the preceding pages as homologous,
for the difficulties apiDear to me insurmountable.

In conclusion, may I again express my appreciation and
admiration of the great work upon which I have ventured
to comment. And if, in my excursus, I have settled more
often and dwelt longer upon points where I have found
myself in opposition to the Professor, it has been in the
belief that such disputations tend towards the elucidation of
the truth, and not because I am in any way insensible of the
magnitude of the work, nor because I am not to a large
extent in accord with the conclusions based upon it ; for no
one can realise more clearly than I do how much of the
intei’est, which we who have the more recently been drawn
to this field of research in mammalian embryology feel in
the subject, is due to the brilliant researches and writings of
Ilubrecht.

List of Papers referred to in the Text.

Assheton, R. (’91‘). — “ A Re-investigation into the Early Stages of the
Development of the Rabbit,” ‘ Quart. Journ. Micr. Sci.,’ vol. 37.

(’94-).—“ On the Phenomenon of the Fusion of the Epiblastic

Layers in the Rabbit and in the Frog,” ‘ Quart. Journ. Micr. Sci.,'
vol. 37.

EARLY ONTOGENETIC PHENOMENA IN MAMMALS. 275

Absheton, R. ('94'^). — “The Primitive Streak of the Rabbit: the Causes
which may Determine its Shape and the Pai't of the Embryo
Formed by its Activity,” ‘ Quart. Journ. Micr. Sci.,’ vol. 37.

('94-'). — “ On the Growth in Length of the Frog Embryo,”

‘ Quart. Journ. Micr. Sci.,’ vol. 37.

('96). — “ On Experimental Examination into the Growth of the

Blastoderm of the Chick,” ‘ Proc. Roy. Soc.,’ vol. lx.

(’98'). — “ The Segmentation of the Ovum of the Sheep, with

Observations on the Hypothesis of a Hypoblastic Origin for the
Trophoblast.” ‘ Quart. Journ. Micr. Sci.,’ vol. 41.

(’98'2). — “ The Development of the Pig during the First Ten

Days,” ‘ Quart. Jouni. Micr. Sci.,’ vol. 41.

('98^). — “ An Account of a Blastodermic Vesicle of the Sheep of

the Seventh Day with Twin Germinal Areas,” ‘ Journ. Anat. and
Phys.,’ 1898.

('05). — “ On Gi'owth Centres in Vertebiaite Embryos,” ‘Anato-

mischen Anzeiger,’ vol. xxvii.

('06).— “ The Morphology of the Ungulate Placenta, Particularly

the Develojniient of that Organ in the Sheep, with Notes on the
Placenta of Hyrax and the Elephant,” ‘ Phil. Trans. Roy. Soc.,’
Series B, vol. cxcviii.

( 07). — “ Certain Features Characteristic of Teleostean Develop-
ment,'’ ‘ Guy’s Hospital Reports,’ vol. Ixi.

( 08). — “The Development of Gymnarchus niloticiis,” ‘The

Budgett Memorial Volume,’ University Press, Cambridge.

( 08). — “ Report \ipon Sundry Teleostean Eggs and Larvae from

the Gambia River,” “The Budgett Memorial Volume,’ University
Press, Cambridge.

(08). — “Notes on the Blastocyst of Capi’a,’’ ‘Guy's Hospital

Rei^orts,' vol. Ixii.

Van Beneden, E. ('80). — ‘Recherches sur I'embryologie du lapin,” ‘Arch,
de Biol.,’ vol. i.

(’99). — “ Recherches sur les premiers stades du developpement

du Murin (Vespertilio murinus),” ‘ Anat. Anz.,’ vol xvi.

et Julin ('80). — “Observations sur la Maturation, etc., de I'ceuf

chez les Chiropteres,” ‘ Arch, de Biol.,’ vol. i.

Bischoff (’42). — “ Entwickelungsgeschichte des Kaninchenies,” Braun-
schweig.

(’45). — “ Die Entwickelungsgeschichte des Hundeies,” Braun-
schweig.

270

EICHAKD ASSHETOX.

Bisclioff {'52). — Eutwickelimgsgeschiclite des Meerscliweincliens,'’
Giessen.

('54). — “ Entwickelungsgeschichte des Relies," Giessen.

Duval, M. ("87). — " Atlas d'Embiyologie,” Paris.

('99)- — ‘‘Etudes sur rEnibryogonie des Clieiropteres,” ‘ Journ.

de I’Aiiat. et Phys.,’ vols. 31, 32.

Gadow, H.

‘ Zeit. f. Morph, u. Phys.,’ vol. iv.

Heape, W. (’83). — “ The Development of the Mole,” ‘ Quart. Journ.
Micr. Sci.,’ vol. 23.

Hertwig, 0. ('06). — “Handbuch,” Jena.

Hill, J. P. ('08). — “ Report by Dr. Ashworth on Zoology at Brit. Assoc.

Meeting Dublin,” ‘ Nature,’ vol. lx.Kviii.

Hubrecht, A. A. W. ('90). — “Studies in Mammalian Embryology; II.
The DeveloiJinent of the Germinal Layers of Sorex vulgaris,”
‘ Quart. Journ. Micr. Sci.,’ vol. 31.

('94). — “ Spolia nemoris,” ‘Quart. Jouni. Micr. Sci.,’ vol. 36.

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{'99). — “ Ueber die Entvvickelung des Placenta von Tarsius mid

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( 02). — “ Furchung und Keimblatt - bildung bei Tarsius

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(05). — “ Gastrulation of Vertebrates,” ‘Quart. Journ. Micr.

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( 08). — “ Early Ontogenetic Phenomena in Mammals and their

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VOI,. o4, PART 2. — NRW SERIES.

20

THE EORMATIOX OP THE LAYERS IX AMPHIOXHS. 279

The Formation of the Layers in Amphioxus and
its Bearing on the Interpretation of the Early
Ontogenetic Processes in Other Vertebrates.

By

E. W. MacBiide, D.Sr., LE.D., F.R.S.,

Strathcona Professor of Zoology in McGill University, MonUeal.

With Plates 18, 19, 20, and 21. and 10 Text-figures.

The reader.s of the Quarterly .Journal of Micro-
scopical Science have been treated in a recent number to
a brilliant essay on the early ontogenetic stages of vertebrates
from the well-known pen of Professor Hubrecht (18), who
has dealt with the subject from the point of view of a
specialist in mammalian embryology, and may be said to
have envisaged the development of vertebrates through
mammalian spectacles.

To every question there are at least two points of view,
and I desire in what follows to lay before the readers of this
journal another way of looking at the early development of
Vertebrata, taking as my starting-point, not the highest
members of the group, as Professor Hubrecht has done, but
the lowest form which all zoologists are agreed in including
within the phylum Vertebrata, viz. Amphioxus. Which of
the two points of view will ultimately gain the greatest
amount of support will depend, I am convinced, upon which
offers the simplest and most natural phylogenetic explana-
tion of the facts to be interpreted.

The first part of this paper will therefore consist of a
re-description of the formation of the layers in Amphioxus.
I say a re-description, because eleven years ago I g’ave a

VOL. 54, PART 3. NEW SERIES. 21

280

E. W. MAC BK IDE.

description of tlie early stnges of the development of this
animal, based on hundreds of series of sections througdi well-
preserved material (26). The conclusions arrived at in that
paper, however, have been combated by several workers
since, and as the figures illustrating it were hastily executed
and somewhat schematic, owing to my impending departure
from England to take up work at McGill, some authors have
taken for granted that the material was badly preserved and
that my observations were therefore of little value. Such a
conclusion is entirely unwarranted, as the figures illustrating
the present paper will, I hope, convince every unprejudiced
observer. I have endeavoured, so far as my powers of draughts-
manship extend, to depict the cellular structure in minute
detail, and I find, on going over the subject ag-ain after eleven
years’ interval, that I must re-affirm all the statements I made
in 1898 with a few trifling exceptions in minor points.

Part I.

The Formation of Layers in Amfhioxus.

Pefoi'e, however, proceeding to the description of the
development of Am phioxus, it is perhaps desirable to give a
brief review of former work on the subject so that the ques-
tions at issue may be clearly grasped. Kowalevsky was the
first who published a paper (21) on the development of
Amphioxus, which he followed up some years later by a
second paper on the same subject (22). In his first paper he
describes the egg as dividing into blastomeres of nearly
equal size, forming a hollow blastula. This blastula, according-
to him, is converted into a gastrula by a flattening of one
pole, followed by an invagination of the flattened area pro-
ceeding equally at every point of its circumference, so that
the blastopore is left as a symmetrically situated posterior
pore. Ill his second paper Kowalevsky describes the forma-
tion of “ folds ” of the inner layer (hypoblast) situated
dorso-laterally. From the context it is clear that by “folds”
he means pouch-like outgrowths. Ke makes the significant

THE FOEMATION OF THE LAYERS IN AMPHIOXUS. 281

statement tliat the cavities of the first pair of these folds ”
remain for a long time widely open to the gut, whilst the
cavities of the others have very narrow communications with
the gut and are soon cut off entirely from it. Further, he
states that whilst the first pair of folds become converted into
hollow epithelial vesicles, the cavities of the other pairs
become quite obliterated. He was not able to trace these
folds into the formation of the body-cavity of the adult, but
he believed that a genetic connection existed between the
two sets of organs.

After an interval of some years Kowalevsky was followed by
Hatschek, who, in a brilliant paper, gave the account of the
development of Amphioxus which has been incorporated in
all the text-books of zoology (15). Hatschek pointed out that
in the formation of the blastula the blastomeres at one pole
are larger that at the other, and that it is the area consisting
of the larger blastomeres which is invaginated when the
gastrula is formed. He observed further that when gastrula-
tion is complete the blastopore was situated at the posterior
end of the dorsal surface. He drew the conclusion that in
the early stages of gastrulation, when there is a wide-open
blastopore, this is directed towards the future dorsal surface
of the animal, and that it becomes reduced in size by the
meeting of its edges, a process which he supposed to take
place from the fi'ont backwards. Hatschek never asserted
that he had seen this union or “concrescence” of the lateral
edges of the blastopore ; and it is hard to see why, if such
a process really took place, some trace of it, in the form of a
seam or raphe, should not be observed. Once the gastrulation
has been completed the processes of formation of the noto-
chord and of the mesoderm are begun. These processes,
according to Hatschek, consist in the production of three
longitudinal folds of the dorsal wall of the gut, of which the
median forms the notochord and the lateral the mesoderm.
These “folds” are genuine longitudinal hollow ridges, not
“folds” in the sense in which the word is employed by
Kowalevsky. It is important to note this because many who

282

E. W. MACBRTDE.

have referred to or transcribed Hatschek’s work appear to
imagine that he described the mesoderm as originating as a
series of pairs of independent pouches, which is not the case.
According to Hatschek the lateral folds, whilst still in open
communication with tlie gut, are divided by constrictions
into four or five “ somites ” on each side, of Avhich, however,
only the first pair are completely separated from the rest,
the others constituting a single moniliform fold on each
side. A little later the front part of this moniliform fold
becomes completely separated from the gut, and its consti-
tuent somites then become completely separated from one
another, the last only retaining its communication with the gut.
As new somites are formed the animal grows in length and
the primary fold is prolonged backwards; from this fold these
somites are cut off, and only the last pair at any moment is in
open communication with the gut. The first pair of somites,
however, which were early separated from the rest, long
retain their communications with the gut. Each of the somites
undergoes segnnentation into a dorsal portion which consti-
tutes the myotome, and a ventral portion, which fuses with
its predecessor and successor to form the splanchnoccele, the
right and left splanchnoceles fusing to form the general
abdominal cavity of the adult. From the most anterior part
of the gut two diverticula grow out which become cut off
from it; these are termed by Hatschek “ head-cavities”; of
these the left becomes a small thick-walled vesicle, which sub-
sequently acquires an opening to the exterior, whilst the right
becomes a large thin-walled vesicle which forms the cavity of
the snout in the young Amphioxus. It is evident that most
of Hatschek’s observations are to be regarded as extensions of
Kowalevsky’s results, due to his more perfect method of
technique. The only point in which they radically differ is
in the position they assign to the blastopore, and in this
matter Hatschek frankly acknowledged that his conclusions
were far from certain.

So clear and simple an account as that given by Hatschek
remained for a long time in favour, but in 1894 it was attacked

THE FOEMATION OF THE LAYERS IN AMPHIOXUS. 283

by Basilius Lvvof¥ (25). According to this observer the pro-
cess oP gastrulatioii consists of two parts ; during the first
stage there is invaginated the genuine large-celled endoderm,
consisting of the larger blastoineres situated at the lower pole
of the blastula; from this the epithelium of the gut is formed.
During the second stage of gastrulation the small-celled ecto-
derm is invaginated round the dorsal lip of the blastopore,
and forms the dorsal wall of the “ archenteron ” or gut of the
completed gastrula. This ectodermal plate, however, is com-
pletely used up in the formation of the notochord and of the
mesodermal folds ; the edges of the endodermal portion of the
gut meet beneath it, and so complete the definitive gut. The
cavities of these folds stand in no relation to the future body-
cavity of the adult, for they are produced by the pressure on
the dorsal wall of the gut of the in-sinking nerve-plate, which
is commencing to be invaginated, and their cavities sub-
sequently disappear entirely, whilst the coelom consists of
spaces wliich arise subsequently as splits in solid masses of cells.
It follows that mesoderm and notochord are of ectodermal
origin, and that Am phioxus is in no sense an “enterocoelous”
animal, nor are the processes of the formation of its layers
capable of being analysed into simple processes of folding in
the wall of an epithelial sac. Lwoff further denies that the
blastopore closes in a slit-like manner from before backwards.

Lwoff’s observations seemed to fit into what had been
observed in the development of the higher Vertebrata. He
emphasises this point, and ingenuously confesses that he would
not have felt certain about his account of the development of
Amphioxus if he had not found it confirmed by his observa-
tions on the higher Vertebrata. An account, however, so
radically distinct from that given by Hatschek needed con-
firmation, and accordingly in the year 1898 four papers
appeared on the subject by Sobotta (32), Samassa (30),
Klaatsch (20), and myself (25). Sobotta deals only with the
gastrulation. He denies Lwoff’s assertion of a difference in
histological character between the cells forming the dorsal
wall of the archenteron and those constituting its ventral wall.

284

E. W. MACBRIDE.

Lwoff had based his conclusion that ectoderm cells ai’e inflected
round the dorsal lip of the blastopore on his observation of
numerous mitoses there ; he asserted that they were wanting
elsewhere. On this point Sobotta joins issue with him, and
points out that mitoses occur frequently in all parts of the
archeuteric wall. Sobotta had not been able to accurately
determine the orientation of his embryos, but Samassa, who
wrote in the same year (29), using the method of double
embedding in celloidin and paraffin, was able to determine
this. Samassa’s results in general confirm Sobotta’s, but they
add several most important points. He points out that mitoses
take place at regular intervals in both endoderm and ecto-
derm, so that it is quite possible in one embryo to find no
mitosis at all, and in another of nearly the same size to find
numerous mitoses. Samassa further asserts that the blasto-
pore closes by the synchronous approach of all its sides, and
he rejects utterly the idea that it closes in a longitudinal
seam. With regard to the formation of the mesoderm he
confirms Hatschek’s results; he points out that the longi-
tudinal grooves which give rise to the mesoderm make their
appearance before the nerve-plate becomes invaginated, and
that therefore Lwoff’s explanation of their formation is
groundless. Klaatsch’s paper on the gastrulation (20) contains
several admirable new points. He remarks that the abundance
of mitoses found by Lwoff in the neighbourhood of the dorsal
lip of the blastopore does not prove an inflection of cells
round this lip, but rather proves that here is to be found a
growing point from which new cells are added to both
ectoderm and endoderm. He denies the possibilit}' of an
inflection of cells round the dorsal lip, because of the difference
in histological character between the ectoderm and the cells
forming the doi-sal wall of the archenteron. With regard,
however, to the ventral lip, he infers that here there is a
genuine inflection of cells, because he found outside this lip
large rounded cells, which in later gastrulm were replaced by
small-celled ectoderm. My own paper (26) was published in
ignorance of the results of the three workers whose observa-

THE FOEMATION OF THE LAYEES IN AJIPHIOXUS. 285

tions I liave just summarised. In it I denied that there was
any histological difference between the cells forming the
dorsal and those forming the ventral wall of the archenteron.
I denied, also, that tliere was any inflection of ectodermal
cells round the dorsal lip of the blastopore, but I asserted
that the gastrulation was due, in the first instance, to a rapid
increase of endoderm cells in this neighbourhood, which pro-
duced a lateral strain on the endoderm to which it yielded by
invagination, that the blastopore was directed posteriorly, not
dorsal ly, and that the closing of the blastopore, or, to speak
more correctly, its reduction to a small pore, was due to the
appearance of a new centre of rapid growth in the ventral lip
of the blastopore. I asserted further that the first pair of
somites were independent formations from the gut-wall, which
I compared to the collar-cavities of Balanoglossus, and
that the other somites owed their origin to the segmentation
of a hinder pair of outgrowths from the gut-wall, which 1
compared to the trunk-cavities of Balanoglossus, and
w’hich grew in len gth pari passu with the growth of the
animal, and that only the hindermost as yet unsegmented
portions of these grooves retained openings into the gut. I
further asserted that whilst the other somites divided into
myotonies above and ventral portions below, which fused to
form a continuous splanchiiocele, this was not true of the first
pair (the collar-cavities J. In these the dorsal myotomic por-
tion remained in open communication with the lower thin-
walled portion of the somite. This latter did not fuse with
its successors to aid in forming the splanchnocele, but on the
contrary extended posteriorly outside the splanchnocele form-
ing the basis of the future atrial fold. In a subsequent paper
(27) I corrected a hasty and ill-founded statement made in
the first paper, that in the spherical blastula the blastomeres
were all equal in size, and I showed that the backward
extensions of the atrial cavity became solid, and apparently
furnished the material for the transverse muscle of the atrial
floor.

In the same year that this second paper Avas published, a

286

E. W. MACBRIDE.

paper appeared by Morgan and Hazen (28), illustrated by
beautiful figures. These authors support Lwoff’s statement
that there is a histological difference between the cells
forming the dorsal and those forming the ventral wall of the
archenteron. Their account of the matter is, liowevei’, very
confusing. In one place they speak of “ yolkless cells
being inflected at the dorsal lip of the blastopore; in other
places, however, they represent the difference between the
roof and floor of the archenteron as merely a difference in the
intensity with which stain is taken up, and further assert that
this difference cannot be seen in specimens preserved by
corrosive sublimate. Their figures certainly show an immense
difference between the ectoderm cells and those forming
the roof of the archenteron, whilst the difference between
the floor and the roof of the archenteron on which they lay
such stress is only indicated by the colour of the yolk-
granules. Morgan and Hazen do not accept Lwoti^s view
that the cells forming the roof of the archenteron ai’e ecto-
derm, and they point out that mitoses in the dorsal lip of the
blastopore are no proof of a migration of cells round the lip,
and that, as matter of fact, mitoses are by no means confined
to this lip but occur everywhere in both ectoderm and
endoderm. They assert that in my earlier paper I had
confused the dorsal and ventral lips of the blastopore, and
they find a gradual passage from ectoderm to endoderm cells
at the dorsal lip. After considering all the views which had
np till then been put forward as to the manner in which the
blastopore is closed, they consider the most probable account
of the matter to be that the blastopore is directed dorsally,
and is closed by the advance over it of its anterior lip. Xo
further paper on the development of Amphioxus appeared
till 1906, when Cerfoutaiue published an exhaustive paper on
the subject, which, largely because it was illustrated by many
beautiful figures, has been accepted as decisive by many
zoologists (9). Cerfontaine is bitterly hostile to the con-
clusions reached in my work, and it is principally in order to
see how far his criticisms are justified that I have undertaken

THE FOEHATIOX OF THE LAYERS IN AHPHIOXL'S. 287

a re-examination of the whole subject, the results of which
are presented in this paper. He gives an elaborate account
of the segmentation, confirming Hatschek’s account, and noting
the presence of two circles of lai’ger blastomeres near the
vegetative pole. When the blastula becomes flattened, he
finds that the invagination commences asymmetrically near
what afterwards becomes the dorsal or anterior lip of the
blastopore. In this he entirely confirms my work, wdthout
apparently being aware of it. But he maintains that small-
celled ectoderm is inflected round the dorsal lip and forms
the loof of the archenteron, thus re-aftirming Lwoff’s asser-
tion. He arrives at this conclusion on two grounds : first,
because of the existence of numerous mitoses near this lip (thus
ignoring the criticisms of Samassa, of Morgan and Hazen,
and of Klaatsch), and secondly, because of a histological
difference between the cells forming the roof and those
forming the floor of the archenteron. When we probe his
figures for evidence of this difference, we find that it consists
in the presence in the cells forming the floor of a few larger
and more deeply staining granules of yolk than are found in
the cells forming the roof of the archenteron ! There is,
however, a profound difference between these latter and the
indubitable ectoderm. Cerfontaine also re-aflirms Hatschek’s
surmise that the blastopoi’e narrows in a seam-like fashion
from in front backwards ; but he no more than Hatschek has
observed such a seam, and the reasoning which leads him to
this conclusion is certainly recondite. In the process of the
narrowing of a wide blastopore the lateral lips must come
together; now he shows by measurement that during this
process the dorsal and anterior lip moves backward, in other
words, that the embryo grows longer. Hence he concludes
that this growth of the dorsal lip must be due to the same
process which narrowed the transverse diameter of the blasto-
pore ! Cerfontaine affirms in accord with Klaatsch that
ectoderm is also invaginated at the ventral lip of the blasto-
pore, basing his conclusion on the existence of mitoses here
during the later stages of gastrulation. Verily Cerfontaine’s

i!88

E. W. MACEHIDE.

conclusions are like a pyramid standing on its apex! In
spite of his hostility to me his figures really suppoi’t my
observations when relieved of the forced interpretation,
which his espousal of Lwoff has led him to put on
them. With regard to the formation of the mesoderm
Cerfontaine has little new to contribute. He dismisses my
observations with the remark that he has never seen any-
thing like them in his preparations and that my material was
badly preserved — -a statement he thinks it necessary to
reiteiate three times in his paper, and which is absolutely"
untrue, and especially uncalled for in this case, as Cerfontaine’s
figures of the formation of the mesoderm are as schematic as
those which I ]mblished in my first paper. Cerfontaine,
however, confirms me an-ainst Lwoff in asserting" that
Amphioxus is an “ enterocoelous” animal so far as the
cavities of the first pair of somites are concerned. Legros, who
had previously published a short paper (23) on the larvae of
Amphioxus, in which he had maintained that the epithelial
vesicle, identified by Hatschek and myself with the left head
cavity, was an ectodermal invagination, and that the club-
.shaped gland had no external opening, followed it up in
1907 (24) with a paper on the formation of the layers
in this animal. Legros’s work differs from that of all
his predecessors in that it is based, not on normal, but on
abnormal development. By fertilising ova with stale sperm
and similar means he obtained some curious pathological
embryos. In one of these there Avas a wide posteriorly directed
blastopore, but the mesodermal grooves had been already
formed and from these, two })airs of somites had already been
separated off. There was also in existence a horizontal ridge
on each side of the archenteron, tending to divide it into an
up[)er section to which the ectodermal grooves belonged, and
a lower one which is l egarded by Legros as consisting of true
endoderm. When the series of sections w"as followed back-
wards the horizontal ridges united and shut off the lower
section of the archenteron, which ended in a cul-de-sac, fiom
the up])er section, which opened by the blastopore. In

THE EOEMATIOX OF THE LAYERS IN AJIPHIOXUS. 289

another embryo wliicli lie describes development is mucli
more advanced, and quite a number ol somites have been
formed. Nevertheless, not only is there a wide blastopore
found at the posterior end of the animal, but also the same
ventral diverticulum of the archenteron, and there is further
evidence to be found that the front part of the blastopore
has healed by “concrescence.” Legros regards his observa-
tions as proving that Lvvott and Cerfontaine are right in
supposing that the roof of the archenteron is ectodermic ;
and the horizontal ridges he regards as conveniently marking
the boundaries of the ectodermal and endodermal portions of
this sac; and he naturally regards his older embryo as
])roving that the blastopore closes by concrescence. It must,
however, be remembered that in normal development the
blastopore is reduced to its normal dimensions before any
trace of mesodermal grooves or somites is formed, and the
processes by which a pathological embryo adjusts itself to
the normal condition of affairs are likely to be widely
different from what occurs in the normal course of develop-
ment. This is obvious even from Legros’s figures; for if
concrescence took place normally the tissue destined to form
the notochord would be formed by the meeting in the middle
line of two svmmetrical halves ; now, in Leyros’s later embrvo
the notochord is seen to belong entirely to one lip of the two
which meet in the middle line. Further, in a normal
Amphioxus so soon as the mesodermal grooves have been
formed a great growth in length takes place largely
localised in the hinder part of the embryo ; now, if through
any reason this growth was unequal we should at once get
Legros’s type of embryo without having any light thrown ou
what took place in normal embryos. Legros’s abnormalities
are, therefore, probably not due to disturbances in the pro-
cesses of formation of the layers, but to disturbances in the
process of growth in length which supervenes very early in
Amphio.xus. Legros agrees with Cerfontaine that Amphi-
oxus is truly an enterocoelous animal, for he finds that the first
pair of somites long retain their open connection with the

290

E. W. MACBRIDE.

gut, and he prudently withdraws his earlier statement
that the left head cavity was of ectodermal origin.

Reviewing the observations which have so far been sum-
marised, we see that two questions stand out as pre-eminently
calling for decision : first, is the roof of the archenteron, and
consequently the mesodeian and notochord Avhich arise from
it, ectodermal or endodermal ? And secondly, does the blasto-
pore close by a seam-like concrescence proceeding from in
front backwards or not ? A third question to be resolved is
Avhether the comparison which I instituted in 1898 between
Amphioxus and Bala nogloss us, so far as the formation
of the mesoderm is concerned, can be sustained.

Material and Methods.

The material at my disposal consisted of a profusion of eggs
and embryos covering all stages from the spherical blastula
up to the period when the mouth, club-shaped gland, and one
gill-slit have been formed. There were also a number of older
larvae, but a considerable gap intervened between them and
the larvie with one gill-slit, and in this paper I consider only
the material which formed a series without any gaps. Next,
as to preservation, the material was preserved partly in cor-
rosive sublimate and acetic acid, partly in Kleinenberg’s
picrosulphuric acid, partly in Fleming’s fluid, and partly in
osmic acid. I had material of all stages preserved in cor-
rosive sublimate and in osmic acid, and in the accuracy and
fineness with which histological detail is preserved there is no
other reagent which will stand comjDarison with osmic acid.
This is, of course, a conclusion that is becoming more and
more widely accepted, and some histologists go so far as to
maintain that osmic acid is the only reagent which retains in
dead protoplasm anything like the organisation it possessed
in life. When, however, yolk is extremely abundant osmic
acid makes the material so brittle that section cutting is
almost impracticable ; and for this reason, in dealing with
the stages of gastrulation I have used material preserved in

THE FORMATION OF THE LAYERS IN AMPHIOXUS. 291

picrosulphuric acid and in corrosive sublimate and acetic acid,
but for all later stages osmic acid material alone has been used.

Samassa and I simultaneously employed the method of
double-embedding in celloidin and paraffin, which, indeed, is
the only method of dealing effectively with such minute and
delicate embryos. As modified by me the procedure was as
follows : The celloidin containing the embi’yos after being
congealed in chloroform, was transferred to cedar oil. In
this oil it became as clear as glass, so that the embedded
embryo could be examined under the microscope and its
orientation determined. An appropriately shaped piece of
celloidin was then cut out and embedded in paraffin. The
sections were stained on the slide. My conclusions so far as
the gastrulatiou is concerned are based only on sagittal
sections, the orientation of which was determined as I have
described, but the accuracy of this orientation was tested by
counting the sections in the series and observing whether the
extreme ones at either side were alike in form. This is pos-
sible only where the direction of the sections is accurately
sagittal. The median section in each series was then figured,
and all the figures are drawn with about the same magnifi-
cation, viz. 900-1000 diameters.

The Gastrulation.

AVe shall commence our study of the development of
Amphioxus with the stage of the flattened blastula, for in
this stage the axes of symmetry can for the first time be
determined. A median sagittal section through an embi’yo
of this stage is represented in PI. 1, fig. 1. The outline is
hemispherical, and for reasons which will presently emerge
the flat surface is dii’ected backwards. It will be observed
that the cells on the convex side are somewhat rectangular
in outline and much smaller than the tall columnar cells on
the flat side. In the upper part of the section where the
convex part of the embryo abuts on the flat side there is an
abrupt passage from the rectangular to the columnar type of

292

E. AV. MACBRIDE.

cell. This spot can be recognised in later stnges, and will
be denominated a’. In the embryo under consideration a
cell can be seen at this point whose nucleus exhibits dot-like
aggreg'ations of chromatin, indicative of one of the prophases
of mitosis. Another case of mitosis can be seen in the middle
of the convex surface. When we examine the lower border of
the section at the point marked y in the figure where the
convex surface passes into the flat surface, we find that there
is a gradual transition from the rectangular to the columnar
type of cell. If wm now examine the nuclei we find that in
all the cells they are of a uniform character so far as their
staining properties are concerned. They present a vesicular
appearance, the nuclear sap being crossed wnth cords of liuin
on which are very minute chromatin granules. Turning our
attention now to the grannies of yolk we find that these are
most numerous in the large cells of the flat side and become
fewer as we pass to the cells of the convex side, and that
there appear in these latter cells smaller granules amongst the
larger ones, but that there is no difference whatever either in
size or in staining properties between the largest yolk-granules
in the cells of the convex side of the blastula and those in
the cells on the flat side. In this stage of development, there-
fore, it is impossible to say that the cells of the embryo are
segregated into two definite tissues, as Lwoff and Cerfontaiue
have maintained, for as we have already remarked, and as a
simple inspection of the figure will prove, the cells of the flat
side pass by insensible gradations into those of the convex
side. If with these authors we call the cells of the convex
side ectoderm and those of the flat side eudoderm, it is im-
possible, if we consider the point y, to say where the one
ends and the other begins. At the point x there is an abrupt
change in the size of the cells, and this is manifestly due to
the beginning of a process of division by which the size of
the cells is reduced. The smaller yolk-granules which appear
amongst the larger ones in the cells of the convex side are
due to the utilisation and fragmentation of these owing to
the process of multiplication of cells which has just taken

THE FORMATION OF THK LAAGERS IN AAIPHIOXUS. 293

place, and of wliicli tlie mitosis shown [k) is evidence. A
slightly later stage is shown in PI. 1, fig. 2. In this embryo
the process of invagination has commenced. At the point
we may recognise the same abrupt transition in the size of the
cells which we remarked at the same place in the preceding
stage. Here an nnmistakable mitosis can be seen. The im-
pression conveyed to the mind is that the new cells are added
to the flat side by the process of cell division which has begun
at .T, and that these have exercised a lateral strain to which
the fiat side has yielded by bending inwards. The convex
side has become moi-e convex than in the preceding stage,
and its cells more numerous. The cell-division at x has
therefoi'e probably added cells also to the convex side, but
there is not the slightest evidence that any cell orginall}' on
the conve.x side has passed on to the flat side or vice versa.

In fig. 3 we have a median sagittal section of a still later
stage. AVe again notice a mitosis at the })oint a;. We obtain
further evidence that the cell division which is taking place
at this point does not result in a passage of cells from the
convex side to the invaginated side of the embryo, because
the cells adjacent to this point on the two sides are exceed-
ingly different from one another. Those belonging to the
convex side are smaller than those on the invaginated side,
and have fewer volk-granules. The new cells on the convex
side are remarkalde for their rounded form ; cells of this
shape have been noticed in the neighbourhood of the blasto-
pore by several authors ; the shape seems to be indicative of
the state immediately following division when the
centric force of the new nucleus is at its maximum strength
(v. TheePs beautiful figures of the development of Echino-
cvainus pusillus [33]). At the point // the large clumsy
cells are continued some little distance beyond the lip of the
invagination on to the convex surface.

A still later stage is represented in fig. 4. As compared
with fig. 3, Ave note that the invagination has become deeper,
and that a decided difference in staining property
between the nuclei belonging to the convex side

294

K. W. ]\[ACBRIDE.

and those belonging to the invaginated area is now
observable. The former stain more deeply, since their nuclear
sap has to some extent acquired staining powers. The transi-
tion between lightly staining and more deeply staining nuclei
takes place exactly at x, but at the lower lip of the blasto-
pore the lai’ge vesicular nuclei are continued round for a
short distance on to the convex side of the embryo. I regard
this difference in staining power as the outward sign of
the physiological differentiation of the cells into
ectoderm and endoderm; it is observable all through
the later stages of gastrulation and during the formation of
the mesoderm. I conclude that at the point x we have a
growing centre, comparable to the meristem of a root or the
cambium of a tree with secondary growth, whence oh the
one side young ectoderm, and on the other side young endo-
derm cells are produced. To the appearance of this centre of
growth I attribute the first impulse towards gastrulation, and
the push which causes the in-bending of the lower cells should
on this view be directed not towards the centre of the lower
surface of the flattened blastula, but more towards its dorsal
edge. As a result the invagination does not at first involve
all the cells which will eventually be included within the
archenteron, but the endodermal vesicular nuclei extend
beyond the lower lip of the blastopore at y. A mitosis can
be seen in fig. 4 in the centre of the convex surface, and also
one at the apex of the invaginated area, and they serve to
emphasise the fact that division of cells is not by any means
confined to the growing point x. According to this view the
ectodermal nuclei become differentiated from a type of
nucleus which we may call endodermal, and this is in accord-
ance with what we should expect, for the type of nucleus
found in an assimilative cell must be the primitive one.

In figs. 5a and hh we have two sections from a sagittal
series through an embryo almost of the same age as, or very
little older than, that represented in fig. 4. Fig. 5 a repre-
sents a median, fig. 5 h a more lateral section. In both
figures the sharp contrast between ectodermal and endodermal

THE EOEMATION OF THE LAYERS lA’' AMPHIOXUS.

295

nuclei is seen, but fig. 5 a deserves our special attention
because of the small rounded cells which are seen on both
sides of the point x. As may be clearly seen by reference to
Cerfontaine’s paper, it was on sections like this that that
author based his conclusion that ectodermal cells were being
added to the vault of the archenteron ; we have already sug-
gested that the rounded form is a passing phase following on
mitosis; now our figure shows that the rounded cells which
are added to the arch enteric wall have vesicular
nuclei, Avhilst those added to the ectoderm have
more deeply staining' nuclei; all question, therefore, of
there being any ^‘inflection” of ectoderm cells is settled in
the negative. I have spoken of “ the point ” x when referring
to a median sagittal section of the gastrula ; the growing
point is really, however, a horizontal arc, and appears in
several sections on each side of the median one; fig. 5 h,
which represents a lateral section in which the cells at the
dorsal lip of the blastopore are quiescent, lies four or five
sections to one side of the median one. The ectoderm cells
immediately beyond the growing point in fig. 5 a are seen to
be specially tall and columnar. This is the first indication of
the nerve-plate [n.'p.), and is of the greatest, importance for
the decision as to the orientation of the blastopore. It is also
seen in the older stage represented in fig. 6 («.p.) ; from these
two figures we may conclude that the diameter of the
wide-open blastopore is at right angles to the long
axis of the nerve-plate, and this conclusion seems to me
to settle in the negative the question of the closing of the
blastopore by a concrescence proceeding from in front back-
wards. For such a theor^q whether advanced by Hatschek,
Legros, or Cerfontaine, has always assumed that the nerve-
plate is formed pari passu as the blastopore closes, and this
could only be possible if the long axis of the nerve-plate
and the diameter of the blastopore coincided in
direction. In fig. 6 we can see sevei'al mitoses in what even
Cerfontaine would admit to be the endodermic portion of the
archenteron, and, what is most interesting, one of the cells
VOL. 54, PART 3. — NEW SERIES. 22

296

E. W. MACBRIDE.

undergoing mitosis shows the same rounded form, which,
when occurring at tlie dorsal lip of the blastopore, Cerfontaine
regarded as an infallible proof of ectodermal origin.

In figs. 7 a aud 7 h (PI. 2) a median and a lateral section
of a still older embryo are represented. The nerve-plate is
now unmistakably present, as the flattened dorsal surface of
the embryo shows. The abrupt passage from ectodermal to
endodermal nuclei at the point x is also evident. Now, how-
ever, we may see the extent of the histological difference
between the cells forming the roof and those forming the
floor of the archenteron, on which Lwoff and Cerfontaine lay
so much stress, and which has also been referred to Morgan
aud Hazen. The yolk-granules in the cells of the roof {ch.)
are becoming fewer, aud there are mixed with the large ones
smaller granules, and they do not stain quite as deeply as
those in the cells forming’ the apex or anterior end of the
archenteron. All these appearances are due to one cause ; the
yolk-granules are being more rapidly used up in the cells
forming the roof of the archenteron than elsewhere, and this
in turn can be referred to two causes : first, these cells have
undergone more rapid multiplication than those forming the
rest of the archenteron; and secondly, it is from these cells
that the notochord will eventually be formed, aud the
incipient disappearance of the yolk-granules may be in part an
anticipation of that vacuolisation which distinguishes the
notochordal cells from those forming tlie I’est of the archen-
teron. It is in any case much less observable in the more
lateral section represented in fig. 7 h.

We have now traced the development of the gastrula step
by step from the stage of the flattened blastula up to what we
may term the thimble stage, in which the nerve-plate is
clearly developed and about the orientation of which there
can be no possible doubt, and I consider that this is the best
answer to the charge of Morgan and Hazen that I confounded
the two lips of the blastopore. No such close series of stages
is represented in their paper. The gastrula, whilst still
retaining its wide-open blastopore, has become deeper and

THE FORMATIOX OF THE LAYERS IX AMPHIOXUS. 297

move cylindvical, due iu all probability to a widely distributed
increase of cells both in ectodertn and endoderm. This is the
beginning of that persistent growth in length which
converts the short plump embryo into the comparatively long
attenuated two-da}"-old larva. The axis of the nerve-plate
is clearly at right angles to the diameter of the blastopore,
and how that can be reconciled with a theory of concrescence
of the dorso-lateral lips of that aperture I leave it to advo-
cates of that theory to settle.

The last stage in the gastrulation with which I shall deal
is represented in fig. 8. Here we see the process of closing,
or more strictly speaking of narrowing the blastopore. A
centre of groAvth and multiplication of cells has
now appeared in the v e n t !• a 1 lip of the b 1 a s t o }) o r e,
and in consequence this lip has grown upwards and pro-
trudes as a rounded mass blocking the cavity of the blasto-
pore. We are forcibly reminded of the yolk-plug in
Amphibian embryos ; and at the same time we note that all
the cells with vesicular nuclei are being brought within the
confines of the archenteron. It was the discovery of this
new centre of growth that led Cerfontaine to imagine that in
the later stages of gastrulation ectodermal cells were being
inflected at the ventral lip of the blastopore; it will be seen
also that Klaatsch was perfectly justified in supposing that
cells passed iu at this lip, only the cells that passed in are
endoderm, not ectoderm.

'I’he remnant of the blastopore is eventually found as a
small pore at the posterior end of the nerve-plate, iu which
position it becomes covered in by the meeting above it of the
ectodennal folds which arise at the sides of this plate.

Figs. 9 and 10 represent sections from series cut through
nearly complete gastrulae in a horizontal direction, in order
that some account may be taken of the condition of the
“lateral” lips of the blastopore. Fig. 9 is a section from the
middle of the series cutting the blastopore near the mid-
horizontal line, whilst fig. 10 is a section which cuts the
blastopore near the doi'sal lip. In both we see the sharp

298

E. W. MAOBEIDE.

dusky sliade the new endoderm added hy the growing points. White isorigina
new ectoderm.

THE FORMATION OF THE LAYERS IN AMPHIOXUS. 299

contrast between the ectodermal and endodermal nuclei at
the lips of the blastopore, but in the more ventral section the
endodermal nuclei protrude beyond the lip somewhat, whereas
in the more dorsal they are pushed inwards by the over-
growth of ectoderm. From these figures we may conclude
that the point y like the point x is really an arc of considerable
lateral extent — in fact, of much greater extent than x. We
should probably be justified in dividing the whole circum-
ference of the blastopore into a dorsal and a ventral arc; by
cell division in the former gastrulation is initiated, by cell
division in the latter the blastopore is closed. Text-fig. 1
represents approximately my view of the steps by which
gastrulation proceeds in Amphioxus. It is impossible, of
course, to define the limits of the new endoderm and of the
original endoderm which formed the flat surface of the hemi-
spherical blastula, because, as we have seen, mitoses take
place in the original endoderm and thus it increases in extent.
Tlie text-fig. gives the most probable view of their relative
extent. On account of this cell-division in the original endo-
derm its cells become reduced in size, and at the conclusion
of gastrulation the archenteron is lined by a layer of cells
which are all of approximately equal size.

The Formation of the Notochord and of the Mesoderm.

We have seen that the blastopore becomes covered in by
the meeting above it of two folds of ectoderm, which arise
at the sides of the nerve-plate. These folds first arise in the
hinder part of the embryo and first meet there, and they then
grow gradually forwards, but for some time the anterior part
of the nerve-plate is uncovered. During this time the embryo
is growing in length and pari passu diminishing in diameter.
The only possible explanation of this phenomenon which I
can make to myself is that the yolk-granules are being used
up, and that coincidently the cells are multiplying all over
the animal, but especially towards the hinder end. If we

300

E. W. MACr.KIDE.

now examine a series of transverse sections through an
embi-yo in wliich the diminution in diameter is only sliglit,
and in whicli a good part of the nerve-plate is still uncovered,
we shall see the first stages in the formation of the mesoderm
and the notochord. As my contention that in the formation
of its mesoderm Amphioxus resembles Balan ogloss u s is
based on an examination of this stage, and as Cerfontaine
appears to have completely missed it, I have thought it wise to
figure in detail five sections from such a series. Figs. 11 a, h,
c, d, and e (PI. 2) represent these sections. In fig. 11 a, the
most anterior, we see the nerve-plate consisting as yet of a quite
horizontal platform of columnar ectoderm cells. In the centre
are two clear cells (or), which I suspect are part of the
rudiment of the eye-spot which is found in this position in
later larvm. The archenteron shows at its dorso - lateral
angles two pouch-like outgrowths {coll.). The form of these
pouches and the sharp angle at which their cavities unite
with the cavity of the archenteron negatives any idea that
they could be due to a simple process of folding of the
archenteric wall; they are clearly due to an active process
of proliferation which takes place at this position in the
endoderm. In fig. 11 h, which is only two sections further
back, we see that the nerve-plate is no longer flat but gently
arched inwards, and the beginnings of the two ectodermic
folds (/), which are later to meet above it, are seen at its
sides. In the archenteric wall we observe the same pouches
that we remarked in the more anterior section, but above
them and situated nearer the mid-dorsal line we find another
pair of similar pouches {fr.). In fig. 11c, which is only two
sections further back still, the ectodertnic folds have almost,
but not quite, met above the nerve-plate, and the first and
more ventral pair of pouches of the archenteric wall have
disappeared, but the second or more dorsal pair persist and
continue through half a dozen sections more, always open to
the gut, though this opening is narrower in one place than
it is in front or behind. Now the first or more ventral pair
of pouches give rise to the first pair of somites, whilst the

THE FOEMATION. OP THE LAYERS IX AMPHIOXUS. 301

more posterior and dorsal pouches give rise to all the other
somites. The first pair of somites therefore stand
in a different category to all the rest. Tliis is true
not only of their origin as independent evagina-
tions of the archenteric wall, but as we shall
presently see of all their subsequent history. I
compared these somites in 1898 to the collar cavities of
Balanoglossus, and all my subsequent research has con-
firmed me in this view. The more posterior pair of pouches
will then be equivalent to the trunk cavities of Balano-
glossus, and the varying diameter which these show as we
folloAv them backwards through this series is the first indica-
tion of their approaching division into the posterior somites,
which is the great point in which Amphioxus exhibits an
advance on the condition represented by Balan oglossu s. In
the stage under consideration there seems to be a constriction
of the trunk cavity in one place, b u t throughout its whole
extent it is freely open to the gut. This constriction
is an indication of the constriction of the first of the posterior
somites from the front end of the trunk cavity. In fig. 11 c
we observe in the mid-dorsal line of the archenteron the first
indication of the formation of the notochord (c/i.). The arch-
enteric wall is here arched slightly upwards, and the yolk-
granules are being rapidly reduced in number so that the cells
have become relatively clear. Here we have the explanation
of that difference between the roof and the fioor of the archen-
teron as seen in median sagittal sections, on Avhich Lwoff and
Cerfontaine founded such top-heavy conclusions. Fig. lid
is taken from near the hind end of the embryo. The nerve-
plate is completely covered in by the union of the ectodermic
folds above it, so that we now have a neural canal (n. c.).
The archenteron shows a division into two storeys, exactly as
Legros has described for his abnormal embryos, but which
is here described for the first time in a normal embryo
at least of this age. The lower division (n) is the anal
diverticulum, which at a much later stage will open to the
exterior. The upper division (fd) leads to the still open

302

K. W. MACBEIDE.

blastopore, Avliich lies beneath the cover of the ectodermic
folds and forms a space at the hinder end of the animal into
which the neural canal also opens, viz. the neurenteric
canal. This is shown in fig. He {ne.) two sections further
back in the series. In fig. 11 cZ Ave see on the mid-dorsal Avail
of the archenterou a band of cells undergoing mitosis.
According to Legros and Cerfontaine the notochord, and the
nerve-plate too for that matter, are formed by the union in
the middle-line of two moieties. There is cei-tainly no trace
of such a process here for the band of dividing cells is con-
tinuous across the middle-line. Of course iu every structure
there is an imaginary middle-line, and if anyone chooses to
say that this baud of dividing cells consists of right and left
halves Avhich unite together as quickly as they groAv, I shall
not Avaste time in arguing against such a metaphysical con-
ception, Avhich is capable neither of pi*oot‘ nor disproof.
Legros further asserts that the horizontal ridges Avhich he
finds iu the sides of the archenterou in his abnormal embryo
mark the limits of the ectodermic and endodermic portions
of this structure. A glance at fig. He Avill show the futility
of such a conception ; the histological character of the cells
forming the upper and the lower portions of the archenterou
is identical, Avhereas the place Avhere the ectoderm abuts on
the tissue of the archenteric Avail is clearly seen. But, indeed,
Legros’s OAvn figures, schematic as they are, refute this con-
tention, for they shoAV the genuine ectoderm to be Avidely
different from the so-called ectoderm forming the roof of the
archenterou. About this stage or a little later the embryo
escapes from the egg-membrane and becomes a free-SAvimmiug
larva, propelling itself by its ciliated ectoderm. In figs. 12 a,
h, c, and d (PI. 3) four sections through a someAvhat older
embryo are represented, and its smaller diameter, when com-
pared Avith the embryo Avhich Ave have just considered, should
be noted (vide the magnifications given in the description
of the plates). The union of the ectodermic folds above the
nerve-plate has proceeded forward, and is seen to be complete
in the most anterior section. In this section Ave see the tAvo

THE FORMATION OF THE LAYERS IN AMPHIOXUS. 303

collar-cavities still in open communication Avith. the gut, and
the beginning of the notochord is indicated between them,
showing that the formation of this structure has also pro-
ceeded in an anterior direction. In fig. 12 b on the left side
the collar-cavity is seen to be closed off from the cavity of
the archenteron, and lying beside it nearer the middle line
is the trunk-cavity equally shut off from the archentei’on.
In fig. 12 c the dying out of the collar-cavity can be seen on
the left side, whilst on the right side the collar-cavity can
be seen to be closed off from the archenteron. Finally in
fig. 12 tZ, which is taken from considerably further back in
the series, the hinder portions of the trunk-cavities or trunk-
folds can be seen opening freely into the archenteron. It
follows that the anterior portions of the collar-cavities and
the ])osterior portions of the trunk-cavities retain their
openings into the archenteron. The trunk-cavity has already
had two somites cut off from it. It will be noticed that the
formation of the notochord is most advanced in those sections
in Avhich the collar-cavities are visible, recalling the con-
ditions found in Bal anoglo ss us.

d'he first indication of the head-cavities is seen when
five somites have been cut off from the anterior part of the
trunk-cavity. A section through the front part of such a
larva is shown in fig. 13. The notochord (c/t.), which by this
time is completely detached from the archenteric wall, has
grown forward towards the extreme front end of the embryo.
Above it we see the nerve-plate grooved and partly covered
by an ectodermic fold, leaving, however, a pore — the neuro-
pore— which persists for a long time in the free-swimming
larva. The archenteron may be seen to be produced above into
a thin bi-lobed vesicle. This is the rudiment of the tAvo head-
cavities, and may be compared to the anterior outgrowth Avhich
gives rise to the proboscis cavity in Balanoglossus.
Already the left lobe may be seen to have a thicker Avail than
that on the right side. In fig. 14 a section through an older
larva in the same region is shown. In this embryo six
somites have been cut off from the front end of the trunk

304

E. AV. MACCRIDE,

cavifc}". By the forward growth of the nerve-plate the neuro-
pore region has been carried forwards, so that in the section
figured it is no longer seen, and not only have the ectodermic
folds met above the nerve-plate, but the nerve-plate itself
has been bent into a nerve-tube. The notochord and the
anterior portions of the collar-cavities have shared in this
forward grosvth, aud these last are seen above the archenteron
at the sides of the notochord. Turning our attention now to
the archenteron we observe that a right thiu-walled vesicle
and a left thick-walletl one are in process of being shut otf from
a central thick-walled gut, but that there is still a narrow seam

Text-fig. 2.

Illustrating the formation of the mesoderm in Amphioxus. H. Head-
cavity. c. Collar-cavity. T. Trunk-cavity, ne. Neurenteric canal.

of communication between right and left moieties. A little
later these are completely separated from the definitive gut
and from each other. The relationship to one another of
head, collar and trunk-cavities is schematically represented in
text-fig. 2, in which an Amphioxus embryo is supposed to
be seen from the side. The embryo is supposed to be trans-
parent, and the formation of the head-cavities is antedated so
as to make it contemporaneous with that of the collar and
trunk-cavities. 'I’he portion of the gut, however, from which
the head-cavities are eventually formed can be clearly seen in
even young embryos, such as that figured in figs. 11 a-d. It
projects in front of the region from which the collar-cavities
are eventually formed.

THE FORMATION OF THE LAYERS JN AMFHIOXUS. 305

The latei’ history of tliese coelomic vesicles aud of the rest
of the archenteron which constitutes the definitive gut may
be followed in larvae, in which the mouth is about to be
formed aud the first gill-slit about to break through. This
stage is almost the latest to which it is possible to rear the
eggs artificially ; for older larvae one has to depend on the
tow-net. A series of seven sections through such an embryo
is represented in figs. 16 a-g, fig. 16 a being, of course, the
most anterior section. In this section the neuropore is seen
and the notochord lying beneath the nerve-cord. At the
sides of the notochord the slit-like anterior ends of the collar-
cavities can be seen, whilst beneath the notochord is seen the
thin-walled right head-cavity which has become shifted
anteriorly to its left fellow. This is one of the first signs of
that curious asymmetry which is one of the peculiarities of
Amphioxus larvae, and which has so puzzled all investigators.
A satisfactory, or at any rate, a plausible explanation of it
has been suggested to lue and will be given later on. In the
more posterior section tig. 16 h, the left head-cavity is seen
lying above the right one as a completely closed thick- walled
vesicle. A vacuolated ectodermal cell (r.) marks the spot
where this vesicle will later acquire an opening to the
exterior. 'L’his section therefore completely refutes the view
of Legros (23) that this vesicle is an ectodermal invagination.
Van ^Vijhe (35) had already decided in my favour after
exiiminiug preparations sent him by both Legros and myself.
Tlie nerve-tube is closed and at the sides of the notochord
the collar-cavities are seen as spacious cavities, the inner
walls of which are, however, becoming thickened. In fig. 16 c
the anterior blind end of the gut is grazed, and in the
thickened inner walls of the collai-caviiies adjacent to the
notochord numerous muscular fibrils {muse.) are seen — these
being the first traces of the formation of the first myotome.
It is of particular importance to note that the lower parts of
the collar-cavities which are wedged in between the ectoderm
and the gut are in open communication with the upper
myotomic portions. Tlie cavity on the right side extends

306

E. W. MACBRIDE.

nearer the mid-ventral line than the cavity on the left side.
Below the most ventral point of the latter the gut is pressed
against the ectoderm, which here exhibits a plate of large
cells; this plate marks the place where the perforation
to form the mouth will take place. In the next section,
shown in fig. 16 d, the enlargement of the anterior end of the
gut to form the pharynx is cleaidy seen, and on the right side
of this there is a pouch-like outgrowth, the rudiment of the
club-shaped gland. This structure has been interpreted
by Willey (36) as the fellow of the first gill-slit, and in this
opinion I am prepai’ed to concur. The posterioi- somites
which have resulted from the segmentation of the collar-
cavity are seen to be completely divided into myotome (my.)
above and splanchuocele (sph) below, but beneath and
external to the right and left rudiments of the
splanchuocele are to be seen the posterior pro-
longations of the right and left collar-cavities (r.
coll., 1. coll.). These extend just as far back and no
further than the enlargement of the gut, which is
the rudiment of the pharynx. In the next section, fig.
16 e, the same features are observable, but here we find the first
gill-slit about to open, and we see that as in all Vertebrata it
is produced by the meeting of an endodermal outgrowth
{hr. end.) and an ectodermal ingrowth [hr. ect.).
Figs. 16 f and g are two sections from the most posteidor
part of the series. In fig. 16/ we see the anal diverti-
culum (a.) which we had already noted in a much younger
embryo, but the anus is not as yet open. In fig. 16 y, two
sections further back, we see the solid cord of cells uniting
gut and nerve-cord (iie.), which is the representative of the
neurenteric canal of earlier forms, and this acts as a
“meristem” for the tail, which in Amphioxus is of rela-
tively small dimensions. Since the division of the gut into
two storeys is still near the hind end of the animal, in spite of
the great growth in length which has taken place, we draw
the conclusion that this growth must take place mainly in
front of this region, and that the band of actively growing

THE FORMATION Ob' THE LAYERS IN AMPHIOXUS. 307

cells is therefore situated just about where it is in the embryo
represented in fig. 11. Throughout the sections of this series
the modifications which the notochordal cells undergo may be
clearly seen ; we can perceive that the greater part of each cell
becomes converted into a vacuole, but that the nucleus and
a ]3iece of the protoplasm immediately surrounding it persists.
In the old free-swimming larvae a peculiar tube, termed
Hatschek’s nephridium, can be seen lying above the front end
of the pharynx and opening into it posteriorly. In 1898 I
asserted that this tube was the persistent connection between
the left collar-cavity and the pharynx. This connection lasts
certainly for a long time, and much longer on the left side
than on the right, and it is found in the same place as
Hatschek’s nephridium. It is shown in an embryo of the
same age as the one we have been considering in fig. 15 as a
cord of cells with virtual cavity connecting pharynx and left
collar-cavity {neph.). Owing, however, to the gap which in
7ny material intervenes between larvm of this age and those
in which an undoubted Hatschek’s nephridium is present, I
am not prepared to dogmatically maintain my assertion of
1898. The oldest stage to which I shall refer in this paper is
represented by fig. 17 a-h (PI. 5) inclusive, which represents
eisflit sections taken frotn the same transverse series. The
specimen which was sectioned was a larva with the mouth and
first gill-slit open and the club-shaped gland fully formed. In
the most anterior section (fig. 17 a) the left head-cavity {Ih.)
is shown to have developed an opening to the exterior. In
fig. 11 h the external opening of the club-sliaped gland is
shown lying just ventral to the thickened plate of cells which
forms the anterior border of the mouth. In fig. 17 c the
mouth is seen opening on the left side of the larva and the
cavity of the club-shaped gland can be seen on the right side
of the pharynx. Beneath the right rudiment of the splauch-
nocele the right collar-cavity can be seen, and the ecto-
derm surrounding it is thickened — this is the first visible
rudiment of the atrial ridge {at.), which eventually meets
its fellow so as to enclose the atrial cavity. Fig’. 17 d-h

308

E. W, MACBRIDE.

represents a, series of five consecutive sections tlirongli tlie
hinder end of the pharynx to show how the collar-cavities
and the pharynx cease simultaneously, and that at the
same time the right and left rudiments of the splanchnocele
meet in the mid-ventral line. If we examine the hinder end of
the pharynx in a much older larva a similar set of appearances
can be made out. Hence we conclude that as new gill-slits
are added in the extraordinary manner described by Willey
(37) the pharynx grows in length, and pari passu the collar-
cavities and their enclosing atrial ridges which flank it. Van
Wijhe (34) has described in the adult animals cavities in the
edges of the oral hood and in the sides of the atrial cavity,
which lie identifies with these inferior portions of the collar-
cavities. He believes, however, that in the adult the upper
portions of the collar-cavities become divided into several
myotonies. His conclusions are very interesting, but owing to
the lack of material of older stages on which I can depend I
cannot either confirm or combat his assertions. I think, how-
ever, that my explanation of the left-sided mouth is simpler
than his.

I said above that a plausible explanation of the asymmetry
of the Amphioxus larvae, which has proved a puzzle to most
investigators, had been suggested to me. The credit of this
is due to my assistant, J. Stafford, Esq., Ph.D., but as he is
unwilling to publish I give it here and take full responsibility
for it. It is as follows: The pelagic larva of Amphioxus
represents a pelagic ancestral condition of the race. Whilst
in this condition the ancestors of Amphioxus were fully
bilaterally symmetrical, and from this stock the rest of
Vertebrata are descended. The adult Amphioxus leads a
burrowing life in sand, a degenerate condition of affairs
compared with its former free-living condition. Nevertheless,
in both conditions of life the food and the mode of obtaining
it are the same. By ciliary action small free-swimming
organisms are whisked into the mouth, and the surplus water
escapes by the gill-slits. Now Amphioxus, like the majority
of fish, is a laterally compi-essed animal, taller than it is

Text-fig.

THE EOEJfATIOX OF THE LAYEES IN AMPHIOXUS. 309

■inial IS the niorpliological mid-ventral line.

310

E. W. MACBRIDE.

broad, and can only maintain its equilibrium so long as it
moves. When it ceases moving it falls on one side, and this
is true of the larvm of Amphioxus, as Willey has pointed
out (37). Between the condition of life when the ancestors
of Amphioxns were entirely pelagic and the condition in
which they burrowed, thei’e intervened in all pi’obability a
transitional condition when they lay on their left
side on the bottom, making only occasional excursions
through the water. In this “ Pleuronectid stage,” as we
may call it, which in all probability represents the mode of
life of the older Am ph ioxu s larvae, it would be advantageous
to twist the originally mid-ventral mouth on to the left side
so as to improve its opportunities of feeding, and at the same
time to twist the left gill-slits on to the upper right side so as
to remove them from the substratum and place them in a
favourable position for evacuating water.

The twist of the mouth seems to have been effected by a
greater growth of the right side of the animal, and hence it
comes about that the endostyle is at first anterior to the
mouth, and so is also the opening of the club-shaped gland
(fig. 17 })). But in the region of the gill-slits the opposite
kind of an inequality of gTowth must have subsisted, for by
greater growth of the left side the left gill-slits are pushed
on to the right side, and the mid-ventral line is displaced
high up on the right side. Very similar conditions prevail
in the head of a developing sole, for in this animal the mouth
is twisted downwards and the left eye upwards. In the
ontogeny of the individual, however, these inequalities in
growth have been hurried on before their time, and the
reason why the left gill-slits only (Willey’s primary gill-slits)
are found in young larvm is that in larvte of small size, when
there ought to be a number of similar structures performing
the same office. Nature reduces the number owing to con-
siderations of want of space (cf. the Nauplius larva of
Crustacea with only three pairs of legs, and the single lateral
cerebral eye in the Ascidian tadpole).

Van Wijhe (35) has given a somewhat similar explanation.

THE FOEMATION OF THE LAYERS IN AMPHIOXUS. 311

He supposes that in one stage of its history Amphioxus
swam in spirals, turning ever from right to left. Under these
circumstances he supposes that the first gill-slit of the left
side, corresponding to the left spiracle of Elasmobranchii,
became enlarged so as to function as a new mouth. He
identifies the pi’esent mouth with the spiracle, because he
supposes that the collar-cavity passes down under it and not
over it, and he quite justly identifies the collar-cavity with
the mandibular cavity of Elasmobranch embryos. In the
supposition that the mouth of Amphioxus lies behind the
lower extension of the collar-cavity he is mistaken, as I hope
the present paper will convince him. I am sorry to differ
from Van Wijhe, for whose generous recognition of my results
I am grateful, but I think that my theory of the twist of
a pre-existing mouth is preferable to any theory which
postulates the formation of a new one, and more in line with
what we know to have occurred in other divisions of the
animal kingdom.

We may sum up our general survey of the formation of
mesoderm and notochord in Amphioxus thus : Both meso-
derm and notochord in Amphioxus are of endodeemal
origin, and the mesoderm originates in a manner closely
recalling that which Bateson described for Balanoglossus;
the differences being that the proboscis cavity divides into
two completely separate head-cavities, that the collai--cavities
have long antero-doi’sal and postero-ventral extensions, ai’e in
a word obliquely orientated, and that the trunk-cavities
become bi’oken up into a series of somites, the dorsal portions
of which form muscular masses, whilst the moi’e ventral por-
tions form an undivided splanchnocele. The notochord, as in
lialanoglossus, becomes mai-ked fii'st in the collar region
(see fig. 12), and grows forward into the head region sub-
sequently. The hindermost part of the gut divides into
neurenteric canal and anal diverticulum.

VOL. 54, PART 3. NEW SERIES.

23

312

E. W. MACBEIDE.

Part II.

Comparison op the Process op the Formation of the
Layers in Amphioxus with the same Process in the
Higher Vertebrata.

Before entering' on a comparison of the formation of the
layers in Amphioxus with the same process amongst the
higher Vertebrata, it is necessary to discuss the position of
Amphioxus amongst the Vertebrata. Hubrecht, and some
others with him, have argued that Amphioxus is not a
primitive but a degenerate form, and hence that its develop-
ment must be disregarded, for this would prove exceedingly
inconvenient for their theories. Now it must be admitted
that in some points Amphioxus is degenerate, and, as we
have seen, its degeneration may be brought into connection
with its burrowing habits in which it has diverged from the
life of its class. But the same is true of all the primitive forms
which Nature has kindly preserved for our inspection, except
those few, like the Sphenodon of New Zealand, which
are confined to very restricted and sheltered areas. All wide-
spread primitive forms must be degenerate, for how could a
primitive animal, whilst retaining its ancestral habits, main-
tain itself in competition with the improved races which have
sprung from its stock ? The thing is absurd ; such an animal
can only maintain itself by getting out of the way, and taking
to a secluded mode of life. Moreover, if Amphioxus is
objected to on this ground, an equal objection must be sus-
tained against Peripatus, against Chiton and the Amphi-
neura amongst Mollusca, against Limulus amongst Arach-
nida, against, in fact, every animal that has thrown any light
on the primitive structure of the class to which it belongs.
All woi’kers in vertebrate morphology, when they have to
deal with the evolution of particular organs, descend to
Amphioxus; and why should we not have recourse to this
foi-m when we seek to give an account of the formation

THE FORMATION OP THE LAYERS IN AMPHIOXUS. 313

of the layers, seeing that it is precisely in i*espect to this
process that the modifications induced by the adaptation of
the stock to a burrowing life do not make themselves felt.
The egg of Amphioxus is relatively free from the factor
that produces the greatest amount of disturbance in the
development of the eggs of the higher Vertebrata, viz. food-
yolk. Moreover, it has an extremely short embryonic life
within the shelter of the egg-shell and a long larval exist-
ence ; and a long larval existence is everywhere a primitive
condition of affairs. As Korschelt and Heider in their
‘ Text-book of Embryology ’ judicially remark, reproduction
by ciliated larvas cannot be interpreted otherwise than as a
primitive feature. But the best proof of all, that the develop-
ment of Amphioxus is really primitive, is to be found in the
way it lends itself to the harmonious explanation of the
development of the higher Vertebrata, assuming as a modify-
ing factor only the disturbing influence of food-yolk, as I shall
now endeavour to set forth.

In our search for other types of development amongst
Vertebrata which may most readily be compared with that of
Am])hioxus, there is one most important consideration to be
borne in mind, viz. that a very large amount of material of
the earliest stnges of development is needed for a successful
analysis of the process of formation of the layers. The closer
to one another are the successive stages available to the
investigator, the less likelihood of error will there be in his
results. Such a condition of affairs is possible in the case of
very few types of Vertebrata. In reviewing the list of
Vertebrata which have been studied, we see that this condition
is fulfilled only in the case of Ascidians, of Urodela and
Anura amongst Amphibia, and to a lesser extent of
Cyclostomata, Dipnoi, Ganoidei and Teleostei amongst
fishes, but these piscine groups have not been studied with
anything like the care that has been lavished on Ascidians
and Amphibia.

In these two last-named gronps thousands of eggs of every
assignable stage can be easily obtained and the processes

314

E. W. MACBRIDE.

tlius followed step by step. But contrast with this the
conditions offered by the development of Elasmobranchs,
Reptiles and Mammals ! Here an adult animal has to be
sacrificed to gain a very small number of eggs; and a long
series of years is requisite before anything like a continuous
series of stages can be obtained. A great many of the
difficulties and disputes which have arisen in connection with
the interpretation of the early developmental stages of the
higher Vertebi’ata are due to this cause alone. Again, if the
theory of evolution be true at all, the developmental stages
of the lower Vertebrata should be less modified than those of
the higher, and should contain the key to them. To quote
the ‘Text-book of Embryology’ of the late Professor F. M.
Balfour on the subject of the development of the fowl, “the
subject itself is by no means commensurate with the atten-
tion it has received. The characters which belong to the
formation of the layers in Sauropsida are secondarily derived
from those in Icthyopsida, and are of but little importance for
the general questions which concern the nature and origin of
the germinal layers.” It is therefore amazing to find
Hubrecht advising students of embryology to tackle Amphi-
bia rather than Amphioxus, and mammals rather than
cartilaginous fishes. The embryology of the higher forms
may show how the process of the formation of germinal layers
may be modified, but not how it originated.

Under these circumstances we may first turn to the
development of the simple Ascidians. The figures given by
van Beneden and Julin for Clavellina and by Kowalevsky for
Ascidia, as reproduced by Korschelt and Heider, are striking-ly
like the figures I have given above for the gastrulation of
Amphioxus. We see the advance of the dorsal anterior lip
of the blastopore, and the coincident formation of the neural
plate above it, and then in the second stage the up-gi’owth of
the lateral and ventral lips and consequent narrowing of the
blastopore. The main differences are ; (1) the contrast between
endoderm and ectoderm cells is greater than in Am phioxus,
and the whole number of cells in both layers is smaller and the

THE EOEMATION OF THE LAYEES IN AMPHIOXUS. 315

individual cells are larger; (2) the up-growth of the ventral
lip is more marked and takes place at an earlier stage.

The whole subject has been fully dealt with by that able
student of cell lineage, Conklin, in his exhaustive description
of the development of Cynthia (19), where he has accounted
for every individual cell and shown its lineage. Conklin
emphatically states that the closing of the blastopore includes
two stages, in the first of which the dorsal lip grows back-
wards, and in the second and subsequent stage the ventral
lip grows upwards and the lateral lips at the same time
coalesce. Conklin admits that mesoblast and notochord are

Text-fig. 4.

4.

Diagrams comparing an advanced gastrula of Amphioxns and of
Cynthia. (According to Conklin.)

differentiated and their component cells recognisable before
they are invaginated, but he utterly declines to call them
ectoderm, “because they are large and full of yolk, and
resemble endoderm.”

When we leave the development of the Ascidian larva,
which in its sense-vesicle and well-developed tail stands higher
in the vertebrate scale than Am phi oxus, the next highest
stage is represented by the Cyclostoine fishes, the develop-
ment of which has been studied by many observers. Unfor-
tunately none of these have been specially interested in the
minute details of gastrulation, but the figures they give of
both segmentation and gastrulation of the egg are extra-

316

E. W. MACBRIDE.

ordinarily like those of Urodele and Amiran Ainpliibia.
Since a very similar method of development is found also in
such old-fashioned types of fish as the Dipnoi, Polypterus,
and the sturgeon, we may with some probability conclude that
this method was once universal amongst Yertebrates, and
persisted till the first invasion of the land had been accom-
plished. Apuultitude of papei’s has appeared on the develop-
ment of the Amphibia, but the careful and exhaustive work
of Bracket ^(6) on the comparison of the development of
Siredon and of the frog* will, I think, for some time represent
the last word on the subject. Bracket finds that at the com-
pletion of segmentation endoderm and ectoderm are not
delineated, that although the lower pole of the egg is occu-
pied by large cells with large yolk-giaunles and devoid of
pigment, whilst the upper pole is the seat of small cells with
small yolk-granules and pigment, yet there is a gradual
passage from tlie one type of cell to the other. As
in Amphioxus the differentiation of ectoderm and endo-
derm accompanies a process of growth and multiplica-
tion of cells, some of the daughter-cells which result there-
from being endoderm with large yolk-granules, and others
ectoderm with small yolk-granules. In this way a skin of small
cells is, as it were, cut out from the surface of the large cells
for a considerable distance below the equator of the egg.
The edge of this advancing skin represents the point x in
Amphioxus. The cavity of the archenteron appears first as
a split within the region of undoubted endoderm
cells. (This is beautifully and clearly shown in the figures
which Bles gives of the development of Xenopus [5].) Then
later the endoderm cells wander inwards in virtue of an
altered cyto-taxis, the arclienteric cavity simultaneously
enlarges, and the segmentation cavity is obliterated. All
question therefore of the roof of the archenteron being
in-turned ectoderm is dis])osed of. The ventral lip of the
blastopore makes its appearance later than the dorsal lip by
a process of multiplication of cells, accompanied by differen-
tiation of the daughters into ectoderm and endoderm, and it

THE FOEMATION OF THE LAYEES IN AMPHIOXUS.

317

slowly advances to meet the dorsal lip. This latter, after the
invagination has taken place, has grown somewhat in
length, and therefore in its edge as in the dorsal lip
of the blastopore inAmphioxus is situated a grow-
ing point by which above new ectoderm and below
new endoderm are produced. This has been proved up
to the hilt by Assheton in his most interesting experimental
studies (2). In Siredon the ventral part of the blastopore
remains open as the anus, whilst the dorsal part is included
within the medullary groove as the neureuteric canal. The

Text-fig. 5.

AMPHIOXUS SIREDON (BRAOHET) STURGEON (deane)

Sagittal sections through the Imlf completed gastrulae of Amphioxus,
Siredon, and Sturgeon.

tail forms as a new growth o n 1 y a f t e r g a s t r u 1 a t i o n is
complete. It will be seen that, making allowance for the
difference in the amount of yolk there is an extraordinary
correspondence between the development of Amphioxus, as
described in this paper, and that of Siredon. The early
formation of the dorsal lip and the later up-growth of the
ventral lip show the same sequence. The difference between
the two embryos lies in the accumulation of yolk in the
ventral wall of the archenteron of Siredon, so that the cells
here are not only more numerous but more massive than those
forming the dorsal wall, and therefore the difference between
roof and floor of the archenteron, which is barely indicated in

318

E. W. MACBRIDE.

Ainpliioxus, is here strongly marked. This difference goes
on accentuating itself as we ascend in the Vertebrate scale, and
it is, of course, this difference which gave rise to the idea that
notochord and mesoderm were ectodermic invaginations. The
reason of this difference is not far to seek. From the dorsal
wall of the archenteron notochord and mesoderm
are formed by processes of folding or by proliferation.
Now, for either of these two processes yolky cells are pre-
eminently un suited, and hence there is an advantage in
storing the yolk necessaiy for development in a portion of
the archenteric wall where formative processes do not occur
until very late.

The work of Kerr (19) on the early stages in the develop-
ment of Lepidosiren is based on a great quantity of material,
and his figures and the general trend of his conclusions
support those of Brachet. I call attention in particular to
the emphasis he lays on the fact that the archenteric cavity
first appears within the region occupied by the endoderm
cells, and on the fact that the ventral lip of the ectoderm is
“ cut out ” from the surface of the large cells of the ventral
half of the egg — that hei’e, as in tlie dorsal lip, growth with
differentiation goes on.

The work of Bashford Dean on the embryology of the three
Ganoid fish, Acipenser (11), Lepidosteus (11), and Amia
(12), enables us to pass gradually from a development like that
of the frog to that of a meroblastic egg. In Acipenser (vide
text-fig. 5) the whole egg is segmented superficially. Above
the whole mass is divided into cells, and there is a spacious
segmentation cavity below ; the segmentation is superficial,
and there is an interior mass of nnsegmented yolk. In Lepi-
dosteus there ai’e but a few superficial furrows at the lower
pole which subsequently completely disappear, whilst in Amia
no furrows are formed at the lower pole at all. In all three
cases at a certain point, which afterwards forms the dorsal
lip of the blastopore, growth and differentiation of cells occur
accompanied by invagination of some of the newly-formed
cells. The ventral lip is formed in the same way, and grows

MORMYRUS MORMYRUS SHARK

THE FORMATION OF THE LAYERS IN AMPHIOXUS. 319

320

E. W. MACBEIDE.

over tlie lower pole and eventually meets the dorsal lip and
so the blastopore is closed. In none of the cases discussed
can there be a question of concrescence of the lateral lips of
the blastopoi’e extending along the neural plate, because
unless such concrescence is to be merely a metaphysical fig-
ment, it must mean that the dorsal lip advances by a pair
of growing points situated to the right and left of tlie
median line, whilst in the mid-dorsal line itself there is a
cessation of growth. Now no such cessation is observable.
A groove found in the floor of the neural canal in Amphibia
has been interpreted as a remnant of this concrescence, but
Brachet has pointed out that this groove is a late secondaiy
phenomenon that does not make its appearance till the blasto-
pore is closed.^ From Am ia we pass naturally to the develop-
ment of a Teleost such as Mormyrus, so well described by
Assheton (3). Here there is, as in Amia (and, of course, in
all Teleostei), a well-marked germinal disc. On the surface
of this the neural plate of the embryo becomes marked out
by its greater thickness. But this neural plate, instead of
extending as in the cases we have just discussed over 180°
or more of the circumference of the egg, extends over quite
a small arc, so that the embryo is reallj’^ differentiated from
but a small part of the egg. At the margin of the germ-
disc as before we find the dorsal lip of the blastopore, but
the ventral lip sweeps completely round the egg and its two
sides close by lateral concrescence on the hinder aspect of
Avhat afterwards becomes the yolk-sac. After this closure
has taken place, and not till then, the growth in length of
the hinder part of the enibryo begins. The alimentary canal
becomes grooved off from the surface of the yolk-sac as a rod
of cells, and the yolk-sac is slowly absorbed.

In the development of Elasmobranchs a very small germ-
disc is formed on the surface of a large yolky egg. Inside

‘ It is true that Brachet in a later papier (7) admits that he is an
adherent of the concrescence theory, but solely on the ground of the
appearances presented by pathological emljryos, which is, I think, a
most illogical position.

THE FORMATION OF THE LAYERS IN AMPHIOXOS. 321

Text-fig. 7.

7.

Median sagittal sections of the gastmlas of tlie Stm-geon, ChiniiBra
and the Shark, s. True segmentation cavity, s'. Inner part
of archenteric cavity sometimes formed by cleavage of the
endoderm. (After Bashford Deane.)

322

E. W. M ACER IDE.

this disc is a segmentation cavity, the Hoor of which at first
contains some cells, but soon becomes unsegmented yolk.
Growth and differentiation of cells leading’ to the formation
of a dorsal lip of the blastopore occur as iisual at the edge of
the germinal disc, and at this point there takes place an
invagination of the lower cells giving rise to an archenteron,
the floor of which is unsegniented yolk containing some free
nuclei. In Chimmra, as Bashford Dean has shown (13), the
invagination takes place, and the dorsal lip is differentiated
not at the edge of the germ-disc, but a little way
inside the edge, and the yolky floor of the archenteron
contains traces of cell division (see text-fig. 7). These facts
point conclusively, as Dean says, to the fact that the unseg-
mented yolky portion of the egg corresponds, not to the whole
of the large cells of the vegetable half of the frog’s egg, but
merely to the lower portion of these.

The lower layer cells of the germ-disc of an Elasmobranch
correspond to the upper half of the yolky cells in the fi’Og’s
egg. The ventral lip of the blastopore, however, is not, as
Dean assumes, situated beneath the open blastopore. It is
the free edge of the sheet of ectoderm which extends from
the opposite anterior side of the germ-disc round the egg,
and wdiich eventually encloses the yolk. The Elasmobranch
peculiarities are two. First, this ventral lip, instead of
extending round the sphere in a great circle,
extends laterally round it in a small circle, and its two sides
meet behind the embryo in a linear streak before the yolk is
covered (see text-fig. 6). Thus the blastopore is divided into
a dorsal and a ventral half ; the latter, called by Balfour the
“yolk-blastopore,” corresponds to a part of the blastopore of
the frog. The dorsal blastopore becomes closed by the medul-
lary folds and gives rise to the neurenteric canal, but the second
Elasmobranch peculiarity is that before this closing takes
place, the growth of the tail region commences and
the hinder end of the embryo presents an open groove
leading from the anal region round to the neural tube. It is
this phenomenon which has given rise to the idea that the

THE FORMATION OF THE EATERS IN AMPHIOXUS. 323

neural plate is progressively formed by the closing of a
slit-like blastopore — an idea entirely negatived by the
examination of those more primitive developmental histories
which we have already described.

Before passing on to consider the development of the
Amniota there is one case which demands special attention,
because a great deal of weight has been laid on it. That is
the description of the development of the Gymnophionan
genus Hy pogeophis by Brauer (8). Here there is a compara-
tively large yolky egg with a germ-disc which alone segments,
though nuclei are found scattered throughout the unseg-
mented yolk. The superficial layer of the germ-disc becomes
differentiated as a well-marked columnar epithelium from the
underlying rounded yolky cells. At one point a raised rim
makes its appearance, and here according to Brauer an
inflexion of the columnar epithelium takes place, leading to
the formation of a sac whose roof is columnar epithelium,
whilst its floor is composed of rounded yolky cells. At the
same time irregular spaces interpreted as a “segmentation
cavity ” have appeared within the yolky cells themselves. In
the next stage the cavity formed by invagination has broken
through into the “segmentation cavity,” and so a quite
roomy archenteron is formed. In the meantime from the
opposite site of the germ-disc the ectoderm has swept
completely round the egg, and so the blastopore is outlined.
The dorsal wall of the archenteron consists partly of
cylindrical cells and partly of rounded cells, and Brauer
maintains that the part composed of cylindrical cells
can be distinctly delineated from the part made of rounded
cells; the part consisting of cylindrical cells becomes then
separated from the rest by the growth under it of the sides
of the yolky layer which constitutes the floor, the sides and
front part of the roof of the archenteron, and this sheet of
columnar cells is entirely used up in forming the lateral
sheets of mesoderm and the notochord, and in this way the
epithelium of the alimentary canal comes to be composed
entirely of “ vegetative ” or lower layer cells. Founding on

324

E. W. MACBRIDE.

these facts, it has been inferred that the columnar epithelium
of the invaginated sac is ectoderm, and even when this sac
has opened into the “segmentation” cavity it is inferred that
a sharp line can be drawn between the two kinds of cells.
Now with reference to this development there are several
points to be noted. First the description labours under the
disadvantage of being founded on a limited number of
specimens. Braner found great difficulty in getting his
material, and his specimens of young stages were few. In
particular a gap occurs about the time of the “breaking
through ” of the invaginate archentei’on, and he has to admit
that it is difficult to fix the precise spot where one sort of
cell leaves off and the other begins. There is no proof that
the place where the “undergrowth” begins corresponds to
this point. Next the invaginate cells are different from the
ectoderm in several points, and in any case a descilption of a
process founded on scanty specimens in which considerable
gaps have to be bridged cannot hold against one like that of
Brachet of Siredon founded on an enormous mass of material.
The whole thing can be interpreted more simply as follows :
lirauer admits that the dorsal lip grows backwards; it is
therefore to be regarded as a growing point from which new
cells are added to the ectoderm above and to the endoderm
below. But we saw that in other Amphibia, before any
invagination had taken place or a dorsal lip had grown out,
the archenteric cavity appeared as a cleavage amongst the
large cells at the lower pole of the egg. Now if we assume
that this cleavage i s represented in Hypogeophis by
the so-called segmentation cavity {/), which, however,
happens to form, not at the surface of the egg, but further in,
and that the “invagination” is the formation of new endo-
derm from the growing point in the dorsal lip, we shall be able
to completely reconcile the development of Hypogeophis
with that of Amphioxus and of Siredon. Indeed, in the
Sturgeon’s egg, as figured by Deane and reproduced in text-
figs. 5, 7 and 8, the archenteric cavity is almost divided into
two parts, such as I have described. The complete separation

THE FOEIIATION OF THE LAYEES IN AMPHIOXUS. 325

Text-fig. 8.

