BIOLOGICAL BULLETIN

OF THE

noarinc Biological laboratory

IP 'i i , MAS

Start

I.. G. ( 'NKi.i\ Princet •

[ACQJJBS I. -.1 M— /'//'• A' Institute \fedicalRcsearch

T. II. M. »RG \N ' -intnl>t\i I :ty.

\\'. M. \\ in i i i it ///'.•//•./ t 'nil-fruity.
\\. I'. . \Vn ON ' • ' "• ' ' I ' :y-

FRANK K. I.n.i.lE- C t 'nirfr*ity

\ .11 Ml XXI.

\V. H IDS 11"! 1 , MASS
JL'NK 1»> \' 'VI MI'-I K, 1911

PRESS OF

THE NEW ERA PRINTING COMPANY
LANCASTER. PA.

CONTENTS OF VOLUME XXI.

No. i . JINK. MI i.
Hii.ins. \\n.i.i.\\i A.

/'• I

Mil I I MUll, Vll

9
CALKINS, GARY N. :<m

j6

in.

7 ;
KING, HELEN Di . . 104

1 1 \- • 1 . 1 1 /

1 1 '-,
Anitoi r, J. F., AND RICH II

.'///r (/I! ] 22

AUGUST, I'M 1 .

sm i i inn-. \ . I . //. /' i-1;

PENNENT, D. H. All' ' ' m in

\ 52
STE YEN s , N . M . //i i > =>

I'INM N . I hi I II. .!

I' -

\«>. .}. SEPTI I'M i .

Ai i i \. \\ I . .1 .v . . . <• ntiat

:•> :• [87

r. \Ki'."i R, TDOMAJ ?«-

M ORGAN, T. H . A •

J 1 -

GEE, WILSON P. ::yf>h \lu\- d

]\'nlkt-r 122

CIIIIM— 1 1 K. I . I . / he .\fati>: the

A''.:, hyu ;

iii

iy COM 1;.M- "I \"LlMi: XXI.

No. 5. OCTOBER, i<ni.

Ti'RNER, C. H. Experiments on Pattern-vision of the Honey Be,.

WODSEDALEK, J. E. Phototactic Reactions and their Reversal

in the May-fly Nymphs He plage n in inter punctata (Say}.... 265

FERGUSON, JEREMIAH S. A Preliminary Xotc on the Relation
of \ormal Living Cells to the Existing Theories of the Ilis-
togenesis of Connective Tissue 272

CHILD, C. M. The Method of Cell Division in Mouiczia 280

PAYNE, FERNANDUS. Drosophila ampelophila Loc\. /-'• ed in tin-
Dark for Sixty-nine Generations 297

STRICKLAND, E. H. Some Parasites of Sinndium Larvae and their

Effects on the Development of the Host 302

NO. <>. XoYKMHER, I9II.

PEARL, RAYMOND. The Personal Equation in Breeding Experi-
ments Invoking Certain Characters of Maize 339

MONTGOMERY, THOMAS H. Differentiation of the Human Cells of

Sertoli 367

RIDDLE, OSCAR. On the Cause of Autotomy in Tubularia 389 ^

Vol. XXI. June, /p//. No. i.

BIOLOGICAL BULLETIN

SOME REMARKS ON I Hi: GASTRULATION OF

DESMOGN Mill - n SCA.

\VII.I.I.\M A. HII. l< i\.

For tin- I.IM three <ir four years I have been trying to

lll.lleri.il In eomjilete < il i-eP \ .1 1 \> HI- Ill.U le -• Ulle I i I11C agO Oil tile

•,M~l filiation ol />, ! ri are many difficulties in

the \\.iy n| dlit. lining tin- ri^ht stages f< >r this Mudy. for the
.in- u it In Hi l |ii^nieiit .1 in 1 eiirl. i-e<l in \ ,-ry tmi-h nieiiilnM'

Ailile'l 1" tile ililtli lilt!' • lei'tinll .lllil i 'Hell M t h HI . i- tile I.U't

that the eggs are large .mil in.t e.i-il\ se< timie.1.

As nieiiii'iiied in .in e.irlier p.iper,1 \\i< • <i thi> Bp ire

lliilnlil.i-tir .ihli'Ul'Ji ;i|i|)fn.n -hilli: the lllenilil.l-tii- t\]>e «'t

FIG. z, i 'pi'iT lull ol .1 h.- .i..ti«-'i « --IN

\\ith -in. ill y.-lk xr-niiil''-

nieiit.ition. A moderate-sized M.i-tul.i r.ixity i- funneil. al'i.\e
it. MIL ill cells \\ith MIL ill yilk ur.mule- .ui<l ln-lnu, large <vll-
\\ith larger granule-. Tlii- r.i\it\ in l.ittT stages f< .iiu^ to In.-
more or le>- tilled in \\ith larger or Miialler i ell- \\liich round off
from the p-neral ma— Figs. I and 2). To \\hat extent the

1 \\'. A. Hilt. .11. ' t.i-n.i.il l-Y.itiii>> .'i an l-'arly 1 )r\ i-l. ipnu-nt nf Dcsmognathiis
• i." J:>nr. Morpli.. \'ul. XX.. 1909.

2 WILLIAM A HILTON.

•iH-ntation cavity may in this way normally become filled in,

m at pri-un unable t«> say, but I have found some specimens

every Xl'-"" t(ir l'K' Past ^cw >"*-'ars m ^'hich the segmentation

ra\ity was totally obliterated. As the cells divide, the small

ones of the animal pole gradually extend
about the larger yolk cells and, at an
early stage, one may recognize at the
point of junction of the two masses and
in from the surface, some of the small
cells which later go to form the meso-
blast. These correspond to those recog-
nized by Morgan, King, Ruffini and
others.1 At a stage when the small cells
begin to migrate over the larger, there
are about three layers of these with
small yolk granules at the animal pole.

This number later becomes reduced to
FIG. 2. Portion of a late .

morula stage of D. fusca. two and then to one, similar to the sin-

gle cell layer found by Lange in Megalo-
batrachns;- this outer layer or ectoblast

becomes more extensive and may be found forming a simple
definite covering over a large part of the egg. The other small
(vIN, main- if not all of which contain small yolk granules, have
l>y this time migrated considerably, but not so extensively as
the ectoblastic. Their exact limits are hard to determine. At
about the time the ectoblastic covering at the animal pole has
become reduced to one cell, the first indication of gastrulation
\\.i^ tound. Fig. 3 is a section of a rather early stage.

As mentioned at an earlier time, the external appearance of
gastrula stages do not differ from those of other amphibia to
.m\ greal derive, but some details of development seem peculiar.
I no\\ h.i\c several series of earlier stages than have been available
and in the earliest of these, 1 find the gastrula mouth

I. II. M«i'.'atl, "Tin- <lrvrl«.|iiin-iit <•! tl.< I 1'Vg." 1897. H. D. KillK.

Che Gastrulation ol tin- LVM »i r>n<~« lent iy.inu^n ." .\ »i. \iit., \'»\. XXXVI.. 1902.

A. Kiiltini, "Coiurilmto alia mil' i-.-i-n/a ili-lla ( >nt..u' "• \mlil>i .mmi rd

li," Anh. It. Di. Aunt. Di. I-'.inb., K>n-.

\>. •!'• I..HIKI-. Jr., "Die Kciiiil>l.i[ti-iliil.lun« <!<•- .\li-i;<il<>l>,iir<n ln<* inn. \inius
Aunt. II- • Bd . •, II. 3, [907.

GASTRULATION OF DESMOGNATHUS FUSCA.

well flown on the yolk mass and appearing as a cleft between
the small cells which have grown down about the larger one-.
This early cleft in all of my specmiens appears to be not entirely
formed by invagination. In later stages, however, there is good
«\idence of ingrowth especially from the small cells of the upper
lip. In ,i numU-r «i -pecimens of this stage, the rounded yolk
cells which were in or ne.ir the segmentation cavity are more or
less artificially rro\\drd to^-ther, probably due to the loss of
fluid bet \\eeii them in tin- pa--a<^e through the alcohols, so that
no clear idea wa- obtained of their position-. It is probable
tli.it about the time of iM-irulation then- i- a con-iderable change
of po-iiion of <rll- at tin- animal pole of the egg, tin- relU may

.
'

FlG. 3 1 In- 1 •

ui.li, • •

lin-ii -ImiiiV.' \li.it

in tin- .!•

11 ilir«ii)Ji -I l.it< : u 11-

' ' tti-.l IK 'III tin- -III-

:ln- .rll- i- ailili'il. X25-

ha\e a looser arrangement, tlu-rr ma\- even b«- con-id. -ral.K-

ana- brt \\rell thrill o. i'll|'ied b> tlllid. exidclirc of which ina\
In- seen in ci-rtain li\ii 1'lu- I"-- "f thi> lluid b\- fixation

Causes tlu- o>lla|>-i- of the thin roof and the croudin^ to-t-tln-r
• .f iln- c.-ll- at the animal pi-!r ! . IIo\\r\i-r. in many

-li-litl> later including both earlier specimens and those

oill\ recently oblaiiud. 1 iieiitatioli ca\it\ -« em- to ha\e

been lauel\- li.-t and -till later than this, new -pace- between the
yolk cell- make their appearance and communic.il.- directly with

4 WILLIAM A. HILTON.

the cavity formed by ua-trulation. Fig. 4 is through a specimen

-uch a lau-r stage where the total cavity, although slight, is

ir and definite. The lining of the first part of this is from

invaginated cells and some of the original ones which were with

small yolk granule- on the animal pole may be distinguished as

part of the lining of the cavity, the total extent of which is shown

in a few sections bevoml this level.

FIG. 5. Longitudinal section through blastopore and archenteron showing
dorsal and ventral lip. Small cells with small granules dotted as in the other
drawing.

In later stages the cavity or archenteron may not be very
extensive. Cells just under the dorsal ectoblastic ones maybe
seen to be composed of small yolk granules and beyond the point
where in vagi nation is apparent these seem to be well organized
and represent a part of the middle germ layer and were probably

-.. ;;7V.-. .•;._•..-.;••.

rvy - • ' ; "'". • ••••
• ' •' ' '

FIG. d. Longitudinal -i-rtinii through the lila-top.ire of a stage a little later
than that oi Fig. 5, showing cells which have grown in from the dorsal lip. X-'5-

formed from some of the earlier sin, ill cells which wen- traced
in their migration-, about the yolk mass.

In other specimens the cavity of the archentcnm is not very
wide, mosl of the d'M-.il cells lining it contain small yolk granules,

GASTRULATIOX OF DESMOGNATHUS FU5CA.

and these may represent some of the early small cells which have
migrated about the yolk. Some of the cells near the opening
of the blastopore have grown in dorsally and the ventral lip
is also to some degree made up of small cells, some of which
may be followed into the floor of the archenteron (Fig. 5).

In later gastrula stages as the enteron becomes more and more
marked, there may IM- noticed a considerable ingrowth of cells,
« -pecially from the dor-al lip, in ><>me -pecimeiis can-ing this
free portion of the roof to !,<•< -ome rolled as Hr.uier1 found to be
the i ase in the < '.ymnophiona !

I- i ;i.ni tli; n <-inlir\

iu-1 \ •••in. -In. \vinu tin- l.i!

• i tin-

\i a time \\lu-n the 1. la-lop,, re i- reduced t<> a \ cr\ -mall hole
and HIM before medullar> fold- are formed, the ca\il\ ha- come
to lie lar^e and e\lell-i\e. Fig. ~ i- from a CTOSS Section of -ueh
a Stage. In it the me-obla-t ha- not j>enet rated \er\ far \eiitrally
e-pecially in tlu- mon- cejihali. ;. Later than thi-, \\ hen the

neural plale has liev^un to close, w»- find that a- \et in the head
end the me-ol.la-i does QOl extend much farther but. in the
caudal region, it ha- ]>enetrated between the ectobla-t and the

\. Mr.ui.-i. .! Ki-nntin- I!,T Knt\vicki-hinj;sK<'sclii' !it<- und

-I. r liyinniiphii'iirii." /.<><>!. J<ilirb. Anal. Abl.. Bd. X.. i

6 WILLIAM A. HILTON.

yolk in all direction- from a i;reat mass of ce!l> surrounding the
bla-topore. extending forward and to a slight extent back of it.
Thi- ma-> of cell-, which is continuous cephalad with the nervous
system and the- mesoblast, is in this more caudal region an un-
ditf'ereniiated -roii]). It is from this that the mesoblast grows
out io meet that which has been formed on each side of the
iiotorhord (Fig. 8). The place where the mesoblast from the
cephalic region and that from this caudal mass meet is ofu-n
distinguishable because of a difference in the staining character
of tin1 cells. In this late stage when the body of the embryo is

FIG. 8. Cross section through the caudal end of a late medullary plate stage,
showing the undifferentiated cell mass, and the small size of the archenteron.

outlined, the cavity of the archenteron is much reduced in size
(Fig. 8). This caudal mass is made up from cells on all sides
of the blastopore, but especially cephalad of it and evidently
represents in large part, the small cells or their progeny which
with the narrowing of the blastopore have been brought to the
dorsal side of the yolk mass.

In the study of these stages in Desmognathus, one is impressed
with the fact that fluids within the egg between the blastomeres
have more significance than has been generally recognized and it
seems possible that this fluid varying with the condition of
development, may have considerable importance in the change
of form and po-hion of the cells. At times this liquid seems to
exert no active influence on the size or condition of the cavitie-
or -hapes of the cells, and at others it seems to definitely force
the cells into an even layer. For instance in early segmentation
Stages, those from the yolk mass and other surrounding cells
round oil to\\.ird the bl.i-tonele and tend to fill it in, while at
other time-, as in late -a-trula Stages, the edij.es of the cell-

GASTRULATION OF DESMOGNATHUS FUSCA. ~

surrounding the archenteron are as smoothly pressed back from
the lumen as though some definite substance occupied the large
cavity.

The variations from time to time of the cavities within the
egg are quite striking. The variations in extent of the segmenta-
tion cavity, its reduction and disappearance in some specimens
at least, the developmen: ally in the later stages of clefts

bet \vrcn cells, the growth in extent of the archenteron and its
reduttion by the time the nervous >\ -tern is developed; all of
ihe-e changes may be dependent upon the large yolk ma-- and
the peculiarities of environment. . If this last be a factor of
importance in the < a\ ity change- within the eggs, then we miuht
expect to find, as ue <1". -Mine variation in individual- of the
-ana- n-lati\< "f d, \ , |, .pmeiit , for females with e-i;- or

yoim- are found in place- that are from wet to almo-t dry at
different seasons <>r during the -aim- -m inner.

- MI < ',l \i k \l ( A INI i.i -f INS.

1. The -mall-^ranuled, -mall cell- < •!' animal pole v^n-w about

the larger with greater or less filling in «i the segmentation cavity.

2. The tell- <>l the allimal pole be.., ill. reduced to olle la\ er.
.v The In -I indication «'t ;Ka-t rulat i< m .ip|>ear- in the VCgel Ltive

liemi-pheie .it a re^i"M bet \\i-en the -mall cell- \\hich ha\e ^i"\\'n
do\\ n and the \<>lk t ell-.

4. There i- an in^n >wtli < -I" -mall cell-, from the d<>r-al lip
e-peciall\ .

5. The e.ul\ -e^meiitatit'ii ca\it\ a- -uch |imbabl\- dm-- lid
bt O >me i"ined tn the in\ agination.

6. The later an hem en. r, is fi .in led parth by in\ agination and
]iai'tl\ b\ -eparatii m betu !!-.

7. The ca\it\ o!" the anhenten.n i- a|iparenll\ made up of
in\a'j.inated cell-. \nlk .ell- and included under either ..lie or
both of these, some oi the -mall cell- which ha\e migrated from
the re^i«'H "I the animal pule.

8. The -mall cell- at the animal pole form ect<>bla-t.

<). The -mall cell- .it the animal pule at lea-t -mile of which
an- under the SUrl Us form me-obla-t and probabK' -ome

eiitt '1 'la-t .

WILLIAM A. HILTON.

10. In later >tages of gastrulation there is a marked ingrowth
from the dm>al lip of the blastopon

11. Dorsal and ventral mesoblast become fused in later stages
due to the ("in-mat it in of the caudal mass in front of and about
the blastopore.

12. The caudal mass furnishes mesoblast in all directions.
That funned raudally and laterally grows between the yolk cells
and ectoblast. Later, the growing edges of the mesoblast fuse
to enclose practically the whole yolk mass.

CORNELL UNIVERSITY,

DEPARTMENT OF HISTOLOGY AND EMBRYOLOGY.

ECOLOGICAL SUCCESSION.

I. STHKAM IVm.- AND mi: MI.THDD OF Pnv-n M.RAI-HIC

ANALYSIS.

VKTOR E. SHKI.FI >RD.

I. Intr-i'liii ti"ii 9

II. Miit'-ri.iU .iii-l !.• • ii'Iy . i }

III. I'n--«-nt;iti'>n "I th'- I). i1.. i . 19

IV. I.M-. ii--i-.ii "l tin- I » 23

I. I Ji-triliiitiii!. _• ;

-\l

'•:r-;iii "t • •iimuniti- V .i.iti"ii .. 24

I >[-. \K!H~ aii'l 1 :i th«- A- i. -in of

-'7
29

I :

V. Siiiiiiii.il \ "! tin- K-
\' I . A- kii' 'U Icdgments and B , i •

[.INI noN.

The \\iilci' i- intric-ii -d |irim.iril\' in c\pi tiiiniu.il \\<>rk. In
(•••iiiu-ciiiiii \\iili . \pri iinciii- mi ilic iini(|ilic.uinn nl" t.iMiiKunii-
characters «\ tin- ti^cr ln-rtl«-- .ind tin- lilV-lii~i<>t\ umk \\hich
QCCessarily .« < •"inp.uiicd it ^lu-ll'ord, '08), certain -trikini; rt-l.i-
ti'Hi^ "I tlic~i- lu't-tlc- in environmental «-niii|tli-\i-- \\cic
in i<)<>5. MM -•• tvl.itii'ii- -|'"iidrd ( l"-rl\ t«i the

ntfiii.il rcl.itiitiis .UK! -iir.(-~i<ni <>t" pi. int-. ,md illu-t r.itrd tin-
.-.inn- pi-incipK-..i«. the pl.int^ tlu-niM-l\»-- Shrlt'nrd. 'i
ilil,- ini|)i.n.iiit ln-.iriii^- "t tin- I'.ict^ di-fd\
\\oik, .in<l the mod,- <•!' att.ickin:; tlu- pmliU-nis of evolu-
lioii, tlu- \\riter undi-rtook to t'.imili.iri/e hini-elf with the prin-
ciples of ph\Mo-iMphv, .ind \\ith the principle- <>l ]il.mt ecology
<-t forth 1>\ \\'. inning. ("o\\le-. Schiniper. ("lenient-, Whitford,
Li\in^-ton, TiMn-e-ui, 1'l.ihault, Schn'iti-r. Mo>s, Schant/,
1 >.ickno\v>ki and in. my other-. The \\<>rk of Ad.un> and of
otlu-r /( n'lli i^i-t - h.i- been followed in detail al-o.

VICTOR E. SHKLFOKD.

It was my hope t<> -ccure guiding principles to be used in the
interpretation of the tiger beetle data, in the fields where zoology
a tril uited only chaos. In 1907 an appointment requiring
that I give- instruction in natural history alone, UMiig the field
method, -erved to stimulate my efforts in this direction in order
to find a basis for the organization of field and natural history
instruction. In 1908, I was compelled, quite against my will.
to drop experimental work for a time and have been left free
to pursue the inquiry to its logical endings.

An early training in zoology which was of the strictest morpho-
logical type, caused me at the outset to share the doubts of
main biologists as to the value of ecological work. However,
because of the circumstances just referred to, I was able to
examine the work of plant ecologists with a large degree of
sympathy, which has grown as the inquiry and accompanying
investigations have progressed. It will be seen that we have
been investigating the question of the value and relations of organized
ecology to absorbing biological problems of today. It is the results
of this investigation that we are concerned in presenting here.
The zoological investigations which have accompanied the work
have been various and only the significant results will be pre-
sented. The investigations have been carried only far enough
to indicate the lines in which they may profitably be directed
further.

The result of our general inquiry, while in the main gratifying,
is in some respects disappointing. \Ve had hoped to find intimate
relations between organizable ecology and the absorbing bio-
logical problems of the day, but everything points to the fact
that animal ecology must be organized independently first, and
related to other problems after organization has been attained.
I or this reason and for the sake of clearness, we- ha\e M -parated
ecology as sharply from other subjects as possible.

It should be noted at the outset that the basis, in principle,
of modern ecology has been developed by botanists quite inde-
|n-ndently of other dubious of the subject of botany. Doubts
as to the \alne of plant ecology once existed among botanists,
but these have disappeared and CCO!M-\ ha> a recognition on a
level with evolution, morphology, and physiology. This is ot

i mere-- 1 here, because llis|or\ often repeal- il-ell.

tCOLOGICAL SUCCESSION. II

In general, animal ecology is concerned with the relation of
animals to their environments. Theirs/ essential is to locate the
animal in its environment.

Our inquiry and the paper- which will deal with the results
have <vnu-rrd around the following purposes:

1. To determine the animal activity which takes place within
the narrowest limit- .md which is most important to the specie-,
with a view to locating motile animals in their environment-.

2. To determine the phi-m.niena in animals which correspond
to gro\\ tli-form in plant-. < '.ro\vth-form in plants is usually a
convenient index of the phy-iolo-ical condition- \\ithin.

3. To determine the validity •logical >ucce--ion a- applied

to animal-.

4. To determine thr v.ilu.- of the principles of physiography
and plant ecolo:_ •. nimals in their environments,

in determining -"iiiethin'c of tin- ph\ -iolo^ical character of
the or-.ini-m .1- a uh«.l.-. .md « in the an.tl\-i- of the or^ani-m

5. To inquire into tin pi .— ib ilit y of or^ani/in^ a -\-tem of
ecological i la— ilication for animal- \\hich -hall lie sufficiently
independent of the Oth( 'i in- and method-, to f. >rm a b.i-i-

for generalization which m.t\ serve as a check on the results oi

tin- held- of evolution, heredity, and phy-iol,

I o inquire int" the relation «-t succession to the quantitative

pro), Inn- ot l.iol. ,-\ .

7. lo in.|iiiiv into tin- relation of succession t«> the origin of
adaptation-.

,s. To inquire into the relation- • . •. to tlu- economic

piol.lein- of zodlogy, and the po— il.Hity of u-ini; ecolo-\ as a
means of lirin^iiu aliout a I'dter uniticalion ot the \ari<ui>
hranche- of /• . pure and applied.

It ha- 1-eeii found impracticalili- to organize the \\«.rk SO that
one pajier \\ill deal \\ith the >ul>jcct of an>' one of the al>o\e
pro|>o-itioii- alone. The di-cu— ion of physiological animal
;ih\- in the \\'hitman Memorial X'olume of the Journal <>J
Morphology i- intended to serve a- an introduction to the -eiu-ral
qtie-tioii-, .md de.il- niainlx with the fir-t t\vo and the tilth
purposes. I'lu- pre-eiit ]t.qier. \\hich i- intended to be the tir-t
of a series "f ti\e or -i\. i- concerned \\ith the (|Ue-lion ot the

12

VICTOR E. SHELFORD.

value of tin- principles of physiography as outlined in the fourth.

\Yith the development of the ideas of genetic physiography

came I lu- recognition of the succession of physiographic conditions

over a ijveii locality (point B of Fig. i). The relations of plants

! animals to physiographic features being recognized, the first

recognition of plant and animal succession came in connection

with physiographic succession. Cowles ('01) carried out a

complete classification of the vegetation near Chicago on the

basis of the plant succession which accompanies physiographic

// G F E D C

B

FIG. i. A diagram showing the successive stages in the profile (general shape
of the bottom) of a very young stream, curved lines, A—B, A—C, A-D, A-E, A-F,
.1 (/. .1 •// representing the successive profiles. The uppermost horizontal line
represents the surface of the land into which the stream is eroding. The horizontal
line with the arrow heads indicates the direction of the migration of the source of
the stream and accordingly of similar stream conditions. The vertical line with
arrow heads when followed downward passess through a succession of stream condi-
tions and represents physiographic succession at the locality B. The point A is the
mouth of the stream. Opposite this are shown three successive sizes of the stream,
and therefore succession at that point. The vertical scale is greatly exaggerated.

change. Adams ('01) also discussed in general geographic terms
the relations of animals to base-leveling and stream development.
He referred to succession of forms in streams. Since his paper,
little has been done in the study of the actual relations of animals
to the various stages of stream development, or the relative
importance of the activities of the animals and the dynamics of
-t reams in determining distribution.

In the study of all phases of distribution, animal activities
have been usually eiiher ignored or taken for granted. Tax-
onomy has been the center around which all such work has rotated
and the taxonomic characters used have been very generally
-tructural. In the study of succession this ha-> been true, only

ECOLOGICAL SUCCESSION. 13

to a lesser degree. \Ye believe that workers in this line have had
in mind the recognition of activities, but the subject is too new
to make such an id< lizable in practice.

It is the purpose of this paper to present some details of the
di-iriliutii'ii of ti-h which -hall throw light on the limitations and
application- of tin- principles which Adams set forth.

The pre-entation "I data and the discussion are based on the
following que-tion-:

1. Are the h>h of the headv. < •!' older -ttvam- the >ame as
the fi-h of younger streams?

2. It SO, did they uei into tin- -tream \\hen it wa- yung and
-imply keep |iacr \\ith tin- advance "t ero-ion into the land.-'

.}. \\'lial i- tin- relation o| the ,i<ti\itie- of animals to their
di-t ributioii in -t ream

t

4. What aspects "!' -i of purely local and what are

o| -eiier.il -i^nitii ai

II. LOCALITIES \\l> M \ i l Kl \l Si i mi .i'.
In l he ii itioii "I problem- in ait\ tield, one ot the thin

rei|uirin- p-eal care is the selection of material. In the Mud\ ot

ecology, if il i- l" be • lied \\iih particular ^puip-, ihi- i-

triii-, and in addition a still wore inifxirtiiti! »n »:u.\t he made,

nameK . //.'.// c/' !/:<• !•>< dtiti ''v.

I . i ' ' '.'/.

I Of ihi- -tlld\ ti-h \\en- -elected bec.m-e lhe\ are probably
ihe onl\ .inimal- that are not introduced \>\ accidental mean-.
'l'he\ inii-t ha\e eiitrred the streams -tudied at their mouth-.

2. The Points of Stn<Iy.
I or the purposes of this study, the ii>h of several -mall >t reams,

\\ithin forty mile- of Chicago, have been collected. These
-tream- are all indicated on the map.

(a) Present Condi! i>

Beginning \\ith the mo-t northerly on the map, the -t reams are:

Util! Creek-Dead River.— It extend- in\\anl about three fourths

of a mile from the boundary of old Lake Chicago ( >ee map), and

divide- into t\\<> main brandies, each of which has a number of

VICTOR E. SHELFORD.

tributarily. The total distance from the old Lake Chicago bluff
t«i the headwaters i> about two and one fourth miles. Its lower

M.i|> i 'f tin' Yi< inity <>!' Chicago.

course which WES added by the falling of the water of the lake,
i- known a^ I )ead Ki\ er.

ECOLOGICAL SUCCESSION. 15

Pettibone Creek is similar to Bull Creek, but a little smaller.
Its length does not exceed one and three fourths miles.

County Line Creek is about twelve miles farther south. It

differs from the others in being smaller. It also divides into

two branches at a distance of only three sixteenths of a mile from

mouth and the length with tributaries is about one half

mile.

Glencoe Brook is about . in; mile south of County Line Creek and
has a total available length, so far as fish are concerned, of only
,i few rod-.

The-e four -tream- arc cl«-el\ < < >mparable. They all rise in
a moraine which -land- 1 « -tu» en i.o and loo feel .ibo\e the lake.
It- height at the Like, or point neare.-t the lake, i- about <>o feet
in everj Case, With the excepti-m of the lower courses of Bull
(nek and iVuibom- ( 'n ek, all are composed of strictly inter-
mittent riffle- ami |n i in. men- pt in umi-ual seasons) pool-.
\\ e will refer to the-c .1- the North Slmre -tream-. In addition
to the-r. t\\o other -tream- uete studie<l for comparison—
Thorn-Hut ti i In Id (i.ek and Ilick"i\ ( 'reek, including it- west

branch.

Ilitkory < ilillcreiit from the other-. It drain- a

number o| mar-lu-- and i- pel maiiciit . Its upper cmir-e i- \.r
than that of the north -lion- -tream-; it i- -lu^ii-h and meander-
through prairie mat-he- uilh only a very .-light fall. The lex el
of tin- part of tin- Mieam n piv-ein- that of a depre— ion betueeii
glacial rid'.. - ' .. >ldth\\ ait . '«" . So far as the pn-eiit c\ cle of
erosion i- coiK-erned ^ali-burx . '07 , erosion ha- iiroceed.d as
lai- as Marlex. lie|o\\ thi- point the -tream i- a typical .-\\ift
brook.

Thorn-Butterfidd (>(•<(• i- intermediate betueeii lliekoiy
l "reek and the north -lime -tream-. The -ouiheru branch of
Butterfield Creek, near Matte-i'ii, 111., i- -hi^gi-h, but ha- inter-
mittent riffle- and permanent pool-.
History of the Region.

\\'h«.'ii the \\'i-con-in i«e -heet retreated from il- maximum
extent, which lax U xoiid the center of the -late of I'linoi-. the
edge of one of it-, lobes Atuo.,.1 and (ioldt hwait . 'oM1 took up
a po-ition a texv mile- oiit-ide of, and parallel x\ith, the prc-eiit

16

VICTOR E. SHELFORD.

>t" Lake Michigan. Here it depo-ited what is known as the
Valparaiso Moraine. When the ice ret reated from this position,
it occupied the basin of Lake Michigan a little to the north of
the present south end of the Lake. \Vateroccupiedthespace
between this lobe and the Valparaiso Moraine. This body of
\\ater is known among geographers and geologists as Lake
Chicago. At its period of maximum extent (see map), it stood
55 to (>o feel above the present lake. The history of Lake Chi-
cago and the other predecessors of Lake Michigan is complicated

TABLE I.

SHOWING THE DISTRIBUTION OF FISH i\' THE NORTH SHORE STREAMS AT THE
TIMKS INDICATED. Thr numl» i- refer in Fig. 2 and Fig. 4.

N.imi- i-t" Stri-;im ami C»m-
iiii-n Nairn- ..f Fish.

1 >.ite and Scientific Vame.

i

2

•

4

5

1

7

Glencoe Brook

August, 1907 . ....

Horned dace . .

Semotilus atromacidalns . .

*

County Line Creek. . . .
Horned dace

1907-8.
Semnlilna alromacultitus . . .

*

*

*

*

Blacknosed dace ....
1 1 '1 1 nii v darter

Riniclitliys utramisus
f>i>lt-n\(>»iii m\nnn

*

*

*
*

Blackhead minnow

I'inii1 phalfs promelas

*

Biuntnosed minn<>w

I'imcph tiles nolalns

*

Common sucker.

Catostomns commersonii. . .

*

Pcttibone Creek1 . ...

September, 1909, and

Horned dace

April, 1910.

SetHotilus atromaculalus . . .

•>

*

*

*

Red bellied dace ....
Blacknosed dace

( hrosomus erithrogaster . .
Riniclitliys atronasus

*

*
*

*
*

Johnnv darter

Boleosoma ni^riint

*

*

Common sucker

Catoslomus commersonii . . .

*

Bull Creek-Dead River .
Horned dace

September, 1909.
Semotilus utrmnm ulatus . . .

*

*

*

*

*

Red bellied dace ....

Cln<>M»nns erithtogasler . . .
l\ i>iit hth \"i atroiui^u't

*

*

*

*

*

Common sucker

( :!t<i\tn»inx i'(»n»ii'rsonii . . .

*

*

Bluntnosod minnow

Pimephales notatus

*

*

I ittle pickerel

E.SOX vertniculatus

*

*

*

Blue gill

Lepontis pallidus

*

*

Large mouthed black
bass

Micxi/'ti-ni,\ stihnniJes ....

*

Pike

Esa\ //«//<?

( i 1 1 ir >ie

Pomovis (DDiitluri'i

Red horse

Mnxostoma aurcolutn

( (ml) sucker

/• i'i ni v . >>i at I'tta

Golden -lnii'-i

Alirmnn crysoleucas

( < >in i uon shiner

Noti'uf'i't i<>rnntii\

Cayuga minnow

\(i//1i«/'/s ii2 VII 1^(1

Tadpole cat

Schilbeodes gyrinus

'Th<- lower part oi ivttiln.in ( n-«-k has been destroyed by the U. S. Naval Schm.1
otherwise tin- table would include- the records for a point 5 and perhaps a point 6.

but probably not 7. 'i indicate iu.uiuplct.i- identification.

ECOLOGICAL SU< CE--ION.

and it will -iifln e, f"r our purposes, to state that the level of the
water fell to a poim al ut 40 feet above the present Lakr Michi-
gan. After -landing in this position for a time, ii fell again to
a point 20 feet lower. Its next period of standstill \\as at a
i2-toot le\el from which it receded to the proem lake level.

The oldest of the north -hore -t reams probably began as mere
gullic- when the lake was al its 6o-foot level. These gullies have
\\-orked their way hack into the morain and deepened thcirchan-
neK in the manner described by Adam- '»>! and diagram-
matically illustrated in Fig. i. Long ago Bull Creek was similar
to the pre-ent ( "oiinty Line Creek. At a recent period County
Line ( "reek v . liki- < il.-n« • <k.

There ha\e liei-n -oine moilitH-atioiir, i if the-e j)r- caused

l.\ i hair.. •- ill lake level. Karh time the 1(\ el of the lake fell, the
-lriain-u. I \\iih • portion near the mouth-. I'nder

l-'i'.. -'. I >i. 1^:1. iinin.it n.irtli

.in- m.i|i|.i-,! • I tli<- ii.

:lil<- ill til.' .'AH

ill lutikrii liin--. \\hiili 1 hich exist m nature.

,H.| tin.- top oi tiir diagram

iHiinlii-i on tl ree <>t' tin- -m-am

i -ont.iiii- fish. .

tin- n-.l-lii-lls. tin.- I'lack-ii. -:thys

<j/r,". ! minnow-; 5. tlio pifki-n-l ami lilunt-no-'-'l ininnou ;

(>. tin- -iinti-li and l>.i-~; 7. the pike. chut'. ~uck '. i- about

60 fret hi.uh. Tin- stipple.! :'.n iu~t al-cvc tl • 1 the la',.

i8

I-:. -HELFORD.

li conditions the Meep p<>rtion> mi-rate hack into the moraine,
/. e., up stream, until the source- is reached. The proce>s of
lowering the water level at the- mouth of a stream and tin- develop-
ment of the steep portion i> known as rejuvenation. Rejuvena-
tion usually affects the distribution of ti>he> in stream-.

However, streams which are made up of intermittent pools
and riffles are not so much at'fected as larger streams. Further-
more, softness of the glacial clay in which the north shore streams
are located would cause the changes in the stream bed to take

TAHLK II.
Tin-: DisTRiuriitiN <•!•• FISH IN HICKORY CREEK (AND ITS WEST BRANCH) IN THE

SIMMER OF IQOg.

Those starred were in the pool nearest the source. I., The first mile of the stream,

mi-. i :n tin tUh pool nearest the source, toward the mouth; II., the third and

fourth miles; III., at the head of erosion, five miles from the pool nearest the source;
IV., six miles from the pool nearest the source; V, nine miles from same; stream
much larger with K""d riffles and one weedy cove.

I.

II.

III.

IV.

V.

1 Ii illlei 1 ( lace*

S<'»i I////N dti'ont itiiltiliis

*

*

*

*

*

< iolden shiner*

l''ji;w/v crysoleucas . .

*

*

*

|ohnn\* darter*

' iniii >i ! '^t u ni

*

*

*

*

Sti me r< iller*

'. iimpostotna tniniiitiliiiii

*

*

*

*

•v col< >red minm >\v*

\otrof>is hlt'tniiici . . . .

*

*

*

*

Blue spotted sunfish*

*

*

*

*

Blunt no~ed minnow

I'nn t'phnli"* ntitiitiis

*

*

*

*

*

( i iiiinn m sucker* . .

( 'ntii\tii»iii v commersonii

*

*

*

Mud minnow

1 inf'i'd limi . . .

*

1 < ' 1 1 minnow

Fundulus Holding

*

*

Rei 1 bellied dace

( lirns/nmi^ erythrogaster

*

*

Chub <ucker

Erimyzon sitcettu

*

*

Black bullhead

1 nit' hint s »it'l<is .

*

*

Black fin

\'i>tr<if>if iimlimtilis

*

*

l< i • ei t hub .

// \'liof>\i\ L-i'iititi kiensis

*

*

*

!• .1 n i .tiled darter

• lumii lliilii'Htirf

*

*

*

Rainb' iw darter

Etheoslonta cceiiilcnm

*

*

*

*

1 i . i - 1 1 1 a 1 1 1 • r

\l ii i'iif't'1'i it punclulata

*

Sucker mouthed minnow

*

iL;a minnow

V,,/ni/);s ((/v«t'<;

*

R' H k ba--

1 iittiltihlilt",' riifii^lri*

*

in ion -1 liner

\ii//'n/>;v tuftiiitll'i

1 . i < ' • ' 1 1 1 1 1 1 1 1 r ' '.\

\'t>tr»f>i'i ri(l>ifn»i<t

Handed darter

I > 1 u '• K i 1 1

•v/i's ("ill i'l n *•

1 i in unti -h

1 .1 I'inni'- nil i iiloti v . . .

sl i me cat

\(>tll> 11 • /!<!. II \

I'll-, ii llii.-i , i i:

!! mi iiit lied 1 ilai k 1 ia --

\l 1 1 t't> filt'f n ^ ill >li ini n'i<

ll'> • - MI ker . . .

!i 'Hill \ til fit ii>:

i 'oinmon red IP '

Mii\i>>ti»ISil il Ill-fill It III

ECOLOGICAL -U< CE--ION. IQ

place rapidly. In tin- exi-ting streams the slope of bed dit<
within a ^ivcn stream and with respect to the different -treams.
The difference- do not appear to be of importance in determining
the relation- of the ti-h pre-ent to a stream as a whole. Ac-
cordingly, we -hall not consider the effect of rejuvenation in the
di-cn--ii in- 1< 'Hi »\\ ii

111. I'K! 51 N I HI!'. IUIA.1

I t.tia of the kind upon which thi- paper i- ba-cd are doiibtK us
familiar to .ill collr. tor- of ti-h. l;rom our point of view the
presence or absence of is nol "t particular impor-

tance, but \\e arc concerned with the arrangement of li-h in the
-tream- and it- causes, Such causes cannot be di-cu--ed here
because th«-\- -hould be -mdicd experimentally. A- \et \\e ha\e
ii( .t been able to do t hi-.

2 -ho\\- the -tie.un- and Table I. the distribution ol the

li-h in the-e streams.

I . .' ' ! I .

\\V note that the li-h honied of the -malle-t Stream

( ,1, I'M... .Is b. long to tin- sam< e at the hea<l-

r

D

B

L. source; J :n..nth: <

,,i ITI.-I..II; D, I 'liiiiin- in T.ilil.- 1 1.

.!!!• ' ll | '.II ! - ! ! • -111 \vllli ll '

Waters Of the lar-et -tream-. The red-bellied dace, \\heil pn-M-iH ,
i- not foil nil SO tar up-lre.im a- the horned da. . I he r« -d- bellied
dace i- ab-eiit from ('oimt\ Line ('reek. Since all the li-h in
all the -tream- niu-t ha\e eiiter.-d from Lake Michigan, one i-

I In- ti-h nl C.nimy I. in.- < reek « : tln-ir .in.iiiu'-ni'-iH -in.-

n, .in tiini- t.. tiiiu- l.i-nvi-rn [907 .mil i';i". I 1"- .iM.mK<-inriH ..I tin- ti-h a- -li..\\n
l.v tin- III.MI ,.i tin- -tn-.mi \\.i- about t!:.- same thruuKh"iit tin- pt-rii..l- ..i -tinly.
ami Tat.li- I.. I-M-I-JU for the drought 1 ln'i-nll.vii..n-; frmii tin- • it her -in-.uiis

\\rii-m.ulrintlu--iimiiu-r ami autumn "i" loop, partly with th- : Mi. S.

1- Mil. li -1.1.111.1. ..I tin- l-i'-M MII-, -urn ol Natural Hi-t.'iy.

VICTOR E. SHELFORD.

-urpri-cd that they have tound so main- of the Breams as is
indicated la-re. The absence i.f the red-hellied dare is no
doubt due ID accident.

'I'hc Mack-noM-d da<v i- present in all the stream- except
C.K-ncoe Brook, Ian lower down than the two just mentioned.
It is usually accompanied by the Johnny darter, which is absent
from Bull Creek. This absence i- probably due to the same
causes as the absence of the red-bellied dace from County Line
Creek. The Johnny darter probably had never become estab-
lished in County Line Creek; only two specimens have been
taken and opportunities to secure a complete collection were good
when the pools dried.

Records of the occurrence of the blunt-nosed and blackhead
minnows in County Line Creek are based on a single specimen
of each taken by a student from the pool nearest the mouth
of the stream. \Ye have taken no others and these were probal >ly
not residents.1 \Ye note that at the points marked 4 (Fig. 4) as
shown in column 4, all the fishes occurring nearer the source of
the stream are present and in addition the young of the common
sucker.

In Bull Creek at point 5, as shown in column 5, the daces are
represented by the horned dace alone ; the other two being absent.
The minnows were present here in numbers and evidently regular
residents. The little pickerel was not abundant.

In columns 6 and 7 (see map also), we find the record of a
number of fishes belonging to large creeks and to ponds. The
distribution of the fishes in Table I. corresponds to the habitat
preferences indicated by Forbes and Richardson ('08), pages
ci.\ to cxiii.

2. Discussion of Table II.

By comparing columns I to 6 in Table I. with column I. of
Table II., we note that nearly all the fish present in the north
-lion- streams are present at the point nearest the source of
Hickory Creek. Considering the first, third and fourth mile-
of this stream we note that, the little pickerel and the black-
no-ed dace are absent in the preen ision part of Hickory Creek

1 I lankinson ha- poiiUrd mil tin- difficulties <>!' si-curing a complete colln tion of
Ii-li. II, I, in tin- water which In- has difficulty in M-CIM in.u with a dip m-t.

ECOLOGICAL SUCCESSION. 21

and present in the north shore streams. In addition, we find
several species in Hickory Creek (among these is a single specimen
of darter, the species probably not resident) not found in the
north shore streams at all. Hickory Creek is better situated
for fish to enter, as they may come upstream directly, while in
the north shore streams they must enter the lake and enter a
given stream by rh.mrr. A« < ordingly, the larger number of
spent-- i- to IP.- . ted in IhVkory ("reek.

In column III. are sho\\ n the fishes at the head of erosion or
th.- rit'tlt-- nearest the sour< e of the stream. A number of swift-
w.uer fish -darter- are -ho\vn here. Thr-e \\etv abundant.

In column IV. tin- tx > u; •, number of lishe- similar in

habit- to tho-e shoun in column III. i- indicated. The stream
i- -imilar in bottom and volume of water at the localities rcpre-
-ented in column- III. aid IV.. and further collecting \\ould
probably lia\<- -ho\\ n the-e t \\ o localities to be inhabited by
practically the same fi-li. Tin- P-ck bass was represented by a

single ill\ellile spr< imell.

1 'lunm \'. sho\\ s the li-h .u a point in the stream \\ltere the
volume of \\ater i- about four times that at point IV. The
>tivam here i- characterized b\ rillles <m,l lar^e pools. This
\\a- more thoroughly studied. ( '. .llectioiis \\ere maile -e\eial
time- during il;< • MI and dail\ for nearly a \\eek in the

month "I ^.-ptember, [908. I It-re was taken a >inv;K- ju\enile
specimen ot the horiu d dat e. The retl-be!lied d.n f and Johnny
daiier were ii"t taken. The atldition of a number of la;
tislu-s lu-re is ,,f int. •• I .itlilition.il >i . in- character-

i-iic of l.ii -,- streams.

;v /' Table III.

Table III. i- introduced to >h, ,\\ conditions intermediate be-
tween the north shore streams and I lickory ( "reek. The localities,
of study \\cre -elected to correspond to localities in Hickory Creek,
just so lai ditioiis in Thom-Butterfield Creek \\ould permit.

In .general, condit i< nis at A in this suvam c.>rn-poiid to I. in
Hickor> Creek, but differ in that the water at A i- c. .11 fined to
po. .U in dry weather and is continuous at I. B corresponds to
111. and C to V.

VICTOR E. SHELFORD.

Aii inspection of column A >h<»\\> that some of the same fishes
arc present as in tin- uppermost pool of Hickory Creek. A-ain
there .ire certain dillerciiecs. These differences are the absence
of certain fish— the Johnny darter, the golden shiner, the straw-
i-olored niinno\v and the common sucker. The presence of tlic-c
.it locality I. of Hickory Creek is probably due to the artificial
exposure of bare bottom.

TABLE III.

THE FISH OF THOKN CREEK COLLECTION MADK AT THE HEADQUARTERS IN 1908
AND IQOQ AND AT OTHER POINTS IN 1909 AND I9IO.

A. The first fish pool; B, four miles down stream; C, ten miles down stream.

\

B

i

Horned dace

.^I'Hlotilll V till < »>!(!( II lilt 11 S

*

*

*

Blunt nosed minnow

I'imi' fliiilt's Hitliilift

*

*

*

Blue spotted «un-tish

l.<'pinnis cvdiit'llia

*

*

*

Stone roller

( amposloma anomalum

*

*

*

\ntrn f>is uniliftitilis

*

•)

Banded darter

Eheostoma zonule

*

p

( i iiiniK >n ~hiner

\<itn>(>is cornutus

*

•)

Striped top minnow

I'ltniinlus ilispar

*

->

Black sided darter

II illh'np/CI'll s l/S/TO

*

*

Johnnv darter

Boleosoma nigrutn

*

*

M ud ininiii >w ....

1 inlit'ii Ihni

*

*

Caviisja minnow

Notfopis t'n\'K^ii

*

*

( ic ildcn shiner

Ihramis CTysoleucos

*

*

Large mouthed black bass. . . .

Mil m pier us salmoides

*

Small mouthed black bas-<

Microplerus ilulomicn

*

Blue trill

l.i'pumis pulliiiiis

*

Crappie .

1'iiniuxis .s/)(;;-<)/(/(-v ...

*

Pirate perch .

. 1 phri'iloili'i'ii \ stiv mi us

*

Yellow perch

I'i'i'i ii ftavescens

*

Carp . .

(.'vfrntns < <ir [>io

*

Black bull head

. 1 ini'ini'iis nii'las . .

*

• i 'in nion sucker

( atostomus < ommersonii

*

Short-headed red horse

\lu\iistnniii l>>'i",'ii i'/>\

*

Pike. .

!•'.<• n.\ I IK his .

*

Comparison of this list with that of the north shore streams
(except 6 and 7) shows that, in so far as the north shore specie:-
are in Thorn-Butterfield at all, they are at the headwaters with

the exception of the Johnny darter.

In column B we note that llie fish are alioiit the same species
as were found in localities III. and IV. in Hickory ("reek, in so
far ,i- they have been found in both streams at all. \'otropis

commas is found further upstream in Thorn-Butterfield Creek
than in I lickory ( 'reek.

ECOLOGICAL SUCCESSION. :S

In column C we note a number of fishes, twenty -pecies and
four doubtful identifications, as against twemy-i\\<> at point \ .
in Hickory Creek. Ten of these are the same in the tw<> stream-.
Again \\e find larger fi-he- in the large parts of the stream.

IV. 1)1-' USSION OF THE D.\TA.

It i- easy to lose on< self in the maze of taxonomic diversity
which one find- in a number of streams separated no further
than tin- one- con-iden-d ht-n-. \\V are not concerned with
taxonomy. Tin- 5U{ i •!)- and inference- herein are chielK to

illn-iraie .1 melhod ot aiial\ -i- and l" -ui^e>t a method of com-
bined experimental and field -tudy. It -lionld be borne in mind
throughout the di-cii--ion that ecolo-y as an or^ani/ed science
i- in much the -am- :n< >rphol- >-\ \\ a- \\ hen it \\ a- in-i < 5-

\ tO dlSCUSS methods which are now taken for granted.

I . m^trilniti in t!n~ .V

\\'e \\ ill i -'ii-ider the IK. rth -In. re streams lir-t. \\\- ha\e in. ted
that the\ .ire -iinilar \« < -.u h other in origin, in material- t i-'-lid,
in their relation- t-. the lake, and in bein^ for the m-.-l part
madi- H|> ot permanent pool- and intermittent ritlle-. An in-;
lion of Table I. -ln.\\- that there i- a definite arrangement ot
li-he- in the-i- Streams. 1 in tlnini-.re. that there i- a cl-'-c
( -oi Te-pondcin i- betui ell the Upper Colll'-c- of t he di Iterell t -tream>

in the matter of the ii-he- pn--ent . a- \\ell a- in their arrangement.

The only -pccie-. the Inn in 1 1 dace, in the youiue-t -iream < '•!» -n-

coe Brook) is the same as the species \\hich i- >n;uc^t tin- source

.// ///c other st'

Tin-re are - .UK- dittei. i,- • •- in the fi-h communilii-- o| corre-
sponding part- of the dittt rent -tre.im-. but tin -e are probably
due to the failure of B -i\eli SDCCieS to i-ntei a ui\eil -iream.
One i- -truck \\ith the >imilarity of the corrc-pondin- li-h com-
munitie- rather than the ditt.-iein,

In making a general compari- m. reference to the diagram <.f
the map- of the -tream- I ig. J shows that the different jish
communities ar« together in the smaller -tream-. I-'i;^. l

-ln.\\- that the -lope of the bed of the voting -tream- i- greater

24 VICTOR E. -HKLFORD.

than that «i the older streams, and the ditYerent conditions accord-
ingly closer to-eiher.

Turning to the other t\vi> >t reams studied (Hickory Creek and
Thorn-Butt uTtield Creek), we see that the same species, the
horned dace, is at the headwaters of these, as at the headwater
of the north shore streams. It is accompanied, however, by
eral other species, a part of which are found in the north shore
>trcams at points further down stream and in a larger volume of
water than some individuals of the horned dace. In other words
the species of the north shore streams are crowded together in
streams \\here the volume of water is greater at the headwaters
than in the north shore streams,
(a) Causes of the Definite Arrangement of the Fishes in the

Streams.

The arrangement of the fish in these streams suggests definite
reactions to some factor or factors in the stream. Rheotaxis
is suggested as a cause of the upstream movement, and water
pressure and size of stream a factor limiting the upward move-
ment. This should be studied experimentally.
(b} Origin of the Fish Communities. — Migration.

A discussion of the mode of origin of the fish communities is
concerned with the mode of entrance of the fish into the habitat.
\Yhile manner of entrance of fish into a stream is not of particular
importance to us, it follows from a reference to Fig. I that fish may
enter when the stream is young and keep pace with erosion. Fish
entering when a stream was at the youngest stage indicated in
Fig. I, need only maintain their position against the current and
they would be carried inland as the source of the stream migrated
inland. In the case of Bull Creek, the horned dace is absent
from the lower portion (Dead River) so far as our collections
show. Did this species enter when Bull Creek was similar to
Glencoe Brook." This is improbable for the condition in Bull
< 'reek, \\ hen it was similar in size to Glencoe Brook, mn>t ha\e
been Hinilar to conditions now found ne.tr glaciers, e. g., Green-
land.

There is further evidence as to the improbability ol such early
entrance in the data of Hickory Creek fishes. \\V have noted that

ECOLOGICAL SUCCESSION. 25

Hirkory Creek rises in a depression between two morainic ridges,
and that its upper course is sluggish and did not originate by
stream en»-i«n. Its profile is shown in Fig. 3, page 19.

The species of fi-.li n<>\v at the source of thi- -tivam could not
have filtered when this stream wa- young and have kept pace
with the migration « .f conditions, because if thi- was a stream at
all during tin- close "f the glacial epoch, it wa- carrying the waters
from melting i( e "r a pond in the same manner a- it is now drain-
a mar-h.

Tin- li-lie- or ~pi , ies of fi-h that are now found near tin- -ouree
of this -l ream wen- obliged to travel a di-tance of \~\\\- miles
through a murky niar-hy -tn-am to reach the headwaters. The

success of mo-i of the - present in the -lu.^i-h upper course

of thi- -ire, nil i- prolial>l\- due al-o \,> l,,c.tl artifici.il i'\po-ure of

vel, since the set tlrmcnt of tlu- country. There ha> been dr.
in- to facilitate drainage. ^i\ of the thirteen >pecir> are known
to u-e bare hottoin lor hreediiv^ I rbes and Kichard-on -how
thai o| the leu . i| these uhich they . on-idered in their taldes
( |)|». ci\ i \iii nine -ln>\\ a preference for the l>ot toin (,|" n,t-k and
-and. ^< Mile .1) lhe-e decided.

Tin- xhou - that li-h may enter the headwater- of -(ream-
\\iihont i»anicnlar reference to plu-io^raphic condition-. Hen-
po-iti\e rlieot.ixi- ma\ 1 »e a fact.ir a-^ain l.\on, '04).

Tin- observations of tin- .,f drought and llood tiirow

further li^ht on the cause and rate of migration.
. ' on ///c . 1 -

;';/

i. /'/.-• fhere was an unusual drought in the autumn of

[908. The ilata on the < li-tril >ut ion of li-hc- in ("ik-ncoe lirook
and ('oiinty Line ( 're.-k \\ ei .• collected before tin'- dale. TaMe

1\". -ho\\^ tin- arrangement the drought.

< >imt\ Line Creek \\a- entirely dr> except the pool neare-t
its mouth in September. ; Thi- i- the locality marked 4 in

. 2. The t"llo\\ini; spring \\a- OIK- of normal rainfall. The
fish proceeded Upi >nil\ three rods. Thi- partially

re-tored the u-iial arrangement. If thi- repre-eiil- tin- rate, ti-li
proceed np-iream -lo\\l\. < ileiicoe Brook ha- not reco\'ered
it- ti-li.

VICTOR E. SHI I I ' iRD.

TABLE IV.

U.IT IKS <>v i in-: BASIS OF THE I)IMKII:I IK.N ,,i FISH BEFORI-: mi. \)H<« <.IIT.

i TABLI-: I.

union

1 >ati ' 3 • ntil'ic N:imc.

i

4

( ,lell . Ill I ti-ll

I ltd iln-i . 1909.

nty Line ( "n-ck.
1 1 1 >i ned dace

September. 1909.
Si'inotilii '• iitnunaciiliitHS

*

*

Black iii i-ed d.u .•

Rinichthys atronasus . . . .

*

iinon sucker.

( '(iti>\!a»!iis I'omniiTsoiiii .

*

2. Floods. — Abbott ('70) collected the fishes from source to
mouth of a small stream. After a flood he found none of the
^une fish but species belonging to larger streams. Unfortunately,
he does not state that he again studied the stream from source
to mouth, and accordingly leaves the question of a general
upstream migration unsettled.1

2. Succession.

Ecologists are frequently asked what is meant by succession
and what is the significance of succession. In answering the
first part of the question there has often been a confusion result-
ing from a lack of distinction bet \veen the different uses of the
term succession. It is used in three distinct senses. We speak of
(i) ecological succession, (2) seasonal succession, and (3) geological
.succession (Adams, '09).

I. Geological succession is primarily succession of species
throughout a period or periods of geological time. It is due

>As evidence of upstream migration of mollusca, the following seems to be

important. Frequent examination of a section of the North Branch of the Chicago

Rivi-r at Kdgelirook, between 1903 and 1907, shoued that l'l< •/<;-... . ->-,i ,-i, -rulum and

(.'iinif'i-luniti occur in this stream, rit-nrucrra \\-a-; not I'ound during tliis period

MIL; .\o\ciiiln-i. 1907) above a certain point. ('inn/<t'li>nn: \\.ts found only

-]iaiini;]>- almvi- this point. Tin- si>ring of 1908 was one of heavy rainfall and the

in- were in tloo.l iiom April to June. On July f) the snail Pli-itnm'ni was lotind

in nu in In i - one t"ui th «.i a mile t'urtln-t up-trt-am, than formerly. < \i»ip,-lomu had

Kone nearly as far. The sea-mi from November to Apiil was not dilien-nt IKUII

LSOns anil then- i- no n-a^'Ui to a— nine that the initiation ln-i;an ln-lon- the

spring floods. Il tlii^ i- tun- tin -nails could make their way towaid the head \\atei -

at the Kite ol at l«-a-t a mile pel year, il they were intioiliiced into a lai.m- -tn-ain.

'I hi- tnii-t lie a te-p.m-e to liotli \\.iti i pii sure and rurrent. The small \alue o!

SUCll -ill^le oli-el \atioll- i- lero^lli/ed, 1)111 they ale ple-e|lted hi'le I.e. .lll-e the

op|ioi tunit\- to • in -111 h data i- -mall.

ECOLOGICAL SUCCESSION. 2~

mainly t<> the dying out of one set of species and (lie evolution of
others which take their places.

2. Seasonal succession is the succession of species or stages in
the lik-hi-to;-' , ies, over a given locality, due to hereditary
and eni'irnnic differences in the life-histories (time of appearance)
of t!u- species living tht

3. Ecolo-i. ,il succession is the succession of ecological type-
< phy-iolo-u ,il t\ •]!(•-. modes of life) over a given point or locality.
duetochai >f environmental conditions at that point. From
thi- point ••(' \ie\v untiring to do with 5/>< • \-cept that
)innii\ an- necessary. There are of rour-e relation- hetweeii
the-, three phenomena hut the-e relation- are of the nature
of rh»-ek- in method of -tudy rather than e— eiitial-. ami -pace
will not permit US to disCUSS them here. Tin re are al-o qualifi-
cation- of a -imilar nature that might IK- made to the
delinilii'ii-. \\iili me to the -tatemeiit thai Species
are inel.-\anl to the que-tion. a \\ord -hould he added. If the
hahit- of a spe< part "I the delmitioii ot that species,
a- the\ iiui-l -ooiier "i later conn- (o In-, tlu-n -peeie- an -i^nili-

( -am . The definition applies to the manner in \vhirh the spe< ies

(|lle-tioll i- Healed ill ]ir.ieli'

\\ . ha\e noted the migration a-|iett of t In- relai ion .>t \\-\\ in
-mall -iieam-. ^ne» e--i< 'ii i- Imt a dittereiit jioint ot \ie\\.
^nei e--ioii of ii-h mean- succession of e-i.iMi-hed torm-. ;. -..
• •I fi'iin- that make tl tin their regular al>ode. The mo-t

ini|)oitallt a-|nel ..| « --t al ili-hlllellt i- hreedillg. AnumluT of

\\riter-ha\edi-iu--ed the breeding habits of the -pet ie- toiind

in the-e -mall streams I ig< nmann, '^5; l;orlu •- and l\ii hard-on.

I Ian kin-oii. '..7. 'lO; \<> i-^hanl. '• 's . Reeves, '07 : ^iniih. '(

:!ctncn( OJ /'• x • J«m.

>ui-et---inii in tin efon u- i- a ret i >n-i met ii m. It i-

1-a-ed on the -uperpo-ition of all the fi-h communities o\er ihe
olde-t part of the olde-t and large-t -tn-.tm. To make this
clearer, \\ e will -late with the aid of the diagram I ig. 4 the
-ucce--ion of li-h in Bull Creek. Thi- -inve— ion \\ill he < -on-
>idered a- taking place over the olde-t jtart of the portion of
Hull Creek \\hich lie- hack of the hluff of the former and higher
lex el- of Lake Michigan. Thi- i- the point de-ignaied a- 5.

VICTOR E. SHELFORD.

\ \

FIG. 4. This figure is based on Fig. i. The profiles of the streams shown there
are here separated vertically at the mouth. The curved lines represent seven
stream stages as follows: B, Glencoe Brook; C, hypothetical stage; D, hypothetical
stage; E, County Line Creek; F, Pettibone Creek; G, hypothetical stajir; //, Bull
Creek-Dead River. The hypothetical stages could, no doubt, be found along the
shore of Lake Michigan, the difficulty arises from the introduction of sewage into
so many streams.

The comparative size of the mouth of each stream stage is representdl by a
stream cross-section at the right.

The direction of reading in succession is indicated by the vertical line with the
arrow heads pointing downward. The oblique lines marked i-i, 2-2, 3-3, etc.,
pass through points in the stream profiles which are in the same physiographic
condition, and which are occupied by similar fish communities.

When Bull Creek was at a stage represented by the first stage
in our diagram (which is now represented by the present Glencoe
Brook) its fish, if any were present, were ecologically similar
to those now in Glencoe Brook in their relations to all factors
except climate. This ecological type is represented by the horned
dace alone. As Bull Creek eroded its bed and became hypo-
thetical stage C of the diagram, the fish community of stage B
was succeeded by a fish community ecologically similar to the
fish communities at the localities marked 2 in Figs. 2 and 4.
The fish now ecologically representing this community are the
horned d.-Mv and the red-bellied dace. The community <>f the
single species, the horned dace, had at such a period moved
inland to the point where line i-i crosses the curved line repre-
-enting the profile of hypothetical stage C.

As erosion continued tin- l'\^\\ community ecologically ivpre-
-i-utrd by the horned d.u .• and red-bellied dace mo\vd gradually
inland and was succeeded by a fish community occup\ing the

ECOLOGICAL SUCCESSION. 2Q

mouth ot hypothetical stage D, ecologically similar to that m>\v
found at the point- v Thi- i- represented by the three daces
and the Johnny darter.

As the hypothetical stage D eroded its bed and became >tage
K, which i- n .u-d by County Line Creek, ti-h community

1 wa- succeeded by a ti-h community ecologically similar to
iln- ti-h community no\\- pn--ent at points 4. This is eco-
.illy rrpre-ciited by the three daces, the Johnny darter and
the \oiin- ot tin common -ticker. The fir- h communitie- di
nated as I, 2. and ; had meanwhile mo\ed inland and \\
arranged in tin- order which their ec, .logical eon-titution required.

Tin- continuation of the process iv-uhed in di-placin- a li-h
<omimmit\ ecologicall) -imilar to the ti>h o.nnnuinity 4 b\ a
!i-h communii\- . . all\ -imilar to the ]ire-ent ti>h i-ommunit\-

5. Thi> i> repre-ciited in the lower \\ater- <•!' Hull Creek by the
bluiit-iio-(-d minnou .ind the little pickerel. HetWe the de-trilC-

ti«,'ii oi its lower course r»-tiibi. ' eek should have contained

ihi- coiiiiiuinit\ and represei 1

I ' . cal succession is one of the few biological fields in which

]m-di(tioii i-, p..-~iMc. \\',- m.,\ r.in-y thi> di-cu — ii.n a little
tmiliri. \\ e ha\e iiotnl that the dt-\elopiu. . .ntinue

to erode their bid-. ^io\\ lar^i-r. and briiu down the surface , .(
the land. The-e ] - h.i\t- not -topju-d in Hull Ctvi-k; it

\\ill bet mile la; -ntain a larger \olume of water at the

!• •• .tlit\ 5, and tin- !i-h commiinit\ o| l.ieality 5 \\ ill be -IK • <-« d< d
b\ a li>h coinimmit\ t -i o|, ,^ie.ill\ -imilar to that now at locality
I ;ire-ented b\ the -unti-h and ba-- pn -< in.

Thi e ha- been d. -unated a- In | n.t lietic.il G in the

diagram, \\ith a further continuation of the process, the li-h
iimillit> of -• g< ( ,. ' . ' 6, \\ill be succeeded b\ a ti-h
community eo>l( . Really -imilar to that now found at the locality
7 Hull Creek-Dead Ki\n-. Sta^e 11 . The ti-h are -ho\\ii iii
column 7, Table I .

.U<7/.'<></ and

The di-cu— ion ab..\e i- intended to illu-tratc a method of
analysis as well as to ill u-t rale the principle- in\ol\vd. In order
to make both clear, more ti-h communities ha\ e been noted than

V> VICTOR E. -UK!. FORD.

otherwi-c would have been justified. It is important to recognize
how much of the reconstruction is method, and how much is
bin]. i^ical principle.

i. The Biological Principle. — The biological principle involved
i- one well ivco^ni/'ed, Inn lacking in detailed and definite experi-
mental confirmation. It may be stated as follows: Ecologically
ci mi iar.d ile animals living under similar conditions possess certain
similarities of physiology, behavior, habits and mode of life.

Indirect evidence for this seems adequate for the purposes of
this paper, but we have as yet been unable to conduct experi-
mental studies in this line. The process of securing data to
illustrate the principles involved here has not been small.

An obvious difficulty arises in securing language suitable for a
brief expression of all that is included in physiology, behavior,
habits and mode of life. The term form covers all matters of
structure, size, proportion and in common usage, color also.
A- opposed to this and as covering all the physiological, and
behavior characters just referred to, we will use the Latin word
mores.1 The mores of a species or community of species are not
independent of the form or forms: the two are correlated, for
every mores, we may expect to find some kind of structure, and
for every form of structures, we must also expect to find some
physiological differences. Our present methods may not detect
these correlations between form and mores and the correlations
may not be important, but the existence of such correlations
seems to be a necessary assumption.

In such a series of streams as the north shore streams, it is a
well-known tact that the conditions in the young streams are
like i hose ( near the headwaters) of the larger and older ones. It
was for this reason that this series of streams was selected for
the study. Since the same species of fish are present in like
condition^, we have evidence for the uniformity ot physiological
make up. Since different species occupy different positions in
the nicely graded Aeries of complexes, we have an index of their
comparative physiological make up which could not be easily
deiei led by other means. We have located the animals in their

\l">, : In -h.i\ im . i 11- ("in-. 'I In- \\"i'i will !»• 11 -'•'! a- a < < .llrct i\ <• IK mil cx.u tly

Form .ni'l forms are n-ni in

ECOLOGICAL SV< < E-MON 31

environment, the fii>t essential in study of their environmental
relations.

2. Method.- -The method of analysis employed in this paper
i- ba-ed on pi tphy- The relation of physiography to

<•< olo",y in thi- nalogou- to the relations of the microtome

to .niatomy, tin- el.-etric nei-dlc t" experimental morphology, and
reagents to phy-iolo-y. Ecology i- in much the same state as
wa- morphology when • papers and treatises on methods

were necessary.

The object of thi- ' > pe of analy-i- and of locating the animal
in it- environment, i- to determine the physiological character of
the animal as a 7. //<*/<•. The method of procedure i- through the
-tud\ of the complex condition- in which the or^ani-m i> nio-t
nearly in ph\ -iolo-i.-.tl equilibrium. >oine p1 in the actual

anal\-i- of the or^aiii-m can dollbtle-- also be made by thi-
method. Variation ami modili. at i« .n of ph\ -iol< .-ical make up
ma\ ,il-o be iletected. I'll'. . :iial\-i- i- then a bio-

. al method. ||oue\er. both th ni-m and the eiuiron-

meiit mii-t be ,mal\/ed mn< h further. In \\\\> further anal\-i-
the succession 01 t he e\ oluiioii of the environmenl i- an important
background and mu-t fr.rm the ba-i- for the ^election of poim-
ot -tud\. lor i oinpari-oii of rr-ult- and oixani/at ion in general.