Median sagittal sections through poi’tions of the eggs of Sturgeon
(after Deane) ,Hypogeophis (after Brauer) . Tropidonotus (after
Bellowitz). in order to show the process of gastrulation. s. The
segmentation cavity, s'. Inner portion of archenteron. a. Outer
portion of archenteron.

326

E. W. MACBRIDE.

of the lumen into two parts is of no more importance than
the solidity of the oesophagus in many Vertebrate embryos.

The development of the lower Amniota has been worked
out in several fortns in great detail. The work of Will on
the European Gecko Platydactylus (36), of Mitsukuri on
the turtle Chelone (29), and by Ballowitz on the snake
Tropidonotus (4) have given very concordant results. In
all three cases a dorsal lip of the blastopore is formed, not at
the extreme edge of tlie germ-disc, but inside it, and we are
apparently justified in assuming that the early growth of the
ectoderm from the opposite and anterior side of the germ-
disc, which corresponds to the point y in the gastrula of
Amphioxus, and its short-circuiting round a small circle
of the sphere represented by the whole egg, has taken place
far earlier than it does in the case of the Elasmobranch.
If this be granted, the subsequent development bears a strong
resemblance to that of Hypogeophis. In all three cases a
sac-like cavity forms by invagination beneath the blastoporal
lip {a, text-fig. 8). Another cavity forms by the separation
from the yolk of the endoderm cells, which have differentiated
themselves from the lowest layer in the germinal disc {s'.,
text-fig. 8). This space, coiTesponding to the so-called seg-
mentation cavity of Hypogeophis, breaks through into the
invaginated sac ; so an archenteron is formed, the roof of
which is formed of the invaginated cells, the floor of yolk, and
the sides and anterior slope of “ secondary endoderm.” After
the floor of the invaginated sac and the roof of the secondary
endoderm cavity have met and undergone absorption, three
parallel folds are formed in the roof of the compound cavity.
The centre one gives rise to the notochord, the two lateral to
the trunk coelom on each side. In this way the roof of the
archenteron is used up, and the sides of the compound
archenteron unite beneath it in the middle line to constitute
the roof of the definitive gut. In the higher Amniota, such
as Aves, the invaginated sac is replaced by an almost or
quite solid in-growth of cells, and the dorsal lip of the blasto-
pore becomes drawn out into a slit-like form, the well-known

THE FOEMATION OF THE LAYERS IN' AMPHIOXUS. 327

primitive streak.^ The division of the solid in-growth into
central notochord and lateral mesoderm masses corresponds
to the system of grooves found in the roof of the archentei'on
of lower Amniota.

When we pass to Mammalia, we find that the exact
sequence of events in the earliest stages of segmentation is
most imperfectly known. The egg is described as forming a
solid mass of cells or morula, in one side of which vacuoles
appear, which lead eventually to the formation of a space
filled with fluid. When the egg reaches the uterus it is in
the form of a vesicle with a wall consisting of a single layer
(Rauber’s layer), attached to one side of which is a mass of
cells, called by Hubrecht (18) the embryonic button.
From the superficial layer of the embryonic button the
ectoderm of the embryo is differentiated, and Rauber’s layer
over the region of the button disappears. The development
of the embryo, once this has taken place, closely resembles
that of the chick, so that the principal new question presented
for our consideration is the meaning of Rauber’s layer. I
suggest that it is nothing else than the sheet of ectoderm
which in the eggs of Amphibia sweeps round the ventral
surface of the egg, and eventually forms the ventral lip of the
blastopore. Owing to the loss of yolk by the mammalian egg
it is absolutely necessary for its survival that at the earliest
possible moment it should be provided with a covering of
cells capable of assimilating nourisliment from the womb-wall.
Hence the normal spreading of ectoderm, i. e. growth of the
ventral and posterior lip of the blastopore, takes place long
before there is a trace of the dorsal lip, and hence there
is differentiated a special layer of ectoderm cells for this
purpose even over the embryonic area. That this is a
secondary phenomenon is shown by Hill’s observation that in
^Marsupials this is not the case, but that here the embryonic

’ Wilson and Hill (38) describe in Ornithorliynclivis a very
anomalous condition of affairs, viz. the co-existence of an open blasto-
pore and of a i)rimitive streak some distance behind it.

VOI-. 54, PART 3. — NEW SERIES.

24

328

E. W. MACBRIDE.

button is exposed (16) d In those forms of mammals in which
the embryonic area is invaginated into the vesicle, Rauber’s
layer does not extend over the embryonic area, but only up
to the edges of the invagination. These edges are thickened
tnasses which cohere and form a great plug of ectoderm called
the “Triiger.”

The formation of the coelom in Amphioxus as five out-
growths from the gut, viz. a median and two paired out-
growths, is not without parallels amongst other Yertebrata,
even in the present state of onr knowledge, and it is more
than probable that if attention were once directed to this
point, a method of development fundamentally similar to that
of Amphioxus would be found to range throughout the
whole of the Yertebrata. In the embryos of the shark
Acanthi as there is found a bilobed outgrowth from the
anterior end of the gut, which becomes separated off as a
single pre-mandibular cavity, which divides into two cavities,
from whose Avails the inferior oblique muscle of the eye and
the superior inferior and internal rectus muscles are developed.
In the mandibular arch lies the so-called mandibular cavity,
the origin of Avhich has not been traced. From its upper
extremity, Avhich extends forwards over the pre-mandibular
cavity just as the collar-cavity of Amphioxus extends over
the head-cavities, the superior oblique muscle is developed,
Avhilst from its lower portion constrictor muscles of the throat
are developed. A pre-mandibular cavity originating fi’om
the gut has been described in the lizard, and by EdgeAvorth
in the chick (14). Pre-mandibular and mandibular cavities
have been described in Urodela, Elasmobranchii, and in all
Arnniota in Avhich they have been looked for; in Anura and
Dipnoi they are apparently represented by solid masses of
cells, as Agar has pointed out (1). That the trunk-cavities

* I had the pleasure of listening to Professor Hill's exposition of this
2>oint and of seeing photographs of his preparations at the Dublin
meeting of the British Association in 1908. On referring to the ‘ Pro-
ceedings ’ of the Association I regret to find that this interesting pa^Jer
is reported by title only.

THE FORMATION OP THE LAYERS IN AMPHIOXUS. 329

are represented by the great solid sheets of mesoderm cut
out from the sides of the archeiiteric roof termed the “ meso-
dermic bands/’ requires no special demonstration.

To sum up^ the differences between the development of
Arnphioxus and the development of the higher Verte-
brata can be explained on the simple assumption that there
has been a progressive increase in food-yolk, and that this yolk
for the most part has been stored in the ventral wall of the
archenteron, which has been thereby rendered relatively inert.
Tliis has led to a modification of the process of invagination,
which retains its primitive features in connection with the
dorsal lip of the blastopore, but ventrally is changed to a
process of slipping over or epibole. At the same time the
processes of folding which give rise to the coelom in Amphi-
oxus become modified so as to give place to the outgrowing
of solid masses of cells.

Turning now to Professor Hubrecht’s account (18) of the
ontogenetic processes in Vertebrata, we find that he entirely
reverses the method which I have followed. Instead of ex-
plaining the more complex development of the higdier forms
as a modification of that of the simpler forms, he takes the
development of mammals as a starting-point, and then pro-
ceeds to read into the development of the lower forms what
he, finds there. Since in the mammalian egg the cells destined
to form notochord and mesoderm arise by invagination, there-
fore they are ectoderm and for them the name “protochordal
wedge” is given. It does not, however, escape Hubrecht that
these invaginated cells come into continuity in fi-ont with
cells which he regards as true endoderm, and therefore in
front of the protochordal plate there is an endodermic proto-
chordal plate, and the notochord, if I understand him ai’ight,
arises from both, and is therefore a compound strnctui’e.
Further, Hubrecht is a convinced believer in the formation
of the neural plate by the gradual closing of a long slit-like
blastopore. This process he dignifies with the name “noto-
genesis.” To read a complex process like this into the develop-
ment of Arnphioxus appears to me a sheer impossibility.

330

E. W. MACBEIDE.

In support of it, it is true, Hubreclit figures from Legros an
oblique longitudinal section of an abnormal Ampliioxus
gastrula, which is utterly unlike the appearance presented
by any normal embryo. Legros admits the section to be
oblique, a fact of which Hubreclit does not apprise his
readers. Hubreclit regards Rauber’s layer as a special larval
envelope and utterly distinct from the true ectoderm. He
imagines that the aquatic ancestor of Mammalia had a larva
in which there was such an envelope which was afterwards
cast off, and cites certain Trochophore larvm as analogous
instances. When the aquatic ancestor took to a land life the
free-swimming larva was retained within the womb of the
mother, and so the peculiar development of Mammals was
attained. According to this reasoning, then either birds
and reptiles arose from a different stock from Mammals, or
else the oviparous method of development which they exhibit
was secondarily developed out of a previous viviparous con-
dition. Now on this view several remarks may be made.
Rauber’s layer is not analogous to the investing layer of the
Sipun cuius larvfe as Hubreclit imagines, because iu the
latter case we have to do with a median belt of larval ecto-
derm which develops into a broad ciliated baud overlapping
the remaining ectoderm before and behind. The loss of this
belt in the Trochophore larva leaves a wound which is closed
by the cicatricial union of the ectoderm produced by the
head and tail blastema respectively, whereas Rauber’s layer
is an outer layer of ectoderm according to Hubreclit. Then,
whichever alternative we take of the ancestry of Mammalia,
we are beset with difficulties. To maintain that they are the
offspring of a distinct stock from that which gave rise to
birds and reptiles is a supposition which may be left to the
tender mercies of comparative anatomists, who will make short
work of it. Every recent discovery in palaeontology tells
against such a supposition ; the mammalian vertebral column
is constructed on the reptilian plan, whilst the amphibian one
is built on a different plan, and so on. But if we admit the
existence of a common ancestral stock of birds, reptiles, and

THE EOEMATION OF THE LAYERS IN AMPHIOXUS. 331

mammals, then the change from a viviparous to an oviparous
method of development, which Hubrecht must postulate, is
totally unthinkable. How should au animal which had once
adopted the habit of carrying the young in the womb — the
safest method of development, and the one which rendered
the parent completely free from the necessity of visiting any
fixed place for parturition — revert to the dangerous and
primitive method of laying eggs ? In every other case in
which viviparity occurs in the animal kingdom we have
evidence that it has developed out of oviparity, not vice
versa. Hubrecht lays great stress on supposed indications
of a specially differentiated layer of ectoderm over the
embryonic areas of the embryos of reptiles, like Sphenodon
and of Echnida, as a proof of their descent from viviparous
forms. Hill expressly denies the existence of such a layer
in Ornithorhy nchus (38) and the marsupials (16), but
even if the facts were as Hubrecht i-epresents them it by no
means proves his case. The ectoderm in all the higher Verte-
brata is typically many-layered, and that an outer layer
should prematurely become differentiated is only au antici-
pation of adult conditions. Even in Elasmobranchii and
Cyclostomata, which Hubrecht classes together with Amphi-
oxus, and separates from all other Yertebrata on account of
tlieir single-layered embryonic ectoderm, eventually develop
a many-layered adult ectoderm. Natural selection may have
seized on this tendency when Rauber’s layer was evolved.

The way 1 have suggested of looking at the evolution
of Yertebrata, which has the audacity in these days of inno-
vation to be commonplace, escapes all these difficulties. All
grades in the development of viviparity are met with amongst
living reptiles, and it does not I’equire a very violent exercise
of the imagination to pass from a condition such as is found
in Zootoca to that found in Ornithorhy nchus, for
example.

Rut Hubrecht claims for his view that it enables him to
explain the origin of the allantois and the amnion as
embryonic envelo2)es in a satisfactory way. He pictures the

332

E. AV. MACBIJIDE.

process as far as I can follow him in some such manner as
this : Starting with a supposed holoblastic egg which had a

larval envelope, the mass of cells destined to form the embryo
does not develop with equal rapidity to the surrounding
envelope, and consequently becomes detached from it every-
whei*e except at one spot, where there is a stalk of connection
between envelope and embryo. The amniotic cavity between
true amnion and embryo was originally a water cushion
developing within the ectoderm to form a protection for the
embryo, which subsequently derived a more complete pro-
tection by the method of detachment just described. Along
the stalk of connection between embryo and vesicle the
bladder subsequently grew, and so the allantois was formed.
From this process the simple folding process seen in the
formation of the amnion in Reptilia is supposed to have
arisen as a secondary modification. Now an amnion has been
developed in Insecta, and Hubrecht will find it hard to
convince any specialist in Ai’thi'opoda that it has come about
otherwise than as a modification of the folding' of the germ-
disc seen in its incipient stages in Myriapoda. But why, if a
protective fold has developed in the Arthropodan egg, should
there be any difficulty about its development in the Verte-
brate egg ? Suppose we apply in both cases the same
hypothesis. The folding of the germ-disc in a Myriapod egg
is due to its great length and the consequent impossibility of
its expansion within the egg-shell. In the insect egg the
germ-disc is proportionately shorter, but the iidierited habit
of folding' has persisted and has led to the formation of the
amniotic fold.

Now ill the Amphibian egg no expansion of the embryo
takes place till it has escaped from the egg membrane, but
when the Reptilian ancestor took more completelj^ to life on
land a longer retention of the embryo within the egg-shell
and a greater supply of yolk would become a necessity.
Hence at a certain stage in the development of land animals
there arose a necessity of bending the head inwards into the
yolk-sac so as to give room. The most flexible part of the

THE FOllMATlON OF THE LAYERS IN AMPHIOXUS. 333

yolk-sac was the so-called pro-amnion, which lay in front of
and beneath the head, and here the flexure took place. 'I'he
inere fact that in Mammalia there is here no meeting of the
lateral sheets of mesoderm, so that the diploblastic head
envelope is called “pro-amnion,” whereas in reptiles the lateral
sheets of mesoderm early meet beneath the head, and so a
“ true head fold of the amnion ” is developed, seems to me to
be a secondary affair. This bending tended, just as a ship
sinking by the head tends to lift her stern, to raise the tail
region and so bring the bladder-like outgrowth of the gut —
already present in Amphibia — near the surface of the egg',
and so to increase its chances of getting oxygen. In this the
foundation was laid for the modification of the allantois into
a breathing organ and for the corresponding development of
the tail-fold of the amnion, the two developing, as Balfour
long ago showed, together, since the tail-fold of the
amnion contains the extension of the bladder or allantois.

Hubreclit scornfully asks if the pro-arnnion was developed
to contain the head, Avhy there is none in the human embryo ?
The answer is easy; in the human embryo the embryo itself
is as long as the yolk-sac from the beginning, the yolk-sac
being in this case a vestigial organ which has suffered great
reduction in size, and hence when the embryo elongates there
is no yolk-sac for it to plunge its head into. Space, however,
will not permit us to pursue this subject further. To an
author like Hubreclit, who finds no difficulty in supposing
that the oviparous mode of development in Echnida is secon-
darily derived from a placental method of development, such
as is found in the rabbit, no change is so unlikely as to seem
impossible.

Apart from Hubrecht’s desire to prove that the ancestors
of Mammals never had yolky eggs, the main result of his
paper is to advocate the view that Vertebrates have developed
from an Actiuian ancestor. The protochordal wedge repre-
sents, according to him, the old stomodaeum which opened
into the true endodermal gut below. The old mouth was
originally surrounded by a nerve ring, but it became closed

THE FORMATION OF THE LAYERS IN AMRHIOXUS. 335

from in front backwards and the anus is the only remnant of
it; the present mouth is a new formation. In this theory
Hubrecht supposes that he is reviving an old theory of
Sedgwick’s as to the origin of the Metazoa (30). But on
Hubrecht’s theory the coelom is of ectodermal origin, and
must have originated from stomodmal pockets which do not
exist in Actinozoa, whilst Sedgwick regards the coelom as
derived from the inter-mesenteric snaces of the true endo-

X

dermal gut. Sedgwick’s theory was published about a quarter
of a century ago, and was based on a review of all the
evidence available at the time. It is a theory of the origin
not of Vertebrata but of all Metazoa, and it traces them back
to an Actinia-like ancestor. The theory of Sedgwick may be
analysed into three parts — for the name Actinia given to the
common ancestor was, of course, only an indication of a very
general resemblance to a modern sea-anemone. The three
parts are: (1) The position that the coelom arose as pockets

of the original gut or archenteron ; (2) the position that
mouth and anus are two separated portions of a long slit-like
mouth ; and (3) the idea that the central nervous system is
homologous throughout the whole of the Metazoa and had
originally the form of a ring round the original mouth. Now
I venture to maintain that the first two parts of the theory
have received more and more support as embryological
research has gone on, but that the third part must be given
u]) and that the comtnon Metazoan ancestor in consequence
takes on more resemblance to a Ctenophore than to an
Ac ti Ilian, for it has become evident that in many types of
larva) the apical plate of neuro-epithelial cells is the first
rudiment of the brain, and that it is independent of post-oral
nervous aggregations. ’Jliat the coelom arises in Amphioxus
as five archenteric outgrowths I trust I have convinced the
readers of this paper. ’J'hat the mouth and anus in all
animals in which two such openings are found, owe their
origin to the division of a long slit-like cceleuterate mouth I
firmly believe. If, however, the affinities of Amphioxus
are to be sought for in the vicinity of Balanoglossus, then

336

E. W. MACBEIDE.

an excellent reason can be given why we cannot expect to
find a trace of the slit-like inoutb in Vertebrate ontogeny,
for there is no trace of it in the development of Balano-
glossus nor in the development of the Echinodermata,
which is the group most nearly allied to Balanoglossus
and the Enteropneusta. In both Enteropneusta and Echino-
dermata the blastopore becomes the anus and the mouth
is formed later as an independent meeting of ectoderm
and endoderm. I am strongly inclined to believe that the
“ seam ” which originally connected mouth and anus was
situated on the ventral and not on the dorsal surface. It
is quite possible tliat the second stage in gastrulation, viz.
the upgrowth of the ventral lip and the coincident union of
its lateral halves, may be a reminiscence of this closure. The
great difficulty in fixing points of reference in Vertebrate
development is that general growth in length supervenes so
early that before the mouth is formed the embryo has altered
in size and shape to a great extent. Nevertheless the place
where the mouth is formed in Amphioxus cannot be far
from the point y in the gastrula in fig. 6, in which the
upgrowth of the ventral lip has not yet begun.

Now Balanoglossus presents us with a condition where
the nervous system, as in some Nemertinea and Echino-
dermata, is practically co-terminous with the ectoderm — a
condition therefore in which a central nervous system could
originate anywhere where stimuli were concentrated. The
local concentration in the dorsal region of the collar which
we believe to be the forerunner of the earliest part of the
A^ertebrate dorsal nerve-tube receives its explanation from
the structure of the other Enteropneusta, Cephalodiscus and
Rhabdopleu ra. In both of these animals the collar region is
produced into a series of arm-like outgrowths beset with
ciliated tentacles. The dorsal nerve cord is a centre co-
ordinating the two halves of this apparatus. The free-
swimming pelagic ancestor of Enteropneusta probably
possessed similar developments of the collar region, and it is
interesting to reflect that the ciliated cirri” which fringe

o o

THE FOEMATION OP THE PAYEES IN AMPHIOXUS. 337

the edge of the oral hood in Ainphioxus occupy a corres-
ponding position to the “ arms” of Cephalodiscns, and are
in all probability homologous therewith. Adding the “arms”
to the collar of a Balanoglossus and shortening the worm-
like trunk, the length of which is a result of burrowing' life,
we arrive at a conception of the primitive Vertebrate stock
to which the free-swimming ancestors of Echinodermata were
allied. In the Echinodermata the fixed habit was adopted
and one of the “collar-cavities” gretv at the expense of the
other and so the water-vascular ring was formed.

Professor llubrecht indicates a belief that the “supposed
Actinian ancestor developed into a vermiform one.” What

Text-fig. 10.

The foimuon ancestor of Vertebrata, Enteropneiista and
Echinodermata.

kind of a “vermiform” ancestor Hubrecht means he does
not indicate in the paper under discussion. To judge from
his previous work one would suppose that the “ worm ” was
a Nemertean. The theory of the Nemertean origin of Verte-
brata was published in this Journal in two editions. In the
later and revised edition (17) we learn that the Vertebrate
nervous system corresponds to an inconspicuous dorsal nerve
in the Nemertinea, whilst the main nervous system of those
“worms” gives rise to the chain of cranial ganglia. The
proboscis of Nemertinea becomes the hypopliysis of Verte-
brata, whilst the proboscis-sheath forms the notochord. The
question of the coelom is passed over as a difficult and obscure
(piestion. As, however, the figure he gives of the vermiform
ancestor in the present paper is adorned with a number of

338

E. W. MACBRIDE.

coeloinic pockets, one is tempted to believe that lie means
to give his adhesion to the famous “ Annelidan ” theoi-y of
the origin of Vertebrates put forward by Dohrn, to which
we owe such a quantity of excellent work. This seems the
more likely, as in the twenty-two years which have elapsed
since its final revision the Nemertinean theory has gained
little or no support. That the Nemertinea are distantly
related to the common stock of Echinodermata and Vertebrata
is quite possible ; but that they stand anywhei*e near the
direct line of descent is negatived by the consideration that
the J^ilidium larva in its development stands near the Trocho-
phore larva of Annelida, and is widely different from the
Tornaria larva of Balanoglossus or the allied Dipleurula
larva of Echinodermata. The Annelidan theory did justice
to a number of remarkable I’esemblances between Annelida
and Vertebrata. In proportion, however, as zoological
research has advanced, it has transpired that most of these
resemblances — cf. ciliated tubes leading from the body-
cavity to the exterior, development of genital organs from
the lining of the body-cavity, etc. — are common to a wide
range of coelomate animals, and only one specific resem-
blance is left, viz. the metameric repetition of organs, and
above all the metameric segmentation of the muscles in the
two groups of animals. On the other hand, if Vertebrata
were derived from Annelida, we are forced to assume the
production of a new mouth and the abandonment of the old
one, and a total change in the manner of developing the
nervous system, whilst gill-slits and notochord must have
been developed entirely de novo.

The great advantage of Bateson’s theory of the origin of
Vertebrata is that we are not forced to make any such violent
reconstruction as the formation of a new mouth, and of noto-
chord and nerve-cord we already have the beginnings. All
we require to postulate is the appearance of metamerism, and
surely this is no unreasonable assumption considering how
often the repetition of similar organs crops up in the most
widely separated phyla of the animal kingdom.

THE FOEMATION OF THE LAYERS IH AMPHIOXHS. 339

Of course Hubreclit may reply that there is no a priori
impossibility in the formation of a new month. There is cer-
tainly no analogy for it, and if it is legitimate morphological
reasoning to assume such changes of function as are implied
in it, no valid objection can be brought against the attempts
to evoke a Vertebrate from an Arachnid like Limulus, in
which Patten and Gaskell display such diabolically brilliant
ingenuity.

The general conclusion of our study may be summed up thus :

The process of gastrulation in Amphioxus leads to the
formation of a single laye'' of invaginated cells, which there
is no valid reason for analysing into two kinds.

The closure of the blastopore in Amphioxus is due to the
concrescence of the lateral lips of the blastopore and to the
upgi'owth of the ventral one. By these processes the venti’al,
not the dorsal surface of the embryo is formed.

The mesoderm owes its origin to the outgrowth of five
coelomic pouches from the archenteron in the same manner
as the coelom of Balanoglossus.

The formation of the layers in other Vertebrata can be
derived from that of Amphioxus by allowing for first the
disturbing influence of the accumulation of food-yolk in the
ventral wall of the archenteron, and, secondly, in Mammalia
the disturbing effect of contact with the maternal uterus.
In starting with Mammalia, and reading their complicated
pi’ocesses into the development of lower Vertebrata, Pi’ofessor
Hubreclit has read the book of Vertebrate development
upside down.

McGill University,

April, 1909.

List op Works referred to in this Paper.

[Except in the case of Amphioxus this list makes no pretence at
completeness. In the case of the higher veiLehrates I have given
references only to the newest work on the subject w'hich I could find.]

1. Agar. — “ The Development of Anterior Mesoderm in Lepidosiren
and Protopterus,” ‘ Trans. Roy. Soc. Edin.,’ vol. xlv, pt. 3, 1907.

340

E. W. MACBRIDE.

2. Assheton. — “ On Grow-th Centres in Vertebrate Embryos,” ‘ Anat.

Anzeiger,' Bd. xxvii. 1905.

3. *• The Development of G y m n a r c h u s n i 1 o t i c n s," Memorial

volume of John Samnel Biidgett, 1907.

4. Ballowitz. — “ Die Gastrulation bei der Ringelnatter,” ‘ Zeit. fiir

IViss. Zool.,’ vol. Ixx, 1901.

5. Bles. — The Life-bistory of Xenopus laevis,” ‘ Trans. Roy. Soc.

Edin.,’ vol. xli, 1905.

6. Bracbet. — “Rechercbes sur I’ontogenese des Ampbibiens U rodeles

et Anonres,” ‘Archives de Biologie,’ tome xix, 1902.

7. “ Gastrulation et formation de I'embiyon cbez les Cbordes,”

‘ Anat. Anz.,’ Bd. xxvii, 1905.

8. Brauer. — ‘‘ Beitriige zur Kenntniss der Entwickelungsgescbicbte

nnd der Anatomie der Gymnopbionen.” ‘Zool. Jabrbiicber
Abt. fiir Anatomie nnd Ontogenie,’ vol. x, Ht. 3, 1897.

9. Cerfontaine. P. — “Rechercbes sur le developpement de EAmpbi-

oxus,” ‘Archives de Biologie,’ tome xxii, 1906.

10. Conklin. — “ The Orientation and Cell-lineage of the Ascidian Egg

(Cynthia partita),” ‘Joum. Acad. Xat. Sci.,’ Philadelphia,
vol. xiii, second series.

11. Dean, Bashford. — “The Early Development of Gai-pike and

Sturgeon." ‘ Journ. of Morphology,’ vol. xi. 1895.

12. Dean, Basliford. — “The Early Development of Amia.” ‘Quart.

Journ. Micr. Sci., vol. 38, 1896.

13. “ Chimseroid Fishes and their Development,” ‘ Publications

of Cai-negie Institution.’ 1906.

14. Edgeworth, F.H. — “The Origin of the Head-muscles in Gallus and

other Sauropsida.’’ ‘ Quart. Jouni. Micr. Sci.,’ Xo. 204, vol. 51, 1907.

15. Hatschek, B. — “ Studien ilber die Entwickelung des Amphioxus,”

‘Arb. alls der Zool. Inst, zu Wien.,’ Bd. iv, 1881.

16. Hill, J. P. — “ The Early Development of Mai’supialia,” Repoi-ts

of British Association Dublin Meeting. 1908.

17. Hubrecht. — "The Relation of the Xemertea to the Vertebi-ata,"

‘ Quart. Journ. Micr. Sci.,’ vol. 27. 1887.

18. “ Early Ontogenetic Phenomena in Mammals and their

Bearing on our Interpietation of the Phylogeny of Vertebrata.”

‘ Quart. Journ. Micr. Sci.,’ vol. 53, pt. 1, 1909.

19. KeiT, J. Graham. — “The Develoinnent of Lepidosiren para-

doxus,” pt. ii. ‘ Quart. Journ. Micr. Sci..’ vol. 45, 1901.

20. Klaatsch. — “ Bemerkungen fiber die Gastrnla des Amiihioxus,”

‘ Moi'ph. Jahrbuch,’ Bd. xxv, 1898.

THE FOKMATION OP THE LAYERS IN AMPHIOXUS. 341

21. Kowalevsky. — “ Entwickelimgsgescliichte cle.s Amphioxns lan-

ceolatus,” ‘Mem, cle I’Acad. imp. des Sc. de S. Petei'sbonrg,’
series, tome xi, 1867.

22. “Weitere Stndien iiber die Entwickelimgsgeschichte des

Ampliioxus lanceolatus,” ‘ Arcbiv fiir Mikr. Anatomie,’
Bd. xiii, 1877.

23. Legros. — “Developpement de la cavite Imccale de I’Amphioxiis,”

‘ Arch, d’ Anatomie micr.,’ tome i and ii, 1897 and 1898.

24. “Sur fpielqiies cas d'asyntaxie blastoporale cliez rAmphi-

oxus,” ‘Mitt, aus der Zool. Station zn Neapel,’ Bd. xviii, 1907.

25. Lwoft'.— “ Die Bildung der primai'en Keimbliitter mid die Entste-

Imng der Chorda mid des Mesoderm bei den Wirbelthieren,"
‘Bull, de la Soc. Imp. des Nataralistes de Moscon,’ 1894.

26. MacBride. — “ Tlie Early Development of Amphioxns,” ‘Quart.

Joiirii. Micr. Sci.,’ vol. 40, 1898.

27. “ Further Remarks on the Development of Amphioxns,"

‘ Quart. Jonrn. Micr. Sci.,’ vol. 43, 1900.

28. Morgan and Hazen. — “The Gastrnlation of Amphioxns," ‘Jonrn.

of Morphology,’ vol. xvi, 1900.

29. Mitsnknri. — “ On the Process of Gastrnlation in the Chelonia,”

‘ Jonni. Coll. Sc. Imp. Univ. Japan,’ vol. vi, 1893.

30. Saniassa. — “ Stiidien fiber den Eiiiflnss des Dotters anf die gastrnla-

tion mid die Bildnng der jirimaren Keimbliitter der Wirbelthiere :
Amphioxns,” ‘ Arcliiv fiir Ent. mechanik,’ Bd. vii, 1898.

31. Sedgwick. — “ On the Origin of Metameric Segmentation and some

other Morphological Questions,” ‘ Quart. Jonrn. Micr. Sci.,’ vol.
24, 1884.

32. Sobotta. — “ Beobachtnngen iiber die gastrnla des Amphioxns,”

‘ Verh. der Phys. mid med. ges. zn Wiirzlmrg,’ Nene Folge, Bd.
xxxi.

33. Theel. — “ The Development of Echinocyamns pnsillns,” ‘Roy.

Soc. Sc. Upsala,’ ser. 3, 1892.

34. Van Wijhe. — ‘Beitriige znr Kopfregion des Amphioxns lanceo-

latns,’ 1901.

35. “Die Homologisirmig des Mmides des Amphioxns mid die

primitive Leibesgliedcrnng der Wirbelthiere,” ‘ Petrus Camper.’
De 4 aft.

36. Will. — “ Beitriige znr Entwickelmigsgeschichte der Reiitilien : die

Anlage der Keimbliitter bei der Gecko (Platydactylns),” ‘Zool.
Jahrlhicher,’ vol. vi, 1892.

37. Willey. — “ The Later Larval Development of Amphioxns,”

‘Quart. Jonrn. Micr. Sci.,’ vol. 32, 1891.

342

E. W. MACBRIDE.

38. Wilson and Hill,
vliynclius,” ‘
1907.

— “Observations on the Development of Ornitho-
Pliil. Trans. Roy. Soc. Loud.,’ series B., vol. cxcix,

EXPLANATION OF PLATES 18—21,

Illustrating Prof. E. W. ^lacBride’s paper on “ The Formation
of Layers in Aniphioxns.”

[All the figures in this paper were drawm under the magnification
afforded by a Zeiss apochromatic immersion objective of 2 mm. focal
distance. The original magnification of 900 to 1000 diameters has l)een
retained in the case of the smaller figiu-es, but the magnification has
been reduced in the case of the larger to 600 diameters for convenience
in i-eproduction.]

List op Abbreviations.

a. Anal diverticulum, a'. Upper section of archenteron. at. Rudi-
ment of atrial ridge, hr.ect. Ectodermic portion of gill-pouch, hr.eud.
Endoderinic portion of gill-pouch, ch. Notochord, chib. Club-shaped
gland, coll. Collar-cavities. /. Ectodermic folds which extend over
the nerve-plate. 7i. Karyokinesis (mitosis). l.coU. Left collar-cavity.
l.h. Left head-cavity. 7.sp7. Left splanchnoccele. m. Place where the
mouth will be formed, liiwsc. Muscular fibres, my. Myocoele. «.c. Neural
canal. n.e. Neurenteric canal. nepli. Hatschek's nephridium. oc.
Clear cells; first indication of eye-spot, r.coll. Right collar-cavity.
r.h. Right head-cavity, r.spl. Right splanchnocele. tr. Trunk-cavity.
V. Vacuole in spot where left head-cavity will acquire an opening to the
exterior, x. Position of centre of growth which initiates gastriilation.
y. Position of centre of growth which closes the blastopore.

PLATE 18.

All the sections on this plate are magnified 600 di.ameters.

Fig. 1. — Median sagittal section through a flattened blastula in which
the centre of growth at x is just making its appearance.

Fig. 2 . — Median sagittal section through a slightly older embryo in
wdiich gastrulation has just begun.

Fig. 3. — Medial sagittal section through a still older einlnyo in which
gastrulation has proceeded further.

THE FOEMATIOX OP THE LAYERS IN AMPHIOXUS. 343

Fig. 4. — Median sagittal section through an embryo in which the
difference in staining quality between ectodermal and endodermal
nuclei is beginning to appear.

Figs. 5 a and h. — Two sagittal sections through an embryo in which
the nerve-plate (n.p.) is just beginning to be recognisable. Fig. 5 a is
a median section, and in it the rounded character of the cells in the
neighbourhood of the growing point x is remarkable. Fig. 5 is a more
lateral section (four sections to the right) in which the neighbourhood
of X is composed of quiescent cells.

Fig. 6. — A median section through a gastrula in which the archenteron
has become hemisi^herical.

PLATE 19.

Figs. 7 a and b. — Two sagittal sections through an embryo in which
the pi'ocess of elongation has begun. Fig. 7 o is a median sagittal section
and fig. 7 h a moi-e lateral section. The long axis of the nerve-plate is
at right angles to the diameter of the wide blastopore. Magnification
fiOO diameters.

Fig. 8. — Median sagittal section through a gastrula in which the
blastopore is in process of being closed through the activity of the
growth-centre at y. ch. The layer of cells which will form the noto-
chord and which is already beginning to lose its yolk-granules. Magni-
fication fiOO diameters.

Fig. 9. — Horizontal section through an almost complete gastrula
taken near the middle of the series. Magnification 600 diameters.

Fig. 10. — Horizontal section through an almost complete gastrula
taken near the upper and anterior lip of the blastopore. Magnification
1)00 diametei’s.

Figs. 11 n-e. — Five transverse sections through a just complete
gastrula in which the first traces of the body-cavities can be seen. In
fig. 11 a, the most anterior section, the collar-cavities (coU.) alone are
visible. In fig. 11 both collar-cavities and trunk-cavities are seen. In
fig. 11c only trunk-cavities are visible. In fig. 11 d the anal divei-ticulum
(«.) is seen still connected with the more dorsal part of the archenteron
(«'.) by a narrow canal. In fig. 11 e the anal divei-ticulum is shut off from
the rest of the archenteron. ne. Neurenteric canal. Magnification 675
diameters.

PLATE 20.

Figs. 12 a-d. — Four transverse sections through a larva in which two
somites have been cut off from the trunk-cavity. In fig. 12 a the
openings of the collar-cavities are seen on both sides. In fig. 12 5
the collar-cavity is closed on the left side and lies on the outer side of
VOL. 54, PART 3. — NEW SERIES. 25

344

K. \V. MACT.KTDK.

tlie closed front end of the trunk-cavity. The collar-cavity is still open
to the gut on the riglit side ; in fig. 12 c it is closed off on both sides. In
fig. 12 d the posterior ends of the trunk-cavities are seen to open into
the ai’chenteron. Magnification 900 diameters.

Fig. 13. — Transverse section through the front end of a larva in
which four somites have been cut off from the front end of the trunk-
cavity to show the origin of the head-cavities, pore. The neuropore.
l.li. Left head-cavity, r.li. Right head-cavity. Magnification 900
diameters.

Fig. 14. — Transverse section through the front end of a larva in
which five somites have been cut off from the front end of the trunk -
cavities. The head-cavities are more advanced in development, but still
open into the gut. r.coll., l.coU. The anterior ends of the right and
left collar-cavities respectively. Magnification 900 diameters.

Fig. 15. — Transverse section through the front end of a larva about
two days old to show the persistent connection of the left collai’-cavity
with the pharynx, out of which it appears probal)le that Hatschek's
neplu-idium {nepli.) developed. Magnification 1000 diameters.

Figs. 10 n-f/. — Seven transverse sections through a larva of about the
same age as that represented in fig. 15 before the mouth is open. In
fig. 16 a the still open neuropore (pore) is seen, and below the notochord
the thin-walled right head-cavity. In fig. 10 h the left head-cavity is seen
completely closed off from the gut. and below it the right head-cavity.
At V. a vacuole is seen in an ectoderm cell, which marks the spot
where this cavity will eventually acquire an opening to the exterior.
In fig. 16 c the thickening of the ectoderm (m.) is seen, which marks the
spot wheie the mouth will break through. The collar-cavities are seen
at the sides of the gut extending towards the mid-ventral line. In fig. 10 d
the club-shaped gland is seen originating as a hollow outgrowth from
the wall of the pharynx, and the posterior extensions of the collar-
cavities are seen below the splaucbnoceles, which have been foianed by
the fusion of the ventral jDarts of the posterior somites, my. The
myocele of one of the myotomes. muse. Muscular fibres on the inner
walls of these myotomes. In fig. 16 e the first gill-slit is seen to originate
by the meeting of an ectodermal in-growth (br.ect.) and an endodermal
outgrowth (br.end.). In fig. 16/ the anal diverticulum is seen, and the
expansion of the ectodermal cells to form the caudal fin (/a), which is
seen as a cuticular rim. In fig. 16 gr the solid seam of cells representing
the neurenteric canal is seen. Magnification lOOO diameters.

PLATE 21.

Figs. 17 a-h. — Eight transverse sections through a larva in which the
month and first gill-slit have been formed, and in which the left head-

THE FOUMATION OF THE LAYERS IN AMFHIOXUS. 345

cavity has acquired an opening to the exterior, slightly older than that
represented in fig. 16. In fig. 17 « the left head-cavity is seen opening
to the exterior. In fig. 17 b the collar-cavities are seen extending do^wn at
the sides of the pharynx, and the external opening of the clnh-shaped
gland is seen at club in front of the mouth. In fig. 17 c the mouth is seen,
and on the j)osterior extension of the right collar-cavity (r.coll.) an
ectodermal thickening (at.), the rudiment of the atriiil ridge. Figs.
1 7 d, e, /, fj, and h are five consecutive sections through the hinder end
of the pharynx hehind the gill-slit, which is grazed in fig. 17 d (br.end.)
to show how the extensions of the collar-cavities thin out and disappear,
whilst right and left .splanchnoceles meet in the mid-ventral line.
Magnification 1000 diameters.

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STEUCTUEE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 347

The Structure, Development, and Bionomics
of the House-fly, Musca domestica, Linn.

Part III. — The Bionomics, Allies, Parasites, and the Relations
of M. domestica to Human Disease.

By

Cior<loii Hewitt, D.Se.,

Late Lectui'er in Economic Zoology, University of Manchester.

With Plate 2±

CONTENTti.

PAGE

I. Introduction . . . . . . 348

II. Distribution ...... 34h

III. Flies occurring as Co-inhabitants of Houses with M.

domestica or as Visitants .... 351

IV. Physiology :

1. Influence of Food, Temperature, and Light . . 382

2. Hibeniation ..... 383

3. Flight ...... 384

4. Regeneration of Lost Parts . . 385

V. Natural Enemies and Occasional Parasites :

1. Chernes nodos\xs, Schrank . . . 387

2. Acarina or Mites borne by House-flies . . 38U

3. Fungal parasite — Empusa muscse, Cohn . . 371

VI. True Parasites :

1. Flagellata — Herpetonionas musca; - domestica) . 374

Crithidia nixisca;-domestica; . 379

2. Nematoda — Habronema uixxsca- . . . 380

3. Dissemination of Parasitic Worms . . . 382

VII. Dissemination of Pathogenic Organisms by M. domestica

and its non-Blood-sucking Allies :

1. Tyj)hoid Fever ..... 385

2. Anthrax ..... 394

3. Cholera ...... 398

4. Tuberculosis ..... 398

348

C. GORDON HEWITT.

PAGE

5. Ophthalmia

6. Plague .

7. Miscellanea

. 399
. 401
. 402
. 404
. 405
. 412

Vlll. Flies and Intestinal Myiasis

IX. Literature .

X. Appendix on the Winter Breeding of M. domestica

I. Introduction.

The present paper concludes this study of the structure,
development, and bionomics of Musca domestica (the
previous parts were published in 1907 and 1908). In it I
have described the bionomics, certain of its allies which may
occur in houses, its parasites, and its relation to man, especially
as the cari’ier of the bacilli of certain infectious diseases.

The last portion of the present paper, in which is described
what is known concerniug the ability of M. domestica and
its allies to carry and disseminate the bacteria of many impor-
tant diseases, shows, I hope, the grave character of its relation
to man. Although its importance in this respect is being
gradually realised in this country, it is not so widely recog-
nised as it should be. In the United States of America it is
proposed to change this insect’s name from the house-fly to
the “ Typhoid fly ” ; notwithstanding certain objections to this
name, it clearly indicates that more attention must be paid to
preventive measures, that is, they must be reduced by the
deprivation of suitable breeding-places. I have not discussed
in the present paper the relation of house-flies to infantile or
summer diarrhoea, chiefly because we are not yet certain as to
the speciflc cause, but this disease may be included for the
present under typhoid or enteric fever in so far as the relation
of flies with it is concerned.

I should like to take this opportunity of thanking those
medical men, whose names I mention later, for the kind
manner in which they have replied to my inquiries concern-
ing their observations on various diseases of which they have
special knowledge.

STRUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 349

II. Distribution.

Musca domstica is probably the most widely distributed
insect to be found ; the animal most commonly associated with
man, whom it appears to have followed over the entire globe.
It extends fi’om the sub-polar regions, where Linnaeus I’efers
to its occurrence in Lapland, and Finmark as “ rara avis in
Lapponia, at in Finmarchia Norwegias integras domos fere
replet,” to the tropics, where it occurs iii enormous numbers.
Referring to its abundance in a house near Para in equatoral
Brazil, Austen (1904) says: “At the mid-day meal they
swarmed on the table in almost inconceivable numbers,” and
other travellers in different tropical countries liave related
similar experiences to me, how they swarm round each piece
of food as it is carried to the mouth.

In the civilised aud populated regions of the world it occurs
commonly, and the British Museum (Natui’al History) collec-
tion and my own contain specimens from the following
localities. Certain of the localities have, in addition, been
obtained from lists of insect faunas :

Asia. — Aden; North West Provinces (India); Calcutta;
Madras; Bombay (it probably occurs over the whole of
India); Ceylon; Central China; Hong-Kong ; Shanghai;
Straits Settlements ; Japan.

Africa. — Port Said; Suez; Egypt; Somaliland; Nyassa-
land ; Uganda; British E. Africa; Rhodesia; Transvaal;
Natal; Cape Colony; Madagascar; Northern and Southern
Nigeria; St. Helena; Madeira.

America. — Distributed over North America; Brazil;
Monte Video (Uruguay); Argentine; Valparaiso; West
Indies.

Australia and New Zealand.

Europe and the isles of the Mediterranean; it is especially
common in Cyprus.

Not only is this world-Avide disti’ibution of interest, but its
distribution in our owu country is noteworthy. From observa-
tions that I have made during a number of years in town and

350

C. GORDON HEWITT.

suburban houses and country houses and cottages, I find that
in the former it is by far the commonest house-fly. But
whei’eas M. domes tica may be almost the only species in
warm places where food is present, such as restaurants and
kitchens, in other rooms of houses Homalomyia cani-
cularis, the small house fly, increases in proportion and
often predominates ; occasionally one may find it to be
commoner than M. domestica. In country houses the
proportions vary by the intrusion of Stomoxys calcitrans,
which I have often found to be the dominant species. In a
certain country cottage, out of the several hundreds captured,
S. calcitrans formed 50 percent, of the total, the rest being
chiefly H. canicular is together with An tho myia radicum,
whose lai’vae, as I have shown (1907), breed in horse-manure
with those of M. domestica. The following records taken
from a “ fly census ” that was made in 1907 may be taken as
illustrative of the proportional abundance of the different
species in different situations ; although the numbers of these
records are small the pi-oportions are more obvious.

M.

domestica.

H . c anicu-
laris.

Other species.

Restaurant, Manchester

18G9

14

2 (M. stabulans.

C. ery throcephala).

Kitchen, detached suh-

1

urban house (six records).

Lancashire

581

265

14

Kitchen, detached sub-

urban house in Manchester

682

7

14

Stable, suburban house

22

153

14

(12. S. calcitrans).

Bedroom, suburban house .

1

33

4 (M. stabulans).

Out of a total of 3856 flies caught in different situations,
such as i-estaurants, kitchens, stables, bedrooms and hotels,
87’5 per cent, were M. domestica, ITS per cent. H. canicu-
laris, and the rest were other species such as S. calcitrans,
Muscina stabulans, C. ery throcephala, and Antho-
myia radicum. These figures are comparatively small, but

STRUCTURE, DEVELOPMENT, AND RIONOMICS OE HOUSE-FLY. 351

are representative of the average occurrence, as I have
observed, of the different species.

For the proportional occurrence in similar localities we have
interesting figures given by Howard (1900) for the United
States. Of 23,087 Hies caught in rooms where food supplies
ai'e exposed he found that 22,808, or 98'8 per cent, of the
whole number, were M. domestica, and of the remaining
J‘2 per cent. H. canicularis was the commonest species.
Hamer (1908) found that more than nine tenths of the Hies
caught in the kitchens and “living-rooms” of houses in the
neighbourhood of depots for horse-refuse, manure, etc., were
M. domestica. In a further report Hamer gives more
details as to the different species that were found. In one
lot of 35,000 flies caught on four fly-papers exposed in similar
positions, 17 per cent. Avere Homalo my i a canicularis, less
than 1 per cent. wereC. ery throcephala, and considerably
less than 1 per cent, were Muscina stabulans, whereas of
nearly 6000 flies caught in another situation in four fly-
balloons 24 per cent. wereH. canicularis, 15 jrer cent, were
C. ery throcephala, and neaily 2 percent, were M. stabu-
lans. He gives an intei’esting diagram showing from counts
of flies the seasonal prevalence which 1 have previously
recorded from observation. The report shows how the pro-
portions of the different species vary in different situations
according to the substances and refuse that are present in
the locality. We may therefore say with certainty that
M. domestica is the commonest species of house-fly, aud
next to this II. canicularis, and that in country houses
8. calcitrans often occurs in large numbers, although
it is not a house-fly in the strict sense of the word.

III. Flies Occurring as Co-inhabitants of Houses with

M. DOMESTICA OR AS ViSITANTS.

We have seen from the preceding section that M. domes-
tica is by far the commonest species which occurs in houses,
and is, in fact, “domesticated” in the true sense of the word

352

C. GORDON HEWITT.

— Liunaeiis never selected a more truly specific title ;
nevertheless^ other species of closely allied Hies are found in
houses. These may be either co-inhabitants, that is, living in
houses, as in the case of H. canicular is and one or two
others to be mentioned subsequently, or they may be
visitants. The visitants normally lead an open-air life, but
sometimes, as in the case of Stomoxys calcitraus, they
spend a portion of their time in houses, when climatic condi-
tions are less favourable for out-door life. Such flies as the
blow-fly, or “blue-bottle,” Calli ph ora ery throcephala,
and its allies, enter houses only in search of suitable sub-
stances upon which to deposit their eggs. The appearance in
houses of certain flies, as, for example, Pollenia rudis, can
only be regarded as accidental, and the cause may be often
traced to the occurrence of climbing plants such as ivy or other
creepers on the walls of the house.

In India two species of flies closely allied toM. domestica
are found — M uscadomestica sub-sp. d e t e r m i n a t a Walker
and M. enteniata, both of which, on account of their close
resemblance to M. domestica and the similarity of their
breeding habits, are frequently mistaken for it.

(1) M. domestica sub-sp. deter mi nata Walker.

Th is Indian variety of the house-fly was described by
Walker (1856) from the East Indies. His description is as
follows : “ Black, with a hoary covering ; head with a white
covering; frontalia broad, black, narrower towards the
feelers ; eyes bare ; palpi and feelers black ; chest with four
black stripes ; abdomen cinereous, with a large tawny spot on
each side at the base ; legs black ; wings slightly grey, with
a tawny tinge at the base ; prtebrachial vein forming a very
obtuse angle at its flexure, very slightly bent inward from
thence to the tip; lower cross-vein almost straight; alulae
whitish, with pale yellow borders ; halteres tawny.”

In appearance and size it is very similar to M . domestica.
Its breeding habits are also similar. Aldridge (1904) states

STRUCTURE, DEVELOPMENT, AND P.IONOMICS OE HOUSE-FLY. 353

that at certain seasons of the year it is pi’esent in enormous
numbers. The method of disposal of the night soil is to bury
it in trenches about one foot or less in depth. From one
sixth of a cubic foot of soil taken from a trench at Meerut
and placed in a cage, 4042 flies were hatched. Lieut. Dwyer
collected 500 from one cage covering three square feet of a
trench at Mhow. Specimens in the British Museum collection
were obtained from the hospital kitchens, and Smith found
them in a ward at Benares.

They have also been recorded from tlie N.W. Provinces,
Kangra Valley (4500 feet), Dersa, and I have received speci-
mens from Aden.

(2) Musca enteniata Bigot.

This fly has a distribution somewhat similar to the last
species, aud like it, has a marked resemblance to M. domes-
tica, as Bigot’s (1887) description indicates :

“ Front tres etroit, les yeux, toutefois, sepnres. Antennis et
palpes noirs ; face et joues blanches; thorax noir avec trois
larges bandes longitudinales grises ; flancs grisatres, ecussou
noir avec deux bandes semblables ; cuillerons et balanciers
d’un jaunfitre tres pale ; abdomen fauve, avec une bande
dorsale noir et quelques reflets blancs ; pieds noirs ; ailes
hyalines; cinquieme nervure longitudinal (Rondin) coudee
suivant un angle legerement arrondi, ensuite un pen concave;
deuxieine transversale (I’extreme) presque perpeudiculaire,
legerement bisinueuse, soudee a la cinquieme longitudinale, a
egale distance du coude et de la premiere nervure transversale
(I’interne).”

M. enteniata measures 4 to 5 mm. in length. The British
Museum collection contains specimens sent by Major F. Smith
from Benares, with these notes : “ Bred from human ordure ;
hospital ward fly ; at an enteric stool ; bred from cow-dung
fuel cakes.” 1 have received specimens from Suez and
Aden, and it is recorded as breeding in human excrement in
Khartoum (Balfour, 1908) and in stable refuse, as also M

354

C. CiOUDON HP^WITT.

domestica and M. corvina. It will be seen, therefore,
that its breeding habits are very similar to those of M.
domestica and the sub-species determinata. It is in-
teresting and important to note tlie rather exceptional choice
of cow-dung as a breeding-place.

(3) Homalomyia canicularis. L.

This species of fly (see ‘Quart. Journ. Micr. Sci.,’ vol. 51,
PI. 22, fig'. 3) is often mistaken by the uninitiated for M.
domestica which are not full grown. Although it may be
called the small or lesser house-fly its differences from M.
domestica are great, as it belongs to a different group of
calypterate Muscidae, namely, the Anthomyidas. One of the
chief distinguishing features of this group is that the fourth
longitudinal vein of the wing (M. 1 -t- 2) goes straight to the
margin of the wing and does not bend upwards at an angle
as in M. domestica.

The male of H. canicularis differs from the female in
some respects. In the male the eyes are close together, and
the frontal region is consequently very narrow ; the sides of
this, these are the inner orbital regions, are silvery white,
separated by a narrow black frontal stripe. In the female
the space between the inner margins of the eyes is about one
third of the width of the head ; the frons is brownish black,
and the inner orbital regions are dark ashy grey. The bristle
of the antenna of H. canicularis is bare; in M. domes-
tica, it will be remembered, the bristle bears a row of setae
on its upper and lower sides. The dorsal side of the thorax
of the male is blackish grey with three rather indistinct longi-
tudinal black lines. In the female it is of a lighter gi'cy, and
the three longitudinal stripes are consequently more distinct.
The abdomen of the male H. canicularis is narrow and
tapering compared with that of M. domestica. It is bronze
black in colour, and each of the three abdominal segments
has a lateral translucent area, so that when it is seen against
the light, as on a window-pane, three, and sometimes four,
pairs of yellow ti'anslucent areas can be seen by the trans-

STRUCTURE, DEVELOPMENT, AND BIONOMICS OP HOUSE-PLY, 355

mitted light. In the female the abdomen is short in propor-
tion to its length, and is of a greenish or brownish-grey colour
H. canicularis appears in houses before M. domestica,
and can be found generally in May and June. In the latter
month its numbers are swamped, as it were, by M . domes-
tica, and it appears to seek the other rooms of a house than
the kitchen, although I have found it frequently in consider-
able numbers in kitchens. The average length is 5'7 mm.

The larva of H. canicularis (PI. 22, fig. 1) is very
distinct from that of M. domestica, as will be seen fi-om
the figure. It is compressed dorso-ventrally, and has a
double row of processes on each side. Owing to the rough
and spinous nature of these processes dirt adheres to the
larva and gives it a dirty-brown appearance. The full-grown
larva measures 5-6 mm. in length. The breeding habits of
H. canicularis are very similar to those of M. domestica.
The larva) feed on waste vegetable substances and also on
various excremental products, but particularly, I have found,
on human excrement, for which they show a great partiality.
I have frequently found excrement in privy middens to be a
moving mass of the larvae of H. canicularis. The larval
period is from three to four weeks, and the insect spends
fourteen to twenty-one days in the pupal stage.

(4) Homalomyia scalar is F.

Newstead (1907) has found this species occurring as a
house-fly. It is slightly larger than, though similar in many
respects to, H. canicularis. The larva is very similar in
appearance. Newstead found the larva) in ash-pit refuse,
and bred the flies from human faeces. The larvae have been
found frequently to be the cause of intestinal myiasis.

(5) Anthomyia radicum Meigen.

This member of the Anthomyidm has been found in houses,
especially those in or near the country. The female has been
illustrated already (Part I, ‘Quart. Jonrn. Micr. Sci.,’ vol. 51,
PI. 22, fig. 2). The male is darker in colour, the dor.sal side

r. CORDON HEWITT.

85f)

of the tliorax being blackish with three black longitudinal
stripes; the frontal region is very narrow; the abdomen is
grey with a dark median stripe. The average length of the
body is 5 mm.

In the summer they are common and may be found in the
neighbourhood of manure. The eggs are laid in this substance,
especially in horse-manure. The Larvse have also been
found feeding on the roots of various cultivated cruciferous
plants, from which the insect has derived the name “root-
maggot.” The eggs hatch out from eighteen to thirty-six
hours after deposition. 'J'he first larval stadium lasts twenty-
four hours, the second forty-eight hours, and five days later
the larva changes into a pupa, the whole larval life occupy-
ing about eight days. The pupal stage lasts ten days, so that
in warm weather the development may be completed in nine-
teen to twenty days. The full-grown larvte measure 8 mm.
in length, and may be distinguished by the tubercles sur-
rounding the caudal extremity. In this species there are six
pairs of spinous tubercles surrounding the posterior end and
a seventh pair is situated on the ventral surface posterior to
the anus. The tubercles of the sixth pair, counting fi’om the
dorsal side, are smaller than the rest and are bifid. The
arrangement of the tubercles can be seen in fig. 2. The
anterior spiracular processes (fig. 3) are yellow in colour and
have thirteen lobes.

(6) Stomoxys calcitrans Linn.

The species is common, especially in the country from
July to October, and during these months it may be often
found in houses, although Hamer’s observations (1908)
appear to indicate that the presence of cowsheds, in which
they occur in large numbers, does not affect their numbers
in houses. I have found 8. calcitrans in large numbers
in the windows of a country house in March and April,
and it may be found frequently out of doors on a sunny
dav in May, and throughout the ensuing summer months.
It is normally an outdoor insect, but appears to seek the shelter

STRUCTIJKE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 357

of houses, especially during wet weather, from which habit it
has no doubt derived the popular name of “ storm-fly ” ; it is
also know as the “stable-fly.” As these names may be equally
applicable to certain other Diptera they should be discarded.

As I have already mentioned this species is frequently mis-
taken by the public for M. domestica, which is supposed to
have adopted the biting habit, although the latter is unable
to inflict the slightest prick. IE examined side by side the
sreat differences between the two will be seen readily (see
Part I in ‘Quart. Journ. Micr. Sci., vol. 51, PI. 22, _fig. 4).
S. calcitrans has an awl-like proboscis for piercing- and
blood-sucking ; this projects horizontally forward from
beneath the surface of the head (fig. 4). It is slightly
larger and more robust than M. domestica; the bristles of
the antennae bear setm on their upper sides only. The colour
is brownish with a greenish tinge ; the dorsal side of the
thorax has four dark longitudinal stripes, the outermost pair
being interrupted. At the anterior end of the dorsal side of
the thorax the medium light-coloiu-ed stripe has a golden
appearance, which is vei-y distinct when the insect is seen
against the light. The abdomen is broad in proportion to its
length, and each of the large second and third segments has
a single median and two lateral brown spots; there is also
a median spot on the fourth segment.

The life-history of S. calcitrans has been studied by
Newstead (1906), and I have been able to confirm his observ'a-
tions during 1907 and 1908. From fifty to seventy eggs,
measuring 1 mm. in length, ai-e laid by the female. The eggs
are laid on warm, decaying vegetable refuse, especially in
heaps of fermenting grass cut from lawns ; I have frequently
confirmed this observation of Newstead’s. The eggs are also
deposited on various exci-emental substances upon which the
larvc© feed. Osborne (1896) reared them in horse-manm-e ;
Howard (1900) states that they live in fresh horse-manure,
and records their occurrence in outdoor privies in some
localities; Newstead reai-ed them in moist sheep’s dung; they
can also be reared in cow-dung.

358

(\ am? DON m<;wiTT.

Tlie larvae are creamy-white in colour and have a shiny,
translucent apj)earance. They are rather similar to those of M.
domestica, but can be distinguished by the character of the
posterior spiracles. These (fig. 5 and 6) are wider apart than
in M. domestica and are triangular in shape with rounded
corners; each of the corners subtends a space in which a
sinuous aperture lies. The centre of the spiracle is occupied
by a circular plate of chitin. The anterior spiracular pro-
cesses are five-lobed. Under warm conditions Newstead
found that the egg state lasted from two to three days ;
the larval stage lasts from fourteen to twenty-one days
and the pupal stage nine to thirteen days. There are
three larval stages. The whole life-history may be complete
in twenty-five to thirty-seven days. Some specimens passed
the winter in the pupal state.

Although S. calcitrans does not frequent to such a great
extent asM. domestica material likely to contain pathogenic
intestinal bacilli, on account of its blood-sucking habits, which
cause it to attack cattle and not infrequently man, it may
occasionally transfer the anthrax bacillus, as many have
believed, and give rise to malignant pustule, etc.

(7) Calliphora erythrocephala Mg.

This is the commoner of the two English blow-flies or
“blue-bottles.” The other species, Calliphora (Musca)
vomitoria, is less common, although the name is frequently
given to both species indiscriminately. They can be dis-
tinguished, however, by the fact that in C. erythrocephala
the genfe are fulvous to golden-yellow and are beset with
black hairs, whereas in C. vomitoria the genm are black
and the hairs are golden-red.

The appearance of C. erythrocephala is sufficiently well
known with its bluish-black thorax and dark metallic blue
abdomen. Its length varies from 7 to 13 mm. The laiwae
are necrophagous. The flies deposit their eggs on any fresh
or decaying meat, nor is such flesh always dead. On one
occasion, when obtaining fresh material in the form of wild

STRUCTURE, DEVELOPMENT, AND P.IONOMTCS OE HOUSE-FLY. 359

rabbits upon wbicli to rear the larvse of C. ery throcephala,
I found the broken leg of alive rabbit, which had been caught
in a spring trap set the previous evening, a living mass of
small larvae, which were devouring the animal while it was
still alive. An enormous number of eggs are laid by a single
insect; Portchinski (' Osten. Sacken,’ 1887) found from 450
to 600 eggs, though I have not found so many. With an
average mean temperature of 23° C. (73‘5° F.) and using fresh
rabbits as food for the larvae, the following were the shortest
times in which I reared C. ery throcephala. The eggs
hatched from ten to twenty hours after deposition. The larv^
underwent the first ecdysis eighteen to twenty-four hours after
hatching ; the second moult took place twenty-four hours later,
and the third larval stage lasted six days, the whole larva life
being passed in seven and a half to eight days. Foui’teen days
were spent in the pupal state ; thus the development was com-
plete in twenty-two to twenty-three day’s. I have no doubt
that this time could be shortened by’ the presence of a very
plentiful supply of food, as an enormous amount, comparatively,
is consumed.

The full-grown larva may measure as much as 18 mm. in
length. The posterior extremity is suiTOunded by^ six pairs
of tubercles arrauged as shown in the figure (fig. 12) ; there
is also a pair of anal tubercles. The anterior spiracular
(fig. 11) processes are nine-lobed. The posterior spiracles
(fig. 10) are circular in shape and contain three slit-like
apertures. In the second larval instar (fig. 9) there are only
two slits in each of the posterior spiracles, and in the first
larval instar (tig. 8) each of the posterior spiracles consists of
a pair of small slit-like orifices. Howard (1900) found the fly
on fresh human faeces, and Riley records it as destroying the
Rocky Mountain locust.

C. ery throcephala is an outdoor fly, but frequently enters
houses in search of material upon which to deposit its eggs
and also for shelter. From its habit of frequenting faeces,
which may be observed in this country especially in-insanitary
court-yards, and such food as meat and fruit, it is not improb-
VOL. 54, PART 3. NEW SERIES. 26

0. (JOnnON IfEWITT.

8C)0

able that it occasionally may bear intestinal bacilli on its
appendages or body and thus carry infection. Its flesh-seek-
ing habits may also render it liable to carry the bacilli of
anthrax should it have access to infected flesh.

(8) Muscina (Cyrtoneura) stabulans Fallen.

This common species is frequently found in and near houses.
I have usually found it occurring with H . canicularis in the
early summer (June) before M. domestica has appeai’ed in
any numbers. It is larger than M. domestica, and more
robust in appearance. Its length varies from 7 to nearly
10 mm. Its general appearance is grey. The head is
whitish-grey with a “ shot ” appearance. The frontal region
of the male is velvety black and narrow ; that of the female
is blackish-brown, and is about a third of the width of the
head. The bristle of the antenna bears set^ on the upper
and lower sides. The dorsal side of the thorax is g’rey and
has four longitudinal black lines ; the scutellum is grey. The
abdomen, as also the thorax, is really black covered with
grey; in places it is tinged with brown, which gives the
abdomen a blotched appearance. The legs are rather slender,
and are reddish-gold or dirty orange and black in colour.

The eggs are laid upon the following substances, on which
the hirvm feed : Decaying vegetable substances such as fungi,
fruit, cucumbers, decaying vegetables, and they sometimes
attack growing vegetables, having been introduced probably
as laivtB with the manure, as they also feed on rotting dung
atid cow-dung. Howard (I. c.) found the fly frequenting
him an excrement, and observed the species breeding in the
same. In the United States it has been reared from the pupm
of the cotton-worm and the gipsy moth ; Riley was of the
opinion that in the first case it fed on the rotten pupm only.
In 1891 it was also reared on the masses of larva) and pupm
of the elm-leaf beetle. Other observers record it as being
reared from the pupa) of such Hymenoptei’a as Lophyrus.
In all these cases of its occurrence in the pupae of insects, it

STRUCTUKK, DEVELOPMENT, AND BIONOMICS OP HOUSE-FLY. 361

is difficult to say whether it is parasitic or whether it feeds on
the rotting pupae only ; many observers are inclined to take
the last view. The larva may reach a length of 11 mm. It
is creamy-white in colour; the anterior spiracular processes
are five-lobed and ai’e like hands from which the fingers have
been amputated at the first joint. The posterior spiracles
are rounded and enclose three triangular-shaped areas, each
containing a slit-like aperture. I have not been able to study
the complete life-history, but Taschenberg (1. c.) states that it
occupies five or six weeks.

(9) Lucilia Cmsar L.

Although it is not a house-fly, this common fly occasionally
occurs in houses, especially those in the country, and it is often
called a “blue-bottle.” It is smaller than C.erythrocephala
and is more brilliant in colouring, being of a burnished gold,
sometimes bluish, and also of a shining green colour.

It frequents the excrement of man and other animals in
which it is able to breed. Howard (1. c.) reared it from human
excrement. It also breeds in carrion, but the chief breeding-
place in which I have found it in this country is on the backs
of sheep. It is one of the destructive species of “ maggots”
of sheep. The larvae ai’e very similar to those of C. erythro-
cephala — in fact, Portchinski considered them indistinguish-
able from the larvae of the latter except in size. The full-
grown larva measures 10-11 mm. in length. The larval life
lasts about fourteen days, and the pupal stage a similar length
of time, but I have reason to believe that under very favour-
able conditions development may take place in a much
shorter time.

(10) Psychoda spp.

There may be found frequently on window-panes small,
grey, inoth-like flies belonging to the family Psychodidse.
The wings of these small flies are large and broad in propor-
tion to the size of the body, and are densely covered with
hair; when the insect is at rest they slope in a roof-like

:3G2

('. aORDOX IfEWITT.

manner. The larvjB of some species breed in human and other
excrement, others breed in decaying vegetable substances,
while certain species breed in water, especially when polluted
Avith sewage, and these aquatic species have the spiracular
apparatus modified accordingly. Although a form, Phlebo-
tamus, which occurs in Southern Europe, has blood-sucking
habits, the British species have no such annoying habits, and
are of little importance in their relation to man.

IV. Physiology.

1. The Influence of Food, Temperature, and Light.

Food. — Mention has already been made in the second part
of this work of the influence of food on the development of the
laiwte; the experiments which Avere carried out showed that
the larvte develop more rapidly in certain kinds of food, such
as horse-manure, than in others. It has yet to be discovered
Avhat are the chemical constituents Avhich faA'our the more
rapid development. It Avas found that insufficient food in the
larval state retarded development and produced flies which
Avere subnormal in size. BogdanoAV (1908), in an interesting
experiment, fed M. domestica through ten generations on
unaccustomed food such as meat and tanacetum in different
proportions, and he found that the resulting flies did not show
any change.

Temperature. — The influence of temperature on the
development of the larvae has been shown also. A high
temperature accelerates the development of the egg, larva
and pupa. Temperatiu’e also affects the adult insect; they
are most active at a high summer temperature, and cold
produces an inactive and torpid condition. They are able,
hoAvever, to withstand a comparatively Ioav temperature.
BachmetjeAv (1906) Avas able to submit M. domestica to as
loAV a temperature as — 10° C., and vitality Avas retained, as
they recovei’ed Avhen brought into ordinary room temperature.
Uonhoff (1872) performed a number of experiments previous

STEUOTUEE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 363

to this with interesting results. He submitted M. domes-
tic a for five hours to a temperature of - 1'5° C., and they
continued to move. Exposed for eight hours to a temperature
of first — 3° C. and then — 2° C. they moved their legs. On
being submitted for twelve hours to a temperature first of
— 3'7° C. and then — 6‘3° C., they appeared to be dead, but
on being warmed they recovered. When exposed for three
hours to a temperature of — 10° C. which was then raised
to — 6° C., they died. These experiments show that M.
domestica is able to withstand a comparatively low degree
of temperature.

Light. — The female of M. domestica deposits the eggs
in dark crevices of the substance chosen for the larval nidus
and as far away from the light as possible. Bedard (1858)
showed that the eggs develop more quickly under blue and
violet glass than under red, yellow, green, or white. The
larva) are negatively heliotropic, as Loeb (1890) has also
proved in the larva) of the blow-fly. As I have pi'cviously
shown, the distinction between light and darkness is probably
appreciated by the larvae by means of the sensory tubercles
of the oral lobes.

2. Hibernation.

'I’his (piestion is intimately connected with the preceding
physiological facts. The disappearance of the flies towards
the end of October and in November is a well-known fact,
and an endeavour to discover the reason for this has been
made in the present investigation.

I have found that the majority of flies observed were killed
off by the fungus Enipusa muscae Cohn which is described
in the ])resent paper. Of the remainder some hibeinate and
some die naturally. This natural death may be compared,
I think, to the like phenomenon that occurs in the case of
the hive-bee Apis mellifica, where many of the workers
die at the end of the season by I’eason of the fact that they
are simply worn out, their function having been fulfilled.
The flies which die naturally have probably lived for many

364

C. GORDON HEWITT.

weeks or mouths during the summer and autumn, and in the
case of the females have deposited many batches of eggs;
their life work, therefore, is complete. Those flies which
hibernate are, I believe, the most recently emerged, and
therefore the youngest and most vigorous. On dissection it
is found that the abdomens of these hibernating individuals
are packed with fat cells, the fat body having developed
enormously. The alimentary canal shrinks correspondingly
and occupies a very small space; this is rendered possible by
the fact that the fly does not take food during this period.
In some females it was found that the ovaries were very well
developed, while in others they were small, and mature
spermatozoa were found in the males. Like most animals in
hibernating, M. domestic a becomes negatively heliotropic
and creeps away into a dark place. In houses they have been
found in various kinds of crevices such as occur between the
woodwork and the walls. A favourite place for hibernation
is between wall-paper which is slightly loose and the wall.
A certain number hibernate in stables, where, owing to the
warmth, they do not become so inactive, and they emerge
earlier at the latter end of spring. During the winter the
hibernating flies are sustained by means of the contents of
the fat body, which is found to be extremely small in hiber-
nating flies if dissected when they first emerge in May and
June. The abdominal cavity is at first considerably decreased
in size, but the fly begins to feed and soon the alimentary
tract regains its normal size, and, together with the develop-
ment of the reproductive organs, causes the abdomen to
regain its normal appearance. The emergence from hiber-
nation appears to be controlled by temperature, as one may
frequently find odd flies emerging from their winter quarters
on exceptionally warm days in the eai’ly months of the year
(see Appendix).

3. Flight.

The distance that M. domes tic a is able to fly is one of
practical importance in connection with their breeding habits

STRUCTURE, ])F]V£LOFMENT, AND BIONOMICS OF HOUSE-FLY. 365

and disease-genn-can-ying powers. Normally they do not fly
great distances. They may be compared to domestic pigeons
which hover about a house and the immediate neighbourhood.
On sunny days they may be found in lai’ge numbers out-of-
doors, but they retire into the houses when it becomes dull or
rains. They are able to fly, however, a considerable distance,
and can be carried by the wind. A few years ago, when
visiting the Channel Islands, I found ]\[. domestica from
ItV to 2 miles from any house or any likely breeding-place,
so far as I was able to discover. Dr. M. B. Arnold has
made some exact experiments at the Monsall Fever Hospital,
Manchester, on the distance travelled by flies.' Three hundred
flies were captured alive, aiad marked with a spot of white
enamel on the back of the thorax. These were liberated in
fine weather. Out of the 300 five were recovered in fly-traps
at distances varying from 30 to 190 yards from the place of
liberation, and all the recoveries were within five days.
1\I. domestica is also able to fly at a considerable height
above ground, and I have found them flying at an altitude of
80 feet above the ground. Such a height would greatly
facilitate their carriage by the wind.

4. Regeneration of Lost Parts.

If the wings or legs of M. domestica are broken off they
do not appear to be able to regenerate the missing portions,
as in the case of some insects, notably certain Orthoptera.
Kanimerer (1908), however, experimenting with M. domes-
tica and C. vomit or ia, has found that if the wing is
extirpated from a recently pupated fly it is occasionally
regenerated. The new wing is at first homogeneous, and con-
tains no veins, but these appear subsequently.

' Recorded on p. 262 of the ‘ RepoiT on the Health of the City of
Manchester for 11*06,' by Janies Niven, 1007.

366

C. GORDON HEWITT.

V. Natural Enemies and Occasional Parasites.

The most important of all the natural enemies of M.
domestica is the parasitic fungus Empusa muscae, which
will be described here ; this is the most potent of the natural
means of destruction. Of animals, apart from the higher
animals such as birds, spiders probably account for the
greatest number, though owing to the uormally clean con-
dition of the modern house these enemies of the house-fly are
refused admittance. I have been unable to rear any insect
parasites, such as ichneumons, from M. domestica. Their
life indoors and the cryptic habits of the larvm no doubt save
them from the attacks of such insects; but Packard (1874)
records the occurreuce of the pupa of what was probably a
Dermestid beetle, which he figures; this was found in a pupa
of M. domestica. Predatory beetles and their larvaB pro-
bably destroy the larvm, and Berg (1898) states that a species
of beetle, Trox suberosus F., known as “ Champi ” in
S. America, is an indirect destructor of the common fly. I
have frequently observed the common wasp, Vespa ger-
man ic a, seize M. domestica and carry it away. In some
places in India it is the custom, so I have been told by resi-
dents, to employ a species of Mantis, one of the predatory
“ praying insects,” to destroy the house-flies.

In view of the fact that the Arachnids Chernes nodosus
and the species of Gamasid are occasionally found actually
attached in a firm manner to M. domestica, they will be
described under this head, but it must be clearly understood
that it is still an open question whether they are external
parasites in the true sense of the word, or whether M. domes-
tica, instead of being the host, is merely the transporting
agent as it appears to be in the majority of cases. For the
present they may be termed for convenience “ occasional
parasites,” in view of the fact that they have been found
occasionally feeding upon M. domestica.

STEUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 367

1. Ch ernes nodes us Schrank.

Thei’e are frequently found attached to tlie legs of the
house-fly small scorpion or lobster-like creatures which are
Arachnids belonging to the order Pseu do-scorpi oni dea;
the term “chelifers^’ is also applied to them on account of the
large pair of chelate appendages which they bear. The
species which is usually found attached to M. domes tic a is
Cher lies iiodosus Schrank (fig. 13). It is very widely
distributed, and my observations agree with those of Pickard-
Cambridge (1892), who has described the group.

The species is 2‘5 mm. in length and Pickard-Caiubridges’s
description of it is as follows :

“ Cephalothorax and pal[)i yellowish red-brown, the former
rather duller than tlie latter. Abdominal segments yellow-
brown ; legs paler. The caput and first segment of the
thorax are of equal width (fi-om back to froiit) ; the second
segment of the thorax is very narrow. The surface of
the cephalothorax and abdominal segments is very finely
shagreeued, the latter granulose on the sides. The hairs on
this part as well as on the palpi and abdomen are simple, but
obtuse. The palpi are rather short and strong. 'I’lie axillary
joint is considerably and somewhat subcouically protuberant
above as well as protuberant near its base underneath. The
humeral joint at its widest part, behind, is consideiably
less broad than long ; the cubital joint is very tumid on its
inner side ■, the bulb of the pincers is distinctly longer, to the
base of the fixed claw, than its width behind ; and the claws
are slightly curved and equal to tlie bulb in length.”

They appear to be commoner in some years than in others.
Godfrey (1909) says: “The ordinary habitat of Ch. nod os us,
as Mr. Wallis Kew has pointed out to me, appears to be among
refuse, that is, accumulations of decaying vegetation, manure-
heaps, frames and hot-beds in gardens. He i-efers to its occur-
rence in a manure-heap in the open air at Lille, and draws my
attention to its abundance in a melon-frame near Hastings in
1898, where it Avas found by Mr. W. 11. Butterfield.” In

368

C. GOKDUN HEWITT.

view of tliese facts it is not ditiicult to understand its frequent
occurrence on the legs of flies, which may have been on the
rubbish heaps either for the purpose of laying eggs, or, what
is more likely, because they have recently emerged from
pupae in those places and in crawling about, during the pro-
cess of di’ying their wings, etc., their legs were seized by the
C. nodosus.

The inter-relation of the Chernes and M. domestica,
however, is one of no little complexity; much has been
written and many diverse views are held concerning it. An
interesting historical account of the occurrences of these
Arachnids on various insects has been given by Kew (1901).
Three views are held in explanation of the association and
they are briefly these : First, that the Chernes, by clinging
passively to the fly, uses it as a means of transmission and
distribution; second, that the Arachnid is predaceous; and
third, that it is parasitic on the fly. Owing to the unfortunate
absence of convincing experimental proof in favour of either
of the last two opinions, it is practically impossible to give
any definite opinion as to the validity of these views ; never-
theless they are worthy of examination.

The dispersal theory was held by Pickard- Cambridge and
Moniez (1894). Whether the other views are held or not
there is no doubt that such an association, even if it were
only accidental, would result in a wider distribution of the
species of C he rues, as the flies are constantly visiting fresh
places suitable as a habitat for the same. Except in one or
two recorded cases the Arachnids are always attached to the
legs of the fly, the chitin of which is hard and could not be
pierced, a fact which is held in support of this theory as the
only explanation of the association.

The parasitic and predaceous views are closely related.
The Pseudo-scorpionidea feed upon small insects, which
they seize with their chela). It is suggested by some that
the Chernes seizes the legs of the fly without I’ealising the
size of the latter. Notwithstanding its size, however, they
remain attached until the fly dies and then feed upon the

STROCTUUE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 369

body. In some cases as many as ten of the Arachnids have
been found on a single Hy, and if the movements of the
insect are impeded by the presence of a number of the
Chernes it will be easily understood that the life of the Hy
will be curtailed thereby. Pseudo-scorpion idea have been
observed feeding on the mites that infest certain species of
Coleoptera, and it has been suggested that they associated
with the flies for the same purpose, although I do not know
of any recorded case of a fly infested with mites carrying
Chernes also. If this were the case the Chernes would be
a friend and not a foe of the Hy, as Hickson (1905) has
pointed out.

There are few records to support the view that the Chernes
is parasitic on the fly. Donovan (1797) mentions the occur-
rence of a pseudo-scorpionid on the body of a blow-fly, and
Kirby and Spence (1826) refer to their being occasionally
pai’asitic on flies, especially the blow-fly, under the wings of
which they fix themselves. It is probable that the Chernes
seldom reaches such a position of comparative security on the
thorax of the fly ; should it succeed in doing so, however, it
could become parasitic in the true sense of the word. As I
have previously pointed out, little experimental evidence is at
present available and further iiivestigation is necessary before
it is possible to maintain more than a tentative opinion with
regard to this association between the Chernes and the
fly. It is obvious that the association will result in the dis-
tribution of the Pseudo-scorpionid, but whether this is
merely incidental and the real meaning lies in a parasitic or
predaceous intention on the part of the Arachnid, as some of
the observations appear to indicate, further experiments alone
will show.

2. Acarina or Mites borne by House-flies.

As early as 1735 de Geer observed small reddish Acari in
large numbers on the head and neck of M. domes tic a.
They ran about actively when touched. The body of this
mite was oval in shape, completely chitiuised, and polished;

730

C. GORDON HEWITT.

tlie dorsal side was couvex and the ventral side flat. Linnfeus
(1758) called this mite Acarus muse arum from de Geer’s
description^ and Geoffrey (1764) found what appears to be
the same, or an allied species of, mite, which he called the
‘‘brown fly-mite.” Murray (1877) describes a form, Trom-
bidium parasitic u m,^ which is a minute blood-red mite
parasitic on the house-fly. He says : “ In this country they
do not seem so prevalent, but Mr. Eiley mentions that in
North America, in some seasons, scarcely a fly can be caught
that is not infested with a number of them clinging tenaciously
round the base of the wings.” As it only possessed six legs
it was doubtless a larval form.