VmOl IstS and -oine bio! . the ple-elice ot the

-ame or -imilai taxoiiomic ^roii|)- i- olleii taki-n to indicate
-imilariix ••! i ondition-. While CCOloglStS mus! -ometime- follow

this method of reasoning, their method of procedure i> in tin- main
reversed. I he,,-.,. i are studied and the presence or absence

of a ^i\en -et ot <'twani-m- noted. In -uch a detailed -et ol
condition- as the-e \\ith uhich ue are dealing, the old line ot
reasoning -hould not be |ollo\\ed. The mod: liability ol beha\ ior
-peak- a^ain-t it.1 Thi- i- not a contribution to ph> -io^raphy.
The ph\ -io-taph\ in\ol\i-d i- well kno\\ n to element a r> -indent-,
of that -llbjecl .

l'h\ -io^i-.iphic anal\-i- i- a met ho. 1 like all other method- ,.|

science to be impio\.-d. modified or rejected, according as it h.t-

l.-l.v,-//:, Mini, Ml : :i !>»th |)cuitl- ;in,l -Ui-.illi-.

Mi \\ i \:i,-,- In- I mi in I th.il tli<- I M-l i.i \ i,,r , liai.i. tera Ol tl"- imli\ iilual- inlial.it-
inv; tin- streams an.l ponds are •hiirn-iit. !!•• has partially i lum^-il tin- -ti.-.ini
pi.nil »i,.ro l>y kn-piiii; tlu-'ii in poivl cniKlilii'ii- ami vie.

32 VICTOR E. SHELFORD.

served its purpose or sh<>\\- it- defects. It is not, however, the
only method of ecology . and it should not be employed alone, but
accompanied by experimentation.

This same method may be employed in the study of the
historical problems of biology, or to the study of evolution, but
the results are not ecology because this method is employed. Nor
is there any intimate dependence between the fundamental things
in ecology so far as its progress as a science is concerned, and the
di\ isions of biology known as evolution, morphology, or faunistic
geography.

Physiographic analysis is only a part of a more general method
of deducing succession and laying a foundation for comparisons.
The general method of successional study has a probable sig-
nificance which lies beyond the recognition of the physiological
characters of organisms as a whole and the analysis of organisms
as far as the method will permit.

(c) The Significance of Succession.

It will be noted that in the above statement of succession no
reference to species is made. Species in the morphological sense
can have only the most local significance in succession. Species
inhabiting similar stages in the physiographic succession of
streams, i. e., similar conditions — will hardly be the same within
a very small area. It appears that the idea of succession has
been regarded as having little significance for this reason. A
point of significance is the character of the -mores. In the matter
of their mores the fish communities of a stream in Europe, a
stream in Japan, and a stream in the United States are probably
similar if the streams are similar — a matter for experimental
\ erification.

A science with only one method of classifying its data must
constantly fall into error. Up to the present we have but one
organized general system of classification for zoological material-,
namely taxonomy. Taxonomy has been over-emphasized and
carried into fields where it does not properly belong. It has given
us, among oilier things, what has been called composite natural
history. Composite natural history constitutes a large part of
the natural history of today, e-peciall\ as contained in general
\\orks on natural history. The dc\ rlopment of a composite

ECOLOGICAL SUCCESSION. 33

natural history generalization consists in discovering that certain
habits are characteristic of a few known (as to habits) members
of a given taxonomic group of animals, in striking a sort of
general average of the known habits of the members in question
and putting such an average into the form of a general statement
to apply to all the group, known or unknown. Such generali/a-
ti«iii- have their place in didactics, but they are the most in-
accurate phase of zoology. Though often used as a basis for
generali/ati"! rulizations based upon them have a i|tie>tion-

able value.

The crucial question before us is, then, ("an we, on the;>a>i>of
ecolo-jc.il succession or other natural ba-i-. classify and compare

animal mores and make ^-nePali/alii >n- . ,n the ban- ol" >ueh
( -la — il"n ation .md comparison.' If in\ tion an-\\er- in

the at!innati\ i . \\r may or^ani/e a general -\ -tern for the
< la--il"u aiion 1 material- \\hich -hall be -ufticiciitly

independent of other method- and s\-tem- i .f cla--ilic.it ion \«
Bervi nu-aii- i.l" thn. \\iiii4 additimial li.^ht on the ^i-nerali/a-

tioii- i,|" other /• il field-. >urh a- ].h\-ioIo-\, lureditx, or

evolution, Indeed, plant eCol(>Ki-t- ha\e ] -«d laf with

Midi a rla.--ilication i\\'arniin. ' 'OI, and main

other-). Theiin -ill; - .in now re< < .-^iii/i <1 a- of much importance
l.\ l>otani>t- i:eiH rall\ . The lo^i, .md method Lack of their
( la--i!n atioli i- the Bailie as ours. The dr 68 lie in the tact

that dilleMiHe- in ph\ >iolo ;^ical make up in plants i- u-ually

indicated \>\ vegetative J 'lilt ilituitint- in physiological

make ii]> in animal- i- indicated \>\ dilh-n -in e- in //

!•".» oi, ,-i( .il succession has i been -mdicd e\perinu-n-

tall\. l-".\]ierimental >tndii->, if properly conducted, \\ill an>\\er
tin- c|iu-Mioii . ,f ili, signii of all the propositions here prc-

>eiited. Such experimentation -lumld be conducted \\ itli reference
to an iittalyzt. :nl >hoiild con>ist of the comparison

of the inimal- of different and of similar environments.

\ . ^l \I\IAK\ .

I. Fi-hc- ha\e detinite habitat preference:- which cati-e them
to be delinitcU arranged in >tream> which have a graded -eries
of condition^ from mouth to source.

34 VICTOR E. SHELFORD.

2. Beginning at the sources of the streams of tin- devi-lo]>-
menta! series considered, we find the same species represented
in essentially the same order in all the streams, in so far as the
series of conditions is present. The only species in the youngest
stream is the same as the species nearest the sources of the larger
stream-.

3. Migration of conditions for breeding is an important cause
of fish migration, but fish reactions outside the breeding season
may lie often more important than movement of conditions
necessary for breeding over the route of migration. Migration
may even be due to reactions to a single factor.

4. Fish entering a stream will take a position in the stream
suited to their ecological constitution without regard to the
time and mode of origin of these conditions.

5. There is a succession of ecological types over a given point.
Ecological succession is based on similar mores (physiology, be-
havior, habits and mode of life) of fish communities as a whole
or comparable species of communities.

6. Physiographic analysis locates the animal in its environ-
ment and is but a method of studying the organism as a whole
and a basis for proceeding to its analysis.1

VI. ACKNOWLEDGMENTS AND RIHLIOGRAPHV.

The identification of the fish discussed in this paper are by
Dr. S. E. Meek and Mr. S. F. Hildebrand, and of the Mollusca
by Mr. F. C. Baker. Some of the seining was done in co-
operation with Mr. Hildebrand. Without the assistance of these
gentlemen this paper would not have been written. I am in-
debted to Professor F. R. Lillie and Mr. Ellis L. Michael for
criticising the manuscript.

BIBLIOGRAPHY

Abbott, C. C.

'70 Notes on the Freshwater Fishes oi NV\\- Jris.-y. Amrriran .Naturalist. 1870,

p. 99-
Adams, C. C.

'OI Ka-rlrvrlini; and it- F.iunal Si«niliranrr with Illustrations I'rom South-
K-rn I'nitcil States. AMI. Nat. XXXV., pp. 839-52.

1 ' 'iiti iliiitii 'ii Mom tin- Hull /oolojjical l.almiatoi irs <>| the I'nivrrsity of
Chicago, Marino Biological Laboratory »t' San Diego, February 20, K;II.

ECOLOGICAL SUCCESSION. 35

'og I-Ie Royale, Biological Survey of Michigan, Landing. Report Board of
Geological Survey for 1908. Succession of Bird*, p. 134. Succession of
Beetl*--. p. n of Mammals, p. 390.

Atwood, W. W.( and Goldthwait, J. W.

'08 The I ; the Evanston-Waukegan Region. Bull. 7. 111.

:
Clements, F. E.

'05 R. -• ar< h M- th i- in Ecology. Lincoln, Nebraska.
Cowles, H. C.

'01 riant So, i'-ti. nity of Chicago. Bull. II. Geographic Soc. of

< hit mi- 1. A1-" • /., 1901.

Eigenmann, C. H.

'95 Turk' ivin.nmciu and the Variation of its Inhabitants.

1 i. in-, hi'l. A
Forbes, S. A., and Richardson, R. E.

'08 The! Illinois. Vol. III. Ichthy.

N'.it. II.
Goldthwait, J. W.

'09 I'i '1,-y. III. Stati- C.eul. Stirv., Bull. I I.

Hankinson, T. L.
'08 Walnut I .r..' Biol. S State B

lo^i' .il - i I'p :

'10 A: i small Stream I i.m<. 111. M.

Ac. Sci., Ill
Lyon, E. P.

'04 i in Rli. i.l.!-' Am ! XII..

iM .

ReiRhard, J.
'08 Methods of Si Fishes. >* it h .m A. . ..in-

li • th( II ; ill. Hur. I : \ ol. XX\ III..

Reeves. C. D.

'07 I :IK Hi R nl.i'W I' I'.'.ill.. l';'>7, \'nl.

Xl\ . ,.
Salisbury. R. D.

'07 l'h\ -I'.^l.ipli'. .X'

Shelford. V. E.
'07 I'n -limit); r on the Distribution of the Tiger Be< .uid

it~ Kcl.itinn tn I'I. mi > n. Mini. Bull.. \'nl. Xl\'.. Nn. i. ^-14.

"08 l.iti--hi-ti>iii-- .ui'l l.n\.il ll.il'it- ni t Linn. So< J"iir.

i.. Vol. XXX . pp
'IO Ecological Succession '- I'.'.irini; mi I-'i-h CliHuri-. III. M.

A \ i. 111. t O8-IO.

'II I'li\ M. .].•-!. .il Aniin.i! phy. Journ. Murph.. \'nl. XXII. '\Vhit-

in.in X'oluiii. ' ; in ]••
Smith, B. G.

'08 Tin- Sp.i\\ninu II I •: Biol. Bull.. \""l.

X\'.. No. i. p-l8.
Warming, E.

'Q5 Plantesamfund. Grundtrak ai di-n m-ki.loui-ki- riantrjjco.urafi. Kji'i\ i •• nliavn.

EFFECTS PRODUCED BY CITTIXG PARAMKCIUM

CELLS.

GARY N. CALKINS.

CONTENTS. , A,.,

M.i l. -rial ami Mrthods 37

1 allies of Experiments 40

Analyses of Tables and Experiments 47

Analysis of Table 1 47

Analysis of Table II 48

Analysis of Table III 51

Analysis of Table IV 52

Monster Forms 53

General 59

Summary 65

Papers Cited 67

Description of Plates 68

In a previous paper on Urouychia I have shown that the
power of regeneration in a hypotrichous ciliate is a factor of cell
age, increasing as the cell grows older after division, until a
maximum power is attained just prior to cell division. In the
present paper I desire to describe the results obtained by similar
methods of experimentation on a much more difficult subject,
the holotrichous ciliate Paranicciuni. Here in forms from
ordinary cultures, the power of regeneration is very poorly de-
veloped and the nuclear apparatus is more concentrated. Similar
experiments on Parameciitm have often been attempted, but
tin- difficulties in technique are so great that, with the exception
of Balbiani's beautiful work in 1893, little or no extensive work
has been done. The endeavor to ascertain what happens in a
cell in which the power of regeneration is slight and when the
physico-chemical equilibrium is suddenly disturbed is a problem
\\ ell \\ orth the patience and repeated failures necessary in getting
I he small percentage of success, and the results obtained at
intervals during the last three years are. here- brought to.m '. hei
for the first time. In a subsequent paper I shall publish my
results with anoihrr race of Paramecium cuudatnm on regenera-
tion in relation to <vll division.

36

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 37

MATERIAL AND METHOD.

Of the thousands of Paramccinm that I have succeeded in
cutting with a scalpel as in the Uronychia work, only about two
hundred, of which one hundred and fifty approximately are
here described, have given complete records. The cell to be
operated i- placed on a glass slide with a drop of water of
minimum size and the cell is cut with a fine scalpel under a
binocular micro- It i- a comparatively simple matter to

actually cut the cell in two parts but if the knife pa--es through
the macronucleu- »r its membrane, the fragment- are u-cless
for the nucleti- i- never retained and both parts quickly collap-e.
The re-ult- -h..\v a marvellously delicate coordination of the
part-; of the cell, a very -li'Jit injury, as the following record- \\ ill
-ho\v, throwing the ph> -iol, ,-ical mechanism out of ;<ear.

Immediately after cuttit '1 the peristoine depiv— ion tills

out, leaving tin- « ell periphery perfectly -niooth; the mouth
ca\ity, ho\\e\er. does not di-appear but per-i-t- as a miniature
cave in the protopla-m. In from fifteen minute- to half an hour
atti-r the operation the part remaining of the old peri-tome ie-
appear-, movements of translation and rotation are continued
as before, but in mosl ell is truncated and the cut

Mirlace ap|>ear- a- « I. .111 < lit and -month a- though a wax model
of /',:>,:»:<•< iuni lia«l been cut \\ith a -harp knil'i-. If the cut cell
li\ed tor four hour- after the operation it ua- recorded as .1

-licce — fill e\|)erimellt and tile re-lilt- llefe dr-eribcd \\ere m.nle
on i ell- that in\ariabl\ recovered from the -hock of the operation
and li\ed at lea-t four hour- afteru ard-. I'la-nient- li\in.L;
alter thi- four hour test \\ere ke]>t i-olated in -mall watch ^la--
culture ili-he- in a nmi-t chamber. Tin- culture fluid was 24-
hour-old hay tea made by boiling a -mall iiuantii> of ha>- in
tap \\ater. The di\ i-ion rate of normal Purnmccinm kept under
-imilar c.mditii'ii- varied from one to three divi-ion- ]ier da\ ac-
cordinc to the race under nb-er\alioii.

In th' of I'rnnychin both fra-meiits of the cell re-ulting

from a -in^le cut. continue to live for at lea-t 24 hours, and
frequently -uch fragment- continue to live for three or four days
\\ithout rei;eiu-ration. With Pdramccinm, on the other hand,
the smaller fragment almo-t always collap-e-. One or t\\o

38 GARY N. CALKINS.

exceptional cases of continued activity may he mentioned here.
One, a giant form, was cut through the mouth at 11.15 A.M.
At 4 P.M. both fragments \\vre alive, the posterior fragment
swimming actively with the truncate end in advance, the anterior
fragment with the truncate end behind. On the following day
the smaller posterior fragment was dead, the other, anterior,
lived for ~2 hours when it was killed (no. 6). A second case,
cut in the same place, resulted in two fragments both of which
were alive after six hours. On the following day the smaller
one was dead but the other lived and gave rise to two types of
cells, one much smaller than the other and with a blunt posterior
end, while the other had the characteristic pointed end of Para-
meciuin caudal itm. In still another case the giant form was cut
posterior to the mouth, the posterior fragment being about
one half the size of the anterior part. Both parts were' alive
after six hours but both were dead on the following day.

In all of these cases the organisms were treated for from fifteen
minutes to one half an hour with dilute neutral red, granules
in the cell being deeply stained at the time of cutting. What
effect the neutral red has upon the consistency of the protoplasm
I do not know, but certainly there was a marked difference in the
resistance to collapse after treating with this dye. In only two
other cases have I succeeded in keeping both fragments alive
after the operation on normal forms; in one case the smaller
fragment lived only ten minutes after the operation; in the other
case (Table II., no. 63) the original cell was treated with nuclein
before cutting, and both parts lived 24 hours. In the majority
of cases the smaller fragment collapses immediately after removal
of the knife.

Several different races of Paramecium have been used, but
giant lonns on the whole have been selected because of the
greater ease of cutting. The smaller races and races of Parn-
»ir< iinn aurelia do not allow sufficient play for the knife edge
and too great a /one is crushed by the operation. One race of
giant forms was obtained from brackish water at Roscoff, France,
for which 1 am indebted to Mile. Lip>ka. The other races have
been obtained from time to time from wild cultures in the
laboratory at Columbia I 'niver>it \ . As there1 is little or no

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 39

difference in the percentage of successful results in the different
races I shall not particularize but will deal with them all as of
OIK- race. I "p to the present in these experiments, no systematic
effort has been made to study the regenerative power at different
period- between division phases, as in the studies on I'ronychia.
Tin- percentagt of cases of regeneration was so small that -uch
.1 stud} -eemed valueless, but with a race now under experi-
mentation more -ati-factory rc-ults are expected. Of the -even
•f regeneration obtained in the one hundred and forty-
nil)- rei.,rded. five \\ere cells that wen- cut while
either di\idin^ or conjugating. re-tilt- indicating tliat not only
di\ i-ion a^e bin ra< ial age i- al-o a potent factor in regeneration.
Further work on tin- pha-e of the -ubject i- now under \\ ay and

uill undoubtedly yield interc-th Its.

In the follouiiu table- the experiment- are grouped according;
to tlie region df the cell on-inally cut. The cell i- by no mean-
hoi! QUS and the different portion- are not equip»teiit. It
\\e imagine the /'.; :ini \»]]<^ diameter to be di\ided into
I oi 11 equal pan - by three cut-, t he cent ral "lie would pa-- through
the |ilaiie of di\i-ioii o| tin cell, the upper one would divide the
anterior hall into t\\o di-Hinilar quarter-, the lower one \\oiild
di\ide the po-terior halt of" the Paramecium into di— imilar
qiiarlei--. In the table-, the /one- indicating the-e -exei'al
(jiiai lei- are numbeie. 1 from One to four, 1 bein^ the in.i-t anterior,
2, the next anterior. ;, the po-terior central /one and
4 tin- terminal p-.-let i-.r /one text li-un t The
mo-t ini|ioi lain - of the cell are contained in
/oiu-- J and 3, /one j coin ainin;; the macro- and
mii-roinicleii-. and /one ^ containing the mouth,
/one- I and 4 do not contain any of' the more im-
portant organs, and yet, as the -equel -how-. th<
pan- it injured are ratel\ replaced and their ab-eiu e ha- a re-
markable elleci on further acti\itie- of the cell.

40

GARY N. CALKINS.

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EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 45

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GARY N. CALKINS.

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EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 47

TABLE V.

-' MMARY OF TABLES I.-IY.

H .urs.

Alive after 48
Hours.

Alive atV
Hours.

Regeneration.

Cut.

~''S.

,P"

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Cent.

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[49

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31

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8.7

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4-7

2. ANALYSIS OF TABLES AND EXPERIMENTS.

I . .1 n<i!y Tnhlc I .

In the experiment- included in thi- table tin.- cell- were cut
in /one I or through the anterior quarter i>t" the- crll, a part
containing «.nly th«- anterior terminu- of tin- | >eri-t< unc- hut none
of the iiM|iiirt.ini cell . irgans

< M tin- J«» cases n < "fded as having lived at lea-t four hours
.ilti-r the o|>ri.ni"ii, som< Mcd \\ithin 24 hour-: other- -

died without regeneration or di\ i-ion \\ ithin 4S hour-: other- ;
li\e<l lor ~2 hour- or lon-i-r \\hhout regeneration; oilier- J
ulliniateK ;^a\e ri-e to normal de-< vndant -, and mie divided
\\ithin 24 hour- giving ii-e to normal cell-.

Number ll i- a -. " 'd example of i In- M - -• - "I death \\ithiii
24 hour-. Here a (lean cut re-ulted in an anterior -mall fi
nieiit \\hich in-tainl\ collap-ed. and a larger rmmded fragment
in uhich the peri-tome ua- . .|. literati •<!. \\iihin half an hour
the peri-tome reappraied exactU as it ua- 1-elore 6XCCp1 lor the-
-mall part renio\ed Fig [). \" lean an^eiiieiit ol I In- pmto-
pla-mic material occurred and the old torm \\a- perlect except
for the mi — in^ part. So far .1- i \iernal appearance- \\eiv con-
ned ! he i ell -llollld ll.l\ c li\ ed.

t M the !i\e -pctimeii- that died \\ithin the next 24 hour-,
number H> i- a good exampk1. Here no rttort \\a- madr to

• Derate and at the did of 24 hours the tra^nient \\a- -mall and
-lu-ui-h \\ith the remainder of the peri-tome pertect. The cell
died before the end of the next 24 hours.

Number 4 i- a -ood example- of a fragment that lived for 7,;
hour- without regeneration or division. The cut -urtace had

48 GARY N. CALKINS.

developed cilia and regeneration had progressed to that extent
but no further (Fig. 2).

The two cases of regeneration were 3 and 6, only one of which
was a vegetative form, the former being cut while dividing the
latter during a vegetative stage. Number 3 was first cut in the
plane of division separating the two cells; one of these was then
cut again in the first zone. On the following day both cells were
normal P. caudal um, but the cell that had been cut was much
smaller than the sister cell. The cut one died on the third day.

The vegetative form that regenerated showed after 24 hours
an anterior end somewhat blunt but rounded and ciliated, and on
the following day it had divided twice, giving rise to four cells
which could not be distinguished from normal cells. The double
division indicates that the original cell was about ready to divide
at the time of cutting and that removal of the anterior part had
no effect on the subsequent divisions.

Analysis of Table II.

The mortality of Paramecium cut through zone 2 is consider-
ably greater than that for zone I. One chief reason for this is
probably the fact of the injury or destruction of the macronucleus
which usually occupies this region. Of the 68 cases 47.4 per
cent, died before the end of the first 24 hours; 79.4 per cent, were
dead before the end of 48 hours; but if the fragments lived
through this period there was a good chance of continued life
for some days, at 72 hours for example there were only 83.8 per
cent, dead, only a slight increase over the mortality at the end
of 48 hours. The majority of these or 8 out of 1 1 were monsters
while 3 out of the 68 regenerated into perfect cells.

Of those that lived 48 hours or more after cutting, some (5)
failed to divide or else formed monsters; others divided in the
plane of the original center of the cell giving rise to (a) cells with
reduced vitality which soon died; (6) one normal cell and one
abnormal which died; (c) one normal cell and one abnormal
which divided more than once; (d) one normal and one ab-
normal cell which formed a monster, and (e) two normal cells.

Of the four cases living 48 hours or more in which no division
occurred, tin- history was varied; the experiments were numbers

I -I I ! ' TS PRODUCED BY CUTTINC, I'ARAM F.CIUM CELLS. 4')

i. J. '.7 and 68 of Tal>le II. Number i \va- killed after 4*
ami lound to h,i\i- only a fexv nuclear fragment:*. Xuml ><
xva- killed ,tt the end (if <><> hour-. and. although cm during the
pro. . u ii ui, xva- found to have a normal macron ucleii-.

Nuiiil- was dead at the end of 72 hour-.no regeneration

:irred luit the cell xva- \ ery lixely and examination -hoxveda
normal m.u ronm leu-. Number o.s \\,t- 3 vegetative I'-nn which
•vd perfectly from the -h'.ckof the oiu-ratioii and li\ed
willioui i ilion or divi-ioii for a period of three da\ 5.

( ': ' - in which the cut fragment divided in the original

tral plane of the cell, the dividii MH-IU \va- killed in two

for tin- dri.Tinination of the nuclear condition- d)\ l«>th

dau^lr :«-d in .> ca-e- (b)\ an al-iioniial cell \\a- formed

\\ hi( h di\ i«led i hit ( I inn - in nuni-tei - were t'-rmed

in 7 : and : itimi occurred in ., .

I lie di\ idii m \\a-killed eil her duriiiL; di\'i-ioii or

imiiiediaii-l\ .ird- b. determine the condition- of the nuclei

in expeiimeiit- 7 .ind *). Nuinlier ~ \- -ho\\ u in 1'i-. 4. The
pn-ierior till \. 'pan-mix normal, the microiuiclcii- i-

dix idin^ not inallx Inn the macnuiiK leu- i- -onn-x\ hat di-toind.
.\IIMI!MI M was exidentlx an ex-coiiin^ant \\lu-n o]ierated and
dividing t<'i tli. ad time. It \va- killed 24 hour- alter

dix i-ion; the alui'-rmal « ell had one of the lour m-\\ macroinn lei
and -exeial ot the d- Mlelit- o| the old Iliacio-

nucleus ; the other cell was a normal exconju^am I ig. 5. I Mate I

lioth daughter eell- died \\ithotit regeneration in experi-
ment- s. J I and . Nil in I ii i J 1 [sag ..... I « \ample o| the-

Here the cell was cut as shown ii On the following day

then- \\a- a -mall tentacle-like a]i|)enda^e on the cm -ur!

i >n tl nd dax the fragment had dixided through the

original .enter o| tin- . ell into di— imilar daughter (ell- • me

of \\hiih \\a- normal in -hap< . luit xxith a collection of Mack
granule- at t he anterior end ' :ie other cell \\ a- -mall and a-

sharply truncated as when cut , On the third day both were

dead.

\umliei j<» \\a- -imilar in hi-ior\ luit the abnormal fragment
loimed li\ dixi-ion of the cut cell was exttemelx' niinute. In in;.
little more than a -mall -phere with a micleti- and a contractile

50 i.ARY X. CALKIN-.

vacuolc. Its -nuill size \\a> due to the prox.mity of the cutting
plane to that of the normal division plane of the cell. It died
on the second day after cutting (Plate I., Fig. 8, a, b, c, d}.

(c) Division of the Ahaomni! Cell. — In one experiment of this
series (no. 54) the abnormal cell, resulting from the division of
the origin. il cut fragment, divided three times. The vegetative
cell was cut as shown in Fig. 7, a. After 24 hours the fragment
was lively and complete as far as the plane of the cut which was
sharply marked (Fig. 7, b). After 48 hours it had grown much
larger and was particularly broad at the anterior or truncate
end. After <>6 hours it had divided into an abnormal truncate
cell (7, c, and 7, d) and a normal cell (7, ?)• On the sixth day
this abnormal cell had grown in size but retained the original
truncate anterior end (~,f) while the normal cell had divided to
form two normal cells of which one was cut (see Table III., no.
38) and the other lost in cutting. On the seventh and eighth
days the abnormal cell had grown very large again and on the
ninth day divided to form two small dissimilar cells (7, g, //),
each, however, with only one vacuole. On the twelfth day each
of these divided once more giving rise to four small abnormal
cells (7, /', j, "k, /), one of these (/) dying on the same day, two of
the others (7, m, «) dying on the seventeenth day and the fourth
(0) on the eighteenth day.

This experiment resulted in two races of cells, one of which
(from the posterior end) were normal, the other abnormal. The
part lost was never regenerated and the abnormality was finally
transmitted to the posterior end of the original fragment.

(d) Monster Forms. — These will be considered together with
those of Table III. (see page 53).

(e) The three cases of regeneration were nos. 4, 11 and 14.
( M these no. 4 was cut during division of a vegetal ive cell. Within
half an hour after the operation the division was completed, one
perfect cell and a small imperfect cell resulting. In 24 hours
both were living and both were of normal -hape but one was
much smaller than the other. At the end of 48 hours the small
one had ^io\\ n t<> \\ normal cell both in size and shape, but on the
third day it was dead.

No. II was al-n a dividing cell when cut through zone 2. ( )n

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 5!

the following day the larger fragment appeared entirely normal
save for an accumulation of crystals at the anterior end <J;ig. 10,
Her cell wa- truncate. On the second day l>otli were
normal.

No. 14 \\a- a large vegetative cell which had been treated with
neutral red prior to cutting. The -mailer fragment, contrary to
the ii-ual hi-tory, remained alive for -i\ hour- hut was dead the
i\. The larger fragment did not regenerate within 4^ hours.
The \\.iti h glass i • 'i Main ing the fragment was not examined again
for a period of • \<-\ en days when it was found to,, .main 12 1.
giant lonn- .md • mall form- normal save f»r -i

.1 ;////;•> /v '.II.

'Ill-- (i-ll- cui in /on,- ill. gave a higher percentage ot li\ing
fragments than tho-e cut in /one 1 1. 4*> per cent. continued to
li\«- afler 24 horn- as against 42. <> p< -r cent.; \\hile 36.6 pet rent.
loiitiinied to H\. 48 hours as again-t Jo.o per rent. The

pencil • iinni-ler> in both was .ibout the same. \i/..

[2.2 per cent, and i.vJ per rent . but there \\a- not ,i -inglr case
ot regeneration. ( M those that li\ ed Inn did not regenerate, -
.•II- ju-t after conjugation; others - \\ere < ..niu

\\heii cut. \\hile the renuiindiT \\ere laro- \<^,tati\e torm-.
Tin- reai limi- tor tin- most part were -imilar to tho . II- cut

in zone 1 1., the con . forms, or tho-r ju-t mu of (-onju^ai ion.

gi\ini4 the oiil\ n,.\cl n -nits. Tin i. 2, ;. }. s and

I ,:; •', III. Ill nos. I. ^ and 4 the cell- u i • • -n infant -

\\hen cut, the fiamiicin- being kilK'd alter <x> hour- foi tin-
determination of tin- nucli-ar londitioii. Fig. u -hn\\- the
-tructure of nos. I. .; and 4 .it thi- perio<l. the ch.n acteri-t ic
fia^meiited nucleii- -ho\\in- .1 lerminal ol i "iiju^at imi.

It i- po— il.le that tin-, c.-ll- \\oiild ha\e continued to li\e and
to di\ide had the\ had a longer time, lor the\ \\i-n a, the and
normal in appearance u hen killed. It was quite Otherwise, ho\\ -
ever, \\ilh ci-11- i ut dmiiu coiiiugation nOS. -. ^ and o . In all
Cases the fragment- died u it hiu four da \ - \\ it limit .my attempt
at regeneration. In one case no. s the cells were -till united
but dead alter J-4 hour-: in another no. <) both li\ed for several
da\- \\ithoiit regeiieraling or dixiding- in another case no. 21
the\ -eparated and li\ed for 4.^ hours \\hen both died.

52 (,ARV N. CALKINS.

Of the vegetative forms the most interesting cases were nos.
7 and 19, the former typical of the majority of cases, the latter
unusual. No. 7 was a small form cut just below the mouth.
It grew very large but did not regenerate nor divide in the four
days of observation, and was finally killed. The nucleus \\a--
normal although main- fragments of macronuclear material in-
dicated that the original cell was a recent ex-con jugant and was
in about the fourth generation when cut.

No. 19 was more interesting. It was cut just below the mouth
as in the preceding case. In 24 hours it had grown very large
and had divided through the original center of the cell, forming
one normal and one truncated cell (Fig. 9, Plate I.). The normal
cell divided again before the third day forming two perfectly
normal paramecia which were then discarded. The truncated
cell grew very large before the third day and divided on the
fourth day into one small abnormal cell (Fig. 9, b, c, e) and one
larger normal cell (Fig. 9, d}. On the fifth clay the larger,
normal form had divided while the small truncated one had
developed a sharp posterior end and was almost normal in appear-
ance; finally on the tenth day, all were living and all normal save
for a noticeable difference in size (Fig. 9, g, Ji, -i, j). Here, thi
fore, there was a gradual return, through four or more generations
to the normal form, although the single cell failed to regenerate.

Analysis of Table IV.

Cells cut in the extreme posterior end were seriously damaged
although the abnormalities were not so marked as in the other
experiments and 10 per cent, of them regenerated. Only 40
per cent, were alive after 24 hours and only 20 per cent, after
48 hours showing that, although not much mutilated, vitality
was nevertheless badly affected by the operation. There were
no monsters, although in one case (no. 7) the cell divided 24
hours after cutting but the daughter cells had great difficulty
in separating, remaining attached by a narrow isthmus of proto-
plasm for one full clay and finally separating. Within a tew
hours after separation, however, both cell- died. This remarkable
effect was produced by cutting off only the most posterior tip
of the original cell. It seems hanIK credible that so marked an

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 53

effect can be produced by such a minor mutilation and it suggests
the possibility pointed out by McClendon, of different electrical
states at different portions of the cell.

A. tin. in no. 12 the fragment lived for five days without
division »r regeneration and finally died. Peristome and mouth
were normal but the loss of the posterior end appeared to be
fatal.

V>. 15 divided 24 hours after cutting forming one truncated

iterior cell and a normal anterior cell. The physiological
bal.tnc,-, ho\\,-\ ,-r, was again disturb, d for both cell- died on the
follou in- day.

No. i') ua- n«>! -o seriously affected by the operation; the cut
fragment i^rrv. I.<uer each day up to ~2 hours when it divided
into one truncat.-d and one normal cell, the latter dividing a^ain on
the fifth day, the former dividing on the -ixth. forming practically
normal i ell-.

<M the form- that regenerated nos. i ami 3) both were con-
jugating uhcn cut. In no. I both cell- \\eiv cut. >eparuion
lo||,, \\.-.l normally ami each cell had tilled out normally by the
toll,, \\im; day. 1 'lie. however, gt>\\ more and more feeble until

it imalU dii-il. The other was entirely normal and was used for

experiment Ji. In no. 3 onl\' one cell of the pair was CUt, \\\>-
other reiiiaininv: uninjured. Alt,r 24 hour- both were per
in lorin. ahli"ii^li one was perceptibly -mailer than the other.
I '. :h died on the -ixih clay.

Monger . — Of the I^ mon.-ter- only one re-tilted from a

specimen that \\a- cut during di\ i-i<>n. all of the ot her- came It "in
tia^niellt- ot Vegetative cell-. !.!>.'ht of tin- thirteen Uel'e cell-
that had been cut in /«ne II., the other live in /one III. In all
cases the po\\er to divide \\a- limited to the nncleii-, the cell

boi I \ appareniK beiir^ incapable of division.

In no. i, I'.iM, II. the ''II ua- dividiiiL; \\lu-n cut through

the anterior half of the posterior cell (Plate III., Fig. [8). The

ti. lament li\ed for three <la\- \\heii it \\.i- killed for nuclear
-inn inre-. T\\ ent> -four hour- alter cutting the po-terior part
developed a nr\\ peri-toiiu1 and mouth on the cut -urface. but
the original div i-i<>n ua- never ("inpleted. \\ hen killed the i rll
had one large nuclei!- which had grown during the three d.
The other nucleti- had apparenilx been cut away.