Anyone who has collected Diptera as they have emerged
from such breeding-places as hot-beds, rubbish and manure
heaps will have noticed the frequently large number of these
insects which are to be found carrying immatui’e forms of the
Acari. These are being transported merely by the flies in
the majority of cases. Mr. Michael tells me that he used to
call such flies “the emigrant waggons ” — a very descriptive
term. Many of these mites belong to the group Gamasidm —
the super-family Gamasoidea of Banks (1905). These mites
have usually a hard coriaceous integument. In shape they are
flat and broad and have rather stout legs. Sometimes imma-
ture forms of these mites swarm on flies emerging’ from rubbish
heaps. Banks holds the opinion that they are not parasitic,
but that the insect is only used as a means of transportation.
It is difficult to decide whether this is so in all cases. I have
illustrated (fig. 14) a specimen of the small house-fly, H.
canicularis, caught in a room ; on the under-side of the fly’s
abdomen a number of immature Gamasids" are attached,

' This species was named Atoma parasiticnm and later Astonia
parasiticnni l>y Latreille (‘ Magazin Encyclopedi(pie.' vol. iv, p. 15.
17!15). Mr. A. D. Michael tells me that the. genus was founded on
Tromhidium parasiticnm of de Geer. They were really larval
Tromhidiida' and Atoma was founded on larval characters ; probably
any larval Tromljidium came under the specific name.

“ Being unable to identify these immature specimens I submitted
them to Mr. Michael, who kindly informs me that it is extremely diffi-

STRUCTURE, DEVELOPMENT, AND P-inNOMTCS OF HOUSE-FLAL 871

apparently by their stomal regions. These specimens may be
truly parasitic, as I am inclined to believe, since many Acari
are parasitic in the immature state, although the adults may
not be so; on the other hand this form of attachment may be
employed as a means of maintaining’ a more secure hold of
the transporting insect.

8. Fungal Disease — Empusa muscae Cohn.

Towards the end of the summer large numbers of Hies may
be found attached in a rigid condition to the ceiling', walls or
window-panes. They have an extremely life-like appearance,
and it is not until one examines them closely or has touched
them that their inanimate, so far as the life of the Hy is con-
cerned, condition is discovered. These flies have been killed by
the fungus Empusa muscae Cohn, and in the later stages of
the disease its fungal nature is recognised by the fact that a
white ring of fungal spores may be seen around the fly on the
substratum to which it is attached. 'I'he abdomen of the fly
is swollen considerably, and white masses of sporogenous
fungal hypha) may be seen projecting for a short distance
from the body of the fly, between the segments, giving the
abdomen a transversely striped black and white appearance.

The majority of flies which die in the late autumn — and it
is then that most of the flies which have been present during
the summer months perish — are killed by this fungus. Its
occurrence, therefore, is of no little economic value, especially
if it were possible to artificially cultivate it and destroy the
flies in the early summer instead of being compelled to wait
until the autumn for the natural course of events.

Empusa muscae belongs to the grouj) Entomophthoreae,
the members of which confine their attacks to insects, and in
many cases, as in the case of the pi'esent species, are produc-
tive of great mortality among the individuals of the species of

cult to identify ininiature Ganiasids owing to the scarcity of knowledge
as to their life-histories, hut he says that they are very like Dinycliella
asperata Berk

372

C, GOKDOX HEWITT.

insect attacked. In this country it may be found from about
the beginning of July to the end of October, and usually
occurs indoors. It appears to be very uncommon out-of-
doors. A case has been recently recorded^ of its occurrence
on Esher Common, where it had attacked a species of Syrphid,
^lelanostomum scalare Fabr. Thaxter (1888) also
mentions two cases of its occurrence out-of-doors in America,
in both of which cases it had attacked, singularly enough,
species of Syrphidte. This author states that Empusa
muscie is probably the only species which occurs in flowers
attractive to insects, but he only observed it on the flowers of
Solidago and certain Umbellifereae.

The development of this species was studied by Brefeld
(1871). An Empusa spore which has fallen ou a fly rests
among the haii’s covering the insect’s body and there adheres.
A small germinating hypha develops, which pierces the
chitin, and after entering the body of the victim penetrates
the fat-body. In this situation, which remains the chief
centre of development, it gives rise to small spherical struc-
tures which germinate in the same manner as yeast cells,
forming gemmae. These separate as they ai’e formed, and
falling into the blood sinus ai’e carried throughout the whole
of the body of the fly. It was probably these bodies that
Colin (1855) found, and he explained their pi’esence as being
due to spontaneous generation ; he believed that the fly first
became diseased and that the fungus followed in consequence.
After a period of two or three days the fly’s body will be
found to be completely penetrated by the fungus, which
destroys all the internal tissues and organs. The whole
body is filled with the gemmae, Avhich germinate and produce
ramifying hyphae (fig. 15). The latter pierce the softer
portions of the body-wall between the segments and produce
the short, stout conidiophores (c.), which ai’e closely packed
together in a palisade-like mass to form a compact white
cushion of conidiophores, wliich is the transverse white ring
that one finds between each of the segments of a diseased, and
‘ ‘ Trans. Ent. Soc, London.' 1908 (“ Proceedings,” p. 57).

STRUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 378

consequently deceased, fly. A conidium now develops
(fig. 16) by the constriction of the apical region of the
conidiophore. When it is ripe the conidium (fig. 17) is
usually bell-shaped, measuring 25-30 n in length ; it generally
contains a single oil-globule (o.y.). In a remai’kable manner it
is now shot off from the conidiophore, often for a distance of
about a centimetre, and in this way the ring or halo of white
spores, which are seen around the dead fly, are formed. In
some cases, although I find that it is not an invariable rule as
some would suggest, the fly, when dead, is attached by its
extended proboscis to the substratum. Griard (1879) found
that blow-flies killed by Entomophthora calliphora were
attached by the posterior end of the body. If the conidia,
having been .shot off, do not encounter another fly, they have
the power of producing a small conidiophore, upon which
another conidium is in turn developed and discharged. If
this is unsuccessful in reaching a fly a third conidium maybe
produced, and so on. By this peculiar arrangement the
conidia may eventually travel some distance, and it is no
doubt a gi-eat factor in the wide distribution of the fungus,
once it occurs. On the fly itself short conidiophores may be
found producing secondary conidia.

Keproduction by conidia appears to be the only form of
generation, as we are still uncertain as to the occurrence of a
resting-spore stage in this species. Winter (1881) states
that he found resting-.spores in specimens of M. domestica
occurring indoors; they also produced conidia which he
identified as E. muse®. These azygospores measured
30-50 n in diameter, and were produced laterally or termin-
ally from hyph® within the infected fly. Giard (1. c.) describes
resting spores which were produced externally and on
specimens found in cool situations. Brefeld, however, is of
the opinion that E. muse® does not produce resting-spores.
The question of the production of resting-spores needs further
investigation, as it is one of some importance. In the absence
of confirmatory evidence it is extremely difficult to understand
how the gap in the history of the Empusa, between the

:174

C. (lORDlW TTKVVTTT.

late autumn of one year and the summer of the next, is filled.
A number of suggestions have been made, many of which
cannot be accepted ; for example, Brefeld believes that the
Empusa is continued over the winter in warmer regions,
migrating northwards with the flies on the return of summer!
In the case of En tomo phthora calliphora, Giard believes
that the cycle is completed by the corpses of the blow-flies
falling to the ground, when the spores might germinate in the
spring a.nd give rise to conidia which infect the larvEe. Olive
(1906) studied the species of Empusa which attacks a species
of Sciara (Diptera) and found the larvae infected. He
accordingly thinks that the disease may be carried over the
winter by those individuals which breed during that period in
stables and other favourable places. As I have shown,
M. domestic a, under such favourable conditions as warmth
and supply of suitable larval food, is able to breed during the
Avinter months, although it is not a normal occurrence so far
as I have been able to discover. If, then, these winter-pro-
duced larvae could become infected they might assist in
carrying over the fungus from one year to the next, and thus
carry on the infection to the early summer broods of flies.
This suggestion and the possible occurrence of a I’esting-spore
stage appears to me to be the probable means by which the
disease may be carried over from one “ fly-season ” to the next.

E. muscfe, besides occuri'ing in M. domestica, has been
found on several species of Syrphidm, upon which it usually
occurs out-of-doors, as I have already mentioned. In addi-
tion to these Thaxter records its occurrence in Lu cilia
ctesar and Calliphora vomitoria.

VI. True Parasites.

1. Flagellata. Herpetomonas muscEe-domesticoe

Burnett.

This flagellate has been known as a parasite of the ali-
mentary tract of M. domestica for many years. Stein
(1878) figures a flagellate which he calls Cercomonas
muscae-domestica, and identifies it with the Bodo muscae-

STRUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 375

dome Stic 00 described by Burnett and the Cercomonas
in use arum of Leidy. For this form figured by Stein, a new
genus, Herpetomonas, was instituted by Kent (1880-81),
and it is taken as the type-species. It was not until the
economic importance of certain of the haemo-flagellates was
recognised that other flagellates, including H. musem-
domesticae, received further attention, and then Prowazek
(1904) described with great detail the development of this
species. In the previous year Leger (1903) had given a short
account of it, and since Prowazek’s memoir Patton (1908,
1909) has given short preliminary accounts of his study of
the life-history. The accounts of both these authors differ in
several respects from that of Prowazek, as will be shown. I
have examined a very large number of the contents of English
specimens of M. domes tica, but, with one or two doubtful
exceptions, unfortunately I have been unable so far to
discover any of these flagellates in my film preparations.

The full-grown flagellate (VIII) measures 30-50 g in
length. The body is flattened and lancet-shaped, the pos-
terior end being pointed and the anterior end bluntly rounded.
The alveolar endoplasm contains two nuclear structures. In
the centre is the large “ trophonucleus ” {tr.) •, it contains
granules of chromatin, but is sometimes difllcult to see. Near
the anterior end the deeply staining rod-shaped “ kineto-
nucleus” (blepharoplast of many authors) (/c.) lies, usually in
a transverse position. The single stout flagellum, which is a
little longer than the body of the flagellate, arises from the
anterior end, near the kinetonucleus. Prowazek describes the
flagellum as being of a double nature and having a double
origin ; this, which is a mistaken interpretation, is repeated
by Lingard and Jennings (1906).

fi'his mistake, as pointed out by Leger and Patton, is due to
the fact that the majority of the adult flagellates have the
appearance of a double flagellum, which represents the
beginning of the longitudinal division of the flagellate (VI).
Patton (1908) figures a stage in H. lygaei with the double
flagellum, and Leger (1902) in a similar .stage in H. jaculum,
VOL. 54, PART 3. NEW SERIES 27

after Lcger, Patton, and Prowazek. I-III. Pref lagellate
stage. IV-VIII. Flagellate stage : V. Young flagellate.
VI. Flagellate beginning to divide, flagellum having already
divided. VII. Advanced stage of division. VIII. Adult
flagellate. IX-XI. Post-flagellate stage: IX. Degene-
ration of flagellum. Xa. Post-flagellate stage completed by
formation of gelatinous covering, containing double row of
granular bodies (Prowazek). f.v. Flagellar vacuole, fc. Kineto-
nucleus. s.t. Spiral chi-omophilous thread, tr. Trophonucleus.

STRUCTUEE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 377

parasitic in the gut of Nepa cinerea, from which figures it
may be understood how the mistake has arisen. Through
this misinterpretation Prowazek was led to consider that the
pai’asite was of a bipolar type, in which the body had been
doubled on itself so that the two ends came together and the
flagellum remained distinct. The flagellum, according to
Leger, is continued into the cytoplasm as a thin thread,
which stains with diflBculty, and terminates in a double
granule above the kinetonucleus ; this double granule is no
doubt the “diplosome” of Prowazek. According to the
latter author another deeply staining’ double thread {s.t.),
that appears to be spirally coiled, runs backwards from the
kinetonucleus and terminates posteriorly in a distinct granule,
shown in fig. VIII.

The flagellates congregate in the proventriculus or in the
posterior region of the intestine, where they become united
by their anterior ends to form rosettes. Prowazek states that
in the rosette condition the living portion of the flagellate
resides, as it were, in the long tail-like process.

Patton divides the life-cycle of H. muscte-domesticte
into three stages — the prefiagellate, flagellate, and post-
flagellate. The last two are common, but the first stage is
not common, and Prowazek appears to have overlooked it.
For convenience I have described the flagellate stage first,
and the process of division in this stage is simple longitudinal
fusion. The nuclei divide independently, and the kineto-
nucleus usually precedes the trophonucleus. The latter
undergoes a primitive type of mitosis, in which Prowazek
recognised eight chrosomes (VII). The flagellum divides
longitudinally, and each of the two halves of the kineto-
nucleus appropriates one of the halves with its basal granule.

The prefiagellate stage, which Patton (1909) describes,
usually occurs in the masses which lie within the peritrophic
membrane.^ They are round or slightly oval bodies (I), their
average breadth being 5’5 p. The protoplasm is granular and

* I assume that Patton refers to this membrane by the term “ peri-
tricheal membrane.”

378

C. GnEDON HEWITT.

contains a troplionucleus and kiuetonucleus. Division takes
place by simple longitudinal division or multiple segmenta-
tion, and in this manner a large number of individuals are
formed (II h and III). These develop into the flagellate stage :
a vacuole, the flagellar vacuole (III, /.r.) appears between the
kinetonucleus and the I’ounded end of the pre-flagellate form,
and in it the flagellum appears as a single coiled thread, which
is extended when the vacuole has approached the surface.

The flagellate form has already been described, and in the
concluding portion of the flagellate stage, which, according
to Prowazek, is found in starved flies, these forms are found
collecting in the rectal region, and attaching themselves by
their flagellar ends in rows to gut epithelium. The more
external ones begin to shorten, during which process the
flagella degenerate (IX) and are shed. Thus a palisade of
pai-asites is formed, the outer ones being rounded and devoid
of flagella, and some of them may be found dividing (X).
Leger (1902) terms these the “formes gregariennes,” and
maintains that the existence of these “gregarine” forms is a
powerful argument in favour of the flagellate origin of the
Sporozoa, which he had previously suggested, and which
Butsclili had put foi-ward in 1884. After the degeneration of
the flagellum a thickened gelatinous covering is formed, con-
taining a double row of granular bodies (Xu), and these cysts
are i-egarded by Patten as the post-flagellate stage. They
pass out with the faeces, and dropping on the moist window-
pane or on food, are taken up by the proboscides of other flies.

Prowazek describes dimorphic forms of the flagellate stage,
which he regards as sexually differentiated forms, but Patton,
in a letter to me, says that he is unable to And any of these
complicated sexual stages. According to Prowazek, one of
these forms is slightly larger than the other, and has a greater
affinity for stain. The dimorphic forms conjugate; their cell
substance and nuclei fuse, and a resting-stage cyst is formed,
but the subsequent stages have not been followed. He
further states that the sexually differentiated forms may force

STEUCTURE, DEVELOPMENT, AND BIONOMICS OE HOUSE-FLY. 379

their way into the ovaries, where they undergo autogamy
and infect the subsequent brood.

In Madras Patton found that 100 per cent, of the flies were
infected with the flagellate ; Prowazek found it in 8 per cent,
of the flies at Rovigno. In the cold season in the plains
(India) Lingard and Jennings (l.c.) found the flagellate in
less than 1 per cent, of the flies examined ; in the hills
(Himalayas), at an elevation of 7500 feet, the flagellates were
most numerous during’ the hottest season of the year, and
gradually deci-eased in number to October and November,
when none were discovered.

One of the chief points of interest in connection with this
flagellate is its similarity to the “ Leishmann-Donovan ”
body, the parasite of kala-azar, as it was this resemblance
that prompted Rogers (1905) to suggest that the latter
parasite was a Herpetomonas, which I think Patton has
now conclusively proved to be tlie case, and he calls it
Herpetomonas donovani (Laveran and Mesnil).

Crithidia M u sc<e- domes tic ae Werner.

This pai’asite has been recently described by Werner (1908),
who found it in the alimentary tracts of four out of eighty-two
flies. It measures 10-13 /n in length, the length of the body
being 5-7 jj. and the flagellum 5-6 ju. As in other membei’S
of the genus Crithidia, which is closely allied to Herpeto-
monas, the breadth of the body is great compared with the
length, and the kinetonucleus and trophonucleus are rather
close together. A short, staining, rod-like body lies between
the kinetonucleus and the base of the flagellum. The flagellum
is single. Dividing forms undergoing longitudinal division
were frecpiently found. The kinetonucleus appears to divide
first, followed in succession by the flagellum and the tropho-
nucleus. Forms undergoing division and showing a single
trophonucleus and double kinetonucleus and flagellum were
also found. Cases occurred in which the fission began at the

380

C. GORDON HEWITT.

non-flagellate end of the body. No conjugating forms were
found, nor any wandering into the ovaries.

Lingard and Jennings (1. c.) describe certain flagellates of a
flag-shaped or rhomboidal nature, which I am strongly of the
opinion are species of Crithidia and not species of Her-
petomonas. Closely following Prowazek’s account of H.
muscse-domesticae they describe and figure all their forms
as having two flagellae in the flagellate stage. If one allows
for the rupture of the flagellum from the bodies of the
organism in making the film, some of their figures are not
unlike those of Crithidia gerridis, parasitic in the alimen-
tary tract of an Indian water-bug, Gerris fossarum Fabr.,
and described by Patton (1908).

2. Nematoda — Habronema muscie (Carter).

Carter (1861) appears to be the first to have described
a parasitic worm in M. domestica. He described a bi-
sexual nematode infesting this insect in Bombay, and found
that ; “ Every third fly contains from two to twenty or more
of these worms, which are chiefly congregated in, and con-
fined to, the proboscis, though occasionally found among the
soft tissues of the head and posterior part of the abdomen.”
His description of this nematode, to which he gave the name
Filaria niuscee, is as follows: “Linear, cylindi’ical, faintly
striated transversely, gradually diminishing towards the
head, which is obtuse and furnished with four papillae at a
little distance from the mouth, two above and two below ;
diminishing also towards the tail, which is short and termi-
nated by a dilated round extremity covered with short spines.
Mouth in the centre of the anterior extremity. Anal orifice
at the root of the tail.” He gives the length as being one
eleventh of an inch and the breadth as one three hundred and
thirteenth of an inch. In his description of his figures of the
worm he calls what is evidently the anterior, region of the
intestine the “liver.” Von Linstow (1875) described a small
nematode, wdiich he calls Filaria stomoxeos-, from the

STEUCTURE, DEVELOPMENT, AND EIONOMICS OP HOUSE-PLY. 3S1

head of S. calcitrans; this larva measured 1‘6 to 2 mm. iu
length. Generali (1886) described a nematode from the
common fly, which he calls N^ematodum spec. It is
highly probable, as my friend Dr. A. B. Shipley has suggested
to me, that Generali^s nematode and the F. muscm of Carter
are identical. Diesing (1861) created the g’enus Habronema
for the Filaria muscae of Carter, and his description is
practically a translation of Carter’s original description.
Piana (1896) describes a nematode from the proboscis of M.
domestica, which, in the occurrence of the male and female
genital organs in the same individual, he says, resembles
Carter’s nematode. He finds that at certain seasons of the
year and in certain localities it is very rare, while at others it
may occur in 20-30 per cent, of the flies. The larva, after
fixation, measured 2'68 mm. in length and 0'08 mm. in breadth.
It was cylindrical and gently tapering off at the extremities,
with the mouth terminal.

Out of the many hundreds of flies which I have dissected I
have only found two specimens of this nematode (fig. 18). From
the descriptions given by Carter and Piana and tlie figures of
the latter I feel convinced that their specimens and mine are
the same species, called by Diesing Habronema inuscae
(Carter). It is linear, cylindrical, tapering gradually towards
both ends. The anterior end is slightly rounded, having the
mouth in the centre. I am unable to confirm the presence of
the four papillae which Carter describes as a little distance
from the mouth, nor are they figured by Piana. The cuticle
is very faintly marked with transverse striations. The
common genital and anal orifice is situated at a short distance
from the posterior end of the body, which tapers off slighly
moi’e than the anterior end aud terminates in a small dilated
extremity, which is covered with minute sjiines (fig. 19). My
specimens appear to be immature adult forms, not having
reached sexual maturity. The species measures 2 mm. in
length and 0'04 mm. in breadth. The specimens that I
obtained were situated in the head region, between the optic
ganglia aud the cephalic air-sacs, from which position they

382

C. GORDON HEWITT.

could easily move down into the cavity of the proboscis. I
am unaware of any previous record of the occurrence of
Habronema muscse in this country, but I have no doubt
that if one searched specially for it it would be found to
occur more commonly than might appear from my experience,
and to be generally distributed with its host throughout the
world.

The occurrence of a parasitic worm in this position is of
great interest, even though M. domestica is not a blood-
sucking species and the nematode is not of the nature of
Filaria bancrofti. There is no reason, however, why M.
domestica should not under certain conditions carry patho-
genic nematodes, which might easily get on to the food of
man.

3. Dissemination of Parasitic Worms.

In this connection reference might be made to the experi-
ments of Grassi (1883) to which reference is made by Nuttall
in his valuable memoir (1899). Grassi broke up segments of
Taenia solium in water; they had previously been preserved
in alcohol for some time. Flies sucked up the eggs in the
water and he found them unaltered in the faeces. Oxyuris eggs
were also passed unaltered. In another experiment flies fed
on the eggs of Trichocephalus and he found the eggs some
hours afterwards in the flies’ fteces, which had been deposited
in the story beneath the laboratory ; he also caught flies in
this kitchen with their intestines full of eggs.

Calandruccio^ examined flies (? species) which had settled
upon faeces containing the ova of Taenia nana. The ova
were found in the flies’ intestines. The excrement deposited
by a fly on sugar contained two or three ova of the Taenia.
By means of such infected sugar a girl was infected, and ova
of T. nana were found in her stools on the twenty-seventh
day.

1 “ Ulteriori ricerche sulla Taenia nana," “Boll. Soc. Zool. Ital.
Roma,’ vol. vii, pp. 65-69 ; also in ‘ Boll. Acad. Gioenia, Catania,' Base.
89, pp. 15-19.

STRUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-ELY. 383

Nuttall (1. c.) records a personal communication of Stiles,
wLo placed the larvte of Musca with female Ascaris lum-
bricoides, which they devoured together with the eggs
contained by the nematodes. The larvae and adult flies con-
tained the eggs of the Ascaris, and as the weatber at the
time of the experiment was very hot the Ascaris eggs
developed rapidly and were found in diiferent stages of
development in the insect, thus proving, as Xuttall points
out, “ that the latter may serve as disseminators of the
parasite.” These experiments of Grassi and Stiles show that
flies can act as carriers of the eggs of these parasitic worms,
and that man could be infected by the fly depositing its
excreta on his food, or being accidentally immersed in food
as flies frequently are.

VII. The Dissemination op Pathogenic Organisms by

31 USCA DOMES TIC A AND ITS NON-BlOOD-SUCKING AlLIES.

Although 31. domestica is unable to act as a carrier of
pathogenic micro-organisms in a manner similar to that of
the mosquito, so far as we know at present, nevertheless its
habits render it a very potent factor in the dissemination of
disease by the mechanical transference of the disease germs.
These habits are the constant frequenting and liking for
substances used by man for food on the one hand and excre-
meutal products, purulent discharges, and moist surfaces on
the other. Should these last contain pathogenic bacilli, the
proboscis, body, and legs of the fly are so densely setaceous
(see flg. 20) that a great opportunity occurs, with a maximum
amouut of probability, for the transference of the organisms
from the infected material to either articles of food or such
moist places as the lips, eyes, etc. As I have already pointed
out (1907), 31. domestica is unable to pierce the skin, as
certain persons have suggested. The structure of the pro-
boscis will not permit the slightest piercing or pricking
action, which fact eliminates such an inoculative method of
infection. It is as a mechanical carrier, briefly, that 31.

384

C. GORDON HKWITT.

domestica and such allies as H. canicular is, etc., though
to a less degree, may be responsible for the spread of in-
fectious disease of a bacillary nature, and an account will
now be given of the role which this insect plays in the
dissemination of certain diseases.^ Before doing so, however,
it should be pointed out that whereas in some of the diseases
the epidemiological evidence adduced in support of the trans-
ference of disease germs by flies is confirmed bacteriologically,
in others only the former evidence exists. Should neither
form of evidence be available in support of the idea that M.
domestica plays a part in the dissemination of the infection
of a particular disease, it is essential, nevertheless, that if
such a method of transference is possible the potency of this
insect should be realised. This potency is governed by such
factors as the presence of M. domestica; its access to the
infected or infective material, this being attractive to the
insect either because it is moist or because it will serve as
food for itself or its progeny ; and a certain power of resist-
ance for a short time against desiccation on the part of the
pathogenic organisms, although, as in the case of the t3"phoid
bacillus, the absence of this factor is not fatal to the idea, as
it may be overcome by the fact that the fly is able to take on
its appendages an amount sufficient to resist desiccation for a
short time. The last factor is the presence of suitable culture
media, such as certain foods, or moist surfaces as the mouth,
eyes, or wounds, for the reception of the organisms which
have been carried on the body or appendages of the 11}’. If
these conditions are satisfled the possibility of M. domestica
or its allies playing- a part in the transference of the infection
should be carefully considered, and this suggestive evidence
will be discussed in certain of the diseases which follow, in
addition to the epidemiological and bacteriological evidence.

* Though it should he unnecessary, I wish to explain, as I have been
occasionally misunderstood by medical men and others, that M.
domestica is not regarded as being the cause of any disease, but as a
carrier of the infection.

STEUCTUEE, DEVELOPMENT, AXD BIONOMICS OF HODSE-FLY. 385

1. Typhoid Fever.

Of all infectious diseases the conditions in this are most
favourable for the transference of infection by M. dom estica,
and it is no doubt on this account that the greatest attention
has been paid to the role of house-flies in the dissemination
of this disease. The chief favourable condition is that the
typhoid bacillus occurs in the stools of typhoid and incipient
typhoid cases. Human excrement attracts flies not only on
account of its moisture but as suitable food for the larvae.
The infected excrement is often accessible to flies, especially
in military camps, as will be shown shortly, and the flies also
frequent articles of food and not infrequently the moist lips of
man. Such are the conditions most suitable for the transfer-
ence of the bacilli, and it is on account of the frequent
coincidence of these conditions that flies can play, and have
played, such an important role in the dissemination of this
disease among communities, in spite of the fact that the
typhoid bacillus cannot survive desiccation, which I think is
an argument against its being carried by dust.

Epidemiological and other evidence. — There is a
very large amount of testimony given as to the role played
by flies in the spread of enteric in military stations and camps,
and especially during the two wars — the Spauish-American
and the Boer War. All the conditions most favoui’able for
the dissemination of the bacilli by flies were, and in many
military stations are still, present; open latrines or filth-
trenches accessible to flies on the one hand and on the other
the men’s food within a short distance of the latrines. I
cannot do better than repeat the evidence in the words of the
witnesses and allow it to speak for itself.

Vaughan, a member of the U.S. Army Typhoid Commis-
sion of 1898, states d

“ 51y reasons for believing that flies were active in the dis-
semination of typhoid fever may be stated as follows :

* In a paper, " Conclusions Reached after a Study of Tj^ihoid Fever
amont' American Soldiei*s.'' read before the American Medical Asso-
ciation at Atlantic City, N.J., in lyuu.

386

C. fiOEDON HEWITT.

“ (a) Flies swarmed over infected faecal matter in the pits and
then visited and fed upon the food prepared for the soldiers
in the mess-tents. In some instances where lime had recently
been sprinkled over the contents of the pits, flies with their
feet whitened with lime wei’e seen walking over the food.

“ h) Officers whose mess-tents were protected by screens
suffered proportionately less from typhoid fever than did
those whose tents were not so protected.

“ (c) Typhoid fever gradually disappeared in the fall of
1898 with the approach of cold weather and the consequent
disabling of the fly.

“ It is possible for the fly to carry the typhoid bacillus in
two ways. In the first place faecal matter containing the
typhoid germs may adhere to the fly and be mechanically
transported. In the second place, it is possible that the
typhoid bacillus ma.y be carried in the digestive organs of the
fly and may be deposited with its excrement.”

One of his conclusions was that infected water was not an
important factor in the dissemination of typhoid in the
national encampments of 1898, since only about one fifth of
the soldiers in the national encampments during the summer
of that year developed typhoid fever, whereas about 80 per
cent, of the total deaths were due to this disease. In the
latter connection Sternberg (1899) refers to a report of Dr.
Reed upon an epidemic in the Cuban W ar, in which it was
stated that the epidemic was clearly not due to water
infection but was transferred from the infected stools of the
patients to the food by means of flies, the conditions being
especially favourable for this means of dissemination. Stern-
berg, as Surgeon-General of the U.S. Army, issued the follow-
ing instructions^ ; Sinks should be dug before a camp is
occupied or as soon after as practicable. The surface of the
faecal matter shonld be covered with fresh earth or quicklime
or ashes three times a day.” I think that the instructions
of that ancient leader of men, Moses, who probabl}^ had

' ‘ Circular No. 1 of the Surgeon-General of the U.S. Army,’ Api-il,
1898.

STEUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 387

experienced the effects of flies, were even better than these.
He said (Dent., Ch. xxiii, v. 12-13) : “ Thou shalt have a
place also without the camp whither thou shalt go forth
abroad; and thou shalt have a paddle [or ^shovel’] among
thy weapons; and it shall be, when thousittest down abroad,
thou shalt dig therewith, and shalt turn back and cover that
which cometh from thee.’^

Sternberg is of the opinion that typhoid fever and camp
diarrhoea are frequently communicated to soldiers through
the ag-ency of flies, “-which swarm about faecal matter and
filth of all kinds deposited upon the ground or in shallow pits,
and directly convey infectious material attached to their feet
or contained in their excreta to the food which is exposed
while being- prepared in the common kitchen, or while being
served in the mess-tent.”

Yeeder (1898), in i-eferring to the conditions existing in the
camps of the Spanish-American war, says that in the latrine
trenches he saw “ faecal matter fresh from the bowel and in
its most dangerous condition, covered with myriads of flies,
and at a short distance there was a tent, equally open to the
air, for dining and cooking. To say that the flies were busy
travelling back and from between these two places is putting
it mildly.” Further, he says, “ There is no doubt that air
and sunlight kill infection, if given time, but their very access
gives opportunity for the flies to do serious mischief as con-
veyers of fresh infection wherever they put their feet. In a
very few minutes they may load themselves with the dejec-
tions from a typhoid or dysenteric patient, not as yet sick
enough to be in hospital or under observation, and carry the
poison so taken up into the very midst of the food and water
ready for use at the next meal. There is no long and round-
about process involved. It is very plain and direct. Yet when
the thousands of lives are at stake in this way the danger-
passes unnoticed, and the consequences are disastrous and
seem mystei-ious until attention is directed to the point; then
it becomes simple enough in all conscience.”

The Commission which investigated the outbreaks of

388

C. GOEDON HEWITT.

enteric fever that occurred in 1898 in the United States
during this war came to the conclusion that “ flies undoubtedly
served as carriers of the infection” under the conditions
which have already been described. Many other authorities
bear witness to the same facts.

In our own South African war, a year or two later, the
same conditions existed, and there was a very heavy loss of
life frotn enteric fever, Writing on the subject, Dunne
(1902) says: “The plague of flies which was present during
the epidemic of enteric at Bloemfontein in 1900 left a deep
impression on my mind, and, as far as I can ascertain from
published reports, on all who had experience on that occasion.
Nothing was more noticeable than the fall in the admissions
from enteric fever coincident Avith the killing off of the flies
on the advent of the cold nights of May and June. In July,
Avhen I had occasion to visit Bloemfontein, the hospitals there
Avere half empty, and had practically become conA^alescent
camps.” A similar experience is related by Tooth (1901).
Referring to the role of flies he says : “As may be expected,
the conditions in these large camps Avere particularly favour-
able to the groAvth and multiplication of flies, Avhich soon
became terrible pests. I Avas told by a resident in Bloem-
fontein that these insects Avere by no means a serious plague
iu ordinary times, but that they came Avith the army. It
Avould be more correct to say that the normal number of flies-
was increased owing to the large quantities of refuse upon
Avhich they could feed and multiply. They Avere all over our
food, and the roofs of our tents Avere at times black Avith
them. It is not unreasonable to look upon flies as a very
possible agency in the spreading of the disease, not only
abroad but at home. It is a Avell-known fact that Avith the
first appearance of the frost enteric fever almost rapidly
disappears. ... It seems hardly credible that the almost
sudden cessation of an epidemic can be due to the effect of
cold upon the enteric bacilli only. But there can be no doubt
in the mind of anybody Avho has been living on the open
veldt, as Ave have for three or four months, that flies are ex-

STEUCTUEE, DEVELOPMENT, AND BIONOMICS OP HOUSE-FLY. 389

tremely sensitive to tlie change of temperature, and that the
cold nights kill them off rapidly.” In the discussion on this
paper Church stated that “many nurses told me that if one
went into a tent or ward in which the patients were suffering
from a variety of diseases, one could tell at once which were
the typhoid patients by the way in which the flies clustered
about their mouths and eyes while in bed.” It was further
stated in the discussion that where the Americans used quick-
lime in their latrines the cooks in the neighbouring kitchens
found that the food became covered with quicklime from the
flies which came from the latrines to the kitchens.

Dr. Tooth, in a letter to me, says : “ I am afraid my written
remarks hardly express strongly enough the importance that
I attach to flies as a medium of spreading infection. Of course
I do not wish to under-rate the water side of the question,
but once get, by that means, enteric into a camp the flies, in
my opinion, are quite capable of converting a sporadic incidence
into an epidemic. A pure water supply is an obvious necessity,
but the prompt destruction of refuse of every description is
every bit as important.”

Smith (1903), in speaking of his experiences in South
Africa, says that : “ On visiting a deserted camp during the
recent campaign it was common to find half a dozen or so
open latrines containing a foetid mass of excreta and maggots.”
Similar observations were made by Austen (1904), who, de-
scribing a latrine that had been left a short time undisturbed,
says : “A buzzing swarm of flies would suddenly arise from it
with a noise faintly suggestive of the bursting of a percussion
shrapnel shell. The latrine was certainly not more than one
hundred yards from the nearest tents, if so much, and at meal-
times men’s mess-tins, etc., were always invaded by flies. A
tin of jam incautiously left open for a few minutes became a
seething mass of flies (chiefly Pycnosoma chloropyga
Wied), completely covering the contents.”

Howard (1900) referring to an American camp, where no
effort was made to cover the faeces in the latrines, says : “ The
camp contained about 1200 men, and flies were extremely

390

C. GORDON HEWITT.

numerous in and around the sinks. Eggs of Musca domes-
tica were seen in large clusters on the feces, and in some
instances the patches were two inches wide and half an inch
in depth, resembling little patches of lime. Some of the sinks
were in a very dirty condition and had a very disagreeable
odour.”

A few examples of the prevalence of conditions favouring the
dissemination of enteric by flies in permanent camps may be
noted. Cockerill (1905), in describing camp conditions in
Bermuda, mentions kitchens within one hundred yai’ds of the
latrines ; the shallow privy, seldom or never cleaned out, and
middens are found which contain masses of filth swarming with
flies. He states that in more recent years the period of greatest
incidence is in the summer, being chiefly due to flies and con-
taminated dust. Quill (1900), reporting on an outbi’eak of
enteric in the Boer camp in Ceylon, states : “ During the
whole period that enteric fever was rife in the Boer camp
flies in that camp amounted to almost a plague, the military
camp being’ similarly infested, though to a less extent. The
outbreak in the Boer camp preceded that among’ the troops ;
the two camps were adjacent, and the migi’ation of the flies
from the one to the other easy.’^ Weir, reporting on an out-
break of enteric fever in the barracks at Umbala, India, ^ says
that most of the pans in the latrines were half or quite full,
and flies were very numerous in them and on the seats, which
latter were soiled by the excreta conveyed by the flies^ l^gs.
The men stated that the plague of flies was so great that
in the morning they could hardly go to the latrines. He
found that the flies were carried from the latrines to the
barrack-rooms on the clothes of the men. This state of affairs
suggests another mode of infection, namely, per rectum.
As Smith has pointed out (l.c.) it is not improbable that
flies under these conditions may be inoculators of dysentery.

Aldridge (1907) gives some interesting statistics showing
the influence of tlie presence of breeding-places of flies. Flies
are found in greater numbers in mounted regiments than in
‘ ‘ Army Medical Department Repoi’t.’ 1902, p. 207.

STRUCTUEE,DEVELOPMEXT,AND BIONOMICS OP HOUSE-PLY. 391

infantry, and he shows how this affects the incidence of enteric
fever. In the British Army in India, 1902-05, the ratios per
1000 per annum of cases admitted were : cavalry 41‘1, and
infantry 15'5 ; and in the U.S. Army were : cavalry 5'74,
and infantry 4’75. He states that : A study of the incidence

of enteric fever shows that stations where there are no filth
trenches, or where they are a considerable distance from the
barracks, all have an admission-rate below the average, and
all but one less than half the average.”

All these facts are equally applicable to the conditions in our
own towns and cities. Where the old conservancy methods are
used, such as pails and privy middens, the incidence of typhoid
fever is greater than in those places where the system of water
disposal has been adopted. I have examined the annual
reports of the medical officers of health of several large towns
where such conversions are being made, and they show a
falling-off of the typhoid fever-rate coincident with this
change. In Nottingham, for example,^ in the ten years 1887-
1896, there was one case of typhoid fever for every 120 houses
that had pail-closets, one case for every 37 houses with privy
middens, and one case for every 558 houses with water-closets.
'Phe last were scattered, and not confined to the prosperous
districts of the town.

One of the most important investigations on the relation
of flies to intestinal disease was that of Jackson (1907).
He investigated the sanitary condition of New York
harbour and found that in many places sewer outfalls had not
been carried below low-water mark, consequently solid matter
from the sewers was exposed on the shores, and that during
the summer months on and near the majority of the docks
in the city a large amount of human excreta was deposited.
This was found to be covered with flies. The report, consi-
dered as a mere catalogue, is a most severe indictment against
the insanitary condition of this great water front. By means
of spot-maps he shows that the cases of typhoid are thickest

‘ ‘‘Typhoid Fever and the Pail System at Nottingham,” ‘Lancet,’
November ‘29tli, 1902, p. 1489.

VOL. 54, PART 3. — NEW SERIES.

28

392

C. GORDON HEWITT,

near the points fonnci to be most insanitary. He shows, as
English investigators have also shown, how the curves of
fatal cases correspond with the temperature curves and with
the curves of the activity and prevalence of flies which were
obtained by actual counts. He also adduced bacteriological
evidence, and it is stated that one fly was found to be carrying
over one hundi’ed thousand fecal bacteria.

Bacteriological evidence. — In addition to the evidence
of Jackson, to which reference has been made, further proof
that flies are able to carry the typhoid bacillus has been
available for some years. Celli (1888) recovered the Bacillus
typhi abdominalis from the dejections of flies which had
been fed on cultures of the same, and he was able to prove
that they passed through the alimentary tract in a virulent
state by subsequent Inoculation experiments. Bicker (1903)
found that wlien flies were fed upon t\'phoid cultui’es they
could contaminate objects upon which they rested. The
typhoid bacilli were present in the head and on the wings
and legs of the fly five days after feeding, and in the alimen-
tary tract nine days after. Firth and Horrocks (1902), in
their experiments, took a small dish containing a rich emul-
sion in sugar made from a twenty-four-hour agar slope of
Bacillus typhosus recently obtained from an enteric stool
and rubbed up with fine soil. This was introduced with some
infected honey into a cage of flies together with sterile litmus
agar plates and dishes containing sterile broth, which were
placed at a short distance from the infected soil and honey.
Flies were seen to settle on the infected matter and on the
agar and broth. The agar plates and broth were removed
after a few days, and after incubation at 37° C. for twenty-
four hours colonies of Bacillus typhosus were found on
the agar plates and the bacillus was recovered from the
broth. In a further experiment the infected material was
dusted over with fine earth to represent superficially buried
dejecta, and the bacillus was isolated from agar plates upon
which the flies had subsequently walked, as in the former
experiment. They also found the bacillus on the heads, wings.

STRUCTUEE, DEVELOPMENT, AND BIONOMICS OF HODSE-ELY. 393

legs and bodies of flies which had been allowed to have access
to infected material. Hamilton (1903) I’ecovered Bacillus
ty phosu s five times in eighteen experiments from flies caught
in two undrained privies, on the fences of two yards, on the
walls of two houses and in the room of an enteric fever
patient. A series of careful experiments were made by
Sellars^ in connection with Niven’s investigations on the
relation of flies to infantile diarrhoea. Out of thirty-one
batches of house-flies carefully collected in sterilised traps in
several thickly populated districts in Manchester he found,
as a result of cultural and inoculatory experiments, that
bacteria having microscopical and cultural characters resem-
bling those of the Bacillus coli group were present in four
instances, but they did not belong to the same kind or
variety. Buchanan (1907) was unable to recover the bacilli
from flies taken from the enteric ward of the Glasgow Fever
Hospital. Flies were allowed to walk over a film of typhoid
stool and then transferred to the medium (Griinbaum and
Hume’s modification of MacConkey’s medium), and subse-
quently allowed to walk over a second and a third film of
medium. Few typhoid bacilli were recovered and none from
the second and third films. Sangree (1899) performed
somewhat similar experiments to those of Buchanan and re-
covered various bacilli in the tracks of the flies. This method
of transferring the flies immediately from the infected material
to the culture plate is not vej-y satisfactory, as I have already
pointed out (1908), as it would be necessary for the flies to
be very peculiarly constructed not to carry the bacilli. The
fly should be allowed some freedom before it has access to the
medium to simulate natural conditions. Experiments of this
kind were carried out in the summer of 1907 by Hr. M. B.
Arnold (superintendent of the Manchester Fever Hospital)
and mysell. Flies were allowed to walk over a film of
typhoid stool and then were transferred to a wire cage, where
they remained for twenty-four hours Avith the opportunity

’ Recorded in the ‘ Report on the Health of the City of Manchester,
1906,’ by James Niven, jjp. 86-96.

394

C. GORDOX HEWITT.

of cleaning' themselves, after which they were allowed to walk
over the films of media. Although we were unable to recover
B. typhosus the pi'esence of B. coli was demonstrated.
B. coli was also obtained from flies obtained on a public tip
upon which the contents of pail-closets had been emptied; the
presence of B. coli, however, may not necessarily indicate
recent contamination with human excrement. Aldridge (1 . c .)
isolated a bacillus apparently belonging to the paratyphoid
group from flies caught in a barrack latrine in India during
an outbreak of enteric fever. In appearance and behaviour
to tests it was very similar to B. typhosus.

Although we ai’e not certain yet as to the specific organism
or organisms which cause the intestinal disease known as
infantile or summer diarrhoea, which is so prevalent during
the summer months and is responsible for so great a mortality
among young children, I think we must consider the relation-
ship of ]\] . domestica and its ally Homalomyia cani-
cularis to this disease epidemiologically similar to typhoid
fever.

2. Anthrax.

In considering the relation of flies to anthrax several facts
should be borne in mind. As early as the eighteenth century
it was believed that anthrax might result from the bite of a
fly, and the idea has been used by Murger in his romance
' Le Sabot Bouge.’ A very complete historical account of
this is given by Nuttall (1899). Most of the instances in
support of this belief, however, that flies may carry the
infection of anthrax, refer to biting flies. As I have already
pointed out, M. domestica and such of its allies as H.
canicular is, C. erythrocephala, C. vo mi tori a, and
Lucilia csesar are not biting or blood-sucking flies. The
nearest allies of i\I. domestica which suck blood in England
are S. calcitrans, Hsematobia stimulans Meigen, and
Lyperosia irritans L. ; the rest of the blood-sucking flies
which may be considered in this connection belong to the
family Tabanidm, including the common genera H£ema-

STEUCTURE, DEVELOPMENT, AND BIONOJIICS OP HOUSE-FLY. 395

lopota, Tabanus, and Chi-ysops. These biting and’ blood-
sucking flies live upon the blood of living rather than dead
animals. But it is from the carcases and skins of animals
which have died of anthrax that infection is more likely to
be obtained, and I believe that such flies as the blow-flies
(Calliphora spp.), and sometimes M. domestica and
Lu cilia csesar, which frequent flesh and the bodies’ of dead
animals for the purpose of depositing their eggs and for the
sake of the juices, are more likely to be concerned in the
carriage of the anthrax bacillus and the causation of malig-
nant pustule than are the blood-sucking flies. Consequently,
as M. domestica and its allies only are under consideration,
and for the sake of brevity, the relation to anthrax of the
non-biting flies only will be considered here.

'J'he earliest bacteriological evidence in support of this
belief was published by Raimbert (1869). He experimentally
pi’oved that the house-fly and the meat-fly were able to carry
the anthrax bacillus, which he found on their probosces and
legs. In one experiment two meat-flies were placed from
twelve to twenty-four hours in a bell-jar with a dish of dried
anthrax blood. One guinea-pig’ was inoculated with a pro-
boscis, two wings and four legs of a fly, and another with a
wing and two legs. Both were dead at the end of sixty
hours, anthrax bacilli being found in their blood, spleen, and
heart. He concludes: “Les mouches qui se posent sur les
cadavres des animaux morts du Charbou sur les depouilles,
et s’en nourissent, ont la faculte de ti’ansporter les virus char-
bonneux depose sur la peau peut en traverser les differeutes
couches.” Havaine (1870) also carried out similar experi-
ments with C. vomitoria, which was able to carry the
anthrax bacillus. Bollinger (1874) found the bacilli in the
alimentary tract of flies that he had caught on the carcase
of a cow dead of anthrax. Buchanan (1. c.) placed C.
vomitoria under a bell-jar with the carcase of a guinea-pig
(deprived of skin and viscera) which had died of anthrax.
He then transferred them to agar medium and a second agar
capsule, both of which subsequently showed a profuse growth

396

C. OORDON HEWITT.

of B. antliracis as one might expect. Specimens of M.
domestica were also given access to the carcase of an ox
which had died of anthrax ; they all subsequently caused
growths of the anthrax bacillus on agar. I entirely agree
with Nuttall, who says: “It does seem high time, though,
after nearly a century and a half of discussion, to see what
would be the result of properly carried out experiments.
That ordinary flies (M. domestica and the like) may carry
about and deposit the bacillus of anthrax in their excrements,
or cause infection through their soiled exterior coming in
contact with wounded surfaces or food, may be accepted as
proven in view of the experimental evidence already pre-
sented.”

3. Cholera.

One of the first to suggest that flies may disseminate the
cholera spirillum was Nicholas (1873), who, in an interesting
and prophetic letter, said : “ In 1849, on an occasion of going
through the wards of the Malta Hospital, where a large
amount of Asiatic cholera was under treatment, my first
impression of the possibilty of the transfer of the disease by
flies was derived from the observation of the manner in which
these voracious creatures, present in great numbers, and
having equal access to the dejections and food of the patients,
gorged themselves indiscriminately, and then disgorged
themselves on the food and drinking utensils. In 1850 the
‘ Superb,’ in common with the rest of the Mediterranean
squadron, was at sea for nearly six months; during the
greater part of the time she had cholera on board. On
putting to sea the flies were in great force, but after a time
the flies gradually disappeared and the epidemic slowly sub-
sided. On going into Malta Harbour, but without com-
municating with the shore, the flies returned in greater
force, and the cholera also with increased violence. After
more cruising at sea the flies disappeared gradually, with the
subsidence of the disease. In the cholera years of 1854 and
1866 in this country the periods of occurrence and disappear-

STRUCTURE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 397

ance of the epidemics were coincident with the fly-season.”
Buchanan (1897), in a description of a gaol epidemic of cholera
which occurred at Burdwan in June, 1896, states that swarms
of flies occurred about the prison, outside which there were a
number of huts containing cholera cases. Numbers of flies
were blown from the sides where the huts lay into the pi’ison
enclosure, where they settled on the food of the prisoners.
Only those prisoners who were fed in the gaol enclosure
nearest the huts acquired cholera, the others remaining
healthy.

Bacteriological evidence. — Maddox (1885) appears to
have been the first to conduct experiments with a view to
demonstrating the ability of flies to carry the cholera spirillum,
or, as it was then called, the “ comma-bacillus.” He fed the
flies C. vomitoria and Eristalis tenax (the “drone-fly”)
on pure and impure cultures of the spirillum, and appears to
have found the motile spirillum in the faeces of the flies. He
concludes that these insects may act as disseminators of
cholera. During a cholera epidemic Tizzoni and Cattaui
(1886) showed experimentally that flies were able to carry the
“comma-bacillus” on their feet. They also obtained, in two
out of three experiments, the spirillum from cultures made
with flies from one of the cholera wards. Sawtchenko (1892)
made a number of careful experiments. Flies were fed on
bouillon culture of the cholera spirillum, and to be certain
that the subsequent results should not be vitiated by the
presence of the spirillum on the exterior of the flies, he dis-
infected them externally and then dissected out the alimentary
canal, with which he made cultures. In the case of flies
which had lived for forty-eight hours after feeding, the
second and third cultures represented pure cultures of the
cholera sjiirillum. Simmonds (1892) placed flies on a fresh
cholera intestine, and afterwards confined them from five to
forty-five minutes to a vessel in which they could fly about.
Boll cultures were then made, and colonies of the cholera
spirillum were obtained after forty-eight hours. Colonies
were also obtained from a fly one and a half hours after having

398

C. GORDON HEWITT.

access to a cholera intestine, and also from flies caught in a
cholera post-mortem room. Uffelraann (1892) fed two flies on
liquefied cultures of the cholera spirillum, and after keeping
one of them for an hour in a glass he obtained 10,500
colonies from it by means of a roll culture ; from the other,
which was kept two hours under the glass, he obtained
twenty-five colonies. In a further experiment he placed one
of the two flies similarly infected with the spirillum in a glass
of sterilised milk, which it was allowed to drink. The milk
was then kept for sixteen hours at a temperature of 20-21° C.,
after which it was shaken, and cultures were made from it;
one drop of milk yielded over one hundred colonies of the
spirillum. The other fly was allowed to touch with its pro-
boscis and feed upon a piece of juicy meat that was sub-
sequently scraped. From one half of the surface twenty
colonies, and from the other half one hundred colonies, of the
spirillum were obtained. These experiments show the danger
which may result if flies having access to a cholera patient, and
bearing the spirillum, have access also to the food. Macrae
(1894) records experiments in which boiled milk was exposed
in different parts of the gaol at Gaya in India, where cholera
and flies were prevalent. Not only did this milk become
infected, but the milk placed in the cowsheds also became
infected. The flies had access both to the cholera stools and
to such food as rice and milk.

These foregoing experiments prove beyond doubt the ability
of flies to carry the cholera spirillum, both internally and
externally, in a virulent condition, and to infect food.

4. Tuberculosis.

Although it may be considered to be hardly necessary to
introduce flies as a means of disseminating the tubercle
bacillus, it has, nevertheless, been proved experimentally
that they ai'e able to carry the bacillus in a virulent condition.
As early as 1887 Spillman and Haushalter carried on experi-
ments in which they found the tubercle bacillus in large

STRUCTUEE, DEYELOraiENT, AND BIONOMICS OF HOUSE-FLY. 399

numbers in the intestines of flies from a hospital ward, and
also in the dejections which occurred on the windows and
walls of the Avard. Hoffmann (1886) also found tubercle
bacilli in the excreta of flies in the room where a patient had
died of tubei’culosis, and he also found the bacilli in the
intestinal contents. One out of three guinea-pigs which were
inoculated Avith the intestines died ; two inoculations Avith the
excreta had no effect, Avhich led him to believe that the bacilli
became less virulent in passing through the alimentary tract.
But Celli (1. c.) records experiments in Avhich tAVO rabbits
inoculated with the excreta of flies fed with tubercular sputum
developed the disease. HayAvard (1904) obtained tubercle
bacilli in ten out of sixteen cultures made from flies Avhich
had been caught feeding on bottles containing tuberculous
sputum. Tubercle bacilli Avere also recovered from cultures
made from the faeces of Hies Avhich had fed in the same
manner, Avhich apparently caused a kind of diarrhoea in the
flies, and they died from tAvo to three days afterwards.
Fmces of flies fed on tubercular sputum Avere rubbed up in
sterile Avater and injected into the peritoneal cavity of guinea-
pigs, which developed tuberculosis. Buchanan (1. c.) alloAved
Hies to Avalk over a Him of tubercular sputum and then over
agar; a guinea-pig died of tuberculosis in thirty-six days by
inoculating it with the resulting culture.

o. Ophthalmia.

Flies have been suggested as playing an important part in
the spread of conjunctivitis, especially Egyptian ophthalmia,
and although, so far as I have been able to discoA^er, Ave have
no bacteriological evidence in favour of the belief, the circum-
stantial evidence is sufficiently strong to Avarrant it.

In speaking of its occurrence at Biskia, Laveran (1880)
says that in the hot season the eyelids of the indigenous
children are covered Avith Hies, to the attentions of Avhich
they submit; iu this AAmy the infectious discharge is carried
on the legs and probosces of Hies to the healthy children.

400

C. GORDON HEWITT.

Dr. Andrew Balfour^ of the Gordon College, Khartoum, in a
letter to me, says that the Koch-Weeks bacillus is generally
recognised as being’ the exciting cause of Egyptian ophthal-
mia. He says, “ Ophthalmia is not nearly so common in the
Sudan as in Egypt, nor are flies so numerous; doubtless the
two facts are associated.’^ Dr. MacCallan, of the Egyptian
Department of Public Health, in answer to my inquiries, says
that acute ophthalmias are more liable to transmission by
Hies than is trachoma. In his opinion the spread of the
latter is, to a comparatively small extent, through the agency
of flies, but it is mainly effected by direct contact of the
fingers, clothes, etc.

The Koch-Weeks bacillus was first seen by Koch (1883) in
Egypt in cases of acute catarrhal ophthalmia. He found that
two distinct diseases were referred to under the name; in the
severe purulent form he found diplococci, which he identified
as very probably Gonococci; in the more catarrhal form he
found small bacilli in the pus-corpuscles. He ascribed the
propagation of the disease to flies, which were often seen
covering the faces of children. Axenfeld (1908) states that
“ almost the only organisms occurring in acute epidemics
of catarrhal conjunctivitis are the Koch-AVeeks bacillus
(perhaps also influenza bacillus), and the pneumococcus (in
Egypt the gonococcus also, rarely sub tills). Other
pathogenic conjunctival organisms^ only exceptionally occur.”
And, further, Gonococci and Koch-Weeks bacilli evi-
dently lose their power of causing a conjunctivitis very slowly
indeed, and are very independent of any disposition.” His
statement that, '‘on account of their great virulence and the
mai’ked susceptibility to them, a very small number suffices,”
is important in considering the relation of flies to the spread
of the disease, although, as he remarks, every infection does
not produce the disease. The fact that the Koch-Weeks
bacillus cannot resist dryness cannot be urged as an argument

' In tliis connection he states (p. 236) : “ We can make the general
statement that the staphylococcus in the conjunctiva is not conta-
gious.’

STRUCTUEE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 401

against the spread of the infection by flies, or the same would
apply to the typhoid bacillus, whose carriage by flies is
proven. Axenfeld mentions L. Muller and Lakah and Khouri
as advocating the view that flies may spread the infection
more readily. In view of the fact that, as the same author
states, “ Koch-Weeks conjunctivitis is’ to be classed with the
most contagious infectious disease which we know of,” it is
important that the role of flies should be fully recognised.
Notwithstanding the occurrence in this country of flies in less
numbers than in such counti'ies as Egypt, it would be well to
bear in mind the j)robable influence of flies in cases of acute
conjunctivitis, such as those desciibed by Stephenson (1897)
in our own country. The sole difference between the disease
in Egypt and here is, as Dr. Bishop Harman points out to me
in a letter, that “ the symptoms produced (in Egypt) are, from
climate and dirtiness of the subjects, more severe, and that
there is found a greater number of cases of gonorrhoeal
disease than in England”; and, I would add, a far greater
number of flies. This disease is eminently suited for dissemi-
nation by flies, both on account of the accessibility of the
infectious matter in the form of a purulent discharge from
the eyes and on account of the flies’ habit of frequenting
the eyes.

6. Plague.

Although fleas are considered to be the chief agents in the
dissemination of the plague bacillus in spite of the fact that
the proof is not absolutely convincing, it is nevertheless
interesting, and certainly not unimportant, to refer to the
series of experiments of Nuttall (1897) on M. domestica.
In these experiments he conclusively proved that flies were
able to carry the plague bacillus, and that they subsequently
died of the disease. Flies were fed upon the crushed organs
of animals which had died of plague. Control flies were fed
in a similar manner on the organs of uninfected animals, and
the control experiments were kept under the same conditions.

402

C. GORDON HEWITT.

In two oE the experiments the flies were all dead on the
seventh and eighth days respectively, at a temperature of
14° 0. At higher temperatures he found that flies died more
lapidly. He was able to show that the flies contained the
bacilli in a virulent condition for about two days after thej’’
had fed on infected organs ; this,, and the fact that the infected
flies can live for several days, are extremely important from
the practical standpoint, as indicating that flies should neither
be allowed to have access to the bodies or excreta of cases of
plague, nor to the food.

7. Miscellanea.

There are on record a number of suggestions that flies may
be responsible for the dissemination of other diseases caused
by bacteria and other micro-organisms, and some account will
now be given of these and the experiments in support of such
beliefs.

If flies have access to wounds of an inflammatory and sup-
purative nature they are liable to transport the kStaphylo-
cocci to other spots. Buchanan (1907) allowed M. domestica
to walk over a Him of Staphylococcus pyogenes aureus
from an abscess, and afterwards over agar ; a mixed growth
resulted, in which S. pyogenes aureus predominated.
Celli (l.c.) records experiments which proved that S.
pyogenes aureus retains its virulence after passing-
through the intestine of the fly.

In the experiments carried out in 1907 by my friend Dr.
M. B. Arnold and myself, he chose B. prodigiosus for
the purposes of the experiment, as it is easily recognisable
and not likely to be accidentally introduced. Flies which had
just emerged from the pupae, and therefore not already con-
taminated with an extensive bacterial flora, were allowed to
walk over a film of the bacillus, after which they were con-
fined to sterile glass tubes. At varying periods they were
taken out and allowed to walk over the culture plates. Those
confined for over twelve hours retained the bacilli on their

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appendages and transferred them subsequently to the culture
media, but they were not recovered from those flies which
were kept in confinement for twenty-four hours ; a large
number of flies, however, were not used.

Dr. Kerr, of Morocco, in a paper on “ Some Prevalent
Diseases in Morocco,” i-ead before the Glasgow Medico-
Chirnrgical Society (December 7th, 1906), described epidemics
of Syphilis where, according to the author, the disease was
spread by flies which had been feeding upon the open sores
of a syphilitic patient.

Howard (1909) calls attention to an important investigation
carried on by Esten and Mason (1908) on the role which flies
play in the carriage of bacteria to milk. The flies were caught
by means of a sterile net; they were then introduced into a
sterile bottle and shaken np in a known quantity of sterilised
water to wash the bacteria from their bodies and to simulate
the number of organisms that would come from a fly falling
into a quantity of milk. They summarised their results in the
table given on p. 403.

From that table it will be seen that the numbers of
bacteria carried by a single fly may range from 550 to
6,600,000, while the average number was about 1,222,000.
Commenting on these results, the authors state that “ early in
the fly-season the numbers of bactei’ia on flies are compara-
tively large. The place where flies live also determines
largely the numbers that they carry.” From these results the
importance of keeping flies away from milk and other food
will readily be seen.

VIII. Flies and Intestinal Myiasis.

The larvae of M. domestica and its allies are frequently
the cause of intestinal myiasis and diarrhoea in children. The
occurrence of the larvae in the human alimentary tract may be
accounted for in several ways. The flies may have deposited
the eggs on the lips or in the nostrils of the patient, or the
eggs may have been deposited on the food, subsequently

STEUCTUEE, DEVELOPMENT, AND BIONOMICS OF HOUSE-FLY. 405

passing uninjured either as eggs or as young larvse into the
alimentary tract owing to insufficient mastication. Or the
larvte may have entered per rectum, the eggs having been
deposited Avhen the patient was visiting one of the old-style
privies where these flies, especially H. canicularis and
H. scalaris, frequently abound. These last two species are
frequently the cause of this intestinal trouble, and it is most
probable that the larvae enter per rectum.

Owing to the inability on the part of the observers to dis-
tinguish the different species of dipterous larvae v/e have
little information as to their occurrence in these cases.
Stephens (1905) records two cases. Two larvae were pi’o-
cured which were stated to have been passed per rectum ;
one was H. canicularis and the other is described as
i\I. corvina. The latter larva was stated to possess eight
lobes on the anterioi- spiracular processes which “ distinguishes
these larvae from M. domestica, which has seven only.” I
suspect this larva was M. domestica, which has six to eight
lobes on the anterior spiracular processes. Some years ago
a number of larvae which had been passed by a child were
sent to this laboratory, and I found that they were M . domes-
tica. In 1905 some eggs taken from the stool of a patient
suffering from diarrhoea were sent to me and on examination
they proved to be the eggs of C. ory throcephala. The
larvae of the small house-fly, H. canicularis, as I have
already mentioned, have occasionally been found in the stools
of patients.

In certain cases the larvae may wander from the mouth or
alimentary tract and get into the nasal passages or other
ducts, in which cases complications may ensue and result in
the death of the patient.

IX. Literature.

A few of the more important references included in the two previous
bibliogi-aphies are repeated here for the sake of convenience.

1909. Ainsworth, R. B. — “The House-fly as a Disease Carrier,” ‘ Journ.

Roy. Army Med. Corps,’ vol. xii, pp. 485-498.

406

C. GORDON HEWITT.

1904. Aldridge. A. R. — “ Tlie Spread of the Infection of Enteric Fever
by Flies,"’ ibid.,vol. iii, pp. 649-651.

1907. “ Honse-llies as Carriers of Enteric Fever Infection,”

ibid, vol. ix. pp. 558-571.

1904. Ansten, E. E. — “ The House-fly and Certain Allied Species as
Disseminators of Enteric Fever among Troops in the Field,”
ibid, vol. ii, pp. 651-668, 2 pis.

1908. Axenfeld. T. — ‘ Tlie Bacteriology of the Eye ’ (Translated by
A. MacNabb London. 402 jjp.. 87 figs., 3 pis.

1901. Bachmetjew, P. — ‘ Experi men telle entomologische Studien. i.
Temperaturverhiiltnisse bei Insekten.’ Leipzig. 160 pp.

1908. Balfour, A. — ‘ Third Report of the Wellcome Research Labora-
toi’ies, Gordon College. Khartoum,’ pp. 218, 219.

1905. Banks, N. — •'* A Treatise on the Acarina or Mites,” ‘ Proc. U.S.
Nat. Mus.,’ vol. xxviii, pp. 1-114, 201 figs.

1858. Bedard, J. — “ Influence de la lumicre sur les animaux,” ‘ C.R. de
I'Acad. d. Sc.,' vol. Ivi, pp. 441-453.

1898. Berg, C. — “ Sobre los enemigos pequenos de la langosta peregrina
Schistocerca paranensis (Bui-m.)," ‘Com. Mus. Buenos
Aires,’ vol. i, pp. 25-30.

1887. Bigot, J. M. F. — “ Dipteres nouveaux on pen connus,” ‘ Bull. Soc.
Zool. France,' vol. xii, pp. 581-617.

1903. Bogdanow. E. A. — " Zehn Generationen der Fliegen (Musca
domestica) in veriinderten Lebensbedingmigen," ‘ Allg. Zeitschr.
f. Entom..' vol. viii, pp. 265-267.

1874. Bollinger, O. — “ Experimentelle Untersuchungen fiber die Ent-
stehung des Milzbrandes,"’ 46 Versammh. d. ‘ D. Naturf. u.
Aerzte zu Wiesbaden,' September, 1873 ; and ” Milzbrand.” in von
Ziemssen's ‘ Handb. d. spec. Patol. u. Therapie,’ vol. iii, pp. 457
and 482.

1871. Brefeld, O. — “Untersuchungen fiber die Entwickelung der
Empusa musca; and E. radicans." ‘ Abh. d. Naturf. Gesellsch.
Halle.,’ vol. xii, pp. 1-50, pis. 1-4.

1897. Buchanan, W. J. — “ Cholera Diffusion and Flies,” ‘ Indian Med.
Gaz.,’ pp. 86. 87.

1907. Buchanan, R. M. — “ The Carriage of Infection by Flies,” ‘Lancet,’
vol. clxxiii. pp. 216-218, 5 figs.

1861. Carter, H. J. — “ On a Bi-sexual Nematoid Worm which Infests
the Common House-fly (Musca domestica) in Bombay,” ‘ Ann.
Mag. Nat. Hist.,' ser. (3), vol. vii, pp. 29-33, 4 figs.

1888. Celli, A. — “ Transmissibilita dei gernii patogeni mediante le

STEUCTURE, DEVELOPMENT, AND DIONOMICS OF HOUSK-FLY. 407

dejecioni delle Mosche,” ‘ Bull. Soc. Lancisiana ospedali di Roma,’
fasc. 1, p. 1.

1905. Cockerill, J. W. — “ Report on the Prevalence of Enteric Fever in
Bermuda,” vs^ith Tables and Diagrams, ‘ Joura. Roy. Army Med.
Corps,’ vol. iv, pp. 762-796.

1855. Cohn, F.— “Empusa muscse und die Krankheit der Stuhen-
fliegen,” ‘ Nova Acti. Acad. Caes. Leop. Carol. Germ. Nat. Cur.,’
vol. XXV, p. 301.

1870. Davaine, C. — “ Etudes sur la contagion du charhon chez les
animaux domestiques,” ‘ Bull. Acad. Med. Paris,’ vol. xxxv, pp.
215-235.

1861. Diesing, K. M.— “ Kleine helminthologische Mittheilungen,”

‘ Sitz. Kais. Akad. d. wiss. Wien.,’ vol. xliii, pp. 269-282.

1872. Donholf. — “ Beitriige zur Physiologic, 1 : Ueber das Yerhalten
Kaltbliitiger Thiere gegen Frosttemperatur,” ‘ Arch. f. Anat. und
Phys. und wiss. med. von Reichert und Du Bois-Raymond,’ p.
724.

1797. Donovan, E. — ‘ Natural History of British Insects,’ vol. vi, p. 84.

1902. Dunne, A. B. — “ Typhoid Fever in Soutli Africa; its Cause and
Prevention,” ‘ Brit. Med. Journ.,’ March 8th, 1902, p. 622,

1908, Esten, W, M., and Mason, C. J, — “ Sources of Bacteria in Milk,”
‘ Bull. No. 51, Storrs Agric. Exp. Sta.,’ Storrs, Conn., U.S.A.

1903. Picker, M. — “ Typhus und Fliegen,” ‘ Arch. f. Hygiene,’ vol. xlvi,
pp. 274-282.

1902. Firth, R. H., and Horrocks, W. H. — “ An Inquiry into the
Influence of Soil, Fabrics, and Flies in the Dissemination of
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1906. Franklin. G. D. — “ Some Obsei-vations on the Breeding Ground
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1752-78. de Geer. C. — ‘ Mcmoires pour sei-vir ii I'histoire des Insectes,’
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1886. Generali, G. — '• Una larva di nematode della mosca commune,”
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1764. Geoffroy, E. L. — -'Histoii'e ahregee des Insectes,’ vol. ii, p. 624.

1879. Giard, A. — “ Deux especes d’Entoniophthora nouveaux pour la
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1909. Godfrey, R. — “The False-scorpions of Scotland,” ‘Ann. Scot.
Nat. Hist.,’ No. 69, January, 1909, pp. 22-26.

VOL. 54j PART 3. NEW SERIES.

29

408

C. GORIION nEWITT.

1883. Gnissi, B. — ‘‘ Les mefaits des Moiiclies," ‘ Arcli. ital. de Liologie,’
vol. iv, pp. 205-228.

1908. Hamer, W. H. — “ Nuisance from Flies." Report Oy the Medical
Officer presenting a report hy Dr. Hamer. Medical Officer (General
Purposes), on the extent to which the fly nuisance is produced
in London hy accmnnlations of offensive matter. 10 i^p., 2 figs.,
3 diagrams. Printed for the London Comity Council (Puhlic
Health Committee), London.

1908. Hamer, W. H. — “ Nuisance from Flies," Report of the Medical
Officer of Health presenting a further report hy Dr. Hamer,
Medical Officer (General Purposes), on tlie extent to which the fly
nuisance is produced in London hy accumulations of offensive
matter. 0 pp., 4 diagrams. Printed for the London County
Council (Puhlic Health Committee). London.

1904. Hayward. E. H. — “ The Fly as a Cai-rier of Tulierculous Infec-
tion," ‘New York Med. Journ.,’ vol. Ixxx, pp. 643, 644.

1907. Hewitt, C. G. — “ On tlie Life-history of the Root-maggot,
Anthomyia radicum, Meigen," ‘Journ. Econ. Biol.,' vol. ii,
pp. 56-63 ; 1 pi.

1907. “The Structure, Development, and Bionomics of the

House-fly, Musca doinestica Linn.; i- The Anatomy of
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1908. Idem., pt. ii. "The Breeding Hahits, Development, and

the Anatomy of the Larva," ihid., vol. 52, 2ip- 495-545 ; 4 jils.

1907. “ The Proboscis of the House-fly," ‘ Brit. Med. Journ..'

November 23rd, 1907, ji. 1558.

1!*08. " The Biology of House-flies in Relation to the Puhlic

Health." ‘ Journ. Roy. Inst. Puhlic Health,' vol. xvi, ^ip- 596-608 ;
3 figs.

1905. Hickson, S. J. — “ A Parasite of the House-fly," ‘ Nature.' October
2t)th, 1905.

1888. Hoffmann, E. — “Ueher die Verhreitung der Tuherculose durch
Stuhenfliegen," ‘ Corres2:>ondenzhl. d. azztl. Krcis und Bezirk-
svereine im Kunigr. Sachsen.,' vol. xliv, j^p. 130-133.

IftOO. Howard, L. O. — " A Contribution to the Study of the Insect
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Spread of Typhoid Fever hy Flies)." ‘ Proc. W ash. Acad. Sciences,'
vol. ii, j)p. 541-604, figs. 17-38, jJs. xxx, xxxi.

1909. “ Economic Loss to the Peoj>le of the United States through

Insects that Carry Disease," ‘Bull. No. 78, U.S. Dej)!. Agric.,
Bureau of Entomology ' ; 40 ^ip-, 3 tables.

1907. Jackson, D. D. — "Pollution of New York Harbour as a Menace

STRUCTURE, DEVELORMENT, AND BIONOM ICS OF UOUSE-FLAA 409

to Health l)y the Dissemina tion of Intestinal Disease through the
Agency of the Common House-fly,” a Rei^ort to the Committee
on Pollution of the Merchants’ Association of Hew York ; 22 pp.,
2 maps, 3 charts, 3 figs.

1908. Kammerer, P. — “ Regeneration cles Dipterenfliigels heim Imago,”
‘ Arch. f. Entwick.,’ vol. xxv, pp. 349-360 ; 4 figs.

1880-1881. Kent, W. S. — ‘A Manual of the Infusoria.’ vol. i, p. 245,
pi. xiii, figs. 29-34.

1901. Kew, H. W. — “ Lincolnshii'e Pseudo-scorpions : with an Accoimt
of the Association of such Animals with other Arthropods,”
‘ Naturalist,’ No. 534, July, 1901, pp. 193-215.

1826. Kirhy and Spence. — ‘ Introduction to Entomology,’ vol. iv, pp.
228-229.

1883. Koch, R. — “ Bericht fiber die Thatigkeit der deutschen Cholera
Komission in .dEgypten und Ostindien,” ‘ Wiener med.
Wochenschr.,’ No. 52, pp. 1548-1551.

1880. Laveran, A.— “ Contribution a I’etude du bouton de Biskra,”
"Ann. d. Dermatologic, ' 2nd ser., vol. i, pp. 173-197.

1902. Lcger, L. — “ Sur la Structure et la mode de multiplication des
Fhigellcs du genre Herpetomouas, Kent,” ‘ C.R. Ac. Sci.,’
vol. cxxxiv, p. 781 ; 7 figs.

1903. “ Sur fiuehpies Cercomonadines nouvelles on peu connues

parasites de I'intestin des Insectes,” ‘ Arch. f. Protistenk.,’ vol. ii,
pp. 180-189 ; 4 figs.

1906. Lingard, A., and Jennings, E. — “Some Flagellate Forms found
in the Intestinal Tracts of Diptera and other Genera.” London
(Adlard & Son) ; 25 pp., 5 pis. (It is hardly necessary to point
out that the Diptera do not constitute a Genus ! — C. G. H.)

1758-9. Linnaais, C. — ‘ Systema naturas’ lOth ed., p. 617.

1875. von Linstow. — “ Beobachtungen an neuen und bekannten
Helminthen,” ‘Arch. f. Naturgesch.,’ pp. 183-207.

1890. Loeb, J. — “ Der Heliotropismus der Tliiere und seine Uberein-
stimmung mit dem Heliotrox)ismus der Pflauzen,” Wurzburg,
118 pp., 6 figs.

1894. Macrae, R. — "Flies and Cholera Diifusion,” ‘Indian Med. Gaz.,’
1894, pp. 407-412.

1885. Maddox, R. L. — " E.xperinients in Feeding some Insects with the
Curved or “ Comma ” Bacillus, and also with another Bacillus
(B. subtilis?),” ‘ Jouru. Roy. Micros. Soc.,’ ser. (2), vol. v, pp.
602-607, 941-952.

1894. Moniez, R. — ‘‘ Apropos des publications rccentes sur le faux paia-

410

C. GORDON HEWITT.

sitisin des Cliernctides sur differents Artliropodes," ‘ Rev. Biol,
du Nord de l:i France,’ vol. vi, pp. 47-54.

1877. Mni-ray, A. — ‘Economic Entomology,’ London, p. 12!).

1906. Newstead, R. — “ On the Life-History of Stomoxys calcitrans,

Linn.,” ‘ Journ. Econ. Biol.,’ vol. i, pp. 157-166, 1 pi.

1907. “ Preliminary Rej^ort on the Hahits, Life-Cycle and

Breeding-Places of the Common Honse-fly (Mnsca domestica,
Linn.) as Observed in Liverpool, with Suggestions as to the Best
Means of Checking its Increase,” Liverpool, 23 pp.

1873. Nicholas, G. E. — “The Fly in its Sanitary Aspect,” ‘Lancet,’
1873, vol. ii, p. 724.

1897. Nuttall, G. H. F. — ‘‘ Zur Aufkliirnng der Rolle, welche Insekten
bei der Verbi-eitung der Pest spielen — Ueber die Empfindlichkeit
verscliiedener Thiere fiir dieselbe,” ‘ Centralbl. f. Bakteriol.,’ vol.
xxii, pp. 87-97.

1899. “On the Role of Insects, Arachnids, and Myriapods, as

Carriers in the Sprend of Bacterial and Parasitic Diseases in Man
and Animals ; A Critical and Historical Study,” ‘ Johns Hopkins
Hosjjital Reports,’ vol. viii, 154 pp., 3 pis. (A vei-y full biblio-
graphy is given.)

19U6. Olive, E. W. — “ Cytological Studies on the Entoniophthoreae :
i. The Morphology and Development of Empusa,” ‘ Bot. Gaz.,’
vol. xli, p. 192, 2 pis.

1887. Osten-Sacken, C. R. — “ On Mr. Portchinski’s publications on the
larvaj of Muscida;, including a detailed abstract of his last
paper: — ‘Comparative Biology of Necrophagous and Copro-
pliagous Larva',’ ” ‘ Berl. Ent. Zeit.,’ vol. xxxi, pp. 17-28.

1874. Packard, A. S. — “ On the Transformations of the Common House-
fly, with Notes on allied forms,” ‘ Proc. Boston Soc. Nat. Hist.,’
vol. xvi, pp. 136-150, 1 pi.

1908. Patton, W. S. — “ The Life-Cycle of a species of Cnthidia jjara-
sitic in the intestinal tract of Gerris f ossaruiu, Fabr.,” ‘Arch,
f. Protistenk.,’ vol. xii, pp. 131-146, 1 pi.

1908. “ Herpetomonas lygai,” ibid., vol. xiii, pp. 1-18,

Ipl.

1909. “ The Parasite of Kala-Azar and Allied Organisms,”

‘Lancet,’ January 30th, 1909, pp. 306-309, 2 figs.

1896. Piana, G. P. — “ Osservazioni sul Dispharagus nasutus, Rud.
dei polli e sidle larve Nematoelmintiche delle mosche e dei
Porcellioni," ‘ Atti della Soc. Ital. d. Sci. Nat.,' vol. xxxvi, pp.
239-262, 21 figs.

1892. Pickard-Cambridge, O. — “ On the British Species of False-Scor-

STRUCTURE, DEVRROri\rENT, AND BIONOMICS OE HOUSE-FLY. 41 1

pions,” ‘ Proc. Dorset Nat. Hist, and Antiq. Field Club,’ vol. xiii,
P23. 199-231, 3 jjls.

1904. Prowazek, S. — “Die Entwicklung von Heiqietomonas eineni
init den TryjJimosoinen verwandten Flagellaten," ‘ Arb. aus deni.
Kaiserl. Gesimdlieitsamte,’ vol. xx, jip- 440-452, 7 figs.

1900. Quill, R. H. — “ Rej)ort on an Outbreak of Enteric Fever at
Diyatalawa Cainji, Ceylon, among the 2nd King’s Royal Rifles,"
‘ Ai'my Med. De^it. Report,’ Ajijiendix 4, ji. 425.

1869. Raimbert, A. — “ Reclierclies exjDerimentales sur la transmission
du Charbon jiar les mouclies,’’ ‘ C.R. Ac. Sci. Paris,’ vol. Ixix, iqi.
805-812.

1905. Rogers, L. — “ The Conditions affecting the develojiment of
Flagellated organisms from Leishman bodies and their bearing
on the jirobable mode of infection,’’ ‘Lancet,’ June 3rd, 1905,
pp. 1484-1487.

1899. Sangree, E. B. — “ Flies and Ty^ihoid Fever,” ‘ New York Med.
Record,’ vol. Iv, jip. 88-89, 4 figs.

1892. Sawtchenko, J. G. — “ Le role des mouclies dans la jiroiiagation de
rcqiidcmic cholerique,” ‘ Vratcli,’ St. Petersburg. (Reviewed in
‘ Ann. de I’lnstitut Pasteur, vol. vii, j). 222.)

1892. Simnionds. M. — “ Fliegen und Choleraubertragen,” “ Deutsch.
med. Wochenschr.,’ No. 41, ji. 931.

1903. Smith, F. — “ Municiiial Sewage,” * Joum. Troji. Med.,’ vol. vi,
p. 285.

1887. Spillmann and Haushalter. — “ Dissemination du bacille de la
tuberculose par les mouclies,” ‘ C.R. Ac. Sci.,’ vol. cv, jiji. 352-
353.

1878. Stein, F. R. — “ Der Organismus des Infusionsthiere, iii, Abthei-
lung — Die Naturgeschichte des Flagellaten oder Geisselin-
fusionen,” 154 jij)-’ 24 jils., Leipzig.

1905. Stejihens, J. W. W. — “ Two Cases of Intestinal Myiasis,” ‘Thoinji-
son Yates and Johnstone Laboratories Rejiort,’ vol. vi, jiart i,
pp. 119-121.

1897. Stephenson, S. — " Rejiort on the Prevalence of Ophthalmia in the
Metroiiolitan Poor-Law Schools,” ‘ Blue-Book,’ October 2nd,
1897. (Reviewed in ‘ Lancet,’ October 16th, 990, 991.)

1899. Steinberg, G. M. — “ Sanitary Lessons of the War,” ‘ Philad. Med.
Journ.,' June 10th and 17th, 1899.

1888. Thaxter, R. — “ The Entoniophthorea; of the United States,”
‘ Mem. Boston Soc. Nat. Hist.,’ vol. iv, 133-201, pis. 14-21.

1886. Tizzoni, G., and J. Cattani. — “ Untersuchungen fiber Cholera,”
• Centralbl. f. d. med. Wissench. Berlin,' vol. xxiv, jijj. 769-771.

412

r. GORDON iiKwirr.

1000. Tooth, H. H.- — •• Enteric Fever in tlie Army in South Africa."
‘Brit. Med. Journ.,’ Novemhei' 10th, 1900.

1901. “ Some Personal Experiences of the Epidemic of Enteric

Fever among the Troops in South Afi'ica, in the Orange River
Colony," ‘ Trans. Clin. Soc.,’ vol. xxxiv, (J4 pp.

1892. Uft'elniann, J. — “ Beitriige zur Biologie der Cholerahacillus,”
‘ Berl. klin. Wochenschr.,’ 189'2, pp. 1213-1214.

1898. Veeder, M. A. — “ Flies as Spreaders of Disease in Camp,” ‘ New
York Med. Record.’ vol. liv, SejAemher 17th, p. 429.

1850. AValker, F. — ‘ Insecta Sanndersiana, i, Diptei'a,’ p. 345.

1908. Werner. H. — “ Uher eine eingeisselige Flagellatenform im Darin
der Stnlienfliege," ‘ Arch. f. Protistenk.,’ vol. xiii, pp. 19-22, 2 pis.

1881. Winter, G. — ‘‘ Zwei nene Entoniophthoreen,” ‘Bot. Centralbl.,’
vol. V, p. 62.

X. Appendix.

Oil the Breeding of M. domestica during the
Winter Months.

In the account that I gave of the breeding habits of M.
domestica in the second part of this monograph, it was
stated (p. 503) that the experiments and observations pointed
to tlie fact that, in the presence of suitable larval food, such
as excrernental matter or decaying and fermenting food
materials in a moist and warm condition, the female flies
would lay their eggs and the larvie would develop if the
temperature of the air was sufficiently high for the prolonged
activity of the flies. Flies are sometimes found under these
conditions in warm restaurants and kitchens, stables, and
cowsheds, and under these conditions are able to breed during
the winter mouths. I am pleased to find that my own observa-
tions and those of Griffith (there referred to) as to the ability of
M. domestica to breed during the winter months has been
confirmed by Jepson^ during the past winter.

Flies were caught in February (1909j in the bakehouse of

' In “ Reports to the Local Government Board on Public Health and
Medical Subjects (New Series, No. 5). Preliminary Reports on Flies as
Carriers of Infection. No. 3. Mr. Jeiisou's Report on the Breeding of
the Common House-Fly during the Winter Mouths,” pp. 5-8, 1909.

STRUCTURE, DEYELOrMENT, AND DIONOMICS OF HOUSE-FLY. 413

one of the colleges (Cambridge), and were transferred to a
small experimental greenhouse in the laboratory where the
temperature was from 65° F. in the morning to 75° F. in the
evening. The flies Avere allowed to oviposit in moist bread in
which the process of fermentation had begun. He found
that the times for the developmental stages approxi-
mately agreed with those obtained by me at about the same
temperature, and that the whole development was completed in
about three weeks. At an average temperature of 70° F. the
eggs Avere all hatched in twenty-four hours. The first larval
stage lasted thirty-six hours, the second larval stage four
days, and the third stage was complete in five and a half
days; the Avhole larval period, therefore, occupied eleven
days, fl'he average period occupied in the pupal stage Avas
ten days; some puprn incubated at a temperature of 77° F.
hatched in three days.

It may be stated uoav, therefore, without fear of contra-
diction, that flies are able to breed during the winter months,
if the necessary conditions of food, temperature, and moisture
are present. It is probablj'^ from these Avinter flies that the
early summer flies are produced, as I have previously sug-
gested.

Corrigendum.

My attention has been very kindly called by Prof. W. A.
Riley to a slight mistake that I have made in my account of
the venation of the Aving (Part I, p. 412). B}' an 0A"ersight

I have termed transverse nervnres the two small veins
m.cu. (medio-cubital) and cu.a. (cubito-anal) . These ai’e
really parts of the original longitudinal veins il/. 3 and Cu. 2.
A study of such a series of dipterous Avings as those figured
by Comstock in the papers there quoted (Comstock and
Needham, 1898), or in his ‘Manual for the Study of Ento-
mology,’ Avill shoAv that these ap])arent transverse or cross-
veins are morphologically equivalent to branches of the
primary veins.

The University;

Manchester.

414

C. GORDON HEWITT.

EXPLANATION OE PLATE 22,

Illustrating- Dr. C. (Jordon Hewitt’s paper on “The Structure,
Development, and Hionoinics of the House-fly, Mu sea
doniestica, Linn. Part III. The Bionomics, Allies,
Parasites, and the Pelations of M. dornestica to
Human Disease.”

Fig-l. — Mature larva of Homalomyia canicularis, L. X 17.
Anterior spiracular processes. Posterior spiracular apertures.

Fig. 2. — Posterior end of mature larva of Antliomyia radicum
Mg. an. Anus.

Fig. 3.— Anterior spiracular process of mature larva of A. radicum.

Fig. 4. — Head of Stonioxys caleitrans, L. ; left lateral aspect.

Fig. 5. — Posterior end of mature larva of S. caleitrans.

Fig. (5. — Posterior spiracle of the same, enlarged.

Fig. 7. — Posterior spii-acle of mature larva of Mu sea dornestica.

Fig. 8 . — Posterior spiracles of first larval stage of CalliiDhora
ery tlirocepliala. Mg.

Fig. 9. — Posterior spiracles of second larval stage of C. erythro-
cephala.

Fig. 10. — Posterior spiracle of mature larva of C. ery tlirocepliala.

Fig. 11.— Anterior spiracular process of mature larva of C. erythro-
cepliala.

Fig. 12. — Posterior end of mature larva of C. ery tlirocepliala.

Fig. 13. — Che rues nodosus, Schr. X 30.

Fig. 14. — Thoraco-ahdominal region of Homalomyia canicu-
lar is, $ . showing Gamasids attached to the ventral side of the ahdoiiien.

Fig.l.'i. — Longitudinal (sagittal) section of ahdonien of M. dornestica.
which has hcen killed liy Enipusa niuscai, showing the feltwork of
fungal hyplia; filling the inside of the ahdomiiial cavity and the pro-
duction of conidia in the intersegniental regions, x 12. c. Coiiidio-
phores producing conidia. /. Fungal liypluc.

Fig. 10. — Four conidioiihoies showing the formation of conidia (c.).
X PlO (approx.).

Fig. 17.— Conidium of Empusa muscai. X 400. o.y. Oil glohule.

Fig. 18. — Hahronema miiscai (Carter). Adult hut immature
specimen. X 85. f/.a. Genito-aiial aperture.

Fig. 19. — Caudal end of Hahronema miiscai. X 360.

Fig. 20.— Tarsal joints of one of posterior pair of legs of Musca
dornestica. Lateral aspect, to show densely setaceous character.

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15

THE DEVELOPMENT OF THE TEMNOCEPHALEvE. 415

The Development of the Temnocephaleee.
Part I.

By

Professor W. A. IfasAvell, Ifl.A., D.Sc., F.R.S.

With Plates 23—25.

I. Introduction.

Though several additions of importance^ have been made
to the literature concerned with Tern noceph ala and its
allies since 1893, when I published an account of the group
under the title “ A Monograph of the Temnoceplialese ” (8),
no attempt has been made hitherto to deal with the embryo-
logy of any of its members. In view of the interest which
attaches to them on account of their isolated character and
problematical relationships with other sections of Platodes,
it is very desirable that something should be done towards
filling this hiatus iu our knowledge.

What follows is by no means a complete or exhaustive
account of this subject. It is concerned mainly with the
developmental history of a single form — viz. Temno-
cephala fasciata — one of the most widely distributed
of the Australian representatives of the group : and in this

■ The most extensive and important of these is that of Wacke (16),
published in 1900. When writing this the author had, apparently, not
liad tlie opportunity of consulting my “ Monograph," and all his refer-
ences, critical and otherwise, concern themselves with a short paper
which I publislied in 1888 in the ‘ Quarterly Journal of Microscopical
Science.’ Hence there is a good deal that goes somewhat wide of the
mark.

VOL. 54, PART 3. NEW SERIES.

30

416

W. A. HASWKLL.

history there are a number of important points that have not
yet been determined — notably the processes of maturation
and fertilisation, and the early segmentation phases. Since^
however, the portions of the development which I have
succeeded in tracing reveal certain phases that are quite
unique in character, it appears to me expedient to publish
these observations as a first contribution to our knowledge
of the subject.

In addition to Temnocephala fasciata, T. minor, T.
dendyi, and T. qnadricorn is, as well as Craspedella
spenceri, have been made the subjects of study; but in
noue of these were the methods adopted with the eggs
sufficiently satisfactory in their results to enable me to do
more at present than state that the general course of the
development is the same in all these forms. Of the somewhat
specialised New Zealand species — T. novim-zealandiae — I
have procured ample material, which is now in course of
preparation.

AVhat has impressed me most strongly in connection with
this investigation has been the complete absence in the
development of any definite evidence of relationships with
the groups looked upon as the most nearly allied. Until
comparatively recently little that could be accepted as well
authenticated had been published on the development either
of the Rhabdocccles or of the Heterocotylean Trematodes.
In the latter group as yet little has been done since the
publication of Zeller’s (17) account of the development of
Polystomnm in 1876. But Bresslau’s valuable work (3)
has furnished us with a much-needed body of information on
certain Rhabdocoeles, and this has enabled me to effect a
comparison with Temnocephala, with the result that the
differences appear to be more numerous and more radical
than the resemblances. So far as can be ascertained in the
present state of our knowledge, a similar result follows from
a comparison with the Heterocotylean Trematodes. The
upshot .seems to be that the study of the development, so far
as it has been carried at present, does not in any way tend to

THE DEVELOPMENT OF THE TEMNOCEPHALE^. 417

bridge over, but ratlier to widen, the gap between the
Temnocephaleae and neighbouring groups.

II. Methods.

Though the eggs are abundant and readily procurable, the
study of the development of Temnocephala presents
considerable technical difficulties, owing to the intractable
character of the material. The egg-shell is toug'h and rela-
tively thick, and not readily j^erineable by reagents. When
it is broken through, the content.s, in the fresh condition,
burst out, and become completely disorganised. When fixed
and hardened in the ordinary way the yolk becomes ex-
tremely hard and brittle. Many methods were experimented
with before the following coui'se of procedure, which has
proved sufficiently satisfactory, was finally arrived at.

The eggs are fixed with sublimate alcohol followed by
iodized alcohol and 90 per cent, alcohol. After hardening
they are treated with a solution of hypochlorite of soda. If
the eggs are transferred dii*ectly from the strong alcohol to
the hypochlorite solution, the shell of most of them splits
longitudinally, and, before the desired effect in softening
and removing the egg shell has been attained, the contents,
completely exposed, are disintegrated. This effect is avoided
by making the transference gradually through downwardly
graded alcohols to the wateiy solution.

A weak solution of the hypochlorite soon dissolves the
cement that attaches the eggs to one another, and begins to
act on the substance of the egg-shell itself. When the action
is judged to have ])roceeded far enough, the eggs are washed
in distilled water, and then dehydrated with alcohol. Double
embedding is essential. From absolute alcohol the eggs are
transferred to a mixture of equal parts of absolute alcohol
and anhydrous ether, in which they remain for twenty-four
hours. They then remain for a like period in | per cent,
solution of photoxyliu (or celloidin) in equal parts of absolute

418

W. A. HA SWELL.

alcohol and ether, followed by a per cent, solution of the
same. The celloidin blocks, hardened in chloroform, are
then finally embedded in the hardest paraffin in the usual
way. The staining agents employed almost exclusively were
Ehrlich’s liaBinatoxylin, or Mayer’s hmmocalcium, followed by
eosin.

III. Formation of the Egg.

The ovary (germarium) in Temnocephala fasciata,
and in all the Australasian species of the genus (PI. 23, fig. 1),
is a solid ellipsoidal mass of ova enclosed in a thin capsule of
muscular fibres. At the right extremity, which is the one
situated nearest to the oviduct, is the largest ovum, which is
more rounded in form than the rest. The remainder decrease
gradually in size towards the left, the largest of them ex-
tending across the entire width of the ovary. At the left
end is a mass of smaller ova, which show evidence of slow
multiplication by mitotic division. The full-grown ovum, at
the right-hand end of the ovary, is about O’ll mm. in long
diameter. Its protoplasm is densely loaded with very fine
granules, and contains, in addition, a number of much larger
rounded masses of very definite spherical form. The nucleus
is large, about one third of the diameter of the ovum itself,
with spherical nucleolus, and a fine, open, achromatin net-
work .

The ripe ovum becomes detached from the others, and
passes into the oviduct, which opens through the capsule of
the ovai’y. It must then pass along the oviduct to the
ootype, where it becomes surrounded by a mass of yolk-cells
and the whole then becomes enclosed in a chitinous shell,
the substance of which is secreted by the shell glands.

Considerable differences in detail distinguish the various
species of Temnocephala as regards not only the male
parts oE the reproductive apparatus, but also the female.
But in all the species which I have had the opportunity of
examining, the essential features of the parts concerned in

THE DEVELOPMENT OF THE TEMNOCEPHALE^. 419

the fonnation of the egg closely correspond. The two main
vitelline ducts, right and left, open together into the oviduct
near the ovary. Close to this the oviduct gives off a short
branch leading to a larg’e sac, with a syncytial epithelium,
lying near the middle line in front of the rest of the female
apparatus. This has been very usually called the recep-
taculum seminis — a name for which, in view of the fact
that its contents consist very largely of yolk matter, I pro-
posed (8) to substitute that of receptaculum vitelli. I
was then of opinion that it serves as a receptacle in which
yolk accumulates until enough has been collected for com-
pleting an egg, when it is discharged into the ootype. A
more thorough examination of the subject has shown me,
however, that, at least as regards the Australasian forms,
both names are inappropriate. The sac usually contains
spermatozoa it is true, but they never form a large propor-
tion of its contents ; and they are spermatozoa which have
lost their activity, and move, when they move at all, with
comparative sluggishness. It also contains yolk matter — the
great bulk of the contents, in fact, consisting of that mate-
rial, but it is yolkinatter which has undergone degeneration ;
the cells have broken up, and the nuclei have, for the most
part, disappeared. Moreover, mingled with the motionless
or sluggish spermatozoa and the broken-down yolk-cells are
shreds and strands of a substance which corresponds exactly
in appearance and behaviour to staining agents with the
secretion of the shell-glands. The conclusion to be arrived
at from these facts is clear enough ; the so-called recep-
taculum seminis, or receptaculum vitelli, is in reality a recep-
tacle for surplus spermatozoa and surplus vitelline matter as
well as shell-gland secretion.

The question may suggest itself — What necessity is there
for such a receptacle ? Why should the surplus matter not
be passed directly out through the female duct ? To this the
answer obviously is that very frequently — whenever, in fact,
an egg is in course of formation in the ootype or is lodged in
the distended atrium — the way to the exterior is blocked,

420

^V. A. HASWELL.

and in order that the foi’mation of the egg may proceed
without interference, the yolk-cells which are being dis-
charged into the oviduct, and the shell-gland secretion which
collects after the shell has become formed, as well as the
surplus spermatozoa, have to be disposed of.

That this is the function discharged by the receptaculum
in Temuocephala there remains, to my mind, not the
slightest doubt. The function of a true receptaculum
semiuis in those animals is discharged by the anterior
part of the oviduct, in which a mass of actively moving
spermatozoa is usually to be found. T. comes has an excep-
tional aiTangement of the parts ; in that species the cavity of
the receptaculum is incompletely divided into two unequal
parts by a partition which is pierced in the middle by a wide
aperture. In all the specimens I have examined, while the
distal larger part of the cavity, the part furthest from the
oviduct, is filled with the usual mixture of effete genital
products, the smaller part is occupied by a mass of normal
spermatozoa.

Whether the relatively large sac known as receptaculum
seminis in other Platodes may perform in some cases the
same function as that above ascribed to it in Tern no-
cep lial a is a question worthy of further investigation.
Where, as in many Distomids, a Laurer’s canal is present
that canal seems to be the natural outlet for the unused and
effete materials; when as in Distomum nodulosum,
D. globiporum, D. isoporum, and others (Looss, 11), a
receptaculum is present as Avell, it may act as a true recep-
taculum seminis. When a Laurer’s canal is absent the
receptaculum in some forms — D. variegatum (Looss, l.c.) —
contains yolk-cells as well as spermatozoa. Looss expresses
the opinion that, in general, the spermatozoa contained in
the receptaculum semiuis of Distomids aie in process of
dissolution, and no longer capable of effecting fertilisation.
If this be true of the Distomids as well as Temn ocephala,
it is at least possible that the same may prove to be true of
the other groups — Polyclads, Heterocotylean Trematodes.

THE DEVELOPMENT OF THE TEl\[NOCEPHALE*E. 421

IV. The completed Egg.

The eggs of all the species of Temnocephala, on being
discharged, are fixed by a chitinoid cement to some part of
the outer surface of the body of their hosts. In some of the
species attachment is effected through the intermediation of
a longer or shorter stalk situated at one end. Such stalked
eggs occur in T. chilensis according to Monticelli (14),
Plate (15), and Wacke (16). Similar stalks occur also in
T. no vaj-zealandise and in T. minor. The cementing
material usually extends between the stalks of neighbouring
eggs, thus uniting them into groups as observed by Monticelli
and by Wacke. In sucli stalked eggs an operculum may be
formed, when the young animal is ready to become free, by
the formation of a circular split in the egg-shell near the
distal end. In both T. novEe-zealandiae and T. minor
there is a short filament attached near the middle of the distal
end. T. fasciata, T. comes, T. dendyi, '1'. semperi, and
T. quadricornis have more or less elliptical eggs which
have no stalk, but are cemented down by one side, a number
being, in most cases, united together by means of the cement-
ing material. Of these the eggs of T. fasciata, T. comes,
T. quadricornis, T. semperi are provided with filaments,
those of T. dendyi are devoid of them.

The size of the egg is, in general, in relation with the size
of the adult. The larger species — T. fasciata, T. quadri-
cornis, and T. n o vae-zealan dim — have comparatively
large eggs, about 5 mm. in length. The minute Craspe-
della speuceri, at the other extreme, has oval sessile eggs,
without filaments, which ai’e only 0'2 mm. in diameter.

Temnocephala, like its allies the Rhabdocoeles and the
II eter ocoty 1 ea, has no larval stage; the young animal,
when it escapes from the egg, differing from the adult only
in its small size, and in the repi’oductive apparatus not having
attained to complete development.

422

W. A. HASWELL.

The older eggs are to be recognised by the eyes, which are
visible through the shell. These may be placed in such a
way as to show that the young Temnocephala is lying with
its long axis parallel with that of the egg, but in a large
number of cases the position is a transverse one, and occa-
sionally an intermediate condition occurs. This variation in
the direction of the long axis is not due to movement of the
larva in its later stages; the direction varies from the outset,
and regulating the direction of sectioning, except in advanced
embryos, is little more than mere guesswork.

After the ovum has become fertilised, and the egg com-
pleted by the addition of the mass of yolk-cells and the
enclosing shell, it appears to pass without much delay into
the genital atrium, which serves the purpose of a uterus.
Here it may remain some little time before passing out
through the genital aperture ; but the stage of development
which has been attained when the deposition takes place
varies. In most cases an egg from the ootype or atrium
contains an ovum in which the process of segmentation has
not yet begun ; and unsegmented ova are occasionally found
among those attached to the surface of the crayfish ; but
occasionally segmentation is found to be well advanced in a
uterine egg. 1 have never found more than one egg in the
uterus.

When the egg is fully formed, the greater part of its mass
consists of yolk-cells. These are polyhedral cells of an aver-
age diameter of about '050 mm., with granular contents.
Each has a nucleus '015 mm. in long diameter, of oval or
elliptical shape, with a single spherical nucleolus with a dia-
meter of about '005 mm. The yolk-cells are at first quite
distinct from one another (PI. 23, fig. 2) ; but, as develop-
ment proceeds, a gradual coalescence takes place, beginning
at the periphery, and eventually (PI. 24, fig. 7) the entire
mass completely fuses to form a syncytium, in Avhich all
trace of cell outlines has become completely lost. When the
formation of the syncytium has begun, the nuclei of the more
superficial cells pass outwards and come to lie close to the

THE DEVELOPMENT OE THE TEMNOCEPHALE.E. 423

surface^ so that they present the appearance, to some extent,
of the nuclei of a superficial epithelium.

The ovum (PI. 23, fig. 2) is embedded in the yolk-cells,
usually towards the middle, sometimes towards one end of
the egg. It is a polyhedral cell, ‘08 mm. in diameter, with a
finely granular protoplasm that stains much more deeply than
the substance of the yolk-cells. In all the specimens I have
had the opportunity of examining the nucleus has undergone
modification, and is represented by a cluster of clear vesicles,
each enclosing a rounded particle with the staining affinities
of chromatin.*

In two cases I found an egg containing two cells. The
nuclei of both had undergone the modification just described.
One of the cells was very much smaller than the other, and
on that account it seems to be more probable that we have
here to do with the first staj^e of segfinentation rather than
with an egg in which two ova had become enclosed.

V. I'Iahly Develoi’men'I'.

'Idle process of segmentation results in the formation of a
blastoderm of irregular shape, which comes to be drawn out
in the direction of the long axis of the future worm — a direc-
tion, as already explained, usually parallel with the long axis
of the egg, but not invariably so. No germinal layers are
recognisable : but from a very early stage (PI. 23, fig. 3) the
blastoderm is found to consist of three sets of cells, which
differ from one another in a very marked manner in their size
and in the character of their nuclei. The cells of one set are
0'03 mm. in diameter, have nuclei about 0‘015 mm. in dia-
meter, each containing a large rounded nucleolus. Those of
the second set are 0‘12o mm. in diameter, have smaller nuclei,
0‘0075 mm. in diameter, usually without nucleoli, but with a
rather close network. The cells of the third or smallest set

‘ Somewhat similar appearances were observed by Zeller (17) in
Polystomum.

424

W. A. HA SWELL.

are 0’0075 mm. in diameter, with correspondingly small
nuclei. No definite arrangement of these cells is recognis-
able until a few of the larger cells become grouped together
(PI. 24, fig. 5) in such a way as to bound a small rounded
cavity. This elongates and widens (PI. 24, fig. 6), the bound-
ing cells meantime increasing in number. Eventually (PI. 24,
figs. 7, 8, 9, and 10) the space dilates very greatly, the cells
which form its walls becoming- correspondingly extended,
and uniting together to form a comparatively thin membrane
with flattened nuclei. As it enlarges, this space becomes
approximated towards the surface of the egg, coming to be
separated from the shell only by a thin layer of yolk. In
apposition with the deeper side of the space lies the main
mass of the blastoderm, which is rapidly increasing in extent,
the increase being mainly due to the multiplication of the
middle-sized cells.

The space above referred to does not correspond, so far as
I have been able to ascertain, to anything that has been
found to occur in any other group of animals. Since it plays
an important part in development, it is necessary to have a
name for it, and I propose the term endocoele as one not
involving any dubious homologies.'

The rudiment of the brain (PI. 24, figs. 8, 9, and 10, hr.)
makes its appearance as the endocoele approaches its maxi-
mum size. It appears first as a bilobed, dense aggregation
of cells on the deeper dorsal side of the endocoele, a little
distance from the lining- membrane. In the middle of this
appears a transversely elongated space filled with finely
fibrillated matter — “ Punktsubstanz.” Nerve-fibres (or nerve-
tubes) are only developed in the latest embi-yonal stages.
From the central mass a pair of processes — the foundations
of the peripheral nervous system — are given off laterally.

About the same time as the beginnings of the nervous
system, appears the first rudiment of the excretory system of
vessels. This takes the form of several speci:illy modified

* Tlie same cavity with the same relations occurs in Craspedella
as well as Temnocephala.

THE DEVELOPMENT OP THE TEMNOCEPHALE.E.

426

large cells^ on either side of the rudiment of the brain, and
somewhat behind it. These cells are situated immediately
below the thin epithelial lining of the endocoele cavity. One
cell becomes considerably enlarged, and a narrow sinuous
channel becomes formed in its substance. This channel
becomes continued thi-ough a second and a third cell placed
in close apposition with the first. As subsequent stages sliow,
these constitute the rudiments of the terminal conti'actile
sacs and the beginnings of the main vessels of the excretory
system.

The pharynx is formed from a number of cells which
become arranged after the manner of an epithelium imme-
diately beneatli (i. e. outside of) the thin syncytial epithelium
of the endoccele on the dorsal side. The time of appearance
of this layer varies somewhat. Usually it is not seen until
both the brain and excretory rudiments have become well
established. The part of the Avail of the cavity on which it
is situated becomes somewhat rounded off, thougrli still re-
maining in wide communication with the rest. Posteriorly a
short prolongation without cellular lining extends for a short
distance backwards into the mass of yolk; this represents
the lumen of the intestine (see PI. 24, fig. 11).

The cavity of the endocoele as a whole decreases much in
size. The thin layer of yolk by which it is separated from
the exterior becomes still more attenuated, but still remains
as a definite septum (Pis. 24 and 25, figs. 11, 12, 13, s.),
cutting off the whole internal cavity from the exterior.

^Vhen the brain and the excretory sacs are first formed,
the blastoderm does not extend anteriorly or posteriorly
beyond the limits of the endocoele. But a little later it begins
to grow backwards to form the foundations of the posterior
parts of the embryo. This baclcAvard extension (PI. 25, fig.
14) is made up, like the main body of the blastoderm, of cells
of three sizes, small, intermediate, and large; and these are
arranged in groups with a marked bilateral symmetry. The
large cells are formed by a proliferation of the membi’ane
lining the endocoele. Outlying small cells are to be found

426

W, A. HASWKLL.

here and there embedded in the yolk, at a little distance from
the main body.

In the next stage observed this posterior elongation of the
blastoderm has reached the ventral surface, in what is destined
to be the genital region, and has become almost completely
separated off from the anterior part, in which the further
development of the brain, the pharynx, and the exci’etory
system is going on. There thus come to be two distinct foci
of development, an anterior and a posterior. From both
superficial cells must be separated off to form the epidermis,
since this layer is to be recognised as a distinct, though very
thin layer, with widely separated nuclei, at the stages when
the tentacles and sucker first begin to be formed. This layer
is early completed on the ventral surface, but on the dorsal
surface it does not make its appearance till a considerably
later stage.

The rudiments of the tentacles (PI. 24, fig. 11) make their
appearance at a stage when certain changes in the cavity
have led to the first differentiation of the pharynx, and when
the rudiments of the eyes have first become distinguishable.
They appear as processes which grow at first straight for-
wards from the anterior extremity of the body; but as they
elongate, they become bent (PI. 25, figs. 12, 13), usually
towards the ventral, but in many cases towards the dorsal,
side. In the former case the cephalic portion of the body
(PI. 25, fig’. 13) becomes strongly flexed ventrally at the same
time. When this takes place the part of the surface covered
by the reflexed tentacles develops a system of minute tooth-
like epidermal papillm, which are apparently of firm con-
sistency.

The rudiment of the sucker makes its appearance about the
same time as those of the tentacles. Epidermal papillae,
similar to those underneath the tentacles, are formed under
the sucker.

We have left the endoccfile as an extensive rounded space,
occupying nearly a third of the length of the egg, with a
floor and a roof. As the cavity reaches its greatest dimen-

THE DEVELOPMENT OF THE TEMNOCEPHALE.E. 427

sions the layer of yolk, which separates it ou the ventral side
from the exterior, becomes greatly reduced in thickness. At
the same time, subsequently to the formation of the embry-
onic brain and excretory sacs, a portion of its wall just behind
the brain undergoes modification, a number of large cells
becoming arranged in an epithelium-like manner beneath the
thin lining membrane. The cavity then becomes much re-
duced in size, and the part with the large cells becomes
rounded off to form the pharynx. Ventrally it continues to
be separated from the exterior by the original roof of the
cavity, which becomes reduced to an extremely thin mem-
brane. Posteriorly a further remnant of the original cavity
is represented by a very short passage which ends blindly in
the yolk. The pharyngeal sac and mouth become formed by
growth of the integumentary and muscular layers around the
thin membrane that represents the original roof of the cavity;
but the membrane remains as a distinct, though very delicate,
partition between the buccal cavity and the pharynx as long
as the young animal remains within the egg.

The intestine remains without lumen in the most advanced
stage observed within the egg, a stage in which all the other
parts, even the male reproductive apparatus, have reached an
advanced stage of development. It would, perhaps, be more
correct to say that the young animal has no intestine at this
stage, the site of the future intestine being occupied by a
solid mass of yolk still containing remains of the original
nuclei of the yolk-cells.

VI. Development op the Excretory System.

The excretory system of the adult Temnocephala,
though constructed on the same type as that of the Platodes
in general, possesses certain features which, so far as our
present knowledge extends, are peculiarly its own. The
presence of contractile terminal sacs through which the
system communicates with the exterior is not peculiar to

428

W. A, HASWELL.

this group, similar structures occurring in some of the
Heterocotylean Trematodes (Braun, 2; Groto, 4). But the
nature of these sacs and their relations to the system of
vessels in Temnocephala are quite unlike anything that is
known to occur in other forms.

The most essential features of this system were described
by me in 1893 (8). But since some of the most important of
these points have been overlooked by recent writers,^ or
their significance not recognised, it seems desirable to give a
brief resume of the facts.

Each of the two dorsally situated excretory apertures leads
into a thick-walled contractile terminal vesicle (PI. 25,
fig. 18), which is of pyriform shape, bent on itself towards
its apex. The contractions of the walls of the vesicle are
effected by the agency of an enclosing’ layer of muscular
fibres, and the external aperture is surrounded by a mus-
cular sphincter. Between the muscular layer and the proper
wall of the vesicle is a layer of loose parenchyma, which
doubtless facilitates freedom of movement. The vesicle
itself consists of two large cells fused together and hollowed
out to form the lumen. The greater part of the wall of the
vesicle is formed by one of these two cells ; the narrower
apical part, and the beginning of the main duct which is
given off from it by the other. The relative position of the
two cells is indicated mainly by the position of their two
nuclei, but tlie texture of the protoplasmic substance of the
two cells differs somewhat in character, and that of the
smaller is much more susceptible to the action of staining
agents.

The inner surface of the vesicle is quite smooth and uni-
form. Where an occasional exception to this appears to
occur, and the usually sharp internal outline appears blurred,

* Plate (15) and Wacke (16) for example. The former states, “An
den Nepliridien der Temnocephaliden ist bis jetzt vergeblicli nacb den
fiir die Platybelmintben so characteristiscben Flimmerzellen gesucbt
worden.” Yet I bad described the system with its flame-cells in a
paper in the ‘ Zoologiscben Anzeiger’ two yeai-s before (1892).

THE DEVELOPMENT OF THE TEMNOCEPHALEiE. 429

and the lumen is occupied by fine filamentous matter that
might be mistaken for cilia, this occurs in such an irregular
way that I have little doubt that this appearance is due to a
rupture or other alteration that has occuri’ed during the
fixing process. In no case, either in Temnocephala
no vae-zealan d iae or any other form, is there an epithelium
with scattered nuclei, as supposed by Wacke. Tn some
preparations the internal contour appears double, as if the
cavity of the vesicle possessed an excessively thin cuticular
lining, but this is always very indefinite, and in many cases
is not to be detected. The only nuclei in the entii-e organ
ai-e the two already referred to as the nuclei of the two
constituent cells.

Arising from the main excretory trunk at a little distance
from the vesicle is a special branch of considerable size — the
vesicular vessel as it may conveniently be termed. This
runs inwards, and enters the wall of the vesicle on its inner
side. Here it breaks up (PI. 2.5, fig. 19) into a number of
branches, which ramify throughout the protoplasmic sub-
stance of the wall of the sac in all directions. In the course
of the system of fine intracellular capillaries which is thus
formed occur numerous ciliary flames of small size, but in
other respects similar to the ciliaiy flames in the flame-cells
of other Platodes. I have counted as many as fifty of these
ciliary flames in movement at one time in the case of
T. no va3-zealandias, and probably many more than that
number are actually present.

In sections of the terminal vesicle in all the Australasian
species the ramifications of the vesicular vessel are very
conspicuous, pervading the protoplasmic wall in all direc-
tions. But the ciliary flames are not to be made out with
any certainty save in the living animal.

In most cases the inner part of the protoplasmic wall in
sections appears regularly divided by fine parallel vertical
lines, and one might be tempted to suppose that these repre-
sent a system of vertical canals forming outlets from the
system of vesicular capillaries into the lumen of the vesicle.

4:30

W. A. HASWELL.

From the unbroken appearance of the limiting line of the
surface ou which the vertical lines terminate, I incline to the
opinion that no such communications exist.

The details of the arrangement of the vessels differ in the
different species. The main trunk soon bifurcates to form
anterior and posterior main vessels, which give off numerous
branches to ail parts of the body. A large vessel runs along
the axis of each of the tentacles.

Given off from the larger vessels in the body is a system of
fine, thin-walled capillaries, which are most abundant near
the dorsal surface, where they form an extensive plexus.

A limited number of ciliary flames are to be detected in
the living animal distributed throughout various parts of the
body and the tentacles. The relation of these to the vessels
of the excretory system still remains undetermined. In no
case was a nucleus observed in close relation with the ciliary
flame.

The walls of the larger vessels consist simply of a fine-
grained, structureless protoplasmic material. Here and there,
usually at long intervals, are the nuclei of the elongated cells
of which the walls are composed. These are comparatively
few in number even in the larger species. Their presence
and their relations to the vessels are best observed in longi-
tudinal sections — most readily in the tentacles, in which snch
appearances as those represented in figs. 16, 17, 18 of pi. x of
my “Monograph ” are i*eadily recognisable.

Some of the excretory vessels end in certain specially
modified large excretory cells. The branch in question,
sometimes fairly thick-walled, sometimes very delicate, enters
the cell and breaks up into a richly ramifying and anasto-
mosing system of minute capillaries within its substance.

The first trace of the system of excretory vessels makes its
appearance at a very early stage in the history of the embryo.
In a blastoderm in which the endocoele has become developed,
bnt is still very small, and is bounded by thick massive cells,
there may be observed (PI. 25, figs, 15 and 16) on each side
in close apposition two cells which have the appearance in

THE DEVELOPMENT OF THE TEMNOC'EPHALEHL 431

section of being' pierced by an exceedingly fine, perfectly
clean-cut canal. How these canals end I have not been able
to determine: but subsequent stages show that these, with
the cells that contain them, are the foundations of the excre-
tory system.

At first these perforated cells are widely separated from
the endocoele cavity and embedded in the thickness of the
blastoderm. As the cavity increases in size their relative
position becomes altered, until they come to lie directly
below the lining meinbi'ane of the cavity. At first the entii'e
structure consists on each side of two cells, a larger and a
smaller, which have fused, and the substance of which has
become perforated by a sinuous canal. This is destined to
give rise to the terminal sac of the excretory system. The
canal extends through several cells situated close to the first
two, and in this way is formed the beginnings of the main
longitudinal vessels. Later (PI. 25, fig. 17), when the rudi-
ment of the brain has become well advanced, the terminal
sacs, while still retaining their position immediately under —
i. e. external to — the membrane lining the cavity, and, while
still continuing each to consist of only two fused cells per-
forated by a canal, assume a more complex structure, and
take on, in all the most essential points, the structure which
we have found to chai-acterise them in the adult. The canal
becomes wider in relation to the thickness of the enclosing'
wall ; and from the main vessel is given off a slender branch
— the future vesicular vessel — which, approaching' the terminal
sac on the inner side, breaks up into a nnmber of excessively
fine capillaries that ramify through the substance of the wall
of the sac.

It will be seen from the above account of its mode of
formation that the excretory system of Tern no cep ha la
is, in the strictest sense, of intracellular character. I am
thus compelled to dissent from Goto’s opinion (4, p. 71) that
I was not justified iu using that term, as well as to his more
general view (p. 74) that “the term intracellular is quite in-

voL. 54, PAirr 3. — xkw sekies. 31

432

W. A. UASWELI..

appropriate to the excretory system of the Trematodes and
the Turbellaria.”

Perhaps the most remarkable event in the history of the
development of the excretory system in Temnocephala is the
change which takes place iu the position of the contractile
terminal sacs. Originally, as we have seen, they are placed
close to the epithelium of the endocoele, and apparently open
into the latter. When the cavity becomes reduced, and the
pharynx begins to become rounded off, the sacs lose their
original connections, and, becoming displaced outwards,
enter into connection with the epidermis, and open on the
exterior on the dorsal surface. Thus, iu a stage in which the
rudiments of the tentacles ai’e being formed, the sacs occupy
the position which they retain in the adult.

VII. Development of the Alimentary System.

The alimentary system of Temnocephala consists of two
principal parts — pharynx and intestine. The mouth, situated
far forwards on the ventral surface, leads, through a very
small cavity representing a pharyngeal sac, into the lumen of
the pharynx. The latter is a large, rounded organ with
thick walls of highly complex structure. The cavity is lined
internally by a layer, the nature of which is by no means
clear. By Weber (18) it has been described as a continua-
tion of the cuticle of the integument. Mouticelli (14) refei’s
to it as a syncytial epithelium. Wacke (16) refers to it as
an epithelium, and gives excellent figures of its minute
structure. It is a non-cellular layer, in which a degenerate
nucleus may sometimes be detected, but only quite excep-
tionally. It is composed of gi’anular material, the granules
of which are arranged in rows or strings, most, at least in
the Australasian species, having a vertical direction. Many
of these strings are traceable in some series into the thick-
ness of the wall of the pharynx. The internal cuticle,
described and figured by both Monticelli and Wacke as

THE DEVELOPMENT OF I'HE TEMNOCEPHALE.E. 433

bounding this layer internally, is not a separate and distinct
layer, but is merely the innermost stratum of the granular
layer, and in most preparations is not differentiated at all.

Externally a. limiting membrane forms a pharyngeal cap-
sule, separating the muscular mass of the pharynx from the
surrounding parenchyma. Between the external capsule
and the internal epithelium, in addition to the elaborate
system of muscular fibres, there are a number of cells and a
system of nei’ve-fibres. The cells are of several kinds, com-
prising bipolar nerve-cells, excretory cells, and unicellular
glands, the ducts of which are usually said to open into the
cavity of the pharynx.

I’osteriorly the pharynx leads through a short passage —
the oesophagus — into the spacious intestine. Bound the
oesophagus are a number of unicellular “ salivary ” glands.

The intestine in all the Australasian species (and also in
T. chile usis according to Wacke) is constricted at intervals
by a number of annular muscular dissepiments, the number
of these varying with the species. The epithelium is com-
posed of long narrow cells, the majority of which, though
probably mainly absorptive in function, contain vacuoles
enclosing granules which are probably excretional, while
others are of the nature of unicellular digestive glands.

The pharyngeal sac, the pharynx with the oesophagus, and
the intestine are all derived from different sources. The
first may be said to be of the nature of a stomodseum. The
pharynx is derived from a portion of the wail of the eudo-
coele. A number of large cells become arranged in a manner
presenting the appearance of an epithelium on the wall of
this region, separated from the internal space by the thin
epithelial liniug of the cavity. This specially modified por-
tion of the wall then becomes rounded off as the wall of the
pharynx, what remains of the cavity, which becomes much
diminished in size, forming a short passage corresponding in
position with the future oesophagus, and terminating abruptly
behind in the mass of yolk.

The large cells in the wall of the embryonic pharynx at

434

W. A. HASWf]LL.

first form its entire thickness, with the exception of the
exceedingly thin internal epithelium. Bnt afterwards mus-
cular fibres ai’e developed both internally and externally.
The mode of formation of these is not clear, but since no
other elements come into play, tliere can be little doubt that
a portion of the cells of the wall of the pharynx are of the
nature of myoblasts. The rest become the nerve-elements
and the glandular and excretory cells.

The mode of formation of the so-called epithelium of the
pharynx is a matter of some interest. It is represented at
first, as already pointed out, by the thin syncytial lining of
that part of the endoccele from which the pharynx becomes
developed ; and at no subsequent stage does its epithelial
character become more pronounced. On the contrary, as
development advances, the cellular character of this layer
becomes almost or completely lost. In fact, were it not for
the occasional occurrence in it of a more or less altered
nucleus, it might be supposed to be entirely non-cell ular.
But, though it loses, or nearly loses, its cellular character,
this layer greatly increases in thickness, and, in the adult,
maintains its thickness in spite of the loss of material to
which it must be subjected as a result of constant wear and
tear during the capture and ingestion of active living prey.

Up to a late stage in Temnocephala fas data the rest
of the digestive system is merely represented by a short
passage, which continues back the lumen of the pharynx,
and ends abruptly within the mass of yolk. This is the
future oesophagus, and in its neighbourhood are a number of
cells destined to become the unicellular or “ salivary ”
glands. Thus, at a stage when the muscular wall of the
pharynx, with its anterior and posterior sphincters, has
reached an advanced stage of development, the intestine is
not yet definitely represented.

At this stage the cavity of the pharynx is still completely
shut off from the exterior by the septum formed, as already
described, from the persistent thin roof of the original endo-
coele.

THE DEVELOPMENT OP THE TEMNOCEPHALE.E.

435

The stages by which the intestine, with its definite epithe-
lium and septa, becomes differentiated, have not yet been
ti’aced. It seems probable, however, that a structure which
makes its appearance below the endocoele, after the brain and
excretory sacs have become formed, may be concerned in the
formation of the primitive endoderm. At this stage (PI. 25,
fig. 20), a cleft or infolding appears among the cells on the
floor of the cavity. Later this takes the form of a group of
cells arranged around a small lumen — the appearance being
very similar to that presented by the first beginnings of the
endocoele itself. Since this is behind the brain and excretory
sacs, and is too far forward to be of the nature of a genital
primordium, it is permissible to suppose that it represents
the earliest rudiment of the endodermal system. Its further
history has not been traced; in fact, it soon disappears
as such ; but if the above supposition as to its nature be
correct the cells to which it gives rise must extend round the
yolk between it and the rest of the developing parts, and
become converted into the intestinal e]uthelium.

In the latest stage observed within the egg — a stage with
fully-developed eyes and abundant pigment, and with the male
part of the reproductive apparatus far advanced — the intes-
tine is still entirely devoid of lumen, and consists of a solid
granular mass with numerous nuclei, which are most abundant
in the peripheral zone. Slender strands passing inwards into
this solid mass represent the future dissepiments. In the
case of T. novm-zealandiae, however, in the latest stages
from the egg ttie intestine has developed a lumen, and the
epithelium is recognisable, though the septum still persists
shutting off the cavity of the pharynx from the exterior. It
is a remarkable fact, which is probably of significance in
connection with the function of this persistent septum, that
in nearly all sections of late stages with fully-developed
pharynx the lumen of the latter is found to be filled with a
mass of yolk-granules wliich have evidently become detached
from the central body of yolk, and have been prevented by

\V. A, 11ASWE1;L.

4^?a

the occluding septum from reaching the exterior, i. e. the
space between the embryo and the shell.

VIII. Development of the Epidermis.

In the integumentai-y system of Temnocephala the most
characteristic feature is the presence of a well-developed
nucleated epidermal layer of a syncytial character. In the
embryo this is not represented by any definite primordium,
such as has been observed to be formed in Polyclads (Lang
and others) and Rhabdocoeles (Caullery and Mesnil [4],
Breslau [3]), and there is no process corresponding to the
process of overgrowing of the embryo by an epidermal layer,
such as occurs in these groups. When an epidermis is first
disceimible, it consists of a very thin membi-ane with wide-
apart flattened nuclei, covering only the ventral surface, and
there is no evidence of any process of proliferation such as
must accompany the spreading of the edge of this la3’er by
cell-division. It would appear, in fact, as if the epidermis
were formed by cells migrating to the surface and there
becoming flattened out and united together to form the
syncytium.

Temnocephala fasciata, unlike T. minor and T.
Dendyi, has no cilia on the surface in the adult, and
I have found no trace of them at any stage in the deve-
lopment.

IX. Development of the Reproductive System.

The details of this process have not yet been followed out.
In all the species examined the male part of the reproductive
apparatus is developed at an earlier stage than the female.
In the later stages within the egg the penis, the vesicula
seminalis, the vasa deferentia, and the testes are all well
advanced. In T. fasciata each testis is represented by a

'I'HK DEVELOPJrKNT OF THE TEMNOCEPHAl.E.E.

437

mass of primordial cells surrounded by indifferent cells. In
T. novae-zealandite before the young animal leaves the egg
the first stages of spermatogenesis have occurred. At this
stage the atrium does not yet open on the exterior. The
definite development of the female part of the apparatus does
not begin till after hatching, so that AVacke’s statement
that Temnocephala is protandrous appears to have some
evidence in its favour, though a study of the post-larval
development will be necessary in order to decide whether
the condition is one of actual functional protandry, and not
mei’ely one of more active development of the male apparatus
in the early stages.

The unique features of the early development of Temno-
cephala are associated with the formation of the remarkable
internal cavity (endocoele) around which the foundations of
various systems of organs are laid down. As nothing parallel
to this cavity has, so far as my knowledge extends, been met
with in other groups,^ it is natural to inquire if its pi’esence
can be associated with any special conditions under which
the development takes place — if its occurrence can be sup-
posed to be of the nature of an adaptation.

The Australian fresh-water Crayfishes, on which live all
the known Australian members of the Temnocephalefe, shelter
or support a great variety of small Invertebrata. A consider-
able proportion of these live in the branchial cavities, but many
adhere to various parts of the outer surface. Among sucli
dependents of the Ci’ayfishes are Protozoans, Nematodes,
Rhabdococles, Rotifers, Stratiodrilus, Phreodrilus, a parasitic
Ilydrachnid, and others. In a country subject at times,
as Australia is in many parts, to long-continued di’oughts,

* It might 1)6 possible to trace some coiniection between the embryonal
pharynx of the Tricladida (Halle/. [6], Metscbnikoff 112], lijima [10] )
and the endocoele of Temnocephala. Both cavities arise as spaces in
rounded groups of cells in the blastoderm ; but the former opens on the
surface and swallows yolk cells, with which the intestine becomes dis-
tended. Moreover it is jmrely provisional, and disappears entirely, a
new pharynx becoming developed in its place.

488

\V. A. UASWEfJ,.

aquatic organisms must often suffer wholesale destruction as
a result of the drying-up of the smaller streams. Crayfishes
are able to avoid such a fate by sheltering between boulders
or burrowing deeply in the bed of the stream. The animals
adhering to them thus have a chance of survival denied to
their free-living relatives. Yet it may, and doubtless does,
often happen that even the Crayfishes are unable to escape
the risk of desiccation. Under such cii’cumstances the
presence of a relatively large space filled with water in the
intei'ior of the egg of the Temnocephalese may make all the
difference in enabling the embryo to retain its vitality until
the dry period passes and the stream fills again.

Literature cited.

1. Benedeu, E. von — “ RechercJies sur la comjjosition et la significa-

tion (le roBnf,” ‘ Mem. Acad. Roy. de Belgique,’ tome 34 (1870).

2. Braun, M. — “ Trematodes ” of Bronn's ‘ Klassen imd Ordnnngen

des Tliierreiclis ’ (1879 — 1893).

3. Bresslan, E. — “ Beitrage znr Entwickelungsgeschiclite der Turhel-

larien I,” ‘ Zeitsclir. f wiss. Zool.,’ 76 Bd. (1904).

4. Goto, S. — “Studies on the ectoparasitic Trematodes of Jai^an,”

‘ Journal College of Science, Imperial University of Japan,'
vol, viii, i^art I (1894).

5. Graff, L. von. — “ Turhellaria.” Bronn's ‘ Thierreich,’ iv (1908).

6. Hallez, P. — ‘ Embryogcnie des dendrocoeles d'eau douce,’ Paris

(1887). ‘

7. “ Sur la nature syncytiale de I'intestin des Rhahdocoeles,”

‘ Comptes Rendues,’ tome 146, pj). 1047-9 (1908).

8. Haswell, W. A. — ‘ A Monograph of the Temnocephaleae,’ Macleay

Memoiual Volume, Linnean Society of New South Wales (1893).

9. “ Note on the Fauna of the Gill Cavities of Freshwater

Crayfishes," ‘ Repoi’t Austral. Assoc. Adv. Sci.,’ vol. 8 (1901).

' I have not been able to see this paper. What I have learnt regarding
its contents has been obtained from Graff (5).

THE DEVELOPMENT OP THE TEMNOCEPHALEHL 439

10. lijima, J. — “ Untersuclnmgen liber den Ban und die Entwickelimgs-

geschichte der s.-w. Dendrocoelen,” ‘ Zeitschr. f. wiss. Zool.,’ Bd. 40
(1884).

11. Loos, A. — “ Die Distomen unserer Fisclie u. Frosclie,” ‘ Bibl. Zool.,'

Heft Hi (1894).

12. MetsclmikofF, E. — “ Die Embi-yologie von Planaria polycbroa,"

‘ Zeitscbr. f. wiss. Zool.,’ Bd. 38 (1883).

13. Monticelli, F. S. — Breve nota sulle nova e sngli einbrioni della

Temnocephala chilenses, Bl.,’ ‘ Atti Soc. Ital. Sci. Nat.,'
vol. 32.

14. “Sulla Temnoceijliala brevicornis. Montie., e sidle

Temnocefale in generale,” ‘ Bull. Soc. Nat. Napoli,’ 12.

15. Plate, — . — “ Mittlieilungen ueber Zool. Studien an der Cliilenisclien

Kuste, VIII,” ‘ S. B. Akad. wiss. Berlin ’ (1894).

16. Wacke, Robert. — “ Beitrage zur Kenntniss der Temnoceplialen,'’

‘ Zool. Jahrb. Supplement,’ Bd. vi, heft 1.

17. Zeller, E. — “Weiterer Beitrag zur Kenntniss der Polystoiuen,”

‘ Zeitscr. f. wiss. Zool.,’ 27 Bd. (1870).

18. Weber, Max. — “Ueber Teninoceiihala, Blanch,” ‘ Zool. Ergebnisse

einer Reise in Niederl. Ostindien,' Heft 1 (1889).

[EXPLANATION OF JM^ATES 23— 25,i

Illustrating Professor W. A. HaswelPs paper on “ The
Ueveloptnent of the Temnocephalea3.”

Lettering.

br. Brain, e. Supposed endoderni primoi’diuni. en. Endocoele. cp.
Epidermis, ep. en. Ejiithelial lining of endocoele. ex. Excretory sac or
excretory cells, ey. Eye. o. Ovum. o. p. Oral papilla', jdi. Pharynx,
s. Roof of endocoele, becoming septum, cutting off lumen of pharynx
from the exterior, t. Tentacle. ;/. Yolk-cells, y'. Yolk syncytium.

PLATE 23.

Fig. 1. — Longitudinal section of the ovary of Temnocei^hala
comes. X 50U.

‘ All the figures have been re-drawn by Mr. A. C. Cronin, of the
Water and Sewerage Department, Sydney, from my original drawings.

440

W. A. JIASWELL.

Fig. ‘2. — Transverse section through uterine ovum of T. fasciata.
passing through the unsegmented ovum. X 250.

Fig. 3. — Section through an early lilastodenn in which three kinds of
cells have become distinguishable. X 500.

PLATE 24.

Fig. 4. — Section through a later stage, with groups of larger, lighter,
surrounded by smaller, more darkly stained, cells. X 50o.

Fig. 5. — Section through a lilastoderm in which the first definite
trace of the cavity has made its appearance. X 500.

Fig. 0. — Later stage in the development of the cavity. X 500.

Fig. 7. Longitudinal section through the entire egg, with a more
advanced cavity. X 170.

Fig. 8. — Portion of a longitudinal section of egg passing through the
cavity, and cutting the blastoderm nearly transversely. X about 330.

Fig. 9. — Similar section of a later stage with the brain more
advanced. X about 330.

Fig. 10. — Similar section, cutting the blastoderm nearly transversely,
Imt with some oblicpiity, so that it passes through the brain and one
excretory sac. X 330.

Fig. 11. — Anterior part of a vertical longitudinal section, approxi-
mately median, at a stage when the rudiments of the tentacles and eyes
are appearing. X aljout 330.

PLATE 25.

Fig. 12. — Entire longitudinal and vertical section of embryo with
the tentacles further developed than at the stage represented in fig. 11,
and flexed backwards over the mouth. X about 130.

Fig. 13. — Anterior part of a longitudinal and approximately median
and vertical section of an emliryo with well-developed, ventrally flexed
tentacles. I

Fig. 14. — Transverse section through the blastoderm in the region
behind the endocoele at a stage somewhat later than that represented in
fig. 10, showing the posterior jjrolongation comprising the genital rudi-
ment. X about 500.

Figs. 15 and 10. — Successive sections of an embryo at an early stage
in the formation of the endocade, showing an early phase in the history
of the cells destined to form one of the excretory sacs. X about 500.

THE DEVELOPMENT OP THK TBMNOOEPHAl.E.E.

44 i

Fig. 17. — Portion of a section passing through the endocoele at a
stage similar to that represented in fig. 9, showing the developing
excretory sac, and its connection with the endocoele.

Figs. 18 and 19. — -The excretory sac in the adult; fig. 19 shows a
f)art of the system of capillaries in which are numerous dame-cells,
given off from the vesicular vessel and ramifying through the wall of
the sac. Reproduced from a ' Monograph of the Temnocephaleas.’
pi. X, dgs. 11 and 12.

Fig. 20. — Portion of a section of an egg with far advanced endocoele
and distinct brain and excretory sacs, showing what is supposed to be
the first trace of an endodermal primordium.

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EXPERIMENTAL OBSEEVATIONS ON PENNATULIDS. 443

Experimental Observations on the Organs of
Circulation and the Powers of Locomotion
in Pennatulids.

By

Editli M. Miisgrave, D.Sc. (nee Pratt),

Late Honorary Research Fellow in the University of Manchester.

With Plates 26 and 27.

The study of Pennatulids has long engaged the atten-
tion of zoological workers. In the publications of the eigh-
teenth century wonderful and fantastic accounts are given of
the habits, form and movements of members of this interesting
family ; of their graceful flight through the water, their
exceeding luminosity by night, and their gymnastic contor-
tions even in captivity. In those early days of zoological
research a “ sea pen ” was regarded as a single individual
provided with lateral “ fins ” for swimming purposes, and
a mouth opening at the base, Avhich served also for the excre-
tion of waste matters.

Later accounts on the subject are written in a more tem-
perate spirit, and in them we observe a general contradiction
of the apparently extravagant statements of early writers.

In the tenth edition of the ‘ Systema Naturm ’ of Linnaeus
(1758) a diagnosis of Pennatula phosphorea is gdven, in
which mention is made of a basal opening to the exterior (os
baseos commune rotundum). Six years later Ellis (1764,
pp. 419-428) published an excellent account of an investigation
of this species, in which he denies the existence of the mouth
or basal opening. Of the habits of this interesting species he
says :

VOL. 54, PART 3. — NEW SERIES.

32

444

EDITH M. MUSGEAVE.

“ It floats and swims about freely in the sea, whereas
corals, corallines, Alcyonia and all that order of beings
adhere firmly by their bases to submarine substances
This species . . . has been found in the ocean from the

coast of Norway to the most remote parts of the Mediter-
ranean Sea, and not only dragged up in trawls from great
depths of the sea., but often found floating near the surface.”

Dr. Shaw, in his ‘ History of Algiei's,’ remarks that they
afford so great a light in the night to the fishermen that they
can plainly discover the fish swimming about in various
depths of the sea. He states that from this extraordinary
property LinnaBus called this species of sea pen '^Pennatula
phosphorea,” and remarks : Habitat in Oceano fundum

illuminans.”

Of the leaves or pinme of the sea pen Ellis says :

“ These are evidently designed by Nature to move the
animal backwards or forwards in the sea, consequently to do
the office of fins, while at the same time the suckers (auto-
zooids) or mouths furnished with filaments or claws (tentacles)
were certainly intended to provide food for its support, for
notwithstanding what Dr. Linngeus has said in regard to its
mouth in his ‘System of Nature,^ viz. os baseos commune
rotundum, I could not, with the aid of the best glasses,
discover that the point of the base was penetrated in the
least.”

Bohadsch (1761) had already denied the existence of a
basal mouth, and stated that the cavity or sinus, which
usually exists at the base of the stalk (fig. 13), is merely a
space caused by the contraction of the axis.

This assertion is supported by Ellis (p. 430), who makes
the following statement :

“ The bone (axis) is fastened to ligaments at both ends,
which are likewise inserted in both ends of the animal. When
the ligament at the base is contracted, it forms that sinus at
a. a. (Ellis, 1764, figs. 1 and 2, pi. xx) which has been taken
for a mouth by most authors.”

So convincing, apparently, was the joint denial by

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 445

Bohadscli and Ellis of the existence of a mouth or of any
basal opening to the exterior that the statement ‘'os baseos
commune rotundum” was omitted in the diagnosis of the
genus in the eleventh and subsequent editions of Linnmus’
‘ Systema Xatnrte.’

Later, when the knowledge of the colonial habit of the
“sea pens” had become established ‘Ellis and Solander’
(1786), in a joint publication, made the following state-
ment :

“ They have no opening at the bottom as was formerly
thought, nor any other passage but through their polyp
mouths”; but notwithstanding this emphatic denial 0. F.
^Miiller (1788) recorded the presence of a pore at the base of
the stalk in Kophobelemnon, and later Della Chiaje (1827)
described, with accompanying illustrations, the interesting
occurrence of a basal pore in specimens of Pteroeides
spinosnm, Pteroeides griseum and Pennatula rubra,
and furthermore stated that the pore communicates with the
canals in the stalk.

These observations were confirmed by Kolliker (1872),
Avho noticed in some cases a single pore, and in others two
pores at the base of the stalk in Pteroeides and Penua-
tula. In 1875 Kolliker’s statement was further confirmed by
Schultze, but Hubrecht (1882, p. 572) was unable to observe
any basal aperture in the case of the new genus Echino-
ptilum,and Marshall (1887, p. 29), who appears to have made
the most recent observation on the subject, remarked that
he was unable to confirm KollikeFs statement with regard to
the presence in Pennatula of a minute orifice at the base of
the stalk opening into the canals.

I have therefore sought experimental evidence, which is
recorded in the present paper (pp. 452-459), in the hope of
obtaining a satisfactory and final solution of this questionable
point.

The early accounts of the active swimming movements of
Pennutalids have been, during recent yeai's, somewhat dis-
credited. There can, however, be little doubt that they

446

EDITH M. MUSGRAVE.

exhibit, under normal conditions, certain controlled move-
ments of the colony apart from the mere expansion and con-
traction of the zooids.

The earliest account of such movement was given by
Lancaster^ (1601) who, on attempting to pluck up a growing
Virgularia, observed it to ‘^shrink down to the ground
uidess held very hard.”

A later account of the movements of the colony was given
by Bohadsch (1761, p. 424) who makes the following state-
ment with regard to the conduct of a living specimen of
Pennatula phosphorea in sea-water;

“ The trunk was contracted circularly at the bottom of the
naked part of the stem, and by this contraction formed a zone
of the most intense purple, which moved upwards and down-
wards successively. When it moved upwards through the
length of the pinnated trunk it there became paler, and at
length terminated at the top ; the motion being scarce finished,
a like zone appeared at the end of the naked trunk, which
finished its motion in the same manner as tlie former.”

The peristaltic dilatation and contraction thus originally,
described by Bohadsch has been confirmed by Kolliker,
who suggests that the boi-ing in the sand is effected by this
means, but the statement also made by Bohadsch in the same
publication that a specimen of Pennatula was seen “swim-
ming freely with all its delicate transparent polypi expanded
sailing through the still and dark abyss by the
delicate and synchronous pulsations of the minute fringed
arms of the whole polypi ” still requires confirmation, although
Ellis and Solander (1786), in the diagnosis of the genus
Pennatula, state:

“This genus of animals differs remarkably from all other
zoophytes by their swimming freely in the sea, and many of
them have a muscular motion as they swim along-. I know
of none of them that fix themselves by their base.”

In his account of the Penuatulidae in the ‘ Cambridge
Natural History’ (Ccel en terates, p. 360), Hickson writes:

' See Darwin, ‘Naturalist's Voyage round the World,’ p. 99, 1845.

EXPEEIMENTAL OBSERYATIOXS ON PENNATULIDS. 447

“ The sea pens ai*e usually found on muddy or sandy sea
bottoms, from a depth of a few fathoms to the greatest depth
of the ocean. It is generally assumed that their normal posi-
tion is one with the peduncle embedded in the mud and the
rachis erect. Positive evidence of this was given by Rumphius
writing in 1741 in the case of Virgularia rumphii and
V. j uucea at Amboina, and by Darwin in the case of
Stylatula darwinii at Bahia Blanca. “At low water,”
writes Darwin, “ hundreds of these zoophytes might be
seen projecting like stubble, with the truncate end upwards,
a few inches above the surface of the muddy sand. When
touched or pulled they suddenly drew themselves in with
force so as to nearly or quite disappear.”

“It is not known whether the Pennatulids have the power
of moving from place to place when the local conditions have
become unfavourable. It is quite probable that they have
this power, but the accounts given of the sea pens lying
flat in the sand do not appear to be founded upon direct
observation.”

This prone })Osition was, unfortunately, the usual condition
of all the Pennatulids observed in captivity in the expei-i-
menting and exhibiting tanks provided with sandy or pebbly
bottoms in the Naples aquarium. I am also informed by
Dr. Lo Bianco, of that institution, that specimens kept thus in
captivity constantly remain prone, and have never been observed
to display any of those active swimming muscular movements
of the leaves so frequently commented upon by earlv writers.
It is very possible that specimens described as swimming
thus through the ocean depths have become voluntarily or
involuntarily removed from their basal attachments, while the
flapping movements of the leaves may have been induced by
water currents and other mechanical external causes. They
are certainly not induced by muscular effort on the part of the
individual colonies. This question is further discussed in
the present paper.

Bohadsch desci-ibed four different movements of the
“ leaves,” then universally regarded as flns, thus : “ They

448

EDITH M. MUSGEAVE.

are moved both towards the naked stem and towards the
pinnated stem ; sometimes they are drawn in very much to
the belly, a little after they are inclined to the back.”

I regret that I can produce no evidence in support of this
statement. The movement of the leaves was observed in all
cases to be extremely slight, and extended over a consider-
able interval of time. In many cases it was only after several
hours that the leaves became slightly inclined veiitrally
or dorsally upwards or downwards, but the change in posi-
tion was invariably accompanied by a change in their contour,
and seemed to be entirely dependent upon the state of hydro-
static dilatation or otherwise of the colony.

Upon internal examination of the rachis, the tissue around
the insertion of the leaves was observed to be composed almost
entirely of that distensible spongy tissue which we have reason
to suppose plays no unimportant part in the dilatations and
contractions of the colony (pp. 463-464). In this region the
musculature is extremely feebly developed, and serves chiefly
for the protraction and retraction of the autozooids, the
coelenteric cavities of which are separated from the large
canals of the rachis merely by a thin connective-tissue layer.
The arrangement of the musculature in connection with the
spongy tissue and controlled movements of the colony is
further discussed on pp. 460-473.

The Canal System.

As very excellent accounts already exist of the form,
arrangement, and general anatomy of the canal system of
Pennatulids it is necessary to add little to what has already
been published on this portion of the subject.

For the earliest account we are indebted to Kolliker (1880),
who described most carefully and thoroughly the arrange-
ment and general anatomy of the canal systems in several
genera: Pen na tula, Pteroeides, Virgularia, etc. He
also states that, with the exception of the presence of the
four large central canals in Pennatulids, the canal system

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 449

is very similar to that of Alcyonium and Corallium
rub rum.

The brothers Marshall (1882) in the classic memoir on the
Oban Pennatulida gave admirable accounts of the canal
systems of Pennatula and Virgularia, in which the obser-
vations of Kblliker are to a great extent confirmed and
supplemented.

Jungersen, 1904, describes for the first time the canal
system of Protoptilum.

Gravier, 1906, gives an account of a curious modification
in the arrangement of the canals in the Virgularian genus
Scytaliopsis.

Notwithstanding the valuable contributions which Kolliker,
Marshall, Jungersen, and Gravier have made with regard to
our knowledge of the general anatomy and arrangement of
the canal systems of Peunatulids, their actual function still
remained a matter of uncertainty. Experimental evidence has
therefore been sought Avith the hope of obtaining some
elucidation of this unknown problem.

All species of the genera Pennatula, Pteroeides, Vir-
gularia and Anthoptilum which 1 have had the oppor-
tunity of examining ag’ree in the following respects :

In the presence of the four large central canals — one
dorsal, one ventral and tAvo lateral canals. The dors.pl canal ^
extends from the extreme base to the distal end of the
colony, and is separated from the ventral canal by the vertical
septum (figs. 12, 13, 14), and the septum terminates inferiorly
by fusion ventrally Avith the connective tissue of the body-
Avall, usually, as iu Pteroeides some little distance above the
basal termination of the stalk (fig. 14), so that the extreme
loAver portion of the stalk is occupied only by the dorsal
canal.

lu Pennatula rubra the vertical septum extends almost
but not quite to the extreme base of tlie stalk (fig. 13).

* I liaA'e used the terms “dorsal” and “A’entral” in accordance
Avith Jungersen’s designation, Avhich is the reverse of that formerly
adopted By Kolliker and other antecedent Awiters.

450

EDITH M. ]MUSGEAYE.

The dorsal aud ventral canals communicate with each
other by means of numerous apertures in, or short canals
whicli traverse the vertical septum.

The two lateral canals are considerably shorter than the
dorsal and ventral canal ; they also communicate with the
latter at fi’equent intervals by means of pores in the inter-
vening septa. In Pteroeides, according to Kolliker, each
of the two lateral canals is in direct communication interiorly
with the lumen of the dorsal canal. In all cases the coelen-
tera of apparently all the zooids communicate by means of
more or less short canals with the four large central
canals. In some cases these connecting canals a.re ciliated as
in Peuuatula inurrayi (Hickson)^ in others they are funnel-
shaped as iu Protoptilum carpeuteri (Jungersen, 1904,
p. 53).

In the stalk, which is usually destitute of zooids, the large
canals communicate with the exceedingly numerous canals of
the spongy tissue which line the internal body-wall of stalk
and rachis, by means of numerous apertures (figs. 13, 14,
arranijed in longitudinal lines between the insertion of the
long oblique muscles controlling the movements of the axis.

In the genus Virgularia, as in other Pennatulids, the
coelentera of the zooids are iu communication with the
large central canals, but according to the brothers Marshall
(1882) supplementary radial canals are also present iu the
rachis, which do not communicate Avith the coelentera of the
zooids, but run outwards towards the peripheral tissues.

In Scytaliopsis (Gravier, 1906) similar tubes arc appa-
rently present, but in this genus they open dii-ectly to the
e.xterior. Gravier states that these tubes, with their external
apertures, are extremely numerous, and contrasts their abun-
dance Avith the paucity of zooids in this genus.

It Avould be extremely interesting to determine Avhether iu

' Professor Hickson has drawn my attention to the presence of
ceiiain “ciliated” canals in the rachis of Pennatula murrayi.
These canals ai-e seen in section to he coiuiected hy means of other
canals which appear to he rmciliated, with the large central canals.

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 451

Virgularia the radial canals described by Marshall have
communications with the exterior as in Scytaliopsis.
These supplementary canals, according to Marshall, do not
occur in Pen na tula or Funiculina.

The occurrence of the “radial canals” in Virgularia
is no doubt correlated with the paucity of the zooids. This
modification has probably been brought about by the insuffi-
ciency of the zooids, to serve hydrostatic and possibly also
nutritive purposes.

Nematocysts,

As a general rule the Pennatulids are less formidably armed
with those weapons of offence and defence — the thread-cells
or nematocysts — than the Alcyonidae.

The tentacles are usually smooth and filamentous, and lack
the characteristic warted appearance of those of Alcyonids,
in which each wart is a battery of poisoning and paralysing
thread-cells. Nematocysts of the usual type are present, but
in considerably smaller numbers, and are more or less uni-
formly distributed in the general ectoderm of the tentacles.

From their constitution and habits Pennatulids would appear
to be less voracious in their habits than are Alcyonids gene-
rally, and are apparently more dependent upon the supply of
nutriment suspended in the sea-water in the form of organic
particles, which may be carried into the zooids and canals by
inflowing curi’ents, where they are ingested and digested by
the mesenterial filaments of the autozooids, as in Alcyonium
(Pratt), or by any epithelial surface of the canal system
(fig. 9).

The FiXPERIMENTS.

The experiments recorded in the present paper were
performed in the Zoological Laboratories of the Marine
Biological Station at Naples during the month of April,
1905.

452

EDITH M. MUSGEAYE.

I.

Several colonies of Pennatula rubra and P. phos-
pliorea were placed in tanks of running, filtered sea-water.
At frequent intervals tiny clouds of finel}’ powdered carmine
were squirted around the pens by means of a small glass
S}’ringe. After four days the colonies w'ere submitted to
microscopic examination when particles of carmine were
observed.

(1) In the coelentera of the autozooids and siphonozooids
and in an ingested condition in the cells of the mesenterial
filaments of the autozooids (Pratt, Alcyonium, 1905).

(2) In the canals connecting the coelentera of the zooids
with the four large central canals of the stalk and rachis.

(3) In the lumen of the four large canals just mentioned.

(4) In an ingested condition in the cells composing the
coelenteric lining of the zooids, and also in many of the cells
composing the epithelial lining of the large canals throughout
the colony (figs. 1, 9).

This experiment, in my opinion, indicates the existence of
a very complete and comprehensive system of communication
between every poi’tion of the colony and the exterior, while
the presence of ingested particles of carmine in the cells lining
the coelentera of the zooids and the lumen of the large canals
throughout the colony confirms the opinion based upon
numerous feeding experiments on other members of the
Order Alcyonaria (Pratt, 1905), that these cells possess at
least a nutritive function (fig. 9), and indicates the possi-
bility of the ingestion and digestion by any epithelial surface
(other than that occupied by the mucus-secreting gland-cells)
of any nutritive matter which may be present in the fiuid
circulating so freely within the colony. Of the nature of this
fluid comparatively little is known, and it was, unfortunately,
impossible to obtain a chemical analysis during experimental
operations, but it is evident that this question was not entirely
ignored even in a period so remote as the eighteenth century,
for Bohadsch (17t5l), in the account of his investigation of
living specimens of Pennatula phosphorea, states :

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 453

“ Wheu tlie trunk is opened lengthways a saltish liquor
flows out of it, so viscid as to hang down an inch.”

The viscidity of the fluid contained in the canals was notice-
able in all the specimens which came under my observation,
and is, in my opinion, attributable to a mucoid secretion
which exudes in considerable quantity from a profusion of
gland-cells in the epithelial linings of the lai'ge canals (figs. 8
and 9) of the stalk and rachis. In this viscid exudation from
the deeply staining gland-cells particles of carmine were
often observed.

After an interval of a hundred and twenty years, during
which period the zoological records are mute on this subject,
Kolliker (1880) published a carefully compiled account of the
canal systems of Pennatulids, but apparently makes no state-
ment as to the nature of their fluid contents. Tlie canals, how-
ever, he describes as “ nourishing canals,” so that doubtless he
considered the probability of the presence of nutrient matter
in the liquids contained in them.

Later the brothers Marshall (1882, p. 33) made a statement
to the effect that the fluid contained in the canal system of Pen-
natulais probably nutritive but mixed largely with sea- water.

After a comparative study of the arrangement and anatomy
of the canal system of Pennatulids and other Alcyouaria
(Alcyouium, Corallium, etc.) and their physiological
beliaviour and condition when subjected to experiments with
nutrient (Pratt, 1905) and non-nutrient substances, as in the
experiment herewith recorded, we have strong evidence for
believing the fluid contents of the canal system to be composed
mainly of sea-water, in which nutrient matter may be dissolved
or suspended in minute particles, and a mucoid secretion (fig.
9) in which foreign particles, harmful or useless to the colony,
may become embedded and carried to the exterior.

II.

A freshly captured specimen of Pen na tula rubra was
placed, as in the former experiment, in a tank of running,
filtered sea-water about 5 p.m., April 14th. By means of a

454

EDITH M. MUSGEAVE.

niiuate incision in the dorsal snrface of the middle of the
rachis, an injection of finely powdered particles of carmine in
suspension in sea-water was made, with the aid of a small
syringe, into the dorsal canal. The injected fluid passed
immediately to the base of the stalk, its presence being re-
vealed by the transparency and tenuity of the body-wall in
this region (fig. 1). Three quarters of an hour later no
apparent change had taken place in any other portion of the
colony, but at 8.15 o’clock the following morning a thin,
viscid stream containing carmine particles was observed to
ooze from a single pore at the base of the stalk. Similar
streams, but more minute in quantity, were also observed to
proceed from four small pores also hitherto unrevealed, in the
median naked streak in the dorsal surface (fig. 5) about
25 mm. from the extreme apex.

About 10 a.m. the same day the ventral canal of the same
specimen was injected from the ventral surface in a
similar manner with a solution of methylene blue in sea-
water, when a portion of the injected fluid was immediately
discharged through the mouths of the more ventrall}^ situated
siphonozooids and autozooids, while a considerable portion
passed almost to the extreme base of the stalk and was dis-
charged through an aperture which then only became
apparent. Two basal apertures were thus revealed : one
slightly dorsal to the other extruding particles of carmine,
the second, slightly ventrally placed, discharging the solution
of methylene blue. Several hours later, however, carmine and
methylene blue were observed to emerge together from the
ventral aperture.

The two basal apertures thus revealed were afterwards
found to be direct openings of the dorsal and ventral canals to
the exterior (figs. 1 and 2). The experimental evidence of
their presence and active physiological function in living
specimens gives an affirmative answer to the controversy
which has existed since the eighteenth century (pp. 443-445)
as to whether Pennatulids have or have not a basal communi-
cation with the exterior (figs. 1 and 2).

EXPERIMENTAL OBSERVATIONS ON PENNATULTDS. 455

A short time after the two basal apertures became
apparent, minute streams of methylene blue were observed
to emerge from six additional apertures, also about the base
of the stalk, four of which were definitely arranged to form
the four corners of a square wlien viewed basally (fig. 2) and
the other two laterally placed but in close proximity to the
dorsal and ventral apertures. Particles of carmine were
observed in a state of extrusion from one only of all the six
apertures (fig. 2, L.Ap.).

Particles of carmine were also observed in a state of extru-
sion from the mouths of the autozooids and of the siphono-
zooids (fig. 7), but only extremely minute particles were seen
to be expelled from the latter ; it may have been impossible
for lai’ger particles to escape that way.

'Idle specimen under consideration differed from all othei’s
which I have had the opportunity of examining in the presence
of an aggregation of unusually larg-e siphonozooids in the
region superior to that occupied by the dorsal pores (fig. 5,
From these zooids I observed no instance during the experi-
ment of the extrusion of carmine or methylene blue. It is
possible that at the time of examination the injecting fluids
had not then penetrated to that particular region; but it was
very evident that currents were passing outwards from these
siphonozooids, for clouds of methylene blue squirted among
them were immediately dispersed in an outward direction.

On the third day the colony was fixed and preserved for
future examination in a moderately hot 7 per cent, aqueous
solution of formalin.

At intervals during the month of April this experiment was
repeated on other freshly captured specimens of Pennatula
rubra, and in three instances in specimens of the species
P. phosphorea.

The results in all cases were very similar in character.
Four dorsal poi’es in the act of extrusion of coloured liquids
and solid carmine particles were observed in all the specimens
examined of both species (fig. 5, D.P.)

The large basal apertures of the dorsal and ventral canals

456

EDITH M. MUSGRAVE.

were also present and also tlie four additional apertures form-
ing tlie four corners of a square (fig. 4, L.ap.), but in the case
of P. pliospliorea tlie two lateral apertures in close
proximity to tlie dorsal and ventral apertures in tlie species
P. rubra were not observed.

III.

Healthy freslily captured specimens of P ter oe ides
spinosum were submitted to injections similar to those
already described inPennatula, and as the results differed
somewhat in the two forms, it will be more convenient to
consider the case of tliis genus separately. As the body-
wall, and, in fact, the texture of the whole colony in Pteroeides,
is much stouter, tougher, and more muscular than in Penna-
tnla, some considerable time elapsed before a carmine injec-
tion of the dorsal canal about the middle of the rachis caused
any appreciable difference in the appearance of the colony;
but after an interval of more than twenty-four hours particles
of carmine were extruded from four minute dorsal pores near
to the apex of the rachis, which, as in Pennatula, only
became discernible after injection. Carmine was also ex-
truded in the form of minute particles from several pores
about the base of the stalk (fig. 3) ; of these, one larger than
the others, and almost medianly situated, had tumid lips, and
proved later to be the dorsal aperture of the large dorsal
canal (fig. 3a, B.D.ap.). Four slightly smaller apertures were
also actively engaged in the extrusion of carmine.

The day following the lower end of the stalk of the same
specimen (hereafter known as specimen No. 1), was placed in
a solution of methylene blue in sea-water, care being exercised
that the remaining portion of the colony, while submerged in
clear sea-water, should not be exposed to any material extent
to the influence of the coloured fluid. This was accomplished
by placing the stalk into a deep, narrow-necked vessel con-
taining the methylene blue solution, and carefully introducing

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 457

tlie whole into a tank of sea-water, so as to entail as little
diffusion of liquids as possible.

After an interval of two hours the colony was examined in
fresh sea- water, when it was found that the coloured fluid had
been absorbed by a single pore only. This proved to be the
basal ventral aperture of the large ventral canal (fig. 3a,
B.V.Ap.). Carmine particles were still observed in the act
of extrusion in considei’able quantity from the basal, dorsal,
and other apertures.

This experiment indicates the simultaneous presence of
an inhalent current into the ventral canal, and several
“exhalent” currents, the principal one being from the large
dorsal canal to the exterior (fig. 3).

It is by no means proved that such conditions always
prevail. It is very probable that under certain conditions the
currents may be entirely inhalent or entirely exhalent; the
direction of the current is doubtless dependent upon the
individual requirements of the colony and responsive to
stimuli from within. This matter is further discussed on pp.
470 and 471 .

The stalk of a second specimen of Pterooides spinosum
was placed in a narrow-necked vessel as before, but the
vessel contained in this instance finely powdered particles of
carmine. After several hours had elapsed the stalk of the
colony was opened and examined, but no trace of carmine
was observed in any of the canals. 'I’lie currents during the
experiment may have been entirely exhalent, but there seems
sotne evidence for the supposition that the colony has the
power to some extent of filtering the sea-water as it enters
the canals basally. AYe have reason to believe that such
is not always the case, for in other Pennatulids particles of
earthy foreign matter are frequently observed in the large
canals near the base of the stalk (cf. Pennatula rubra,
as indicated in Plate 26, fig. 1).

On the third day specimen No. 1, which still remained
apparently healthy and responsive to stimulation, was injected
with a solution of methylene blue from the ventral canal about

458

EDITH M. MUSGRAYE.

the middle of the rachis. As in the case of Pennatula
the coloured solution was immediately ejected in continuous
streams from the autozooids and siphonozooids, indicating a
very brisk circulation of fluid within the colony, the direction
of the currents being mainly exhalent.

Within a very few minutes the methylene blue streamed
from several basal pores which then became evident. These
pores had taken no part previously in the expulsion of the solid
particles of carmine. Several of the pores are indicated in fig. 3.

In all cases the injection of a coloured fluid into the
dorsal canal was immediately followed by its distribution to
all parts of the colony and expulsion from autozooids and
siphonozooids, and, indeed, after a very few minutes from the
basal and dosal pores.

Upon examining a dilatation in the region of the stalk,
which appeared in a specimen of Pteroeides shortly after
an injection had been made, the coloured fluid was found in
considerable quantity in the “spongy tissue” of this
particular region of the colony, giving evidence in support
of the supposition of its distensibility, in assisting- in the
dilatation and contraction of the colony (pp. 463, 464).

These facts afford an interesting illustration of the extreme
rapidity and comprehensiveness of the circulation of fluids
within the colony under what appear to be normal conditions.

In the case of the injection into the dorsal canal of the
rachis of powdered carmine suspended in sea-water, the distri-
bution of these solid but minute particles proceeded much
more slowly. In the case of Pennatula the injecting fluid
was transferred immediately to the base of the stalk, but the
subsequent ejection of carmine particles did not take place
from autozooids or siphonozooids until several hours had
elapsed, and after a still further interval from the basal
apertures and dorsal pores.