54 i.ARV X. CALKINS.

No. 6 (Table III.) was a similar monster, hut was derived from
a normal vegetative cell cut in zone III. After three days the
fragment had attempted to divide; a constriction was piv-un
but the posterior cell was only a swelling with a mouth opening
and no peristome (Plate III., Fig. 19, a, b, c).

In the remaining eleven cases of monster formation many
degrees of malformation resulted from the operation. Of these
the least monstrous was no. 23 (Table III.). The original cell
was cut as shown in Fig. 13, Plate II. There was no sign of
regeneration during the next three days but the cell grew larger
and finally attempted to divide (Fig. 13, b, c). One of the cells
was normal in shape and size; the other, posterior, was smaller
and truncated. The two remained attached for a period of
three days swimming about actively, bending and twisting with
the various cilia in action, until they finally died, undivided, nine
days after the operation. The nuclear apparatus was not deter-
mined. In this case two complete peristomes and mouths were
developed and the cells were attached by only a delicate strand
of protoplasm. The general result was similar to that of experi-
ment no. 7 (Table IV.) in which the daughter cells remained
attached for 24 hours and finally separated, dying shortly after
the separation (see page 52).

Other monsters formed without division of the cut cell were
nos. 35, 39, 40, 51 and 68 of Table II., and nos. n and 41 of
Table III. Nos. 35 and 39 were killed after 4 and 5 days respec-
tively, while nos. 68, n and 41 died in from 4 to 8 days. The
other two, nos. 40 and 51, lived for 20 and 17 days respectively,
and developed into relatively huge protoplasmic masses.

The remaining three monsters, nos. 22 and 25 of Table II., and
no. 17 of Table III., all came from an original form that divided
after the operation into one normal and one abnormal cell, the
monster in all cases coming from the abnormal cell.

The history of only the most interesting of these monster-
need be given, all matters of importance in the other cases being
included in the history of these.

The simplest case is no. 51 which lived, after cutting, for 17
days undergoing two attempts to divide in that time. The cell
was cut as shown in Fig. 14, Plate II., in /one II., and the- frag-

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 55

merit was very lively for 72 hours growing enormously during
that period (Fig. 14, b). Between the third and the fifth day it
divided giving ri-e to a monster as shown in Fig. 14, c. The
mass had two peristomes and two mouths placed as shown in the
figure, and continued to grow in size changing form from day to
day and adding a new mouth on the sixteenth day. It died on
tin- -e\eiiteenth day. Tin.- nuclear apparatus had disintegrated
with death of tin- ma-- and could not be determined. Thi- < ase
gave th>- lar^i -' mass of protoplasm obtained, with tin- tc\\e-t
cell organ-, only one i • mt Tactile vacuole being O!>MT\ »-d and \\ ith
onl\ two mouth- for mo-t of the linn-. The power to gro\\ was
intai i and the in . at enough for -i\ cell-.

No. 17 Table 1 1 1. is another simple case of monster formation
\\hiili dillered from the foregoing in appearing after the lir-t
di\i-i"ii -ub-ei|iieiii to the operation. The cell wa- cut in /one
III. a- -ho\\ n in Fig l -. •:. < »n l he foil- >\\ing day the fragment
had di\ided inio ,m abiionnal and a normal cell (b, C, </). < >n
the -et ..lid da\ the normal cell di\ ided \\ hile the abnormal one
remained as b - gn>\\ ih. < >n the third da y the normal

cell dixiiled once .uain into i \\ o normal celU, and the abnormal
One attempted to divide bin formed a nion-ter with a -mall

anterior and a larger pi • portion. There were two mouths,

t\\o peri-tome- and l \\ o coinraciile \acnole-. ( >n the tourth
<la\ the mon-ier had changed -ome\\hat in axial relation- so
that the form- ol t \\ o paraiin . ia coiil-1 be ma-le mit. The mass
lio\\e\er \\a- ue.iker than on the |)reci-din- day and died on the
follouin;^ tilth' da\. A ] >re] laralion >ho\\ed the pre-eiice o|
t\\o macroiuiclei and t\\o micronuclei in their a|)propriati- po-i-
tion- Fig. [5,

|-".\|terimellt 40 Table II. ;^a\e one of the I)lo-t illtere-tillg

of the monsters Ph< cell was cut in /one II. on the fourth of

February. Fori \-eight hour- later it hail attempted to ilixide
in tin- original center of the cell but formed a nion-ter with a
-mall anterior cell and a full >i/e posterior cell Fig. 17, a and 6).
T\\o complete peri-tome- uith mouth- were piv-ent and two
contractile vacuoles. The-e condition- \\ere retained \\ithout
further morphological change >a\ e growth in size until Feb-
ruar\ lo. or the -i\th da\ , \\ hen a drop of nuclei n was added to

56 GARY N. CALKINS.

the ten drops of hay infusion medium. On the seventh d.iy the
monster had grown enormously ' Fig. 17, c ); on the ninth day it
gave rise to a small free cell which \\as an exact miniature copy
of the original cut fragment (17, e). This cell arose from the
upper end of the monster as shown in 17, d. On the tenth day
the isolated fragment was killed for the determination of the
nuclear apparatus and was found to have a normal full size
macronucleus. On this day also, the monster divided at the
point .v forming two monsters, one having three mouths (17, g)
the other with only two (17, /), but the latter at the time of
division, was also dividing to form a small terminal deformed cell.
Thus two divisions were going on at the same time in the proto-
plasmic mass. The small deformed cell was detached from the
parent monster on the same day and appeared as a minute
reversed duplicate of the original fragment (Fig. 17, //). This,
like the first detached fragment, had a single contractile vacuole.
On the eleventh day the small fragment was still alive and active
but had not regenerated nor grown in size while the two monsters
had grown to look more like paramecia fused (17, i and j). On
the twelfth day the small fragment was dead while the others
remained as before. On the thirteenth day the smaller monster
had grown much weaker and was killed on the eighteenth of
February or the fourteenth day (17, k). The remaining monster
was very plastic changing shape from day to day until on the
twenty-fourth of February appearing very weak, it was killed,
twenty days after the original operation during which time no
more than three mouths were formed in this part of the divided
monster. The original fragment thus developed seven mouths
indicating at least seven attempts to divide while the monster
divided once to form two monsters. Preparations (17, k and /)
show relatively enormous nuclear masses.

One interesting feature of this experiment was the form. it ion
of free-living but abnormal fragments separated off from the
protoplasmic mass. In experiments nos. 22 and 25 (Table II.)
the mon-trrs gave rise to similar cells either attached (22) or
free (25), but in ever) case the^e offshoots were normal in struc-
ture although abnormal and weak in function.

In No. 22 the cell was cut in /one II. as shown in Fig. 16, a,

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 57

leaving the lower part of the peristome, the nucleus and the
mouth intact. Twenty-four hours later the truncate cell divided
in the plane of the original center into a small truncated anterior
fragment (b, c, Fig. 16), and a normal cell (16, d). The abnormal
fragment had one contractile vacuole, the normal cell t\\<>.
Seventy-two hour- after the original cut the normal cell
divided into t\. :ectly normal paramecia. which divided

again on the following day into normal forms, thus establishing
the entirely normal ronditioii of this product of the original
truncate fragment. The almormul fragment (c) neither re-
generated nor divided for the lir-t live days after the operation,
althoii insiderably in size (16, e). < >n this fifth day,

howe\er, it atteni|)ted todixide, hut cytopla-mir di\i-ion failed
and a mon-ter with two mouth-, two contractile vacuole- and

two blunt ends was formed i(>. ''/ 1. The power of di\ i-ion seemed

to : ..red for on the nxth day the ma-- attempted a

••nd di\ i-ion. an«l tin- time a perfect Paranicciiini wa- formed,
but. as in experiment no. 7. Table I\ .. it could not -eparate from
the parent ma— and remained attached by a delicate connection
throughout life of tin- mon-- . < >n the -e\eiith,

eighth, and ninth da\ - the mon-ter gradually a — unied the in-
definite outline- of three paramecia Fig. M>. ;i hut when finally
killed on the ninth day tin -e ••utlim •- WC1 in l"-t (j). The

macronuclei were properly distributed for three indi\ idu.ils bill

the micioiilK lei Were ilK KM-ed in number, -ix |ierfect oile- being
loimd. -eparated h\ c"ii-iderablr di-taii(f from the macronuclei.
In expel inn lit no. - s not »nl\- \\ere normal indi\ iduals fornn -1
but lhe\ were detached and li\e<| for -onie da\ - a- onlinar\
paramecia. Tli> .,etati\e cell \\a- cut in /one II. on

JanuaiA Js, lea\ing the nucleus, the lower part of the peri-tome
and the mouth intact 1 ig. Jo. n, 6). < >n the t \\ cut y-nint h the
ti. lament di\ided in the original renter of the cell forming
-mall truncated anterior fragment and a normal po-terior cell
1 . 2O, r, </ and (•). The normal cell, a- in experiment 22
above, divided on the following da\ forming normal cell-, and
the-e continued to dixide normally on the axerage of once a
da\ throughout the life of the other fragment and wen- discarded
on death of the latter. The anterior truncate cell (e] grew

i.ARY N. CALKINS.

much larger during the 48 hours after the operation and tin- 24
h<mrs after division, and attempted to divide on the thirtieth,
i he same day that its normal sister cell divided. The result of
this effort was a monster with two mouths and two vucuoles
l-ig. 20, /). On the first of February, or the fourth day after
tin- operation, the monster attempted a second division, this
time a double division, the result being one free individual and a
monster with three mouths (Fig. 20, /;, g). This free individual
\\.is smaller than the normal but it lived for 48 hours and died
during the process of division. It was detached from the parent
mass at the point corresponding to the upper end of Fig. 20, g.
The monster, meanwhile, continued to grow in size and on the
sixth day had two perfect Paramecium buds attached by their
posterior ends, and several papilla-like outgrowths which con-
tinued to grow like buds until on the ninth day well-defined
Paramecium cells were formed but remained attached to the
mass of protoplasm (Fig. 20, i, j). On the ninth day a second
normally formed cell (k) was detached, this being the upper one
figured in 20, i. This free form did not attempt to divide but
died before the end of 24 hours. On the twelfth day, still another
free form was given off (Fig. 20, n~), this being the individual on
the left side of Fig. 20, /. This one was small like all of the
detached forms, but appeared to be normal, living for 36 hours.
On the tenth and eleventh days the outlines of the previously
distinct paramecia buds on the parent mass, became obscure and
the individuals seemed to fuse with the protoplasmic mass while
the latter twisted and turned with the various ciliary movements.
At one period on the tenth day the mass became drawn out into
two main lobes (/) with a connecting strand between them and
appeared to be dividing, but the figure was due to the mechanical
pressure caused by the presence of a girdle of zooglcea about the
middle. When this girdle was removed with needles, the proto-
plasm returned to the more solidly massed state (m). A similar
constriction appeared on the thirteenth and fourteenth days and
/ooglu-a was once more removed and the obscure outlines o!
ghostly paramecia again appeared, no less than eight bizarre
forms stivtrhini; out from the parent mass at one time while six
more mouths could be counted on the general surface (Fig. 20, o).

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 59

On the following day or fifteenth day after the operation, t\v<>
dead masses of protoplasm were found; the monster (Fig. 20, 0)
had divided in the middle and the two separated ma--e- evi-
dently licid n«.t been physiologically balanced, and had died.

In preparations made with the dead ma--c- no definite nuclei
could 1,.- (li-iin-ni-hed, ih,- matt-rial had evidently died during
tin- ni^ht and had liei-n dead too many hours for the preservation
of the nuclei. A large granular ma-- in one of the fragment.- may
ha\e I.e. -11 tin- remain- ol mi' lei perhaps aggregated into a mul-
berry mass as <le-. ribed l>y Balbiani.

Gl ' i RAL.

I'.all.iani'- coiiclu-ioii that / •• inn; doe- not regenerate as

do oilier |iroto/.,a i- too -ueepii,. < nerali/at ion. A small

p'-P enlace i.| cases do regenerate. Lilt the percentage xaries

with dim-rent i In the e\|)eriment- hen- ..utlined. three

diltereiit ^iaiit races "I I'urii n:fiium «iudatum were u-fl. ( >ne
race, ln>m bracki-h \\.iter in R..-C. .1: approximately IO

pel cent. "I regeneration-; another race, from New York. -a\e
approximately I per cent.: another race, al-> from New York.

gave a relati\el\ hi'Ji percentage of regenerations, approximately

JO per Cent., and in a foiinh race now under examination CVCTJ

one »r KM) per cent, regenerates. The method eim>l..\e<l by

Halliiani \\a- l" in-lead of individual- one by one.

This method made it impossible t,, determine iweniy-ioiir hours
atleru anl-. \\lu-iher the perte. i d-11- had not been cut at all. "i

had been rut and had regenerated.

I'roin my re-uli-it appear-, therefore, that diltereiil raie.-haM
dilten-nt p«. \\ers • >f te.-eiier.uion. ..r, st.iti-d in .mother \\a\ , h.i\e
\ar\im; decrees of \itality. This po\\er of regeneration is con-
nected in some wax . apparently, With the phy.sical make-up of the
protoplasm. In oi her forms of |ir«to/oa, notabK in Stcntor i man\
observers, including lialbiani. ( iruber. Nu--!Miim. \\-rworn.
Tioua/ek. Lillie. l'o|»ot't and other- . in Loxophyllum (Holni'
in StyliiH \rhin, O.\-y!rii'li<:. +{->; >t>s!<nniini , l:rontonia, Trachcloi,-r(a,
Dili-ptns and Sputhidiitm, all of which, sa\-e Loxophyllum,
I ha\e experimented with in tin- connection, the protoplasmic
walls come together more or le-- ([iiirkh' alter the operation of

GARY N. CALKINS.

cutting, thus leaving no endoplasm exposed. In Stentor, Spiro-
stomum, Trachclocerca, Spathidium, Dileptns and Froutonia, the
walls come together in a relatively short time, but in Stylonychia
and Oxytricha, the union is delayed and sometimes never occurs.
In Paramecium, on the other hand, with the exception of the
fourth race mentioned above, the cut fragments form a new
im-mbrune upon the cut surface and the normal form is not re-
gained, the cell dividing asymmetrically. Where the vitality is
sufficiently strong for regeneration the normal form is occasionally
gained before division of the fragment, but more often after
a gradual change requiring several generations. In the fourth
race, on the other hand, the walls of the fragments behave exactly
as in Stentor or Lo.vopliyllnni and regeneration is perfect.

The difference in regenerative power may be due to the dif-
ferences in potential at different periods of the life cycle, and
correlated with differences in the physical make-up of the proto-
plasm at such periods. This certainly appears to be the case
with the power to divide, abnormal divisions or monster forma-
tion taking place at the end of a long series of generations (see
Calkins, 1904), and in those races of Paramecium where regenera-
tion occurs in a small percentage of cases.

Child and others have concluded that movements have much
to do with the restoration of form in regenerating animals.
Holmes shows that this is of secondary importance in the re-
generation of cut fragments of Loxophyttum. So too in Para-
mecinni, movement apparently plays no part whatsoever in the
restoration of form. The old peristome is always the site of the
new mouth formation, but in the majority of cases, if the mouth
is cut away, the anterior fragment with the bulk of the peristome
fails entirely to form a new mouth. The general shape of the
cell is usually retained; there is no rounding out of the cut frag-
inciit any more than there would be were the Paramecium made
of cheese. Form, therefore, is a highly stable characteristic of
Paramet i 'n»i. This is demonstrated with remarkable clearness
in the case of UK Misters. A two mouth monster is, at first, little
more i han an elongate amorphous mass of protoplasm (Figs. 14,
[5, Jo). As it grow- older, however, the typical shape of the
mi-m begins to appear. Still more remarkable is the budding

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 6l

out ot individual forms in monsters of older growth. H<
the mass is watched from day to day, fairly complete cells make
their appearance only to be absorbed again in the general m.
This absorption i- .-analogous to that resulting from a \v<nmded
P(ininn'finni in which the cut failed to extend entirely through
the diameter ot the cell. In such cases the wound is closed and
the cut -urfaie- fuse within a short time. Experiment no. 25
•d illustration of the appearance and disappear-
ance ot individuals Inun the common proiopla-ni'V ma— of -uch
a mon-ter. tin- bud- Jway- growing into a -etnblance .it lea-t
of the typical I'nr<. m.

\ orm in Paramc> iuni, therefore, i- a per-i-teiit characteri-tic,
apparently independent of movement, and connected in -ome
\\.iv \\iih I In- phy-ical make-up of tin- protoplasm.

Regeneration oi tli .i-m which occur- in a -mall percent.

- \ar\ing in ditlereiit races, i- probably due \» a perfectly

balanced plu-i'-lo:.!. il condition of the cell. This conclusion is

dedmed ironi the tact obiained in main experiment-, that a
coni|>arati\el\ in-i^niln ant i lit \\ -hereby only an e\uvmit\ of a
I'uriinn-i'inni i- r<-ni"\ed. ha- a profound effect upon the further

activities of the eel!. Kvunple- are J\ell in eXpelimelll- of

Table I. and of Table IV.. \\ here "in- or the other end of tin- i ell
\\a- remo\ed. lii the inajontN ..I '•-iilnal fragment

died \\ilhin a -hoiM period; in a te\\ cases tin- le-idual tia^im-nt

di\ided asymmetrically, ^i\in^ ri-e to on«- normal Paramecium

\\hich coiitinui-d to |j\e and to multiply normally, ami an ab-
normal cell \\hich - i died: in a leu • :n, the fragment

• neiated either i >v -ooii alter di\ i-ioii.

l! tlii- phenomenon could be explained \ve -Innild be wi-11 on
the \\a\- to an e\| ilan.it ion of that ^ho-t- train of processes u hich

we designate vitality. I k of a balancing of pr , or

of a -table and un-table equilibrium. ln>\\e\-er, i- no explanation
of what take- place. N.me recent ob-er\ ei> ha\e imdert.iken
the ta-k of explaining cell ph\ siology, iiu hiding di\ i-ion. through
the \ar\ingratioof nuclear ma-- to c\ topla-mic ma — . Balbiani
bi'lie\ed that nion-ter.- are due to some slight injury to the maCTO-
nucleus in cutting, and made tin- !i/atioii thai tlu-\- are

"alway- indi\ idual- mutilated in the anterior part who-e de-

62 i.ARY N. CAI.K1.V-.

scendants give rise to abnormalities. . . . Those which lose
the posterior part only, always multiply normally by fission. . . .

A sort of paralysis attacks cells thus injured in their nuclei"

('92- P- 78).

The number of monsters (four) obtained by Balbiani is too
-mall to support such a generalization. My experiments, in
which live of the thirteen monsters were derived from cells cut
in the posterior half throw this suggestion out of further con-
sideration.

A more modern view of the nature of physiological processes
of the cell was suggested by R. Hertwig ('03) and has been
elaborated by himself and his school in numerous publications.
This theory according to Popoff's ('08) presentation, involves the
view that the quotient obtained by dividing the cytoplasmic
mass by the nuclear mass is fairly constant under "normal"
conditions, and just as long as this relation varies only within
narrow limits, the cell functions are normal. But if by dis-
proportionate growth of either cytoplasm or nucleus, the nucleus-
protoplasm relation is changed to favor either one, then the cell
gets into an "abnormal" condition. In order to become "nor-
mal" again, the "normal" nucleus-protoplasm relation must be
reestablished. Cell division, Hertwig believes, is the means
whereby normal relations are reestablished. He postulates,
furthermore, two periods in the growth of the nucleus; one
"functional." the other "divisional," the former beginning shortly
alter division of the cell and lasting until shortly before the
following division, the nucleus growing less rapidly than the
cytoplasm and disturbing the nucleus-protoplasm relation in
fax or of the cytoplasm. This disturbance of "normal" relations
persists up to a certain point which Hertwig calls the nucleus-
protoplasm -tension -moment i Kernplasmaspannungsmoment),
u In'di In- Hoards as the immediate inciting cause of cell division,
tin- first effect being the rapid "division growth" ot the nuclei!-.
The result of the division 1 1 HI- started is a return to the " normal "

A

nucleus-protoplasm relation.

The use ol terms normal and abnormal in this connection i-
hardlx appropriate, for cell division is certainly a normal process
and .ill Stages leading to it must likewise be normal, hence an

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 63

abnormal nucleus-protoplasm relation in a normal vegetati\e
tell, cannot exi-r. Waiving this verbal matter. however, and
e\ iminiiig the principle involved, \ve admit the change in the
relative masses of nm leu- and cytoplasm in periods of gn>\vth.
As to the gro\\ t h of the nucleus prior to cell division the evidence
i- not -o clear. In I'nniychia transfu°a at division, for example.
Hot only i- ihe ' i -lirface of the nuclear element- lc— etled

Imt the mass as well is decreased, and the two -mall cells resulting
from the divi-ion <oin.iin -mall nuclei which -ub-e(|Uently
break up into fragment- before growth commence- and with
-i"\\ th there i- a constantly increasing -ur face of nuclear matter
1 ilkin-, 'll . On the Hertuig theory u e -hould expect a
re-iii ution b\ divi-ion, of tin- "normal" ma-- relation- in
flagmen!- of idl- containing cither a preponderance of nuclear
matter or of t \ topla-mit . I igi < • oi cell-. ho\\e\ cr. -ho\\
no -inh regulator} ; . for tutting th«- fell- iiixariably

po-tpoiic- di\i-ion mile-- the di\i-ion i- already undi-r \\a\.
I ndci abnormal t oin lit idi- of temperature Pop. iff find- that
the volume i,f the niulcii- i' - more rapidly than that o|

the c\topla-m. \arioii- observers ha\e noieil that under ab-
normal i < -in lit ion-, geiieialK . it u I u< ling t hat of \\ <-akt net 1 \ itality
tioin .m\ cause, the mil leu- i- relali\c-l\ larger than under
normal condition^ and the enlai Cement i- bi-tter interpreted
as an ellcct o| lc--cin-d \italit\ than ii- « an-. . p. .poll liki-\\i-e
|>ci!oiincd i lilting cxpciimcin- "ii l-'<<nitnnin . and louinl. a-

lialbiaiii ami other- ha\e loimd. that the cell, it cut late in an
inter-tli\ i-ional period. di\ ide- in the original plane of <li\i-ioii.

lint if the cdl i- cut i •• hi- period the cell tir-t regenerates

before di\ iding. 1 le t oiu ludt - " 1 >!. 56 \ < T-IH h /eigeil, da— tier
AnstOSSgebende Moment d«-r rheilung in dein . \iigeiiblit k der

Kernplasmaspannung /u -m hen i-t. ^o \\ie die /.die die-en

M.-ment uber-t hrit ten hat. i-t der Teilung dci-elben -choii cinge-
leitet und kein ausserei I ingrill kann -ie mehr \erhindcrn"

'08, p J37). It i- tlitfit nli • this conclusion from the facts

pre-cntcd ami -till nu>re dilticult to interpret Mich ca-e- a- my
experiment U), Table III., \\here t \\ « • SUCCCSSlve di\ i-ion- of a
fragment were entire!) a-\ mmct rir.d SCCp. 52 . Inthi-ta-e
furthermore there \\a- a long period bet \\eeii di\ i-ion- \\hen

64 <.ARY N. CALKINS.

"normal" condition- ot the divi-ion energy might have been
reestablished.

In reference to my Paramccinm work of 1904 Popoff states that
at tin- final period of depression the nucleus-protoplasm relation
was changed to the ad\ .image of the nucleus and he argues that
this condition may have been the cause of the depression (he. cit.,
p. ,VV)|- As my paper of 1904 makes no reference to the
nucleus-protoplasmic relation he drew his conclusion from the
photographs of the cells at the end period of depression, but nat-
urally, he did not realize that when the cells thus photographed
were killed, they had lived for days without dividing. Under
-uch conditions, like any other Paramccinm cell under similar
conditions, they were "abnormal" so far as the nucleus-proto-
plasm relation is concerned. '

In the latter part of the first section of Popoff 's paper of 1908
he gives an inkling of a possible interpretation of the nucleus-
protoplasm relation in the statement: "Die Kernplasmarelation
wird ein morphologischer fassbarer Ausbruck der jeweiligen
Chemismus der Zelle bleiben." With this interpretation we are
inclined to agree, and we look upon the changeable nucleus-
protoplasm relation as an unstable effect produced by varying
conditions of nutrition, temperature, or vitality and not at all
as a cause of division or depression or of vitality. In a mutilated
Paramecium the nucleus divides equally, the cell unequally; the
smaller fragment has a full size nucleus and a much deranged
"nucleus-protoplasm" relation; yet, in some cases at least, it
behaves like a normal cell, dividing at the proper time and again
forming dissimilar products, one of which is perfectly normal,
the other again abnormal (expt. 25, p. 57).

By removing a portion of a cell there can be little doubt that the
ordinary chemical interchange, or perhaps the physical or
electrical potential, of cell and nucleus is violently disturbed.
In forms with a labile protoplasm, as manife-tcd in some fonii-
of Paramccinm and as in Stentor, Loxophyllum Spathidium, etc.,
the wounded cell regenerates quickly, but in other races of
Pnninn-i inm and in non-nucleated fragments of other protozoa,
ihe more -table protoplasm does not respond and regeneration
fail-. l)ivi-ion of the fragment indicate- that the power of

EFFECTS PRODUCED BY CUTTIX<. PAKAMKCIl M < ELLS. 65

division and the power of regeneration arc entirely independent
phenomena and are alike independent of the nucleus-protoplasm
relation BO far as mass is concerned. The failure of a small
fragment to divide completely might be due to the fact that the
divi-ion energy i- not great enough to overcome the -ur:
ten-ion of the body \\.dl-. Hut t hi- interpretation would hardly
account for the incomplete divi-ion of the cell in which only
a -mall portion of the terminal protoplasm is removed experi-
ment 7. Table IV.. p. 5J

^l MMAHN. .

1. I'nriinn i ;n»: miulatnm ma\ be ea-ily rut \\ith a -calpel.
one. tin- larger, part, continue- to li\-e in about ^^ per cent, of

and both part- ne\ er lor more than 24 hour-. If the
inn Icn- is injured by the knife neither part live- for more than
a te\\ hour-. The piv-ein paper i- a -ummar\ of about 15-'
rec< -idi d experimei

2. ( ell- cut at either cxtiemitv are ju-t a- much deinorali/cd
a- iho-e cm in apparent!) mme vital part-.

3. MM po\\er i.t rej 'i"ii \arie- in diftefeiit races "I

"giant" P In die race "iil> about i per cent.

^•iieiaicd : in another bout lo |>er cent., in a third i

about v ]„•! cent, n venerated, and in a tourth race all, or loo
IHI cent, n generated. Thi- la-t I.MC i- not included in the
ple-elit paper.

4. There i- Strong t \ idem t "I a di\ i-ion /OIH- in l\irnmci nnn

which lies in the center of tin- i ell. It tin- cell i- cm anterior

or po-trii.-i- in tin- /<nu' the fragment di\i(K-- in the original
]i!ane into a truncated abnormal !<>rm and a normal form. The
truncated form ma\ divide a^ain not through it- center, but
through the center ot ilu- cell \\en- it perlect.

5. A Iragment. \\hethei anterior or |.o-trrinr. die- \\ilhout
di\i-ion in tin- majorit\ of . in cases "I li--i»n. it di\id«--
a-\ mnu'IiicalK a- -tated above. Tlu* di\ i-ion. ho\\ e\ er. i- re-
tarded. In a feu cases the di\ i-ion i- abt>rii\e and a nioii-ter

results.
o. After such a division into an abnormal and a normal cell.

the normal cell continue- to divide normallv and form- a race of

GARY N. CALKINS.

individuals entirely unaffected by the operation. The abnormal
cell in some cases, divides again asymmetrically and forms
another normal cell and an abnormal cell; in other cases the
second division is abortive and monsters are formed; in a fra-
cases it continues to divide with a gradually decreasing abnormal-
ity until normal forms are regained; in many cases, finally, it
dies without further division.

7. The monsters represent as many individuals as there are
mouths. In one monster as many as fourteen mouths were
present at one time. There is a well-marked tendency of the
protoplasm to assume the form of a normal Paramecium about
each of the mouths, and such individuals bud out of the proto-
plasmic mass, remain for several hours, and may be absorbed
again into it.

8. Free cells, complete in all respects, may be given off from
the protoplasm of a monster. These may live for days and may
even divide, but vitality is weak and they invariably die.

9. Cell division and cell regeneration are entirely independent
phenomena. A fragment divides without regenerating and the
abnormal product of this division may divide again without
regenerating. In other cases the fragment divides asymmetrically
and the resulting abnormal cell regenerates before the following
division. The cell need not regenerate to divide, and may or
may not regenerate before dividing.

10. The Paramecium cell acts as a unit; cytoplasm and nucleus
are equally important, a small loss at either extremity is usually
enough to throw the physiological mechanism out of gear and
lead to death, to asymmetrical division, or to monster formation.

1 1 . The phenomena resulting from cutting Paramecium cannot
be explained on the Kernplasmarelation theory of the Munich
5( hool. Mass of nucleus in relation to mass of protoplasm is
only a morphological index of the physiological (chemical,
electrical) activity of the cell— an effect and not a cause of the
\ .moiis vital reactions. •

12. Regeneration of a cut cell in race> with a limited po\\ ei ot
regeneration, is more often observed in cells that have recently
conjugated, or in cells that were cut while in conjugation, than
in ordinary \ei;etaiive cells.

EFFECTS PRODUCED BY CUTTING PARAMECIUM CELLS. 67

PAPERS CITED.
Balbiani, E. G.

'93 Merotomie des Infusoires cilie?. Ann. de Micrographie. Vol. V.
Calkins, G. N.

'04 Studif-s on the Life History of Protozoa. IV. Jour. Expt. Zool.. Vol. I.

"ii Regeneration and Cell-division in Uronychia. Jour. Expt. Zodl., Vol. 10.
Hertwig, R.

'03 l'f\jf:r K'.rr«-!ation von Zell- und Kerngrosse und ihre Bedeutung filr die
< 111'-' lil: :erenzierung und die Teilung der Zelle. Biol. Cent.,

Holmes, S. J.

'07 'I In- Bi-ha\i ••. "phyllum ami its Relation to Regeneration. Jour.

Yul. 4.
Popoff, M.

'08 I-;.\inTiiiii-iit«-lI'- /••ll-tii'li.-n. Art h. I. /. llli >r-ch., \'i>l. I.

68 GARY N. CAI.KIN>.

DESCRIPTION OF PLATES.

Unless otherwise stated, all figurr- an- made from camera diawings or -keu ln--
from the living cells. The black line? indicate the planes of cutting. All tigmes
are of Parameeium caiidatuin.

PLATE I.

FIG. i. a, vegetative cell cut in zone i ; b, the fragment at the end of four hours.
It tiled within 24 hours. Kxp< i iinent II, Table I.

!•!<,. 2. a, a similar cell cut as above; b, fragment 72 hours afterwards showing
neither regeneration nor growth. Experiment 4, Table I.

Fi<;. 3. a, conjugating ]'in-<i»n-i iitm cut in zone 2; b, fragment 24 hours after
the operation; c, rlie other, normal, cxconjugant; d, the fragment fixed and stained
after 96 hours, showing normal vegetative nuclei indicating that conjugation had
just begun when cut. Experiment 2, Table II.

FIG. 4. a, vegetative cell cut in zone 2; b, the fragment 6 hours after cutting;
c. the fragment dividing 24 hours after cutting and killed during division to show
the nuclear apparatus. Experiment 7, Table II.

FH.. 5. a, vegetative cell cut in zone 2; b, fragment dividing, 16 hours later;
c, anterior, and d, posterior, cells killed 24 hours after division for the nuclear
structures. The disintegrated fragments of macronuclei indicate recent conju-
gation. Experiment 9, Table II.

FIG. 6. a, vegetative cell cut in zone 2; b, fragment 24 hours after cutting, with
tentacle-like process; c, fragment dividing in the original center of the cell 48 hours
after cutting; d, e, resulting daughter cells 6 hours after division. Both of these
cells died on the following day. Experiment 21, Table II.

FIG. 7. a, vegetative cell cut in zone 2; b, fragment 72 hours after cutting;
c, fragment in division on the fourth day; d, e, daughter cells resulting from this
division;/, cell d on the sixth day, much grown; g, h, daughter cells of /on the ninth
day; /, j, daughter-cells of /; on the twelfth day; k, I, daughter cells of g on tin-
twelfth day; m, n, figures of i and k on the seventeeth day when they died; o, figure
of j on the eighteenth day after the operation when it, too, died. Experiment 54.
Table II.

FIG. 8. a, vegetative cell cut in zone 2 near the plane of division; b, division
of the fragment 24 hours after cutting; c, d, resulting cells, the former minute, the
latter normal. Both died on the second day. Experiment 29, Table II.

FIG. 9. a, vegetative cell cut in zone 3; b, fragment 12 hours after cutting;
c, same dividing 24 hours after cutting; d, e, daughter cells resulting from division
of b;f, division of truncated cell e on the fourth day; g, figure of truncated posterior
half of /on the eighth day; h. i, daughter cells of the normal half of/, on the tenth
day, j. figure of g on the tenth day. Experiment 19, Table III.

FIG. 10. a, dividing cell cut in zone 3; b, division figure aiiei removal of tin
anterior end; c, small anterior cell alter division; </, normal posterior cell alter
division of b; e, figure of c 24 hours latei ; f, li-me oi </, J4 hours alter di\ ision of b.
Experiment n, Table II.

Fit;. 11. a, dividing cell cut in zone 4; b, posterior liagment with mass of
crystals in the anterior end. This died on tin ii.llnwing day. Experiment 5,
Table IV.

FIG. 12. a, ex-conjugant cut in zone 3; b and c, tin anterior fragment 4S and
96 hours after cutting.

BIOLOGICAL BULLETIN, VOL. XXI.

PL*TE I.

a b

2

an

•

• «> . V».'

*

,/ ' - • * m "

'•

v

.

<•

•

10

fl

1 1

,7 /,

**

a

12
b

G JVC

C. N. C

GARY X. CALKINS.

PLATE II.

FIG. 13. a. vegetative c.-ll cut in zone 3; />. fragment three days after the oper-
ation; c, a<ymm<?trical division on the fourth day; ,1. same on^the'"seventh'day; e,
same on thr eighth day. Dead on the ninth day. Kxpriimnu 23. Table III.

FIG. 14. a, vegetative cell cut in zone 2; b, fragment 3 days after the operation;
c, monster from asymmetrical division on the fourth day; <l. -ame on the fourteenth
day. Dead on the seventeenth clay. Experiment 5, Table II.

FIG. 15. a, vegetative cell cut in zone 3; b, asymmetrical division of the frag-
ment on the following clay; c, d, normal and abnormal cells resulting from the
division; e, monster formed by the cell don the third day after the operation. Dead
on the fifth day with nuclear apparatus normal. Experiment 17, Table II.

FIG. 16. a, vegetative cell cut obliquely in zone 2; b, asymmetrical division of
the fragment 24 hours later; c, d, abnormal and normal cells resulting from tin-
division; e, abnormal cell c on the fourth day;/, g, normal cells resulting from normal
ti-Mon of d on the third day; h, monster formed from e on fifth day; i. second at-
tempted division of the monster on the the sixth day; j, same monster after fixation
and staining on the ninth day. Experiment 22, Table 1 1 .

FIG. 17. a, vegetative cell cut in zone 2; b, attempted division of the fragment
48 hours after the operation; c, same monster on the seventh day, 24 hours n'.i<\
addition of nuclein; d, same monster on the ninth day after a second (double) at-
tempted division; e, free, truncate fragment derived from the upper extremity of
the monster figured in d; f, g, two monsters -derivi-d from the division of monster d
at the middle point (x) on the tenth day; in / the lower branch is in the process of
division; h, small, reversed, truncate fragment 24 hours after being formed from
division of the lower branch of/; i,j, the two monster- on tin- eleventh day; k, the
smaller monster after fixation and staining on the fourteenth day; /, the larger sister
monster after fixation and staining on the twentieth day alter the original operation.
Experiment 40, Table II.

BIOLO3ICAL BULLETIN, VOL. XXI.

PLATE II.

13

.

15

L6

17

•

\.
b

16

:•

*• » •

••

i

•

'.

*

-

G NC

0. N. CAIKINS.

/2 GARY N. CALKINS.

PLATE III.

FIG. 18. a, dividing cell cut in zone 2 of the posterior cell; b, fragment 24
hours after the operation; c, fragment 3 days after the operation. Experiment 6,
Table II.

1 1 1.. 19. a, vegetative cell cut in zone 3; b, anterior fragment 48 hours after
the operation; c, attempted division resulting in monster formation mi the third day
aiit-r cutting. Experiment 6, Table III.

FH.. 20. a. vegetative cell cut in zone 2; b, posterior fragment 12 hours after
the operation; c. asymmetrical division of b, 24 hours after the operation; d, e, normal
and abnormal cells resulting from the division of c; f, attempted division of c 48
In mis alter the operation; g, h, results of a second (double) division of / on the
fourth day: /;. a normal Parameciitm derived from the upper branch of g; this
li\i-d for 48 hours and attempted to divide but died in the process; i, mon-trr
derived from g on the sixth day, with two practically complete paramecia and
several papilliform buds;,;', same monster on the ninth day; k. a detached free
cell on the ninth day derived from the upper cell shown in figure i\ I, same monster
on the twelfth day with constriction due to a girdle of zooglcea; m, same monster
on the thirteenth day with individuals partially withdrawn; n, a third, free indi-
vidual given off on the twelfth day, this being the attached individual on the left
side of Fig. 1; o, same monster on the fourteenth day with eight Ptirnmcdnm buds,
six mouths additional (not shown) and ten vacuoles. As the figure shows the mass
\\.i-againconstrictedand this time the constriction resulted in division but the
daughter monsters were dead on the next day. Experiment 25, Table II.

BIOLOGICAL BULLETIN, VOL. XXI.

PLATE III.

18

i

N.

d

b

19

_

.

20

d

f

.
\'"

g

•

•

•

•

- :•

m

'

GNC

C. S. CAIKISS.

Vol. XXI. July, /p//. No. 2.

BIOLOGICAL BULLETIN

I. TRUSTE1 -

AL'GL'ST. 1910
. OFFICIO

i-. 1\ 1. 11 i i, . /':/•, ctor LJnivei ;ty of Chic:.
(in MAN \ I IKI \\ nit I'ircctor, Vniver-itv <>f Maine.

I ). l!i. \KKIY HiiAK1. Treasurer, n>i 1 >r\i>nsln're Street, Boston. M
TIIM-. ||. M..- ,;v. JR.. C/c-rfc o/ //it- C> •r/'.'/-<i//1»/. University of

I ' inisylvania.

XT 1 1. ]<JI4

E. G CON KI IN .......... l'rinrrt"i] I'niver-ity.

I I \K'I N aK- I'liiver-ity.

KIMUKK. . . . j- \\ illiam Strn-t. \\ w ^"ork City.

M. M. Mi ........ ' '!., rlin Co!' 'ln-rlin. < >!:•

\\'IIII\M I'VIIKN ........ I 'artinouth ' . HaiMver. \t-\\ Hani|)-

shire.
i\ M CLAP? ..... Mount Holyokr • -nli I la'lIi-y.Mass.

I M MI; Ri i(,ii \KII ......... CniviT>ity of Micliiijan, Ann Arlior. Mich.

\\'. I1.. S<MM ............ I'riiu-i-toii L'niver^ity.

\ i n.
\\III\MM L. I'.KII heeatur Avenue, Bronx. Xe\v York

•y-

S !•'. ('I\KKK ........... William- William-town. Mass.

CHARLES I ..... [DGI ....... \me- Buililint;; Bo-t<'ii. Mass.

i . R. ( K\NI ............ 2559 Mirhi^aii I'loulevard. Chicago, 111.,

TreMilent of the Boanl.
A i i KI -n i '•. M AVKK ....... ( 'an; • uie Institution. \\"n«.hin^ton, I). C.

T. II. MMK.;A\ .......... Columbia University.

l'k\\i\ !•". SMITH ........ l'nite-1 States Department of Agriculture,

\\'a>hiii!4ton, D. C.
I . I'.. \\'n -o\ ........... Cohnnhia I'nivcrsity.

73

74 MAKIXE BIOLOGICAL LABORATORY.

TO SERVE UNTIL IC)12

M. J. GREENMAN ........ \\~istar Institute of Anatomy and Biology,

Philadelphia.
C. \Y. I IAKI.ITT .......... Syracuse University.

C. A. HEKTEK' ........... University and Bellevue Medical School,

Xew York City.
H. S. JENNINCS ......... Johns Hopkins University.

1 ii <>KGE LEFE-VRE ......... University of Missouri.

D. P. PENH ALLOW' ...... Mcdill University, Montreal, Canada.

A. P. MATHEWS ......... University of Chicago.

G. H. PAKKKR ........... Harvard University.

TO SERVE UNTIL

H. C. BUMPUS .......... University of \\'isconsin, Madison, Wis.

\\'. A. LOCY ............. Northwestern University, Evanston, 111.

JACQUES LOEH ........... Rockefeller Institute for Medical Research,

New York City.
F. P. MALL ............. Johns Hopkins University.

GEORGE T. MOORE ........ \Yashington University, St. Louis.

L. L. NUNN ............. Telluride, Colo.

JOHN C. PHILLIPS ....... 299 Berkeley Street, Boston, Mass.

C. O. WHITMAN' ........ University of Chicago.

1 Deceased.

II. ACT OF INCORPORATION

No. 3170.

' MI.\\\ KAI.I II OF M.\>SAClir-i

Be It Known. That wherea- Alpheus Hyatt, \Villiam Sanford
Stevens, \\'illiani T. Sed^uick. Kdward '",. < iardiner. Su-an Minns,
Charles Scd^wick MiiK-t. Samuel Well-, \\"illiani ( i. l-"arh)\v, Anna 1 '.
I'liillip- ami I'. II \"an Yleck h.. • 'dated themselves with the

intention iif forming a ('> ir\»> ration under the nanu- of the Marine
Biological J.ahoratory. fur the pur- -tahl^liin^ and inaintain-

inj; a lalioratnry <>r -tatinn f(,r -.»•;< ntifu- >tiidy and invoti^atioii. and
a M-lii'i'l for iii-trurtii'ii in hiolo^v and natural hi>tory, and have
>•.. in], lied \\ith the pn.\- tUtCS «'f thi> ( '.unin.in\\ ealth

in -ui-h i-a-e made and pr.>vi.'. rs fnnu the certifu-ate of

the I're-idint. I r. a-urer. and Trn-t. , •. ,.f ^ai-1 Ci>rpnratii>n. duly
a|i|ir. .vi-d I iy the Ci.nimi->ii.ner i.f ('.-rpi- ration-, and recorded in

tlll^ nil:

Now, .'/;• I. HENR\ 1'. 1' - retarj oi tin- c,,ininnn-

•!th of MaSSachuSi I v that -aid A. ll\att. \\ . S.

<n-. \\ . \\iek. 1 G ' lardiner. S. Minn^. ( '. S. Miimt, S.

Wells, W.G Farlow, \ l> Phillips, and B. H. Van VIcck, their a

eiate» and -u. and e-taMi^hed a-, and

are hereh\ made, an existing •', .rpn ration, tinder the nanir .if the
M \KI\I. I'.ioi in. n \i LABORATORYj \\ith the jmuers, ri-ht-. and privi-
legCS, and Mihjeet to the liinilatioii>. dntie-. and restriction-, which \<:
l.iu ajipertain thereto.

I/'/'//;.-\.N- my official M-natnre hereunto >nh-crilied. and t! of

tile ( 'omnionwealth of MaSSacl 'lereiinto artixed. thi> twentieth

d.t\ of March, in the \ear of • nr I .ord < IN K Tiior>.\\ii. I-.U.H i llr\-
\\n I n.ii i ^ -ElGB r.

IlKXRY I1,. I'll 10 I .
N. t-;v/(jry <'/" the Ci»)ii>i»>i\'ni/tli.
[SEAL.]

-

III. I',Y-LA\YS OF THE CORPORATION OF
Till- MARINE BIOLOGICAL LABORATORY

I. The annual meeting of the members shall be held on the second
Tuesday in August, at the Laboratory, in Woods Hole, Mass., at 12
o'clock noon, in each year, and at such meeting the members shall
choose by ballot a Treasurer and a Clerk, who shall be, c.r officio,
members of the Board of Trustees, and Trustees as hereinafter pro-
vided. At the annual meeting to be held in 1897, not more than
twenty-four Trustees shall be chosen, who shall be divided into four
classes, to serve one, two, three, and four years, respectively, and
thereafter not more than eight Trustees shall be chosen annually for
the term of four years. These officers shall hold their respective
offices until others are chosen and qualified in their stead. The Direc-
tor and Assistant Director, who shall be chosen by the Trustees, shall
also be Trustees, ex officio.

H. Special meetings of the members may be called by the Trustees,
to be held in Boston or in Woods Hole at such time and place as may
be designated.

III. The Clerk shall give notice of meetings of the members by
publication in some daily newspaper published in Boston at least
fifteen days before such meeting, and in case of a special meeting
the notice shall state the purpose for'which it is called.

IV. Twenty-five members shall constitute a quorum at any meeting.

V. The Trustees shall have the control and management of the
affairs of the Corporation; they shall present a report of its condition
at every annual meeting; they shall elect one of their number Presi-
dent and may choose such other officers and agents as they may think
best; they may fix the compensation and define the duties of all the
officers and agents; and may remove them, or any of them, excej.i
those chosen by the members, at any time; they may fill vacancies
occurring in any manner in their own number or in any of the offices.
They shall from time to time elect members to the Corporation ujn >n
such terms and conditions as they may think best.

VI. Meetings of the Trustees shall be called by the President, or
by any two Trustees, and the Secretary shall give imtice thereof by

76

BY-LAWS OF THE CORPORATION*. 77

written or printed notice sent to each Trustee by mail, postpaid.
Seven Trustees shall constitute a quorum for the transaction of busi-
ness. The Board of Trustees shall have power to choose an Execu-
tive Committee from their own number, and to delegate to such Com-
mittee such of their own powers as they may deem expedient.

VII. The President shall annually appoint two Trustees, who shall
constitute a committee on finance, to examine from time to time the
books and accounts of the Treasurer, and to audit his accounts at the
close of the year. Xo investments of the funds of the Corporation
-hall be made by the Treasurer except approved by the finance com-
mittee in writ:

VIII. The consent of every Tr dl be nece--ary b -olu-
tion of the Marine Biological Laboratory. In case • 'inii>n, the
property -ball be k'iven to the I'.o-ton Society of Natural History, or
some similar public in->titut:< h terms as may then reed
upon.

IX I hes« I'-;. I.a\s- may be altered at any mee1 the Tr

provide. 1 that the notice of such :i -hall -tate that an alteration

of the I'.y I .a\\ - \\ ill ''• '. upon.

X. Any meinb'-r in - lay v< either

in per-on or '

[V. TREASURER'S REPORT

FOR THE YKAK K.NIMNC, DK.CKM P.F.R 31. 1910

INCOM i:

Annual dues $ 606.00

Donations (Charles R. Crane) 8,500.00

Miscellaneous:

Interest on deposit $ 38.89

Rent of microscopes 14.00

I'ath house 52.40

Div. on 35 shs. Woods Hole

Yacht Club 350.00

Mosquito survey 400.00

Check not used .82 856.11

Supply department 9,300.58

Mess (net) 55T-97

Tuitions 4,150.00

$23,964.66

EXPENSES

Administration $ 900.00

Advertising 99.3 1

Biological Bulletin I net ) 512.70

Boats and launch 1.502.77

Botany 10.05

Chemicals, chemist supplies, etc 783. -3

Fish trap 88.35

Instructors' salaries 3,180.00

Insurance -'X .j .35

Interest 150.00

Labor 3,872.52

Mosquito survey 50.00

78

TREASURER'S REPORT. / 9

Periodicals and library supplies ~ 1.60

Postage 89.06

Repairs i .377.09

Sundries 1,900.20

Supply department 6.Q5S.j; Sjj. 547.48

Surplu- $ 1,417.18

III-..MIXI.II Li-i "i- SrxiiRY EXPENSES FOR THE YI.AR 1010

.\nn"unceinent- and expense mailing SilO.So

Adi can> u.oo

Care - if cemetery lot - <XD

Carting ashes 33 ' -;

f'-r ice 8.'"'

I >\-. I )iv\\ 's < ifti.-, nses j_\:

1 >r. Lillie'- «.tVh-e expenses 104.00

l;.\c!i.in-e • Hi elleek- 3 1

Mxpre— 47 -T

l-'rei-ltt and hauliui; ' >.8o

machine expense 55 . i J

I [owes1 bill %V'.(I4

I lubbard. f. t ).. SCI fio.oo

In-trunicnt-, lalmra: • ;>plie> i 7^.40

Laundry 21.36

I ,ectur< expense --

X'eu \"ik ( 'alcium I.i^ln < "Ui|>an\ 7

Notary fee- O

iVtty cadi at V. 1 I ok- 63.62

I'mniinn on Ixmd 750

Kent of microscopes 30.00

Kent nf typewriter 'K>

Spring \\ater 10.511

Stationery, ottice -upplie- 57

Steel band cart IO.OO

Sundry -upplie- 37-57

Teaming -'.2O

Telepb' MIC 31

Travelling expense- 3'

SO MARINE BIOLOGICAL LABORATORY.

Treasurer's office, clerical services 520.00

Trustees' dinner 16.00

Yeeder's expenses 24.76

Water 65.00

Wooden cases 8.00

Sr. 900.20

MARIXK BIOLOGICAL LABORATORY
INVESTMENTS

JANUARY i. 1911

Frxn

Amount c f fun<l I Kvi-mlii-r i. i^'jo .............. $4.55^.14

• I from life mi-mU-r-hiji- ................ 600.00

IIH-OIIH- !" January I. I'jil .................... I ;

•i from -al' .................... 37 _•

7.61549
! for curn tory ........ 6,000.00 ^1.01:

I\YM r\ e fuiul nn\\ i- follo\\ i:

oo Am. Trl. \- I\ 1. ( "o. 4 ........... $2,921.25

isi-finin- ' • I'''1'. Ci Si 7,>-.OO
.......... 7:(l -'.-

-h ........... ...... .........

4,615.49

The :I!H • k- aii'l !lati-ral

for loan of ............................... .}.' OO.OO $1,615

I.ll'.KAkV

Amount ' f fmi'l 1 Uvi-mU-r i. |S'^<) .............. S S'ifi.i;

IiK-oiiu- to January I, i-ji I .................... 7<>l-A7

l lain from -aK- "f -i O;M :' :> - .................... 74

of ri-ht^ ................................ ,8l

I.ilirar\ fund no\\ •! tlu- following;

3 shs. Am. Tel. i\ .............. .S ;>s^ 25

4 ; of >iooo Am. Ti-1. \- Tel. Co. 4- cost.... 779.00
I sh. Am. Snu-ltinv; \ i\tfmin<; Co. Pfd. cost 122.00

icral Kln-tric Co. cost ............ 302.50

sh ..................................... 56-56

81

82 MAKIXF. BIOLOGICAL LABORATORY.

LUCRETIA CROCKER FUND

Amount <>f fund December i, 1899 $2,500.00

Income after paying students' fees 522.41

Sale of rights 1.90 $3,024.31

Lucretia Crocker Fund now consists of the following:

18 shs. Vermont & Mass. R. R. Co. ccst $2,416.50

I sh. West End St. Ry. Co. cost 83.00

I sh. Am. Tel. & Tel. Co. cost 127.75

1/5 of $1000 Am. Tel. & Tel. Co. 45 cost 194-75

I sh. General Electric Co. cost I5I-25

Cash 51.06 $3,024.31

V. THE DIRECTOR'S REPORT

THE Ti<r.-u-i- OF nil. MAKINK P.io[.or,i< \i. I. \r.oi< ATORV;

Gentlemen: I have the honor t.> -uhmit herewith a report of
the twenty-third se--i»M of the Marine I'.i.'l, -gical Laboratory for
the \ear I'/ID. ami ?<une recommendations fur future develop-
ment.

\Ve record with - -rn >\v the loss hy death during the year of

three member- ••!" the board of trustees: D \. I lerter. Pro-

• r I). P. l'enhall"\v and P: 0. Whitman. It is

fitting that due n of tile SCfl -lumld be

en in the form of resolution- to l>e -pread upon the minutes

of the lizard and -ent t" the fainili. -ir lamented colleague-.

The death -f Pr^f \Vliittnan wa- Midden and unexpected.

lie \\a- taken with pneuiii"iiia • >u \.\einher .^ • and died . >n

December ''. Mrinorial - in hi- h"ii«>r \\ere held at the

LJniversit) of ( hicago -n Decembei •s;. and hi- h»d\ \\a- laid in

the lal>"rat<>ry l"t in the Mpi-c- -pal ' ry at W'«>d- ||"!«- OH

December ID in the pt . a -mall jjatheriiiL,' "f his former

atCS and friend-. wh«> almie \\ere able t" attend.

Professor Whitman'- death may be -aid t-» mark the cl<»sc of
the tir-t period of development of the Marine I '. il Labora-

tory. A- director for the tir-t twenty •- of the exi-tence

of the Laboratory, more than an} other per-on he i;ave the Labora-
tory it- -tatn- and it- ideal-. In a very fitting way. therefore,
\\e max -ax that the Laboratory i- a monument to hi- memory.
lUit it i- de-irable an«l due that -onie permanent memorial should
mark hi- la-t re-tin.^ ].lace near the iii-titutioii that he loved so
well, and it i- recommended that a committee be appointed to
take thi- matter in charge.

The -tatY of instruction in [910 comprised 29 member-, repre-
-eiitini,T eighteen in-tituti"ii<. Acknowledgment -hould again be
made to the member- of the staff of investigation who rendered
their .-ervice- freely.

84 M. \K1.\K mol.oC.ICAL I.AI'.oR \TOKY.

The attendance of workers at the laboratory comprised 62
investigators and <>4 -Indents — a total of 126 as compared with
66 investigators and 63 students, total, 129, in 1909, and 52
investigators, 48 student?, total 100, in 1908. There was a grati-
fying increase in the number of subscribing institutions, from 19
in 1909 to 24 in 1910. One of the subscribing institutions of
1909 dropped out this year. The new subscribing institutions
were Dartmouth College, Massachusetts Agricultural College,
The Rockefeller Institute for Medical Research, The University
of Illinois and the Washington University Alumni Association.
The membership of the corporation was also somewhat in-
creased. The receipts of the supply department increased from
$8.54';. 55 i» |('°9> to $9.300.58, in 1910.

Xo definite progress was made in the plan for a permanent
building during the year, except that steps have been taken to
secure the possession of more land that is necessary for the
protection of our present holdings. Success has been attained
in this direction and I think we may say that we are distinctly
nearer the realization of the permanent building, not only in
point of time but in preparedness. This raises definitely two
questions: first, as concerns the nature and use of the building,
and second, concerning administration affairs of the laboratory.

As regards the building, I believe that the committee that was
appointed December 30, 1909, was fully agreed that the new
building should provide for the library and for a certain number
of research rooms. This committee has not formally met and I
do not present this as an official report from the committee.
But on the assumption that provision would be made for re-
search rooms on a more adequate scale and with better equip-
ment than the present quarters, I think that we should begin to
ascertain the attitude of certain universities and research institu-
tions with reference to the maintenance of quarters for investi-
gation to be available throughout the year. Such an inquiry
may ^eem premature; but when we consider that the character
of the building to be erected must depend to a certain extent
upon the financial -upport of institutions for its maintenance. I
think it will be admitted that we can not move too soon in [In-
direction of receiving assurances.

THE DIRECTOR'S REPORT. 85

The second question concerns administration. A- matters
now stand the burden of administering the affairs of the labora-
tory falls rather heavily on men who already have a full assign-
ment of work in their own universities. There i- n» question but
that the work of the laboratory could be made more effective in
all respects if we had an administrative officer who dev»ted
hi- entire time to the affair- . .f the laboratory: it is also question-
able whether the laboratory can continuously retain the servi
of men under present condition.-, when their work is i^iven at the
expense of cffectiveiu -- in their university work and their re-
search. Jt i- therefore recommended, that, a- 5OOD a- may l>e.
the office of assistant direct "i- IK made a resident po-iti-n for
the entire \car. l"nder -uch an arrangement all our relation-
should prosper better: with the inve-tii,ra:»r-. with the student-,
and with in-titutioii-. The e\peii-c involved would probably be
a dimini-liini; amount «• \\in.i,' to improvement in laboratory condi-
tion- generally.

As SOOn as plai tely under way for a permanent

buildin ;i an officer \\onld be !!• I • the

detail- decided on by t' • •!) the building, and to make

ime-ti^ati'.n- . .n behalf of the committee; and later when the
laboratory \\a- ere at other seasons

of the \ear ' • immer. lie -hould be furnished with

-tanre ade-|i ire him -nffu'ient time for research.

Th< • appended a- part- of th'~ report (l) a h-t of the

-tatV. i j ' h-t- of inve-- .;id -tir I I n attendance. ' .^ ' a

comparative tabular view of attendance, 14' the name- of -ub-
scribini,' in-titution-. 151 the evening lecture-. (6) a li-t of mem-
ber- of the

i. THE STAFF, 1910

F. R. ULLIE, DIRECTOR,
Professor of Embryology, UniverHty of Chicago.

GILMAX A. DREW, ASSISTANT DIRECTOR,
Professor of Biology, University of Maine.

ZOOLOGY

I. I \VESTIGATION

Zoology and Embryology

E. G. CONKLIN Professor of Zoology, Princeton University

(Absent in 1910.)

GILMAN A. DREW Professor of Biology, University of Maine.

GEORGE LEFEVRE Professor of Zoology, University of Mis-
souri. (Absent in 1910.)

FRANK R. LILLIE Professor of Embryology, University of

Chicago.

THOS. H. MONTGOMERY, jR.Professor of Zoology, University of Penn-
sylvania.

T. H. MORGAN Professor of Experimental Zoology, Colum-
bia University.

E. Pi. \Yii.sox Professor of Zoology, Columbia University.

II. INSTRUCTION

\YINTERTON C. CURTIS. .. Professor of Zoology, University of Mi--

souri.

PAUL M. RKA Professor of Biology, College of Charleston,

and Director of the Charleston Museum.

KnwAiui !•".. WlLDMAN .... Central High School, Philadelphia.

JOHN \\". SCOTT \\YMport High School, K;m>as City.

G. S. DODUS Fellow in Zoology, University of Pennsyl-
vania.

TI. J. Si'Kxci.u \s-i-tant in Zoology. Columbia University.

86

THE DIRECTOR'S REPORT. 87

EMBRYOLOGY

I. INVESTIGATION. (See Zoology)
II. INSTRUCTION

OILMAN A. DREW Professor of Biology, University of Maine.

LoRAMiF. L. \V FF. .. Assistant Professor of Biology, Vale Uni-

versity.

\VII.I.IAM K. KEI.LICOTT. . I'r< aVssor of Biology, Goucher College.

Kor.KkT A. I!riii\.;-mx. .. Associate Professor of Zoology, Oberlin

College.

PHYSIOLOGY
i.

AM-.IKI I'. M A i ii KU - . . . . Prnfi ~-..r nf Physiological I'lK-miMry. Uni-

I ( "hicago.

1 I' LVOM PrniY--nr nf I'hy-iiili'gy. I'nivi-rsity of St.

lis.

K. S. 1. in ii In-iriH-tnr in * 'mnparativi- l'h\ >iology, Uni-

of iVnti-vlvaniu.

•

ii. i ••

11. II \IUM\\ !'• ir of Zi I'liiviT-ity nf Ti-xa-.

NK I' KNOWLTON. ... Professor nf I'liy-ii-In^y. Syracu^i- I'ni-

:ty.
!•'. II I'IKK Iiistnu-tiir in I'lr. . l"n:\ iT-ity of

PHILOSOPHICAL ASPECTS OF BIOLOGY AND
ALLIED SCIENCES

I- nwAKii < I. SPAULDING... Assistant I'- r nf Philosophy, Prince-

tmi I'liivcrsity.

BOTANY

•KI.K 'I". MOORE ProlY — 'T nf Plant Ph\ .-iolngy ami Applied

Botany. \\'ashington University.

GEORGE K. I.YMVN \^:-tant Professor of Botany, Dartmouth

College.

MARIXE BIOLOGICAL LA in >!•• ATORV.

B. M. DUGGAR Professor of Plant Physiology. Cornell Uni-
versity.

I. F. LEWIS Professor of Biology, Randolph-Macon

College.

LEWIS KNUDSOX Instructor in Plant Physiology. Cornell Uni-
versity.

LIBRARY

H. McE. KXOWER University of Cincinnati, Librarian.

CHEMICAL SUPPLIES

OLIVER S. STRONG College of Physicians and Surgeons,

Chemist.

G. M. GRAY Curator of Supply Department.

THOMAS M. DOUTHART. . Collector in Zoology.

ARTIIUK \V. TAYLOR Collector in Botany. Salem High School,

Salem, Mass.
JOHN VEEDER Cockswain.

2. INVESTIGATOR-

1910
OCCUPYING ROOMS

i. ZOOLOGY

iN, \V. H I .. Uni - nia.

KWiiii. (OKA .1., Iiistni(.-t«T ii B i,-e.

rk.

ge.
, I . k.. I'nr.i < ton . .1.

ty.
iri.

!.cal School,
St. i 01

i

> . Vale

HAK\ BY, E. NEWTON, < ^ City.

M \s M of Texas.

Ki . ucher (

' ' ivcrsitv mnati.

.i-rsity ol ^o.

: . r.-ity of
l'<
Mi I ylvania.

. 1 1. .|K. ;. Ivania.

1 1 . . I ' i • . • .

IN, J. 1 MUM A.S Instria-i

l\i \

i . Juii N \\ .. \\ City, 1

Hi NKV .1.. Assistant •• I 'nivcr-

KI>\\AI.' I'liiloMijihy. Princeton

University.

Instriicti>r in A 'liege of 1'i - and Surgeons,

.\\-\\ \ .irk '

\\MKI lACl I'... Coluni! .cr^ity.

\\;:;itiK. KKMM I.. Assistant in AiKitmny, Johns Hopkins University.
WMIS. 11. i,.. Ass "t l'ath"l".i;y, 1'ni vi-r-ity of Chicago.

\VIIHM\\ l-".n\\ \K-i« i:il Hiuh School, Philadelphia, Pa.

89

QO MARIXK BIOLOGICAL LABORATORY.

WILSON, E. B., Professor of Zoology, Columbia University.
WOODRUFF, I.. I... Assistant Professor of Biology, Yale University.

2. PHYSIOLOGY

DONALDSON*, H. H., Professor of Neurology. \Yistar Institute.

HIRSCHFELDER, ARTHUR D., Associate in Medicine. Johns Hopkins Medical

School.
KNOWLTON, FRANK P., Professor of Physiology, Syracuse University, College

of Medicine.
KIM;. W. O. REDMAN, Rockefeller Institute for Medical Research. Xevv York

City.

LOEB, JACQUES, Rockefeller Institute for Medical Research. New York City.
LILLIE, R. S., Instructor in Physiological Zoology. University of Pennsylvania.
MATHEWS, A. P., Professor of Physiological Chemistry, University of Chicago.
XEWMAN, H. H., Professor of Zoology, University of Texas.
PIKE, FRANK H., Instructor in Physiology, University of Chicago.
ROGERS. CHAKLES G.. Associate Professor of Physiology, Syracuse University. .
\Y.\STENEYS, HAKDOLPH, Rockefeller Institute for Medical Research, New York

City.

3. BOTANY

BESSEY, ERNEST A., Professor of Botany, Michigan Agricultural College.

DUGGAR, BENJAMIN M., Professor of Plant Physiology, Cornell University.

K \UDSON, LEWIS, Instructor in Plant Physiology, Cornell University.

LEWIS, IVEY FOREMAN, Professor of Biology, Randolph-Macon College.

LYMAN. GEORGE R., Assistant Professor of Botany, Dartmouth College.

MOORE, GEORGE T., Professor of Plant Physiology and Applied Botany, Wash-
ington University, St. Louis, Mo.

THOMAS, MASON B., Professor of Botany. Wahash College.

OSTERHOUT, W. J. V., Assistant Professor of Botany, Harvard University.

SMITH, ERWIN F., Pathologist in charge Laboratory of Plant Pathology, U. S.
Department of Agriculture, Washington, D. C.

OCCUPYING TABLES

i. ZOOLOGY

ALLYN, HARRIET, University of Chicago.

DAVIS, SARAH ELLEN, Student, Columbia University.

LINTON, ELEANOR A., Washington, Pa.

KELLEY, FRANK, J., University of Pennsylvania.

WILSON, HERRICK E., Assistant in Geology, Oberlin Colleg-.

SPOONER, GEORGIANA B., Palo Alto, Cal.

2. PHYSIOLOGY

DONALDSON, JOHN CALVERT, 3310 Race Street. Philadelphia, i- --
TASHIRO, SIIIRO, Fellow, University of Chicago.

3. BOTANY
TAYLOR. ARTHUR W., Warren, Mass.

STUDENTS

igio

INVERTEBRATE ZOOLOGY

BAKKK, RITA < i.. 72 liimtiiu nue. Boston, Mass.

P.M.!.. MAKY M.. < >bcrlin Collegi n, Ohio.

l!nii\v i i.i., H . i ;,," Main SIP

CASIIM '.' Syrncu- York.

•;h College. Ham.vir. N". H.

KI>IM>\\ ' tin ore, Md.

A. I!.. ersity.

•
I-'IM ; ' ;. V

« w York i ity.
i . . Syracu-i-. N". V.

Orono,
M. I '.^riculti;

M \ K 1 1 % . I : M.- 1 1 •

•'••.'•'
•

MOOK, CHAI ty, New

M>" ' ,n Aven • City.

M \-, i -t. Xew N "fk (ity.

M \M \s S., M

1'Ai- ' t;c.

I' AM i N . II v • Mil.

I'l \ I, I'lula.'.i ']iln:..

S i \ : •• GEBTBI

V \\'.. ! ' ' • "

p, EMILY, 4

\\ M KI s - \\ illi;ini-|M,rt,

\\ 11 l i \\'

\Vi.s\\ i ii . \sn I'.. I a-i M

\\i<i..in ( Oil \ i'. Slu-ridan. \iu \ ^rk.

,-SKII Hi i i s M.. Vassal « bkeepsie, X. V.

EMBRYOLOGY

Aiuiorr. M\> • l"r..nl Strict, Plainfield. X. .i.

\ "\. JuM rn I... St. ii< ' liege, I'.t.

ENNIS \ •- - A.. I'.arnaril i 'i.]li-.i;c. Columbia Univcrsit>.

. I-'.KMM ]•'.. H..\v.ini I'nivi-rsity, Washington, D. C.
Krsiv. Aimnr. State University of Iowa.

•'

MARINK IlIiiU M.ICAL I. Al'.i iKATuKY.

LEAVITT, GEORGE C., University of Maine. Orono. Maine.
MELLEN, IDA M., 530 Carleton Avenue, Brooklyn, Xe\v York.
PRENTISS, HENRIETTA, Normal College, New York City.
REDELINGS, LESLIE R., Northwestern University, Evanston, 111.
SMITH, ISABEL S., Illinois College, Jacksonville, 111.
ULLMAN, HI.NKV J.. Rush Medical College, Chicago, 111.