In Pteroeides the interval between the injection and
extrusion of carmine extended over twenty-four hours. Its
ingestion in minute quantities and subsequent protrusion
from the cells lining the canals may to a great extent be

EXPERIMEN'L’AL OBSERVATIONS ON PENNATULTDS. 459

responsible for the delayed progression. The metabolic
influence of the contents of the circulating fluids upon the
tissues bathed by them is not without interest to the physio-
logist.

DuriTig fixing operations the contour of the colony and
especially that of the stalk usuall}^ changed somewhat.

This was particularly noticeable in the case of Pteroeides,
in which the fixative re-agent penetrated the thick and tough
body-walls very slowly, allowing the internal axis to become
considerably shortened by muscular contractions, and bring-
ing about a slight invagination of the extreme base of the
stalk. The normal basal apertures were thus completely
obliterated, but the slight invagination of the body-wall gave
the appearance of a single aperture, which is so usual in pre-
served specimens of Pennatulids that it has been frequently
described as a month opening by early writers (fig. 3b),
(Linnajus, Della Chiaje, etc.), and as frequently denied by later
ones (Marshall, Pennatnla; Hubrecht, Echiuoptilum).

In the case of Pennatnla it was possible to fix and pre-
serve specimens in which the two apertures of the dorsal and
ventral canals were discernible, but in Pteroeides and other
forms all the basal and dorsal pores were obliterated during
the operations of fixing and preserving.

The peristaltic dilatations and contractions described by
Bohadsch and Kblliker were observed in the case of Pteroe-
ides and Penjiatula, but in all cases the movements were
confined to the stalk and rachis, and were somewhat lethargic
even in freshly captured and apparently healthy specimens,
in some instances being so slight as to be barely perceptible.

3’here can be no doubt that specimens examined in captivity
suffer considerably in this respect from their change of
environment, but a study of the internal anatomy of the stalk
aTid rachis of Pteroeides, Anthoptilum, Virgularia, and
Pennatnla furnishes indisputable evidence at least of the
Tnuscular activity of the axis as a boring organ, and suggests
also the probability of its function as a propelling organ in
these genera.

VOL. 54, PART 3. NICW SERIES.

33

460

EDITH M. MUSGEAYE.

The Musculature of the Stalk and Eachis.

The musculature of the “body-wall” of Peunatulids has
been carefully worked out by Kolliker (1872) and the
brothers Marsliall (1882). For descriptive accounts, there-
fore, the reader is referred to these authorities.

In Pteroeides the body-wall is generally dense, opaque,
and of tough, fibrous texture. In the specimens of this genus
the musculature is correspondingly more sti'ongly developed
than in those of Virg’ularia, Anthoptilum and Penna-
tula. In the species “p ho sphere a” of the last-named
genus an extreme condition in the opposite direction is
attained, for in this form the body-wall is very feebly
developed, and the stalk is characterised by its delicacy,
fragility, and transparency. The musculature is, in accord-
ance, composed of less numerous and more delicate fibres.

In all cases the body-wall of the stalk contains two series
of muscles : an “outer layer of longitudinal muscles,”
and an inner layer of transverse and frequently
circular muscles.

The “ longitudinal muscles ” lie immediately below the
dermis, and are composed of comparatively long muscle-
fibres whose direction is parallel Avith the longitudinal axis
of the colony. These muscles form a continuous layer in the
stalk, and pass upAvards into the dorsal and Amntral portions
of the rachis, but are discontinued laterally, the muscula-
ture of the sides of the rachis, and particularly at the insertion
of the leaves and pinnse, being so feebh’^ developed as to
serve only for the protraction and retraction of the zooids,
and can take no part Avhatsoever in the movements of the
leaves. We have absolutely no evidence for the statements
made by Ellis, Bohadsch, and others, of the muscular sAvim-
ming movements of the leaves or “ fins,” as they Avere
formerly termed.

The longitudinal muscles of the body-Avall assist in the
support of the colony by strengthening the tissues of the

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 461

body-wall, aud are also actively concerned in the extension
and contraction of the colony in a vertical direction, and
thereby assist in directing the circulating currents within
the canals.

The “transverse muscles” of the body -wall are more
deeply seated than the longitudinal muscles, and usually lie
on the inner side of the spongy tissue. In the stalk their
general direction is usually at right angles to the longitudinal
axis of the colony, and in this region the muscles are usually
arranged in more or less complete rings. In the rachis the
fibres are less definitely arranged and are also less numerous.
These muscles materially assist in the support of the colony
by strengtliening the central axis, but their main function is
apparently that of instituting' and maintaining, by peristaltic
action, a series of dilatations and contractions of the stalk
and to a less degree of the rachis, thereby assisting in con-
trolling the quantity of fluids contained in the CfTnals and also
in directinof the circulating currents.

Marshall believed the cii'culation of fluids within the colony
to be largely dependent upon the muscular system of the
body-wall. He wrote of Pennatula phosphorea: “The

well developed muscular system (i. e. of the body-wall) may
be supposed to have for its main function the maintaining, b}'
compression of portions of the spongy mesh-work, of currents
from one part of the pen to another.” As this system of
muscles appears to be more feebly developed in this than in
any of the other forms under discussion, Marshall’s statement
would apply with greater force to Pennatula rubra,
A n t h o p t i 1 i u m , V i r g u 1 a r i a, and with special emphasis to
Pterceides. We have, however, evidence for believing
that, with the exception of the “axis-less” form Eenilla
(p. 402), the musculature of the body-wall is not the sole
force in inducing, maintaining, and couti'olling the circulat-
ing currents.

Wilson observed certain creeping movements of living
specimens of Renilla — a Peunatulid without a central cal-
careous axis — which he atti'ibuted to the movement of fluids

462

EDITH M. JIUSGEAVE.

within the “ gastric cavity.” ^ Of this interesting Pennatulid
he writes : “The anterior part of the body being well dis-
tended, an active peristaltic contraction of the circular
muscles takes place, and the fluid is thus forced backwards
into the posterior region (peduncle). The latter consequently
becomes elongated, somewhat as the ambulacral ‘ foot ’ of an
echinoderm is protruded, and the body is pushed forward a
short distance. The circular muscles then relax and the longi-
tudinal ones contract in such a manner as to pull the posterior
region forward towards the anterior part which adheres to
the bottom. By the constant repetition of this process the
whole organism moves slowly forwards. The ci*eeping’ move-
ments are very irregular since the action of the muscles is
not uniform.”

In large and robust Pennatulids having a central, more or
less rigid, calcareous axis, certain modifications of structure
have arisen which apparently play an important part in
inducing and maintaining a comprehensive circulation of
fluids throughout the colony.

In all the genera examined the musculature of the body-
wall becomes attenuated towards the base of the stalk (figs.
12, 13, 14), its place being occupied by the spongy distensible
tissue which is so abundantly provided with apertures into
the large canals, and also with a definite number of apertures
(which appears to be constant on each species) to the exterior
(figs. 1, 2, 3, 4). In living specimens this region is often
considerably dilated by the absorption of fluids by the canals
of the spongy tissue, so that the stalk terminates basally in a
small distended bulb, which, if buried in the mud of the sea
bottom, may materially assist in supporting the colony in a
vertical position.

The distension may also represent the initial stage of the
peristaltic movements of the stalk and rachis commented upon
elsewhere (p. 473).

* By the teim “ gastric cavity ” Wilson, no doubt, means the spaces
occupied by the large central canals of the stalk and rachis.

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 463

The Spongy Tissue.

The spongy tissue varies considerably in consistency
among the members of the Pennatulids examined.

In all the specimens of Pteroeides it is extremely dense
and tough; in Pennatula phosphorea it is slightly trans-
parent and delicate in texture^ while specimens of Pennatula
rubra, Authoptilum, and Virgularia represent an inter-
mediate condition in these and other respects. In all cases
the spongy tissue appears upon microscopic examination to be
made up of a more or less homogeneous deeply staining
matrix, traversed in all directions by numerous canals
having epithelial linings. They communicate by means of
numerous apertures — usually ari’anged in longitudinal rows
(figs. 13, 14, np.), with the large dorsal and ventral canals of
the stalk and rachis, and at the base with the'exterior by
means of a definite number of pores which appear to be
constant for any given species (figs. 1, 2, 3, 4).

The “ spongy tissue ” is capable of becoming rigid and
turgid by the absorption of li(piids by its canals. On occasion
these may be filled to distension, or the reverse, by the com-
bined peristaltic action of the transverse and the longitudinal
muscles of the body- wall.

The “ epithelium ” lining the small canals of the “ spongy
tissue ” is somewhat different from that lining the large
canals of the stalk and rachis in that its ceils are more cubical
in shape. This tissue is apparently characterised by an
absence of the deeply staining mucus, the vacuolated, and the
food-ingesting cells which form the main portion of the
epithelium of the large canals.

Kolliker (1870, p. 10) regarded the small canals of the
spongy tissue as “ nutritive canals.” We have experimental
evidence of the possession of a nutritive function on the part
of the “ epithelium ” lining the hu’ge canals, but it is extremely
])robable that the small canals of the “ spongy tissue ” are
developed primarily for distension purposes.

464

EDITH M. HUSGEAVE.

Structural Modifications in the Stalk.

Several extremely large and robust specimens of Autbo-
ptilum grandiflorum present a peculiar modification of
the upper portion of the stalk in the form of a curiously
folded and thickened, ring-like zone (fig. 10, T.Z.). This was
found upon internal investigation to be caused by an extra-
ordinary growth of the spongy tissue in this region (fig. 11,
L.M.P.) in the form of numerous laminate vertical processes
with free edges directed inwards, and lined internally by
a moderately well-defined musculature comparable to the
transverse musculature of the body-wall in other forms, but
in this instance produced into the folds and convolutions
caused by the remarkable development of the spongy tissue.

The laininm are traversed by numerous canals, Avhich
ramify within their substance and communicate by numerous
apertui'es with the interlaminate interstices of the body
cavity.

It was unfortunate that the state of preservation of the
specimens would not permit a detailed investigation of the
histology of the lamime, but a microscopic study of the body-
wall in the lower region of the thickened zone proved of
great interest. Its outer surface was observed to be studded
with numerous minute papillae, which proved upon examina-
tion to be siphon ozooids (fig. 10, si.) — extremely small, but
of the normal Peniiatulid type, with mouth apertures and the
usual number of mesenteries. Their coelentera communicate
as in other forms with the large central canals by means of
the connecting canals of the spongy tissue.

The curiously folded epithelium of the outer body-wall, and
also that of the siphonozooids, contained numerous deeply
staining gland cells from which a mucus-like secretion was
seen exuding in considerable quantity.

The occurrence of siphonozooids^ in this unusual region

' The presence of siphonozooids in the stalk of Pennatulids is not
quite unique, for Hickson (1907, p. 1.3) has also observed them in the
stalk of “Unihellula carpenter!. ”

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 465

is of great interest, for it confirms the supposition that these
modifications of the stalk in this extremely large Pennatulid
have arisen for hydrostatic purposes. The curiously folded,
appearance of the tissue composing the thickened zone
suggests the possibility during life of enormous dilatations
of the stalk in this region, while the presence of the
musculature on the inner surface of the laminae gives evidence
of its function as an extremely powerful pumping apparatus.
By its operations currents could be driven upwards with
considerable force into the rachis and. thence distributed
throughout the colony.

In the extremely lai’ge and robust species of Penn at u la —
P. naresi and P. borealis, a slight thickening and dis-
tension of the body-wall was observed in the stalk, in the
region corresponding to the folded and thickened zone in
Anthoptilum (fig. 10). This was found, to be ‘due to a
slightly increased development of the “spongy tissue” and
“transverse musculature” of the body-wall in this region.

It is evident that in the case of these enormous species
greater force wouhl be required to induce and jnaintain the
vertical currents from the stalk to the rachis than would be
the case in more delicate and smaller species of Penn a tula,
e. g. P. rubra and P. phosphorea. This modification has
probably arisen in the species “ naresi ” and “borealis”
in response to the increased, demands of the colony in this
respect. In the presence of the slightly thickened zone in
the stalk these species represent an intermediate condition
between the comparatively small forms P. rubra and P.
phosphorea and the more magnificent Anthoptilum
g r a n d i fl o r u m .

The Sphincter Muscle.

In the genus Pteroeides the modification has proceeded
still further. As in the forms just considered, the upper
portion of the stalk in all species presents a pronounced,
distension, due in this instance to the development of an

4(36

EDITH M. MUSGRAYE,

internal transverse musculature, which assumes the form of an
extremely powerful “sphincter muscle” (fig. 14, Sph.^I.),
whose dense and numerous fibres are attached internally to
the integument of the axis and externally to the body-wall,
where a slightly increased development of the musculature
has arisen in that region to support this extraordinary
development of muscular tissue.

Between the attachment of the muscular fibres to the body-
Avall are the numerous apertures {Ap.) of the canals of the
spongy tissue.

The chief function of the “Sphincter muscles” is
obviously that of dilating and contracting the diameter of
the stalk in the region immediatel}^ inferior to the rachis, and
thereby opening and closing the lumen of the canals. It
may therefore be regarded ‘ as a very important factor in
instituting and maintaining the circulation of fluids within
the colony. This matter is further discussed on p. 473.

It apparently also serves as a double fulcrum in supporting
the axis in a vertical position (p. 468), and assists also in the
support of the “oblique musculature” of the stalk, with
which it doubtless acts in conjunction (fig. 14), p. 470.

4'he Axis.

The axis of Pennatulids is usually mainly calcareous; but
in Pteroeides it is not wholly so, but contains in addition a
considerable proportion of dense,hard, horn-like matter, which,
owing to its insolubility in acid and other solutions, renders
the cutting of satisfactory sections an extremely difficult and
patience-exhausting task. In all cases the axis is enveloped
in a sheath-like integument (figs. 11, 12, 13), delicate and
membranous in smaller forms (e. g. Pennatula phos-
phor e a), but usually extremely tough and of considerable
density in robust types (Anthoptilum and Pteroeides).

The axis varies according to the magnitude and density of
the colony in length, breadth and thickness. In Pennatula
phosphorea it is remarkable for its exceeding tenuity. In

EXPERIMENTAL OBSERVATIONS ON PENNATULIOS. 467

this species it is usually thickest at junction of the stalk and
rachis, where it is quadrangular in cross section, and tapers
gradually above and below, becoming* cylindrical as it
proceeds upwards and downwards. It usually extends almost
to the base of the stalk, but terminates in the rachis some
little distance below the apex of the colony. It is rigid in the
middle but extremely flexible at each end ; and in the con-
tracted condition of preserved specimens the basal end is
usually bent upwards to form a hook as in the drawing of
Pennatula rubra (fig*. 13). The apical portion of the axis
is also usually hooked when the colony is contracted, but to
a less degree.

In the specimens of Virgularia the axis in the stalk
becomes extremely attenuated towards the base, tapering
gradually to a fine needle-like point, as in fig. 12. In this
specimen, however, the axis does not extend to the extreme
base of the stalk, as, according to the accounts given by
i\Iarshall and others, is usually the case in this genus.
The unusual length of the stalk, its extreme tenuity and
flexibility of the lower portion of the axis, may account to a
considerable degree for the extraordinary powers of con-
tractility for which this genus is remarkable.

The axis of Pennatula rubra is larger and stronger than
that of the more delicate species P. phospliorea, and the
investing integument is correspondingly stronger and thicker.

In Pteroeides and other robust forms the axis is corre-
spondingly stout and strong (figs. 11 and 13), and is less
pointed above and below. In a state of complete extension
the axis is continued from the base to the extreme tip of the
colony. It is rigid throughout the greater portion of its
length, but in a state of contraction it is shortened by the
two flexible ends becoming hooked and slightly twisted
spirally (figs. 14 and 15), the hook being more pronounced
at the basal end. The spiral twist of the two ends of the axis
was also observed in a dissection of Pennatula phos-
phor e a .

408

EDITH M. MUSGEAYE.

The Musculature of the Axis.

In all Pennatnlids possessing a calcareous axis, with
apparently the exception of Virgularia,^ in which the apical
portion of the rachis is generally missing in captured speci-
mens, the axis is supported and its movements manipulated
by two series of muscles :

The apical musculature in the rachis, which supports
the upper and apical portions of the axis, and the oblique
musculature in the stalk, which supports the lower and
basal portions.

The concerted action of the two series of muscles would
extend and contract the axis superiorly and interiorly. Their
probable function is discussed later.

In Ptero3ides the axis is further supported by the powerful
“sphincter muscle’^ of the stalk (fig. 14, S2^h.M.) and by two
strong muscle-bands to which are attached certain fibres of
the inferior portion of the sphincter muscle, and numerous
strong muscle-fibres of the superior portion of the oblique
musculature (fig. 14, M.B.).

The apical musculature of the axis is most powerfully
developed in Pteroeides (fig. 15), in which the whole of the
apical portion of the axis is enveloped in a muscular sheath,
from which jiroceed numerous stout and tough fibres which
run obliquely downwards and are attached to the tissues
lining the bod}'-wall. In the retracted condition they are
lightly twisted in a spiral (fig. 15). This spiral twist was
also indicated, but to a less degree in a dissection of Penna-
tula phosphorea.

The chief function of this musculature is obviously that of

’ This curious and apparently general mutilation of specimens of
Virgularia, so frequently commented upon by writers, is believed by
J ungersen to be due to natural causes, and not, as was generally believed,
to accident. He attributes the absence of the upper leaves and apical
portion of the rachis to a gradual dying off and atrophy of the older
apical portion of the colony.

EXPERIMENTAL ORSERVATIONS ON PENNATULIDS. 469

supporting, extending and contracting the flexible apical
portion of the axis. By the co-operation of these muscles
with the longitudinal muscles of the body-wall the length of
rachis would be materially increased or diminished.

The somewhat oblique insertion of the apical muscles,
coupled with the action of the transverse muscles of the body-
wall, although less well developed in this region than in the
stalk, would nevertheless induce a slight muscular dilatation
and contraction of the central canals of the rachis.

The spiral twisting of the apical portion of the axis and its
ensheathing musculatiu'e, so noticeable in Pteroeides (fig.
15) in a state of contraction, may possibly serve the purpose
of a spring, which when released by the protraction of the
muscles would tend to propel the colony in an upward
vertical direction or vice versa.

The oblique musculature is extremely well developed
in Pteroeides and other large and robust Pennatulids (fig.
14, Oh.M.).

It consists of a series of “ obliquely set muscles-fibres ”
whose direction is mainly longitudinal. T’hey are attached
internally to the integument of the axis in such a manner
that its basal portion is completely enclosed by a muscular
sheath (figs. 11, 12, 13, 14), and are attached externally to the
tissues lining the body-wall.

Many of these muscles-fibres are embedded throughout the
greater jjortion of their course in the tissues of the vertical
se])turn. In Pteroeides the fibres are extremely stout and
strong (fig. 14, M.A.A.). Those of the upper portion of the
musculature are attached superiorly to the extremely stout,
obliquely set “muscle bands” (fig. 14, M.B.) situated
immediately below the sphincter muscle, with which it is
also connected by obliquely set fibres. The muscle-bands are
attached internally to the integument of the axis and to the
tissues lining the body-wall ; the plane of their direction is
slightly inclined to the horizontal.

In the presence of the powerful “sphincter muscle
and the two stout obliquely set muscle-bands” sup-

470

EDITH M. MUSGRAVE.

porting the oblique musculature of the stalk Pteroeides
appears to be unique.

The “oblique musculature” of the stalk is present
in all the forms examined, but its development varies con-
siderably among the members of the family. It is more
powerfully developed in Pteroeides than in any other genus
(fig. 14, and apparently most feebly developed in Pennatula
phosphorea. The specimens of Anthoptilum grandi-
florum, Virgularia juncea, and Pennatula rubra
(figs. 11, 12, 13) in which the oblique musculature is very
similar, iudicate an intermediate condition between these two
extreme forms.

The “sphincter muscle,” so prouounced a feature in
the stalk of Pteroeides and the well-marked “muscle-bands”
(fig. 14, and M.B.) are absent in the other forms, but

ill Virgularia the oblique muscle-fibres gradually assume
a horizontal inclination as they ascend the stalk (hg. 12,
M.A.A.). While the chief function of the oblique musculature
of the stalk appears to be that of supporting and controlling
the movements of the inferior and basal portion of the axis,
its protraction and retraction would apparently have a
manifold purpose.

With the co-operated action of the longitudinal muscles of
the body-wall, the protraction of the oblique muscles would
straighten out the hooked ends until the axis assumed a
vertical position, and the length of the stalk would thereby
be increased.

The oblique action of these muscles, simultaneously assisted
by the ringed transverse musculature of the body-wall, would
also increase the diameter of the stalk.

The extension of the stalk in length and diameter thus
accomplished would be accompanied by extensions in the
same direction of the four large central canals, whose fluid
capacity would thereby be considerably increased. To main-
tain the internal equilibrium of the colony an inhalent current
would therefore be induced through the basal pores of the
stalk so as to fill to distension the large central canals of the

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 471

stalk4 Through the iunumerable apertures in the walls of
the canals curz-ents could be directed into the distensible
spongy tissue (figs. 13, 14, Ap.). By this means the lower
portion of the stalk would become considerably dilated.

The concerted retraction of these znuscles would be accom-
panied by a simultaneous shortening of the stalk, and the
hooking up of the axis in its muscular sheath in the doi’sal
canal would give an upward impetus with a piston-like move-
ment to the fluids contained in it; simultaneously the i-etrac-
tion of the oblique muscles embedded in the vei’tical septum
would shorten the ventral and latei’al canals, which would
increase the basal impetus ah-eady mentioned of the fluid in
an upward vei-tical direction. This upward cui’rent would be
maintained by the peristaltic action of the musculature of the
body-wall of the stalk and to a less degi'ee in the rachis.

In accordance with the needs of the colony the dii-ection
of the cuiment may be reversed, as in the observatioizs by
Wilson (1893) in the case of llenilla.

In an experiment on Pterceides spinosum (fig. 3) an
inhaleut cui-rent was observed to enter the basal poi’e of the
venti’al canal, while several exhzilent currents were obsei-ved
to issue simultaneously from the pores of the dorsal and
other canals in the neighboui-hood of the base of the stalk.
In this instance, however, the coloity appeared to be in a
perfectly quiescent condition, and during the expei-iment
exhibited zione of the muscular movements observed in
otlier forms.

In the case of small and comparatively delicate species of
Penn atu lids (e. g. Penn at nl a rubra and P. phosphor ea)
the concerted action of the apical musculatui-e of the I’achis,
the oblique musculatuz-e of the stalk, and the longitudinal and
'transverse musculature of the body-wall in stalk and rachis
ai’e appzii'ently sufficient to induce and maintain a complete
cii’culation of fluids thi’oughout the colony.

* In the case of Anthoptilum the canals of the stalk would be
I’apidly filled by inhalent cuzTents through the siphonozooids specially
developed in that region.

472

EDITH M. MUSGRAYE.

In extremely larg’e and robust Pennatulids such means are
apparently insufficient for that purpose. In Anthoptilum
grandiflorum (fig’s. 10 and 11) we find a modification of the
upper portion of the stalk in preserved specimens in the form
of a thickened and much-folded zone, caused by an extra-
ordinary development of the distensile spongy tissue and
transverse musculature of the body-wall. This region is
also chai’acterised by the unusual presence of numerous
siphonozooids (fig. 10, si.).

This structural modification doubtless serv'es as a powerful
pumping apparatus for the institution and maintenance of
currents tohigher altitudesand more distant partsof thecolony.

An intermediate condition between the small species of
Pennatula(P. rubra and P. phosphorea) and the magni-
ficent species Anthoptilum grandiflorum is indicated in
the comparatively large species of Penuatula (P. naresi
and P. borealis). In these forms a slight protuberance of
the body-wall occurs in the region of the thickened zone,
and in this instance also is due to an increased growth, but
to a less degree than in Anthoptilum of the distensile
spong-y tissue and transverse musculature of the body-wall.

In Pteroeides structural modifications of the stalk, to serve
the same purpose but undoubtedly with greater efficiency, has
proceeded still further. This genus is characterised by the
development in the stalk of an extremely powerful sphincter
muscle (fig. 14), whose pmnp-like actions, co-operated with
those of the powerfully built “ oblique musculature ” of the
stalk and the musculature of the body- wall are apparently
necessary to institute and complete the circulation of fluids
within the large, densely built, and compact colony.

We have incontrovertible evidence of the distensile function
of autozooids and siphonozooids in inducing inhalent and
exhalent currents in tlie region of the rachis, and we have
also experimental evidence of the function of the four dorsal
pores for exhalent, and probably also for inhalent, purposes
(figs. 5 and G), but from the evidence based upon a study
of the musculature in connection with the canal systems of

EXPERIMENTAL OBSERV'ATIONS OX” PENX^ATULIDS. 473

Pennatnlids it seems extremely probable that the
main impetus of the circulating circuit has its
origin in the stalk.

The Powers op Locomotion.

It is generally assumed that the usual position of a liviug-
Pennatulid is an erect one with the stalk buried in the mud
of the sea bottom. Such is undoubtedly the case with forms
inhabiting shallow water like Virgularia, but we have
reason to believe from records of observation by early writers
(Bohadsch, Ellis, etc.) and from a study of the general
anatomy of the Pennatnlids, that the stationary erect con-
dition is not necessarily alwa3’s maintained.

It is extremely probable that many deep-sea Pennatnlids
are endowed with certain controlled powers of locomotion,
and may transfer themselves from place to place under suit-
able conditions.

In Pterccides, for example, the protraction of the
“oblique musculature” of the axis would enable the stalk
with a spiral twist to bore into the sand or mud of the sea
bottom in true screw-like fashion, when the bulb-like dilata-
tion of the base would assist in supporting the colony in
a vertical position. In this erect position the basal pores
of the stalk (fig. 3) would, it is assumed, remain closed and
inactive, and the circulatoiy current would now be dependent
upon the activity of the autozooids, siphonozooids, and dorsal
pore.s, for in this position the remaining portion of the colon)"
must necessarily remain more or less quiescent, for it is
obvious that any activity on the part of the muscles would
dislodge it from its basal burrow.

The retraction of the oblique musculature with the co-
operative activit)" of the apical and sphincter muscles, com-
bined with the peristaltic action of the musculature of the
body-wall Avould not only Avithdraw the colony from its
basal support, but propel it Avith such considerable force
through the Avater as. to accomplish a swimming excursion
of no small maguitude. These muscular movements Avhich

474

EDITH M. MUSGEAYE.

liave tlieir origin in the stalk and rachis have doubtless
been observed by early writers, and have given foundation
for the belief that Pennatulids have been observed “to swim
freely by means of their 'fins,’” although the “fins” or
“ leaves,” from their very constitution cannot possibly
take any part in the muscular activity and propulsion of the
colony.

This publication is the outcome of certain experiments
attempted on living Pennatulids in the Zoological Laboratories
of the IMarine Biological Station at Naples during the month
of April, 1905, during which period I occupied the British
Association table in that institution. I must express my in-
debtedness to the committee of the British Association for
the grant of the table.

The greater portion of the research in connection with this
paper has been accomplished in the Zoological Laboratories of
the Victoria University of Manchester. I must express my
very cordial thanks to Professor Hickson for much valuable
information on the snbject of Pennatulids and assistance in
my work.

Literature.

1741. Rumph. — ‘ D’Ainhoinsche Rareit kamer.,’ Amsterdam.

1758. LimiEeus.— ‘ Systema Naturae,’ lOtli edition.

17dl. Bohadsch. J. B. — ‘ De quibusdam animalibus marinis, etc.’

1764. Ellis. — “ An Account of the Sea Pen, etc.," ‘ Phil. Trans.,’ vol. liii,
pp. 419-428, pis. 19-21.

1786. and Solander. — ‘Nat. Hist. Zoophytes,’ 1786.

1788. Midler, O. P. — “ Zoologia Danica Sen Animal, Dan et Norveg,
etc.,’’ ‘ Descriptiones et Historia.’

1827. Chiaje. Stefano della. — ‘ Animali senza vertebrae del Regno si
Napoli,’ vol. iii.

1834. Ehrenbei’g, C. — “ Die corallenthiere des Rothen Meeres,” ‘ Abhand.
der Akad. Wiss.,’ Berlin.

1845. Darwin, C. — ‘ Naturalist’s Voyage Round the Woild.’

1847. Johnston. — ‘ British Zoophytes.’

EXPERIMENTAL OBSERVATIONS ON PENNATULIDS. 475

1848. Dalyell. J. G. — ‘ Rare and Renaarkable Animals of Scotland.’

1885. Hnbrecht. A. A.— “ On a NewPennatulid from the Japanese Sea,”
‘Proc. Zool. Soc., London,’ 1885, p. 512.

1869. Richiardi, S. — “ Monografia della Pamiglia dei Pennatularii,”
‘ Archiv per la Zoologia I’Anatomia e le Fisiologia,’ ser. 2. vol. i.

1870. Gray, J. E. — ‘ Catalogue of Sea Pens or Penuatiilai-iidse in the
Collection of the British Museum.’

1872. Kolliker, A — “ Anatomisch-systematische,” ‘ Beschreibung der
Alcyonai'ien, Frankfoi-t — Pennatuliden.’

1875. Schultze, F. E. — “ Coelenterata ” in ‘Jahresber. der Kommission
zur Wissench. Untersuchimg deutscher Meere. f.d.j.,’ 1872-3.

1880. Kolliker, A. — “Pennatulida,” ‘ Challenger Reports, Zool.,’ vol. i,
pt. ii.

1882. Marshall. A. M., and W. P. — ‘ Report on the Oban Pennatulida.’

1883. Wilson, E. B. — "The Development of Renilla,” ‘Phil. Trans.
Roy. Soc.,’ 1883.

1887. Marshall, A. M. — *• Report on the Pennatulida dredged by
H.M.S. ‘ Triton,’ ” ‘ Trans. Roy. Soc., Edinburgh,’ vol. xxxii,
pp. 119-152.

1887. and Fowler, G. H. — “Report on the Pennatulida dredged

by H.M.S. ‘ Porcupine,’ ” ‘ Trans. Roy. Soc.. Edinburgh,’ 1887.

1888. Jungersen, H. F. E. — ‘ Uber Ban und Entwicklung der Kolonie
von Pennatula j) h osphorea.’

1890. Bell, J. F. — " Remarks on the Mode of Life of the Penuatulids,”
‘ Proc. Zool. Soc., London,’ p. 462.

1891. V. Koch, G. — "Terminal polyj) xmd-zooid bei Pennatula und
Pterceides,” “ Moii^h. Jahrb.,’ Bd. U, 1891.

1900. Bourne, G. — “ Anthozoa.” ‘ Treatise of Zoology,’ Lankester.

1901. Bujor, P. — " Sur I’organisation de la A^’eretille,” ‘Arch de Zool.
Exper.,’ No 4, 1901.

1902. Kiikenthal, W. — “ Eine neue Familie der Pennatuliden,” ‘Zool.
Anzeig.,’ Bd. 25.

1904. Jungersen, H. F. E. — “ Pennatulida.” ‘ Danish Ingolf Expedn.’

1905. Pratt, E. M.— ‘‘The Digestive Organs of the Alcyonaria,”

‘ Quart. Jouni. Micr. Sci.,’ vol. 49, pt. 2.

1906. Gravier, C. — “ Un type Nouveaii de Virgulaire,” ‘Bull. du. Mus.
D’Hist. Nat.’

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1907. “ Ccelentera — Alcyonaria,” ‘ National Antarctic Expedi-

tion, Natural History,’ vol. iii.

VOL. 54, PART 3. NEW SERIES.

34

476

KDITH M. MUSOUAVE.

EXPLANATION OF PLATES 26 and 27,

Illustrating Edith M. Musgrave’s paper on “ Experimental
Observations on the Organs of Circulation and the
Power of Locomotion in Pennatulids.”

Abreviations.

Ap. Apertm-e. Ax. Axis. Ax. 8. Axial sheath. B.D.Ap. Basal
dorsal aperture. B.V.Ap. Basal ventral aperture. B.W. Body- wall.
C. and Ca. Canals. D. Dennis. D.Ap. Apertui’e leading from dorsal
canal to exterior. D.C. Dorsal canal. D.M.S. Delicate musciilar
sheath encircling the stalk near the base. D.P. Dorsal pores. Ep.
Epidermis. Ex.C. Exhalent current. D.S.T. Vacuolated tissue with
deeply staining cells. F.Ap. False aperture often seen in preseiwed
specimens. G.Ax. Growing tip of axis. Iii.C. Inhalent cun-ent.
L. Polyp bearing leaf. L.Ap. Lateral aperture near the base of the
stalk. L.C. Lateral canal. L.M. Longitudinal muscles. L.M.P.
Laminate muscular processes. M. Brownish earth-like matter on dorso-
ventral septum. M.A.A. Muscular apparatus controlling movements
of axis. M.B. Muscle-bands. Mg. Mesogloea. M.S.Ax. Muscular
sheath encircling the axis near the apex of the I'achis. Muc.C. Mucus
cells. N.D.S. Naked dorsal streak. Oh.M. Oblique musculature
supported by dorso-ventral sejAum. if. Rachis. S.C.C. Small canals
connecting dorsal and ventral canals. Si. Siphonozooids. Sp>- Spicule.
Sph.M. Sphincter muscle regulating dilatation and contraction of
stalk. Sp.T. Spongy tissue. St. Stalk. T.Ax. Termination of axis.
Th.B.W. Ring-like thickening of the body-wall in the region of the
insertion of the oblique muscles. T.Z. Thickened zone. T.Zo. Terminal
zooid. V.Ap. Aperture from ventral canal to exterior. V.C. Ventral
canal. V.L. Young leaves. F.S. Vertical septxmi. Y.St. Young siphono-
zooids.

PLATE 26.

Figs. 1-9

Fig. 1. — Pennatula luibra; x 8. Basal portion of the stalk with
cut edge as transverse section. While living the specimen had been
injected in the middle of the dorsal region of the rachis with finely
powdered particles of carmine in suspension in sea-water and later
by a second injection of methylene blue in solution in sea-water.
The deposition of carmine particles on the walls and in the lumen of
the canals is shown. The distribution of the methylene blue solution in

EXPEUIMEKTAL OBSERVATIONS ON PENNATULIDS. 477

this portion of the colony is also seen and also its extrusion from the
basal apertures (B.D.Ap. and B.V.Ap.) of the dorsal and ventral
canals. The brownish mass (M.) of earth-like matter, which appears to be
deposited on the vertical septum, may represent an aggregation of
foi’eign matter sucked in by the sea pen, or waste matter on the point
of extrusion. In this genus the ventral canal extends to the base of
the stalk. The four canals of the stalk are shown in ci’oss-section. The
dorsal canal (D.C.) is the largest, and contains in this region the two
smaller lateral canals (L.C.), between which the basal end of the axis
(Ax.) is shown by transparency of the walls of the canals.

Fig. 2.- — Pennatula rubra. Basal vein of the termination of the
stalk after injection with carmine particles and a solution of methylene
blue as in fig. 1 ; x 7. The two large apertures (B.D.Ap. and B.V.Ap.)
of the dorsal and ventral canals are shoum. Six other apertures are
also indicated, whicli only became perceptible after injection. Of these,
two are in close i^roximity to the large dorsal and ventral apertiu'es and
the othei' four definitely arranged to form the four corners of a square
(Ap.^„^^). The solid particles were exj^elled from the large apertures
of the dorsal and ventral canals, and the methylene blue solution
from the other six ; occasionally solid particles and coloured liquid
were expelled from a common pore (L.Ap.i.).

Fig. 3. — Pterceides spinosum. («) Drawing of the basal jDortion of
the stalk from the left side of a living specimen twenty-four hours after
an injection of caimine and methylene blue, as in fig. 1 ; x 3. Numerous
apertures are indicated which before the experiment were impercejjtible
to the naked eye, from which issued streams of methylene blue solution,
indicated in the dniwing. Carmine particles are also shown in a state
of extrusion. The arrows indicate the direction of the currents. An
inhalent cuiTent into the ventral canal was revealed by a prolonged
immersion of the base of the stalk in methylene blue solution. It is
possible that the direction of the currents may be reversed or otherwise
on occasion, (h) The base of the same colony after fixing with formalin
solution. The contraction of the axial tissues has caused an invagination
of the basal portion of the stalk, giving the appearance of a single pore
at the base {F.Ap>.). This appears to be the usual condition in preserved
specimens, and the apparent presence of a single pore has given rise to
considerable difference of opinion in the past as to the presence or other-
wise of a so-called mouth in this region.

Fig. 4. — Pennatula phosphorea. Drawing of the base of the
stalk showing inhalent and exhalent apertures; X 30. The two
apertures in close proximity to the basal, dorsal and ventral apertiu’es
in the species “ rubra ” were not observed in this species.

Fig. 5. — Pennatula rubra. Drawing of the dorsal surface of the

478

EDITH M. IMTTSGRAVE,

racliis of an injected specimen to show the extrusion of carmine particles
from four dorsal pores (D.P.) ; X 2. Tliese are the apertures of small
transverse canals, which in this portion of the rachis establish com-
munications dorsally between the large dorsal canal and the exterior.
The particular specimen differed from any other which I have examined
in the presence of a group of unusually large siijhonozooids (Si.) in the
region superior to that of the dorsal pores. Elsewhere the siphonozooids
(Si.) are quite normal in character.

Fig. 6. — Pennatiila phosphorea. Drawing of the dorsal surface
of the uiiper portion of the rachis of a young colony, x 15, showing the
terminal zooid and four dorsal pores (D.P.) from which, after injection,
as in fig 1, carmine particles were extruded.

Fig. 7. — Pennatula rubra. A portion of the dorsal surface of an
injected specimen to show the extrusion of carmine particles from the
mouths of the siphonozooids twenty-four hours after the injection was
made. X 30.

Fig. 8. — Pennatula rubra. Transverse section through the stalk
near to the base ; X 15. On the left side two of the smaller apertures are
shown (Ap.) which communicate by means of the spaces in the spongy
tissue witli the ventral canal (V.C.). In the upper portion of the section
a small canal (C.) is shown in the dorso-ventral septum (D.V.S.), which
establishes communication between the lumen of the dorsal and
ventral canals. Small canals (Ca.) are also shown leading from the
large canals into the spaces of the spongy tissue (Sjj.T.). A very thorough
and complete system of communication is established by means of similar
canals throughout the colony, which is thus brought into communication
with the external sea-water. The epithelial tissue lining the canals is
much vacuolated (Fig. 0) and contains numerous deeply staining cells,
which are prol>ably mucus-secreting, and are similar in character to cells
composing the greater portion of the periphery of the base of the stalk.
Immediately below this layer of vacuolated and deeply staining tissue,
parallel with the periphery and outlining the large dorsal and ventral
canals, is an extremely delicate sheath of transverse muscular fibi'es
(T.M.F.), which becomes, however, much moi'e pronounced in the
superior regions of the stalk. The spongy tissue occupies the whole
of the space between the canals and the body-wall.

Fig. 9. — Pennatula rubra. A portion of the vacuolated and
deeply staining epithelial tissue which lines the dorsal and ventral
canals as seen in a transverse section of the stalk ; X 800. A similar
epithelial tissue covers externally the basal portion of the stalk. The
endoderm in this portion of the canal is curioiisly papillate in form,
and exti'emely vacuolated. It contains numerous deeply staining cells,
which probably have for their function the secretion of mucus, and also
send off into a supporting mesogloeal core numerous muscle-fibi’es

EXPERIMENTAL OBSERVATIONS ON PENNATULTDS. 479

(M.F.), which probably play an important part in the expansion and
contraction of the canals. Many of the endodernial cells also contain
ingested particles of the injected carmine, which have Ijeen conveyed to
this 2)ortion of the colony by means of the circulating currents. The
presence of carmine in the cells may indicate their possession of
nutritive and excretory functions. The mucous secretion from the cells,
which is no doubt responsible for the viscid character of the contents
of the canals, may be lubricating in function, and may from its hygro-
scopic nature play an important part in the dilatation and contraction
of the colony.

PLATE 27.

Figs. 10-15.

Fig. 10. — Anthoptilum grandiflorum. Drawing of the ventral
surface of the stalk to show the folded thickened zone {I’.Z.), princi^^ally
due in this instance to an extraordinary growth of the spongy (disten-
sible) tissue composing the internal body-wall, and not to the special
development of an internal sphincter muscle as in Pte roe ides (fig. 14,
Spli.M.). Both modifications of structure doubtless serve the same
function in bringing about the dilatations and contractions of the
colony. The lower portion of the thickened zone and the region imme-
diately inferior to it is studded with numerous minute siphonozooids
(Si.), which play an important part in maintaining the hydrostatic
equililn'ium of the colony. (Natural size.)

Fig. 1 1.— Anthopti him grandiflorum. Dissection of the stalk
from the ventral surface (two thirds natural size), showing the hook-like
termination of the axis induced by the contraction of tlie muscular
apparatus controlling it (M.A.A.), the terminal basal aperture of the
dorsal canal (B.D.Ap.), and the thickened zone (T.Z.) in tlie upper
portion of the stalk — composed of hydrostatic and muscular spongy
tissue, which is produced into laminate processes (L.M.P.) presenting a
free edge to the body-cavity. The outer surface of the thickened zone
was observed, on microscopical examination, to be studded witli minute
siphonozooids of the normal Pennatulid type (fig. lU). This extra-
ordinary development of the laminate processes appears to be unique
and has not been (observed in any other genus.

Fig. 12. — Virgularia juncea. Dissection of the stalk from the
dorsal surface exposing the lumen of the dorsal canal (three quarters
natural size). The stalk in this genus is characterised by its unusual
length, tenuity, brittleness and extreme powers of contractility. In
elemental constitution it is similar to Penna tula (fig. 13) differing only
from that genus in proportionate development. As in Penna tula the
muscular body-wall becomes attenuated towards the base, where it

180

EDITH ^r. MUSfHIAVE.

l)ecomes delicate and meiubranons in character. In the specimen
represented in the drawing the slender, needle-like, hut slightly flexible
axis (Ax.), supported by the axial sheath (Ax. S.), does not extend down
to the base of the stalk as is usually the case in this genus (Marshall,
p. 56). A close similarity exists between Pennatula and Virgularia
in the musculature (31. A. A.) controlling the movements of the axis.

Fig. 13. — Pennatula rubra. Dissection of the stalk from the
dorsal surface exposing the lumen of the dorsal canal ; X 2. The stalk
is shorter and of comparative greater thickness than in Virgularia
(fig. 12), but it is very similar in constitution. In Pennatula the four
large canals of the stalk are shorter and broader, and the musculature
controlling the axis less strongly developed. Pennatula appears to be
less sensitive to contact, and therefore less contractile than Virgularia.
In the drawing the vertical septum is seen to extend to the extreme base
of the stalk, so that the ventral canal is equal in length in this region to
the doi’sal canal (compare Pterceides [fig. 14], Anthoptilum [fig.
10], etc.). The transverse muscular tissue (T.3I.F.) of the body-wall in
the neighbourhood of the insertion of the musculature controlling the
axis is strongly developed to give the necessary additional support.
The thickening of muscular tissue in this region is more strongly
marked in P. naresi and P. borealis. In this respect Pennatula
approaches Pterceides.

Fig. 14. — Pterceides griseum var. longispinosum. Dissection
of the stalk from the dorsal surface exposing the lumen of the dorsal
canal ; X 1|-. The musculature of the body-wall is very pronoimced in
this genus. The thickened zone in the upper portion of the stalk is due
in this genus to an extraordinary development of a si^hincter muscle
(Sph.3I.), in which I’espect it apparently dilfers from all other genera.
The fibres of the sphincter run almost transversely, and are attached on
the inner side to the axial sheath and on the outer side to the body- wall.
This powerful muscle has probably a double function — it may assist in
the support of the axis, but its chief function seems to be that of con-
trolling the dilatations and contractions of the stalk, and therefore
materially assists in regulating the quantity of fluids within the canals.
At the base of the sphincter muscle are powerful muscle-bands
(M.B.), which are connected with the axial sheath and body-wall, and
in addition assist in supporting and controlling the extremely power-
ful muscular apparatus (31. A. A.) governing the action of the axis
(Ax.). The calcai'eous axis with its accompanying muscular sheath
is slightly twisted spirally at its apical and basal termination (fig. 15).
The whole conformation of the stalk seems to indicate that it is
specially adapted as a boring organ, working, in this instance, in a
screw-like fashion, and a’so as a pump woi'king with a slightly piston-

EXPERIMENTAL ORSERVATIONS ON PENNATULIDS. 481

like movement in the cavity of the dorsal canal when the mnscula-
ture contracts. This would give an impetus in an upward direction
to the ciuTents entering at the basal apertures. As in other genera
the musculature of the body-wall is less well developed near the
base, its ^jlace being occiipied by the spongy hydrostatic tissue,
which is abundantly supplied with canals having numerous apertm’es
into the lumen of the canals and also to the exterior (fig. 3). In living
specimens this portion of the stalk is often considerably dilated. In the
drawing numerous apertures (Ap.) are indicated, which are arranged in
two vertical rows, one on the left the other on the right of the drawing,
between the insertion of the muscular fibres of the muscular apparatus
controlling the axis, which establish communications between the lumen
of the large canals and the canal of the spongy tissue lining the body-
wall.

Fig. 1.5. — -Pteroeides caledonicum. Dissection of the upper
poi’tion of the rachis from the dorsal sitrface exposing the lumen of
the dorsal canal (D.C.) to show the musculature conti’olling the ajDical
portion of the axis, which is slightly twisted spirally when the muscles
are contracted, as indicated in the drawing; X 1^. The corresponding
twist of the basal portion of the axis in this genus is indicated in fig. 14.
The musculature and thickness of the body-wall is moie jjronounced in
this specimen than in that of the species Pt. griseum (fig. 14).

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I

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTERUS. 483

On Certain Features in the Development of the
Alimentary Canal in Lepidosiren and
Protopterus.

By

J. Oraliaiii Kerr,

Professor in the University of Glasgow.

Witli 13 Text-figures.

Contents.

PAGE

I. Introduction ......

II. Diiferentiation of the Main Regions of the Alimentai’y Canal

III. Buccal Cavity .....

IV. Lung ......

V. Pancreas ......

VI. Sunimaiy ......

483

484
494
501
513
517

I. Introduction.

The following pages contain a short description of certain
features which have seemed of special interest in the develop-
ment of the alimentary canal of Lepidosiren and Proto-
pterus. 'The technical methods used have been the same as
those used in earlier stages of my work. In particular, I have
made constant use of the method of reconstruction by means
of ground glass plates, and I have endeavoured to use both
the celloidin and the paraffin methods of embedding. As I
have already had occasion to point out more than once, it is,
in my opinion, essential in embryological investigations to
use these methods side by side. Both are liable to mislead,
both have their faults, but the faults are different in the two
VOD. 54, PART 4. NEW SERIES. 35

484

,T. GRAHAM KERR.

cases, and by a careful and critical use of the two, error can
be to a great extent eliminated.

I have, as on other occasions, to acknowledge the valuable
assistance which I have received from Mr. Maxwell, to whom
I owe the drawings illustrating this paper, and to Mr. P.
Jamieson, who has done the necessary section cutting.

II. Differentiation of the main regions of the Alimentary

Canal.

The al imeutary canal of the adult Protopterus or Lepido-
siren becomes developed out of the mass of primitive endo-
derm, characterised by its large cells and by the large size of
the yolk granules with which their protoplasm is laden. The
almost spherical mass of endoderm of the early embryo
becomes fashioned into the tubular alimentary canal of the
adult by a complicated process of what I have somewhat
loosely termed modelling, the general course of which is
illustrated by the figures in this paper. It is necessary to
state quite definitely that, in using the term modelling, I do
not for a moment mean to suggest that this process is carried
out by the active agency of the surrounding tissues upon a
plastic and passive endoderm. On the contrary, I believe it
to be essential, in all embryological work, to bear constantly
in mind that the organ is only a part of the organism ; that
any organ or piece of tissue is throughout its development in
intimate physiological connection with its surrounding tissues,
and that to consider it by itself, without reference to these
surrounding structures, is to make use of a method which is
almost certain to lead to grave error. The alimentary canal,
then, of the developing Lepidosiren or Protopterus is, during
its process of “ modelling,” by no means to be assumed to be
passive; the whole process is one of co-operative activity
between the endodermal rudiment and the mesodermal struc-
tures in relation with it.

The first stage of the modelling of the alimentary canal is
that in which a narrow anterior part — the fore-gut — becomes

ALIMENTAEY CANAL IN LEPIDOSIEEN AND PEOTOPTEEUS. 485
Text-fig. 1 a.

Text-fig. 1 b.

Text-figs. 1 a, b. — Sagittal sections illustrating the folding off of
the fore-gut. a. Protopterus, stage XXIII. b. Lepido-
siren, stage XXV. br. Brain, f.g. Fore-gut.

486

J. GRAHAM KERR.

differentiated from tlie larger hinder region (mid-gut and
hind-gut) which forms the main storehouse for the yolk.
The commencing development of the fore-gut is illustrated
by text-fig. 1 A, in which the fore-gut rudiment is beginning
to be nipped oft' from the rest of the endoderm by the de-
velopment of a chink beneath it. This chink then gradually
spreads backwards, as shown in text-fig. 1 B, and also later-
ally, and in this way the fore-gut becomes demarcated, the

Text-fig. 2.

Section through junction of fore- and mid-gut of a Lepidosiren
larva of stage XXXIV, showing the origin of the pyloric valve,
c. 9)1. g. Cavity of mid-gut. /. g. Hind end of fore-gut.

ventral part of the space alluded to becoming occupied by the
pei’icardiac cavity with its mesodermal lining. The folding
off of the fore-gut is continued backwards until, about stage
XXXII, it reaches the level of the pylorus, after which it
increases in length by its own growth, and undergoes gradual
histological differentiation until the adult condition is reached.
About stage XXXIV the active growth in length of the fore-
gut causes it to push its hinder end into the cavity of the
raid-gut, the wall of which is relatively thin at the point of
junction of fore- and mid-gut (see text-fig. 2). The
flattened spout-like projection of the hinder end of the fore-

ALIMENTARY CANAL IN LEPIDOSIEEN AND PROTOPTERUS. 487

gut into the cavity of the mid-gut persists throughout life,
and forms the characteristic “ pyloric valve.”

The main features in the topographical evolution of the
mid-gut and hind-gut will be gathered from an inspection of
the figures of the external features (see Keibehs “Nor-
mentafeln,” Heft x) together with text-figs. 3 A — d, 4 A — d,
and 5.

In correlation with the function of this part of the gut
as the storehouse of food material on which the young
animal has to subsist for a prolonged period, its cells remain
for long laden with yolk, and its developmental progress is
correspondingly retarded. The first conspicuous change
consists in the rapid elongation of the hinder part of the gut
which accompanies the rapid growth of the hinder trunk
region (see figures of external features, stages XXV — XXX).
The anterior region of the mid-gut for a considerable time
retains its spheroidal shape, and it is this which gives the
characteristic tadpole-like appearance to these stages. About
stage XXXIII in Lepidosiren, but not till about stage
XXXV in Pro top ter us, the prominent bulging of the ante-
rior part of the mid-gut disappears, and the tadpole-like
appearance is finally lost.

The general appearance of the mid- and hind-gut as seen in
a dissection of a young Lepidosiren of stage XXXII is shown
in text-figs 3 a and 4 a. The prominent rounded bulging
of the anterior end is already at this stage giving place to a
gradual tapering. Towards the hinder end a faint spiral
marking caused by a shallow groove traversing the surface
of the gut rudiment foreshadows the development of the
spiral valve.

In the mid-dorsal line a broad valley passes back for some
distance, and in this lies the rudiment of the lungs. Text-
fig. 4 A shows how the fore-gut in the I’egion of the glottis
bends abruptly towards the left side, passing into the mid-
gut far to the left of the mesial plane.

In the dissection of stage XXXV (text-figs. 3 b, 4 b and 5) it
is seen that the swollen mass of yolk in front has been greatly

488

J. GEAHAM KERR.

reduced, and this part of the gut no longer bulges con-
spicuously. The spiral valve is now indicated by a deep

Text-fig. .3 A. Text-fig. 3 b.

Text-figs. 3 a, b, c, d.^ — Dissection of young Lepidosirens of stages
XXXII, XXXV, XXXVI, and XXXVII, from the ventral side,
illustrating the development of the alimentary canal, g. b. Gall
bladder, ht. Heart. 1. a. Left auricle, li. Liver, pc. Pericardium.
r. a. Eight auricle.

“ incision,” which traverses the mid-gut right to its anterior
end, but is absent for a short distance posteriorly.

ALIMENTAEY CANAL IN LEPIDOSIREN AND PROTOPTERUS. 489

Text-fig. 3 c.

490

■T. GRAHAM KEEK.

At stage XXXVI (text-figs. 3 c and 4 c) the intestinal
rudiment forms a spirally coiled structure, the turns of the

Text-fig. 4

Text-fig. 4 b.

Text-Figs. 4 a, b, c, d. — Dissections of the mid-o-iit of young Lopi-
dosirens seen from the dorsal side. a. Stage XXXII. b. Stage
XXXV^. c. Stage XXXVI. d. Stage XXXVII. c.c. Cloacal
ciBCum. f.fl. Fore-gut. 1. Lung. li. Liver, m. n. d. Mesonephric
duct. pa. Pancreas, pa. d. Dorsal iiancreas. p?i. Pharynx,
sjj. Spleen, t. Tongue.

spiral being separated by the deep incision mentioned in the
preceding stage. In all probability this spirally coiled condi-

ALIMENTAEY CANAL IN LEPIDOSIBEN AND PEOTOPTEEUS. 491

Text-fig. 4 c.

'i

I

,1

I

I

Text-fig. 4 d.

492

J. GRAHA!^[ KERR.

tion of the intestine may be looked ou as a repetition of a
phylog’enetic stage,' in whicli the intestine had assumed a
spiral coiling owing to its relatively great length in com-
parison with the length of the splanchnocoele. As was shown
especially by Riickert ^ the spiral valve in certain Elasmo-
branchs is similarly preceded by a spirally coiled condition
of the endodermal gut rudiment, in this case definitely asso-
ciated with actual growth in its length. As, further, the
spiral valve is characteristic of all the more primitive groups
of fish-like Guathostomata, Elasmobranchs, Crossopterygians,
and Lung-fishes, not to mention Actinopterygians and ancient
groups of amphibians and reptiles, we may take it as fairly
piobable that the Gnathostomata were as a whole character-
ised during an early period of their evolution, a period ante-
cedent to the splitting up into the groups above named, by
the possession of a long spirally coiled gut.'”'

In comparing the stage under discussion (XXXVI) with
the preceding stages it will be seen that the using up of the
yolk is still taking place most actively in the anterior region.
As a consequence it is seen that the first turn of the spiral
has become greatly reduced in size, so tliat, as shown in
ventral view, it is decidedly smaller than the succeeding
turn instead of being much larger, as was the case in
stage XXXV. Now that the turns of the spiral are distinct
it may be seen that there are in all most usually nine or ten
turns.

In the last stage figured (stage XXXVII, text-figs. 3 D and
4 d) the yolk has been used up to such an extent as no longer
to influence the outward form of the intestine. The turns of
the spiral are now of approximately uniform diameter, they
are closely bound together by mesenchyme, and are enclosed

‘ Graham Kerr, ‘Phil. Trans. Roy. Soc.,’ B. cxcii, 1900, p. 325.

- ‘ Arch. f. Entwick. Mech.,’ iv. 1890, p. 298.

^ In all probability the gut has varied much in length from time to
time during the evolution of the vertehrata. changes in length being
associated with changes in the nature of the diet, e.g. from vegetable to
animal.

ALIMENTARY CANAL IN LEPIDOSIEEN AND PROTOPTERUS. 493

U. Liver, o.c. .Auditory capsule, oc. r. Occipital ril). p.ji. Pronephros. L o. Tectum opticum.

494

.T. GRAHAM KERR.

in a cylindrical slieatli of splanchnic mesoderm dotted with
scattered chromatophores. From being a spirally coiled
structure the intestine has therefore now assumed the out-
ward form of a straight cylinder, only the cavity in its
interior betraying its once coiled condition.

III. Buccal Cavity.

The Dipnoi share with various Amphibians the peculiarity
that the main part of the buccal cavity arises in place of the
anterior portion of the yolk-laden enteric rudiment derived
from the macromeres of the segmented egg, i.e. its wall is in
great part deidved from a mass of cells which would ordi-
narily be called endoderm. Those who are sticklers for the
sanctity of the germ layer theory find it difficult to accept this
statement, and would rather believe that, although the defini-
tive buccal cavity comes into existence in place of a “ primitive
endoderm ” structure, yet its lining is formed by a definite
ingrowth of ectoderm. Thus the buccal cavity of the forms
mentioned would be not merely in theoi’y, but as regards its
actual ontogenetic development a typical stomodaeum. Greil
in particular, who has investigated the development in C era-
tod us, appears to have no doubt that actual ingrowth of
cells from the outside takes place to form the lining of the
buccal cavity. Personally I see no reason to depart from
the statement which I made some years ago^ that the main
part of the buccal lining arises in situ by actual transforma-
tion of the originally yolk-laden cells. The active metabo-
lism associated with this process of transformation is hei’e as
elsewhere accompanied by a breaking up of the yolk granules
into very fine particles so that they may be more easily
assimilable, and it is this assumption of a finely yolked and
richly protoplasmic character that causes the cells to assume
an ectoderm-like appearance. “ It is,” as I put it in my
former paper, as if an influence were spreading inwards

' ‘ Quarterly Journal of Microscopical Science,’ vol. 46, p. 423.

ALIMENTARY CANAL IN LEPIDOSIEEN AND PROTOPTEEDS. 495

from the external epiblast, gradually transforming the
original ^'endoderm” yolk-laden cells into ectoderm like
itself/’ Apart from the difficulty of believing in the possi-
bilitj' of a layer of soft protoplasmic ectoderm cells growing
inwards and pushing aside compact masses of yolk-granules
without even producing any signs of mechanical disturbance
of the tissues, the study of carefully prepared celloidin
sections is, I think, sufficient to convince anyone that it is
really a process of conversion in situ which is taking place.
In such sections there is frequently visible a quite broad zone
of transition in which the richly protoplasmic '^ectoderm”
cells pass by imperceptible gradations into the typical yolk-
cells, there being no trace of the absolutely sharp boundary
which must be present were the ingrowth hypothesis cor-
rect.

To the main pai-t of the buccal cavity, which arises by
cytolysis in the midst of an originally solid mass of yolk-
cells, and the walls of which give rise to the teeth, as has
been described in Part III, there becomes added in later
stages of development the antero-lateral part of the definitive
buccal cavity, in the roof of which are situated the narial
openings. This additional part of the buccal cavity arises in
ontogeny in the same kind of way as the whole cavity does
in Polypterus, i.e. by the walling in of a space on the
lower side of the head, through the development of the upper
lip and the forward growth of the lower jaw. The mode of
development of this part of the buccal cavity is made clear
by text-fig. () A — F. In stages XXXI and XXXII of Protop-
terus (text-fig. 6 A and b) the position of the front end of
the alimentary canal is marked out in a ventral view of the
larva by a transverse line — the line of junction of the yolk-
laden enteric cells with the ectoderm. Some little distance in
front of the outer end of this line upon each side is seen a
dimple, which marks the olfactory rudiment. In a specimen
rather younger than stage XXXIV in its general features the
olfactory dimple (text-fig. 6 c) is seen to have become elon-
gated in an oblique direction, so that its long axis passes from

Text-fio. Oa. Text. -fig. 6 b. Text. -fig. 6 c.

496

J. GRAHAM KERR

Text-figs. 6 A, B, c, D, E, F. — Views of under-siirface of head in Protopterus of stages XXXI, XXXII, XXXIV — , XXXI\ ,
XXXV and XXXVI — , to sliow the relations of the olfactory organs. In e and p the floor of the buccal cavity has been
partially cut away. c. o. Cement organ, e. g. External gill. olf. Olfactory rudiment, olj. a. and o?/. p. Anterior and
posterior nares. op. Operculum, to. Tooth.

Text-fig. 6 D. Text-fig. 6 e. Text-fig. 6 f.

ALIMENTAEY CANAL IN LEPIDOSIEEN AND PEOTOPTEEUS. 497

•gi'iu 1

498

J. CiEAHAM KERR.

in front backwards and outwards. It now forms a deep cleft
leading right into the interior of the olfactory oi'gan along
nearly its whole length. (The internal cavity of the olfactory
organ is at first closed, and arises in the midst of the originally
solid rudiment, as explained in Part

The area containing the two olfactory clefts is now marked
off from the rest of the under surface of the head, behind by
a sharp fold — the first indication of the lower lip, and in
front by the much less sharply marked rudiment of the upper
lip. The area within these folds, and having an olfactory
cleft upon each side, is the rudiment of that additional antero-
lateral part of the buccal roof which becomes added on to the
posterior and larger portion derived from the solid mass of
yolk-cells.

By stage XXXIV (text-fig. 6 d) the delimitation of this
additional part of the buccal roof from the rest of the under
surface of the head has become more sharply mai’ked, the
upper lip being now more prominent, and the lower lip or
lower jaw having commenced to grow forwards to form its
floor. The olfactory cleft is more elongated. It has become
drawn out and narrowed in its middle part to a fine slit,
which connects the dilated anterior and posterior ends — the
rudiments of the anterior and posterior nares. Of these the
posterior naris is now hidden in a ventral view of the head,
owing to the forward growth of the lower jaw.

In stage XXXV it is necessary to cut away part of the
lower jaw to see the olfactory clefts. It is seen (text-fig. 6 e)
that the lips of the cleft are now in close apposition except
at their ends, and between this stage and stage XXXVI the
lips undergo complete fusion, so that the anterior and poste-
rior nares are now distinct openings (text-fig. 6 p).

Keibel has remarked:^ “The so-called upper lip of the
Dipnoi lies morphologically further outwards than does the
mouth margin in any other Vertebrates.” This character is
specially expressed in the fact that the margin of the mouth

' ‘ Quarterly Journal of Microscopical Science,’ vol. 46, p. 438.

2 ‘ Anat. Anz.,’ vol. viii, p. 487.

ALIMENTARY CANAL IN LEPIDOSIEEN AND PEOTOPTEEUS. 499

encloses anterior as well as posterior nares, and forms one
of the most distinctive features of the Dipnoi. How this
ai’rangement has come about in Phylogeny does not seem
quite certain, but most probably it has been by a backward
migration of the narial rudiment, rather than by an extension
forwai’ds of the mouth boundary. It will be noticed that the
upper lip, while very prominent laterally, can hardly be said
to exist in the region near the mesial plane. This gives a
characteristic gaping, almost cyclostomatous, appearance to
the mouth of the young Dipnoan,^ and if it be assumed that
this is a repetition of a phylogenetic condition, it is clear that
a backward migration of the narial openings into the buccal
cavity could readily have taken place. The physiological
significance of the iutrabuccal position of the narial openings
is clearly in all probability adaptive to the mud-burrowing
habits. The olfactory organ is in the living Dipnoan used,
so far as my observations go, entirely as a sense organ, its
respiratory function not yet having developed. The sense of
“smell” affords the principal means by which the living
Lepidosiren or Protopterus finds its food; a little colouring
matter, e. g. blood, in the water, shows how they actively
“sniff” about, with snout sharply bent down, in search of
food particles at the bottom of the water.

Glands and Sense Organs. — Unicellular glands and
sense buds are scattered, as already shown by Parker for
Protopterus, over the lining of mouth and pharynx as on the
outer skin, while the flask-shaped glands so characteristic of
the external ectoderm are normally absent from the buccal
cavity.

Thyroid. — The thyroid makes its appearance about stage
XXX in Lepidosiren as a solid keel-like projection from
the ventral side of the solid buccopharyngeal rudiment. The
study of sagittal sections (text-fig. 7) show that the thyroid
rudiment becomes gradually cut off from the buccopharyn-
geal mass from behind.

‘ Even in the adult Lepidosiren the month is actively suctorial, and
food is drawn into the mouth by a strong sucking action.

VOL. 54, PART 4. NEW SERIES.

36

500

J. C4RAHAM KEER

Text-fig. 7 a.

Text-fig. 7 b.

Text-fig. 7 c.

Text-fig. 7 d.

Text-figs. 7 a, b, c, d. — Sagittal sections showing origin of thyroid in
Lepidosiren. a, b, and c. Stage XXX. d. Stage XXXI. th.
Thyroid, t. Tongue.

ALTMEXTARY CANAL IN LEPIDOSIREN AND PROTOPTERUS, 501

About stage XXXI (Lepidosiren) the narrow isthmus
which still unites the thyroid to the buccal floor in front
of the tongue becomes severed, and the organ lies free as a
rounded mass of coarsely yolked cells.

Between stages XXXIV and XXXV the thyroid rudiment
becomes broken up into strands by intruding mesoderm with
blood-vessels, and a little later the strands are broken up
into typical rounded follicles with colloidal secretion in their
interior.

Tongue. — Reference to the text-figures (7, a — d) illus-
trating the development of the thyroid is sufficient to show
that the tongue of the Dipneumona is a primary tongue, like
that of Urodele Amphibians, except that in this case no gland
field develops in connection with the tongue, at least up to
stage XXXVIII. The downgrowth from the solid buccal
rudiment, from the posterior side of which the thyroid is
developed, becomes split about stage XXXI, and it is this
splitting which causes the portion of buccal floor behind it
to be bounded in front and laterally by a deep cleft so as to
form a distinct tongue.

IV. Lung.

The first rudiment of the lung is seen in text-fig. 8, A,
which is a ventral view of the pharyngeal region of a Pro-
topterus of stage XXXII.

The lung rudiment is seen to form a rounded bulging from
the pharynx in the mid-ventral line just at the level of cleft
VI. Pharynx and lung rudiment are alike solid at this
stage.

The endodermal lung rudiment grows at first ventralwards,
and slightly towards the right side (text-figs. 8 and 12, a,
pages 502,514). Meanwhile the modelling of the oesophagus
is proceeding ; it becomes more elongated, more slender, and
becomes displaced more and more towards the left side of
the body. The lung rudiment soon begins to bend somewhat
dorsally, and then continues to grow directly backwards. It

502

J. GEAHAM KEEE.

Text-fig. 8 a.

Text-fig. 8 b.

Text-pig. 8 c.

Text-fig. 8 d.

I to::!).

Text-figs. 8 a, b, c, d. —Reconstruction seen from the ventral side of a thick horizontal
slice through the pharyngeal region of Pro top ter us, illustrating the early develop-
ment of the lung. a. Stage XXXII. b. Stage XXXIV. c. Stage XXXIV. d. Stage
XXXV. e. g. External gill. /. g. Fore-gut. 1. Lung. op. Operculum, pa. d. Dorsal
pancreas, p.b. Post-branchial body. p.f. Pectoral limb. th. Thyroid, v.c. Visceral
cleft rudiment. (Cut surfaces are indicated by the light tone.)

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTERUS. 503

is enabled to do this by the already mentioned displacement
of the oesophageal rudiment towards the left side. In speci-
mens where the displacement of the oesophagus has not

Text-fig. 9.

Transverse section of a young Lepidosiren of stage XXXIV
through the region of the glottis, a. Aorta, at. Atrium, c. o.
Cement organ, gl. Glottis, n. Notochord, oc. r. Occipital rib.
p./. Pectoral limb. ph. Pharynx, v. s. Ventricular septum.

taken place to so great an extent, the lung grows round it in
the manner to be described later.

By stage XXXIV the hind end of the lung rudiment is dis-
tinctly bilobed, the right lobe being for some time relatively

504

J. GEAHAM KERR.

small and inconspicuous as compared with the left (text-fig. 8,
B and c).

By stage XXXV (text-fig. 8, d)^ the two cusps of the lung
rudiment are seen to be growing actively. They are now
approximately of equal length, and they extend back for a
short distance along the dorsal side of the main mass of yolk,
their tips lying in the shallow valley upon its dorsal surface.

Subsequent stages in the development of the lungs will be
made clear by text-figs. 4, a — d (pages 490, 491) repi-esenting
dissections of young Lepidosirens of stages XXXII, XXXV,
XXXVI, and XXXVII. It will be noticed how the lungs
gradually extend backwards, at first in the shallow valley,
already mentioned, on the dorsal side of the enteron. Later
on they lie well above the surface of the enteron and, indeed,
eventually, as will be shown later, dorsal to the entire
splanchnoccele.

The lung rudiment is at first quite solid, but a cavity soon
begins to develop in it, the pharynx at this level still remain-
ing solid. By about stage XXXII in Lepidosiren the lung
has become hollow throughout, although the pharynx at the
level of the glottis is still solid. It is not till about stage XXXV
that the lumen of the pharynx is completed, and there is an
open glottis leading into the lung. It was at this same stage
that the young Lepidosii-ens were observed first to SAvallow air,
so we may take it that the lung is functional practically from
the time at which its communication Avith the exterior is
established. Con-elated Avith this the mesodermal sheath of the
lung is highly vascular by this stage (XXXV), and in places
the blood-vessels are seen to penetrate the endodermal lining.

Torsion of the lung. — During the course of its develop-
ment the lung undergoes a complicated process of torsion,
Avhich introduces considerable difficulties in the Avay of its
investigation. Indeed, had not extensive material been
available, it would in all probability have proved impossible
to make out exactly Avhat happens. The difficulty is due to
the fact that torsion of the lung rudiment takes place suc-
cessively in two opposite directioiis.

ALDIENTAEY CANAL IN LEPIDOSIREN AND PROTOPTEEUS. 505

Text-fig. 10 a. Text-fig. 10 b.

Text-figs. 10 a, b, o, i>. — Sections from the same series as that shown in
text-fig !) to illustrate the changing relations of lung to gut as it passes
backwards, a. Aorta, cjlom. Glomus. 1. Lung. n. Notochord, ces. (Eso-
phagus.

506

J. GRAHAM KERR.

There is first what may be called the primary torsion of
the lung, which is illustrated by the camera drawing in text-
figs. 9 and 10, A — D. During the early stages of its develop-
ment the lung rudiment, while increasing in length, describes
a spiral curve ^ round the oesophagus. Starting from the
mid-ventral glottis it grows first ventralwards, tailwards,
and towards the right side; then dorsalwards and tailwards;
and, finally, tailwards and towards the mesial plane, until it
attains its mid-dorsal position over the gut, after which it
grows directly tailwards. Now, during the spiral part of this
course, in addition to changing its position relative to the gut
(being first ventral, then on the right, and finally dorsal),
the lung undergoes what I have called the primary torsion —
torsion about its own long axis in a counter-clockwise
direction, as seen from the tailward direction. This torsion
takes place through 180°, so as to cause a complete reversal
in position of the hinder part of the lung rudiment at this
stage, its morphologically ventral aspect becoming dorsal, its
originally right side becoming left. The process of torsion
is continued still further, however, in the majority of speci-
mens, for it may be as much as 30° or 40°. The result is that
in such specimens, when the tip of the lung begins to
bifurcate, the (actual) left lung is seen to be considerably
displaced towards the ventral side, as compared with the
right (see text-figs. 8, B and c). This difference in level soon
becomes corrected by processes of differential growth, in
which a secondary torsion takes place, so that the two lungs
are brought to the same level. It will be seen that this
secondary torsion is in a direction the reverse of the primary
torsion (i. e. it is clockwise, as seen from the tail end).^ The

' Only visible in occasional individuals. More usually the oesophagus
is sufficiently out of the way to the left side to render the cuiwing no
longer necessary.

" The primary torsion of the oesophagus is clearly of the type which
would naturally be associated with a spiral coiling of a dextral type
like that of the inid-giit, while the secondary torsion may express a
tendency to return to the original condition. The possibility is

ALIMENTARY CANAL IN LEPIDOSIEEN AND PROTOPTEEUS. 507

occurrence of these two torsional processes in opposite direc-
tions introduce, as will readily be understood, very puzzling
and deceptive appearances into the sections of larvae of such
stages of development.

Topographical Relations of Lung to Coelom. — In
its earliest stages the endodermal lung rudiment is naturally
enclosed within the mesenchymatous tissue of the splanchno-
pleure. In a Protopterus of stage XXXIV the root of the
lung retains these relations. If, however, sections farther back
towards the hind end of the lung rudiment are examined it is
found that the oesophagus has, in this region, bent away
towards the left side, and has become freed from the dorsal
mesentery, which passes direct from the under surface of the
dorsal aorta to the upper surface of the liver. The lungs
grow directly backwards in the substance of this dorsal
mesentery. (It may be mentioned incidentally that in Poly-
pterus the hind portion of the large right lung retains
throughout life this relatively primitive position in the sub-
stance of the mesentery.)

The dorsal mesentery undergoes a remarkable process of
thickening from side to side, forming a broad mass of spongy
connective-tissue bounded superficially by ccelomic epithelium.
It is specially broad dorsally, and it is in this specially bi'oad
dorsal region that the lungs are situated, so that as the
broadening of the base of attachment of mesentei’y to dorsal
body-wall goes on the originally dorsal part of the mesentery
(containing the lungs) becomes gradually completely merged
in the roof of the splanchnocoele. The lungs thus come to be
situated outside of and completely dorsal to the splanchnocoele.

General Discussion of the Morphology of the
Lungs in Lepidosiren and Protopterus. — I feel com-
pelled to accept the general homology of the organs known
in various subdivisions of the Vertebrata under the name of
lung and swim-bladder or air-bladder. The early stages of

obviously suggested that the spiral coiling may once have extended
forwards into the region of the oesophagus, and that the primary torsion
has persisted on the straightening out of the spiral coils.

508

J. GRAHAM KERR.

development of this organ in the ordiuai*y lung-breathing
animals, in Lung fishes, and in the most archaic existing
Teleostome — Polyp ter us — are identical in their main
features, the differences which are so conspicuous in the
fully-developed organ of the adult being of a purely secondaiy
character. For convenience I will use the term lung to
express the organ alluded to in whatever form it occurs.

It must further, I think, be conceded that the original
position of the lung was ventral. This can hardly be denied
in view of the fact that in both Crossop terygians and Lung
fishes — both of them archaic groups in which the lung plays
an important hydrostatic function and has, in correlation
with this, assumed in the adult, partially or completely, a
position dorsal to the other viscera of the splanchnocoele — the
whole lung rudiment is in early stages of development, as is
the glottis throughout life, ventral in position.

But, admitting these two points, there at once ai-ises the
further question as to the exact method by which the dorsal
position of the lung, e. g. of a Teleostean fish, has come about
in phylogeny, and this carries with it other questions as to
the precise hoinology of the parts of the dorsal lung- of such
forms with those of the ventral lung.

It was Sagemehl ^ who first propounded the view which, in
its main features, finds a striking corroboration in the facts
of ontogeny and adult anatomy of the Dipnoi. Taking the
bilobed mid-ventral condition of the lung as a relatively
primitive one, Sagemehl points to the condition in Poly-
pterus in which the left lobe or left lung has become
greatly reduced, as is so frequently the case in lung-breathing
Vertebrates with elongated bodies. Were this lopsided con-
dition of the lung can-ied further, and the left lobe reduced
to relatively insignificant dimensions, it is obvious that there
would no longer be any insuperable difficulty in imagining a
dorsalward shifting of the remaining right lung round the
right side of the oesophagus, so that eventually a dorsal
position might be attained, as in e. g. Ceratodus. Sagemehl
* ‘ Morpliol. Jalirh.,’ x, 1885, p. 108.

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTERUS. 509

believed that the arrangement in Lung-fishes had actually
come about in this way, and that in Actinopterygians matters
had gone a step farther in that the glottis had become dorsal
as well as the lung itself. Sagemehl, it is true, greatly
impressed by the discovery that in Ery thrinus, Macrodon,
and Lebiasina the glottis was situated on the left side of
the pharynx, took the view that in the Actinopterygians the
migration of lung apparatus had been up the left side of the
pharynx instead of up the right side, as in Lung-fishes. No
one, however, who is familiar with the wide range of variation
in the position of the glottis, now to the right, now to the left
side of the median plane in e. g. Characinids and Siluroids^
will probably see any reason to suppose that the lung migra-
tion has not taken place in exactly the same fashion in the
Actinopterygians and in the Lung-fishes.

The Lung of Polyp ter us. — The condition of the lung
in the adult Polypterus (text-fig. 11, a) must be noticed,
especially in regard to three points. Firstly, the lung is here
an important hydrostatic organ, and correlated with this we
find that the lung apparatus as a whole shows a symmetrical
arrangement. The hinder half of the large right lung being
without any fellow on the left side to balance it, has assumed
a median position lying in the dorsal mesentery, and is prac-
tically symmetrical about the mesial plane; " it is only in its
anterior half, where it is still balanced by the remains of the
left lung, that it bends away from the mesial plane towards
the right. It is obvious then that, were the left lung to
undergo still further reduction, we should expect more and
more of the right lung to assume the mesial position, until at
last, when the left lobe had approached vanishing point, the
right lobe would tend to be symmetrical about the mesial
plane right to its front end. The whole lung apparatus

’ More especially if we Lear in mind the fact, chronicled above, of the
definitely dorsal side of the lung in the young lung-fish being carried
temiiorarily to the left side by the primary torsion.

- In the dissection shown in text-fig. 11a the hinder i)art of the right
lung has been displaced towards the right side and the symmetry is
consequently destroyed.

Text-fig. 11a.

Text-fig. 11 b.

Text-figs. 11a, b. — View from the dorsal side of the lung of
Polypterns (a) and Lepidosiren (b) to show nerve
sujiply. ini. Intestine. 1. 1. Left lung. l.p.a. Left pul-
monary artery, as. (Esophagus, ph. Pharynx, r. 1. Eight
lung. st. Stomach, r.p.a. Eight pulmonary artery. XL Pul-
monary branch of left vagus. Xr. Pulmonary branch of right
vagus.

ALIMENTARY CANAL IN LEPIDOSIEEN AND PEOTOPTEEUS. 511

would thus become medio-dorsal except the pneumatic duct
or trachea, which would lead round the right side of the
oesophagus or pharynx to the still ventral glottis.

(2) Correlated with the increased size of the right lung in
Polypterus we find that the left vagus takes a share in its
innervation, a stout, in fact the main, branch of this nerve
passing across to the right lung dorsal to the oesophagus.

Bearing in mind the mode of development of the lung from
a ventrally placed rudiment it need hardly be pointed out
that this dorsally placed connection between left vagus and
right lung must be a secondary development. How it has
come about does not concern the argument, but it may well
have been owing to the short circuiting of nerve impulses
through the nerve plexus of the pharyngeal wall having
caused that part of the plexus which formed the path of the
impulses to become enlarged so as to form a distinct nerve
trunk.

(3) The glottis or opening from pharynx is no longer
symmetrical in the adult Polypterus in relation to the two
lungs. It has undergone a shifting towards the right side,
and is in line with the larger right lung.

The two first of the three points just established have an
important bearing upon the comprehension of the conditions
seen in the Lung-fishes. (1) shows us how the condition of
the Lung-fishes with their dorsal lung communicating with a
ventral glottis round the right side of the alimentary canal is
just a natural step beyond the condition actually existing in
Polypterus. (2) does away entirely with the at first sight
apparently insuperable objection to adopting this as a phylo-
genetic hypothesis involved in the peculiar nerve supply of
the lung in the Lung-fishes — where (text-fig. 11, b) the pul-
monary branch of the left vagus extends on to the actually
right lung by a path which is dorsal to the oesophagus — for
we see that in Polypterus (in which the lung retains the
assumed ancestral condition) there already exists a similar
prolongation of the left vagus dorsal to the oesophagus to the

512

J. GRAHAM KERR.

lung on the ihght side of the body, an arrangement that must
necessarily have been developed secondarily.

It will be seen that neglecting the discordant evidence of
the left vagus as we are now justified in doing, the course of
the right vagus, right pulmonary artery, and left pulmonary
artery agree in testifying that the lung has undergone a
twisting on its long axis in a counter clockwise direction
(seen from behind) during its movement to the dorsal posi-
tion. The X-like crossing of the two pulmonary nerves
would of course indicate that this twistina; of the lunsf
apparatus had taken place previous to the establishment of
the nervous “shoi't circuit” dorsal to the oesophagus.

Having’ now shown (1) the clear probability of an ancestral
arrangement like that of Polyp ter us becoming evolved
into an arrangement like that of existing Lung-fishes, and
(2) that the obstacle formed by the course of the left vagus is
of no importance, it remains now to pass back to the evidence
afforded by the ontogeny of Lepidosiren and Pro-
top ter us.

These embryological phenomena show clearly (1) that the
lung rudiment is originally ventral in position ; (2) that
during its development distinct twisting of the lung in a
counter-clockwise direction takes place ; and (3) that during
early stages of development the actual right lung, i.e. the
lung which on Sagemehl’s hypothesis is homologous with the
small left lung of Polyp ter us is actually much smaller than
its fellow.

Taking into account these various considerations we are,
I think, irresistibly driven to the conclusion that, so far as
regards the Dipnoi, Sagemehl’s hypothesis must be accorded
a very high degree of probability.^

Actinopterygii. — The support to SagemehPs hypothesis
which has been adduced in the foregoing paragraphs lends
increased probability to a similar view being applicable to
the Actinopterygians. It has been indicated how in Poly-

* In this I agree with N EUM aye, Semon’s ‘ Zoolog. Forschungsreisen,’
I, p. 407.

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTERDS. 513

p ter US the lung apparatus is in process of attaining to a
mediodorsal position, the hinder half of the large right lung
having already done so. In Ceratodus the whole of the
lung except glottis and air duct has attained to the mid-
dorsal position, and the original left lobe has apparently been
completely withdrawn into the lung, so that the latter is a
single structure without any paired appearance. It is clearly
but a step from the Ceratodus condition for the air duct to
become shortened and the glottis to reach the neighbourhood
of the mesial plane dorsally, such dorsalward migration of the
glottis being aided by the tendency of the gut to become
rotated in a counter-clockwise direction, as seen from the
tailward end.^

V. Pancreas.

There are in the Dipneumona, as in Ceratodus and the
majority of Vertebrates, three pancreatic rudiments, one
dorsal and two ventral.

Protopterus. — The dorsal rudiment is the first to make
its appearance (about stage XXXII) in the form of a solid
projection from the dorsal surface of the yolk practically in
the mesial plane. In some embryos of this stage the dorsal
pancreatic rudiment is greatly elongated in an aiitero-posterior
dii’ection — possibly a reminiscence of some unknown earlier
phylogenetic condition (text-fig. 12, a). In most embryos,
however, the dorsal pancreas is of compact and rounded form
(cf. text-figs. 8, B, and 12, b), and is situated in about the
same transverse plane as the hinder nephrostome of the
pronephros. The attachment of the dorsal pancreas becomes
rapidly constricted to form a narrow stalk, and a small,
irregular cavity appears in the interior of the organ.

By stage XXXIII the ventral rudiments have made their
appearance in the form of a yolky projection from the gut on

' Moser (‘Arch, iiiikr. Anat.,’ Ixiii, 1904, p. 562) has demonstrated the
existence of this in ontogeny. See also the interesting paper by H. Marcus
(‘ Arch, inikr. Anat.,’ Ixxi, 1908), who is led by his studies on the kmg of
Gymnophiona to a similar conclusion regarding the hmg of Ceratodus.

514

J. GRAHAM KERR.

eitliei’ side of the attachment of the bile-duct rudiment. The
right is more bulky than the left. Both right and left
rudiment grow rapidly in a dorsal direction, one on either
side of the bile-duct, and finally the left arches towards the
right side, dorsal to the bile duct, comes in contact with the
tip of the right rudiment, and undergoes fusion with it
(between stages XXXIII and XXXIV most usually). The

Text-fig. 12 a.

Text-fig. 12 b.

,Tr, I.

Text-fig. 12 a, b. — Reconstruction of thick horizontal slice through
Protopterus, showing rudiments of lung and (dorsal) pancreas.
A. Stage XXXII. b. Stage XXXIV. 1. Lung. op. Operculum.
pa. d. Dorsal pancreas. p. b. Post-branchial body. p.f. Pectoral
limb. V. c. Visceral cleft rudiment.

dorsal pancreatic rudiment has meanwhile been growing
rapidly. Its wall is still thick and yolk-laden, but its cavity
has increased in size, and has become more regular in shape.
With the gradual modelling of the surface of the gut a
change has been brought about in the attachment of the
dorsal pancreas ; it now springs from the right side of the
gut just where the mid-gut is continued forwards into the
fore-gut. It projects towards the right side of the body, and
at the same time slightly forwards.

Up till now the dorsal pancreas has been separate from the

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTERUS. 515

fused ventral rudiments, but between stages XXXIV and
XXXV the dorsal surface of the right ventral pancreas comes
in contact with the ventral surface of the dorsal pancreas,
and fusion promptly takes place, so that from now onwards
there is a single pancreatic complex.

By stage XXXV the pancreatic complex forms a voluminous
organ, the dorsal end of which is visible in a dissection from
the dorsal side (cf. text-fig. 13 of Lepidosiren at this stage),
lying immediately to the left of the dorsal lobe of the liver — •
between it and the junction of fore- and mid-gut, and ex-
tending back in the spiral groove. This dorsal pai-t of the
complex extends ventrally downwards into the posterior limb
of a saddle-shaped mass, which bestrides the bile-duct, and
which represents the fused right and left pancreatic rudi-
ments, the original right rudiment having now become pos-
terior in position. The threefold origin of the pancreas is
still betrayed by its three attachments to the gut, the original
right and left ventral being now immediately posterior and
anterior to the point of junction of bile duct and gut, while
the long fine duct, which represents the long drawn-out stalk
of the dorsal pancreas, opens in the posterior angle between
fore-gut and mid-gut. The right and left ventral attach-
ments are still solid. Tlie substance of the gland itself is
seen in sections to be undergoing obvious histological diffe-
rentiation, and to be penetrated by a rich network of blood-
vessels.

By stage XXXVI the pancreas Inis become actively func-
tional, and its cells have attained their definitive character
with nucleus at outer end, and the main mass of the cell
protoplasm being packed with zymogen granules. With the
tucking in of the gut wall to form the spout-like pyloric
valve, the opening of the dorsal pancreatic duct is carried
inwards, and is now found to open into the cavity of the
spout-like structure, the actual opening being on the dorsal
wall of the spout.

Lepidosiren. — In Lepidosiren the general features of
pancreas development are as in Protopterus, with minor

VOL. 54, HART 4. — NEW SERIES. 37

516

J. CxEAHAJI KERR.

differences in topographical relations in early stages. In
a larva of stage 32 (text-fig. 4, a) the dorsal pancreas forms
a rounded projection from the gut^ as in Protopterus, but
situated farther back, well behind the level of the hinder
pronephritic nephrostome. The rudiment in Lepidosiren
has already a wide cavity opening into the cavity of the gut.
By stage XXXIV the yolk in the rudiment is practically used
up, and by stage XXXV the gland cells and duct lumiua are
distinguishable; the whole organ is penetrated by a rich net-
work of blood-vessels, and the outer surface of the organ
presents a distinctly lobed appearance (see text-fig. 13).

Text-fig. 13.

Part of the dissection shown in text-fig. 4, b (p. 490), with the
kings removed so as to show the pancreas. /, g. Fore-gut.
ht. Heart. H. Liver, pa. Pancreas, ph. Pharynx, v. Vein.

As regards the later development of the pancreas in
Lepidosiren and Protopterus, the only point requiring
special notice is that in the adult (cf. text-fig. 4, d), as was

ALIMENTARY CANAL IN LEPIDOSIREN AND PROTOPTEEUS. 517

shown by W. N. Parker, it remains enclosed within the
splanchnic mesoderm ensheathing the gut, neither bulging
into coelotn nor spreading in the substance of the mesentery.
As a consequence the pancreas remained undetected by the
earlier investigators.

VI. Summary.

1. The fore-gut first becomes folded off from the main mass
of yolk-cells.

2. The pyloric valve arises by the hind end of the fore-gut
being pushed back into the cavity of the mid-gut.

3. The main mass of yolk-cells becomes gradually
“ modelled” into a spirally-coiled intestinal rudiment.

4. The main part of the buccal lining- is developed in situ
from large yolk-cells.

5. The part of the ventral side of the head, on which are
the olfactory rudiments, becomes enclosed in the buccal
cavity by the development of the upper lips and by the for-
ward growth of the lower jaw.

6. The olfactory opening becomes divided into anterior and
posterior nares by the apposition and fusion of the inter-
mediate portion of its lips.

7. The thyroid arises as a solid downgrowth from the
buccopharyngeal floor, which gradually becomes cut off from
behind forwards.

8. The tongue is a primary tongue like that of Urodeles,
but without gland-field.

9. The lung arises from a solid mid-ventral rudiment.

10. When the lung becomes bilobed, the (actual) right
lobe is for a time small in size as compared with its fellow.

11. Complicated torsional processes take place during the
development of the lung.

12. Through the dorsal mesentery becoming partially
merged in the splanchnocoele roof, the lungs come to lie
outside the splanchnocoele.

518

J. GRAHAM KERR

13. The general facts of lung development go to support
the view that the lung of Polypterus shows a persistence
of the condition ancestral to that of Dipnoi and Actino-
pterygii.

14. The pancreas arises from a dorsal and two ventral
rudiments.

THE PHYLOGENT OP THE TEACHEiE IN AEANE^. 519

The Phylogeny of the Tracheae in Araneae.

By

W. F. Purcell, Pli.D.,

Bergvliet, Diep River, near Cape Town.

With Plate 28, and 21 Text-figures.

Introduction.

In an excellent paper on the tracheae of spiders E. Lamy
(:02) has given an account of the tracheae of thirty families
of Araneae, so that only four small and comparatively rare
families, comprising 1 — 3 genera each, remain, of which the
tracheae are still unknown. It is now possible, therefore, to
consider the tracheal systems of the Araneae as a whole
from a phylogenetic point of view, and as I barely touched
upon this point in my paper (:09) on the development and
origin of the respiratory organs in spiders, I propose to make
it the subject of the present paper.

Lamy has made it perfectly clear that the degree of com-
plication of a tracheal system, as regards the manner and
extent of the branching and the structure of the internal
armature (spines, spiral thread, etc.), cannot be used as a
family character, since we may find the most varied degrees
of complication amongst the different genera of one and the
same family (e. g. in the Uloboridee, Thomisidae, Age-
1 enidae, Clubionidm, Attidae, etc.). Lamy concludes from
this that the tracheal apparatus is evolved separately in each
family and not in the Araneae as a whole (p. 265) and this
statement may, I think, be accepted as in general correct.

520

W. F. PURCELL.

provided that it be not interpreted to mean the development
of the tracheae out of lung-books separately in each family
but merely the development of more complicated tracheal
systems from simpler ones or vice versa.

Another important point which Lamy has emphasised
(p. 264) is that the number of the lung-leaves is in inverse
ratio to the size of the tracheal apparatus. Thus^ forms with
highly-developed tracheee have a small number of lung-
leaves, as Dictyna, with 4 — 5 leaves (Bertkau), Segestria,
with 10 — 12 (Bertkau), etc., while forms with a feebl3'-deve-
loped tracheal system have a comparatively large number of
lung-leaves, as Bpeira and Agelena, with 60 — 70 leaves
(Bertkau).

Since the i-elative size of the tracheal apparatus in general
increases proportionately with its degree of complication it
further follows from the above paragraphs that the number
of the lung-leaves in forms having both tracheae and lung-
books can have no greater ph3dogenetic value than that pos-
sessed by the degree of complication (in respect to branching
and internal armature) of the tracheal apparatus and this
Lamy has shown to be of subordinate phylogenetic value,
without even the importance of a family character. We
cannot, in fact, use either of these characters in comparing
widely remote families ph3dogenetically, as, for instance, the
Dysderidae and Argiopidm (Epeiridse). Thus, assuming
that the tracheae were derived from lung-books, it would be
incorrect to argue that an Argiopid, with its numerous luiig-
leaves and simple tracheae is, on that account, a more primitive
form than a Dysderid, with its few lung-leaves and large
complicated tracheae ; or, conversely, if we admit that the
Dysderidae are more primitive than the A rgiopidae, we
could not argue that because of the different relative develop-
tnent of these two organs in the two families, the lung-books
must have been derived from trachefe.

It does not follow, however, from any of Lamy’s arguments,
that the two characters just discussed ai’e of no ph3dogenetic
importance at all, e. g. amongst allied genera in one and the

THE PHYLOGENY OF THE TEACHEiE IN AEANEH:. 521

same family, as in the Agelenidae, or between allied families,
such as the Dysde ridge and Oonopidge — nor that other
tracheal characters, such as the acquisition of the respiratory
function by the ectodermal tendons of the tracheal segment,
which I have shown to have taken place in most spiders, may
not have a much higher phylogenetic value.

Before entering upon this subject I wish to consider a
certain remai’kable conclusion drawn by Lamy, viz. that in
spiders neither the lung-books nor the trachete are the more
primitive organs (p. 264), both having been produced simul-
taneously and replacing one another (p. 265). The sole
difference Lamy sees between these two organs lies in their
special mode of branching, lamellate branches pi'oducing
lung-books and tubular ones trachejB (p. 267). The branchial
origin of the lung-books is discarded by him as unnecessary
and the formation of the tracheal organs is considered to be
the consequence of the respiratory function taking place in
the same conditions in all air-breathing Arthropods (pp. 266
and 267).

Lamy arrives at the above conclusion by the following
ai’guments (p. 264): — (1) The Dysderidm and the Capo-
niidm come very near together, approaching one another in
several characters, and ought, therefore, to be regarded as
equally primitive. Nevertheless, in the latter, the first pair
of lung-books is replaced by a pair of tracheae, which strongly
resemble those which replace the second pair of lung-books
in the Dysderidm. The fact that one sees the trachete
indifferently replacing the lung-books in these two somewhat
primitive families indicates that neither organ is to be re-
garded as more primitive than the other. (2) The same con-
clusion results fi’om the fact that we find amongst the
Araneae verae' another family, the Hypochilidae, which,

* Simon divides the spiders as follows: Aranese tlieraphosai (=
Mygalomorphai, Pocock -t- Liphistius), including all 4-limged
foi’ms except the H ypochilida?. Araneai verse (Arachnoniorphse.
Pocock), including the Hypochilidai and all dipneumonous and
apneuinonous spiders.

522

AV. F. PURCELL.

although it resembles the Aranem verae and not the
Theraphosae in all other respects, nevertheless, has the
tracheae replaced by a second pair of lung-books.

Neither of these arguments, however, wan-ant the conclu-
sion that Lamy has drawn from them, since both cases may
be readily explained, even when we assume that the lung-
books were in all cases the primitive organ and that the
tracheae were derived from them. Lamy’s assumption that
the Dysderidae and the Caponiidae are equally primitive
is certainly incorrect, since, as I shall presently show, the
Caponiidae differ from the Dysderidae, as well as from all
other spiders (so far as I know), in several important anato-
mical characters. They are in fact, in these respects, a
highly-specialised group, compared with which the Dys-
deridae are much more primitive. But even if these two
families were equally primitive they are by no means the
most primitive spiders, the vast host of mygalomorphous
forms being all more primitive than they and all provided
with lung-books only. Moreover, the highly-developed
tracheae of the D3’sderid£e, which present no obvious resem-
blance to lung-books, do not so sti’ongly resemble the anterior
trachem of the Caponiidae as Lamy makes out, since these
latter are very similar to the lung-books of a Dysderid. In
fact, these anterior tracheae may be most readil}^ explained,
as I shall presently show, as lung-books which have been
transformed into tracheae more recently than those of the
second pair and which have retained the primitive shape more
nearly than has been the case with the tracheae of any other
Arachnid known. They have evidently been evolved out of
a few-leaved lung-book like that of a D\^sderid, and their
presence merel}’ proves that tracheae have been evolved out of
lung-books within the Aranefe at least on two occasions, but
it does not prove that the tracheae and lung-books are equally
primitive.

Similarly, the presence of a second pair of lung-books in
the Hypochilidae may be quite readily explained by
assuming that this family is an arachnomorphous form in

THE PHYLOGENY OP THE TRACHEA IN ARANE^. 523

which the primitive lung-books have been retained, whereas
they have been replaced by tracheae or lost in all other
members of the group. It is not at all necessary to assume
that the Hypochilidse once possessed tracheae, nor that the
lung-books and the tracheae must necessarily be equally
primitive organs.

Lamy’s conception of a lung-book as developing from an
ectodermal invagination with lamellate branches (p. 256) is
incorrect, since, as I had already shown some years previously
(’95), the two oldest saccules^ are formed as independent
invaginations on the free posterior side of an embryonic
abdominal appendage, quite outside of the basal pulmonary
sac (vestibule) in the anterior wall of which the remaining
saccules appear, — the two oldest saccules being only later on
included within the pulmonary sac, when the sinking of the
appendage takes place. Lamy puts this observation aside
with the remark that I am the only observer who mentions it,
but it is none the less a fact.

I have already (:09) fully discussed the question of the
primitiveness of the lung-books, and have shown on purely
embryological grounds that the typical form of trachem found
in most spiders must have been derived in part from lung-
books and in part from ectodermal tendons (entapophyses of
Eay Lankester, apodemes), the lateral pair of tracheal trunks
being metamorphosed lung-books and the medial pair meta-
morphosed entapophyses.

Starting from this as a basis, the tetrapiieumonous group,
Araneso theraphosse, appears the most primitive of living
Araneas, a view which has, in fact, long been generally
recognised on account of other primitive characters of the
group, such as the presence of a free nervous ganglion
behind the central nervous mass in the cephalo-thorax, the

I have given the term “saccules” to the hollow air-containing
leaves of a lung-book and “septa” to the j)artitions or lamellaj
separating the cavities of adjacent air chambers (: 09).

524

W. P. PURCELL.

simple form of the external sexual organs, the presence of
four spiracles, etc.

The remarkable genus Liphistius, which I have had no
opportunity of carefully examining, appears, as Pocock (’92)
has pointed out, to be much more primitive than the rest of
the group, at least in some of its characters. One of the most
interesting among these is the mesial position of the spinners
on the under side of the abdomen, so that in this genus
none of the ventral abdominal segments have been exces-
sively elongated (text-fig. 1). Considering the apparently
primitive structure of this genus, which has also its abdomen

Tkxt-pig. 1. — Abdomen of Liphistius (after Pocock, ’92).

Tkxt-fig. 2. — Abdomen of a mygalomorphous spider.

ec. t. 8 and 9. Depressions in the integument to which the ventral
longitudinal muscles are attached at the posterior margins of
somites 8 and 9 g. o. External genital opening. IV., lb".
First and second pair of lung-books, sp'., f^p". Spiracles of the
first and second pair of lung-books. 7 — 11 denote the extent
of the seventh to eleventh somites.

segmented dorsally like a Pedipalp, it is somewhat peculiar
that both respiratory segments (judging from the figures
and descriptions given by Simon and Pocock) evidently
possess a deep interpulmonary or epigastric fold, like the
Pedipalpi and the Aranese verm.

In the rest of the group (Pocock’s Mygalomorphm) the
fourth abdominal (tenth post-oral) segment has greatly

1

2

THE PHYL0C4ENY OF THE TRACHEH: IN ARANEAl. 525

elongated at the expense of the following segments, being,
in fact, as long as or longer than the second and third
segments taken together, so as to bring the spinners to the
hinder end of the abdomen (text-fig. 2). The positions of
the spiracles {sp'., sp" .), muscular stigmata (ec. t. 8 and 9,
representing rudimentary entapophyses to which the longi-
tudinal muscles are attached), and the genital opening {g. o.)
are very primitive, at least in all the forms which I have
been able to examine. All these openings are frequently
perfectl}’ exposed and separate from each other, especially in
distended abdomens, as in text-fig. 2. There is at most a
shallow, open, transverse depression behind the posterior
edge of the segments, and the skin in this groove behind
the genital segment is frequently soft and flexible, like the
soft skin between the hard plates of a segmented body.
AVhen the abdomen is distended the spiracles and muscular
stigmata in this soft skin are exposed, but in a contracted
abdomen (such as that of a female after the deposition of the
eggs) these openings may become somewhat hidden from
view owing to the infolding of the flexible skin. Such a
groove is, however, very different from the typical, deep,
and more or less rigid infolding found behind the second and
third abdominal segments in the Pedipalpi (see Tarnani, ’89,
p. 377, fig. 1, and Lankester, ;04, fig. hh), nearly all arach-
nomorphous spidei s, and in Liphistius. The longitudinal
muscles are attached to shallow ectodermal depressions (see
my paper, ;09, fig. 36), which lie either free or in the larger
transverse grooves mentioned above, and there are, so far as
I know, no deep invaginations or ectodermal tendons (enta-
pophyses) like those found in l*edipalps and arachnomorphous
spiders. Can this and the absence of interpulmonary folds
perhaps be a secondary condition in the Mygalomorphas ?
Or have these folds been acquired independently in the
Pedipalpi, Liphistius, and Arach no m or ph as ^ ?

‘ Two interesting drawings by R. I. Pocock are given by Ray
Lankester (:04. figs. .56 and 64) showing the genital segments of a
male Thelyphonns assamensis and a female Liphistius de-

526

W. F. PURCELL.

Turning now to the Araneae verse or Arachnomorph se,
we find that in nearly every case the second pair of respira-
tory organs (and in one family the first pair as well) have
been replaced by tracheae, the exceptions being the small
family Hypochilidae with two pairs of lung-books and the
Pliolcidae, in which there are no other respiratory organs
besides the single pair of lung-books of the genital segment.
Moreover, there is a deep infolding along the hind edge of
each of the respiratory segments between the spiracles, so as
to hide from view the genital opening and the external
openings of the well-developed ectodermal tendons or enta-
pophyses of the ventral longitudinal muscles. There are a

suitor, with the epigastric fold drawn apart so as to expose the
genital opening and the edges of the septa of the lung-books. These
two figures are remarkaljle for showing that in these two Arachnids
the pulmonary saccules of the genital segment open directly into the
cleft of the epigastric fold, being, in fact, attached to the anterior
wall of the fold. I examined a female of Thelyphonus caudatus,
and found the conditions exactly as depicted by Pocock. The pul-
monary ante-chamber opens along its entire medial side into the
median part of the epigastric fold, and cannot, therefore, be said to
form a separate chamber, except in its dorso-lateral prolongation or
jjortion containing the youngest saccules. In the male (i.e. specimens
with a sjjine on the second abdominal sternite, teste Kraepelin) of this
sjjecies, however, I found the conditions different. Here there is a
longitudinal fold of integaiment on each side between the deep median
part of the epigastric fold and the anterior pulmonary chambei's, so
that the latter may be said to form separate chambers op)ening by the
ventral slit only into the epigastric fold, as is usual in dipneumonous
spiders. The condition depicted by Pocock in Liphistius is not
known to occui- in any dipneumonous spider, and may indicate that the
epigastric fold of this form is directly connected with that of Thely-
phonus and not of independent origin, in which case the absence of
the fold in the My galomorphaj would be a secondary condition. Its
presence in the four-lunged arachnomorphous family Hypochilidae
is also an interesting circumstance.

These two figures of Pocock’s should have been included in the his-
torical list of papers concerning the lung-books of Arachnids given at
the end of my previous paper (:09). They iinfortunately did not come
to my notice until after the paper had been sent to the press.

THE PHYLOGENY OE THE TEACHEiE IN ARANEJ:. 527

few rare exceptions — thus in the Dysderidse the tracheal
segment has no infolding and the genital duct sometimes
opens free on the ventral surface of the pulmonary segment
(male of Harpactes; see my paper :09, fig. 40).

Leaving the four-lunged Hypochilidm, with which I am
unacquainted, out of account, there appears in the first place
a small group of three families (Dysderidm, Oonopidae,
and Caponiidse) which possess some very primitive features
in connection with their respiratory segments. As these
segments are of peculiar interest in connection with the
phytogeny of the tracheae, I shall give some account of their
anatomy before proceeding to more general conclusions.

Material and Treatment. — The material used was the
same as that given in my previous paper (:09) with the addi-
tion of specimens of a species^ of Oonopidae fi’om the
neighbourhood of Cape Town.

For following the muscles, whicli are often very slender,
suitable differential staining is very necessary, and for this
purpose I found very old Delafield’s haematoxylin (mine was
thirteen years old) most excellent, even for old museum
specimens. The sections are stained on the slide for four to
five hours, placed in acidulated alcohol for three to four

* As this species is a new one, I append the following description :
Calculus n. g. Cephalotliorax broadly ovate. Ocular area transverse,
the eyes arranged as in Orchestina. Labium short and broad, as in
Oonops. Coxa; of pedij^alps parallel, their anterior ends widely sepa-
rated and not converging. — C. bicolor n. sj). Pale yellowish, abdomen
with a ]>road infuscate patch behind above, and narrowly blackened on
each side of the spinners as well. Clypeus l^arely as wide as an anterior
lateral eye. Anterior row of eyes, seen from above, almost straight,
the median eyes large, a trifle longer than their distance from the
anterior margin of cephalotliorax ; anterior lateral eyes the smallest of
the six, distant about half their own width from the median eyes ; pos-
terior eyes forming a row which is only very slightly wider than the
anterior row, their distance from the median eyes greater than their
own width. Tibia and metatarsus of first leg with 0-2 spines near the
middle below, tibia and especially the metatarsus of fourth leg more
numerously spined. Several females from the Cape Flats, near Princess
and Zeekoe Vleis. Length 4 mm. Allied to Tele hius, E. Sim.

528

W. V. PUECELL.

minuteSj washed with spirits, and then held inverted over a
vessel containing a drop of ammonia in some water until the
sections change colour. After this they should be mounted
in balsam without delay. The nuclei become blue and
the muscles reddish and easily distinguishable from other
tissues.

In preparing the tracheae by the caustic potash method I
obtained the best results for such highly complicated systems
as those of Caponia by first allowing the object, after
removal of a part of the dorsal integument, to remain in cold
concentrated caustic potash for twelve hours or longer. If
the solution be then gently heated and some water added the
soft parts remaining will rapidly disappear without injury to
the delicate trachem. These should be examined in water or
weak alcohol, and not in glycerine or acetate of potash, since
these latter cause the tubes to collapse and become distorted.

The Kespieatory Segments op the Dysderid.®, Oono-

PIDA5, AND CaPONIID.E.

These spiders more nearly resemble the My gal o morph a?
than they do the rest of the A rachnoniorphge in the
anterior position and wide separation of the second pair of
spiracles (sp"., text-figs. 3 — 5) and the more rudimentary
condition of the ectodermal tendons (where present) of the
second respiratory segment.

Further, the transverse epigastric fold, lying between the
two anterior spiracles, although present, never encloses a
spinous canal of communication connecting the lumens of the
two anterior respiratory organs, while the Dysderidm are
unique amongst arachnomorphous spiders in having no inter-
tracheal fold between the two posterior spiracles (text-fig. 3),
a primitive character only met with elsewhere in the
Mygalomorphae (p. 524, text-fig. 2). Owing to the presence
of trachem instead of lung-books in the ninth somite this
segment is somewhat shortened, but in other respects the

THE PHYLOGENY OP THE TRACHEJ5 IN AfiANE.E. 529

extent of the abdominal segments in the tliree families much
resembles that in the Mygalomorphse.

Dysderidfe. — The lung-books in the Dysderidse have
few leaves. I counted about thirteen in Harpactes (in
sections), but there are more in Dysdera and Segestria
senoculata (Bertkau [’72] records only ten to twelve for
Segestria). The ante-chamber {pulm. a.) is strongly
inclined forwards from the base at an angle of 40° — 50°,
and is evenly curved forwards in Harpactes (PI. 28, fig. 5),
but almost straight in Dysdera and Segestria (p. 530, text-
fig. 7). It is spined on its posterior wall except quite
interiorly, where a muscle (text-fig. 7, No. 11) is attached.

Text-fig. 3. — Abdomen of Dysdera sp., ad. ? (magn. 3).
Text-fig. 4. — Abdomen of Calculus bicolor, ad. ? (magn.

10).

Text-fig. 5. — Abdomen of Caponia spiralifera, ad. 9
(magn. 3.)

sp'., sp". Spiracles of the first and second respiratory segments.

7 — 10 denote the extent of the seventh to tenth somites.

The peculiarities of the epigastric fold have already been
described (:09).

The well-known tracheae (p. 551, text-fig. 19) have been
described by several authors (see Lamy [:02, pp. 180 — 183]
for some excellent figures of Dysdera and Segestria), and
I have given a summary of their structure with some addi-
tional observations on the entapophyses and muscles con-
nected with them (:09).

In order to ascertain anatomically whether a trachea or a

530

W. F. PURCELL.

THE PHYLOGENY OF THE TRACHEA! IX ARAXE.E. 531

part of one is homologous Avith a lung-book or with an
entapophysis it is necessary first of all to identify the
entapophyses of the great longitudinal muscles. These
entapophyses, as I have shown for Attus and Agelena
(’95, :09), arise in various abdominal segments as invagina-
tions on the posterior side of the provisional appendages,
while the invaginations Avhich form, or correspond to, a
pulmonary sac or ante-chamber always lie to the lateral side
of the entapophyses. In the two segments bearing the
spinners in Attus and Agelena the entapophyses are
attached at the posterior, medial, basal corners of the
anterior and posterior spinners. For the identification of
the entapophyses anatomically a knowledge of the abdominal
muscles connected with the respiratory segments is necessary,
and I have given the two accompanying diagrams (text-figs.
G and 7) to illustrate these muscles and their entochondrites
in a typical Dysderid.

Ijist of the Entochondrites and Muscles in Text-figs. G and 7.

t. Small entochondrite on the lateral side of the trachea
and attached to the fold of the integument, /‘d. 2.

t'. Fntochondrite between the muscles 23 and 21, etc., but
not attached to the integument.

/. H. Large entochondrite situated on the medial side of the
pulmonary aute-charnber and attached to the epigastric- fold.

t. 9. Corresponding entochondrite of the tracheal segment,
situated on the medial side of the trachea.

Muscles.

1 and 2. From the entochondrite C8 to ujtper and middle
part of side of abdominal pedicel.

3. Longitudinal from the entochondrite L8 to the cephalo-
thorax.

4. From the entochondrite L8 to ventral integument of
pulmonary segment.

VOL. 54, PART 4. — Xltw SERIES.

38

532

W. F. PUECEJiL.

5. From upper part of side of abdominal pedicel to venti al
integument of pulmonary segment. (1 and 5 are inserted
together anteriorly.)

0. Dorso-ventral on side of abdominal pedicel. (2 and 6
are attached to the same ectodermal infolding on the medial
side of 5.)

7. Oblique dorso-ventral from hinder end of the ectodermal
tendon ec.t. to the ventral fold/d.l of abdominal pedicel.

8. Oblique dorso-ventral from hinder end of the ectodermal
tendon ec..t. to the entochoudrite t.

9. From the ectodermal tendon ec.t. to anterior intesjument
of abdomen.

10. From the ectodermal tendon ec.t. to dorsal integument
of abdomen.

11. From latei’al part of posterior side of pulmonary ante-
chamber in a postero-dorsal direction to the entochoudrite t.
(This muscle can widen the ante-chamber in Segestria.)

12. Longitudinal parietal along ventral integument of
abdomen.

13. From the entochondrite t.9 to ventral integument of
abdomen (some strands apparently continuous with 12.)

14. From the entochondrite t. to ventral integument of
abdomen.

15. Longitudinal connecting the entochondrites t.S and

^.9.

16. From posterior side of entapophysis (cc.f.8) of pul-
monary segment to anterior side of the integumental fold/d.2.

17. Subtransverse from the entochondrite t.9 in a medial
direction to posterior side of base of epigastric fold.

18. Longitudinal from the entochondrite i.9 to the spinners,
breaking up posteriorly into 19, 20, 21 and 22.

19. Three muscles from 18 (one large one to medial side of
large posterior spinner and two smaller ones to small mesial
spinner).

20. From 18 to posterior medial side of anterior spinner.

21. From 18 to medial side of same spinner.

22. From 18 to anterior side of same spinner.

THE PHYLOGENY OF THE TEACHE.T: IN ARANEH:. 533

23. Longitudinal connecting the entocliondrites t. and t' .

24. Longitudinal from the entochondrite t' . to the spinners,
dividing posteriorly into 25 and 26.

25. Two muscles from 24 to anterior lateral angle of large
posterior spinner.

26. From 24 to posterior lateral angle of anterior spinner.

27. From the entochondrite t'. to anterior lateral side of
anterior spinner.

28. From the entochondrite t'. to anterior side of anterior
spinner (inserted behind 22).

29. From posterior end of the cephalothoracic tracheal
trunk ceph.. tr. (inserted just below the origin of the short
abdominal branch) to ventral integument of abdomen.

30. Subtransverse connecting the entocliondrites t.S and t.

31. Connecting the entochondrites ^9 and t'.

(Tsvo other muscles of the female of Harpactes are given
in PI. 28, fig. 3).

3’he part of the epigastric fold (text-fig. 6, ec.t.S) to which
the entochondrite t.S is attached, plainly corresponds to the
entapophysis of the pulmonary segment in other dipneu-
monous spiders (see my paper :09, fig. 41, ec.t.S), but the
identity of the corresponding entapophysis of the tracheal
segment is not at first sight so evident, since there are two
entochondrites, one on each side of the trachea, and two sets
of longitudinal muscles, both connected with the entochon-
drite t.S. In fact, the whole muscular system of the ninth
and tenth somites is very different to that of Attus, Age-
lena, Epeira, etc., which is, of course, due to the circum-
stance that in the latter the relative lengths of the two
somites are exactly reversed. From the fact that the muscles
19 — 21 of the medial set in Segestria (text-fig. 6) are con-
nected with the medial and postero-medial sides of the
anterior and posterior spinners, while the muscles 25 — 27 of
the lateral set (text-fig. 7) are connected with the lateral
side of the same spinners, it is evident that the entochon-
drite t.9 is the one which in the tracheal segment corresponds
to the entochondrite t.S of the pulmonary segment. The

534

W. F. PURCELI,.

entocliondvite ^.9 is attached (in Segesti'ia at least) to a
small lobe of the medial side of the tracheal pedicel. This
lobe (the entapophysis of the tracheal segment), which I
have already described and figured (:09, figs. 32 and 33,
ee.f.9), is not really a part of the trachea proper, being merely
connected with the base of the pedicel and unspined internally,
and I am not even sure that it is present in Dysdera or
Harpactes, in which genera the entochondrite may possibly
be attached directly to the integument at the medial basal
angle of the tracheal pedicel. From the position of the
trachea on the lateral side of the entochondrite t.9, it is
evident that the whole of the trachea must be considered as
homologous with a pulmonary sac or lung-book, as I have
already pointed out (’95) and Lamy also agrees with this view
by reason of the position and appearance of the tracheae and
the complete separation of the spiracles (:02, p. 259).

Oonopidae. — This family is very closely allied to the Dy s-
deridae as was pointed out by Bertkau (’78), who included
Oonops in the latter. The only anatomical difference of any
importance connected with the respiratory segments appears
to be the presence of an intertracheal fold observed by Lamy
(:02), joining the two tracheae in the Oonopida3.

I found the muscular system connected with these seg-
7uents to be practically identical in the two families, all the
characteristic muscles of the Dysderidm being present in
the Oonopid, Calculus bicolor, examined by me, although
sometimes in a somewhat modified form.^ Thus the muscle
29 (text-fig. 7) is much shorter and 15 (text-fig. 6) somewhat
longer in the Oonopid, as the posterior trachefe ai-e here
placed further back. Tlie lateral entochondrite t. is particu-
larly conspicuous, much more so than in Segestria.

The tracheal trunks are quite similar in both families.
Those of Oonops were first described by Bertkau (’78) and
later in greater detail by Lamy (:02), who also examined a

* Three additional muscles not noticed in Segestria are given in
figs. 1 and 2, hut two of these (ru. 38 and 40) are also found in the
female of Harpactes (fig. 3).

THE THYLOGENY OE THE TEACHE.E IN ARANE.H. 535

Dysderina. Neither o£ these authors, however, observed
the anterior ending of the ceplialothoracic trunks, but quite
correctly supposed them to eud, as in the Dysderidm, in a
bundle of tubules. In Calculus bicolor these trunks are
short and very much as in text-fig. 6. They do not enter the
abdominal pedicel, but break up at the anterior end into a
dense cluster of fine tubules, which then pass through the
pedicel into the cephalothorax. 'J'he short posterior branch,
fii’st found in this family by Lamy, is also present, and
corresponds exactly to the similar branch in the Dysderidae.
The anastomosing ends of the internal spines form a simple
network, like that in Harpactes, but in the forms examined
by Bertkau and Lamy they are said to form a spiral thread.
The cavities of the two tracheal trunks are directly connected
by a spinous canal of communication (fig. 1, can.), enclosed
within the iutertracheal fold [tr. fd.). As in the Bys-
d e r i d a3, the tracheal trunks and their branches
are to be considered as entirely homologous with
lung- books.

Another important point of resemblance to the Dys-
deridte is the presence in the female of a single median
receptaculum seminis, pointed out by Bertkau (’78), who
observed that such a receptaculum is not found in any other
family of spiders besides these two (p. 374). In Calculus
bicolor the receptaculum forms an elongate, narrow, curved,
median pouch (fig. 2, r. s.), placed horizontally with the
concavity of the curvature upwards, and opening into the
anterior wall of the epigastric fold {ep. fd.). From the under
side of the pouch a large vertical keel {Ic.) hangs downwaids,
reaching to the body hypodermis. Fach side of this keel
serves for the attachment of a powerful muscle (/a. 38), which
runs obliquely backwards and outwards to the anterior surface
of the entapophysis of the epigastric fold (tig. 1). There is
also a median muscle {ui. 4Uj running’ from the under side of
the pouch along the posterior edge of the keel to the ventral
body integument. In Harpactes Hombergi I found a
very similar receptaculum, provided with a similar remark-

536

W. F. PUFCELL.

able keel and pair of muscles (6g. 3). Bertkau (’78) pointed
out the similarity between Oonops and Harpactes as
regards their female sexual organs (p. 371), but he does not
describe or figure the keel and muscles (’78, pi. xii, fig. 8).
In Segestria and Dysdera the receptaculum seems to be
differently formed. (See Bertkau, ’75, pi. vii, fig. 12, and
’78, pi. xii, fig. 6.)

The lung-book in the Oonopid which I examined has
nearly twenty leaves. Its ante-chamber differs from that of
the Dysderidas and has the normal shape found in many
other dipneumonous spiders, that is to say, it rises vertically
from its pedicel but soon curves gradually forwards to form
a long “ horn,” which is nearly horizontal in its anterior part.
The ante-chamber is densely spined on its posterior wall,
except along its lateral edge interiorly, Avhere a small muscle
(corresponding to No. 11 in text-fig’. 7, p. 530) is attached.

There is a distinct and deep epigastric (interpulmonary)
fold, which ends laterally just behind the medial ends of the
pulmonary spiracles, but is not continuous with them. There
is, therefore, no canal of communication between the lung-
books and no part of the fold is lined with spines (see fig. 2).
As in Dysdera and Segestria, the portion of the fold
between the eutochondrites is much deeper than the portions
which lie laterally to these. The lateral coruers of this
deepened part of the fold form the entapophyses to which
the entochondrites (fig. 1, t. 8) of the ventral longitudinal
muscles are attached. These entapophyses are somewhat
unusual in form, their deeper part forming a solid, darkly
staining plate (fig. 1, ec. t. 8), the anterior face of which
serves for the attachment of the obliquely transverse muscle
{■m. 38) connected with the keel of the receptaculum seminis,
while the entochondrite {t. 8) is attached to the upper lateral
edge of the plate.

Caponiidse. — This small but very interesting family bears
some external resemblance to the Dysderidse and Oono-
pi dae but it differs from these and, so far as I know, from all
other spiders as well, in four unique and remarkable anato-

THE PHYTOGENY OF THE TEACHE.E JN ARANE.E. 537

mical characters connected with the respiratory segments.
These are (1) the presence of an anterior pair of trachem in
place of the lung-books (apneumonous spiders), (2) the pre-
sence of a peculiar sense-organ within the second pair of
ti’achese, (3) the absence of the segments (corresponding to
15 and 18, text-fig. 6) of the great, ventral, longitudinal
muscles, so conspicuous in other spiders, belonging to somites
9 and 10 and of the entochondrites connected with them, and
(4) the replacement in the female of the usual receptacula
seminis of the epigastric fold by a pair of great chamber-like
dilations of the oviducts in the upper anterior part of the
abdomen.

The trachea} of Caponia and Nops have been very well
described and figured in Simon (’93, pp. 326 and 327, figs. 294
and 295) from drawings made by Bertkau (also reproduced by
Lamy [:02, p. 184, figs. 24 and 25]). The following descrip-
tion was made from a number of sections and other prepara-
tions of Caponia spiralifera. Pure., specimens of which
Avere collected for me at Hanover, Cape Colony, and well
preserved in spirits by my friend, Mr. S. C. Cronwright
Schreiner.

The anterior pair of tracheae (p. 545, text-fig. 17, and
fig. 9, a.tr.) are situated in precisely the same place which is
occupied by the lung-books in dipneurnonous spiders, and
they are evidently merely a pair of lung-books of which the
saccules have been metamorphosed into branched tracheal
tubules. Fig. 10 shows one of these trachem, prepared in
caustic potash and seen from the medial side. Figs. 11 and
12 are from sagittal sections.

The spineless, thick-Avalled pedicel (fig. 11, ped.), which is
continuous with the adjacent epigastric fold and much re-
sembles it in sagittal sections, leads from the spiracle
into an ante-chamber {tr.a.). The latter is shaped much like
that of the lung-book of a Dysderid, being strongly inclined
forwards and slightly outAvards, broadest near the base and
tapering towards the higher anterior end or apex (figs. 10 and
12). It is, hoAAever, somewhat more capacious, owing to the

538

w. r. ruRCELi,.

THE PHYLOGENY OF THE TRACHE.E IX ARANE.E. 539

Text-figs. 8 — 15.— Eight consecutive transverse sections through
an anterior trachea of Caponia spiralifera. commencing
with the most posterior one of the eight (see PI. 28, fig. 11).
spi. Anastomosing spines of ante chamber. tr. a. Ante-
chamber. (Tlie lateral side of the trachea is on the right of
each figure.) Magn. 192.

greater elongation of its ventral side. Internally the ante-
chamber is lined with anastomosing sjiines [■'‘pi.), except
laterally near the base of the upper (posterior) wall, where
there is a fold [J'd.) to which a short muscle (No. 11 in text-
fig. 17, p. 545) is attached, exactly as in the Dysderidie.
Nearly all the tracheal tubules, except a medial group of
four, spring from the ascending anterior side of the ante-
chamber and run forwards. They occupy exactly the posi-
tion of the saccules of a lung-book (c.f . text-figs. 7 and 17)
from which they are plaiidy derived. In fact, if the 13 or 14
saccules of the lung-book of a Harpactes (fig. 5) weie each
divided longitudinally into several tubules, Ave should have
the condition found in Caponia. To illustrate this, as well
as the arrangement and origin of the tubules on the anterior
wall of the ante-chamber, 1 have given a series of consecutive
transverse sections (text-figs. 8 — 15), of which text-fig. 8
through the ante-chamber represents the most posterior of
the eight. It will be observed that the tubes are frequently

540

W. F. PUECELL.

somewhat flattened dorso-ventrally at their origiu, only to
divide into a couple or more cylindrical tubes in the next
section, e.g. the lowest tube in text-fig. 9. The tracheal tubes
are not long, being only about three or four times the length
of the ante-chamber, and they do not enter the abdominal
pedicel. All, or nearly all, are several times branched, the
branches becoming slenderer towards the apex where they
frequently end in a short fork (fig. 10). They are either
cylindrical or compressed, being then mostly flattened dorso-
ventrally, like the saccules of lung-books, and are lined inter-
nally, except quite at the base, with a tine, probably spiral,
thickening of the cuticle, just like the tracheal tubules of the
Dy sd erid ae, etc. The bi-nucleate columns of the original sac-
cules have, of course, disappeared. The anastomosing spines
of the ante-chamber enter the bases of the tubes for a
short distance and the free edges of the tubes bordering on
the cavity of the ante-chamber have very much the appear-
ance of those of pulmonary saccules in sagittal sections (cf.
fig. 5 of the lung-book of Harpactes with figs. 11 and 12
of the trachete of Caponia). In a young specimen examined
the tubules were much fewer than in any of the adult speci-
mens, hence it is evident that the}'^ increase in number with
the growth of the spider.

In addition to the above there is a small group of tubules
which start from a slightly projecting portion of the ante-
chamber at its base on the medial side. This group is com-
posed of a bunch of four tubules, which may, however, sub-
divide into several more. They take at first a transverse
medial direction and then bend and run some distance back-
wards on the lateral side of the second pair of trachem
(fig. 9, med. tub.). This group of tubules has no equivalent
in the lung-book of theDysderidm and is to be looked npon
as a new formation. They may, perhaps, be the posterior
group of six tubules represented in one of Bertkau’s figures
(Simon, fig. 294, or Lamy, fig. 25).

The second pair of trachea? of Caponia forms per-
haps the most complete and extensive tracheal system known

THE PHYLOGENY OE THE TEACHEHI IN AEANEiE. 541

in any spider. It furnishes the abdomen, cephalothorax and
appendages Avith a very great number of fine air-tubes, but
only a portion of these are represented in fig. 9 in which the
terminal parts of the numerous bi-anching tubules, often
measuring only 3 fx in diameter, have not been drawn in.

The pedicel of the posterior tracheae opens into a capacious
tracheal trunk (p. 645, text-fig. 16, c.tr.) lined with spines,
which runs forwards at an inclination of about 45° for a very
short distance only before dividing into two sub-cylindrical
branches of equal length and thickness, which may be called
the cephalothoracic trunks (fig. 9, ceplh.tr.), and run
horizontally forwards into the cephalothorax, becoming
thinner anteriorly. They remain in contact one above the
other but the upper one a little more to the lateral side, and
are somewhat flattened along- the place of contact. Soon
after passing through the pedicel of the abdomen they each
break up into a bunch of fine tubules which then spread in
various directions. Those of the right upper trunk are drawn
in fig. 9, from Avhich it will be seen that most of the tubules
of the right trunk cross over to the left side and, generally
remaining unbranched, enter the coxse of the left appen-
dages, whereas only the posterior appendages of the right
side receive tubules from this trunk. Several of the tubules
give off dendritic branches {d.hr.), which ramify Avithin the
cephalothorax and supply its upper part Avith air. While the
inferior cephalothoracic trunks remain unbranched except at
apex, the two upper ones each give off a small branch (fig-. 9,
hr.) from the upper side near the middle. This branch is
arborescent in form and divides into a number of tubules
Avhich supply the region above the anterior pair of tracheae.

In addition to the tAvo anterior trunks each of the short
main trunks also gives off from its posterior side at base a
cylindrical posterior trunk of half the diameter of either of
the anterior ones. This trunk, which may be called the
abdominal trunk, runs first in an upward and medial
direction, and then curves and runs horizontally toAvards the

542

W. F. rUECELL.

posterior part of the body. It is of a pronounced arborescent
form, but only its larger branches are provided with spines.

Tlie spines which line the various trunks within are
arranged in longitudinal rows (fig. 4, S2)i.), and are connected
at apex by transverse threads {thr.), which, however, also
anastomose with adjacent threads, the whole arrangement
being' very much as in Dysdera (see Lamy :02, pp. 180 and
181, figs. 20 and 21). The larger tracheal branches iu the
abdomen have similar spines, but the finer branches or
tubules in the abdomen and all the tubules in the cephalo-
thorax have the transverse threads only, but no spines.

Each of the short main trunks (text-fig. 16, c. tr.) is also
produced laterally, together with the pedicel and spiracle,
beyond the points of origin of the three principal branch
trunks to form a small but very curious, lateral pocket
(fig. 9, 1. p.), indicated by Bertkau in his two figures. This
pocket is divided into a higher posterior and a lower anterior
compartment, each compressed from before and behind. The
posterior compartment (fig. 8, p. c.) is provided with anasto-
mosing spines, directly continuous with those of the main
trunk, along its upper and medial edges only (■spb), the rest
of its surface being spineless but much crumpled. It gives
off tlu'ee small branches, viz. one from the upper edge in an
antero-lateral direction, and one each from the upper lateral
and medial angles. These soon subdivide and end in fine
tubules; they are shown in fig. 9, and have also been indi-
cated in Bertkau’s figures.

The anterior compartment (figs. 6 — 8, a. c.) of the lateral
pocket is lined with anastomosing spines on its anterior side
(fig. 8, tipi.), but the upper part of this side and that of the
posterior side is furnished with short sharp spines, the rest
of the posterior surface being spineless but much crumpled.
From the upper edge of the compartment two (iu the male)
or three (in the female) peculiar stout rods or processes {rd.)
of the cuticula project downwards into the lumen of the
compartment, each being armed at the base with some

THE THYLOGENY OP THE TRACHE.E IX AEAXE.E. 543

minute, sharp, conical spines, and. with some longer ones
towards the apex.

The hypodermis of the upper and posterior sides of the
anterior compartment, and especially that of the anterior
side of the posterior compartment, is much thicker than
elsewhei’e, and its cuticula has a corrugated appearance
in sagittal sections, and stains more deeply than the adjacent
cuticula does (fig. 8). Plainly the whole of this structure
has some function other than respiratory. The hypodermis
bearing the three rods is connected at base by means of a
strand with some cells or fibrous tissue, which may be a
nerve (fig. 8, nv.). As, however, the specimens were not
especially preserved for histologdcal purposes, it is impossible
to say anything definite about the character of these struc-
tures, except that the rods certainly strongly resemble sense
organs.

There are well-developed, transverse, intertracheal folds of
the integument connecting the spiracles of each pair and
already indicated by Bertkau in his figures. That of the
posterior pair (fig. 0, ir.fd.) encloses a spinous canal of com-
munication (p. 545, text-fig. IG, can.), which connects the
lumens of the short main trunks {c.tr.) with one another. In
the anterior segment there is no spinous canal of communica-
tion, although the lateral parts of the fold are directly con-
tinuous with the pedicels of the anterior pair of tracliete.

'J'he well-developed anterior (epigastric) fold (text-figs. 16
and 18, ep.fd.) is strongly inclined forwards or even hori-
zontal, especially throughout the median half, where the
genital duct opens into its anterior wall. Each lateral fourth
of the fold, lying (in the female) between the opening of the
genital duct and the tracheal pedicels, appears twice bent
(text-fig. 18), first upwards or slightly backwards, and then
more sharply forwards and downwards, the whole of the
anterior deflected portion (1.) serving for the attachment of a
broad and powerful muscle (Xo. 4 in text-figs. 16 and 18).
Near to the trachea the upper part of the fold is somewhat
inflated, and produced upwards to form a conspicuous enta-

544

F. PURCELL.

pophysis (text-fig'. 16 and fig. 8, ec.t. 8) for tlie attachment of
the muscles 1 — 3. The hollow entapophysis of Caponia
much resembles the solid one of the Oonopidge (fig. 1), and
muscle N^o. 4 in the former corresponds exactly as regards
its place of attachment to the muscle 38 in the latter, although
not homologous with it.

The muscular system of the abdomen of Caponia is in
some respects very peculiar. That connected with the respi-
ratory segments (with the exception of the muscles of the
oviducts) is given in text-figs. 16 — 18 and explained in the
following list :

List of the Muscles in Text-figs. 16 — 18.’

1 and 2. From the entapophysis ec.t. 8 to upper and middle
part of side of abdominal pedicel.

3. Longitudinal from the entapophysis ec.t. 8 to the cephalo-
thorax.

4. From lower lobe of the entapophysis ec.t. 8 to ventral
integument of anterior respiratory segment.

5. From upper part of side of abdominal pedicel to ventral
integument of anterior respiratory segment. (1 and 5 are
inserted together anteriorly.)

6. Dorso- ventral on side of pedicel. (2 and 6 are attached
to the same ectodermal infolding on the medial side of 5.)

7. Oblique dorso-ventral from hinder end of the ectodermal
tendon ec.t. to the ventral fold/d. 1 of abdominal pedicel.

8. Oblique doi’so-ventral from hinder end of the ectodermal
tendon ec.t. to the integumental fold fid. 2 on lateral side of
posterior spiracles.

9. From the ectodermal tendon ec.t. to anterior integument
of abdomen.

10. Two muscles from the ectodermal tendon ec.t. to dorsal
integument of abdomen.

11. Short muscle from lateral part of posterior (upper) wall

‘ Throughout this paper homologous muscles ai’e indicated by the
same numbers.

33 iit.i JOCL

54G

W. F. PUECELL.

of anterior tracheal ante-chamber in a dorso-lateral direction
to tlie body integument on the lateral side.

12. Longitudinal parietal along ventral integument of
abdomen.

13. From posterior intertracheal fold to integument of
abdomen (many or most of the strands continuous with 12).

23(7. Longitudinal from x to the spinners, running along-
side of the abdominal tracheal trunk for the greater part of
its course and dividing posteriorly into several muscles, the
lowest of which is attached to the lateral or middle part of
the anterior side of the mesial anterior spinner (thus appa-
7-ently corresponding to 27 or 28 in Segestria).

30. From the apex of the entapophysis pcJ. 8 in a dorso-
lateral direction to integument of abdomen on lateral side of
anterior spiracles (inserted immediately behind 11).

30(7. Longitudinal from the apex of the entapopli3’sis ec.L 8
to X. (See also PI. 28, fig. 8, m. 30 a.)

32. From x to posterior side of posterior intertracheal fold
(inserted at extreme medial end of the spiracle).

33. From x in an antero-dorsal direction to the oviduct,
meeting the latter at the point where the muscle ends in the
figure. (The muscles 23«, 30n, 32, and 33 are continuous
with each other at x on the lateral side of the abdominal
tracheal trunk.)

34. Oblique from medial and posterior side of the common
basal part {c.fr.) of tracheal trunk to anterior intertracheal
fold (inserted on medial side of the entapophysis ec f. 8).

35. From upper edge of posterior intertracheal fold to
base of anterior intertracheal fold (many strands apparently
continuous with 13).

36. Subtransverse from posterior medial edge of anterior
tracheal pedicel (in a transverse line with the cross x in
text-fig. 17) to between the upper and lower cephalothoracic
trunks of the posterior trachea. (See also fig'. 8, m. 36.)

37. From the spineless, basal, posterior part of the second
trachea and its lateral pocket in a dorso-lateral direction to
the integument on the lateral side of the second spiracle

THE PHYTOGENY OP THE TRACHEAE IN ARENEJE. o47

(inserted next to 8, and just beliind but sligditly lower than
dO). (See also fig. 8, m. 37.)

The most remarkable features of tliis muscular system are
the complete absence of all entochondrites, and as well as of
those segments of the great, ventral, longitudinal muscles
which belong- to the second respiratory and the anterior
spinner segments (somites 9 and 10).

The muscles 1 — 10 are identical in the Dysderidm and
Caponiidm, 1 — 4, however, being attached in the latter
directly to the anterior side of the entapophysis {ec.t. 8)
without the interposition of an entochondrite. No. 4 is a
very broad atid powerful muscle, being attached, as already
explained, to the whole anterior side of the lateral deflected
lobe (1.) of the epigastric fold, and is represented in the
IJysderidm by several feeble strands only. The muscles
1 — 3 are attached in Caponia to the prominent lateral
entapophysis (ecJ. 8), which is, therefore, plainly homologous
with the entapophysis of the pulmonary segment of the
Dysde ridm (p. 530, text-fig. G, ec.t. 8) and other dipneu-
monous spiders. It may be noticed that in Caponia this
entapophysis has completely taken the place of the ento-
chondrite (t. 8) of the Dysderidae, its anterior surface being
correspondingly expanded to take the four large muscles.

The three muscles, 8, 11, and 30, which in the Dysde-
ridm are attached to the enchondrite t. (p. 530, text-fig. 7),
are also represented in Caponia, only here the lateral ends
of these muscles are attached separately to the body integu-
ment, and are more dispersed (although still quite close
together) owing to the absence of the entochondrite, and 11
runs parallel to the transverse plane, while in Segestria its
fibres lie in sagittal planes.

The parietal muscle 12 is a part of the abdominal muscular
sac which lies immediately within the outer hypodermis and
envelops the intestines. The ventral strands of the sac are
here longitudinal, and form a continuous layer from side to
side, where they are again continuous with the lateral walls
of the sac. Anteriorly the ventral strands of the sac ascend

VOL. 54, I’ART 4. NEW SERIES. 39

548

\V. F. rUKCELL.

(No. 13) to the upper edge of the posterior intertracheal
fold along its whole ex^^tent, but the descending strands
(No. 35) on the anterior side of this fold are only met with in
the lateral part, being absent from the median part of tlie
fold. A similar parietal muscle is met with in the Dys-
deridm and Oonopidm, differing only in so far that the
lateral ascending’ strands (No. 13, p. 530, text-fig. 6) are
attached to the large entochondrite (/. 9) of the tracheal
segment.^

I could find no trace of the usual medial longitudinal
muscles corresponding to 15 a,nd 18 in the Dysderidte, and
connecting the anterior entapophysis {ec.t. 8) with the spin-
ners on the medial side of the trachem. In fact, the only
muscle connecting- the respiratory segments with the spin-
ners, and lying inside of the abdominal muscular sac, is the
slender muscle 23n, which, however, lies on the lateral side
of the trachea, and is, I think, probably homologous with
muscle 23 of the Dysderidae. This little muscle in
Cap on i a divides posteriorly into at least two muscles and
the most ventral of these branches,” corresponding to 27 or
28 of Segestria, is attached to the lateral (S') or middle
( ?) part of the anterior side of the mesial anterior spinners,
which I take to represent the anterior pair of the Dysde-
ridm. Anterioidy 23a unites with three other small muscles,
30a, 32, and 33, at a ])oint .r at the base of the lateral side of
the abdominal tracheal trunk (ahd.tr.). The four muscles
are here in contact with the trachea, and their fibres inter-
mingle without forming an entochondrite. One of them,
30a, passes on to the apex of the entapophysis (ec.t. 8), and

’ The alxloniinal muscular sac in other spiders has Ijeen descril)ed
l>y various authors, particuhirly l)y Causard ('96, pp. 22-24, pi. iii, figs.
1 and 2). and more recently liy Lamy (: 02, p. 158, pi. vii). Of the
muscular nature of its fibres there can be no doubt whatever, as the
typical transverse striations may he frequently observed in the
Oaponiida“. the Dysderida, and other forms.

- I could not make out where the posterior ends of the dorsal branch
or branches were attached.

THE I’HYLOGEXY OF THE TRACHE.E IX AFvAXE.E. 549

may possibly represent a strand which became separated
from 30 when the lateral entochondrite (p. 530, text-fig. 7, t.)
disappeared.

From a comparison with the Dysderidm it is evident
that the trachece of the second respiratory segment in
Caponia correspond to those of the Dysderidte, and, like
the latter, are to be considered as entirely homo-
logous with lung-books, for there is no evidence that
entapophyses took any part in their formation.

Tlie receptacula seminis are paired, and consist of an
enormous dilation of each oviduct apparently at the point
where the ectodermal and mesodermal elements of the duct
meet, Tliey are placed nearer the upper part of the abdomen
just above the area enclosed between the four spiracles, but
extend for some distance to the front and behind this area as
well. The ventral wall of each dilation has a cuticular
lining, but the dorsal wall and the greater part of the side
walls have none. Apparently the latter represent the meso-
dermal part of the oviduct and the former the ectodermal
part. The ventral wall with its cuticula is continuous with
the basal portions of the oviducts, which open into the lateral
ends of the unpaired median portion of the duct. This
latter again opens into the epigastric fold along a wide cleft
occupying about one half of the distance between the two
anterior trachem. The spacious lumen of each receptaculum
contains coagulated stainable matter and mimerous sperma-
tophors. Tills form of receptacula is apparently quite unique
amongst spiders, the usual ones which open directly into the
epigastric fold, and are evidently invaginations of the body
integument, being quite absent in Caponia.

Ckxeral Cox'clusioxs.

A tracheal system may be imagined to have been evolved
out of a lung-book in either of the following ways :

(1) The pulmonary saccules may have been converted into

550

W. F. rUECELL.

more or less cylindrical tubes (say, by longitudinal division),
accompanied by tlie disappearance of the bicellular columns
of the septa. The trachea thus produced would be composed
of an ante-chamber formed out of the pulmonary ante-
chamber, with a bunch of tubules on its anterior surface,
formed out of metamorphosed pulmonary saccules. This
metamorphosis does not involve a reduction in the effective-
ness of the respiratory organ, and I imagine the trachea so
produced to be in no way inferior, but rather superior, to the
lung-book. Accoi-dingly it would doubtless increase in size
and tfike over the mnin respiratory functions, and this
would be accompanied, in the case of the second respiratory
segment, by a corresponding reduction in the number of the
leaves of the anterior lung-books, in accordance with Lamy’s
law of the inverse correlation between the size of the tracheae
and the number of the lutig-leaves.

(2) The saccules may have disappeared, leaving only the
pulmonary sac or ante-chamber, which would then constitute
a trachea, and may subsequently elongate or even acquire
secondary branches. These latter, however, would not be
homologous with pulmonary saccules. This method of origin
really consists in a reduction in the effectiveness of tiie
respiratory organ of the somite, and would be accompanied,
in the case of the second respiratory segment, by- a corre-
sponding increase in the number of the leaves of the anterior
lung-books, which would then become the principal organ of
respiration. This, then, would be exactly the opposite pro-
cess to that which would have taken place in the first case.

It appears to me very probable that both these methods of
origin have actually occurred in the Aranem, the first method
being applicable to the Dysderidte and their allies, and the
second to the rest of the tracheate spiders. It will be con-
venient to take these two sections of spiders in turn.

(1) The Dysderidm, Oonopidm, and Caponiidm,
being those forms with the tracheal spiracles far
apart and not moved backwards, i. e. still nearly
in their primitive positions. — If, after the metamor-

THE PHYLOGENY OF THE TRACHEAE IN ARANE.E.

551

phosis of tlie saccules into the tubules in the case of the first
of the two methods given above, we further imagined the
ventral part of tlie ante-chamber to lengthen slightly, we
should have almost exactly the condition found in the ante-
rior pair of tracheae of Capon ia, which differs from this ideal
case only in two minor points, viz. in the pi-esence of trans-
verse or spiral thickenings in the tubules instead of small
spines, and of a medial group of tubules at the base of the
ante-chamber. In fact, as I have shown above, the anterior
pair of tracheae of Capon ia may be taken to represent the
most primitive form of metamorphosed lung-books known in
Avhich the saccules still persist as tubules.

Diagram of a tracliea of Harpactes Hoinl)ergi (ad. J*), seen
ill section. Magn. H14. ahd. ir. Alidomiiial branch of the
trachea. ceph. tr. Ceplialothoracic trunk. s})". Siiiracle.
spt. Anastoinosing spines, tub. Tubules.

The second pair of tracheae in Capouiai, being wholly
homologous with lung-books, plainly belong to the same type
as the tracheae of the Dysderidac and Oonopidae, and are
merel}^ somewhat more complicated by the duplication of
eaich of the cephailothoracic trunks and the elongation of the
aibdominal branch of the hitter. The simplest form of this
tyjie, such as that found in Harpactes (text-fig. 19) and in
Calculus (in both of which the anastomosing spines of the
trunks still form ai simple network and do not bear a spiral
thread or inner perforated tube), niaiy be easily derived from
the anterior tracheae of Caponia by merely exaggerating the
tubular elongation of the ante-chamber, already commenced

552

W. F. PUItCELL.

in the anterior tracheee, and. by the addition of the short
posterior branch in place of the medial basal group of tubules.
In such case the bunch of tubules at the anterior end of the
cephalothoracic trunks {ceplb.tr.) would represent metamor-
phosed pulmonary saccules, but those of the posterior abdo-
minal branch {ahd.tr.) would be, of course, new formations.

In a previous paper (:09) 1 had already indicated the
possibility of the anterior bunch of tubules being derived
from saccules, but after studying the tracheae of Capouia
more thoroughly, I am now much more strongly inclined to
believe that such has actuall}" been their origin. A study of
the embryology would, however, be necessary to settle this
interesting point.

If, as I have assumed, the posterior pair of trachete in
Capouia aud those in the Dy sderidie had a common origin,
it follows that the auteilor pair of trachea) in the former must
have developed later and independently of the posterior pair,
and that, therefore, trachea) must have originated from
lung-books at least twice in the Aranem. The same
conclusion would follow even if we assumed that both pairs
of trachea) iu Capouia originated at the same time and not
as separate metamorphoses, for in that case both pairs of
trachem must have originated independently of those of other
tracheate spiders, since these latter still possess the anterior
pair of lung-books.

The mor])hology of the respiratory segments bears out the
view that the three families discussed above are intermediate
in position between the mygalomorphous spiders and the rest
of the arachnomorphous forms. This view was demonstrated
by Bertkau (’78) a good while ago for the Dysderidm and
the Oonopidae, and this author even went so far as to
include these families Avith the mygalomorphous forms in a
common group, the Tetrasticta (i. e. Avith four stigmata).
No doubt these tAvo families are the most primitive of the
three, but the Caponiidae may be considered as an allied
but in several respects a very aberrant t} pe, standing apart
from the other tAvo families.

THE PHYLOGEXY OF THE TRACHE.E IN APvAXE-E. 553

(2) Forms with the tracheal spiracles approxi-
mated and moved more or less toward the hinder
end of the body. — All the remaining ti*acheate spiders
come under this heading,^ and may be considered in two
groups, viz. group A, those in which the entapophyses of
the tracheal system are non-respiratory (Filistatidas,
Sicariida?, and Palpiinanidfe), and, gToup B, those in

Traubeal apparatus of Filistata capitata (after Lamy). ec. t. 9.
X' on-respiratory eiitapopliysis. 1. tr. Lateral or ti-aelieal sac.
Magn. lUO.

which tliese entapophyses have been transformed into
trachete (including all the remaining families).

A very simple and interesting type of trachea3 of the first
group is that of Filistata (text-fig. 20), which is known to
us from Lamy’s description.

The simplest and most usual type of the second group, a

* In some cases, e. g. Argyroneta, tlie common tracheal spiracle
ap2iears to have secondarily moved forward again.

554

AV. F. rUFCFLL.

type found, accordiug to Lamy, in about half of the total
number of genera examined, consists of four simple tracheal
trunks united at base, as in text-fig. 21. It is known from
the embryology that the lateral trunks of this type were
derived from the pulmonary sac or ante-chamber of a lung-
book, and are, therefore, homologous with the lateral trunks

Traolieal apparatus of Liiiypliia triangularis. Cl. (act. 5-
caustic potash). Magn. 80. can. Canal of coniinunication
between the tracheal trunks, hy'. Terminal chitinous filires
T)y which the medial or tendinal trunks (ni. tr.) ai'e attached to
the entochondrites. I. tr. Basal portion of lateral trunk.

Pedicel of trachea, rd. Lateral supporting rod. .sp".
Spiracle.

in Filistata, — while the medial trunks represent meta-
morphosed entapophyses (ectodermal tendons of muscles),
and are, as Lamy (p. 172) has pointed out, homologous "with

THE PHYLOCiENY OF THE TPACHE.E IN APANIOE

555

the medial trunks of Filistata^ -wliicli this author has shown
to be entapophyses.

In both groups more complicated types than the two simple
ones just described are frequently found, and it is important
to notice that this complication takes place along different
lines in each group. Thus in group A some Sicariidae and
Palpimanidae were found by Lamy (p. 176, fig. 15, and
p. 188, fig. 30) to possess branched tracheae, the branching
being confined to the lateral or tracheal trunks. In group
B, on the other hand, in all cases where the lateral and
medial trunks can be identified from Latny’s figures and
show different degrees of development, it is invariably the
medial trunks which show the greatest complexity and the
highest degree of development as respiratory organs.^ This
rule appears to me to furnish the key to the phytogeny of
the tracheae in these spiders. We may also fairly deduce
fi’oni it that the medial tracheae must be more efficient as
respiratory organs than the lateral tracheae are, and the
reason for this, as I have already pointed out (:09), may
be their position in the large ventral sinus containing venous

' This is self-evident from Lamy's excellent figures in many cases
e. g. CEcobiidie (Lamy, p. 170, fig. 10). Argiojjida; (p)). 107 — 100, figs.
38 — 1'2), TliomisidiK (pp. 20f)anJ207, figs. 40 and oO), and Agelenida;
(pp. 214 — 216, figs. 59 — 61). In arborescent types of tracheae (see my
paper, :09) it is not so self-evident, but the same conclusion may
Ije deduced from the great similarity which this form of trachea shows
to that of the Attidai, of which the identity of the parts is known from
the embryology. There remain, however, certain Dictynidae and
Agelenidse, the homology of whose trachea} cannot be ascertained with
any degree of certainty from Lamy’s figures. In Argyroneta 1 found
(:09), from the position of the imiscles and entochondrites, that the
entire trachea appears to have been derived from the medial trunks, but
1 have had no oj^portunity of examining any of the other forms, viz.
Dictyna (Lamy, p. 169, fig. 8), Antistea (p. 213, fig. 57), Cybaeus
(p. 217, fig. 62), and Chorizomma (p. 219, fig. 64). If these, too. coidd be
l)i'Oved to follow the rule given above, the arguments in the following-
pages would be greatly strengthened. I may add here that in the
marine Agelenid, Desis tubicola, Poc., the trachea}, which have not
been hitherto described, closely resemble those of Attus.

556

AV. F. PURCELL.

blood. This greater efficiency would account for the liiglier
de gree of development of tlie medial trunks in many forms.

Now out of the twenty-four families in which the medial
entapophyses have been converted into trachemg twenty-two,
according to Lamy’s investigations, possess tracheal systems
consisting of four simple tubes (p. 554, text-Kg. 21) in some
of their genera at least, while eight of these families possess
both this simple type and more complicated types as well.
In fact, only two very small families (G*lcobiida3 and Pro-
didomida3) have the more complicated type ouly. And
since the type with branched medial trunks must have been
derived from the type with simple trunks, as the medial ones
were originally simple entapophyses, we may fairly conclude
that the comtnou type with four simple tubes is the primitive
one for the entire group, and that the more complicated
types must have been developed from the simpler types
within each family separately and independently of similar
complicated types in other families. This statement is in
agreement with Lamy’s view referred to in the introduction,
except that this author does not consider any particular type
as more primitive than another.

Again, it is evident that the type of trachea in which the
entapophyses are not respiratory must be considered as more
primitive than those in which they are respiratory, since the
more efficient medial tracheal trunks would not be likely to
revert to their original function after once being metamor-
phosed. Hence the trachea? of the Fi listatida?, Sicar-
iidte, and Palpimanidm must be looked upon as more
primitive than those of group B with metamorphosed medial
trunks, and it seems to me very probable that the trachea? of
the latter group were originally derived from some such form
as that found in Filistata (p. 553, text-tig. 20). In this
spider the trachete are placed, according to Lamy (;02, p. 172,
tig. 11), about midway between the spinners and the inter-
pulmonary fold. The anterior end of each of the tracheal
entapoj)hyses is situated near this fold, and consequently the
segments of the longitudinal muscles between the entochon-

THE PHYLOHENY OP THE TEACHE.E IN AEANE.E. 557

drites of tlie pulmonary and tracheal segments are doubtless
quite short. If now the tracheal spiracle moved to the
hinder end of the body and the entapophyses elongated
correspondingly and became converted into a trachea we
should get the type represented in text-fig. 21 (p. 554), which
I consider to be the primitive type of all forms with meta-
morphosed entapophyses. The anterior ends of the entapo-
physes would still be near the iuterpulmonary fold, and the
connecting muscular segment would still be quite short, as it
always is in the spiders of group B.

In Scytodes and Palpimanus the spiracle has moved
to the hinder part of the body without any additional
lengthening of the entapophysis. Hence in these two forms
the segments of the longitudinal muscles belonging to the
tracheal somite are greatly elongated, and iu this respect
these forms (and allied genera) are apparently unique.

Tlie Pholcidm, which Bertkau found to have no trachem
at all, were perhaps derived from some form with a type of
trachea similar to that of Pilistata, since according to
Ijamy’s investigations a pair of entapophyses persists in some
Pholcidse iu the same position iu which those of Pilistata
are found (:02, pp. 191 and 192, figs. 32 and 33). The
IMiolcidte, therefore, should perhaps belong, as regards the
structure of their ninth somite (the tracheal segment iu
other spiders), to the same group as the Pilistatidm,
S i c a r i i d to, and P a 1 ]) i m a n i d m.

In a previous paragraph two ])ossible solutions were
suggested for the derivation of a tracheal system from lung-
books, one of which appeared particularly applicable to the
tracheal .system of the Hysderidm, etc. Now the second
method suggested, which consists in the reduction of the
respiratory functions of the lung-books by the abortion of
the .saccules, appears to me to exactly meet the conditions
found in the spiders with four simple tracheal trunks (or
Avith two tracheal trunks and two entapophyses), iu Avhich
the lung-books have numerous leaves, and obviously play the
most important part iu the respiration. The size of the

5oS

W. F. PURCELL.

lung-books in FilisLata is not known, but judging from
Lamy’s figure (:02, p. 172, fig. 11) they appear here, too, to
be very large to compensate for the feeble development of
the trachem. This relatively greater size of the anterior
lung-books is exactly what 1 have explained should take
place if the posterior lung-books became reduced to their
ante-chambers only.

If we imagined a tetrapneumonous spider with both
pairs of lung-books connected by interpulmonary folds (the
arachnomorphous spider Hypochilus appears to be such a
form), and the entapophyses prominently developed in the
second respiratory segment, as well as in the first, it would
be perfectly simple to derive from it a form with ti-acheae
exactly resembling those of Filistata. All that would be
necessary would be that the saccules of the second pair of
lung-books should disappear, leaving the two ante-chambers
only; and that the two spiracles should come a little nearer
together so as to form practically one opening with the
intertracheal fold. It appears to me very probable that the
tracheae of Filistata and of all other spiders (except the
Dysderidm and tlieir allies) had this mode of origin, which
is in entire agreement with the account given by Lamy of
the structure of the tracheae in Filistata. The two short
lateral tracheal sacs (p. 553, text-fig. 20, l.tr.) of this form
are lined with spines and triangular in shape, exactly resem-
bling a pulmonary sac deprived of its saccules. A study of
the Hypochilidm would probably throw some further light
upon this subject, since here the second pair of lung-books
are placed, according to Simon’s figure ('Hist. Araign.,’
2nd ed., i, p. 201, fig. 145), far back, about midway between
the anterior pair and the spinners, corresponding exactly in
position to the tracheal system of the Filistatid le.

I have made no attempt to explain the origin of those
tracheal tubules, which cannot by any line of argument be
derived from pulmonary saccules. The numerous tubules
emitted from the large tracheal trunks in the Attidrn are a
case in point, since these trunks, with the exception of their

THE PHYLOCxENY OP THE TEACHER IN AEANE^. 559

lateral basal lobes, are metamorphosed entapophyses. These
tubules may have originated simply as outgrowths of the
trunks, and would then, of course, be of ectodermal origiia.
Ray Lankester is of opinion (:04, p. 223) that the tracheal
tubules in Arachnida (and in all other Tracheata) have
developed by adaptation of the vasifactive tissue of the
blood-vessels,” which have come to open in the case of the
Araclinids into the lung-chambers (and elsewhere). Instances
of mesodermal tubes attaching themselves to, and opening
into ectodermal invaginations are, of course, well known, e. g.
the genital ducts. No actual embryological observations,
however, exist, so far as I am aware, regarding the develop-
ment of the fine tracheal tubules in Arachnida. In Attus
floricola no trace of these tubules was found up to the stage
formed at the second moult, and I had no later stages at my
disposal.^

Summary. — The theoretical suggestions in the preceding
paragraphs may be summed up as follows :

In the first place I suppose the saccules of the second pair
of lung-books to have been converted into tracheal tubules in
the common ancestor of the Dysderidse, Oonopidse, and
Caponiidae. The resultant trachem then increased in size,
and, as the number of the leavms of the anterior lung-
books decreased in inverse ratio, the former became the
principal organs of respiration. The second pair of spiracles
retained their position, or may even have moved slightly for-
wards, and the conversion of the entapophyses into tracheae
could not take place here, and would, moreover, be quite
unnecessary. In the Caponiidae the anterior pair of lung-
books were converted into tracheae in ai similar manner, but
at a. later period, and independently of the conversion of the
posterior pair; but as the latter already provided almost the

' A paper by R. Janeck entitled “ Entwickluiig der Blattertraclieen
and der Traclieen hei den Splnnen” lias recently a2ipeared ("Jena
Zeitsclir. Naturw..’ xliv, Hft. 2 — 1, 1909), hut I have not hitherto had
access to this iiublication.

560

W. F. PURCELL.

entire body witli tracheoe, the anterior pair did not further
increase in size.

In the second place, in the progenitor (or progenitors) of
the remaining tracheate spiders, the posterior lung-books
became reduced in size and eifectiveness by the disappear-
ance of their saccules, accompanied by an increase* in the
number of the leaves of the anterior lung-books. Further,
the posterior spiracles became approximated and united to a
single spiracle, and moved towards the hinder end of the
body, thereby causing the entapophyses of the tracheal
segment to elongate. In this condition the Filistatidae,
8 icariid U3, and Palpimanida) have remained, with slight
modifications, such as the division of the tracheal ante-
chambers into branches in some forms. In the great majority
of the families, however, the elongated entapophyses became
transformed into a pair of medial tracheal trunks, thus pro-
ducing a tracheal system consisting of four simple nnbranched
trunks, which is still found in some genera at least, in nearly
all the families. A new factor having been introduced, viz.
the presence of the respiratory entapophyses lying in the
large ventral sinus containing venous blood requiring aeration,
we accordingly find the second respiratory segment again
taking a prominent part in the respiration in many forms,
owinar to the increase in size and the braiichinof of the medial
trunks, accompanied ultimately by a corresponding reduction
in the size of the anterior lung-books, e. g. in the Attidm.
This method of origin of the tracheaa is independent of that
of the Dysderida; and its allies, and the tracheal tubules,
when present, would here not be derived from saccules, but
be new formations.

List of Literatoke.

’72. Bertkau, P. — “ Ueher die Respirationsorgane der Araneen,” ‘ Arch,
f. Naturg.,’ xxxvdii, Bd. i, pp. 203-2.33, 1 PL, 1872.

“Ueher den Generationsapparat der Araneulen," ‘Arch. f.

Naturg.,’ xli, Bd. i, pp. 23.j-2(52, PI. vii, 187-5.

’75.

THE PHYLOGEXY OF THE TliACHE.E IX AEAXEHE 561

’78. “ Vei’sucli einer natiivliclien Aiiorilmmg dei- Spinnen iiebst

Bemerkimgen zu einzelnen Gattnngen," ‘ Arch. f. Natiu’g.,’ xliv,
Bd. i. pp. .351-410, PI. xii, 1878.

’98. Caiisard, M. — “ Recherches sur Fappareil circulatoii-e des Ara-
neides,” ‘ Bull. Sc. France Belg.,’ xxix, pjD. 1-109, Pis. i-vi, 1896.

:02. Laniy, E. — “Recherches anatomiques sur les Trachees des
Araignees,” ‘ Ann. Sci. Nat. Zool.,' (8) xv, pp. 119-280, 71 figs..
Pis. v-viii, 1902.

:04. Lankester, E. Ray. — “ The Structure and Classification of the
Arachnida," ‘ Quart. Journ. Micr. Sci.,' (2) xlviii, pp. 165-269.
78 figs., 1904 (a reprint of the article “ Arachnida," ‘ Encyclo-
piedia Britannica,' 10th ed., xxv, 1902).

’92. Pocock, R. I. — “Liphistius and its bearing upon the Classifica-
tion of Spiders." ‘Ann. Mag. N. H.,‘ (6) x, pp. 306-314, 2 figs.,
1892.

’95. Pui-cell, W. F. — “ Note on the Development of the Lungs, Entapo-
l^liyses, Tracheae, and Genital D^icts in Spiders,” ‘ Zool. Anz..’
xviii. pp>. .396-40(1, 2 figs.. 1895.

:09. “ Development and Origin of the Respiratoiy Organs in

Araneae," ‘Quart. .Tourn. Micr. Sci.,' (2), liv, pp. l-llO, Pis.
1-7, and 7 text-figs.. 1909.

’93. Simon. E. — ‘Histoire Naturelle des Araignees,’ 2nd ed.. i. pp. 257-
488, Paris. 1893.

’89. Tarnani, .1. K. — ‘"Die Genitalorgane der Thelyphonus,” ‘Biol.
Centi-albl..' ix, no. 12. pp. 376-382, 5 figs.. 1889.