PHYSIOLOGY

DAVIS, NATHAN SMITH III. 8 E. Huron Street, Chicago, Illinois.
MARKS, HENRY S., University of Rochester, Rochester, N. \ .
STARK, MARY B., St. Olaf College.

STEPHENSON, J. C., University of Chicago, Chicago, 111.
WHITENTON, ROBERT O., University of Chicago, Chicago, 111.

BOTANY
i. Plant Structures and Responses

CHANDLER, JEAN FORREST, 4651 North Lincoln Street, Chicago, 111.

CLEMENT, FRANK H. P., Tilton, N. H.

WELLS, COLLIN, Dartmouth College, Hanover, N. H.

JENNISON, HARRY M., Massachusetts Agricultural College.

2. Morphology and Taxonomy of the Algae

BRVGGER, HELEN F., Mount Holyoke College, South Hadley, Mass.
COLLEY, REGINALD H., Dartmouth College, Hanover, N. H.
CURTIS, OTIS F., Oberlin College, Oberlin, Ohio.
DAWSON, AVA B., 97 Mountfort Street, Boston, Mass.
HILL, ALBERT F., Dartmouth College, Hanover, N. H.
HOIIBS, ETHELYN, Wellesley College, Wellesley, Mass.
IRWIN, JAMES M., Dartmouth College, Hanover, N. II.
LEIGHTON, HELEN, 68 Park Avenue, Canandaigua, New York.
ROBERTS, EDITH A., Mount Holyoke College, South Hadley, Mass.
RUTHERFORD, ELIZABETH E., 624 Fullerton Parkway, Chicago, 111.
SPARGO, MILDRED W., 5050 Maple Avenue, St. Louis, Mo.
WALLBURG, AMY B., 54 Dale Street, Roxbury, Mass.
WILLISTON, Ri'Tii. 297 Crown Street, New Haven, Conn.

. TABULAR VIEW OF ATTKNOANVK

1907 1908 1909

INVKMH..VIUK — Total ..................... 60 :_• 66 Q2

Zoology ...... ..... 3_> 32 3.) 33

I'll .......................... 8 6 7 9

I'.' t.-iiiy ............................. 10 8 4 9

Occupying 1

/"••I- . ..... •» 6 14 5

1'liyMnlnjjy .......................... I 2

I'.iit.-iny .............................. I

STUDENTS T-t.-il ......................... 47 4S 63 64

ZoolO) 34 i-> 3'. 31

gy .......................... 15 i _• 10

........................... 4 3 .) 5

<> I I '• 17

QNIVERSITIES \M •' IVM-KESENTKH. . . . 47 51 47 4-)

l'.\ [nvestigators ..................... -•'• 27 26

I '.v Stmlrllts .......................... j | 26 2O 24

93

4. SUBSCRIBING INSTITUTIONS, 1910

ACADEMY OF NATURAL SCIENCES, PHILADELPHIA.

COLUMBIA UNIVERSITY.

DARTMOUTH COLLEGE.

GOUCHER COLLEGE.

LUCRETIA CROCKER SCHOLARSHIP, BOSTON PUBLIC SCHOOLS.

MASSACHUSETTS AGRICULTURAL COLLEGE.

MOUNT HOLYOKE COLLEGE.

NORTHWESTERN UNIVERSITY.

OBERLIN COLLEGE.

PRINCETON UNIVERSITY.

ROCKEFELLER INSTITUTE FOR MEDICAL RESEARCH.

SMITH COLLEGE.

SYRACUSE UNIVERSITY.

UNIVERSITY OF CHICAGO.

UNIVERSITY OF ILLINOIS.

UNIVERSITY OF KANSAS WOMAN'S TABLE SUPPORTED BY MRS. ROMIN

SON.

UNIVERSITY OF MICHIGAN.
UNIVERSITY OF PENNSYLVANIA.
UNIVERSITY OF ROCHESTER.
VASSAR COLLEGE.

WASHINGTON UNIVERSITY ALUMNI ASSOCIATION.
\YISTAR INSTITUTE OF ANATOMY AND BIOLOGY.
WELLESLEY COLLEGE.
YALE UNIVERSITY.

94

5- EVENING LECTt'KKS. 1910

I. <>KH " I low I >oe- the Spermatozoon ("ause

the Development of the 1 u-; "'"•-• July i.

C. P.OWYER VAUX " Uo\v to Prepare a Paper for Pub-
lication " July 5.

JOHN- I',. SMITH " M-p>i|uitot-> ami Mo-quito C'am-

pai^n- " July 8

I I II. DO\\> ..."The iVrcet:' :' Water in the

( "entral N : Tin of \\-rte-
l>ratc- " July u.

\\". J. V. OSTEBHOUI Cts of the Action of Min-

upon Plants " July I ;.

I P.. WILSON "Hen • and the < hromo-omes ". . July \>>

P.. M. DUGGAB " 'I he Ivelation of Plant- to Toxic

i.il Refer, nc. •

the Mo.litu-atii'ii of Toxicity Ky
( "liciircal aii'l I'lu-ieal Assent- ".. Ju'
1\ NP N'KKKI - " The l\elation of psychology to '.

July j,.

J. T. P.\ I I i-:i.--o\ "Th' of Specific Polyein-

hryony and the I >etenninatioii of

in the Armadillo" \"'^r 4

I\ \IIMI P.. PI-KKY ••\\heii is a '1 liini,' I:.xplained for the

I'urpo-e of Science" \i:.

l\ \YMOMI Pi \ui . ..."The lnheritai!> • "f |-\-cundity in the

Domestic l"o\vl " \u^. (>.

95

6. MEMBERS OF THE CORPORATION OF THE
MAR I. XI-: I'.IOLOGICAL LABORATORY

T. LIFE MKMKERS

ALLIS, MR. EDWARD PIIELPS, JR., Palais Carnoles, Menton,
France.

ANDREWS, MRS. GWENDOLEN FOULKE, 821 St. Paul St., Balti-
more, Md.

BILLINGS, MR. R. C., 66 Franklin Street, Boston, Mass.

CAREY, MR. ARTIITR ASTOR, Fayerweather Street, Boston, Mass.

CLARKE, PROF. S. F., \\rilliams College, Williamstown, Mass.

CONKI.IN, DR. E. G., Princeton University, Princeton, New

•* '

Jersey.

CRANE, MR. C. R., 2559 Michigan Boulevard, Chicago, 111.

DAVIS, MAJOR HENRY M., Syracuse, New York.

ENDICOTT, WILLIAM, JR., 31 Beacon St., Boston, Mass.

EVANS, MRS. GLENDOWER, 12 Otis Place, Boston, Mass.

FARLOW, PROF. W. G., Harvard University, Cambridge, Mass.

FAY, Miss S. B., 88 Mt. Yernon Street, Boston, Mass.

FOLSOM, Miss AMY, 88 Marlborough St., Boston, Mass.

FOOT, Miss KATHARINE, 80 Madison Avenue, New York City.

GARDINER, Miss EUGENIA, 15 West Cedar Street, Boston, Mass.

HAMMOND, MR. G. W., Hotel Hamilton, Boston, Mass.

HANNAMAN, MR. CHARLES E., 103 First Street, Troy, Nr\v
York.

HARRISON, PROVOST C. C., University of Pennsylvania, Phila-
delphia, Pa.

JACKSON, Miss M. C., 88 Marlborough Street, Boston, Mass.

JACKSON, MR. CHARLES C., 24 Congress Street, Boston, Mass. .

KENNEDY, MR. GEORGE G., 284 Warren Street, Roxlmrv, Mass.

KIDDER, MR. C. G., 27 William Street, New York City.

KIDDER, MR. NAT 1 1 \MI.I. T., Milton, Ma--.

KING, MR. CHARLES A.

96

THE DIRECTOR'S REPORT. 97

LEE, MR-. FREDERK S., 2~<> Madison Avenue, Xe\v York City.
LoWELLj MR. A. LAWRENCE, 171 Marlborough Street, Bos;
Mass.

MASON. Miss K. F., i \Yalnut Street, Boston, Ma--.
MASON. Miss IDA M.. i \Valnut Street. Boston. Ma--.
Mi:.\\~. MR. JAMKS HOWARD. \<><< l',eacon Street. Boston, M.
MLKKIMAN. MRS. DANIKI.. \V. >rci -ter. Ma — .
MINN-. M ^ ., 14 I. ;r^ Sijuare, Boston. Ma--.

MINNS. MR. THOMAS. 14 I.<>uisburg Square, Boston, Ma--.
MINIM. I )u. I'HAKI.KS S.. Harvard Medical School. Bost-m. M.
Mi. vi i R. Mi-- M. ( .. 241 Mar!b(ir<'U^h Street. r.i>-t«n. Ma--.
Mok«;AN. MR. I. I'liRi'-'Ni. JR., \\'all and, P.p-ad Street-. New
Vork

MORI, AN, I'lo-i. T. II., ( '"himhia I'niversity. Xe\v ^"ork C'ity.

M OKI .AN. MR-. T. II.. \\-\v Y'>rk ("ity.

\o; LACK •iiMMiiwcalth Avenue. r.o-t"ii. M

\OM -. M : nth \Yill.uv Street. Montclair. X. I.

N"i \ N. MR. LUCIAN I... Telhirid.

< »-I:ORN. l'R"i. lli\R\ I".. American Museum of Natural Hi-

tory. New \»r\< ( 'ity.

Pi i i. MR. \i i RI 1 1. Highland l;al!-. < >rani,'e l". unity. X. J.
I'lllllli'-. I >K. |on\ ' . .: M I'erkle; :. r,<>-t<.n. Ma--.

PHILLIPS, MRS. JOHN C, 299 I'.erk'.ey Street. I'.. .-t'-n. Ma-

I'MRIIR. I )R. II. ('.. I "i ;" l'enn-\ Kania, Philadelphia, I'.i.

1'ri-iriR. MR. \\ . II . Newton ('enter. Ma--.

KIM, I KV-. M [SS A. I'.. 5 I' 13 ' 'II. Ma--.

KIH.IR-. MR-. \\'II.LI\M I1... 117 Marllior. ni^li Street. Boston,

Mass.

Si \R-. I'R. Hi \R\ !•".. 4_'D I'.eac- -n Street. r..-)«tnii. Mass.

Sin hi'. M R. I'. A.

SMITH. MR-, i '. i '.. 286 Marlborough Street. Bi'-tmi. Ma--.

STROBELLj MlSS £. C, So Mad.i-.'ii Avenue. Xe\v \'ork City.

TiioKNi'iKi. I>R. l;.n\\ \RH L.. Teacher-' i"i 'liege, Columbia I'ni-

ver->ity. Xew \«rk >
TRI i r \.-K. I'ROI. \\'II.I.!\M, Mi--ouri Botanical darden-, St.

Louis, Mo.

\\ \RI. Miss M \KV I... 41 Brimmer Street, Boston. Mass.
\Y\KKi N. MRS. S. D.. o- Mt. X'ernon Street, Boston, Mass.

98 MARINE BIOLOGICAL LABORATORY.

WHITNEY. MR. HENRY M., Brookline. Mass.
WILLCOX, Miss MARY A., Wellesley College, Wellesley, Mass.
WILMATH, MRS. H. D., Elliott Street, Jamaica Plain. Mass.
WILLIAMS. MRS. ANNA P., 505 Beacon Street, Boston, Mass.
\\ ILSOX, DR. E. B., Columbia University, New York City.
WILSON PROF. W. P., Philadelphia Museum, Philadelphia, Pa.

II. MEMBERS, AIV.TST 9, 1910

ABBOI T. MARGARET B., 413 W. Front St., Plainfield, X. J.
ADAMS, C. F., University of Arkansas, Fayetteville, Arkansas.
ADDISON. W. H. F., University of Pennsylvania, Philadelphia,

Pa.

ALSBURG, CARL S., U. S. Dept. Agriculture, Washington, D. C.
BAKER, E. II., 5444 Catherine St., Philadelphia, Pa.
I'. \RDEEN, C. R., University of Wisconsin, Madison, Wis.
BECKWITH, CORA J., Yassar College, Poughkeepsie, New York.
BIGELOW, MAURICE A., Teachers College, Xew York City.
BIGELOW, ROBERT P., Massachusetts Institute of Technology,

Boston, Mass.

BLATCHFORD, E. W., mi LaSalle Ave., Chicago, 111.
BROOKOVER, CHARLES, Buchtel College, Akron, Ohio.
P.K-owxE. ETIIKI. X., Bennett School, Millbrook, Xew York.
I '.i < KIXGHAM, KIMTII X.. 342 Marlborough St., Boston, Mass.
P.rmxcToN, R()i:i-:kT A., ( )l)(.-rlin College, Oberlin, Ohio.
BUMPUS, H. C., University of Wisconsin, Madison, Wis.
I'.YRXKS, ESTHER F., 193 Jefferson Ave., Brooklyn, Xew York.
CALKIXS, ( IAKY X., Columbia L'niversity, New York City.
('\IAT.RT, riiii.ii' P., University of Pennsylvania, Pliikuk-lphia,

Pa.

CARLSON, A. J., l*nivi-r-ity of Chicago, Chicago, 111.
CARY, L. K., Princeton University. Princeton, X. J.
CAT-II LI., |. Mi KKKX, Garrison-on-Hudson, Xew York.
("in ~n K, WEBSTER, Colby College, \\'aterville. Mainr.
CHIDESTER, FLOYD E.. Clark University. \\"orcester, Mass.
CHILD, C. M., University of Chicago, Chicago, 111.
CLAPP, CORNELIA M., Mount I lolyoke College, South Iladley,

Mass.

THE DIRECTOR'S REPORT. 99

CLARK, ELIOT R.. Johns Hopkins University. Baltimore. Md.
COF., \V. R.. Vale University. Xe\v Haven. Conn.
CoL'i'i.v. II. S.. 3409 Powellton Ave.. Philadelphia. Pa.

o< K. J. If.. Cornell University. Ithaca. Xew York.
COOLIDG) . CHAKI.KS A.. Ann.- P.nilding, Boston. Ma — .

LTIS, \\'. C, Univer-ity of Mi-suuri. Columbia. MO.
I)IMO\, Ai:n;Air. C.. y •- < ien< see St., Utica. X. V.
iMii--. ( i. S.. St. L..iii- University Medical Sch«Mil. St. L"iiU. M >.
DONALDSON, II. H., \Yi-tar Institute of Anatomy and l'.iolo-y.

Philadelphia.
DORKAXU.. ANN. I )orraiuv;..n. i

\< \x< i . I-'K \x< i 5, I >orrancet"ii. Pa.

I )KI \\ . i III.M \.\ A.. I "ni\ <.-r~:ty of Maiiu-. < >r> 'ii' >. Maine.

I-*A PON, I 1 I.. I Ioli;irt ' . ' irlK'Va. X. Y.

I-'.II.I.NM \- KI. II.. I : t) of Indiana, P.l.» .niin^ton. Ind.

l-'iii.n. IUVINC, \.. \\Y-u-rn Maryland College. \\\-<tmin>-ter. Md.
I-'IK-'.I SON, I. S.. ( oriu-11 Timer-ity Medical School, Xe\v \»rk

' ity.

l-'iv-i i M \N. M i-- 1 I \ivi' ^- I 'nion Park St.. Boston, Ma — .

S. II.. » oniell Univer-ity. Ithara, Xe\v York.
(In-. \Viiii\M I . .r. iVpt. I'.iolo^ical ('hemi>try. C«>lum-

hia I 'ni\ er-ity, X. \'.

G LASER j O. C., l"n:vn-it\ of Mii-hi^an. Ann Arhor. Mich.
GOLDFARB, A. I . ' •'•imliia rniver-ity. Xew \«rk I "ity.
(iuu \.M\.\. M. I., \\i-i.Li- In-tituu of Anatomy and I'.io]

Philadelphia.

<iivi..«'Kv. I.'TI-I II.. Barnard i . Xew \»\-k ("ity.

HALL, IV-IUKI \\'.. i;j South Linden St.. P.ethlehem. Pa.

II AKi.ii i. C. \\ .. S\racu-e I'niver-ity. Syracn-e. Xew \«rk.

I I tRRISON, \ C., Wood I lole. Ma — .

II \KKI-ON . K . Yale Univer-ity. Xew Haven. I '..mi.
II \K\IV. 1'.. ( '. II.. University of Chicago. Chica-o. 111.

II \K\IY. I-'.. X.. hepartment of /o.-l.^y. Columhia Univer-ity.
HAYES, S. P.. Mount llolyoke College, South lladley. Mass.
11; MM. ir.\k(ii.i'. Stanford University. California.
II" \K. D. I'.i. \Kn.v. i"i hevoii-hire St., Boston, Ma — .
HOLMES, S. I.. i;v^ East < iorham St.. Madi-.n. \\"i-con-in.
Ni i i Y, ]•'. I'... < 'klahoina Academy of Science. Toiikawa, ( >kla.

TOO MARINE i:i(U.tM,H-.\I. I.AP.MR.vn >RY.

JACKSON, C. M., University of Missouri, Columbia, Mo.

JAYXE, HORACE, Wistar Institute of Anatomy and Biology, Phil-
adelphia.

JENNINGS, H. S., Johns Hopkins I'nivcrsity, Baltimore, Mel.

JONES. LYNDS, Oberlin College, Oberlin, Ohio.

JORDAN, H. E., University of Virginia, Charlottesville, Va.

\\\ LLEY, 1;. J.. Department of Zoology, University of Pennsyl-
vania.

Ki LLICOTT, W. E., Goucher College, Baltimore, Md.

KENNEDY, HARRIS, Readville, Mass.

KING, HELEN D., Wistar Institute of Anatomy and Biology,
1 'hiladelphia.

KiNGsr.rRY, B. F., Cornell University Medical School, New York
City.

KINC.SI.KY. J. S., Tufts College, Mass.

KIRKHAM. \V. B., Yale University, New Haven, Conn.

KMAVI.K, H. McE., University of Cincinnati, Cincinnati, Ohio.

KNOWLTON, F. P., Syracuse University, Syracuse, New York.

KRAEMER. HENRY, 424 South 44th St., Philadelphia, Pa.

KRII-.S, HERI'.ERT, University of Pennsylvania, Philadelphia, Pa.

LEWIS. I. F., Randolph Macon College, Ashland, Va.

LEE, F. S., 437 West 59th St., New York City.

LEFKYKK, GEORGE, University of Missouri, Columbia, Mo.

LEWIS, WARREN H., Johns Hopkins University, Baltimore, Md.

LIBBEY. \YII.LIAM, Princeton University, Princeton, N. J.

LILLIE, FRANK R., University of Chicago, Chicago, 111.

LINTON, EDWIN*. Washington and Jefferson College, Washington,
Pennsylvania.

LOEB, JACQUES, Rockefeller Institute for Medical Research, New
York City.

LOEB, LEO, St. Louis University Medical School, St. Louis, Mo.

LUSCOMBE, WALTER O., Woods Hole. Mass.

LYMAN, GEORGE R., Dartmouth College, Hanover, X. II.

LYON, E. P., St. Louis University Medical School. St. Louis, Mo.

MCCLENDON, J. F., Cornell University Medical School, New York
City.

McGiLL, CAROLINE. University of Missouri. Columbia, Mo.

MCGREGOR, J. H., Columbia University, New York City.

THE DIRECTORS REPORT. IO1

M< Ixnoo, X. E., Department of Zoology. University . .f lVnn>yl-

vania.

M.u Kh.vziE, MARY D.. \Ye>tern College, Oxford. < »hio.
Mcl\n:i:i.v, r.\n. S.. University of Chicago. Chica-<>. 111.
M< Mi-RRicir. J. P.. University of Toronto. Torom •. Panada.
MALL, F. P., John-. Hopkins University. Baltimore. Md.
M A- i. S. ' '.. Johns Hopkins University. Baltimore. Mil.
M.viniA\s, Ai.m.kT 1'.. University of Chicago. Chica^". 111.
M \YI-.K. A. <i.. Map!e\V" - !. X. J.

Mi K,S, I-".. p.. Harvard Medical Scho«il. I '."-1.111. Ma
Mi 1. 1 /IK. S. P. i.} West uist St.. Xew \»rk I'ity.
Mi H \i r. M. M., Oberlil ' ' 'lierlin. < 'hi...

Mi.\"K. M \kn L., Wadlei( S '. i 14111 St. and 7th Ave..

Xe\v ^"'.rk ( 'ity.
MOENKHAUSj \\'. I.. LJniversit) of Indiana. I '.1- •• •min-:"ii. hid.

MONTGOMERY, T. II.. |i>.. LJi f IVnn-\l\-ania.

MOORE, G. T., Missouri r.-ia: Card as, St. Louis, Mo.

M IE, J. PERCY, l"m\ i" l'enns\lvania.

11. A. . '.tural l;..\periment n, Kn"\villc.

I rim.
I i.. A. D.. llami!' e, l'lint<m. X. N .

I I . I'll \UI ! s \ .. of Medii'ille.

\e\\ \"<>rk.

M iki: \i II. I .• \v. line. I >rtn>it. Mich.

XATII i Nil r.. II. !•'.. I 'i : Mini: Minneapolis. Minn.

Xl • \l . I I. \ .. Ki: C <

XIAVMAN. 11. II., Uni\er-i:\ of I e\a~. Austin. I
Xh HOLSj Miss M. I Dimmer St.. Philadelphia. Pa.

EVEE, I S.,] • 'n. 111.
( 'KIM \\\. \. I' . i an . Pittslmr^, Pa.

.1 KX. l\\VM"\i- ( . P.arnard ( 'olle^e. Xew V.>rk City.
I'M K \un. I 'n \NI.I S, \\ illianis • . \Villiani-to\vn. Ma — .

PAIKXKP. \\ . IP. P.r.idley 1 '"l\'technic Institute. Pe--ria, 111.

PARKER, G. H., 16 Berl . ( u bi

PATTEN, Mi>sJ. P.. Simnvns d.lle^v. l'..'~t"n.
PATTEN, WILLIAM, Dartmouth College, llan-'ver. \. IP

I'M M RSON, J. 'I'.. University of Texas, Austin. Texas.

PANM . FfiRNANDUSj University of Indiana. l'.l< •• imin^t'-n. Ind.

IO2 MARINE BIOLOGICAL LAW iRAToKY.

PEAK si-:. A. S.. University of Michigan. Ann Arbor. Mich.

PIKE. FRANK H.. University of Chicago, Chicago, 111.

PORTER. WILEIXM T.. 688 Boylston St., Boston, Mass.

PKENTISS. HENRIETTA. Normal College. City of New York.

QfACKENiirsii. L. S.. 27 West ~^\ St.. Xew York City.

RANDOLPH, HARRIET, Bryn Mawr College, Pa.

RANKIN. WALTER M.. Princeton University, Princeton. N. J.

REA. PAUL M., The Charleston Museum, Charleston, S. C.

REIGHAKD, JACOB, University of Michigan, Ann Arbor. Mich.

RICE, EDWARD L., Ohio Wesleyan University, Delaware, Ohio.

ROC-EKS. CHARLES G.. Syracuse University, Syracuse, N. Y.

ROM INK, A. P., 1801 "I" St., P.ellingham, Wash.

S' OTT, G. G., College of the City of New York, New York City.

SCOTT, JOHN G., Westport High School, Kansas City, Mo.

SCOTT, W. B., Princeton University, Princeton, N. J.

SHOREV. MARI \\ L., Milwaukee-Downer College, Milwaukee,

Wisconsin.

S.MN ii, I'.I.KTRAM G., University of Wisconsin. Madison, Wis.
SMITH. KRWIN F., U. S. Dept. Agriculture, Washington, D. C.
SOLI. MAN. TORAI.D, Western Reserve University, Cleveland, Ohio.
SPENCER, HENRY J., Columbia University, New York City.
SPOOXER, GEORGIXA B., Columbia University. Xew York City.
STOCKARD. CHARLES R., Cornell University Medical School, New

York City.

SIREETER, GEORGE L., University of Michigan, Ann Arbor, Mich.
STRONG, O. S., College of Ph\ -icians and Surgeons, New York

City.

STRONG, R. M., Univer-ity of Chicago, Chicago, 111.
SIMXI.R, I;. 15.. Woods Hole, Mass.

TAYLOR. KATIIEKINE A.. Cascade. Washington Co.. Maryland.
Ti N\I:NT, D. H., Bryn Mawr College, Pa.
I i i'RY, < >. 1'., Purdue University. Lafayette. Tnd.
TnoMi'S'.N. CAROLINE I'.., i<»5 \\c-ton Road, WellesU-y. Ma — .
TIIOM \s. MASON B., \\'abash College. Crawfordsville, Ind.
TIN K HAM, FLORE\( i L.. 56 Temple St., Springfield. Mass.
TOMPKINS, l;.i.i/Ar.i.Tii M.. Nigh School, \\"hite Plains, \\-\\-

York.
TOWER, \\'n.i i \M L., University of t'liira-o, C])ii-ago. 111.

THE DIRECTOR'S REPORT. 103

TRKAIAVKI.L. A. L., Ya^sar College, Poughkeepsie, Xc\v York.

I '-HER, SUSANNAH. 1007 \Y. Illinois St.. Urbana, 111.

\VAIIK. FREDERICK C, Western Reserve Medical School, CK

land, Ohio.

WATSON, FRANK F... _"( Maywood St.. Y\"..rc<.--ter. M.
\\"KKI:I-;K. F. I.. Anatomical Laboratory, Johns Hopkins Uni-

versity.

\YHKKF.EK. ISAI-.I r.. X... i i. '\". n, T. .k-«l«>. < 'hi.-.

\YIEMAN. II. I... I'liivcr-ity of < 'incinnati. C'incinnati. Ohio.
WlLCOX, ALICE W., I'>5 Pros] . I'r-'vidciu-i.-. R. I.

Wii.iiMAN. I;.i>w\ivi. I;...4^^i ' . We., I'hiladt-lphia. I'a.
\YII.I.I\M-. ANNA \Y.. ?4>> Riversi l( Drive, N\'\v ^^.rk City.
WlLSON, II. Y.. I'liiviT-ity of North Carolina. Chapd Hill. \. C.
\\'(M,|,KI ri'. I.. I. . Y"alc I'niv Mew Havi-n. Conn.

\\""i.ri . J \MI - I . Trinil . I >urham. X

\\'I<II.IIT. R. R \.v •:: . T- >r< mt< •. * "anada.

ES, ROBERI M.. Harvard I'nivf ambrid-r, M.

THE SEX RATIO IX HYBRID RATS.

HKLEN DEAN KING,
THE WISTAR INSTITUTE OF ANATOMY AND BIOLOGY.

It has frequently been stated that the sex ratio in mammals
can be altered by hybridizing, although there are but few series
of observations that give support to such a view. Buffon (1709-
1788) seems to have been one of the first to note the apparent
excess of males among hybrid offspring; but as his records com-
prise only a comparatively small number of cases they cannot be
considered to afford conclusive evidence that hybridizing changes
the normal proportion of the sexes.

In recent times little attention has been paid to the question
of the sex ratio in hybrid offspring. Davenport ('06) ascertained
the sex of 377 out of a total of 950 hybrid fowls obtained in an
extensive series of investigations on inheritance in poultry, and
found 204 males and 173 females. Taking the sex ratio for
any given lot of individuals as the number of males to each 100
females, it is found that among these hybrids the sexes exist
in the ratio of 117.91 males to 100 females. This sex ratio seems
to indicate that there is a pronounced excess of males among
hybrid fowls, yet Davenport states that the proportion of the
sexes in hybrids is normal, and that "the exceptions to the law
of equality of sexes in hybrid offspring are individual and not of
general significance." Davenport attributes the excess of males
among his 377 hybrid fowls to a difference in the death rate of
the two sexes, yet he gives no figures to show that among young
poultry more female^ (lie- than do males. Human statistic-.
as well as the records I have been collecting for the albino rat,
indie, ue that mortality i> greater among young males than among
\ on ii- female--. I )a\ enport'> conclusion does not appear to apply
to hybrid birds in general, as < iuyer ('03, '09) found a great excess
of male^ among hyl>rid pigeon^, and the data which he ha>
collected regarding the sex of other hybrid binl- ^ho\\ a \ er\
much greater number of male- than of female^ in practical!)

e\ cry case.

104

THE SEX RATIO IN HYBRID RATS. IO5

In order to -tudy the color combinations in hybrid mice von
(.uaita ('98, 'ooj crossed albino mice with Japaiu-e waltzing
mice and inbred their descendants through six generation-. In
hi- various table- von ' '.uaita gives the sex of the great majority
of the hybrid oiT-prin- obtained in each generation, yet he maki •-
no comment whate\er "ii tin- relative proportion ot lib
in thi-e individual-. For the purpose of comparing the
ratio in hybrid mice \\ith that in hybrid rat-, \on Guaita'-
iv< ord- ha\e been collected and the -e\ ratio- calculated tor the
indi\ idual- in each ^ein -r.n ion. The data thus obtained
-ho\\ n in tin- l"<>lln\\ in- table.

i |

I Al;i : I .

. I » \ I \ I • 'K II V1IKII' \1

N

1

i j

1

1

1-

9

|2<

I<

1 1 N

it

According tO these : M'li (".uaila oblained a total

407 h\ briil miie; but he gives the sex of only ;vv> indi\ idual-.
ot \\hich Is'i were male- and K.; u ere female-. In ihi- !<>•

lubrid-, thei< n 113.17 males to 100 females. Thi-

ratio i- iindoiibtedK higher than that \\hich i- normal
cither nl" the t\\o species \\ith \\liirh the e\| >erinieiit - be^an.
Sclnill 'lind a neaiK f|iial | u'o] »oiM i-'ll <>!" tin - in

over i ,000 albino mice that had been bred under dittermt en\ iron-
mental condition-: a -iniilar n-latioii betueeii the .\i-ted
in the l.o ualt/in- mil e bred b> \'erk«-- '07

lxa\nioiid and Maud I >. Pearl '08 have tabulated over two

hundred tlmu-and records ••!' legitimate birth- i.ccurriiu in the
cit\ ul" Hueiio- .\\re- during the years [896 [905 inclu-i\e. in
order to ascertain \\ he t her tin re i- a teiideiirv toward- an •
i\e inoductioii of ott'-prin- o|" one -e\ in CTOSS a- rompared \\ith
pure mating-. Their table- -how that in pure matings, either
anioti- Argentine, Italian or ^pani-h -lock, the number ot m

106 HELEN DEAN KIN',.

to each 100 females ranges from 100.77 to 105.55 in various
cases. Among these races, tl KMT I ore, the same relation of tlic
sexes exist as is normally found \\ hen any large number of human
births are examined. When the parent^ \\ere of different racial
stock there is a marked increase in the relative' number of male-;
the sex ratio showing from 105.72 to 106.74 males for each 100
females. The general conclusion reached by these investigators
i- that "there is a definite tendency towards an excessive pro-
duction of male offspring in cross as compared with pure matin- -
in the data considered. Further, it appears that this tendency
i- uniformly exhibited in all the mating."

The records given above comprise all the sex data for consider-
able numbers of hybrid offspring that I have been able to find.
All of these records are in complete accord, since in each case
there is an excess of males that is much greater than that which
i- probably normal for the parent stock.

For several years past Dr. S. Hatai, of the Wistar Institute, has
been crossing the wild Norway rat (Mus noroegicus] with the
albino rat (Mus noroegicus albinits] and breeding their descend-
ants, in order to obtain material fora study of the central nervous
-\~tvm in hybrid rats. The records for the many hybrid off-
spring that have been examined include the sex data, and Dr.
Hatai has generously offered me the use of these records for the
study of the sex ratios that is given in the present paper. A-
far as I am aware, there are no published statistics regarding
ilif proportion of the sexes among hybrid rats, although three
investigators, von Fischer ('74), Crampe ('^4) and Bos ('1)4),
have carried on extensive hybridizing and inbreeding experiment.^
with these animals.

Cuenot ('99) examined 30 litters of voting albino rats containing
a total of 255 individuals, and found among them a sex ratio of
105.6 males to loo females. Records which I have made of the
sex of 452 young albino rats belonging \<> NO litter-- uive a sex
ratio of 107.33 males to 100 females. The sex ratio in the
albino rat, therefore, agrees with that for man and variou- other
mammals, since the number of the male-, i- ^li^htK greater than
that of the females.

No statistics have as yet been collected re-anlin- the normal

THE SEX RATIO IN HYBRID RA 1 - I(>7

proportion of thi - amon- wild \..r\va\ -rat-. JtnL , >ni

ill- ;tio found in general among the mammal-, it i- very

probable ih.it there i- approximately an equality of th-

in the wild Norway rat as in the albino rat. In the total of

albino rai s i-\amined by Ciu'-noi and by myself there \va- found a

ratio of io(). 40 mal.- to i oo female- Thi- -ex ratio i- con-id-

'. throughout thi- |ia])cr. to expre-s the probable normal n-la-

tion between th( \\ ild rat a- well a- in tin- albino rat.

I "in series of experiments \\i-n- m.nle in which albino rat-

\\ei ised \\ith \\ild Norway ; - R( rds an- a\ ailable

l"i d of \i,$ individuals belon^ini; to tin- 1 ^ciu-ralion.

ot \\hich .so \\rn- in...' d 74 \\.-iv h-malr-. Amon- tlu-c

lubrid-, ili«: t sex i • [20.26 males to too

I ••male-. Although i hi- -. o is murh hi.chrr than that \\ Inch

i- probabK normal |o- r of tl). nt -prcir-. il i- nol

hiwl) .1- that t'-imd in \oii i . . m-ral ion of h> brid

mil •• \\ hen- t In in the ratio . male- to loo

Irlllalf- l.lblr I

All ot tin- h\bi'd i g to thi I g< iii-raljon f( .r \\hich

irds were made wen . months old when the) \>.

killed and examined; tin- leu indisidiial- lh.it died be!. :<'h-

malmitN \\eie iioi iiu luded in il id-. li cannoi be

deleiniined uith aii\ . to \\hal

propi,rii,,n ol the < n:n, numbei • prin^ brl. to tin I ,

generation the abo> o applie-. \.i\ t. \\ of the young

rat- died, as i- kiio\\M, and there i- il" reason to -up;

t hat l he mm talit \ \\ .1 n( it.-iii.tle- t h.in

amour tin- ymir^ male-. It seems probable, then-fore, that the
-ex ratio in the l<>, indixidual- for \\hich data an- a\ailable i-
lairlx repiv-entati\e lor iln t ntire number of oiNprin^ produced

in ihe coir the variou riments to which the) b«-loir^eil.