KXl’LANATION OF VLA.TE 28,

Illustrating’ Mr. W. F. Purcell’s paper on “The Pliylogeny
of the Tracheae in A ranea'.”

Abbreviations.

fi. c. Anterior compai-tment of lateral pocket of trachea, a. tr.
Anterior trachea, alxl. tv. Abdominal trunk of trachea, ant. Anterior
side. hit. c. Blood corpuscles, hr. Branch of trachea, can. Canal of
communication between the tracheal trunks, cejjh. tr. Cephalothoracic
trunks of trachea, cii. Cuticula. d. hr. Dendritic tracheal branches.

562

AV. F. PURCELL.

dors. Dorsal side. ec. t. 8. Ectodermal tendon (entapophysis) of the
anterior respiratory segment. ep. fd. Epigastric fold (along hinder
margin of anterior respiratory segment), fd. Fold in doi'sal wall of
tracheal ante-chamher, to the lateral part of which the muscle 11 (text-
fig. 17) is attached. <). o. Opening of the genital duct into the epi-
gastric fold. hy. Hypoderniis. I'. Median keel on ventral side of
receptacnlum seminis. 1. p. Lateral pocket of trachea, lat. Lateral
side. m. 1-37. Muscles (see numbered list in text). m. 38. Ob-
liquely transverse muscle from the anterior sui-face of the entapo-
physis. ec. t. 8 to the sides of the keel of the receptacnlum seminis.
m. 39. Short muscle from the entochondrite t. 8 in a medio-ventral
direction to posterior side of epigastric fold. m. 40. Median muscle
from venti'al side of i-eceijtacnlum seminis to ventral integument of
hodv. running along posterior ventral edge of the keel of the recep-
tacnlnm. med. Medial side. med. tub. Medial hunch of four tubules
at base of anterior trachea, nv. Nerve? p. c. Posterior compartment
of lateral pocket of trachea. p>ed. Pedicel. post. Posterior side.
pithii. (I. Pulmonary ante-chamber, r. .s. Receptacnlum seminis. rd.
Sensory rods in trachea, sept. Septa (lamellae) of lung-book. sp'.
Spiracle of anterior respiratory segment, sp". Spiracle of posterior
respiratory segment, spi. Anastomosing spines, spi'. Points at which
the tracheal spines are attached, .spm. Sperniatophors. t. 8. Ento-
chondrite at posterior end of the segment of the ventral longitudinal
muscle of first respiratory somite. thr. Transverse anastomosing
threads Ijorne liy Uie ti-acheal spines, tr. a. Tracheal ante-chamber.
tr. fd. Intertracheal fold along hinder margin of second respiratory
segment, tab. Tracheal tubules, reat. Ventral side.

All the figures, except fig. 9, were drawn with the aid of a drawing
apparatus. Transverse and sagittal sections are so arranged that
the horizontal plane of the body is parallel to the lower edge of the
papei'.

Fig. 1. — (Zeiss, objective C. ocular II, spirits.) Calculus bicolor,
adult $ . Sagittal section through the entapophysis of the inilmonary
segment.

Fig. 2. — (Zeiss, C, II. spirits.) Median .section through the recep-
taculum seminis and ejiigastric fold, from the same series as fig. 1.

Fig. 3. — (Zeiss, C, II. warm Flemming's solution + alcohol.) Har-
pactes Hombergi. ad. $ . Similar .section to fig. 2.

Fig. 4.— (Zeiss. yV oil iminers., IV.) Caponia spiralifera. Internal
chitinous threads of cephalotlioracic tracheal trunks.

THE PHYLOGENY OF THE TRACHEH] IN AEANEiE. 563

Fig. 5. — (Zeiss, C, IV, warm Flemming's solution + alcohol.) Har-
l^actes Hombergi, ad. $ . Sagittal section through a lung-book.

Fig. 6. — (Zeiss, A, IV, spirits.) Caponia spiralifera, ad. $ .
Transverse section thi-ough the basal part of the trachea of the second
resiiiratory segment.

Fig. 7. — (Zeiss, C, IV, spirits.) Lateral part of fig. 6.

Fig. 8. — (Zeiss, C, IV, spirits.) Caponia spiralifera, ad. ?.
Sagittal section through the entapophysis of the first and the lateral
tracheal pocket of the second respiratory segments.

Fig. 9. — (Caustic potash.) Caponia spiralifera, ad. showing
tracheal system (the ends of the tubules of the ti'acheaj of the second
resiiiratory segment are not drawn in).

Fig. 10. — (Zeiss, A, IV, caustic potash.) Caponia spiralifera,
ad. $ . Right anterior trachea from the medial side.

Figs. 11 and 12 (Zeiss, C, IV, spirits.) Same series as fig. 8. Sagittal
sections through the left and right anterior trachea? respectively. In
fig. 12 the cuticula of the posterior side and the hypodermis of both
sides of tlie pedicel have not been di-awn in.

VOL. 45, PART 4. — NEW SERIES.

40

ERRATi\.

5f)4

Errata to Mr. W. F. Purcell’s Paper, “Develop-
ment and Origin of the Respiratory Organs
in Aranese.”

Pul)lished in the Quart. Jonrn. Micv. Sci.. vol. 54, Part 1,
September, 1909.

Page 2, line nineteen from top, for Scytodidai read Sicariidse.
Page 26, line five from top, for pp. 17-20 read p. 24.

Page 43, line twenty-one from top, for 49 read 35.

Page 54, line seven from top, for 20 read 46.

Page 59, line three from top, for left read right.

Page 71, line nine from top, for Scytodidas read Sicariidte.

Page 79, line four from hottom, for 33 read 40.

Page 82, line sixteen from bottom, for 17-44 read 25-28.

Page 103, line nineteen from bottom, for yellow read grey.

Page 105, line three from top, for anterior and posterior read
ventral and dorsal.

Page 106, line thirteen from bottom, for anterior read posterior.
Page 108, line two from top, for 22 a read 23a. Line eleven from
top, strikeout Adult or snb-adiilt spiders.

Plate 1, figs. 6 and 6a, for ab. opp. 8-11 read ah. app. 1-4.

^a/rt.^oumv Jl/liiyr" Sau'//o^.54'MS^ii ci H.

\k

12. *1“

sl.’f

I

WF. Puroell i«l,

HuUi,Iuthr London

KEPliODUCTIOX OF KALPIDORH YXCHUS ARENICOL^. 565

On the Reproduction of Kalpidorhynchus
arenicolae (Cnghm.).

By

i^[arg:ai'ct Roltiiisoii,

University College, London.

With Plate 29.

Introduction.

In 1907 Mr. Cnnniiigliam described and gave a life-
liistory of this gregarine in the ^ Archiv fiir Protistenkunde.’
The parasite was first noticed by Mr. De Morgan while dis-
secting some .specimens of Arenicola ecandata in this
laboratory. Owing to pressure of other work Mr. Cunning-
ham was unable to give a complete account of the repro-
duction, and he therefore suggested that I should, at some
future time, try to find the first division nucleus with a view
to ascertaining where the chromatin of its chromosomes
came from. The latter half of this problem still remains
unsolved; but after cutting many cysts into sections I did
find the first spindle in a very early state, and I have been
able to make one or two other observations which may prove
to be not without interest.

]\lETnODS.

The cysts Avere fixed Avith various fluids, Hermann’s,
Flemming’s, corrosive sublimate and acetic acid, Avith and
Avithout the addition of formaline, Bonin’s picro-formol and
Brasil’s picro-formol. The best results Avere obtained by
using the picro-formol mixtures. The sections Avere stained

566

MARGARET ROBINSON.

by Heidenliaiu’s method. Following the directions given by
Brasil (1905), I left them for twenty-fonr hours in the
mordant, and for thirty-six or more in the hyematoxylin. As
plasma stains after Heidenhain I used a mixture of Licht-
Gri'm and picric acid in equal parts dissolved in absolute
alcohol, orange G., and eosin.

Two other stains of which I made use were Delaheld’s
hfematoxylin and that of Kleinenberg.

The First Spindle.

The nuclear membrane disappears gradually bit by bit,
and the nucleus becomes more or less diffuse, assumino- an
irregular outline in the parts no longer contained by the
membrane. Close to this uneven edge there appears a little
cluster of rod-like pieces of very darkly staining chromatin.
'I'liese small rods are connected together by a tine thread,
which stains more faintly than they do, so as to form a small,
loosely folded skein. Part of this skein rests on the achro-
matic fibres of the spindle. These fibres are doubtless intra-
nuclear in origin. The spindle has centrioles which stain
deeply with Heidenhain’s haematoxylin ; and there are the
usual terminal radiations in the cytoplasm (fig. 1).

In my next stage, which follows very closely upon the
above, the nucleus is almost completely dissolved, all that
remains of it being two or three karyosomes still uuabsorbed
by the cytoplasm. The spindle is now in anaphase (fig. 2).
It shows a good number of deflected radiations, in this much
resembling one figured by Brasil (1905), but hei’e one can
see a well-marked centrosome. I could not demonstrate a
centriole. 'Hie number of the chromosomes is four. In
metaphase (fig. 3) it is impossible to count the clu'omosomes,
and there appear to be more than four, but in the anaphase
they can generally be counted, and as I have seen numbers of
nuclei in this phase I am in no doubt as to the number.

This appearance of the first spindle agrees in all essentials
with the events described by Cuenot (1900), Brasil and

EEPROBUCTION OF KALPIDORHYNCHUS ARENICOL/E. 567

others in the formation of the first spindle in the Gregarines,
but most closely with the facts in Monocystis ascidice
(R. Lank.)j Siedlecki (1900), and those in G. ovata as
described by Schnitzler (1905), for here there is no vesicle.

The divisions proceed very rapidly; still, I was fortunate
enough to find a cyst with only two nuclei, both, however,
dividing (fig. 3).

In a stage with a small number of nuclei it could be seen
that in the late anaphase the spindles stretch out, and become,
consequently, very much attenuated in the middle. Thus the
two daughter-nuclei arising from a division seem, at any
rate during the rearrangement of the chromatin, to be
surrounded and supported by the wide ends of the spindle
of the mother-nucleus (figs. 4 a and h). Siedlecki (1900) and
Brasil record the same method of division in Monocystids,
and Brasil says that it seems as though the fibres of the
spindle helped largely in forming the membranes of the
daughter-nuclei. I did not notice that division went on
more rapidly at the periphery of the cyst than elsewhere.

In these early stages the centrosomes could be easily
demonstrated, and sometimes a sphere could be seen lying
completely within the nuclear membrane showing its intra-
nuclear origin (fig. 5). It would seem, too, that the spindle
in these divisions is formed within the nuclear membrane,
which only disappears when the spindle is fully formed
(figs. 5 a and 5 h). Most of the nuclei contain two or three,
some four, spherules of chromatin. These are probably
karyosomes, though on account of their small size it is
impossible to demonstrate two layers in even the largest of
them.

In a few of the cysts, all of about the same age, i.e.
having approximately the same number of nuclei, a small
sphere of chromatin could be seen being thrust out of each
nuclear spindle during division. It is worthy of note that if
one spindle in a cyst showed this every other spindle in that
cyst did so too. This cannot be regarded as a case of
Reduction, for the number of the chromosomes remained

568

MARGARET EORINSOX.

unchanged. 1 look upou it merely as the casting out of
superfluous chromatin, most probably one of the spherules
mentioned above, which is quickly dissolved and then absorbed
by the surrounding protoplasm (6g. o,s2)h.).

In some of the cysts intermediate in age between that
shown in fig. 5 and the pearl stage there are to be seen some
large nuclei at least twice the size of the others. I have
never seen these nuclei dividing, but some of them are in
a state of degeneration. In the earlier stages all the nuclei
divide in exactly the same way. There are not two methods
of division as in S ty lor hy nch u s (Leger, 1903). These
nuclei in the earlier stages seem to be not in any way
different from their neighbours; but after having divided
a certain number of times they divide no more, then
degenerate and die. In the process of degeneration they
naturally swell up a little, but their largeness in size as
compared with the others is mainly due to their not having
undergone so many divisions.

It seems to me that mitotic divisions are continued right
on until the pearl stage,” or, as Mr. Cunningham calls it,
the “convolution stage” is reached.

Mr. Cunningham, when he wrote his account of Kalpido-
rhynchus, was inclined to think that he was dealing- with
a case of isogamy. Nevertheless, he found a slight dift'erence
between the contents of the gametocytes enclosed in one
cyst. I, too, am convinced that there must be an inherent
difference between the gametocytes, but in the cysts which
I cut this difference Avas not appreciable till just before the
pearl stage was reached. Then, while the dividing-wall
between the two gametocytes Avas still intact, it could be
seen that the protoplasm on one side of the Avail stained
more deeply than that on the other, and that its mesh work
Avas slightly, but only very slightly, finer. It could also be
seen that the nuclei on the darker side Avere rounder, smaller,
and darker than the others, showing a more concentrated
chromatin. In sections through cysts at the pearl stage a
great accentuation of these differences can be noticed. Ou

liEPKOUUCTION OP KALriBOliHYNCHUS AliPNlOOL.E. 569

the dark or female side the chains of pearls lie on the edges
of convoluted bauds, which have about half the depth of
those on the male side, showing that in the male gametocyte
there is much more protoplasm left over after the formation
of the gametes than in the female. The nuclei also can be
seen in many cases to be about twice as large on the pale as
on the dark side (fig. 6). This convinced me that we have
anisogamy here, the chief thing which led me to this
conclusion being the difference in size between the nuclei
of the gametes in the respective gametocytes. But it would
not have been easy to prove this difference by means of
sections only, for tlie cysts were cut in all mauner of planes.
I therefore broke numbers of cysts at random on different
slides, and was fortunate enough to isolate in this way a few
female gametes fi-om the “pearl stage” and a number of
conjugation stages from cysts containing conjugating gametes.
Unfortunately i did not so isolate a male gamete, but in the
conjugation stages one could see its shape perfectly well.

The female gamete is nearly spherical in shape with a
spherical nucleus, which, as a rule, has darker and more
concentrated chromatin than that of the male gamete. This
nucleus has a volume equal to about one fourth of that of the
whole gamete.

The male gamete, on the other hand, consists almost
entirely of an oval, sometimes of a pyriform nucleus, which
is surrounded by a very thin layer of protoplasm. This
nucleus is generally about twice as large as tliat of the
female gamete. The oval nuclei are the more common

(fjg- ^)-

While the nucleus of the female gamete is surmounted by
a wide, low cone, the cone on the male nucleus is high and
narrow. In both cases tlie centrioles can often be seen to be
double.

In the act of conjugation it seems as though the male
nucleus with its cone forces itself through its own protoplasm,
which it casts off like a sheath as it enters the female gamete
(fig. 7, h, c, d, etc.). This is what happens most frequently.

570

MARGARET ROBINSON.

but I found some conjugations in which there was apparently
a fusion of the cytoplasm of the gametes, as well as of their
nuclei.

In neither gamete nor zygote could I demonstrate a cell-
wall by the use of Delafield’s hfematoxylin, but preparations
stained with Licht-Griin and picric acid showed a delicate
outline to the cells. This outline was more easily shown in
sections than in whole cell preparations.

The z}"gote is at first pyriform with very little in the way
of a stalk, but with one end a good deal thicker and rounder
than the other. The cell-wall is slightly more pronounced
than it was in the conjugation stage. It is at the narrow,
pointed end of the zygote that its nucleus lies. This nucleus
is also pyriform and has its wide end directed towards the
wide end of the zygote. Its chromatin is loosely arranged
in large thick rods and lumps, and is not surrounded by a
membrane. The absence of a nuclear membrane here is
probably not merely a result of the fusion of the nuclei, but
also a means of aiding the expulsion of a vacuole from the
nucleus (fig 8, h).

Brasil (1905) also notes the expulsion of a vacuole (sphere
hyaline) from the nucleus of every zygote in a cyst; and, as
well as the vacuole, he saw extruded a small globule of
chromatin, which he conjectures may form part of that
chromatin which is subsequently to be seen at both ends of
the spores of Monocystis after the first nuclear division.
I saw no such extrusion of a grain of chromatin here, but on
the assumption that it is merely superfluous chromatin this
is not to be wondered at, for a small globule of superfluous
chromatin was ejected at an earlier stage (see above and
fig. 8).

The absence of a nuclear membrane may possibly facilitate
the movements of the nucleus, for it certaiidy does move.
One can see the vacuole forming, and after its extrusion the
nucleus has not only acquired a membrane, but now lies at
the wide end of the zygote. Nuclei can be seen in inter-
mediate positions during' the formation of the vacnole. Its

UEPRODUCTIOX .(DF KALPIDORHYNCHUS AliENICOLiE. 571

extrusion and the formation of the memhi’ane seem to take
place simultaneously.

After this the nucleus becomes approximately spherical,
and its chromatin appears to be more finely divided and more
closely packed than it was. The zygote at this stage often
has a stalk-like projection at its narrow end, and this stalk
persists so that the spore has the shape of a pear with a
thickened stalk. Sometimes this stalk-like projection or
elongation does not appear till later, but it is invariably
present in the stage with four nuclei. The nucleus now
divides into two, then into four, and ultimately into the eight
nuclei of the sporozoites. It is my belief that at any rate the
earlier of these divisions are mitotic, but I have not been
able to prove this satisfactorily, and Mr. Cunningham is not
of my opinion. It is at the stage with one spherical nucleus
at the wide end of the cell that the cell-wall becomes
thickened to form the sporocyst, and the zygote thus becomes
a spore. Mr. Cunningham has mentioned the transparency
of this sporocyst. I was able to see it, in preparations stained
and mounted in spirit, as a dark line which follows the
outline of the cytoplastn very closely. Karely, until the
spores are fully ripe, i.e. until the cyto|)lasm is segregated
round each of the sporozoite nuclei, does it leave the little
stalk-like projection of the sporocyst (fia\ 8).

In some of my preparations tlie ripe spores are burst.
This may be due to reagents, the withdrawal of the cytoplasm
from the stalk having left a spot vulnerable to pressure in
the sporocyst. But it may be the natural course of events,
for the sporocyst seems to fit the cytoplasm fairly tigditly,
and the withdrawal of some of the cytoplasm from the stalk
into the body of the spore may have caused the sporocyst
to split.

The sporozoites are vermiform, with pointed ends. The
long nucleus occupies at least half the volume of each
individual. The chromatin is finely divided and evenly
distributed throughout the whole nucleus (tig. 8).

It seems to me that it is only by following this chromatin

JIAKGAKET EOIJINSOX.

572

iu the nuclei of the stages between sporozoite and trophozoite,
and in watching the evolution of the karyosonies, that we cati
arrive at any conclusion as to the origin of the chromatin of
the chromosomes in the first spindle. With this end in view
1 examined the alimentary tract of several infected specimens
of Arenicola ecaudata, and found cysts in the oesophagus
and in the intestine, which shows that swallowing is a possible
method of infection. I have also cut sections through the
gut walls of several specimens in the hopes of finding
sporozoites in transit, but always without success. It
seems most hkely that the sporozoites make their way very
speedily through the gut walls and then carry on their
further development in the coelomic fluid. 1 did find in the
coelom specimens of a very young trophozoite without an
epimerite (fig. 9) ; but this, unfortunately for my purposes,
had already several large and small karyosonies in its nucleus.

Two other points on which 1 have been able to supplement
Mr. Cunningham^s observations are multiple association and
the structure aud reproduction of the karyosonies.

Multijjle Associations. — In preparing the cysts for
embedding 1 noticed many cases of multiple association —
sometimes there were as many as five individuals together,
sometimes four, and very frequently three. Mr. Ciiuninghain
has figured four tro])hozoites coming together (19U7). On
cutting the sections 1 found a cyst containing five gametocyies,
each of which had many nuclei ; and i found several cysts
with three ganietocytes in like condition, liut since 1 have
never found a cyst containing more than two ganietocytes iu
the “pearl stage” or furtlier advanced, i am forced on
to iJr. \Voodcock’s conclusion (1900) that these muhiple
associations come to nothing.

Stkuctuke or thk Kauyosume.

Unfortunately 1 have not succeeded in tracing the origin of
the first karyosoine. In the youngest trophozoite seen by me
(fig. 9) there are already several karyosonies in the nucleus.

EKPEODUCTION OF KALFIDOEHYNCHUS ARENiCOL.E. 573

3’lie larger karyosomes all cousist of two layers — an outer
dense layer which stains deeply and strongly with Heiden-
hain’s hgeniatoxylin and other chromatin stains, and is, in fact,
hasophile, and an inner part which has not so strong an
attraction for basic stains, stains palely with Heidenhain,
and strongly with acid stains such as orange G., Licht-Griin
and picric acid, etc. The outer layer is, of com-se, chromatin,
and the inner is nucleolar substance or plastin. The smaller
karyosomes contain no plastin. They consist wholly of
basophile chromatin. This, it seems to me, is only an
expression of the fact that, as the karyosome increases in age
and size, it becomes by degrees converted from a basophile
into an acidopliile substance — i.e. from chromatin into
plastin. The larger karyosomes are often divided up into
a number of small chambers, each chamber being surrounded
by a wall or walls of chromatin. In fig. 10 there is a karyo-
some in which this process is beginning. This drawing
shows the nucleus of a gainetocyte, but the nuclei of tropho-
zoites often contain karyosomes in the same condition.
There can be little doubt that the chromatin of the partitions
and of the little knob-like thickenings is derived from the
dense outer layer. 1’his turning in, so to speak, of the
chromatin may take j)lace in order to increase its area of
action, for the result is an increase in the quantity of the
])lastin, and ultimately this increase is at the expense of the
chromatin. 1 have seen karyosomes in Avhich there Avere
larger chambers Avith thinner walls, and others from Avhich
the outer rim of chromatin had gone completely. It seems
as though finally the Avhole karyosome becomes converted
into plastin.

The karyosome divided up into chambers resembles on a
small scale the karyosome of Aggregata as described by
I\Iorott' (loots).

Kei’KOduction or the Kauyosoiaies.

Increase in the number of kaiyosomes takes place by a
kind of internal budding from the chromatin layer. I have

574

MARGARET RORIXSOX.

not seen a single case of scissi parity. The buds (fig. 10)
consist entirely of chromatin, and it is not until some time
after their escape from the parent karyosome that the inner
layer (plastin) makes its appearance in them.

On first noticing these internal buds I was puzzled as to
how they made their escape, but soon came to the conclusion
that an exit was made as occasion demanded and then closed
up again. I was therefore muck pleased to find that
Schneider had described the same kind of internal budding
in his account of Klossia (Aggregata) eberthi as long
ago as 1883. See also Schellack (1907).

At present, affinity for different stains is our usual criterion
for differentiating the contents of the cell, and we make a
broad distinction between chromatin and cytoplasm by sayiiig
that one is basophile and the other acidophile.

In working at this gregarine my first staining operation
was generally the nse of Heidenhain’s hmmatoxylin, and I
could not help noticing that in staining strongly with an acid
stain, after using Heidenhain, the inner part of the karyo-
some, the linin ineshwork and the centrosomes all took up
the acid stain (eosin, orange (x., or picric and Licht-Griin),
appearing to be stained by that and by nothing else. The
centrioles and outer layer of the karyosomes and the chro-
mosomes, however, kept black and were not affected by the
acid stain at all. If I stained weakly with the acid stain the
meshwork, inner part of the karyosome, and the centrosomes
all retained the black stain of the Heideidiain, though the
black on the inner part of the kai’yosome might with more
accuracy be called grey. I could, in fact, vary the amount of
greenness or blackness by varying the intensity of my acid
stain, but one or other always predominated. Now the linin
meshwork is known to consist partly of protoplasm and partly
of chromatin. In the inner pa it of the karyosome chromatin
is being converted into plastin, presumably for the nourish-
ment of the nucleus and ultimately of the cytoplasm. Does
it not seem that Avhile this process of conversion in going on,
there must be in the inner part of the karyosome a mixture of

EEPEODUCTION OF KALFIDOEHYNCHUS AEENICOL/E. 575

acidophile plastin and basophile cliromatin, and tliat in the
centrosonies also there is a mixture of chromatin and an
acidophile substance ? On this supposition we can, at least,
give our explanation of the results of staining-; for it would
seem that when (after using Heidenhain) we stain strongly
with an acid stain, then in the resulting preparation the
protoplasm masks the chromatin ; on the other hand, when
(after Heidenhain) we stain weakly with an acid stain or do
not use one at all, the chromatin masks the protoplasm. The
chromosomes, centrioles and outer part of the karyosomes,
since they consist entirely of chromatin, when once stained
with Heidenhain keep their black appearance unaltered by
any subsequent treatment with acid stains.

Tn conclusion, I w'isli to express my thanks to Professor
jMinchin for his friendly advice as to literature, and also for
his criticism of this paper.

Eefek?:nc'es to Litekature.

1905. Brasil. L. — “ Sur la reproduction des Gregarines Monocystidees,”

‘ Arch, de Zool. experimentale et generale.' 4 icnie ser., T. 4.

1900. Cncnot, L. — “ Reclierches snr revolution des Gregarines,'’ ‘ Arch,
de Biol..’ T. 17.

1907. Cunningham, J. T. — “On Kaliiidorhynchus arenicolse — a
new Gregarine Parasiticin Arenicola ecaudata,’’ -'Arch, fiir
Protistenk.,’ Bd. x.

1904. Leger, L. — “ La reproduction sexuee chez les Stylorhynchus,”

‘ Arch, fill- Protistenk.,' Bd. iii.

1908. Morolf, Th. — “ Die hei den Cephalopoden vorkommenden Aggre-
gata Arten — als Grundlage einer kritischen Studie liher die
Physiologie des Zellkerns,'' ‘ Ai-ch. fiir Protistenk.,’ Bd. x.

1907. Schellack, C. — Uel;er die Entwickluiig und Foi-tjiflanzung von
Echinomera hispida (A. Schn.),” ‘Arcli. fiir Protistenk.,’
Bd. ix.

1883. Schneider, A. — “Nouvelles ohsei-vations sur la sporulation du
Klossia Octopiana,” ‘Arch, de Zool. experimentale et gene-
rale,’ 2 leme scr., T. 1.

1905. Schnitzler, H. — Uber die Eortpflanzung von Clepsidrina

ovata,” ‘ Ai-ch. fiir Protistenk.,’ Bd. vi.

576

^[AliOAKET ROBIXSOX.

1S99 anti 1909. Siedlecki. M. — ‘‘Ueber die geschlechtliche Vermehiamg
tier Monocystis ascidise (R. Lank.)," ‘Bull. Acad. Sci.
Cracovie.’ xi and xii.

1900. AVoodcock. H. M. — "The Life Cycle of "Cystobia" irregularis
(Minch.), together with Observations on other Neogamous
Gregai'ines." ‘ Quart. Joiirn. Micr. Sci..’ vol. 50.

EXPLAXATION OF PLATE 29.

Illnstratiiig ]\riss M. Rohiuson’s paper “On the Reproduction
of Kalpidorhynchns arenieolte (Cnghtn.).”

Fig. 1 . — The first nuclear spindle and the breaking up nucleus of the
gainetocyte. X 1200.

Fig. 2. — The first nuclear spindle. Anaphase. X 1050.

Fig. 3. — The second and third nuclei. Metaphase. X 1050.

Fig. 4. — (a) Spindle drawn out during late anaphase. (b) Two
daughter-nuclei resulting from above. X 1200.

Fig. 5. — Part of a cyst in section, showing nuclei in different states of
division. X 1200.

Fig. 5rt and 5b. — Two niiclei from same cyst as fig. 5. showing
formation of the .spindle within the nuclear membrane. X 1200.

Fig. 0. — Portion of cyst in section at the pearl stage. X 500.

Fig. 7.— (d) Female gamete. X 1250. (b — </) Conjugation stages
stained with Heidenhain. Licht-Griin and picric acid. X 1250. (b — m)
Conjugation stages stained with Delafield’s liEematoxylin. X 1250.

Fig. 8. — (a) Zygote soon after conjugation, (b) Zygote with nucleus
extruding a vacuole. (c) Spore — one nucleus stage. (d) Spore with
two nuclei. (e) Spore with four nuclei. (/) Spore with eight nuclei,
(y) Spore with eight n\iclei. (b) Sjjoi’ozoite. X 2000.

Fig. 9. — Young trophozoite without an epimerite. X 500.

Fig. 10. — Nucleus of a ganietocyte showing multiplication of the
karyosome. and division of a karyosome into chambers. X 2000.

\

I

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3mxiM-. Sou/r^. JKit/r:Sa,. Vol SU, N.i' <3Pl P9

9

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■ram(p»*r5|

STUDIES IN THE EXPERTHENTAL ANALYSIS OF SEX. 577

Studies in the Experimental Analysis of Sex.

By

Geoffrey Smith,

Fellow of New College, Oxford.

With Plate 30.

1. On Mendelian Theories op Sex.

The re-discovery of Mendel’s observations on lieredity, and
the extended application of his ideas by such writers as
Correns, Tscliermalc, and Bateson to every branch of life, has
had a very profound influence on contemporary biological
conceptions, and it is not surprising that the pi’oblem of sex,
which has occasioned so many speculative theories in past
times, has been brought under the focus of Mendelian
research and subjected to its analysis. The conceptions of
segregation, of allelomorphism, of heterozygotism, to employ
the accepted terminology of Professor Bateson (1), seem
admirably suited in their application to the phenomena of
sex, because in .sexual reproduction we actually see that
the sexual characters do segregate into two sharply separated
sets of individuals, the males and the females, as if malcness
and fenialeness were in some way allelomorphic to one another,
while the occurrence of hermaphrodite forms and the latent
presence in one sex of characters proper to the opposite
sex indicate the phenomenon of heterozygotism or sex-
liybridism.

There is a further reason why Mendelian speculation has
naturally turned in the direction of sex. So long as it was
held that the sex of any animal or plant was not a question
of inheritance or of a pre-determiued quality of the germ,

578

GEOFFKEY SMITH.

investigators principally occupied themselves witli statistics,
and with the supposed influence of various external factors,
such as food or temperature, upon the production of one sex
or the other. But the almost wholly negative result of such
investigations, and the positive evidence afforded by such
cases as that of the bee, and of the rotifer, Hydatina, in
the latter of which two structurally different kinds of eggs
are produced destined to give rise to males and females
respectively, have influenced biologists to look for the cause
of sex-determination in the mechanism of heredity.

'Phe first to definitely formulate a Mendelian theory of sex
was Castle in 1903 (2). He supposed that in all cases each
sex IS a sex-hybrid or heterozygote of the composition $ ,
but that in the male sex maleuess was dominant, and in the
female fenialeness. As evidence that in each sex the opposite
sex-character is present in a latent state, he adduces the
undoubted transmission of male characters through the
female and vice versa, and the appearance of the secondary
sexual characters of the opposite sex in animals, as the result
of injury or disease of the gonads.

fl’lie process of segregation and fertilisation he conceives as
follows : Each sex being a heterozygote produces male and
female gametes in equal proportions ; but among them there
is selective fertilisation of such a kind that only male-bearing
spermatozoa can fertilise female-bearing eggs and vice versa.
Thus :

Mule gametes (spermatozoa). Female gametes (ova).

A ? ^ ?

Both the zygotes produced have thus the composition J' ? ,
and we are given to understand that for some reason in half
the zygotes maleness dominates, and in tlie other femaleness.

It must be admitted that this theory, according to which
every individual is potentially hermaphrodite, is sufficiently

STUDIES IN THE EXPERIMENTAL ANALYSIS OF SEX. 579

comprehensive to cover the fact, but it involves two large
assumptions for which there is little or no evidence, and its
very comprehensiveness prevents its affording any satisfactory
explanation of the undoubted variations in sexual constitution
which occur in Nature.

The two assumptions, firstly, that selective fertilisation
occurs, and secondly, that ’dominance is reversed in two sets
of individuals in the same species, for some unknown reason,
are not beyond the bounds of possibility, but they are
unwarranted and cumbrous additions to Mendelian theory
which we would gladly avoid, at any rate until all attempts
at a simpler explanation have failed.

The essence of a simpler interpretation was first hit upon
by McClung in 1902 (3), althoug-h he did not express him-
self in Mendelian language or definitely formulate a Mendelian
theory of sex. McClung observed that in certain kinds of
insect the males produced two forms of spermatozoa in equal
numbers, one half the spermatozoa containing an accessory
chromosome and the other half lacking it. This peculiar
distribution of the accessory chromosome was found to be
effected in the maturation division of the spermacytes, the
accessory chromosome, instead of dividing, passing bodily
over into half the secondary spermacytes. In the maturation
of the ova no accessory chromosome was observed. McClung
clearly pointed out that the behaviour of this chromosome in
the male was in some way connected with the determination
of sex. He considered that the spermatozoa containing the
accessory chromosome on fertilising an egg gave rise to a
male, while the normal spermatozoa gave rise to females.

It is clear that, although McClung did not state it in this
way, we have here the essence of a Mendelian theory of sex,
according to which the male is a heterozygote of composi-
tion, ^ ? , giving rise to ^ and ? gametes, while the female
is a recessive ? ? , giving rise to pui-e ? gametes. This
would further account for the production of the two sexes in
equal proportions according to the laws of chance. Unfortu-
nately this simple account of the matter has been proved
VOL. 54, PART 4. — NEW SERIES. 41

580

GEOFFEEY SMITH.

erroneous, especially by E. B. Wilson and his pupils. Wilson
(4) has shown that although the dimorphism of the sperma-
tozoa does occur, and is, indeed, a frequent phenomenon
among insects, yet that where an “ accessory ” chromosome
is present in the males it is due to half the spermatozoa con-
taining one chromosome less than the eggs, while those with
the “^accessory” chromosome contain the same number as
the eggs. It is therefore impossible to say that the sperma-
tozoa with the accessory chromosome give rise to males, as it
is evidently those without the accessory chromosome which,
in conjunction with an egg, produce an animal with an odd
number of chromosomes, i.e. a male. This fact has pre-
vented Wilson hitherto from accepting McClung’s simple
interpretation, since he is convinced that the sex-character
must be represented by a definite chromosome in the cell.
Of course, if we are content to give up the idea of the sex-
character being necessarily represented by a definite chromo-
some, an idea which, indeed, has not vei’y much to support it,
McClung’s explanation would still hold, except that the male-
producing spermatozoa would be those without the accessory
chromosome, while the female-producing spermatozoa would
be those that possessed it. Professor Wilson, however, will
not readily accept this, and he has endeavoured to get over
the difficulty by framing several different theoi’ies of sex of
very great complication. The latest of these theories not
only assumes a great complication of gametic representatives,
but also involves selective fertilisation, so that it belongs to
the same category as Castle’s theory and need not be dis-
cussed further at present.

During the years 1904-6 I was occupied at Naples in
studying the effect of the parasitic Cirripede Sacculina on
its host, a species of spider crab (Inachus), and from this
study I was led shortly to formulate a Mendelian theoiy of
sex. As the result of the examination of many thousand
specimens of the crab at various stages of infection, I con-
cluded that whereas the males, under the influence of the
parasite, were capable of assuming all the female secondary

STUDIES IN THE EXPERIMENTAL ANALYSIS OF SEX. 581

sexual characters, and under certain conditions might even
develop typical ova in their testes, the infected females, on
the other hand, although the ovaries in many cases completely
disappeared, never approached to the male primary or secondary
sexual characters in the slightest degree. After citing and
examining other cases, especially among the Crustacea, in
which the male sex showed undoubted signs of latent herma-
phroditism, I concluded that, at any rate in these forms, it
would appear that the male was a potential hermaphrodite,
and the female purely female. The significant bearing of
this conclusion upon a Mendelian theory of sex was obvious,
and since at the time the only definitely formulated Mendelian
theory of sex was that of Castle, according to which both
sexes of any species were potentially hermaphrodite, I was
led to formulate an alternative theory, which did not require
the assumption of selective fertilisation, in the following’
words : “ There is a final topic for discussion, namely, the
connection between the theory of sex here adopted and con-
temporary l\Iendelian theory. It is interesting to observe
that where an attempt has been successfully made to find
structural differences in the germ-cells as possible indications
of this early sexual diffei’entiation, the manner of this differen-
tiation is in harmony with the results we have obtained from
the study of parasitic castration. The discoveries of Henking,
McClung, Wilson and others have shown that in many insects
two kinds of spermatozoa exist, differing in the constitution
of their chromosomes, while the eggs are apparently^ all the
same. If we suppose that the two kinds of spermatozoa
represent the male and female sex respectively, while the
eggs are purely female, we ’would obtain in the process of
sexual generation ^ c? ? + h ? ? , in which the male

spermatozoa united with female eggs give rise to males of
really hermaphrodite constitution, while the female sperma-
tozoa united with female eggs give rise to females of pui’e
female constitution. It is obvious that this interpretation is
in strict agreement with the main conclusion brought out in
this chapter, viz. that males are potentially’ hermaphrodites.

582

GEOFFREY SMITH.

while females are incapable of assuming male characters. It
is doubtful, however, whether this particular ‘ Meudelian ’
interpretation can be applied generally, because in some
animals, e. g. the bee, it appears that the egg by itself is male
and only becomes female through fertilisation, while in many
Cladoceraand Aphidesfemales gave rise parthenogenetically to
males. It appears, therefore, that the primary mechanism of
sex determination may be vai'iouslj’’ distributed in the germ-
cells” ([5] ‘‘ Rhizocephala,” ‘Fauna and Flora des Golfes von
Neapel,’ Monogr. 29, p. 89, 1906).

In the above words I believe that the following Mendeliau
theory of sex is clearly stated :

(1) That in certain species of animals (e. g. Inachus and
other Crustacea) the male is a heterozygote of the composition
(J ? , while the female is a pure recessive of the composition
? ? .

(2) That the sexual constitution is not necessai-ily the same
in all species of animals, e. g. in the case of the Cladocera,
where the female is proved to be a heterozygote owing to her
capacity for producing both males and females partheno-
genetically.

In excuse for recapitulating these statements it may be
pleaded that, owing to their appearance in a publication
principally devoted to morphological studies, they have not
been referred to by subsequent writers who have indepen-
dently arrived at the same Meudelian theory of sex.

Thus, Professor Bateson and Mr. Punnett (1908), in offer-
ing an interpretation of the striking results obtained by
Doncaster in his breeding experiments with the currant moth,
have suggested that in this case the female is a heterozygote
(c^ $) and the male a homozygote (c^ (?). The reason for
this interpretation is as follows : The common currant moth.
Abraxas gross u la riata, occasionally gives rise to a pale-
coloured vai’iety, lacticolor, and this variety has been
hitherto supposed to be confined to the female sex. Doncaster
made, among others, the following crosses :

(1) Lacticolor ? x grossulariata gave cl and ?

STUDIES IN THE EXPERIMENTAL ANALYSIS OF SEX. 583

all Grossu lariata. This is the generation used in
succeeding crosses.

(2) F^ grossnlariata ? x Fj gross u lariata S gave
Fj grossular iata s and $ s and lacticolor ? s, but no
lacticolor s.

(3) Lacticolor ? x F;^ grossulariata ^ gave grossu-
lariata d* s and ? s and lacticolor cJsand ? s, the lacti-
color J s being the first ever seen.

The explanation of these results offered by Bateson and
Punnett depends on the two assumptions —

(1) Tliat the female is heterozygous in sex, femaleness
being dominant, and the male a homozygous I’ecessive.

(2) That when in Fj the two dominant characters, female-
ness and the grossulariata factor co-exist, there is spurious
allelomorphism or repulsion between them, such that each
gamete takes one or other of these factors, not both.

The three crosses described above read, therefore :

(1) Lac. ? X (fros. S
Produce gametes —

Lac. ? . Lac. x Gros. S
Produce zygotes —

Fi Lac. ? Gros. S • Gros. S •

(2) Fi Gros. ? X Fj Gros. S

Produce gametes —

Lac. ?. Gros. J (Lac. d' ) (Gros. ?) x Lac. J (iros. S
Produce zygotes —

Lac. ? Lac. cJ . Lac. ? Gros. J . Gros. d* J . Gros. c?

Gros. J .

The gametes in brackets are not formed owing to spurious
allelomorphism.

(3) Lac. ? X Fj Gros. J

Produce <>:amete.s —

Lac. ? Lac. ^ x Gros. c? Lac. c?

Produce zygotes —

Lac. ? Gros. J . Lac. ? Lac. . Lac. J Gros. .
Lac. cJ Lac. cJ .

This remarkable case has been given in full, as it illustrates

584

GEOFFREY SMITH.

the kind of way in which light is thrown on the constitution
of sex by breeding experiments. This is by no means the
only interpretation of the facts that could be offered, but it is
the simplest and the most in accordance with other results in
which the phenomenon of spurious alleloTuorphism appears to
occur. It must be remembered, however, that in cases of
this kind we ai*e not dealing with the sex characters directly,
but only through the medium of an assumption which cer-
tainly gives a simple though not the sole possible explanation
of the results.

A similar interpretation is given by Bateson (1) in the case
of the cinnamon canary and the brown Leghorn fowl.

Professor Correns (8), on the other hand, as the result of
hybridisation experiments with Bryonia, comes to the con-
clusion that in this case the male is heterozygous and the
female homozygous, and he inclines to give this interpretation
a wide application both in the animal and plant kingdom.

In would appear, therefore, that a considerable body of
evidence is accumulating, drawn from very various fields of
research, which tends to show the justness of the view that
in sex we are dealing Avith a phenomenon Avhich may be
termed “ half-hybridism,” i. e. one sex, either male or female,
is always a sex-hybrid, while the other is pure.

We may noAV examine some of the serious difficulties which
this view encounters. In the first place it may appear very
strange that the sex-hybrid individual (c? ?) should appear
in one case as a male and in another as a female, in other
words that there should be such a complete reversal of
dominance. But it may be pointed out that dominance is one
of the least constant phenomena in cases which have yielded
satisfactorily to Mendelian analysis. Let us, moreover, con-
sider what happens in the case of functionally hermaphrodite
animals, in which there can be no doubt at all as to their
heterozygous nature. We may divide such animals into
protandrous, simultaneous, and protogynous hermaphrodites,
of which the first category is by far the commonest. To take
a typical instance of protandry, in the parasitic Isopoda Epi-

STUDIES IN THE EXPERIMENTAL ANALYSIS OF SEX. 585

carida, the hermaphrodite individuals are at first, in the larval
state, apparently pure males. They then settle down as
parasites and lose every trace of their male organisation and
become converted into what are apparently pure females. In
fact for a very long period of time they were considered by
naturalists to be typically dioecious animals with very marked
sexual dimorphism. Now let us suppose that for some reason
or other certain of these individuals failed to develop further
than the male larval state. They would then be constitution-
ally hermaphrodites, in which the male condition was dominant,
and they would be put down as males. Then let us suppose
that other individuals for some reason left out the male period
of their history, possibly by becoming fixed parasites at an
earlier period before the testes were developed. These indi-
viduals would then be constitutionally hermaphrodites, in
which the female condition was dominant, and they would
be considei’ed with equal confidence as females. In this way,
by shifting the period at which the sexual organisation
matures, a process which may be very easily conceived to
occur, we would arrive at an apparently complete reversal of
dominance.

That this shifting of the period of maturity actually occurs
is shown by the existence of the three classes of hermaphro-
ditism noted above ; thus in the single class of simple Ascidians
we meet with both protandry and protogyny. Let us take
another slightly different instance, the case of the spider crab
Inachus, parasitised by Sacculina. The male parasitically
castrated crabs may show every degx’ee of modification towards
the female state, until finally we obtain male crabs which have
been so completely transformed as to retain only a single male
character, viz. the copulatory style in so i-educed a state as
to be invisible except with a lens. These crabs, besides
exhibiting in a typical condition the broad abdomen, reduced
chelae, and abdominal swirnmerets of the female, may under
certain conditions develop ova from the remains of their
testes, and these ova may grow to a very large size and
become filled with the reddish-coloured food-yolk character-

586

GEOFFEEY SMITH.

istic of the species. In these cases, althougli a certain
amount of sperm was always present as well, the female part
of the hei’inaphrodite gland greatly preponderated. Here,
then, we have individuals of hermaphrodite constitution,
which normally only show the male characters throughout
life, i.e. in which maleness is dominant; but when the
presence of the parasitic Sacculina sets up a distm-bance
this dominance is almost completely reversed, and the hitherto
recessive female characters appear in all completeness.

Again, to take the opposite case. In deer and pheasants it
is well known that certain individuals which have actually
bred as females, may in old age develop the male secondary
sexual characters in a very complete manner. Such indi-
viduals prove themselves to have been heterozygotes in which
the dominant female character is replaced for some reason by
the recessive male.

It is clear, therefore, from the foregoing' instances, that
individuals of hermaphrodite constitution may exhibit any
of a whole series of modifications from apparently pure male-
ness, through simultaneous hermaphroditism, to apparently
pure femaleness. This being the case, the difficulty of con-
sidering that in a normally dioecious animal either one
sex or the other is a sex-hybrid, according to the species or
group of species we are dealing with, is materially lessened.
We may, in fact, state the case as follows : that three types
of individuals exist in respect of sex, pure males, hermaphro-
dites, and pure females, and that the hermaphrodites may
appear as males, hermaphrodites, or females according to a
physiological condition which is confessedly not understood.

We have, so far, formulated the Mendelian theory of sex,
so as to account for the existence within a species of indi-
viduals having' either the constitution $ and $ ? or of
? and c? c? j in the former case maleness being dominant,
in the latter femaleness. This is the simple half-hybrid
theory of sex. We must, however, consider the possibility
of the existence of three types of individuals within the same
species, viz. J* c? > c? ? > and $ ? . The diflSculty of this

STUDIES IN THE EXPERIMENTAL ANALYSIS OF SEX. 587

conception lies in the tact that we would have to assume that
some of the heterozygous individuals ( c? ? ) would have to
function as males and others as females iu order to secure
a continuous output of pure males and females. For if all
the heterozygous individuals were functionally either males
or females exclusively, they could only produce one form of
pure zygote iu combination with the gametes of the pure sex.
The supposition tliat the heterozygote may appear as either
male or female in the same species of animal is by no means
impossible, since it only leaves us face to face with the same
problem, which is at the root of the whole sex question,
namely. What is the physiological state which brings about
the suppression or development of the male or female char-
acters in animals which undoubtedly possess the potentiality
of both? It is held by a number of observers, for instance
Mr. ^Valter Heape (9), that the proportional output of the
Sexes is iuHueuced to a very large extent by external condi-
tions of feeding, temperature, breeding-seasons, etc., and it
is quite possible that these influences are sufticient to give a
bias to the heterozygous embryo to appear as either male or
female.

However this may be, the assumption that three types of
individuals may exist with distinct gametic output would
account for the marked disproportion in number of the sexes,
which is known to occur in the offspiing of certain families,
both among animals (especially butterflies) and human beings.

There is anotlier class of facts which offers an interesting
but difficult field for interpretation upon Mendelian lines, viz.
the sex of parthenogenetically jjroduced ofi^spring. In the
case of such animals we get every variety of product, the
parthenogenetically produced young being either only males
or only females or a mixed brood of both sexes. But there
are certain common features in these cases. Thus, in the
case of the bee, in Cladocera, and in certain Eotifera the ferti-
lised eggs always give rise to females, the males being only
produced parthenogenetically. The females iu some of these
cases, e.g.the Cladocera, are proved beyond doubt to be

588

GEOFFEEY SMITH.

lieterozygotes, as one and the same female may give rise par-
tlienogenetically to males and females. It is also proved in
these cases that no segregation occurs in the pi'oduction of pai'-
thenogenetic females, since a parthenogenetically produced
female may give rise by parthenogenesis to a mixed brood of
males and females. We may be certain, therefore, that the
females in these cases are heterozygotes ((J $ ). With regard
to the parthenog’enetically produced males we are naturally
more in the dark, since they produce no parthenogenetic
young by which they can be judged. The most obvious
supposition is that, since they, like the females, are produced
parthenogenetically, there is no segregation in their produc-
tion, and that they also are heterozygous (c?" ? ) and produce
and ? spermatozoa. If, however, we are going to main-
tain the half-hybridism theory of sex, since the females are
certainly ? the males must be pure ^ (5',and some process
of segregation must occur. But if this is the case why do
the eggs, when fertilised with such purely male spermatozoa,
invariably give rise to females ? It has been held, especially
for the bee, that the mere act of fertilisation in itself is the
cause of the production of females, and if this is the case it
is very difficult to bring the phenomenon into any relation
with Mendelian theory. There is, however, an alternative to
this explanation. AVe may legitimately hold that the female
gives rise to two different kinds of eggs, male and female,
of which only the female egg’s are capable of being fertilised.
Such female eggs, being fertilised by the male spermatozoa,
will give rise to heterozygotes of the compo.sition ? , which
ex hypothesis will appear as females, while the unfertilised
male eggs will give rise to males of pure male constitution.

In the Rotifer Hydatina and in the worm DinoiDliilus
we know that two different kinds of eggs are produced,
large eggs which give rise to females and small eggs which
give males, and this fact seems at first to favour the theory
proposed above. In the case of Dinophilus, however.
Dr. Shearer, in a recent unpublished research, has shown
that the female Dinophilus is fertilised while still immature

STUDIES IN THE EXPEEIMENTAL ANALYSIS OF SEX. 589

and before any visible differentiation of the eggs into male-
producing and female-producing forms has occurred, so that
this differentiation may be the result of fertilisation and not
due to the inherent heterozygotism of the female. In the
case of Hydatina,* as clearly shown in R. C. Punnett’s
interesting' paper (10), and in the Cladocera, there is no doubt
that the female can give rise parthenogenetically to both males
and females, so that in these cases the female sex is certainly
heterozygous; but we have no certain means of judging
whether it is the female eggs alone which are capable of
being fertilised.

Interesting and suggestive as the evidence drawn from
breeding experiments and from the cytology of maturation is,
it appears that the most cogent and unassailable evidence for
sex heterozygotism is afforded firstly by Inachus parasitised
by Sacculina, in which the male sex is proved to possess the
secondary and primary female characters in a latent state,
and secondly by the Cladocera, in which the female sex is
proved to be heterozygous owing to the parthenogenetic
females giving rise to both males and females.

The foregoing arguments and considerations have shown
us that while a number of facts definitely support the half-
hybrid Mendelian theory of sex, there is nothing which
definitely controverts it. The theory has the merit of being
a simple one, and it accounts for the facts without the
necessity of making any additional assumptions such as that
of selective fertilisation, an assumption which may in the
future prove necessary, but which would seriously impair
the validity of Mendelian analysis.

And it may be retnarked that the half-hybrid theory of
sex not only alters our view of the sexual constitution of
animals and plants, but it indicates, if it is well founded, the
real ground upon which the problem of sex must be attacked.
This, as has been already stated, is the inquiry as to the
physiological conditions under which one sex or the other
gains the upper hand, i.e. becomes dominant in a heterozygous
individual which contains potentially the elements of both sexes.

590

GEOFFREY SMITH.

It must be remembered, moreover^tliat sex is not necessax’ily
a simple unit character, inherited in its entirety as such ;
thus sexual characters fall into two main divisions, primary
and secondary, and the latter again may variously affect any
of the organs or parts of the bod}\ A^ie will give reasons,
however, in the next section, for assuming the existence of a
sexual formative substance, male or female, which controls
the development of both primaiy and secondary sexual
characters, and for the present it is assumed that the male
and female modifications of this substance are the allelo-
morphs which segregate in the manner described above, and
give rise to the half-hybrid nature of sex.

2. On the Correlation between Primary and Secondary
Sexual Characters.

Various definitions have been given of primary and
secondary sexual characters. In these studies the term
“primary sexual characters” is applied to those characters
which affect the differentiation of the actual generative organ,
testis or ovary, in which the ova and spermatozoa are pro-
duced, while all those sexual characters are considered
secondary which affect the otlier parts of the body, e.g.
generative ducts, copulatory or any other organs, external or
internal, which differ in the two sexes.

The fact that there is a physiological correlation between
the state of development of the secondary sexual characters
and the primary rejiroductive gland has been vaguely
recognised from time immemoidal. Thus the knowledge that
the castrated males of the liuman sjiecies and of many races
of domestic animals show in various degrees an arrested
development of the secondary sexual characters goes back to
periods long antecedent to scientific biology. Put despite
this long familiarity with certain fundamental facts, there
does not exist even at the [iresent time a clear conception of
the nature and limits of this correlation. Careful observation
and a certain amount of experimental work have revealed a

STUDIES IN THE EXPERIMENTAfi ANALYSIS OF SEX. 591

very large body of facts bearing upon the question, but they
have chiefly served to emphasise the irreg’ularity of the
phenomenon, and it is certainly impossible at present to
formulate any definite theory to connect the known facts in
a comprehensive and satisfactory manner. It is not the
purpose of this essay to attempt a review of the recorded
cases of so-called hermaphroditism and of the abnormal
condition of tlie sexual system which throw a somewliat fitful
light on the problem, but reference may be made to the
critical work of Herbst ('Formative Reize’ [H]), in wliich he
discusses a large body of conflicting evidence and draws
certain wide conclusions. He shows that, while the evidence
in favour of some causal correlation existing between the
primary and secondary sexual characters is oversvhelming,
yet this correlation is not of so definite a nature as to sanction
the simple view that the development of tlie secondary
sexual characters as a whole is directly dependent on the
development of the primai-y characters ; he concludes, how-
ever, definitely, that in the vast majority of cases the full
development of the secondary sexual characters in either
sex is conditioned by the presence of the corresponding
primary organ in a functional state. Even this very cautious
and limited acceptance of correlation breaks down in certain
exceptional cases. Thus Kellogg (12) has .shown that the
gonad of the silkworm can be extirpated in the larval stage, so
that no trace of this organ is to be found in the adult, and yet
the moth develops its marked secondary sexual characters to
the full, while Meisenheimer (13) has performed the ingenious
experiment of transplanting the young gonad from one sex
into the other where it may develop to maturity, and yet no
change is to be observed in the secondary sexual characters
of the adult insect. In the case of those particular insects,
therefore, it appears that there is no connection whatever
between the development of the secondary sexual characters
and the presence of a differentiated gonad, and though it is
true that this is the only case known in which this entire
independence is to be observed, yet we can trace a series of

592

CxEOPEREY SMITH.

instances in wliicli tlie removal of the gonad inhibits in
greatly varying degrees the development of the secondary
characters.

In attempting, therefore, to frame a theory which shall
give a satisfactory account of the undoubted correlation which
exists in various degrees between the primary and secondary
sexual characters, we must bear in mind the variability of
this correlation, and even in certain cases its non-existence.
Mr. J. T. Cunningham, in a recent interesting paper (14), has
put forward a theory which appears to me to fail in this
respect. According to his theory the development of the
secondary sexual chai’acters is due to the action of an internal
secretion produced by the gonad, principally at its maturity.
There can be no doubt that this statement is partially true,
but it does not cover all the facts. If it represented the
whole truth theabseuceof a differentiated gonad should in all
cases be accompanied by the entire absence of all secondary
sexual characters usually connected with it, and this is certainly
not the case.

We will now examine in some detail a particular instance
which appears to throw a more definite light on the subject
than any that has hitherto been obtained. The discovery of
the phenomenon of parasitic castration Avas made by the late
Professor Giard, and it always seemed to me very surprising
that no one had followed up his discovery, since it affords a
very obvious and simple Avay of gaining an insight into the
nature of sex, without the necessity of performing a delicate
operation Avith the clumsy means at our disposal. For in this
case, instead of performing the operation ourselves, Ave find
that Nature employs for the purpose some of the loAver classes
of creation, Avho, though not endowed with the great intelli-
gence Avhich is sometimes reported to be characteristic of man-
kind, yet accomplish a thing' Avhich is not only impossible at
present for a man to do, but also A^ery difficult indeed to under-
stand. The spider crab, Inachus mauritanicus,^ is very fre-

* By an imfortAinate error in nonienclatnre I. man ri tan icus (Lucas)
was called I. scorpio (Fabr.) throngliout my monograph.

STUDIES IN THE EXPERIMENTAL ANALYSIS OE SEX. 593

quently infected with a species of rhizocephalous Cirripede
called Sacculina neglecta (5). This parasite at first lives
a free existence as a minute larva; it then fixes itself to a hair
on the outside of its host and passes into the body of the
latter a small gi'oup of cells which find their way to the blood-
space round the intestine. Here they begin to gi'ow very
rapidly into a branched tumoui--like body which sends its
ramifications into every part of the body-cavity of the crab.
A certain part of the tumour becomes applied to the ventral
body-wall of the crab at the junction of thorax and abdomen,
and at this point the reproductive organs, etc., of the adult
Sacculina ai’e developed and finally thrust to the outside in
a muscular bag which remains attached to the crab and
swells to a large size, gaining its nutriment from the system
of branching roots which continue to multiply and grow
inside the crab’s body.

Now the chief effect which the parasite exerts on the crab
is to cause the complete or partial atrophy of the internal
generative organs, with their ducts, while remarkable changes
take place in the structure of the external secondary sexual
characters. Of 1000 specimens of Inachus infected with
Sacculina examined by me at Naples, 70 per cent, of both
males and females showed very distinct alteration in their
secondary sexual characters, while all showed some degree of
reduction or atrophy of the gonad. Of the many thousands,
at present well over 5000 specimens, of uninfected Inachus
examined, only one specimen showed any trace of the changes
such as were inet with in the infected individuals. This speci-
men, which was a perfect hermaphrodite both externally and
internally, may have been an instance, such as occurs with
extreme rarity in decapod Crustacea, of hermaphroditism
apart from parasitic castration, but it is equally possible that
it was really a crab that had recovered from an infection with
Sacculina, and had undergone several moults so as to lose
the scar characteristic of crabs that have been once infected.
I mention these facts with regard to the numbers of specimens
examined, because it is important to realise not only the

594

GEOFFREY SMEPH.

extent of the material upon which my conclusions are based,
but also the invariable certainty and regularity of the effect
observed.

'File sexes of normal uninfected Inachus mauritanicus
differ in that the adult male (PI. 30, figs. 1 and 2) possesses
greatly elongated and swollen chelae, while the abdomen is
small in size, is carried flatly opposed to the thorax, and is
furnished with only two pairs of appendages — viz. a large
pair of stout copulatory styles and a greatly reduced pair of
appendages behind them. The adult female (PI. 30,
figs. 10 and 11) has small, slender chelae and an exceedingly
broad, trough-shaped abdomen which is furnished with four
pairs of bi-ramous appendages. These appendages are clothed
with long hairs, some of which are used for attaching the
eggs.

The sexual difference in the chelae is not developed until
maturity, but the differences in the abdomen are marked
soon after the Megalopa larval stage and long before maturity.
The female, however, ait first goes through a stage in which
the abdomen is comparatively small and flat, and the
appendages are short and rod-like without the filamentous
haiirs characteristic of the adult (PI. 30, figs. 13 and 14).

The males infected with Sacculina show every degree of
modification towards the female type (PI. 30, figs. 3 to 9).
In some the only change to be observed externally is the
reduction in size of the chelae, and perhaps a slightly tapering
form induced on the usually stout and blunt copulatory style
(PI. 30, fig. 5). In others the abdomen is somewhat
broadened (PI. 30, figs. 3 and 4), and in a further stage
the abdomen is distinctly broadened and somewhat trough-
shaped, while perhaps one or two additional appendages are
developed in a rudimentary condition behind the reduced
copulatory styles (PI. 30, fig. 6). If such forms are carefully
dissected the gonad is observed to be greatly shrivelled, but
it can still be clearly recognised as a testis, while a few
clumps of spermatozoa may be found in the vasa deferentia.
Finally we obtain forms (PI. 30, figs. 7, 8 and 9), usually

STUDIES IN THE EXPEEIMENTAL ANALYSIS OF SEX. 595

among the smaller and medium-sized individuals^ in which
the chelse and abdomen have taken on the complete adult
female appearance, the only male character remaining being
the copulatory style, which is sometimes reduced to a minute
knob (fig. 8). I was for a long time in doubt as to which
sex these highly modified crabs belonged, as the majority of
them on dissection proved to possess no remains of the gonad
and gonoducts, except in certain cases some small shreds of
germinal epithelium. In a few, however, the remains of the
gonoducts were found, and in all cases they were in the
position of the vasa defei’entia, proving the animals to be
males in which the modification towards the female type had
gone very far. Further reflection showed that all these
highly modified crabs were originally males, and this was
proved Avithout doubt by their invariable possession of the
copulatory style in either a complete or greatly aborted state.
For in the case of the females, although they, too, undergo
characteristic changes, they never make any approach either
in the chelae or abdomen towards the male state, and there is
never any trace in them of the development of the copulatory
style.

It is therefore altogether impossible that other crabs, in
which the sex cannot be determined by the internal gonad or
its ducts, should be females which, contrary to all experience,
had suddenly developed the single male character of the
copulatory styles. That they were originally males, however,
is shown by the perfect gradational series, which can be
traced from hardly modified males up to those specimens in
which the only male character retained is the presence of the
copulatory styles.

It is important to notice that the males, when they develop
the female characters to any great extent, invariably exhibit
these characters in the form in which they occur in the adult
female, the abdomen assuming the trough-like appearance,
and the appendages being slender and provided with filamen-
tous hairs. I emphasise this point because certain people
have argued with me that the modification of the male was

VOL. 54, PART 4. NEW SERIES. 42

596

GEOFFREY SMITH.

not due to the assumption of definite female characters, but a
“ reversion” to an “ undifferentiated ancestral condition,” or
to “ an embryonic state.” But if this were so, the male
should at any rate assume the comparatively undifferentiated
state which is actually passed through by the young imme-
diately after the megalopa stage, and which is retained by
the female until the first brood of eggs is produced, viz. the
small, flat, plate-like abdomen and the rod-like form of the
appendages. Now, as a matter of fact this form is never
assumed by the male as the result of parasitic castration, the
female chai-acters being acquired, if imperfectly, yet with the
definite characters only found in adult females which have
produced a brood of eggs. This fact alone seems to me to
demolish the above-mentioned argument, but it is even more
completely answered by the fact, soon to be described, that
certain of these male specimens may, on recovery from the
disease, actually produce ova as well as spermatozoa in their
regenerated gonads, thus proving that they actually have
developed true female characters and have not merely re-
turned to an undifferentiated condition of an altogether sup-
posititious nature. The infected females, as we have already
stated, do not show in any character any approach towards
the male secondai’y sexual characters, though dissection
proves that in all cases the ovary is arrested in development,
or even completely aborted. The only secondary sexual
character affected is the condition of the abdominal appen-
dages, which may be greatly reduced in size (PI. 30, fig. 12).
There is never any approach to the male either in the chelae,
or in the shape of the abdomen, or in the development of an
appendage corresponding to the copulatory style of the
male.

We have now to consider the case of the highly modified
males which have developed the external female characters
but retain the copulatory styles. We have seen that in all
these specimens the gonad is reduced to a few shreds of un-
differentiated germinal epithelium, and in some cases the
remains of the vasa deferentia. In a very few cases such

STUDIES IN THE EXPEEIMENTAL ANALYSIS OF SEX, 597

crabs were found in a state of nature to have recovered from
the disease, the Sacculina having dropped off and left the
characteristic circular scar on the abdomen where it was pre-
viously attached. Of a large number of crabs experimentally
freed of the parasite a few also survived for several months.
Of these specimens, three which had recovered naturally and
one which had been experimented on, were found to have re-
generated the gonad, which had grown to a considerable size.
The gonad was found to contain a certain amount of adult
spermatozoa and a number of ova, some of them small and
immature, others of a very large size and distended with the
reddish-coloured yolk, which normally appears in the eggs as
they approach maturity.

The fact that the alteration of the male, under the influence
of the parasite, is consummated by the final assumption of
complete internal as well as external hermaphroditism is, I
believe, without parallel, and confers on this case a peculiar
definiteness and value which we cannot obtain elsewhere.
The phenomenon appeared to me so strange and so little
likely to gain credit from people who had not actually investi-
gated the matter, that I was greatly pleased when Mr. F. A.
Potts undertook, at my suggestion, to examine the matter in
a parallel instance, namely the effect of the parasite Pelto-
gaster on the hermit-crab, Eupagurus (15). The investi-
gation of this case offered considerably more difficulty than
the case I had examined, but he was able to obtain a series of
results which happily place the main conclusions outlined
above on a very sure footing. Stated concisely he found
that, as in Inachus, so in Eupagurus the infected males
assumed to varying degrees of perfection the female charac-
teristics,^ but that the females, as in Inachus, never acquired
any male characters, although they might show reduction of
their own secondary sexual characters. The most remarkable
result, however, obtained by him consists in the discovery
that in a very large number of modified males, while the

' The female characters assumed by the male are here, as in Inachus,
those of the adult bi-eeding female.

598

CxEOFFREY SMITH.

parasite was still on them, small ova were developed in the
testes. This observation, while differing in an interesting
manner from what occurs in Inachus, where the ova are not
found until after recovery, yet confirms the account I have
given for Inachus in a very convincing way.

If we consider the facts related above in their bearing on
the problem of the correlation of the primary and secondary
sexual characters, it is evident that we are provided with
some instructive evidence. In the first place we observe the
male developing the secondary sexual characters of the
female, and this it does, not mei’ely in a negative manner by
returning to some intermediate, indifferent condition, as
usually happens in the case of ordinary castration, but by
positively acquiring characters which normally only appear
in the adult breeding female. Now, we may hold two
opinions with regard to these males — either that their
resemblance to the female is a spurious one, and that the
development of the female secondary sexual characters is
due in them to a different cause to that which conditions
their development in the female, or else it is a true resem-
blance due to the same cause. That the latter alternative is
correct is shown by the fact that these males may subse-
quently develop typical ova, because we cannot requii’e more
of an animal to prove its female nature than that it should
produce ova and exhibit all the secondary characters of the
female as well. The infected males, therefore, develop the
female secondary sexual characters for the same reason that
the female does. Now what is this reason in the female ?
In the female the development of the secondary sexual
characters is correlated with that of the ovary. Thus the
adult form of the abdomen and the form of the abdominal
appendages is not assumed until the ovary is ripe, while the
atrophy of the ovary, as a result of the presence of Sacculina,
causes the atrophy to some degree of the appendages. In
the case of the female, therefore, we might assume that the
ovary produces a substance or internal secretion which
causes the development of the secondary sexual characters.

STUDIES IN THE EXPEEIMENTAL ANALYSIS OP SEX. 599

But this caunot possibly apply to the infected males, because
they develop the same female secondary sexual characters
before there is any ovary present at all, much less a mature
ovary ready to produce ripe eggs. Now these males,
although they develop the female secondary sexual characters
when there is no ovary present, yet subsequently they may
regenei’ate an ovary from the shreds of germinal epithelium
remaining from the degenerated testis. In other words
they have the potentiality to produce an ovary, and we may
safely argue that it is this potentiality which enables them
to produce the secondary sexual characters befoi’e the actual
ovary is there.

In this conception it appears to me lies the solution of the
uncertain nature of the correlation existing between primai’y
and secondary sexual characters in general. It is not
necessarily the pi-esence of a differentiated gonad producing
some internal secretion which causes the development of the
corresponding secondary sexual characters, but it is the
potentiality to form that gonad. Thus the development of
the secondary sexual characters is not primarily dependent on
the gonad, but the development of both is dependent on a
third factor. If we attempt to formulate what this factor
actually is, it appears to me legitimate to represent it as the
presence of a substance having the nature of an internal
secretion, which circulates thi’ough the body and controls
the differentiation of the primary and secondary sexual
characters. I have called this hypothetical substance the
“sexual formative substance,” and we must suppose that two
kinds of it exist, male and female. By this theory we can
account for the imperfect nature of the correlation between
primary and secondary sexual characters, and also for the
development of the female secondary sexual characters in
infected male crabs before the development of an ovary,
which is unaccountable on the theory that the ovary produces
the substance necessary for the development of the secondary
character. s.

It is, however, a notorious fact that the mere removal of

600

GEOFI'EEY SMITH.

the gonad in the great majority of animals directly inhibits
the full development of the secondary characters, and it may
appear that the theory outlined above gives no explanation
of this fact. I clearly realised this in my first statement of
the theory, and put forward the suggestion that the -sexual
formative substance accumulated, especially at maturity, in
the gonad, and that the removal of the gonad removed a
large quantity of the substance and so inhibited the growth
of the secondary sexual characters. I do not feel, however,
that this explanation is at all adequate, principally for the
reason that the removal of the gonad in the young immature
animal has usually a more pronounced effect than its removal
in the adult. It is therefore more probable that the sexual
formative substance is in many cases actually worked up and
qualitatively altered by the gonad, and that its presence in
this altered state is essential in most cases for the full
development of all the sexual characters.

We may indeed hold, with the highest degree of probability,
that tlie sexual formative substance, both male and female,
is by no means a single simple substance, but that it consists
of numei’ous substances continually changing dui-ing develop-
ment, and both acting and acted on by the various organs of
the body. A view very similar to this is held by Mr. Walter
Heape, as the result of his experiments (16). He considers
that there is present a generative ferment” which is pro-
duced somewhere in the body and which governs the activity
of the generative glands, and another substance, “ gonadin,”
secreted by ovary or testis, which controls the other sexual
characters, but he is clearly of opinion that in certain cases it
may be the generative ferment which controls the secondary
sexual characters, and this would bring his view into close
agreement with my own.

The theory which has been outlined above, and which
differs from other theories chiefly in that it attempts to
include those cases in which the correlation between primary
and secondary sexual characters is of an uncertain and per-
plexing nature, has been attacked by Mr. Cunningham (14)

STUDIES IN THE EXPERIMENTAL ANALYSIS OP SEX. 601

who I’egards my views as illogical, self-contradictory, and
inconsistent with the state of modern biology ; indeed, he
lays so many charges to my account that modesty compels me
to suspect that some of them may be true ; but I can hardly
think that the difference between our respective views is
proportional to tlie severity of his indictment. The only
diffe rence of importance which I can discover is that, whereas
he believes that the internal secretion controlling the develop-
ment of the secondai-y sexual characters is always produced
by the differentiated gonad, I do not believe that this theory
covers all the essential facts, but that we must assume some
common factor at the back of both primary and secondary
characters which may act to a certain extent on either
separately, in an independent manner. On the other hand, I
have never denied the direct influence of the primary character
on the secondary to a limited extent, as it appears to me to
be clearly proved by a very large body of facts.

In conclusion, Mr. Cunningham agrees with me in believing
that the explanation of the development of the secondary
sexual characters, and of their correlation with the primary,
depends on the presence of a substance, sexual formative
substance or internal secretion, circulating in the body, which
in some manner activates the cells of various organs and parts
of the body and causes them to develop and to become
differentiated according to sex, and I think that he would
agree with me that a great deal of experiment and observa-
tion is necessary before we can decide with any certainty as
to the nature of this substance or substances.

That we are dealing with the presence of an internal
secretion is strongly suggested by the analogy of the internal
secretions produced by other organs, such as the thyroid and
other ductless glands. In the development of the sexual
characters we perceive distant parts of the body being
affected in a parallel manner at the same time, while a removal
of part of the system may profoundly modify other distant
parts. This inter-connection can only be accounted for in
one of two ways — either by the supposition that it is due to

602

GEOPi'REY SMITH.

nervous communication, or else by means of substances con-
veyed in the blood or body-fluids. The former supposition is
ruled out by a number of experiments, such as the severance
of the nerves to the reproductive organs, etc., so that we are
perforce thrown back on the second supposition of internal
seci’etious, although the participation of the nervous system
is not altogether precluded.

It would also seem probable that sexual difi^erentiation does
not solely depend on the presence and nature of these sub-
stances, but rather in the interaction of these substances
with the cells of the organism, which may themselves be
differentiated beforehand in the two sexes. The attempt to
analyse the nature of the sexual formative substance and its
relation to the primary and secondary characters will occupy
us in succeeding parts.

Literature.

1. Bateson, W. — Mendel’s ‘ Principles of Heredity,’ 1909.

2. Castle, W. — “ The Heredity of Sex,” ‘ Bull. Mus. Comp. Zool.

Harvard,’ vol. xi, 1903.

3. McClung, C. — “ The Accessory Chroniosoine — Sex Determinant ? ”

‘ Biol. Bull.,’ iii, 1902, p. 43.

4. Wilson, E. B. — “ Studies on Chromosomes,” ‘ Journ. Exp. Zool.,’

vols. ii and iii, 1906 and 1907, and other papers.

5. Smith, G. — ‘ Fauna and Flora des Golfes von Neapel,’ Monogr. 29,

1906.

6. Bateson, W., and Punnett, R. — “ The Heredity of Sex,” ‘ Science,'

N.S., xxvii, 1908, p. 785.

7. Doncaster, L. — “ Sex Inheritance in the moth, Abraxas grossu-

lariata,” ‘Rep. Evol. Comm.,’ iv, 1908.

8. Correns, C. — ‘Die Bestiniung und Vererbung des Geschlechtes,’

Berlin, 1907.

9. Heape, W. — “Note on the Proportion of the Sexes in Dogs,” ‘Proc.

Camb. Phil. Soc.,’ vol. xiv, 1907, and ‘ Phil. Trans. Roy. Soc.
London,’ vol. cc, 1908, p. 271.

10. Punnett, R. — “Sex Determination in Hydatina,” ‘Proc. Roy.
Soc. London,’ vol. Ixxviii, 1906, p. 223.

STUDIES IN THE EXPERIMENTAL ANALYSIS OP SEX. 603

11. Herbst, C. — ‘ Formative Reize,’ Leipzig, 1901.

12. Kellogg, V. — ‘ Journ. Exp. Zool.,’ vol. i, 1905, p. 601.

13. Meisenheimer, E. — ‘Verliandl. Dentscli. Zool. Gesellscb.,’ June,

1908, p. 84.

14. Cunningham, J. T. — “The Heredity of Secondaiy Sexual Cha-

racters,” ‘ Arch. Entwicklungs-mechanik. der Organismen,’ xxvi,

1908, p. 372.

15. Potts, F. A. — ‘ Quai-t. Journ. Micr. Sci.,’ vol. 50, 1906, p. 599.

16. Heape, W. — ‘Proc. Physiol. Soc.,’ December, 1905; ‘Quai’t. Joum.

Micr. Sci.,’ vol. 44, 1900, p. 1 ; ‘ Proc. Roy. Soc. London,’ Ixxvi,
1905, p. 260.

EXPLANATION OE PLATE 30,

Illustrating Mr. Geoffrey Smith’s paper on “ Studies in the
Experimental Analysis of Sex.”

In figs. 1, 3, 4, 7, 10 and 13 only the right chela is shown, the other
thoracic limbs being omitted.

All figures are of Inachus mauritanicus.

Fig. 1 . — Adult normal male.

Fig. 2. — Under-side of abdomen of normal adult male.

Fig. 3. — Male infected with Sacculina, showing reduction of chela
and slight broadening of abdomen.

Fig. 4. — Male infected with Sacculina, showing reduction of chela
and increased broadening of abdomen.

Fig. 5. — Under-side of abdomen of tig. 4, showing attenuated copu-
latory styles and slight hollowing-out of abdomen.

Fig. 6.— Under-side of abdomen of a similar male specimen, showing
reduction of copulatory styles and presence of asymmetrically placed
swimmerets, characteristic of female.

Fig. 7. — Male infected with Sacculina, which has assumed com-
plete female appearance.

Fig. 8. — Under-side of abdomen of fig. 7, showing much reduced
copulatory styles and reduced swimmerets.

Fig. 9. — Under-side of abdomen of a similar male specimen, showing
well-developed copulatory styles and swimmerets.

604

GEOFFREY SMITH.

Fig. 10. — Adult female, normal.

Fig. 11. — Under-side of abdomen of fig. 10, showing swimmei’ets
and trough-shaped abdomen.

Fig. 12. — Under-side of abdomen of female infected with Sacculina,
showing reduction of swimmerets.

Fig. 13. — Immature female, showing small flat abdomen.

Fig. 14. — Under-side of abdomen of fig. 13, showing flat surface
and rod-like swimmerets.

M =-

PHYSIOLOGY OF LAMELLIBRANCH BLOOD-CORPUSCLES. 605

Some Points in the Physiology of Lamellibranch
Blood-Corpuscles.

By

G. H. Drew, B.A.Cantab

With Plate 31.

Table op Contents.

PAGE

Introduction ...... 605

Historical ...... 606

Methods —

Collection and Preservation of Living Animals . . 608

Collection of Blood ..... 608

Fixing and Staining Methods .... 609

Methods of Observing the Process of Agglutination of the

Corpuscles . . . . .610

Methods of Studying Phagocytosis . . .611

Description of the Blood ..... 612

The Process of Agglutination and its Relation to the Healing of

Woimds ...... 614

Phagocytosis ...... 618

Bibliography . . . . . 620

Explanation of Plate .621

Introduction,

The investigations described in this paper were carried out
on the blood of Cardiuin norvegicum. This animal was
chosen as a type as it presents several features which render
it especially suitable for haematological work. Among these
may be mentioned the ease with which the blood can be
obtained in large quantities, the relatively large size of the

606

G. H. DREW.

corpuscles, the great vitality of the animal, and its large and
readily protruded foot.

One object of the present work was to investigate the
“ clotting ” of Lamellibrauch blood-corpuscles, and to follow
the relation this process bears to the natural cessation of
haeinorrhage from a wound and its subsequent healing. Another
object was to investigate the phagocytic action of the cor-
puscles on bacteria, and to find whether they showed any
chemiotactic action towards cultures of bacteria or extracts
of dead tissues.

My results show that some change takes place in the
corpuscles when the blood is shed, which causes them to
agglutinate round the edges of a wound, and that these
masses of corpuscles are connected by thin protoplasmic pro-
cesses running across tlie wound. These processes thicken
and contract, and so draw together the edges of the wound.
There is some evidence to show that the change in the cor-
puscles, which makes them agglutinate, is produced by a con-
tact stimulus imparted on contact with a foreign body or
with injured tissues. Some of the coi'puscles have a phago-
cytic action on bacteria, and show a positive chemiotactic
attraction towards cultures of bacteria and extracts of dead
tissues, so that their protective function appears to be the
same as that of the leucocytes of mammalian blood.

My thanks are due to the Marine Biological Association of
the United Kingdom for their kindness in granting me a
table at their Plymouth Laboratory.

Historical.

The chemistry of the blood of the Lamellibranchiata has
been thoroughly investigated by Ray Lankester (9, 10, 11,
and 12), Cuenot (2), Griffiths (8), and others, but a number of
observers have described the organised elements of the blood
with very varying results. Their conclusions are summarised
by de Bruyne (3). He himself recognises seven varieties of
corpuscles in Lamellibranch blood, taking as his types

PHYSIOLOGY OP LAMELLIBRANCH BLOOD-CORPUSCLES. 607

Mytilus edulis, Ostrea edulis, Unio pictorum, and
Anodonta cygnea. Cuenot (2) describes a “ glande
lympbatique ” at the base of the gills, where the corpuscles
originate, and recognises coarsely and finely granular cor-
puscles, and a third variety consisting of a nucleus with very
little surrounding protoplasm. He considers that these are
all derived by degenerative changes from the coarsely granular
form. In the case of Cardium norvegicum I agree with
Cuenot’s classification, but am inclined to consider that he
has scarcely published sufficient evidence to warrant the
statement that the other varieties are degenerated forms of
the coarsely granular corpuscles.

The fact that the plasma of Lamellibranch blood does not
coagulate has long been known, and Geddes (7), in 1879,
described the plasmodial masses formed by the agglutination
of the corpuscles in shed blood. Dakin (4), in his monograph
on Pecten, suggests that these plasmodia may act as plugs by
which haemorrhage from a wound may be checked, but does
not enter into the causes which determine this agglutination,
nor its function in the healing of wounds.

The question of phagocytosis has not been fully investigated.
Ray Lankester (11 and 12) first recognised the phagocytic
action of certain green amoeboid cells which he found on the
surface of the gills of “green” oysters, and later de Bruyne
(3) described the way in which certain wandering corpuscles
invade and destroy the epithelium of the gills, and finally
escape. De Bruyne attributes an excretory function to these
cells, and considers the “aqui poriferi,” which were formerly
supposed to communicate directly between the blood-spaces
and the surrounding water, to be due to erosions of the
epithelium caused by the emigration of these cells. In the
same paper he mentions the ingestion of carmine granules by
the phagocytes. No account has yet been published of the
phagocytic action of the corpuscles on bacteria, though,
owing to the fact that the blood as a whole does not coagu-
late, many difficulties in the way of such investigations are
eliminated.