I WO Ol the lour -erii •llllellt- Illelltiolled ,lbo\e U ere

exteiidi d b\ mating \aiiou- individuals belong iir^ to the lir-t
lieiieration o| lubrid-. In pairing ihe-e animal- no attention
ua- paid to their blood relationship, and imdoiibtedK nearly
relati-d indixidual- \\ere mated in in. my cases.

In "in- series "f experiment- only a \er\ -mall number of
indixiilual- beloiisjn- to the -econd generation of lubrid- died

I()8 HELEN DEAN KING.

before reaching maturity, and the 114 rats for which n-o.nl- are
at hand form a very great majority of all of UK- offspring pro-
duced. Of these individuals 61 \\eiv males and 5.^ \\erc femaK •-:
this gives a sex ratio of 115. 0() male- io 100 female-.

In the other series of experiments the majority of the indi-
viduals belonging to the F2 generation died while immature, and
records were made for only 27 individuals. Since twice as many
females as males reached maturity, it is evident that in this
instance either the mortality was much greater among the young
males than among the- young females or that there was a very
unequal distribution of the sexes in the newborn rats.

Individuals from only one lot of hybrids belonging to the
second generation were mated. The animals were paired accord-
ing to color, their possible blood relationship being entirely dis-
regarded. Dr. Hatai found, as have other investigators who
have bred hybrid rodents, that there is increasing infertility
among individuals belonging to succeeding generations of hybrids.
The total number of hybrid offspring belonging to the F3 genera-
tion was relatively small: many of the rats were stunted in their
development, and the majority of them died before reaching
maturity. Since records are available for only 23 of these
individuals the sex ratio among them can give no idea of the
probable ratio in a large number of hybrids belonging to tin-
third generation.

Since in mating individuals belonging to the first and to the
second generation of hybrids no attention was paid to their
blood relationship, it is very probable that in >ome cases closely
related individuals were paired. There is the po— ibilitv, there-
fore, that inbreeding might have had some influence on the sex
of the descendants. According to Diising ('84), a noticeable
increase in the number of male offspring i> produced 1>\ inbreed-
ing, in man as well as in various mammals. Inbreeding could
have had little, if any, influence on the sex ratio- in the-e \ ,u ion-
lots of hybrid rats. The sex ratio found among the 114 hybrid.-
of the Fo generation which comprise practical!) all of the off-
spring produced in the series of experiment- to \\hich they be-
longed, is somewhat less than that found among the offspring
produced by crossing pure Mock (.Table II.). The data tm

THE SEX RATIO IN HYBRID RATS. IOQ

tin- other individuals belonging to the F? generation, a- \\e!:
those for the hybrid- of the F3 generation. -ho\v an excess of
female-. The-e latter record-, cannot be considered to turni-h

lence against Dti-in:;'- theory, since they include -uch a
very -mail number of individual-. There seem to be no record-,

• •pt th' en by I Hi-ing, that -ho\v that inbreeding alone

prodni e- a relaii\eh . r number of male than ot temale

offspril . -• liull/<- \\a- unable to de!r. t any change in the

rat io <if albino mice that had been inbred for four generation-,
and I have found no unu-ual « •!' male- anioii- some 500

individual- beloii^in^ io ti\e generations of inbred albino rat-,
keeoid- are available for ui other hybrid rat-, nio-t ot uhii'h
\\ere ihe oti-prin^ "f reciprocal Cl I'he-e indixidiial- all

belonged to the I _ or to the I ^-neralion of hybrid-; and amon-
them were 7- male- and 4<i tVnul. 5. Thr-e In brid- \\ ere -elect .d .
b«-< au-e of their -i/e and condition, from a considerable number ot
indi\ iilual- for \\liich im record- \\eiv made. There \va-. theie-
fore, lindoiibtedK all lllicoll-cii -U- -. leetjoii of male- in choo-in-
the rat- to be , \amined. -in«v the adult male rat i- considerably
larger and hea\ ier than the female. The data for the-e Nation-
lot- of -(lei ted indi\idual- ha\e been coinbine<l in one ;;ioiip,
-iiii-e the\ are of \.ilm- onl\ 1 • -ho\\ the -e\ o| a

( on-iderable number -.1 h\ bi id-.

The |o|lo\\ iii'^ table ;J\e- i -imim.HA of the 5CX record- |o|-
all ot t he h\ briil rat- te. . nded.

TABU II.

llshkli' 1<

.
1

S

1

114

1

.Ml

!•". aii'i 1

i .• i

( -

49

•

194

II'

In the total of 425 lubiid rat- lor \vhich ri'cord- \\ere made,
j;i \\eiemale-and I «i \ \\ el e lelliale-. .\mon.uthe-eilldi\idual-,
therefore, there i- found a -e\ ratio \\hieh i- con-ider.ibl\ higher
than that \\hich i- probably normal for either the albino or the
\\ild \oi\\a\ rat, -ince there are \\<>.<») male- to loo female-.

I IO III I EN Dl-.AN KING.

If from the above table we omit the records for tin- 27 indi-
viduals belonging to tin- Kj generation of hybrid:- \\hirh formed
only a very small percentage of tin- total number of offspring
produced in tin- experiment to which they belonged and also
the records for the selected individuals belonging to tin- F- and
to the F3 generation, there remain the data for 277 individuals
which comprise the very great majority of tin- hybrid offspring
obtained in several series of experiments. It would seem as if
the sex ratio in these individuals might justly be taken to repre-
sent the probable sex ratio in any large lot of hybrid rats. Of
these individuals 150 were males and 127 \\ere females; this
gives a sex ratio of nS.i i males to 100 females. This sex ratio
is but very little lower than that found in the total number of
hybrids for which records are at hand, and it is not very much
higher than that in the 356 hybrid mice bred by von Guaita
(Table I.).

The excess of males among these hybrid rats is seemingly
beyond the limits of normal variation in the proportion of the
sexes in the pure stock, and it is too uniform in the various series
of experiments to be attributed to chance. It appears, therefore,
that hybridizing alters the sex ratio by producing a marked
increase in the relative proportion of males. This conclusion is
in essential agreement with that reached by Buffon, by R. and M.
Pearl and by Guyer.

Guyer ('09) has offered an explanation for tin- excess of males
among hybrid offspring which accords with the theory, advocated
by a number of investigators, that sex is determined in the ovary
chiefly by nutritive conditions. Guyer suggests that in the /y-
gote produced by cross fertilization there would probably be
"more or less default in the metabolic processes because of the
incompatibilities which must necessarily exist bet wren t\\o gcrm-
plasms so dissimilar." An interference with the metabolic
processes would naturally retard the constructive phases of
metabolism in the fertilized ovum, and therefore tend to tin-
production of relatively more males, since tin- theory assumes
that females are produced only when the conditions are most
favorable for constructive metabolism.

To explain the sex ratio in hybrids according to the ciirreni

THE SKX RATIO IN HYBRID RAT-. Ill

liyp<>the>i- that tin- male i- tin- -e\-deierminin- factor seem? t<>
necessitate tin.- further a— umption that fertili/atinn is selectixe
when individuals belonging to different race- are cn---ed, the
egg offering greater n-i-tancr to the entrance <>t a -permatn/oan
ilia; i- female-producing than in <me tliat i- male-producing.
Then- i- link- r\ idi t, that fertilization is ever selective.

Tin- pn.li.diility of it- <>< rurrciice as a normal phcm >meii« m ha>
ently been advocated bj \\< .i\« although Wil-m '10

considers ii "SO iinpr-'haMc as alum-l 1" inxalidalr an> inti-r-
l»|-ciaii«»n into which it i-nti-:

Guy< lia- -h(i\\n that thnv i- t •• .n-idi-ralilr amount nf

dr^i-iu-iMiidii in tin- t. •: many luhrid i>ii;i-('n-: almonnal

mitoses .md mi>-hap»-n -jn-nnal< •/( >a lx iiu "t trniiu-nt occnrrciicr.

( )tln-r nli-rr\ IT- li.r. • d thai the gonads in hybrids are

o|- It--- drfci ii\i-- lull in-ill- "1 them ha\c made a lii-li -l«
in\r-iiv;alion in "idi r i-> a-« rlain \\hat -liiKMir.il i hani^cv ha\i
n pr..dinrd. Guyer's «'!--• i \ alii-n- .-tn.inj\ -nc^r-l that
ill- «-| \.ilm- ma\ In .-lit.iiiu-d h\ a -lu<l\ <-t the ^miad- in
otlitT li\ lirid-. M. iii-rial i- ln-in^ collected for .m inxf-ti^ation
of tin- cmiad- in h\l>rid rat-, in the hope that it \\ill at lea-l i;i\c
--me (hie i-» the i.ui-e fi -r the iiu • iilit\ in -nci ei diir^

^eiieratii-n- <>! li\liri-l-. e\eii it ii .ill"nl- n«- e\ ideiice that \\ill
en.il-le i.ne [» off( \planati«-n f- -r the altered

ratin in h\ hrid fi -rm-.

I I II-K \ 1 l Kl- i I I I
Bos. Rit/cma.
'.J4 LJntei • '• : ^.ui.!-. I

\1\

Crampe, Dr.
'84 /in in '. .iiiiiifii \v ii R.W iltati dei K-

/. ili in. i, K.iiti-ii mil \\il.l. n. l.in.luin! fahrl . ••• IM.XIII.

Cu^not, L.

'99 Sur la .li-triiiiiii.iti-'n <lu * . I'-ui':

ilc l.i M«-i^ii|i:.-. I
Davenport. C. B.

'06 Inliniuiiii «• in l'"iilti\. I'.il' ' .;-'.

Dusing, K.

'84 I>i«- K-xiilH-iuiii; -\.-iluilf.. i -li-ii \'C-IIII--|IIIIIIK

Menachen, riere, un.l I'll.m/'-n. Ji-n. /'-it-iin. Naturwiss., H-l. X\ll.
von Fischer, J.

'74 Hi-nli.i. lit iir.^'-n uebei Ki' : "ini;i-ii \-rr-iliii--lt-iii-r l-'.ti l» ii-pii-Lu t< 11 innci-
Imlli ei] -'I t''ii. H'l- X\

112 HELEN DEAN KING.

von Guaita, G.

'98 Yersuche init Kivu/im.ni-n von \ n -, hir.l.-nrn R.I--CH ,ln

Ber. Xaturforsch. Gesellsch. zu Freiburg, lid. X.

'OO Zweite Mitthcilung uebcr Vrrsiu h<- init Kn-ii/uniirn von v<TM-hird<-ncn
Hausmausra^sen. Ber. Xaturforsch. Gesellsch. zu Freiburg, Bd. XII.
Guyer, M. F.

'03 Spermatogensis of Xormal and of Hybrid Pigeons. Bull. I'niv. Cincin-
nati. Xo. 22.

'09 On the Sex of Hybrid Birds. Biol. Bull., Vol. XVI.
Heape, W.

'09 1 In- Proportion ot the Sexes Produced by \Yhitr and Coloured Peopli--

in Cuba. Phil. Trans. Royal Soc., London, Vol. 200.
Pearl, M. D. and R.

'08 On the Relation of Race Crossing to Sex Ratio. Biol. Bull., Vol. XV.
Schultze, O.

'03 Zur Frage von den Geschlechtsbildenden Ursachen. Arch. mikr. Anat.,

Bd. LXIII.
Wilson, E. B.

'10 Selective Fertilization and the Relation of the Chromosomes to Sex-Pro-
duction. Science, Vol. XXXII.
Yerkes, R. M.

'07 The Dancing Mouse. The Macmillan Co., Xew York City.

THK RHYTHMICAL MOVEMENTS <>F LITORINA
LITnKKA SYNCHRONOUS WITH OCEAN TIDES

J. I). HA-KMAX.
l\ I Kcil.l • riON.

Through the kindnt — ol 1 >r. Snnim-r, tin- \vritrr \\a- ^ivcii an

(i]i])urt unii y to \\ork in the l.il >or.ui >ry of the Bureau of Fi-herie-
,ii Wood- Hole. M :--.. duriiu the -umin.T of MO;. He i- in-
debted to I >r. ^tinnier for many valuable sug{ >ns.

Tin- plirp"-e o| ||u- pre-ellt paper 1- In -llo\V tll.lt tile IlK'N e-

meiit- ol /- '. \\liich arc -yiH-linuioii-> \vith oivan

tiilc-. an- not diir. ilin-i il\ . io riilu-r ;<• oi r< .pi-in or phoi- .i.txi-
luii io i In- .it lion of tin- til in of \\att-r on the moiv or K--- i-\po-( d
-nail-. In ani\in^ al llii- lomlu-ion main rxprrinu-ni- were
noi onl\ in tin- lal'oi.ilor\ Inn al-o mnli-r natural
In addition, ill, hal»ilai-o| i hrrr -| n vii- o|

f.iturimi um- -itidicd in ddail. ^-iich oli-ri \ al ion- arc nciv--arv
\\hcii tin- naliir.il o-rillai ion- of Litorinn. \\liirh rorrc-pond to

the rise and the fall of the tid< .n-i-lcn-d.

l.a-il\ ihc \\iiu-r \\ill aiu-inpi to -lio\\ ihal Litorinn lite
doc- not n-iain a rlnihin, \\liidi is synchronous with tin- tide-.
cilhcr in ijiii<-i .u|iiaria or in naiun- \\licii lip ts of lilni- o|

\\alci ha\c I .ecu renio\ed 1>\ ki epin- the -nail- pcrm.iiicntly
-ill. UK 1-^1 d.

I . I'm II \i-.i i \ i ' 'i Litorinn.

lndi\ idtial- W an- a- a rule foinxl on n>. k-

\\liich lie I .el \\ceii ihc tidal mark-, 1. nl ..rca-ion.ilK an indi\ idnal
OCCUrS on rock \\ecd- ,nid c\ en on the muddy -lion-. Thi-
Bpecies \\a- alumdanl everywhere in lln- region of \\'..od- Hole,

excepting in Grape and Tashmo pond-. \»>\\\ of which are l>ra<-ki-h.

i I ndi\ idnal- • >f Litorinn pallintn li\ c as a rule < m n >ck \\ cci I -

\\hich ;^ro\\ l.ci \\ccii ihc ii<lal mark-. Thi- -pci ie- \\a-noi cii-

i.iuntcn-d al V.l.-ka I'-.inl or in ('.rape. Lagoon and Ta-hmo

1 Tin- liii.il pir|..ii.itii'ti "I t: "lint •>! ilic writ

1-im trip in S..iith Ann-iii'.i i"i tin- ( '.iin. .ui.- Mu-ciini. • >nly tin- in. in- imp. .1 t.int
1 .tn.l in.my "I tin- .l.-t.iil- liu\ •• l.<-t-n ..inittfl.

i '3

I 14 J. D. HAM-MAN.

ponds. Ii is always more abundant along protected ci >a-t- and
hence its absence I'niin Nobska Point, which i- exposed to the
open -c-a, was to be expected.

1 Individual- ot Litorina li!on-a i -aid to have been imported
to tin- American coast from Europe) usually live on rock) -nrfaces
\\ Inch lie bet \\i-en the high-lide mark and one loot below the lo\\ -
lide mark. The specie- \\a- not loiind in 1 a-hmo I'oiid but a
l'e\\ \oung individuals \\ere -ecu in ('.rape Pond living (»ii trau--
plauted oysters. This observation, taken in connection with
other- described belou , indicates that the absence ol Litorina from
these bracki-h pond- \\.i- not due, direct ly. to the lower specific
gl avity that t'xists there.

In order to explain the restricted distribution of Litorinn the
following obserxations and experiments \\ere made:

A. l-'ood.- — Litorina fxillid/u is neail\ a!\\a\s found on two
t I'ininon species of rock weeds, and its supply of food is doubt-
less associated with these plants, while Litorina rudis and
litorea eat small green al^a- which grow more abundantly on
the rocks which are situated between the tidal marks.

When individual- of Litorina litorea were placed in shallow
aquaria, which contained rocks from the sea shore, from Lagoon
Pond or from the street and contained also pieces of glass and fish
they came to nest for the most part on the rocks taken from
normal sea water. The snails, which were taken from the
entrance to Lagoon Pond with their accompanying rocks, a!-o
-ettled on the sea rocks.

B. Salinity. When indi\ iduals of Litorina litorca were placed
in a mixture of equal part- of fresh and sea water, they were
able to crawl about; but when they were placed into a mix-
lure of two parts fresh and one part sea \\ater, all died within
ei-ht days. When the\ \\ere placed in sea water, to which a-
much as fourteen grams of sun e\aporated sea -all had been
added to each Xoo c.c. of sea water, all died within eight da\-.
The specific gra\ityof the various u-st solutions indicated that
Litorina could li\e in water which has a louer specific gra\ii\

t han was obsei \ ed in < .rape and T.ishmo ponds.

('. '/'<-nif><-rali<r<-. -The \aiiation in the tempera! tire along the

coast did not appeal t»,itlect the distribution ol Litorina. Snails

RHYTHMICAL MOVEMKNT5 OF I.ITOR1NA. 115

\\cre -ubjt/cted i" ini'.ch greater cluing - of tempcrat ure than
ir alon^ tin- coast and no ol>-er\al>le effect- were detected.

D. Pressure.— Severs.] individual- of Litorina litorca were-ul>-
mer.ucd in cages, which contained MM -hore rock-. in a depth uf
."-. !>> and .V feel of \\atcr. T\\o week- l.lUT the -nail- appeared
to l>e perfectly normal.

L ! .:!inn. — Litorina litor*;: i- \ ery -en-iti\e \» -lisjit chaiio -
in aeration and e, >n-e(|iiently it i- \ er> 'difficult to arrange t IK-
Condition in an aquarium SO that tlu-\ l>eha\c normal 1> .

I . \o direi ot li-hi (,n the di-irilunion of Li.'orina

(ould IM- drif. ic«l citht-r IA Miifiit or din-ri observations

made in nal i;

I i :' \\hen indixiduaU ol . •:<[ litotn: are \ioleiitl\
>pla-hed l'\ \\a\i--.lln-\ 1 •«•( •• -me t |llie-cell t .

II CharG lndi\ idual- of Litorina litor,,i are
raiel\ toimd on either -and or mini, due in part to the alienee
ot tood ill -in h plai «•- and in par I to the difficult \ eiicountereil in
<ra\\!iiiK over -in h -urfa. . - I hi- i- e-|ieciall\ true \\lu-n the
waves ton^taiiiK < han^e the |io-ition oi the -aim- and hence
lumlde the -nail- al-out at ramloin.

I Mm-!:, Litorina litorca doe- not craul on dr\ -in la.

II. FACTORS WHICH DETERMINI i HI M< >\ i \n \ i- «>i

It a|>|>eai - ( \ ideiil Inun the preceding data that thecharai
lei o| the -urlaie. nioi-iure and I ..... I are the chief ta« tors vliich
detei'tnine the litoral di-t t il nit ion of the-e -nail-. Hut a- I hope
to demon-uale in the tollouin^ pai;e- the-e three factor- pro-
duce landom mo\emeiit- ol the -nail- and -Inuild not he con-
ton nded \\ ith the tai tor- \\ Inch direct l\ [irodm e rh\ ! limit al mo\ r-
nieni- cori-e-poinlin^ to the rise and fall of the tide-.

The lollo\\ in^ experiments and ol>-ei \ at ion- \\ j||, | think, deter-
mine \\hat the "«lirecii\e lone" ot riuthmiial movement- is.1

I. I'he -nail- \\hich are located on Mat hori/oiital -urLc
ln't\\eeii tin- tidal mark- do not -how rhythmical moxement-
\\hich corre-poilil to tho-e of the tide. The -nail- \\hi.h are
iled or placed l>e!o\v the low-tide m. irk on i-ilhrr a \erti<al
or a hori/ontal surface do not -how rhythmical mo\ nnrnt-.

MM m.uiiiiiv ••! t In •• \]" -niiii-iH - riin-il ali.ni; tin- -i-a-lmn- \>\

inaikin.n tin- -nail- an-l tin-

1 if, J. D. HASEMAN.

But tin- -nails which are- located mi an\ more or le-s vertical
-u rl act • bei ween tin- tidal marks do exhibit rhythmical niovemeiu-
which correspond to those of the tides.

2. \Yln-n a stone, which has snails crawling on a llat surface
belo\\ ihc low-tide mark, is raised and lowered in the sea, tin-
-nail- do not -how rhythmical movement- \\hich correspond with
those of the stone, /'. c., their movements were not directhe.
The same tv-iilts are obtained when, with the llat surface hori-
/ontal, the stone is raised out of and lowered into the sea. But
when, with the surface \eriical, the stone is raised out ol and
lowered into t he sea, the snails at once showed rhythmical mo\e-
meni - \\ hich correspond to those of the stone. These same snail-
were known (from observations made on snails placed bei \\een
tin- tidal marks and below the low-tide mark) to have shown in
some cases and not to have shown in other cases rhythmical
movements during several days prc\ ions to the experiment-.
Therefore an\ alleged retained rhythm has nothing to do with
the abo\ e results.

Kvcn more interesting is the fact that when, with a vertical
-m-faiH', a stone, upon which snails were crawling at random, was
rai-ed out of the sea, the snails alwax- followed the vanish! M.:
film of water even when the vertical surface was rotated through
an angle of 180°. In this case the rotation of the vertical surface
would reverse the direction of motion of the film of water and
the snail- would at once turn around and follow it. But if most
of the \\ater was previously removed from the surface ol the
-tone, in order that the film might entirely disappear before tin-
snails (which 'were crawling downward in the direction o! the
\ani-liing film i had reached the lower surface, and if, as the
film was drying up, the vertical surface was rotated through
an angle of lSo°, tin- -nails continued to crawl tor some time-
in the direction in \\hich they had started. In other words,
the -nail- crauled upward instead of do\\ n\\ ard. They con-
tinued io crawl thii- until the rough -urlace, lood and moisture
either delleded or -lopped iheir movements. In the above ex-
periment, the mere turning of the moi-1 but lilmle— surface
through an anije of I So doc- not si-em adequate to n-\er-e ai
once ihe n-a< lion to gr.i\il\ and light, if either ol these ha\e a

direct inthieiice on i he rh\ ihmical movements of Litorina.

RHMHMK AI. M< .\KMI MS OF LITORINA. Ii;

V When tin- eves of Litorina !i(o>m ,iiv dc-tro\i-d. thcv -till

« ' - •

-ho\\ rhytliinic.il mox vim-nt-. <m xvrtical -urface- Let \\eeii the
tidal mark-. I ndi\ idual- of Litorina litorca al-» o-cillate with
the tide- during dark niidit-.

4. I hiring the month- <>f |nl\ aii<l Aui;u-t. tin- \\riter could

• any tvlatixe cli n tin- po-ition of -n.iil- , ,n 1.

l.oulder-. It" phototaxi- i- an ini|M .riant factor, a marked change
tt-.iild In- expected 1..-. all-e the dail\ 1. 'he tide- a- \\e!l

as tin- diurnal and nmiuhh change- »\ the relative x.lar pn-itinn
r.t" the earth. c-.ii-tanll\ change the relatixe ail-le "|" the ra\ -
«.!' li'^ht.

5. \\hen it rain- <ui e\im-ed -nail-. tln\ 1 u • • nt

just as they do when splashed \>\ the water. There!. .ie dmin^

-ic.rm\ as i"iii|iared \\ith fair da\-. a < < ui-idetal >le ditlefeiK e in
the ,1111. .unt i.l ..-(illation- n mil: was noted.

Sul MI -nail- ci ui Id I >«• <>nl> -liv;htl\ «.r n«t at all d.in -cted

l.\ current- »\ water. Tin- .intent- \\cte |.i.'du.ed artituialK
an'dal-..|.\ movil :ie- \\ it h -nail- t hi • ui'ch t he u atei . Mm.,

the ln-haxii.r »t /. -m xertical -urlacr luM\\«-eii the tidal

111.11 k- i- not ol .i i heotiopi. natun .

7. \\heii indix idual- are 1« H hi's'" and dtx on x«-rti«al -ml.i.
dlliin- l..\\ tide. th«x ...me to i. "direcu-d ii|.uard." /.
\sith their head end tou.ird the -k\ . I hi- i- true lot all -id«-
of the -ton.-- and i- ..Lxiou-ly due tn the -h.ij.. o| the aperture
of the -hell- \\hich make- it tar e.i-ier for e\pn-ed indix idual-
lini; ihu- to x ertii al -url.f

In. li\ idual- o| >KI liturfi are on an average found

^liiditly higher ..n the north -ide of l.ir^e l.oiihler- than on the
-oinh -ide. Thi- appeal- to In- due ier nioi-ture and a

higher :<n>\\ th ol ln.th al^.e and barnacles Over u hii 1 1 -nail- < rau 1
\\ ith dit'tictilt \ .

<i. SOIIH- Lottie- p.utlx tilled \\ith \\ater and air plus hydro-
and other- partlx lilleil \\ith carlion dio\id \\ere u-ed to te-t

the effect of crawling out of the water into a gaseous medium and

x i. e versa. N" difference- x\ ere detected from that of craulinv;
out of \\ater into the atmo-phen- mile-- SO much hydrogen ,md
carl .on dioxid were u-ed that the -nail- \\.-ie a-| >hx xiatei 1 .

lo. Thin tilm- of olixe oil and kcn.-eUe Were placed on \\ater

Il8 J. D. IIA-I.MAN.

in a<|uaria to test the effect of -urface icn~i.ui. I'ndcr these
< onditions the -nail- had ci m-iderablc difficulty in breaking the
-urface tilni and alwa\- he-itated ,n the surface pii-hin^ them-

selves to the right and to the left until their bodie- wen elevated

-ufticieiilly to break the -urface tension. Their lieha\ior under
the above condition -hovxed that the lu'sitation on entering and
lea\ ini^ the -urface wa- due to tin- -urface til in and noi to photo-
taxis, hydrotaxi- <»r clieniotaxis.

M. \\'lien individuals of LitnriiHi litorca, nidis and pxilliata
\\ere placed in a -mall round a(|iiariuin e<|iial areas of \\ Inch were
painted brown, yellow and dark olive, they as a rule came to
re-t on the dark olive sector, rareh' stopped on the bn>\\ n sector
and still more- rarely on the yellow sector. When the same
-naiK \\ere |>laced in a similar unpainted ai|iiarium before a
window, about one half of them came to rest on the lighter >ide
and half on the darker side. The position of the aquaria were
interchanged and the -.a me results were obtained. Various
-hading experiments were tried and neither positive or negative
phoiota\i> could be detected. In nature one sees snails crawling
at random from lighter areas into shaded areas and \ ice
\ ersa.

The writer wa^ unable to dc-tect \\h\- the snails stopped on
the dark olive sector, but inasmuch as such conditions are not
met on vertical sin-face- of stones between the tidal mark, the
results appear to have nothing to do with the rhythmical move-
ment- of Liiorina.

12. Main- -nails \\cre placed in quiet aquaria and aquaria in
which running water \\a- kept at a constant le\ el and they never
exhibited an\ -i^n- of o-cillations which corre-pondi'd to those
ot i he tide-. In the-e experiments. snail> \\hich were kno\\n to
have be.-n pre\iou-ly o-cillatin;^ with the tide- \\ere u-ed. I
also |)laced snail-, which had been oscillating with the tide-,
on \erlic.il -urface- bt lo\\ the low tide mark and an average of
many count- did not -how rhythmical movement- \\hich corre-
sponded to iho-e of the tide-.1 ( )nly when the -nail- are touched
by the -urface film of water are directive movement- called
forth.

1 M -nt l\ ol milai i'-Milt.

RHYTHMICAL MOVEMENTS OF LITORINA. IIQ

| )|-.( i SSI< ,\

Mit-nkuri 'oi found that the routine— of tin- -urface scatters
the -nail- .IIKJ th;it the\ m-vt-r crawl on dry »urface-. He al-o
stated that Litnriua cxi'^Ha lie-itated mi entering the water and
con eluded that their oscillations with the tide- \\< -re due to photo-
taxis. In thi- connection tin- writer found that the -nail-
he-itated a- n ui cli on leaving the \\ater a- they did on entering ii .
Tin- fact, taken in comieciiim \\ith -e\eral of my e\|)eriuient -
and e-pe<ially tho-e on -urface tension, -hnw that the < •! i~i-r\'ed

*

he-it. ition \\a- due to the <-t'|. -he -urlai >• tilm ot \\ater and

not in photoiaxi-.

liohn '05 .'1- IK hide- that ]>lx>i»n.i\i-. i- the important

lai tor \\lmli determine- the "-< illatory mo\fmeiit- o| Litorinn.
Hi states thai !.il»rin.i nul. nd ///«;•.•<; h.i\e lor all

their li\e- liei-n desiccated a^ain and av;a;n. and \\heii |. laced in
(|iiiet aijii.uia. all three -|>e(ie- retained .1 period ol' rh\thm
-\ IK hioiimi- \\ith the tide- and la-tin^; liom thirteen hour- to
loin it en d.i\ -. M\ oli-ei \ .ition- -hdu th.it l.ilorhni lit»" <; do, ~
not ha\ e -IK li .in i -laNi-hed rh\ thin.

Ilie l.eha\io|- •.) , mi in e\| n-i inieiit 2 i- -tittK ieiil to -ho\\

th.it the "directive foi in their rh\ thmical nio\i-nieiit-

-\ IK hi i- \\ iih tlii tide- i- t he ill m o! \\ater ami not phototaxi-.

\l-o the fact, lli.il onl\ -n.iil- oil \erlic.il -url'.ice- |iet\\eell the
I id 1 1 mark- ha \ e o~eill.it or\ movements, shows thai the direct i\ r
force i- not phototaxi- lu-cau-r man\ -nail- on flat hori/i nu al
-ml .mid ciaul to t '• arlace and de-cvnd to

the base of the -tone. l\-pecia'l\ the ol p-er\ at ion- made during
dark niuhl- a- \\ell a- maiix other (\perimetil- alieadx cited
indicate that Molm'- \ i< \\ i- nnteiiaMe.

.> 1 I -!' >\.

Ill ( -oiu lii-ion. theii.it a|>pear- that -ulmier.ued indix idu;'.!- of
Tina litorcn ciaul almni on i it random, perha|>-

in search ol looil. The\ ne\ er cra\\l on dr\ -url.ice- and con-
-e«|Uentl\ are not found above the h;;Ji-tide mark. When the
tide ri-e-. the en\ ironnii'iit i- temporarily enlarged and con-
sequently man\ -uhmer-ed -nail- on more or less vertical surfao -
1>\ mere chanci will crawl up higher.

J. D. HASKMAX.

The individual^ which \\ere previously left high and dry are
already directed upttard and hence lollow tin- rising tide. Be-
fore inaxinunn high tide is reached, the unevenness of the surface
alt in- with the presence and absence of food scatter the snails
at random because the rapidly rising tide soon submerges them
and then then' is no "specific directive force."

\\ In D the tide begins to fall some of the snails are already
crawling downward and those that are not will be directed thus
by the Mirface film. I hiring a falling tide, some snails are
ng do\\ n beneath the surface, some with the surface, and
above the surface. As the tide falls lower and lower,
main snails lag behind more and more until they are finally left
on filmless surfaces under which conditions there is no specific
directive force and such snails are left exposed. This lagging
appears to be due to feeding, rough surfaces, rough sea, rapid
fall of the tide and the more rapid desiccation on certain days.
As a result of all these factors the amount of oscillation varies
from day to day.

The snails on either flat horizontal surfaces between the tidal
marks or on any kind of a surface below the low-tide mark have
no ''specific directive force" and consequently do not exhibit
"M illatory movements synchronous with the ocean tides.

SUMMARY.

1. Litorina rndis, pallia! a and lilorca are found in definite
zones

2. Individuals of Litorina lilnrea, located on vertical surfaces
I iet \\een the tidal marks, exhibit oscillatory moxeinents which
<Mrn-pond to those of the tide- but they do not exhibit such
movements \\hen they are located either on Hat hori/ontal sur-
face- beitteen the tidal marks or on any kind of a surface below
the lo\\-tide mark.

J. The primary direct i\e force for rhythmical movements is
the -iirface film of water. The -econdary directive forces are
the r|uiesceni position of desiccated individuals, character ol the
moist lire and food.

l. Liturhid litorni ha- no e-tablished rhvlhm in the absence
of t ida! chair

RHYTHMICA1 MOVEMENTS Ol LITORINA. I J I

KKFKKKM I -

1. Bohn, Georges

'05 Attrai tivcin--- i-t ( )-t illati"iii-- '!<•- . \niinaux Mai in- -ur I'mlliR-ncr dc la
I.uinii TI-. In-t. Generale Psychologiqm-. I'.ni-.

2. Mitsukuri, K.

'01 Negative Pbototaxis and other Properties of Litorina .1- l-'actor-^ in Drtrr-
miniiiK it- Habitat. . \nimtatinnr- Xnnlc i-i( a- Japanr-f-. \\<\. IV.. pt. i.

3. Morse, Max Withrow

'10 :\lli-«c<l Rliythm in Pin.- hnmou- \\iili < >o-an 1 i'lr-. 1'

l.\p. Hi<.l. aii-1 M<-i!., Ma\ [8, i<;i". \,-\\ Vi>rk.