608

G. H. DREW.

Methods.

Collection and Preservation of Living Animals.

Cardium norvegicum can be obtained in the neighbour-
hood of Plymouth by dredging on several grounds at a depth
of twenty to thirty fathoms. It was formerly quite plentiful,
but the supply has been falling off somewhat during the last
few years. The animal will live for many hours out of water,
and for some days if packed in damp seaweed. I have kept
a number for months in the Laboratory in basins into which
a small jet of sea-water flowed. Sufficient food is obtained
from the minute forms of life present in the water circulating
in the experimental tanks, and any artiflcial method of
aeration is unnecessary. The vitality of the animal is so
great that a relatively large volume of blood may be with-
drawn, and complete recovery ensue, in the course of a few
days, after which the blood appears to be normal in consti-
tuents and quantity.

Collection of Blood.

The blood can be most conveniently obtained from the
anterior adductor muscle. When the valves of the shell are
slightly apart, a small wedge is introduced between them to
prevent closing of the shell ; this usually causes the protru-
sion of the large and powerful foot, which is violently waved
about, and may displace the wedge unless it has been inserted
near the anterior adductor muscle, where it is beyond reach
of the foot. A clean, fine-pointed glass pipette, fitted with
a rubber teat, is then introduced between the fibres of the
adductor muscle, and the blood slowly withdrawn.

The following precautions should be taken :

The pipette should not be forced through the adductor
muscle so as to rupture any of the viscera ; it should pene-
trate about half-way through the muscle, and then be slightly
withdrawn to free the end.

PHYSIOLOGY OP LAMELLIBEANCH BLOOD-COEPUSCLES. 609

A pipette with a jagged or sharp point should not be used,
as this tends to cause agglutination of the corpuscles.

By this method I have obtained as much as 20 c.c. of blood
from a large specimen of Cardium norvegicum.

Fixing and Staining Methods,

By far the most satisfactory results were obtained by
simple fixation with corrosive sublimate, and staining with
aqueous eosin and methylene blue.

A drop of blood is allowed to fall on a slide from the
collecting pipette, and is then left in a moist chamber for from
half an hour to an hour to allow the corpuscles to expand.
Two or three drops of a saturated solution of corrosive subli-
mate in sea-water should then be added, and left for five
minutes. This is drained off, and the slide washed in 90 per
cent, alcohol. After this the slides may be placed in water
for a few minutes, and then stained for some time in a dilute
aqueous solution of eosin ; this is washed off, and Loefflei*’s
methylene blue, diluted 1 in 100, is added, and left for from
one to two minutes. The slide is finally washed in distilled
water, allowed to dry (not blotted), and mounted in xylol
balsam.

Almost equally good results were obtained by fixation with
osmic acid, but no other fixatives employed were really
satisfactory. No variety of the Romanowsky stain gave such
good results as treatment with eosin and methylene blue,
one after the other. Haematoxylin stained the nuclei well,
but interfered with the differentiation of the eosinophil
granules.

Treatment of the fresh blood with 1 per cent, acetic acid
and methylene green differentiates the nuclei of the corpuscles,
but fixation is not sufficiently rapid to enable the corpuscles
to be examined in the expanded condition.

Sections of the tissues showing wounds, etc., were fixed in
corrosive sublimate, and stained with Ehrlich’s haematoxylin
and erythrosin, or van Gieson’s stain.

610

G. H. DREW.

Methods of Observing the Process of Agglutination
of the Corpuscles.

The plasmodial masses, formed by agglutination of the cor-
puscles, can be produced by shaking the blood in a tube, or
by stirring a drop on a slide with a needle, and can then be
directly examined under the microscope.

To study the exact process by which they were formed
I employed the following method :

Two extremely thin strips of plasticine were placed on a
slide, inclined to each other at about an angle of 45°, and
arranged so as to leave a very narrow opening between their
convergent ends. A cover-slip was applied, and pressed firmly
down, so as to prevent the escape of liquid under the bands

of plasticine. A drop of blood can then be run under the
cover-slip, and can only escape from the cell at the narrow
end, though which it may be drawn with a piece of filter
paper. In some experiments a few strands of cotton-wool,
or glass-wool, were placed in the narrow opening, the fibres
being arranged as far as possible parallel to one another and
in the direction of the flow of the blood. Agglutination of
the corpuscles occurs readily as the blood passes through the
narrow end of the cell, and the process can be obsei’ved under
the microscope.

The way in which these agglutinated masses of corpuscles
close the opening of a wound, and so stop haemorrhage, was
studied by making small incisions in the foot when extended,
and then fixing by immersing the whole foot in saturated
corrosive sublimate, and finally cutting serial sections.

PHYSIOLOGY OF LAMELLIBEANCH BLOOD-COEPUSCLES. 611

Where the healing of a wound in later stages was followed,
the animal was narcotised with the foot in an extended con-
dition by a 1 per cent, solution of cocaine in sea-water; the
required portion of the foot was then cut off, and fixed, and
sectionised as before. In these experiments it is advisable
to keep the animals in separate basins with a good flow of
watei’, as if overcrowded, a number die, presumably from
infection through the wound.

Methods of Studying Phagocytosis.

The bacteria used for this purpose were obtained by inocu-
lating peptonised fish-broth, made with sea-water, with a
platinum loop which had been passed along the edge of the
mantle. This resulted in a mixed culture of bacteria, in
which a rather large non-motile bacillus with rounded ends,
and a tendency to form diplo-bacilli, much predominated.

The actual process of phagocytosis was observed by adding
a loopful of the diluted culture to a drop of blood. Permanent
slides, showing ingested bacteria, were prepared by leaving a
drop of blood, to which bacteria had been added, for from
one to two hours in a moist chamber, and then fixing and
staining by the same method as that employed in the prepara-
tion of stained blood-films.

Further observations were made by filling thin-walled
capillary tubes with cultures of bacteria, sealing them at one
end, and introducing the open end into a drop of blood on a
slide. A cover-glass with a small quantity of wax at each
corner was then placed on the drop, and the tube containing
the bacteria kept under observation.

Similar experiments were conducted by introducing capillary
tubes containing cultures, etc., into the adductor inuscle of
the animal, which may be replaced in the water and left for
some hours. The tubes were then withdrawn, and the number
of corpuscles which had entered the tubes were noted under
the microscope.

In these experiments the capillary tubes should be marked

VOL. 54, PART 4. NEW SERIES. 43

612

G. H. DREW.

with a diamond, broken clean across, and passed through the
flame, so as to obviate jagged ends, which tend to cause an
agglutination of the corpuscles around the end of the tube.

Description op the Blood.

The blood of Cardium norvegicum is a somewhat
opalescent fluid, appearing slightly yellow by transmitted,
and blue by reflected, light. On shaking, the corpuscles
stick together, forming small white floccular masses which
fall to the bottom of the liquid.

The plasma can be obtained free from the corpuscles by
filtering, and appears of tlie same colour as the blood. It
contains hmmocyaniu (Cuenot) (2). Its reaction to litmus is
neutral. On heating to 74° C. it becomes distinctly cloudy,
and in time a floccular precipitate of coagulated proteid is
produced, which is soluble in alkalies and acids, but is re-
precipitated on neutralisation. The addition of an equal
volume of 90 per cent, alcohol produces a white precipitate.
The plasma gives the usual proteid reactions, such as the
xanthoproteic and Biuret reactions, a brick-red precipitate
with Millon’s reagent, a white ring on the addition of nitric
acid (which does not disappear on warming), and a white
precipitate with potassium ferrocyanide and acetic acid.

An analysis of the blood for salinity, by titration with
silver nitrate, using potassium chromate as an indicator, gave
a chlorine value very slightly higher than that of the sea-
water in which the animal was living’. This small increase
was probably due to evaporation while collecting the blood,
boiling to remove the hmuiocyanin, and filtering. Without
very specialised apparatus it is almost impossible to go
through these operations without a small loss due to evapora-
tion.

Cuenot has estimated the total salts in the blood of a
number of marine invertebrates living in waters of various
salinities. He evapoi-ates and incinerates the blood, and
estimates the salts gravimetrically. By this method he

PHYSIOLOGY OP LAMBLLIBRANCH BLOOD-CORPUSCLES. 613

invai'iably found that the total salts in the blood was slightly
less than that in the water in which the animal lived, but his
published researches do not extend to any species of Cardium.

The organised elements in the blood of Cardium norve-
gicum consist entirely of amoebocytes. As seen in the freshly
drawn blood, they appear as slightly granular, colourless
corpuscles showing a number of short pseudopodia ; they
vary considerably in size, but the different varieties of the
corpuscles cannot be easily distinguished in unstained prepa-
rations, nor can the nucleus be clearly made out.

If a slide in which the cover-glass is supported at the
corners with wax, so as to ensure a fairly thick film of blood,
be kept under observation, it will be noticed that the cor-
puscles perform slow amoeboid movements. After the lapse
of about half an hour many of the corpuscles can be seen fully
extended, and may then measure three or four times their
diameter in the contracted state. In this condition they are
very thin and ti'ansparent, and can be most conveniently
observed under a nari*ow cone of illumination. The pseudo-
podia are often remarkably long and slender.

Stained preparations show that the corpuscles can be
divided into three classes :

(1) Finely granular eosinophil corpuscles (fig. 1). These
are the lai’gest corpuscles present. They possess a single
round or oval nucleus, and a relatively large amount of proto-
plasm, which contains a number of exti-etnely small eosinophil
granules, chiefly concentrated round the nucleus. The longer
pseudopodia are as a rule free from these granules at their
extremities, but there is usually a line extending round the
periphery of the corpuscle, which stains slightly with eosin.

(2) Coarsely granular eosinophil corpuscles (fig. 2), similar
to the finely granular variety, but slightly smaller, and
possessing large, well-defined, eosinophil granules.

(3) Basophil corpuscles (fig. 3). These are much smaller
than the two preceding varieties, and do not take on the
eosin stain at all. They possess a sing’le round nucleus, and
a very small amount of protoplasm, which also takes on the

614

(i. H. DREW.

blue stain. In fresli unstained preparations they can often
be distinguished by their small size, almost spherical shape,
and somewhat high power of refraction.

Differential counts gave the following as the average rela-
tive proportion of the corpuscles :

Finely granular eosinophil . . .48 per cent.

Coarsely granular eosinophil . . . 44 ,,

Basophil ...... 8 ,,

The Process of Agglutination of the Corpuscles, and its
Relation to the Healing of Wounds.

The small white floccular masses produced when the freshly
drawn blood is shaken in a tube, consist of a number of
corpuscles which have coalesced to form a compact mass ; the
individual corpuscles are still distinguishable, and those
round the periphery of the mass protrude pseudopodia. This
coalescence of the corpuscles also occurs when the fresh blood
is stirred with any foreign body, or when the corpuscles come
into contact with any rough surface. If the blood be care-
fully collected by a pipette with well-rounded ends, and
dropped on a slide, there will be comparatively little aggluti-
nation of the corpuscles, but if the pipette be at all dirty, so
as to present a rough surface to the blood, or if the end of
the pipette be at all jagged, there will be considerable
coalescence.

When the blood is allowed to flow through a meshwork of
cotton-wool, nearly all the corpuscles will stick to the strands
and form dense agglutinated masses; if glass-wool be employed
instead of cotton-wool, this result is not so marked.

It seems possible that this power of agglutination depends
on some change produced in the corpuscle by the stimulus of
contact or friction with a non-living body.

This agglutination can be studied more fully in the plasti-
cine cell already described. If the blood be drawn through
the small opening, a clot of corpuscles will soon be formed,
which will close the opening, and so prevent further escape

PHYSIOLOGY OP LAMELLIBRANCH BLOOD-CORPUSCLES. 615

of blood. Individual corpuscles can be watched as they near
the opening, and it will be seen that the majority touch the
bands of plasticine at least once before they adhere to its
surface. When the stream of blood is extremely slow, so
that the momentum of the corpuscles on contact is slight, a
corpuscle may touch any foi’eign body three or four times
before adhering, but if the momentum of the corpuscle be
greater, it may adhere on the first or second contact. In the
case of contact with a polished surface, such as glass, the
power of agglutination is not so marked. Once a corpuscle
has adhered, it possesses the power of sticking to any other
corpuscles that may come in contact with it, and these, in
their turn, can then adhere to others. That this power of
mutual adhesion is not possessed by the corpuscles in the
freshly drawn blood, is proved by the fact that corpuscles
may often be seen impinging without showing any tendency
to adhere, until one of them touches some foreign body, or
meets with other agglutinated corpuscles. Two corpuscles,
one or both of which have developed this power of aggluti-
tion, may frequently adhere to each other by their pseudo-
podia, and then become separated by the blood-current ; when
this occurs, a thin protoplasmic connection seems always to
remain between the corpuscles.

This phenomenon can best be studied by placing a few
strands of cotton-wool between the convergent bands of
plasticine. In this case, the corpuscles can be seen to form
agglutinated masses along the strands of cotton-wool, and
frequently thin protoplasmic connections are visible between
masses of corpuscles situated on different strands of cotton
(fig. 4). At first these bands may be so thin as to present
considerable difficulty in resolution, unless oblique illumina-
tion be employed. Even when invisible, their presence is
often shown by their power of arresting and adhering to
passing corpuscles. I have frequently followed the course of
two corpuscles, which had adhered by their pseudopodia, and
then become separated in the blood-stream, and settled down
on neighbouring strands of cotton-wool. In such cases a

616

G. H. DREW.

thin protoplasmic connection, often of remarkable length
relative to the size of the corpuscles, was either directly
visible, or was demonstrated by the adhesion of passing
corpuscles.

If two agglutinated masses of corpuscles, connected by one
or more such protoplasmic strands, be kept under observation
for several hours, it will be seen that the strands slowly
thicken. Corpuscles from each end may travel up a strand
and appear to become merged in it, as do also any fi’ee cor-
puscles which may have adhered to it. The corpuscles at
each end also send out pseudopodia along the strand, and
may be drawn up into it as it thickens.

During this process the strand contracts, increasing pro-
portionately in breadth. The force of this contraction is
often sufficient to draw together the two neighbouring fibres
of cotton to which the corpuscles have adhered, and by this
means the two original masses of agglutinated corpuscles
may finally be fused into one.

This phenomenon was studied as far as possible in life by
making small incised wounds in the foot when under water,
fixing after varying intervals, and sectionising. Unfortu-
nately the extremely delicate protoplasmic strands, formed
between adjacent masses of corpuscles in the earliest stages
of agglutination, did not withstand the fixing and embedding
process, but sections of a wound, that had been left for from
one to two hours before fixation, showed agglutinated masses
of leucocytes on the edges of the wound, with connecting
bands running in all directions, thus forming a plug which
would at least prevent the escape of corpuscles through the
wound, and probably much hinder the escape of the plasma.
Sections of wounds a few hours older show'ed the wound
completely blocked by the agglutinated corpuscles. Sections
of still older wounds showed that the process of healing in
many ways resembled that in Mammalia. The agglutinated
masses of corpuscles soon become more or less structureless,
and much resemble a mass of fibrin. This is then invaded
by other corpuscles which have a phagocytic action, and

PHYSIOLOGY UP LAMELLIBRANCH BLOOD-CORPUSCLES. 617

they are accompanied by connective-tissue corpuscles with
elongated nuclei, which form connective tissue. At the same
time, in the case of small incised wounds of the foot, the cut
muscle-fibres grow across, and the surface epithelium joins
up over the surface of the wound, so that after the lapse of
about three weeks the site of the wound may be almost
indistinguishable in sections.

I consider that the process as observed in the plasticine
cell is probably identical with that in Nature. When a wound
is made, the tissues along the immediate margins of the
wound have their vitality impaired, and though perhaps not
dead, are at least in an abnormal state. As the blood
escapes, the corpuscles impinge on these tissues and agglu-
tinate, and connecting strands of protoplasm may be formed
between masses of corpuscles on opposite sides of the wound,
in the same tnanner as that described when the blood is
drawn through cotton-wool. The subsequent thickening and
contraction of these bands of protoplasm would tend to draw
the edges of the wound together, and cause complete fusion
between the neighbouring masses of agglutinated corpuscles.

It is obvious that some change takes place in the corpuscles
of blood which has been withdrawn from the animal, which
confers on them the power of agglutination. Of the nature
of this change I have no evidence. Arguing by analogy, it
seems possible that it is due to the liberation of some enzyme
from the corpuscle. There is no visible change in the con-
tents of the cells after agglutination, and stained preparations
show that both the large and small eosinophil granules are
still present.

The effect of exposure to air, as a predisposing cause of
agglutination, may be eliminated by the fact that a small
wound, made under water, soon becomes plugged with agglu-
tinated corpuscles, and also by the fact that agglutination
occurs when the blood has been collected under water in a
narrow pipette already partially full of sea-water. This does
not dispose of the action of dissolved air, but it is reasonable
to assume that the blood during its passage through the gills

618

G. H. GREW.

contains approximately the same amount of dissolved air as
the surrounding water.

The corpuscles also aggdutinate in the case when the
animal, with the shell-valves wedged open, is washed with
distilled water, and the blood withdrawn with a dry pipette.
This disposes of the possibility of agglutination being caused
by admixture with sea-water. Mixture of the blood with
hypertonic or hypotonic salt solutions also does not hinder
agglutination.

'Phe possibility that some stimulus, conveyed to the corpuscle
by contact or friction with some foreign body, is the predis-
posing cause of agglutination, is suggested by the fact that
corpuscles can be seen to agglutinate after contact with a
foreign body, and more especially by the fact that the rapidity
of the change in the corpuscle appears to depend on its
momentum when impinging on the body. The fact that
agglutination does not occur so readily after contact with a
polished surface, such as glass, where there would be less
friction, is also in favour of this theory.

Phagocytosis.

Idle phagocytic action of the corpuscles on bacteria can be
watched by placing a di’op of blood on a slide, and adding a
loopful of a culture of bacteria in broth diluted with sea-
water. Tlie corpuscles can be seen to send out pseudopodia
in the direction of the bacteria, and engulf them. They may
then be fixed and stained as before (figs. 5 and 6). Agglu-
tinated corpuscles do not appear to possess this power, but
motile bacteria, in the course of their movements, may touch
and adhere to them; this is probably a purely passive action
on the part of the corpuscles. No phagocytic action on the
part of the basophil corpuscles was observed, nor did stained
preparations show that this had taken place. Experiments
were tried by introducing capillary tubes containing cultures
into a di'op of blood under a cover-slip, supported at the
corners by wax; in this case a certain number of corpuscles

PHYSIOLOGY OF LAMELLIBEANCH BLOOD-COEPUSCLES. 619

could usually be seen to enter the tube, and there was an
apparent concentration of the corpuscles about the mouth of
the tube after the lapse of from half an hour to an hour, but
though negative results were given in check experiments, in
which capillary tubes filled with sea-water were employed, yet
the number of corpuscles were usually so few (four to eight
on an average in half an hour), that the results cannot be con-
sidered conclusive. Much more conclusive results were given
by introducing similar capillary tubes into the anterior
adductor muscle, and leaving them there about two hours,
having inserted a small wedge between the valves of the shell
to prevent the breaking of the tubes. Tubes filled with the
following- fluids were employed, and all introduced at the same
time into the same animal :

(1) Culture of non-motile bacilli in peptonised fish-broth
made with sea-water.

(2) Peptonised fish-broth made with sea-water.

(3j Culture of the same bacilli in the blood of Cardium
norvegicum.

(4) Filtered extract of the tissues of Cardium nor-
vegicum which had been killed by heat and then minced in
sea-water.

(5) The fresh blood of Cardium norvegicum.

(6) Sea-water.

It was assumed that a positive result was given when over
twenty corpuscles were seen free in the tube, and a mass of
agglutinated corpuscles was found at the mouth of the tube
and extending a little way up it. On this assumption, jjositive
results were given with the culture in broth (1), the culture
in blood (3), and the extract of the tissues (4), and negative
results with the fresh blood (5), and sea- water (6). Varying
and inconclusive results were given with the sterile fish-
broth (2). These conclusions represent the mean of a large
number of experiments. The chief experimental errors, which
are liable to cause false results, are due to employing capillary
tubes which are rough at the open end ; in this case the
opening becomes rapidly closed by a mass of agglutinated

620

G. H. DREW.

corpuscles. Another source of error consists in using tubes
containing’ air at the closed end, when the fluid may be
expelled, or blood sucked in, as a result of changes of tempera-
ture. Even when the tube is completely full, time must be
allowed after sealing the end, for it to take up the room
temperature. The water in which the animal is kept should
also be at the I’oom temperature, and the tube should never
be touched with the hand, to avoid warming.

I consider that these experiments show that cultures of
bacteria and extracts of dead tissues have a positive chemio-
tactic attraction for the coi'puscles.

Bibliography.

A full and exhaustive bibliography of the subject up to 1896 is given
by De Bruyne in his “ Contribution a I’etude de la Phagocytose (1),”
published in the ‘ Arch, de Biol.,’ Tome xiv, pp. 231-236.

The following references only include those papers directly bearing
on the subject in hand.

1. Broun, H. G. — ‘Das Tier-Reich. MoUusca.,’ Abtlilg. ii, Leipsic,

1907, p. 604.

2. Cuenot, L. — “ Etudes sur le Sang et les Glandes Lymphatiques,”

‘ Arch, de Zool. Exper. et Gen.,’ Deuxieme Serie, Tome ix. Paris,

1891.

3. De Bruyne, C. — “ Contribution a I’etude de la Phagocytose (1),”

‘ Arch, de Biol.,’ Tome xiv, Paris, 1896, p. 161.

4. Dakin, W. J. — " Pecten.” ‘ Liverpool Marine Biological Committee

Memoirs,’ xvii, London, 1909, p. 73.

5. Frederique, L. — “ Sur rhemocyanine,” ‘ Comptes Rendus,’ Bd. 115,

Paris, 1892, p. 61.

6. von Fiirth. 0. — ‘ Vergleichende chemische Physiologie der neideren

Tiere,’ Jena, 1903, p. 60.

7. Geddes, P. — “ On the Coalescence of Amcfiboid Cells into Plasmodia,

and the So-called Coagulation of Invertebrate Fluids,” ‘Proc.

Roy. Soc.,’ vol. XXX, London, 1880.

8. Griffiths, A. B. — ‘ Respiratory Proteids,’ pp. 29-.S6, 50-67, 71-73,

London, 1897.

9. Lankester, E. Ray. — “Preliminaiy Notice of Some Observations

with the Spectroscope on Animal Substances,” ‘ J ourn. Anat. and

Phys.,’ Bd. 2, 1867, pp. 114-116.

PHYSIOLOGY OP LAMELLIBRANCH BLOOD-CORPUSCLES. 621

10. Lankester, E. Ray. — “Abstract of a Repoi’t on the Spectroscopic

Examination of Certain Animal Substances,” ‘ Journ. Anat. and
Phys.,’ Bd. 3, 1870, pp. 119-192.

11. “On Green Oysters,” ‘Quart. Journ. Micr. Sci.,’ vol. 26,

1886, p. 71.

12. “ Phagocytes of Green Oysters,” ‘ Nature,’ vol. xlviii, 1893,

p. 75.

EXPLANATION OF PLATE 31,

Illustrating Mr. Gr. H. Drew’s paper on “ Some Points in the
Physiology of Lamellibrauch Blood-Corpuscles.”

Fig. 1. — Finely gi’anular eosinophil corpuscles in expansion and con-
traction. Stained with eosin and methylene blue. X 750.

Fig. 2. — Coarsely granular eosinophil corpuscles in expansion and
conti'action. Stained with eosin and methylene blue. X 750.

Fig. 3. — Basophil corpuscles in expansion and contraction. Stained
with methylene blue. X 750.

Fig. 4. — Agglutinated masses of corpiiscles adherent to strands of
cotton fibre, showing thin coimecting bands of protoplasm, x 100.

Fig. 5. — Later stage of fig. 4, showing thickening and contraction of
the connecting bands of protoplasm. The cotton fibres have been
drawn closer together by the contraction, x 100.

Fig. 6. — Ingested bacteria in the corpuscles. X 750.

,^AJ.am. (Myu/r'rv a.iI/Ujco-: dau . iJoL oU, fl S. <l)%. C>I.

NOTE ON THE CYTOLOGY OF CALOTHEIX FESCA. 623

Note on the Cytology of Calothrix fusca.

By

Dr. !V. H. 8wclleng;rel>el,

Amsterdam.

With Plate 32.

While studying the cytolog}' of several TrichohacterinEe I
came across a representative of the group of Cyanophyceae, the
study of which may cast perhaps some light on the question
of relationship between Cyanophycese and Bacteria.

I found Calothrix fusca in aquaria among large quanti-
ties of Gloeocapsa ; I never found it not associated with those
algse. It seems not impossible that a symbiotic relationship
exists between those two algae, a relationship which would be
obviously beneficial to Calothrix, this species being more or
less deprived of chlorophyll. This question, however, must
remain for the Avhile a mere hypothesis, because I was not
able to study the question more thoroughly.

'I'he dimensions of the cells are very variable. At the end
of the cell-filaments the cells are rather short (from 3‘6 ju —
7‘2ju), but lengthen towards the base (from 7'8 /j — 10'8^);
their breadth is from 8-6 fi— 7-2 fi. The filaments which are
enclosed in thick hyaline sheaths are pseudo-ramified. Each
pseudo-ramification possesses at its base a heterocyst and
some concave cells.

To study the cytological details the cell -filaments were
fixed in Pfeiffer’s solution, washed in alcohol 60 per cent.,
after the ordinary passages through alcohol embedded in

624

N. H. SWELLENGREBEL.

paraffin (52°), and cut in sections from 4 — 6 fi thick. The
sections were washed in xylol and stained with iron-hema-
toxylin (Heidenhain).

I shall not enumerate here the different papei’S which have
been published about the cytology of Cyanophyceae. For
reference to them the very complete works of Kohl,^ Fischer,*
and Guilliermond ® may be consulted. According to Guillier-
mond, who recently studied various representatives of this
group, the central mass of stainable matter (the “ central
body” of Biitschli^) is composed of chromatin which is sup-
ported by an achromatic substratum of alveolar structure;
the whole central body is to be regarded as a primordial
nucleus, a view which was already held by Biitschli and his
followers. A contrary opinion is upheld by A. Fischer, who
does not believe in the nuclear nature of the central body ;
according to this author it represents only the central part of
the cytoplasma free from chlorophyll. The chromatophil
bodies within it consist of a peculiar hydrocarbon called “ ana-
bsenine.” His strongest argument against the chromatic
nature of those granules consists in the fact that they ai'e
dissolved in water. It must be remembered, however, that
this argument no longer holds good, since Oes ® showed that
the chromosomes of Spirogyra are equally dissolved in water.
Kohl (loc. cit.) showed that the chromatophil granules of
the central body give all the characteristic reactions of
true chromatin. I also have carefully examined the micro-
chemical reactions of those granules, and have arrived at the
same conclusion as Kohl’s. I do not describe them here, since
Kohl has done this in ex ten so. There cannot consequently
be the least doubt that the granules within the central body

' Kohl, ‘ tiber der Organisation luid Pliysiologie der Cyanophyceen,’
Jena, 1903.

" Fischer, ‘ Botan. Zeitimg,’ 1905.

’ Guilliermond, ‘ Revue general de botanique,’ 1907.

< Biitschli, ‘ fiber die Bander Cyanophyceen und Bacterien,’ Leipzig,
1826.

* Oes, ‘ Botan. Zeitimg,’ 1908.

NOTE ON THE CYTOLOGY OP CALOTHEIX FUSCA. 625

consist of real chromatiu. I also observed in the cells of
Calothrix fusca the metachroinatic granules (volutine
granules) ; they always were found iu the central body.

The cells of Calothrix fusca do not possess a well de-
veloped chromatophore, surrounding the central body, as is
found in other members of the Cyanophycem. Often the
cells are not at all green coloured, and when this is the case
the green colour is diffusely spread thi’oughout the cell
without a well-marked differentiation between coloured and
non-coloured cytoplasma. This is perhaps the reason why
the central body of Calothrix fusca is never so compactly
built as in other Cyanophycem, and why it becomes so easily
diffuse.

The cytoplasma of the young cells (at the end of the fila-
ments) contains few or no inclusions, and has an alveolar
structure (PI. 32, figs. 1 — o) which is not always distinctly
visible. The central body is generally of normal shape. It
is formed of an achromatic sub.stratum (which, however,
stains more deeply than the surrounding cytoplasma), in which
are embedded the chromatic granules and filaments. The
achromatic substratum determines the form of the central
body ; this form is rather variable, often the central body is
star-shaped, and resembles the same organ of Toly pot hr ix
lanata described by Kohl (loc. cit.). The chromatin is
not always distributed throughout the whole central body ;
often several parts of it are free from chromatin (PI. 32,
fig. 5). Such specimens are very favourable for the study of
the relation between the achromatic substratum of the central
body and the surrounding cytoplasma. Both have an alveolar
structure, and by carefully examining the places where cyto-
plasma and achi-omatic substratum come together, one can
often observe that the septa of the cytoplasmic alveoli are
continued without interruption into those of the achromatic
substratum, the only difference consisting in the different
avidity with which stains are absorbed (PI. 32, figs. 5, 6).

Cell division is performed in the ordinary way ; in the
middle of the cell an imperfect ring-shaped transverse mem-

626

N. H. SWELLEXGREBEL,

brane is formed, which becomes afterwards closed. The
central body divides by simple fissure, chromosome-like
masses of chromatin not being found as is the case in other
Cyanophycete (Kohl, Guilliermond).

Often it can be observed that the central body loses more
and more its ordinary shape. It becomes elongated with
more or less developed ramifications ; often a slight curvature
or zigzag form is to be observed (PI. 32, figs. 2, 6, 7, 8 c),
After carefully staining, one can always observe that the
central body is normally formed by its two components, the
chromatin and the chi’omatic substratum. In other specimens,
however, the distinction between cytoplasma and achromatic
substratum becomes more and more indistinct, and it is
impossible at last to trace a distinction between the two
(PI. 32, figs. 8 d, e, 9). The chromatin is in such cases spread
diffusely throughout the whole cell. The protoplasma of the
latter is built after the ordinary pattern. The cells resemble
very much those of some sporogenic bacteria, i*ecently de-
scribed by Guilliermond.'

The dissolution of the central body described here was also
observed by Guilliermond (loc. cit.) in Scytonema cin-
cinnatum,but it occurred there only in old vacuolated cells,
so it is highly probable that the dissolution was of a patho-
logic origin. This, however, cannot be the case in Calo-
thrix fusca, as very young cells (at the ends of young cell-
filaments) show already this phenomenon (PI. 32, fig. 8). In
grown-up cells there appear in the cytoplasma large hyaline
granules. They surround the central body at first, and
seem afterwards to invade the latter, so causing its dissolu-
tion. I was at first deluded by this phenomenon, thinking
that the dissolution of the centi’al body had a purely mecha-
nical cause, due to an auto-destruction by the formation of
the hyaline granules. But a closer observation made clear
that the diffusion of chromatin is equally found in cells which
are not provided with hyaline granules (PI. 32, figs. 8, 9), so

Guilliermond, ‘ Arch. f. Prot. kunde,’ 1908.

NOTE ON THE CYTOLOGY OF OALOTHRIX PUSOA. 627

the destruction of the central body has not a merely mecha-
nical cause.

I vainly tried to make out the chemical nature of these
hyaline granules. They are not identical with the “ cyano-
phycinkornchen ” of the Glerman authors, nor do they con-
sist of fat. They are only a little to be stained with eosine
and carbolic fuchsin, they are dissolved in diluted acids and
in pepsine, not in diluted alkalies.

The changes in cell-structure in the microtome sections
were controlled by the study of toto-stained preparations.
After fixation the cells were placed on a cover-glass, and were
stained and imbedded in the ordinary way. The normal
central bodies had the same aspect as in the sections (PI. 32,
fig. 10). The begitming of dissolution of the central body
was also very clearly to be seen (PI. 32, fig. 12) in these pre-
parations, also the cells with diffuse chromatin (PI. 32, fig. 11).
Generally the distinction between the protoplasma of the
central body and the surrounding parts was not very clearly
to be seen. Except this point the toto-preparations had the
same value as the sections.

I will now shortly discuss the results of the observations
described here. The normally built central body of Oalo-
thrix fusca contains chromatin granules imbedded in the
alveoli of the plasma of the central body (the “achromatic
substratum”). The latter is easily differentiated from the
cytoplasma ; the alveoli of the latter are the continuation of
those of the aclu’omatic substratum. I think therefore with
Guilliermondthat the central body of theCyanophycesemust be
regarded as a primordial nucleus, the difference of cytoplasma
and nuclear plasma already existing, but being not yet very
distinctly marked. Under certain circumstances, unknown
to me, the central body becomes at first irregularly shaped
(in this stage the central body resembles strikingly the
“ diffuse nuclei ” of Opalinopsis and Poettingeria), after
which the difference between cytoplasma and nuclear plasma
(s. achromatic substratum) disappears, and the chromatin
granules ai’e spread throughout the whole cell.

VOL. 54, PART 4. — NEW SERIES.

44

628

N. H. SWELLBNGREBEL.

The stages with diffuse chromatin resemble very much the
Bacteria with chromatic granules spread throughout the pro-
toplasma. There is not yet differentiation between cytoplasma
and nuclear plasma. Other forms of Bacteria, with their
chromatin condensed into a more or less compact central mass
(Sphaerotilus [SwellengrebeP], Bac. spirogyra [Dobell^]),
find their match in those forms of Calothrix f usca, where a
well-marked differentiation between cytoplasma and nuclear
plasma does not yet exist, but where the chromatin is no
longer spread throughout the whole cell, but takes a central
position (PI. 32, fig. 8 c). The stage with a well developed
central body is not yet found in the group of Bacteria.

Biitschli (loc. cit.) has already observed the resemblance of
the structure of Cyanophycem and larger Bacteria, the latter
showing a central agglomeration of chromatin suggesting a
central body. I think that the stages with diffuse chromatin
come much nearer to the structures described in Bacteria,
and that these observations may aid to support the view con-
cerning the relationship between Cyanophyceffi and Bacteria.

Hygienic Institute,

Amsterdam ;

June, 1909.

EXPLANATION OF PLATE 32,

Illustrating Dr. N. H. Swellengrebel’s paper “ Note on
the Cytology of Calothrix fusca.”

(Drawings made under a Zeiss 2 mm. homog. oil immersion apochromatic

comp. 6c. 18.)

Figs. 1 — 9. — Sections from 4 — 6 n thick.

Figs. 1 and 3. — Three cells showing a well-developed central body with
differentiation into chromatin and achromatic substratum.

* Swellengrebel, ‘ C. R. Soc. de biol.,’ Juin, 1908.

“ C. C. Dobell, ‘Quart. Journ. Micros. Science,’ vol. 53, May, 1909.

NOTE ON THE CYTOLOGY OF CALOTHEIX FUSCA. 629

Pigs. 2 and 4. — Central body in the act of becoming diffuse.

Pig. 5. — Showing the star-shaped achromatic part of the central body.

Pig. 6. — Idem. Achromatic part zigzag shaped.

Pig. 7. — As fig. 5, but achromatic substratum no longer visible.

Pig. 8. — Showing the different stages of dissociation of the central
body.

a, b. Normal central body.

c. Central body become diffuse. Achromatic substratum no longer
differentiated.

d, e. Complete dissolution of the central body. Chromatin in the form
of granules and filaments spread throughout the proto-
plasma.

Pig. 9.— Same stages as fig. 8d and 8e.

Pigs. 10 — 12. — Preparations of cell-filaments stained in toto.

The figures show the same peculiarities as the microtome sections.

THE “AEGHENTEEIC KNOT ” OF OENITHOEHYNCHUS. 631

Tropidonotus and the “Archenteric Knot ” of
Ornithorhynchus.

By

Kicliai'd Asslietoii, M.A.

With Plate 33.

In my paper in a recent number of this Jouimal, in which
I discussed Professor Hubrecht’s memoir upon the ontogenetic
phases of mammalia, I referred to the peculiar condition of
the egg of Ornithorhynchus as described by Wilson and Hill
in the ' Philosophical Transactions of the Eoyal Society,’
vol. cxcix, and tlie interpretation placed by them thereon,
A remarkable spot found by these authors and named by
them “primitive or archenteric knot” exists at an eaidy stage
of the blastocyst of Ornithorhynchus, a stage before the
upraising of the neural folds, at some distance in front of
tlie primitive streak which is present in a form perfectly
typical of mammalia.

The whole blastocyst at this period is “occupied by mainly
fluid contents,” the more solid yolk of the previous stage
(6 mm. in diameter) having become partly disintegrated by
absorption of fluid from the uterus by this time, when the
diameter of the blastocyst has attained 10 mm,

Wilson and Hill regarded this spot as representing an early
stage in the development of the archenteron, and attempted
to identify it with the “Hensen knot” of the later period.
Considerable gaps exist in the material, so that they were not
able to trace accurately either the origin or the fate of this
structure.

632

RICHARD ASSHETON.

I ventured to suggest that this spot had nothing to do with
archenteron formation, and that it might be a quite erroneous
conclusion to identify it with the anterior end of the primitive
streak of later times. I argued at some length to show from
their description that their interpretation is not tenable. As
an alternative I offered the suggestion that the spot in question
is the morphological lower pole of the egg, and gave figures
illustrating a comparison between Ornithorhynchus as de-
scribed by Wilson and Hill and a Sauropsidan, such as the
sparrow, postulating a complete growth round by the edge of
the blastoderm at an earlier stage in the Prototherian than in
the Sauropsidan egg in correlation with a smaller quantity of
yolk. The fact that there is a very well-marked eccentricity —
I mean that this spot is not diametrically opposite to the
centre of the “embryo” in Ornithorhynchus as it is in the
sparrow, but as it is not in the rabbit — did not seem to me to
be a serious objection.

Since making this criticism I have come across a series of
sections which I cut through the lower pole of the egg of
Tropidonotus natrix some years ago, but which I had
forgotten — at a stage represented by the outline drawing
(PI. 33, fig. 1).

I find I have also in toto the same spot indicating the
coalesced edges of the blastoderm of another egg of the same
snake, and the appearances of these specimens are such as to
add very considerably to the degree of probability that the
suggestion I made will turn out to have been well founded.

The resemblance between the drawings of sections taken
through the centre of this area (fig. 2) and Wilson and Hill’s
text-fig. 4 (p. 51) of their section through the archeuteric
knot of Oniithorhynchus is most striking. It must be
remembered that at this stage in the snake’s egg the “ blasto-
cyst cavity” is still nearly filled with yolk, which is not
shown in my drawing, as the loose yolk has been all washed
off during the preparation of the specimen, this loose yolk
corresponding to the “tolerably large remainder of the
original yolk as a more or less coherent mass, lying free within

THE “ARCHENTEEIC KNOT ” OE OENITHOEHYNCHUS. 633

the cavity of the vesicle of the Ornithorhynchus egg of
10-12 mm, in diameter (p. 42).

Fig’. 2 is the 239th section of the series of 440 sections
passing through the spot, and so is nearly central.

This shows that the lower pole of the egg of Tropidouotns,
after the yolk has become completely enveloped by the edge
of the blastoderm, consists of the following parts : On the
outside there is the epiblast (ep.), a thin layer, except at the
centre where its free edge has concentrated forming the
dense mass of cells (c. p.). Beueath this there is, towards
the periphery, the hypoblast [hy.), consisting of a reticulum
containing many yolk-spherules {y. y.), and small nuclei on
the surface next the epiblast. This hypoblast layer is seen
to be much thicker near the centre of the figure, and a few
more deeply placed nuclei maybe found. This is comparable
to that part which, in the chick, was termed by Balfour “ the
germinal wall” [g.w.]. Beneath the plug of epiblast {c. p.)
is a deeply staining and very finely granular matei’ial (y.),
which is the just-covered superficial layer of the lower pole
of the yolk. Even as it is, the resemblance between this
structure and the so-called “ archenteric knot of Oruitho-
rhynchus” is sufficiently marked to cause one to view with
suspicion the interpretation placed upon that spot by Wilson
and Hill. But if we imagine a slightly more advanced con-
dition, if we imagine the coalescence of the germinal wall,
either by a gradual closing of the ring or by a differentiation
of the smalt remaining piece of pui’e yolk into germinal wall,
the resemblance between the two structures would be even
more marked.

In fig. 3 I have drawn a diagram from this figure and
adopted the same method of indicating the layers as Wilson
and Hill use in their text-fig, 4, p. 51, using the same
lettering but attaching to them my interpretation. Thus ect.
is the epiblast or ectoderm in each case, ent. the hypoblast.

The epiblast {ect.) is obviously in continuation with the
cell-plug (c.p.), which is seen in Tropidonotus to be the
thickened coalesced margin of the epiblast of the blastoderm

634

lUCHARD ASSHETON.

The part which pi’ojects above the surface, the actual cell-
plug, is probably partly due to a growth of the epiblast cells
after the coalescence of the margin, as I have found certainly
one mitotic figure within this mass. In some sections, e. g.
fig. 4, the distinction into “ cell-plug ” (c. j>.) and “ central
more cellular zone ” (c. z.) is still more marked than it is in
fig. 2.

The part called by Wilson and Hill the “marginal or
cortical zone of the knot tissue ” (m. z.) is represented by the
germinal wall part of the hypoblast of Tropidonotus, which,
like the author^s marginal zone, can be well described as
showing “ coarsely reticular, indefinite and feebly staining
characters,” and being “ poor in nuclei which are chiefly met
with near its entodermal aspect.”

Fig. 4 is another section of the same specimen which passes
through a little cleft still remaining, which shows the relations
of the thin epiblast to the thickened rim very clearly.

It is difficult to resist the conclusion that a condition
similar to this has given rise to the state of affairs in Orni-
thorhynchus, described by Wilson and Hill and indicated in
their text-fig. 5, p. 52, and figs. 9 and 10, pi. 4.

If my comparison is a correct one, the archenteric knot of
Ornithorhynchus with its anterior and posterior lips of the
blastopore and its “commencement of true archenteric in-
vagination ” may be dismissed, and another stumbling-block
will be removed from the path of the student of mammalian
embryology.

As regards the question of the eccentricity of this spot in
Ornithorhynchus I have no further evidence to bring forward.
I have no note on this point and cannot recollect whether
tlie point of coalescence of the blastoderm is diametrically
opposite to the mid-dorsal surface or not. The egg of Tropi-
donotus is long. Even if the point of coalescence in the
snake’s egg is exactly opposite to the upper pole I do not think
that it invalidates in any way the argument from the morpho-
logical point of view.

Whether, if this is the correct interpi’etation, it supports

THE “ AECHEINTEEIC KNOT ” OF OENITHOEHYNCHUS. 635

or opposes tlie view that the Prototherian egg is derived
from an egg of the Sauropsidan type may be open to question.
There can be very little doubt that, if such has been its
origin, there would be a condition as suggested. But it
would be hazardous to assert that those are the only circum-
stances under which such a condition could have arisen.

One can only say that the condition in all essential features
is identical with that of the Saui’opsidan type, and is totally
unlike that of Amphibia, Teleosteans, Dipnoi, Elasmobranchs,
or Cyclostomes. To my mind it appears to form strong
evidence in confirmation of the validity of the association of
the Reptilia, Aves, and Mammalia in one distinct group —
the Amniota.

Fig. 5 shows the cellular character of the cell-plug, the
thin general layer of epiblast continuous with the thickened
margin, and the hypoblast and yolk-spherules, many dis-
integrated, foi’ming the germinal wall or marginal zone of
Wilson and Hill.

Aros,

Isle of Mull,

August, 1909.

Papers referred to in the Test.

1. Assheton, R. — “ Professor Hubreclit's Paper on the Ontogenetic

Phases of Mammalian Development ; An Appreciation and
Respectful Criticism,” ‘ Quart. Journ. Micr. Sci.,’ vol. 51, 1909.

2. Wilson, J. T., and Hill, J. P.— “ Primitive Knot and Early Gastru-

lation Cavity Coexisting with Independent Primitive Streak in
Oniithorhynchus,” ‘ Proc. Roy. Soc. London,’ vol. Ixxi, 1903.

— “ Observations on the Development of Ornithorhyn-

chus,” ‘Phil. Trans. Roy. Soc. London,’ ser. B, vol. cxcix, 1907.

8.

636

EICHAED ASSHETON.

EXPLANATION OF PLATE 33,

Illustrating Mr. Eichard Asslieton’s paper on “ Tropidonotus
and the ‘ Arclienteric Knot ’ of Ornithorhynchus.”

Fig. 1. — Outline figure of Tropidonotus embryo, showing three gill-
cdefts, allantois, etc. all. Allantois, lit. Heart.

Fig. 2. — Transverse section theough the central region of the lower
pole of the egg of Tropidonotus after complete envelopment of the yolk
liy the blastoderm of the stage of fig. 1. The loose yolk has been
washed away, leaving hypoblast and germinal wall as a thickening
round the fused edge of the epiblast. c. p. Heaped-up rim of coalesced
ei^iblast. e.p. Epiblast. l\y. Hypoblast with much yolk. (j. w. Genninal
wall. y. Pure yolk not yet covered by, or converted into gemiinal
wall.

Fig. 3.^ — Diagram of the above, lettered and drawm after the manner
of Wilson and Hill's text-fig. 4, with which it should be compared,
c.p. Cell-plug of ectoderm. ecA Ectoderm. e«t. Endoderm. c.z. (Central
zone) part of central phig of ectoderm obviously in continuity with the
ectoderm, m. z. (Marginal or cortical zone) -- germinal wall, or hypo-
blasts and yolk. y. Yolk.

Fig. 4. — Another section of the same series of transverse sections
through the central region of the lower pole of Tropidonotus. The
section passes through a little cleft still remaining between the edges
of the coalesced blastodenn rim. c. p. Plug of cells derived from the
coalesced rim of the blastoderm, e. p. Epiblast. hy. Hypoblast, g. w.
Germinal wall. y. Yolk.

Fig. 5. — A portion of the section shown in fig. 4, to illustrate the
cellular character of the “ cell-plug ” (c. p.) (coalesced blastoderm edge),
and e.p. epiblast, hy. germinal wall (hypoblast).

Ik .A

k- -ik

Sofj/rmt.oMijr/r' S<x. Vol. 54-,NS. <0^.33.

4

RiC.Afl^Leton del

INDEX TO YOL. 54,

NEW SERIES.

Amphioxus, excretory organs of, by
Goodrich, 185

„ formation of the layers in, by

E. W. MacBride, F.R.S., 279
Aranese, development and origin of

the respiratory organs in, by W.

F. Purcell, Ph.D., 1

,, phylogeny of the tracheae in, by
W. F. Purcell, Ph.D., 519
Arnold on digestive processes in
Planariae, 207

Assheton, R. J., a criticism of
Hubrecht’s paper on early onto-
genetic phenomena in mammals,
221

„ on Tropidonotus and the Archen-
teric knot of Ornithorhynchus,
631

Blood-corpuscles of Lamellibranchs,
by Drew, 605

Calothrix fusca, cytology of, by
Sw ellengrebel, 623
Cyanophyceae, Swellengrebel, on
Calothrix, a representative of that
group, 623

Dendrosoma radians, bySydney
J. Hickson, D.Sc., 141

Digestive processes, intra-cellular
and general, in Planariae, by
I Arnold, 207

Dinophilus, nephridia of, by Good-
rich, 1 .1

I Drew, G. H., on the physiology of
Lamellibranch blood-corpuscles,
605

Echiurus, nephridia of, by Good-
rich, 111

Goodrich, E. S., on the structure of
the excretory organs of Amphi-
j oxus, 185

I „ on the nephridia of Dinophilus
I and of the larvae of Polygordius,
Echiurus, and Phoronis, 111
j Gregarinae, Margaret Robinson on
I Kalpidorhynchus, a representative
of that grouj), 565

I Haswell, Professor, on the develop-
I ment of the Temnocephaleae, 415
j Hewitt, Gordon, on the Bionomics,
Allies, Parasites and relations to
' disease, of Musca domestica,
347

I Hickson, S. J., on Dendrosoma
radians, 141

638

INDEX.

House-fly, Part III, by Gordon j
Hewitt, D.Sc., 347

Hubrecht, criticism of, by Assheton,
221

Kalpidorhynchus, reproduction of,
by Margaret Robinson, 565

Kerr, Professor J. Graham, on the
development of the alimentary
canal in Lepidosiren and Pro-
topterus, 483

Lamellibranch blood-corpuscles,
by Drew, 605

Lepidosiren, alimentary canal of, by
Professor Graham Kerr, F.R.S.,
483

MacBride, E. W., on the ,‘ormation
of the layers in Amphio; ns, 279

Mrisca domes tica, Pait III, by
Gordon Hewitt, D.Sc., 347

Musgrave, Edith M. (nee Pratt), on
Pennatulids, 443

Nephrididium of adult and tarsal
Amphioxus, by Goodrich, 185
„ of Dinophilus and of larvae of
Polygordius, Echiurus, and Pho-
ronis, by E. S. Goodrich, F.R.S.,
111

„ Hatschek’s, in Amphioxus, by
Goodrich, 185

Ornithorhynchus, archenteric knot
of, by Richard Assheton, M.A.,
631

Pennatulids, experiments on the
organs of circulation and locomo-
tion of, by Edith M. Musgrave
(nee Pratt), 443

Phoronis, nephridia of, by Goodrich,
111

Planariae, intra-cellular digestive
processes in, by G. Arnold, 207

Polygordius, nephridia of, by Good-
rich, 111

Pontobdella, trypanosome found in,
by Muriel Robertson, 119

Pratt (E. M. Musgrave), on Penna-
tulids, 443

Protopterus, alimentary canal of, by
Professor Graham Kerr, F.R.S.,
483

Purcell on the development and
origin of the respiratory organs in
Araneae, 1

„ on the phytogeny of the tracheae
in Araneae, 519

Robinson, Margaret, on the repro-
duction of the Gregarine Kal-
pidorhynchus araneae, 565

Robertson, Muriel, on a Trypano-
some found in the alimentary
tract of Pontobdella, 119

Sex, studies in the experimental
analysis of, by Geoffrey Smith,
577

i Smith, Geoffrey, on the experimental
analysis of sex, 577

I Swellengrebel on the cytology of
Calothrix, one of the Cyanophyceae,
623

I Temnocephaleae, development of the,
by Professor W. A. Haswell,

I F.R.S., 415

Tracheae of Araneae, phytogeny of,
by W. F. Purcell, Ph.D., 519

Tropidonotus and the archenteric
knot of Ornithorhynchus, by
Richard Assheton, M.A., 631

Trypanosome in the alimentary
tract of Pontobdella muri-
cata, by Muriel Robertson, 119

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BEYER KKOKBSSOK OK ZOOLOGY IN TUP. UNIVERSITY O MANCIlESTKk;

AND

E. A. MINCHIN, M.A.,

PROFESSOR OK PROTOZOOLOGY l.N THE UNIVERSITY OK LONDON.

WITH LITHOGRAPHIC PLATES AND TEXT-FIGURES.

LONDON.

J. A A. CHUKCHILL, 7 GREAT MARLBOROUGH STREET.

1909.

.\dlard and Son, Impr.,]

[London and Dorking.

CONTENTS OF No. 213.-New Series.

MEMOIRS :

PAGE

Development and Origin of the Eespiratory Organs in Arane®. By
W. F. Purcell, Ph.D., Bergvliet, Diep Eiver, near Cape Town.
(With Plates 1—7, and 7 Text-figures) . . . 1

Notes on the Nephridia of Dinophilus and of the Larv® of Poly-
gordius, Echiurus, and Phoronis. By E. S. Goodrich, F.E.S.,
Fellow of Merton College, Oxford. (With Plate 8) . .Ill

Further Notes on a Trypanosome found in the Alimentary Tract of
Pontohdella muricata. By Muriel Eobertson, M.A., Car-
negie Eesearch Fellow ; Junior Assistant to the Professor of
Protozoology in the University of London. (With Plate 9, and
5 Text-figures) ...... 119

New Series, No. 214 (Vol. 54, Part 2). Price lOs. net.

Subscription per volume (of 4 parts) 40s. net.

OCTOBER, 1909.

THE

QUARTERLY JOURNAL

OF

MICROSCOPICAL SCIENCE.

EDlTRh HY

Sir ray LANKESTER, K.C.B., M.A., D.Sc., LL.D., F.R.S.,

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FOREIGN ASSOCIATE OK TUB NATIONAL ACADEMY OK SCIENCES. U>8.> AND MEMBER OK TUB
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PHYSIOLOGY IN TUR ROYAL INSTITUTION OP OBRAT BRITAIN ;

LATR LINACHR PROPRSSOR OF COMPARATIVE ANATOMY AND FELLOW OF MERTON COI.f.R GR , OX PU R D ;
RMERITU8 PROFESSOR OF ZOOLOGY AND COMPARATIVE ANATOMY IN UNIVERSITY COLLEGE, UNIVERSITY OF LONDON.

WITH THE CO-OPERATION OP

ADAM SEDGWICK, M.A., F.R.S.,

PROFESSOR OP ZOOLOGY AND COMPARATIVE ANATOMY IN THE UNIVERSITY OF CAMBRIDGE

SYDNEY J. HICKSON, M.A., F.R.S.,

BEYER PROFESSOR OK ZOOLOGY IN THE UNIVERSITY OF MANCHRSTKR;

AND

E. A. MINCHIN, M.A.,

PROFESSOR OF PROTOZOOLOGY IN THE UNIVERSITY OK LONDON.

WITH LITHOGRAPHIC PLATES AND TEXT-FIGURES.

LONDON.

J. & A. CHURCHILL, 7 GREAT MARLBOROUGH STREET.

1909.

Adlanl and Son, Impr.,]

[London and Dorking.

CONTENTS OF No. 214.-New Series.

MEMOIRS :

PAGE

Dendrosoma radians, Ehrenberg. By Sydney J. Hickson, D.Sc.,

F. E.S., Beyer Professor of Zoology in the University of Manchester,

and J. T. Wadsworth. (With Plate 10) . . . 141

On the Structure of the Excretory Organs of Amphioxus. Part 2. —

The Nephridium in the Adult. Part 3. — Hatschek’s Nephridiuni.

Part 4. — The Nephridium in the Larva. By Edwin S. Goodrich,
P.R.S., Fellow of Merton College, Oxford. (With Plates 11 — 16
and 1 Text-figure) ...... 185

Intra-cellular and General Digestive Processes in Planariae. By

G. Arnold, from the Cytological Laboratory of the University of
Livei-pool. (With Plate 17) ..... 207

Professor Huhrecht’s Paper on the Early Ontogenetic Phenomena
in Mammals ; an Appreciation and a Criticism. By Richard
Assheton, M.A., Trinity College, Cambridge ; Lecturer on Biology
in the Medical School of Guy’s Hospital, in the University of
London. (With 5 Text-figures) .... 221

New Series, No. 215 (Vol. 54, Part 3). Price 10s. net.

Subscription per volume (of 4 parts) 40s. net.

1909.

JOURNAL

MICROSCOPICAL SCIENCE.

EUlTKh KY

I Sir ray LANKESTER, K.C.B., M.A., D.Sc., LL.D., F.R.S.,

I HOr«UMAtlT VBLLOW or BXKTBR C OLI.R GB , OX KO K D ; C<' K R E8 P UN D B X T OK TUB INSTITUTB OK KHANCB*

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I OP 8CI8NCB8 OF FBILADEI.PIIIA, AND OP THE ROYAL ACADEMY OP SCIENCES

I OK TURIN; koreign member ok tub royal society op sciences op

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I OP TUS ACADBMY OP THE MNCEI OP ROME« AND OP TUR AMERICAN

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LATE DIRECTOR OP TUB NATURAL HISTORY DBPARTMBNT8 OP TUB BRITISH MU8KU.M; LATE PRESIDENT OP THE
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WITH THE CO-OPEUATION OF

ADAM SEDGWICK, M.A., F.R.S.,

FELLOW OP TRINITY COLLEGE, CAMBRIDGE. AND PKOPP.SSOR OP ZOOLOGY IN TUB IMPERIAL COLLEGE OP
SCIENCE AND TECIINOLOGY, LONDON.

SYDNEY J. HICKSON, M.A., F.R.S.,

BEYER PROFESSOR OP ZOOLOGY IN THE UNIVERSITY OP MANCIIKSTKK’,

AND

E. A. MINCHIN, M.A.,

PROFESSOR OP PROTOZOOLOGY IN THE UNIVERSITY OP LONDON.

WITH LITHOGRAPHIC PLATES AND TEXT-FIGURES.

LONDON.

J. & A. CHURCHILL, 7 GREAT MARLBOROUGH STREET.

1909.

DECEMBER,

THE

QUARTERLY

OF

Afllard and Son, Impr.,]

[London and Dorking,

CONTENTS OF No. 215.-New Series.

MEMOIRS :

PAGE

The Formation of the Layers in Aniphioxus, and its bearing on the
Interpretation of the Early Ontogenetic Processes in other Verte-
brates. By E. W. MacBride, D.Sc., LL.D., F.R.S., Strathcona
Professor of Zoology in McGill University, Montreal. (With
Plates 18—21, and 10 Text-figures) .... 279

The Structui'e, Development, and Bionomics of the House-fly, Musca
domestica, Linn. Part III. — The Bionomics, Allies, Parasites,
and the Relations of M. domestica to Human Disease. By

C. Gordon Hewitt, D.Sc., Late Lecturer in Economic Zoology,

University, of Manchester. (With Plate 22) . 347

The Development of the Temnocephalese. Part I. By Professor
W. A. Haswell, M.A., D.Sc., F.R.S. (With Plates 23 — 25) . 415

Experimental Observations on the Organs of Circulation and the
Powers of Locomotion in Pennatulids. By Edith M. Mdsorave,

D. Sc. (nee Pratt), Late Honorary Research Fellow in the Univer-
sity of Manchester. (With Plates 26 and 27) . . 443

New Series, No. 216 (Vol. 54, Part 4). Price lOs. net.

Subscription per volume (of 4 parts) 40s. net.

FEBRUAEY, 1910.

THl^

QUARTERLY JOURNAL

OF

MICROSCOPICAL SCIENCE.

KDIIKI* HY

Sje ray LANKESTER, K.C.B., M.A., D.Sc., LL.D., F.R.S.,

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CORRKSPUNDING MEMBER OF THE SRNKKNBEKG ACADKMY OP K R AN K FU RT*A- M ;

FUKKIGN ASSOCIATK OF TUK NATIONAL ACADKMY OF SCIENCES. U.8.. AND MEMBER UK TUB
AMERICAN rilll.OSOPIIlCAI. SOCIEl Y ;

HONORARY KELLOW OF THE ROYAL SOCIETY OF EDINBURGH;

LATE lilllRrtOH OK TUK NAtl'RAL IliSTUllY DRPARTMKNTS OK THE BRITISH MUSKUM : LATE PRRSIDENT OF THE
BRITISH ASSOCIATION FOR THE ADVANCEMENT OF SCIENCE.’ l.ATK Kt'l.LKKIAN PUUKKS80R OF
FIIYSIULOGY IN THE ROYAL INSTITUTION OF QRRAT BRITAIN ;
l.ATK MNACRK FROFKRSOR OF COMKAHATIVE ANATOMY AND FEM-OW op MERTON C O LLF GE , O X FO R l> .
EMERITUS PROFESSOR OF ZOOLOGY AND COMPARATIVE AXATOMY IN UNIVERSITY COLIEGE. UNIVERSITY OK LONDON

WITH THK CO-OPEUATION OP

ADAM SEDGWICK, M.A., F.R.S.,

FELLOW OK TRINITY COLLEGE. CAMBRIDGE. AND PROFESSOR OK ZOOLOGY IN TUB IMPERIAL COLLEGE OK
SCIENCE AND IBCUNOLOOY, LONDON.

SYDNEY J. HICKSON, M.A., F.R.S.,

BKTRR PKOKRSSOK OK ZOOLOGY IN TUK UNIVERSITY OK MANCMKsrKR;

AND

E. A. MINCHIN, M.A.,

PKOKESSOn OK PROT OZOOLOGY IN TUB UNIVERSITY OK LONDON.

WITH LITHOGRAPHIC PLATES AND TEXT-FIGURES

LONDON.

& A. OHUllOHlLL, 7 GEEAT MAKLBOROUGH STREET.

1910.

Adlanl and Son, Impr.,]

[London and Dorkin]

CONTENTS OF No. 216.-New Series.

MEMOIRS

PAGE

On Certain Features in the Development of the Alimentary Canal in
Lepidosiren and Protopterus. By J. Graham Kerr, Pro-
fessor in the University of Glasgow. (With 13 Text-figures) 483

The Phylogeny of the Tracheae in Araneae. By W. F. Purcell,
Ph.D., Bergvliet, Diep River, near Cape Town. (With Plate 28,
and 21 Text-figures) ..... 519

On the Reproduction of Kalpidorhynchus arenicolae (Cnghm.).

By Margaret Robinson, University College, London. (With
Plate 29) . . . . . . .565

Studies in the Experimental Analysis of Sex. By Geoffrey Smith.
(With Plate 30) ...... 577

Some Points in the Physiology of Lamellibranch Blood-Corpuscles.

By G. H. Drew, B. A. Cantab. (With Plate 31) . . 605

Note on the Cytology of Calothrix fuse a. By Dr. N. H. Swel-
LENGREBEL, Amsterdam. (With Plate 32) . . 623

Tropidonotus and the “ Archenteric Knot ” of Ornithorhynchus. By
Richard Assheton, M. A. (AVith Plate 33) . . 631

Title, Index, and Contents.

With Ten Plates, Royal 4to, 5s.

CONTRIBUTIONS TO THE KNOWLEDGE OF RHABDOPLEURA
AND AMPHIOXUS.

By E. EAY LANKESTER, M.A., LL.D., F.R.S.

London : J. & A. Chuechill, 7 Great Marlborough Street.

Quarterly Journal of Microscopical

Science.

The SUBSCiilP'iTON is £2 for the Volume of Four Numbers;
for this sum (prepaid) the Journal is sent Post Free to any part
of the world.

BACK NUMBERS of the Journal, which remain in print, are
now sold at an uniform price of 10/- net.

The issue of Supplkment Numbers being found inconvenient,
and there being often in the Editor’s hands an accumulation of
valuable material, it has been decided to publish this Journal at
such intervals as may seem desirable, rather than delay the appear-
ance of Memoirs for a regular quarterly publication.

The title remains unaltered, though more than Four Numbers
may be published in the course of a year.

Each Number is sold at 10/- net, and Four Numbers make
up a Volume.

London : J. & A. CHURCHILL, 7 Great Marlborough Street.
TO CORRESPONDENTS.

Authors of original papers published in the Quarterly .Journal
of Microscopical Science receive fifty copies of their communica-
tion gratis.

All expenses of publication and illustration are paid by the
publishers.

Lithographic plates and text-figures are used in illustration.
Shaded drawings intended for photographic reproduction as half-
tone blocks should be executed in “ Process Black ” diluted with
water as required. Half-tone reproduction is recommended for
uncoloured drawings of sections and of Protozoa.

Drawings for text-figures should not be inserted in the MS.,
but sent in a separate envelope to the Editor.

Contributors to this .Journal requiring extra copies of their
communications at their own expense can have them by applying
to the Printers,

Messrs. Adlard & Son, 22 i, Bartholomew Close, E.C., on

the following terms :

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. V-

Plates, 2/- per 25 if uncoloured; if coloured, at the same rate for

every colour.

Prepayment by P.O. Order is requested.

All Communications fop. the Editors to be addressed to the care
OP Messrs. .J. & A. Churchill, 7 Great Marlborough Street,
London, W.

THE MARINE BIOtOClGAL ASSOCIATION

()!•’ THE

UNITED KINGDOM.

:o:

Patron— HIS MAJESTY THE KING.

President— Sir RAY LANKESTER, K.C.B., LL.D., F.R.S.

:o:

The Association was fodnoed “ to establish and maintain Labokatokiks on
THE coast of the UNITED KINGDOM, WIIEKE ACCUEATE EESEABCHES MAT BE CAEEIED
ON, LEADING TO THE IMPEOVEMENT OF ZOOLOGICAL AND BOTANICAL SCIENCE, AND TO
AN INCEEASEOF ODE KNOWLEDGE AS EEGAEDS THE FOOD, LIFE CONDITIONS, AND HABITS
OF BeITISH FOOD-FISHES AND MOLLDSCS.”

The Laboratory at Plymouth

iv.is opened in 18S8. Since that time investigations, practical and scientific, have
been constantly pursued by naturalists appointed by tlie Association, as well as by
those from England and abroad who have carried on independent researches.

Naturalists desiring to work at the Laboratory

should couimunirate with the Director, who will supply all information as to
terms, etc.

Works published by the Association

include the following : — ‘A Treatise on the Common Sole,’ .1. T. Cunningham, ALA.,
4to, 25/-. ‘ The Natural History of the Marketable Marine Fishes of the British

Islands,’ J. '1'. Cunningham, M.A., 7/6 net (published for the Association by
Messrs. Macmillan & Co.).

The Journal of the Marine Biological Association

is issued half-yearly, price 3/6 each number.

In addition to these publications, the results of work done in the Laboratory
are recorded in the ‘Quarterly Journal of Alicroscopieal Science,’ and in other
scientific journals, British and foreign.

Specimens of Marine Animals and Plants,

both living and preserved, according to the best methods, are supplied to the
principal British Laboratories and Aluseums. Detailed price lists will be forwarded
on application.

TERMS OF MEMBERSHIP.

Annual Membeus . . . .£110 per annum.

Life Membees 15 15 0 Composition Fee.

Foundees 100 0 0 „ „

Goteenoes (Life Members of Council) 500 0 0

Members have the following rights and privileges : — They elect annually the
Officers and Council; they receive the Journal free by post ; they are admitted to
view the Laboratory at any time, and may introduce friends with them ; they have the
first claim to rent a table in the Laboratory for research, with use of tanks, boats, etc.;
and have access to the Library at Plymouth. Special privileges are granted to Governors,
Founders, and Life Alembers.

Persons desirous of becoming members, or of obtaining any information with
regard to the Association, should communicate with —

The DIRECTOR,

The Laboratory,

Plymouth.

With Ten Plates, Royal Ato, 5s.

CONTRIBUTIONS TO THE KNOWLEDGE OF RHABDOPLEURA
AND AMPHIOXUS.

By E. KAY LANKESTER, M.A., LL.D., F.R.S.

London: J. & A. Chuechii.l, 7 Great Marlborough Street.

Quarterly Journal of Microscopical

Science.

The SUBSCRIPTION is £2 for the Volume of Four Numbers;
for this sum (prepaid) the Journal is sent Post Free to any part
of the world.

BACK NUMBERS of the Journal, which remain in print, are
now sold at an uniform price of 10/- net.

The issue of Supplement Numbers being found inconvenient,
and there being often in the Editor’s hands an accumulation of
valuable material, it has been decided to publish this Journal at
such intervals as may seem desirable, rather than delay the appear-
ance of Memoirs for a regular quarterly publication.

The title remains unaltered, though more than Four Numbers
may be published in the course of a year.

Each Number is sold at 10/- net, and Four Numbers make
up a Volume.

London : J. & A. CHURCHILL, 7 Great Marlborough Street.
TO CORRESPONDENTS.

Authors of original papers published in the Quarterly .Journal
of Microscopical Science receive fifty copies of their communica-
tion gratis.

All expenses of publication and illustration are paid by the
publishers.

Lithographic plates and text-figures are used in illustration.
Shaded drawings intended for photographic reproduction as half-
tone blocks should be executed in “ Process Black ” diluted with
water as required. Half-tone reproduction is recommended for
uncoloured drawings of sections and of Protozoa.

Drawings for text-figures should not be inserted in the MS.,
but sent in a separate envelope to the Editor.

Contributors to this Journal requiring extra copies of their
communications at their own expense can have them by applying
to the Printers,

Messrs. Adlard & Son, 221, Bartholomew Close, E.C., on
the following terms :

For every four pages or less —

25 copies

. 5/-

50 „ . .

6/-

75 „ . .

6/6

100 „

. . 7/-

Plates, 2/- per 25 if uncoloured ; if coloured, at the same rate for

every colour.

Prepayment by P.O. Order is requested.

.\ll Communications for the Editors to be addressed to the care
OP Messrs. J. & A. Churchill, 7 Great Marlborough Street,
London, \V.

THE MARINE BIOLOGICAL ASSOCIATION

OV THK

UNITED KINGDOM.

:o:

Patron— HIS MAJESTY THE KING.

President— Sir RAY LANKESTER, K.C.B., LL.D., F.R.S.

:o:

The Association was founded “ to establish and maintain Labobatobibs on

THE COAST OF THE UNITED KINGDOM, WUEBE ACCUEATE BESEAECHES MAY BE CABBIED
ON, LEADING TO THE IMPEOVEMENT OF ZOOLOGICAL AND BOTANICAL SCIENCE, AND TO
AN INCEEASE OF ODE KNOWLEDGE AS EEGAEDS THE FOOD, LIFE CONDITIONS, AND HABITS
OF BbITISH FOOD-FISHES AND MOLLUSCS.”

The Laboratory at Plymouth

was opened in 1888. Since that time investigations, practical and scientific, have
been constantly pursued by naturalists appointed by the Association, as well as by
those from England and abroad who have carried on independent researches.

Naturalists desiring to work at the Laboratory

should communicate with .the Director, who will supply all informatiou as to
terms, etc.

Works published by the Association

include the following : — ‘A Treatise on the Common Sole,’ J. T. Cunningham, M.A.,
4to, 25/-. ‘ The Natural History of the Marketable Marine Fishes of the British

Islands,’ J. T. Cunuiugham, M.A., 7/6 net (published for the Association by
Messrs. Macmillan & Co.).

The Journal of the Marine Biological Association

is issued half-yearly, price 3/6 eacli number.

In addition to these publications, the results of work done in the Laboratory
are recorded in the ‘ Quarterly .lournal of Microscopical Science,’ and in other
scieutific journals, British and foreign.

Specimens of Marine Animals and Plants,

both living and preserved, according to the best methods, are supplied to the
principal British Laboratories and Museums. Detailed price lists will be forwarded
on application.

TERMS OF MEMBERSHIP.

Annual Members . . . .£110 per annum.

Life Members 15 15 0 Composition Fee.

Founders 100 0 0 „ „

Governors (Life Members of Council) 500 0 0

Members have the following rights and privileges : — They elect aunnally the
OflScers and Council; they receive the Journal free by post; they are admitted to
view the Laboratory at any time, and may introduce friends with them ; they have the
first claim to rent a table in the Laboratory for research, with use of tanks, boats, etc. ;
and have access to the Library at Plymouth. Special privileges are granted to Governors,
P’ounders, and Life Members.

Persons desirous of becoming members, or of obtaining any information with
regard to the Association, should communicate with —

The DIRECTOR,

The Laboratory,

Plymouth.

lyuh Ten Plates, Royal ^to, 5s.

CONTRIBUTIONS TO THE KNOWLEDGE OF RHABDOPLEURA
AND AMPHIOXUS.

Hy E. HAY LANKESl'ER, M.A., LL.D., F.ll.S.

Eondoii : J. & A. CHnKCHIl.L, 7 Great Marlborough Street;

Quarterly Journal of Microscopical

5cience.

The SUBSCRIPTION is £2 for the Volume of Four Numbers;
for this sum (prepaid) the Journal is sent Post Free to any part
of the world.

BACK NUMBERS of the Journal, which remain in print, are
now sold at an uniform price of 10/- net.

The issue of Supplement Numbers being found inconvenient,
and there being often in the Editor’s hands an accumulation of
valuable material, it has been decided to publish this Journal at
such intervals as may seem desirable, rather than delay the appear-
ance of Memoirs for a regular quarterly publication.

The title remains unaltered, though more than Four Numbers
may be published in the course of a year.

Each Number is sold at 10/- net, and Four Numbers make
up a Volume.

London : J. & A. CHURCHILL, 7 Great Marlborough Street.
TO CORRESPONDENTS.

Authors of original papers published in the Quarterly -Journal
of Microscopical Science receive fifty copies of their communica-
tion gratis.

All expenses of publication and illustration are paid by the
publishers.

Lithographic plates and text-figures are used in illustration.
Shaded drawings intended for photographic reproduction as half-
tone blocks should be executed in “Process Black” diluted with
water as required. Half-tone reproduction is I’ecommended for
uncoloured drawings of sections aud of Protozoa.

Drawings for text-figures should not be inserted in the MS.,
but sent in a separate envelope to the Editor.

Contributors to this .Journal requiring extra copies of their
communications at their own expense can have them by applying
to the Printers,

Messrs. Adlard & Son, 22.^, Bartliolomew Close, E.C., on
the following terms :

For every four pages or less —

25 copies .... 5/-

50 „ .... 6/-

75 „ .... 6/6

100 „ .... 7/-

Plates, 2/- per 25 if uncoloui-ed; if coloured, at the same rate for

every colour.

Prepayment by P.O. Order is requested.

All Communications for the Editors to be addressed to the care

OP Messrs. -J. & A. Churchill, 7 Great Marlborough Street,

London, W.

THE MARINE BIOLOGICAL ASSOCIATION

OF THK

UNITED KINGDOM.

:o:

Patron— HIS MAJESTY THE KING.

President— Sir RAY LANKESTER, K.C.B., LL.D., F.R.S.

:o:

TjiE Association was founded “to establish and maintain liABOuATOiiiEs on

THE COAST OF THE UNITED KINGDOM, WHEUE ACCUUATE EESEAKCHES MAT BE CABBIED
ON, LEADING TO THE IMPIIOVEMENT OF ZOOLOGICAL AND BOTANICAL SCIENCE, AND TO
AN INCKEASE OF ODE KNOWLEDGE AS EEGAEDS THE FOOD, LIFE CONDITIONS, AND HABITS
OF BeITISH FOOD-FISHES AND MOLLUSCS.”

The Laboratory at Plymouth

was opened in 1888. Since that time investigations, practical and scientific, have
been constantly pnrsned by natnnilists appointed by the Association, as well as by
those from Ungland and abroad who have carried on independent researches.

Naturalists desiring to work at the Laboratory

should communicate with the Director, who will supply all information as to
terms, etc.

Works published by the Association

include the following ; — ‘A Treatise on the Common Sole,’ .1. T. Cunningham, M. A.,
4to, 25/-. ‘The Natural History of the Marketable Marine Fishes of the British
Islands,’ J. T. Cunningham, M.A., 7/6 net (published for the Association by
Messrs. Macmillan & Co.).

The Journal of the Marine Biological Association

is issued half-yearly, price 3/6 each number.

In addition to these publications, the results of work done in the Laboratory
are recorded in the ‘Quarterly .lonrnal of Microscopical Science,’ and in other
seientific journals, British and foreign.

Specimens of Marine Animals and Plants,

both living and preserved, according to the best methods, are supplied to the
principal British Laboratories and Museums. Detailed price lists will be forw.nded
ou application.

TERMS OF MEMBERSHIP.

Annual Membehs . . . .£110 per annum.

Life Membees 15 15 0 Composition Fee.

Foundees 100 0 0 „ „

Goveenoes ( Life Members of Council) 500 0 0

Members liave the following rights and privileges: — They elect annually the
Officers and Council; they receive the Journal free by post; they are admitted to
view the Laboratory at any time, and may introduce friends with them ; they have tlie
first claim to rent a table in the Lal)or.atory for research, with use of tanks, boats, etc. ;
and have access to the Library at Plymontii. Special privileges are granted to Governors,
Founders, and Life Members.

Persons desirous of becoming members, or of obtaining any information with
regard to the Association, should communicate with —

The DIRECTOR,

The Laboratory,

Plymouth.

With Ten Plates, Royal -ito, 5s.

CONTRIBUTIONS TO THE KNOWLEDGE OF RHABDOPLEURA
AND AMPHIOXUS.

By E. RAY LANKESTER, M.A., LL.D., F.R.S.

London : J. & A. Chuechill, 7 Great Marlborough Street.

Quarterly Journal of Microscopical

Science.

The SUBSCRIPTION is £2 for the Volume of Four Numbers;
for this sum {'prepaid) the Journal is sent Post Free to any part
of the world.

BACK NUMBERS of the Journal, which remain in print, are
now sold at an uniform price of 10/- net.

The issue of Supplement Numbers being found inconvenient,
and there being often in the Editor’s hands an accumulation of
valuable material, it has been decided to publish this Journal at
such intervals as may seem desirable, rather than delay the appear-
ance of Memoirs for a I’egular quarterly publication.

The title remains unaltered, though more than Four Numbers
may be published in the course of a year.

Each Number is sold at 10/- net, and Four Numbers make
up a Volume.

London : J. & A. CHURCHILL, 7 Great Marlborough Street.
TO CORRESPONDENTS.

Authors of original papers published in the Quarterly Journal
of Microscopical Science receive fifty copies of their communica-
tion gratis.

All expenses of publication and illustration are paid by the
publishers.

Lithographic plates and te.xt-figures are used in illustration.
Shaded drawings intended for photographic reproduction as half-
tone blocks should be executed in “Process Black” diluted with
water as required. Half-tone reproduction is recommended for
uncoloui’ed drawings of sections and of Protozoa.

Drawings for text-figures should not be inserted in the MS.,
but sent in a separate envelope to the Editor.

Contributors to this Journal requiring extra copies of their
communications at their own expense can have them by applying
to the Printers,

Messrs. Adlard & Son, 22.1, Bartholomew Close, E.C., on
the following terms :

For every four pages or less —

25 copies .... 5/-

50 „ .... 6/-

100 „ .... 7/-

Plates, 2/- per 25 if uncoloured; if coloured, at the same rate for

every colour.

Prepayment by P.O. Order is requested.

All Communications for the Editors to be addressed to the care

OP Messrs. J. & A. Churchill, 7 GreAt Marlborough Street,

London, W.

THE MARINE BIOLOGICAL ASSOCIATION

OF THK

UNITED KINGDOM.

:o:

Patron— HIS MAJESTY THE KING.

President— Sir RAY LANKESTER, K.C.B., LL.D., F.R.S.

:o:

•

The Association was founded “ to establish and maintain Labobatobies on
THE coast of the UNITED KINGDOM, WHEBE ACCUBATE HESEAECHES MAX BE CABBIED
ON, LEADING TO THE IMPEOVEMENT OF ZOOLOGICAL AND BOTANICAL SCIENCE, AND TO
AN INCEEASEOF OUE KNOWLEDGE AS BEGAEDS THE FOOD, LIFE CONDITIONS, AND HABITS
OF BeITTSH FOOD-FISHES AND MOLLUSCS.”

The Laboratory at Plymouth

was opened in 1888. Since tliat time investigations, practical and scientific, have
been constantly pursued by naturalists appointed by tlie Association, as well as by
those from England and abroad who have carried on independent researches.

Naturalists desiring to work at the Laboratory

should communicate with the Director, who will supply all information as to
terms, etc.

Works published by the Association

include the following : — ‘A Treatise on the Common Sole,’ J. T. Cunningham, M.A.,
4to, 25/-. ‘ The Natural History of the JIarketable Marine Fishes of the British

Islands,’ J. T. Cunningham, M.A., 7/6 net (published for the Association by
Messrs. Macmillan & Co.).

The Journal of the Marine Biological Association

is issued half-yearly, price 3/6 each number.

In addition to these publications, the results of work done in the Laboratory
are recorded iu the ‘Quarterly .lournal of Microscopical Science,’ and in other
scientific journals, British and foreign.

Specimens of Marine Animals and Plants,

both living and preserved, according to the best methods, arc supplied to the
principal British Laboratories and Museums. Detailed price lists will be forwarded
on application.

TERMS OF MEMBERSHIP.

Annual Membees . . . . f 1 1 0 per annum.

Life Membees . . . . . 15 15 0 Composition Fee.

Foundees ...... 100 00 „ „

Govebnors (Life Members of Council) 500 0 0

Members have the following rights and privileges: — They elect annually the
Officers and Council; they receive the Journal free by post; they are admitted to
view the Laboratory at any time, and may introduce friends with them ; they have the
first claim to rent a table in the Laboratory for research, with use of tanks, boats, etc. ;
and have access to the Library at Plymouth. Special privileges are granted to Governors,
Founders, and Life Members.

Persons desirous of becoming members, or of obtaining any information with
regard to the Association, should communicate with —

The DIRECTOR,

The Laboratorj%

Plymouth.

Quarterly journal of mil
V.54 1910 i

amnh library

1001 13604