I'lll-: LKTHAL EFFECT ()K IM kl DISTILLED

VVATKR ON THF VINKC.AK KKL

ANCHTLLl'LA ACKTI).1

JAMES FRANCIS ABBOTT AND KIIIH1. I KH.I1 KM HARDS

The question of the toxicity of distilled \\ater for aquatic-
organisms appears to be still an open one. An excellent summary
of the literature of the subject has been given by Bullot.-' Ringer
first noted that distilled water is toxic for a variety of organisms,
and attributed this toxicity to (i) an abstraction from the organ-
ism of salts which are necessary for its life, or (2) the penetration
of water into the cells through osmosis, or (3) the imbibition of
water by the "intercellular substance." About the same- time
a contrary opinion was advanced by Nageli1'5 who claimed that
it is not the purity of the water but the presence of the slightest
trace of copper (even so small a quantity as 1/77,000,000) thai
gives water its toxic qualities, whereas if water of this sort is
redistilled in glass it loses its toxicity. Shortly after this, Locke
questioned Ringer's conclusions and decided that the latter's
results too were due to the presence of copper rather than the
toxicity of the water and in i8954 confirmed his belief by experi-
ments with the tadpole and Tubifex. Ringer5 then retestcd his
experiments and partially recalled his former conclusions, agree-
ing with Locke that "pure water (i. e., that distilled in glass) is
completely harmless for the animals in question. This was con-
firmed by Jennings for Paramecium, Miss A. Moore for trout
and tadpole and F. K. Lillie for Planaria. On the other hand
Lyon6 investigated the action of distilled water on developing
Arbacia larvae and found that artificial sea water made up <>! tap
water or ordinary distilled water, whether vapori/ed in copper or
glass, is very toxic for these larva' although tap water \\lnVh ha-

•From the Zoological Laboratory, Wa-liiii^iun University, M. l.miU. M<>.
2 Hullot. Univ. of Calif. Pub.. I'liy^inl.. I.. io<>|. i

XiiKfli, Dcnksch. d. Schweiz. Naturfor^li. (,< \\<\. 33, [893.

4 Locke, Jour. Phys., 18, 1895, 319.
6 Kinder, Jour. Phys., 22, 1897.
'Lyon, Hun.. Mei.i... 6, 190.}, lyS.

122

I-.KFECT OF DISTILLF-D \VAIKR U\ Till VINEGAR EEL. 123

l>een boiled long enough to concentrate it one third is much le--
toxic than ordinary di-iilk-d water. For this and other reasons
he concluded that the chid toxic agent in such waters is the
ammonia \\ hich they hold in. solution. In fact he found that the
larva- often de\ el< .|)i-d better in ammonia-free artificial sea \\ater
i han in normal sea \\ater, a result that may \x.- due, as he sui^e-t -.
to the presence »t' ammonia in natural sea water.

About the same time Bullot1 puMi-hed a thorough in\ v-tigation
of ihe toxicity of di-tilled water tor the fresh \\ater ('mmmarns
and found that the \\aU-r di-tilled OVi 3S \\ith all the pre-

caution employed in a phy-ical chcmi-try laboratory \vas toxic
for tin- form allhou-h t«i a !• ml than \\atrr distilled over

coppi-r. lit- found, ho\\r\rr, that the ])re-nice of \a< I in Mich
.1 dilution ,i- i coii.-i-ni ralioil ot o.ooooS.\' enable^

ihf Gant1 to !i\c indflmiicK in >uch a medium. I.oi-|,-'

suggest - thai, in -ucli a • I • •;• % m e "f a trace "f \a("l in

the di-tilled water po--iM\ ves tin- mcm!>ranc ln'ttrr or

maintain- U-itrr tl .cti\il\ of the cell-. >o that the

animal can I .«• !;. -<-d troiu tin - o( \\alcr \\hich dillu-e- into

it."

The :<n>np "f \i-m itodes Is noted !'"' the e\lraordinar\ re-i-t-
anCC uhich the cnticli to external media. The t oiumon

vinegar or ]• nilliiln aceti, i- \\ell kno\\n to occur

iiormalK in \\eak vinegar, although acetic acid i-, to IIK^I or.^an-

isms a rapidl) actin »n. Anguillula aceti has been observed

in our lalioralo!\ to live m.ire than 24 hour- in Tell\ e-nick\ '-
lixin:; llnid and three or four hour- in < ,il-on'-> llnid. The cuticle
of ihi- \\orm miulil theii-!'-re I «• i-x|iected to I.e <|uile unallecled
1)\ Mich ion- as .-idinar\ di-tllled uater max contain. It i- of
inlere-l to iinte in tin- ...nnectioii that l)e\aine" in hi- lijolo-ic.il

ilixe-li^alioll ,,t the vinegar eel o|.-er\eil thai the \\onil- li\e

.it most l>ui eight in di-tilled water. Thi- effe< i mrjn In-

due to the lack of the acelic acid uhich ordinarily form- a pan
of their normal en\ ironmcnt or to the din. t effect of the uater
it -elf. It ua- --.on found thai although ihe \\orms li\e I.e-t in

• Bullol

; i : M.uiri ." |>.

•»;;•/ A', n,l.. !.<•!. it

Uj JAMES F. AUr.oii \NU liTIIEL L. Kim.\KI».

an environment of weak vinegar, it i> the sugar rather than the
acetic acid which appears to lie the essential element of the
medium.

With the hope of anaK^ing the relations of this organism to
its environment, a number ol lines ot investigation have been
taken up in connection with tin- reactions of the nematodes
to various media and to different food substances. The present
paper deals only with the eflect produced on the worms by
distilled water. The worms used in the>e experiments were
found in rather -tale weak vinegar of an acidity of <S 10 N. No
difficulty was experienced in breeding them abundantly in the
laboratoi y both in this medium and in cider. Two kinds of dis-
; illed water were employed. One of these was such as is u>ed in
the ordinary work of the laboratory. This was distilled in an
automatic copper still and will be designated "A." The- >econd
water, which will be designated " B." was obtained by distilling
the first kind over potassium dichromate and sulphuric acid, con-
den-Miig in block tin out of cc.ntact with the air and redistilling
over barium hydrate. The first six tenths of this distillate was
rejected and the remainder gave an extremely pure sample,
CO-j-frec and showing no ammonia reaction i Noler test) even
after the lapse ot twelve hours. This water when tested in a
conductivity apparatus of the department of physical chemistry
gave no measurable conductivity with a resistance of 20,000
ohms and may be considered to be practically free from electro-
lytes. It was kept in well steamed, hard glass bottles. In each
experiment every precaution was taken to remove all traces of
the medium in which the worms were living. The \\omis were
washed 6 to 8 times with distilled \\ater in a centrifuge1, and in tin-
cases where the distilled water "B" was used, two or three times
more in this. They \\ere then allo\\r<| to remain in the "B"
water lor about 24 hours to injure a thorough ringing. Finally
they were again washed in pure- \\ater previous to their in-eriion
in the new medium. The bottles and tot tube-- ii-M'd to contain
the worms in the first two sets of experiments were ol ordinary
soft glass and in the latter ones, .^< liolt (ind (lamsscu, Jena resist-
ance glass was employed. The --e containers \\ere thoroughly
soaked in chromic acid, and previous to n^e were repeatedly

EFFE< I "l DISTILLED WATER ON 1111 V1NEGAK EEL. 125

rin-ed in distilled water. They \vciv then -teamed for .1 greater
or les^ length of time under pre— lire of 150 pound-. An attempt
wa- made to put .1- nearly as po— ible the -ame number of
worms in each culture tube but owing la their minute size it
\va- of cour-e impo— .ible to do more than approximate the
number.

Contrary to expectation- the \\onn- lived much longer in the
onlinary laboratory water "A." pre-umably full of copper and
ammonia than in that distilled over potassium dichromate and
free from CO Ml copper and other ions. In "A" and -ol t
glass tube- they li\.-d +\ days, in "H" \vater and -oft tube-
they lived -ixteen da\ s ami in "B" water and Jena in-oluble
tube- the\ li\ed but si\ da\-. The-e re-ult- uollld seem to
indicate that the pi of the elect rolyte- in ordinary dist illed

water and of the ion- di--ol\ed from the :J.i — prolonged the
life of the \\orm- Be< • -f t he fai t t hat t he | Hirer t he water

the .Kent po\\er il i- -till \» be determined

tthether tin- . i- due !<» the -ub-tance- di— »l\ed from the

•^1 tsa or to the copper and other ion- in the di-tilled water it-elf.

Iniht writh " B " water and Jena j ;n \\hi.-li the \\orms

li\ed but -i\ da\-. then- \\eif no impnritie- in the \\ater and

prat ticall> no solvents i mm the glass, ^o \\e are ton-eti <<•
conclude that the toxic chara< the water it -elf was the cause

,,1 ihe de.dh ol the \\orm-. I'lij- i- boriu- out b\ « •' >-er\ at ions
on i he beh a \ j. .r "1 t he \\orin- umler -uch condition-. The outer
internment of tin- neinatode in contra-t to it- normal hardne--
and re-i-tam f. app. .in d to become verj visCOUS after a -ojoiirn in

pure \\ait i. \\henan\ obstacle was encountered in -\\immin^.

the -itle of the tube or the -nrface of t he \\ a 1 1 i , <>r oi her \\ orm-.
the organi-m- -tuck fast. Tin- alteration of the natural condi-
tion of the CUtide was SO marked that after a time the \\ornis
bunched lo-ether in masses, unable to free t hcin-elve- or el-t
coining in contact \\ith the -nrl'ace ..f the water, the\ hnn^
-ll-pciided. 'rili-eliect \\ollltl -eem l«> bear out Loeb's Suggestion

(|ut»ted above. That there i- a market! taking up of water on the
part t.f the oruani-m seems quite e\i<lent. The l>o<|\ becomes
-ttoiien and the \\orm- 1. -e their elasticity, becoming more or
It -- lim'd through turgor and swimming with the bodj almo-t in

126 JAMES I. ABBOTT ANM> I IIII'.L L. UK HARD-.

a straight line- without the characteristic twitching of the cud-.
The worm- ^n>\\ more and more impotent until at la-t iu-
Stead of -wimminii directly towards the surface as i- usually the
case, they swim in a hapha/ard m.inucr and in spite of instinctive
efforts to reach the top of the water are unable to advance more
than an inch or two, falling back towards tin- bottom with
ever\ effort . This may be due to decrea-cd vitalit y but max also
be due t<> a greater \\ei.nht oil a<-count of the imbibition of water.
Repetitions ol the.-c experiments have confirmed the results in
every instance. Experiments have been undertaken to determine
what ions control or effect the permeability of the cuticle to
the water.

Si \t\t \KV.

1. Anguillula <u~cti succumbs in six days to very pure water
free from electrolytes when-a- in ordinary distilled water it \\ ill live
for much longer periods.

2. The toxic qualities of the pure water appear to reside in
its solvent effect on the normally resistant cuticle of the worm
which is rendered much more permeable to water.

3. Death is accompanied by physical manifestations suggestive
of great imbibition of water by the tissues.

4. The presence of electrolytes in the medium, derived either
from solution of the glass culture tube or carried by "impure"
samples of distilled water, prolongs the life of the organisms in
<|iicMion, apparently by preserving the character of the membrane
so that the tissues will not imbibe water.

XXI. August, /y//. M

BIOLOGICAL BULLETIN

i ' GICA1 51 CCESSK >\.
I ! POM FISHI s.

!•; -H!-i

I. I
1 1

I . ' i j s

III

IV h •

. . .
V I

VI •
V 1 1 Summan

V! 1 1

I. 1\ I l<" -Hi i I I- >\.

Ill ilu- lir-i p.iprr nf tin'- ^ i l"i-ln •- .ind I'h\-in-

ihi( \n.il\-i-" . \\c (ininitd mil tli 'ii^ l"i»r uixlcri.ikin^

thi- investigation and ed iln- inu-p" ••'iini! \\liidi tlu-
\\ru-k h.i- Tlicrc \\.- were I-.>IU-.TMC<I \\iih ilu- \.iliu-

nl" ilu- |)|-iiici|)!i-- ••!' pliN- •!)>• in a nu-tlunl of l« ic.iiin- ilu-

.iniin.il in ilu- t n\ inuiiiii-in .uul of (k-irrminin^ sniiu-lliin;; <•!' ii-
character as a whole. Here we are to di-«-u>s ilu- \ .ihu- «>f pi. mi
-11. . i --inn in .1 -imil.ir \\.t\- .uul to h.i\v .1 1-i-iu-r opportunity i"
-hn\\ ilu- \.ilidii\- <•! ilu- principlr ol succession as .ipplird to
.iiiiin.il-. l-'iirilu-rninn-. .1- \vr pointi-d mil in the O|!UT p.i;

•

logical -ii' »n i- to be differentiated fn'm -colo-ii .,1 -uc-

128 VICTOR E. SHELFORD.

MOII. Ecolojiif.il Micce— ion is succession of ecological types
regardless of species, while geological succession is the succession
of species. Tin- data presented here affords an excellent op-
portunity to bring out the differences and relations of these two
types of succession.

\Ve noted also in the first paper that the first recognition of
plant and animal succession came with the development of
genetic physiography. It was mainly the successsion which ac-
companies physiographic change. Cowles ('01) also clearly
recogni/ed succession due to the action of plants themselves.
Tin- l.n ler idea has been elaborated by Clements ('05) and essen-
tially demonstrated by Schantz ('06) and Dacknowski ('08).
Animals must obviously play an important role in this type of
succe— ion. |,ut unfortunately this has not been investigated.

The succession with which we will deal in this paper is that
n--ulting fron the action of organisms on their own environment.
For all practical purposes the area selected for this study has
been in a condition of physiographic stability for a considerable
period. The selection and analysis of the place of study is the
most important step in the whole investigation. Indeed there
are only a few suitable localities in North America.

II. AREA OF STUDY.

Owing to the fact that succession is always either dependent
upon, or modified by changes in conditions, a correct interpreta-
tion of this phenomenon depends largely upon accurate kno\\ ledge
of the area under consideration.

I. Location and General Character. — The ponds which are tin-
subject of this study lie in the sand area at the south end of Lake
Michigan, within the corporate limits of the city ot (iary, I ml.
They maybe reached from stations known as Pine, Buffing ton, or
Clark Junction. This locality is characterized by a large series
of sand ridges, for the most part nearly parallel with the lake shore
(Map I.). Their average width is about 100 feet. They are M-pa-
rated by pond- \\liich are someuhai narrower (Map II.. p. 131).
Most of these ponds are several miles long. The\ \ar\ in depth
during the spring high water, from a few inches to four or live
feet. They describe an an- somewhat longer than the lake shore
(Map I.), and are farthest from it about midwas of their lengths.

ECOLOGICAL SUCCESSION.

129

- - -
-

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0

3 1 *
3i

- =

r •=

- - Z

1 :.

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Ij

'<* —

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=

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I in VICTOR E. SHELFORD.

Originally, there were probably a number of outlets to the
;em of ponds which were joined to one another by the-e
outlets and through various lo\v pl.«-t - in the sand rid^e-. In
the area of our concentrated study then' is an outlet (Map II
which has served to connect all the younger por.ds in an intimate
fashion.

The building of sew< -ociatcd with the growth of northern

Indiana towns (\Vhiting, Ka-.t Chicago, Hammond and Gary)
has drained la rue portions of the pond-, u hile roads and railroad-
have isolated other portions.

2. Origin of the Ponds. — During the linal retreat of the North
Americ;m ice sheet, its Lake Michigan lobe stood for a time with
-outhern end at the cresl of the Valparaiso moraine which
live.- concentrically around the -oulhern end of Lake Michigan.
When the ice retreated from this position, water occupied the
ce between this lobe of the sheet and the moraine. Thiswa-
Lake Chicago, the forerunner of Lake Michigan. Affr having
stood respectively 60 feet, 40 feet and 20 feet above its present
level, long enough to deposit conspicuous beaches, it took up a
position at a 12-foot level. The waters appear to have fallen
dually Irom this level. At present one or two ridges and
depressions similar to those found above the water on old I
Chicago plain, are below the surface of the water along the shore
at the south end of Lake Michigan. The retr< a1 of the water-
has evidently exposed such ridges as fast as they were formed.
This has left a series of parallel ridges and ponds urnui^ed in tin-
order of their age— the oldest furthest from the hike, the youngest
nearest the luke. '

These ponds are not all of the same size. The largest ones
were selected lor study and will be rel'-rrcd to in the paper by
number as the entire series is counted inland from the lake shore.
There are between seventy-five and ninety of these ponds or
depre--ion- between the lake shore and the Jo-foot beach level.
This is the maximum number. Map l.-h<>\\sihat as \\epass in
cither direction from the area "I" stud their numlur decreases.

1 For a trr.it iin-iit Mt iiii- -ul.; cited in the bibliography i>ui nol

specifically n-i<-n«-cl tn ln-if. l'i i1 ; < - i!i-lnn\ ti'lU im- tli:it tli'

question conccniinu tin- n-l;iti\ •• m p!i\ -i'>^i.i|iliii r\ -ji

ECOLOGICAL SUCCE--H >\ .

•J. M

- —
— — -

r- T •=: "

= r
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VICTOR E. SHELFORD.

M.ip II. shows the relations of the fii>t t\\enty-four of tlu>e ponds.
The recent changes of this smaller area are indicated <>n this map
by the dates of the building of the various road and railroad
grades.

III. THE DATA.

All methods <>t collecting have been employed. The dip net
has been found most elTecthe, but the drought of 1908 and drain-
ing b> sewers have yielded cruical results in Ponds i , the outlet .
Pond 5</, .v , i-|</ .ind ~h.

\\'ith tin- exception of Anna caha, tin- records are for adults,
or for young where the pond is known to be isolated, and the
presence of adults is therefore necessarily implied.

The record of Ameiurns melas in Pond 56 is based upon the
presence of young and adults; identification of the adults
depended upon identification of the young, because the adults
\\ere too badly macerated to make identification practicable.

i. nistrihntion of the Species of Fish. — The distribution of the
fish in the various isolated parts of the ponds is of much impor-
tance. Turning to Map II., we note that the extension of the
ponds to the east and west is not shown. Toward the east Ponds
i to 17 become shallow near the Gary Steel Plant and are not-
connected with other bodies of water. Parts of Ponds I to 7
have been directly connected with the lake by the outlet (Map
II.) until within the past few years. Excepting in the case of
pond 7, other connections with the lake, to the nort Invest of the
area shown on our map, have probably been closed for a much
longer period. \Ye may concern ourselves largely with the
parts of the ponds to the east of the outlet, \\hich have been
isolated by railroad grades, and with their relation to the outlet.

Table I., taken with Map II., presents the exact data on the
distribution of the fish in the various parts of the ponds. All
fish nomenclature is after Forbes and Richardson, '08.

In Table I. we note first that the large-mouthed black bass,
the suntishrx. the pumpkin seed, and tin- warmouth, are all con-
fined to the outermost pond, or Pond I. The pa^a.^e Irom pond
l to the outlet and to pond 5 was open until 1906. The study
began in 1907. In the autumn of n(oS the drought reduced the
water in the outlet to a minimum, but none of the-r -pecie> \\a>

ECOLOGICAL SUCCESS!* >N .

133

found there, though the passage between the outlet and pond I
had been open two \ears before. \Yhy have tin- ti-h vacated or
avoided the outlet? One can only suggest a behavior reaction
is the cause.

TABLE I.

I >IMKII'.I I I' >v- i>K THE FlMI.

The letter-; and number- at tin- In -ad- «il the column- reier t. > tin- vai i< ni" i-' ilated
part i- and. i-xo-ptini; <> which n-ii-i- tn tin- "iitl'-t. may l>e Incated an Map

II. Tin- la-t column indi. at'-- tin- occurrence <>t fish in the older ponds <>t the
-'•ri'--. uhiili ari- iii.t imjud "H thi- map; the nuiii i i« the number of

th<- p«.iifl in whirh the fi-h \\i-n- lOund. ' indicate" an incmnplete identiticatinn.

1

-

111. ii k !...--

*

Bin-- '.'ill

*

Hlii<

*

:(,

Pumpkin

\ . -Ili .u pi ii !.

( Illlli -Hi ke[

• d l.iilll,
I .id;

•
*
*
* *
* *

f

li KCI '-1
niiiiin iv

• » * * *

*

< .. ild> '

Yellow luilllli .id

HI. ii k luilllii -.1 !

* * * *

*
•

•

»

'I'lit- IM-I-I-II i- di-iribinril ill a -till nmn- [n-culiai way. Ii is
I'liiind in I'-'iiil- i and 5i.bin ;/"/ in tin- (.mid i, r in r.nnl- 5</
and .V-. 5-.' ainl 5 •! \\itli tin- outlet until [907

but 5- ha- |»robabl\ been separated -imv I s.^ I , \\heii the I. S. \
M . S. \\ \\ \\.i- built . I ha\ e no »-\ ideiiee I h.it .1 >luire \\.i- CVCf
tuaintaineil under llii- railio.ul in I'oiid 5. The perrh i- then
di-tributed in -u« h a \\.i\ .1- to necessitate the eonclu-ioii, cither
that it \\.i- artitieJ.tlh int roduei-d or that il \\.i- once in the
outlet and in I'ond 5-.' and .V'. because ihe-e make the only
pa-^agi- to I'ond ,v u here it uo\v oi-cui

The chub Mirker i> in all the pond- up to ~: . excepting 5«;
and ~ii. The passage to. from, and through 1'ond 5..'. and oilier
point- \\liere the rhub -ueker occurs, \\a- open until 11)07 .mil
the erueial rolleetin^ \\a~- done in [908. Il \\ill be noted ihat

134 VICTOR E. SHI I.I i >kl>.

the distribution of tin- chub Mickt r in Pond- ~a and ~'< \^ \ ntircly
similar to the distribution of the perch in Ponds 50 and 6 and 5 .

The chub sucker is not in tin- part of tin- Ion- Pond 7 which li. -
been recently connect. -d with tin- outlet.

It will be noted al>o that tin.- tadpole cat and the -potted bull
head have not been taken in 5</. The yellow bullhead ha-dum
taken from ~a but once, but we may expect that ii i- a re-id, nt
of the locality. One would expect to tind it in Ponds 56 and 5c.

With the exci ption of Pond- 14(1 and 14/1. there arc- no non--
worthy peculiarities. The study was begun in 1411. Thi- was
partially drained in the winter of i<n>X <> and it was therefore
necessary to turn attention to i _}./;. This was probably partially
drained previous to i8<>2 by the East Chicago shin canal, but
again renewed through a dam made by the building of tin- YVaba-h
railroad grade between the point of draining and the part
pond studied about 1892. This probably accounts for the pres-
ence of only a single species in Pond 14/7. A single juvenile !>'
spotted sunfish was found in Pond 56 which is directly conm-cu d
with the Calumet River where it is common.

2. Ecological Age of Ponds. — The ecological age of the pond:*
is determined by an inspection of (a) the amount of bare bot torn.
(b) the amount and kind of vegetation, and (r) the amount of
humus. It is a well-established fact that an entirely new pond
(in the matter of recent separation from a lake, like Lake Michi-
gan) has little vegetation and very little or no humus. Both
vegetation and humus come only with ag< . Ag< -determination
is so simple that no difficulty usually is experienced by •
trained in plant ecology, in arranging a »l pond-, in the

order of their ecological age. In the mailer of the kind of
vegetation we have had the advice of Dr. II. ('. ("owl

Pond i is the youngest, because it has the- kind of vegetation
that grows in young ponds, more bai I bottom, h ,.-i humu>,

and least vegetation. For similar n I'on-i- ->•'• and ,v -land

second in the matter of age. Because of lnini.ui interfi rence,
which has kept the vegetation down in I'ond %v, ii i^ probabh-
ecologically >'ounger than 5/7. The oui! orobabK- inter-

mediate between 5 aiul 7. Pond '..ml next, but with-

out anv dilTei'ence as far as on< •

1 ( ci.nGK AL >rrCE--I< X.

135

From the -landpoint of bare bottom and hunni- I'oiid ?<i \>

very much advanced. I'. ;ed mar ihe outlet, i; ha-

!•< rome filled \\ith (k-<\i\-:: n»n and in- IHI-. now

Toyed, indicau-d an adxanr. d sfc 'inj\iral >lc with 14.

tt differs from 14 in ] •_• qualitatively "\ -tation

[ some other eh; - of youth. Tin- di!"tVivn<-t - l>i.t \\rfii

1411 and i4//ha\' idy been discussed.

In T.-'.lili- 1 1. • ponds are arrangi in order

•d th«- di 'ril-ution of ti-h i- shov n.

I!.

i

1

1
I

* * *

\\ hen the ponds an ! age,

i noti. the distribution of tlu- li-h. and \\t

al on.

I\'. l\ 1 1 '
\\ V li. 'ii of 1 i hat

ill'' ]>|. :.[l(d \\illi A the

of (In- p. : ' i nd (B) tl of 'inn lhal

clianiii 1- ha\ e been I

h i- a \\i-ll-kno\\ii |iond> till with plant dctritu^.

\\"ilh i In- lillin- "I ponds, tlu rmidii i« -n- ••! | ; . 1\

in a drlinitr diivr;i»n. I ' \\hich ]•• in the >anu-

136 VICTOR E. SHELFORD.

general ph\ -iographic relations during .1 o m-idcr.il >le period of
time, there i- .1 succession of conditions due to the accumulation
of detritus just as there is a succession <it conditions in a si ; earn
due to phx sjographic changes.

Since the channel^ of c< >mmtmicali< m between the different
ponds have been open until recently, the present arrangement
of the species of tish is \ ery probably a helnreior adjustment to the
changed and changing conditions just so far as barriers have
permitted.

\\ •• see from the arrangement and mode of origin of the ponds
that our oldest pond — number 14. Map I., and Fig. I, was once
in tin- -a me relation to the lake as Pond I is now. At such a time
it was in a condition similar to that of the present Pond I.
Ponds 5 and 7 are intermediate in conditions between Ponds i
and 14. We ha\e the same general basis for the discussion of
ecological succession as in the streams. The changes in these
ponds have depended mainly upon physiographic stability within
each pond, rather than upon physiographic changes, and have
been due to tin- action of the nryuiisnis present on their o\cn cm-iron-
vnent.1 This is true because after a given pond is once separated
from the lake (Fig. i) the changes due to the organisms go on
\\ithotit regard to further separation and the lowering of the
lake lexvl. Fvidently the level of the water has remained much
the same in the ponds after their separation from the lake proper
regardless (,| the lowering of the lake level (Fig. l).

The method of deducing succession herein employed is similar
to that used in the case of the streams. The easily obsen. able
Facl thai animals occupying similar conditions are ecologicallv
similar (i. e., similar in habits and some main features of their
phx sjologv of external relations) is used as a starting point, and
the conclusions drawn are to ihe effecl th.it \\hen the older
habit a i \\.i-in the stage of a younger habitat, it \\.is occupied by
fishes i-inli>"t/iil/y similar to those now in the younger halntat.
Whether lhe\ \\ere the same or different species is often of little
import. nice lo ecological succession. With this simple explana-
tion as a background, and \\ith the use ot Figure i, xve will

1 The- change--; \vlii. 1 1 are caused \<\ I li>- tilling • it i hr |» >n>l \\ it li plant iii.i 1 1 -i i;i I
are pli\ -iMjuplii. , hut tin' hint . »ecl is the more im|m[ t.mt . and \M- m. iv

i-;s the < 1 I hy hiuldvi, al I. .1

ECOLOGICAL SUCCESSION.

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state, as well as practicable, tin- succession in these ponds with
particular reference to fish.

I. Statement of I:.colo':icdl Succession.- A Mateim-nt of sin
sion can In- best made in the form of a hi-tory of Pond 14 (Fig. i ,
From our knowing- of" ilu- origin of tin- pond- and ihcir present
topography, it is reasonably certain th.i! ill. n- was < lime when
I'oiid 14 was nio'-f clost !\- associated with the lake than Pond i
'•in Fk. i. hypothetical stai;e B). Ai such a time
I'oiid 14 contained le-s vegetation than we now find in Pond I.
For a kno\\lrd-e of the ecological character of tin- fish which
inhabit such pond-, \\ i have collected from a pond of tin same
origin as tho-M- made' tin subject of tin- present study. This
one has mainiaiiied a close connection with Lake Michigan and

eives the waves of the lake during the spriiiL' and winter
-tornis. ll flows into the- lake during every hi-hu ater period.
This pond is at Beach Sta.tion, four miles north of \Yaukcgan. 111.
Nf( ar its outer end it presents a clear bottom of sand and gravel,
little vegetation and no humus. In this outer portion we have
collected the pike (Esox Inciiix'i, which prefers clear, clean, cool
water (Forbes and Kichardson, '08) ; the red horse ( Moxostoma
(iitri'oliihi), which dies quickly in the aquarium if the \\ater i-- the
least impure and Miccumbs to impure conditions in its natixe
wai. •- ! orbes and Richardson, '08); Xntrnpix cdyit^i, coninu n
in < "a\ -u^a Lake and the lower course- of its tributaries i Keed and
\\ii-ht, '09). As compared with Illinois waters, the-.- streams
\\ould be counted clear. \Yealso found Xolrnpis cornntus, which
shows a marked preference fcr clear waters (Forbes and Kichanl-

-oll, 'O?

\Yhcn Pond 14 was in a very earl) stage (hypothetical stagi B,

Fig. I) it must ha\'e been occupied b\ fishe- which \\eri eco-
logically similar to the red hors,-, the pike, the < tyuga minnou,
and the common shiner. This i- a comimmit\ of species which
may be characteri/ed as requiring clear, ck-an waters, clean
bare bottom (especially during ilu- breeding season), and littk-
vegetation. Such fishes may be designated as pioneer ecological
types. To this group might be added such fishes as the common
perch, which is hardy and lives in a \\id< rang< ol conditio
Following the history of Pond 1.1 furiln r, \\ i note thai as the

.

fetation -n-w, humus accumulated in tin- deeper part-, and
forms ecologically similar to the black ba>s, the blue ^ill, the
pumpkin-' :id the white crappie {Pomoxis annularis) whicli

i- widely distributed i Forbc- and Kichaid-on. '08 . but which
Meek and Hildi brand f'loi record only from streams and pond-
which are ecologically youn.u. mu-t have ma.de their appearance,
il any such fi-he- wen All but one of thc-c:-pt

were taken from the pond at Beach but chiefly from the older
parts. They were IK. i dearly -eparated from the other sped
but condition- \ d and no barriers prc-ent. \\'e \\oiild

expect the ai i .f tin- pioneer ecological P uch

'he pike and tin nd horse, from -uch a community.

\Yhen the condition- in 1'ond 14 \\hich made the habitat

•r th:- la-t meiiiii>iii-d d-h community had pro-re— rd

a little InrthiT, nil ///• ust //</, ntly

:/>itdt :- 'niiK-il > the

(>H" ;rcd

'i as tin- blue -p. .tied -unli-h,

the little i dub sucker, the warmouth bass, the spotted

bullhead ami imk' mud of the deeper portions a

ft u fishes M\ -imilar to miicl minno\\- ami tadp"|.

I'hi- i- lh<- coiid; our 1'oml I at pn -nit a -hallou pond

\\ith « imulaiion- of huinn- in i he deeper part-,

and much d \\ iili ml other plain-.

In this Connection it -lloiiM IK- noted tha' \b « k ami I lildebi.illd

I" record and top mimiou [900 from 1'ond I

M.ip II ; 'in the\ are m • Imu'er found thei

It -hoiild tl< nibend th it the part- of a pond which arc

-itnated a- to i ||ow the accumillatioii of plain

detritu- \\ill be lir-t to become unfa\or.ible to pioneer li-ln -.

while the parts which are swept clean by wave action remain in a
d condition much longer perio<l of time. Stages in

pond development -itch a- the prc-ciH 1'ond 1 are the most
coni],l( \ of all, and -.re more like the larger lake-, ^uch staj

in a coiiiliiioii to >up|io greater di\erM't\- of ecolo-i.al

type- than the latir - \\liile >till retaining a part of ii-

pioiicer li-he-, 1'ond l -nppori- the forms which belong to
older condition- (mud minno\\ ami tadpole cat).

Ylt I"R E. SHELFORD.

Up to such a stage, Pond 14 must have become from the fir-t
more and more favorable to di\cr-itv of ecological types, and
accordingly possessed .it Midi .1 time its greatest number of species
of fish. When Pond 14 was at a stage comparable to the present
Pond i, tlif li-li community present and all the other organisms
"ciatcd \\ith it, so acied on their environment (just as they
are acting on their environment in Pond I at present) as to
make the habitat less favorable to the fish of the earlier pond
M.I-CS, and more and more favorable for those dependent upon
and tolerating deii^- vegetation, absence of bare bottom, and
lower oxygen content. As a result of this action of the biota on
its eii\ ironmeiit, tlu- fishes of ecological constitution similar to
that of the sunfishes and basses now present in Pond I, disap-
peared either by emigration or death. In the absence of these,
and in the more favorable conditions of competition and denser
vegetation, fish, such as the golden shiner, were able to find a
suitable habitat. At such a stage Pond 14 possessed a fish
community of the ecological character of that now found in
1 '< >nd $c. Here the i >ioneer element is reduced to a single species,
tin perch, which is very hardy i llankinson, '07).

The same process continued and caused the disappearance of
the perch and like ecological types. A fish community ecologically
like that now in Pond 56 (Table I., column 3) then existed. The
absenceof the perch in 5/>and its presence in 5c may be explained,
judging from the general habits of the perch (Forbes and Rich-
ardson, '08), by the fact that neither pond appears to be favor-
able for perch and they have been able to move out of Pond 56,
but not out of Pond 5*;. An experimental comparison of tin-
behavior of perch from I'ond 5c and other perch habit. its would
have an important bearing on our problem.

The fish community of Pond 56 is made up of the chub sucker
and the golden shiner, which are abundant, the spot ted bullhead.
the tadpole cat and the mud minnow. The spotted bullhe.nl
is the only one known to use bare bottom for nesting. There i-
only a little bare bottom in Pond 56. Tin- spotted bullhead
usually builds its nest under «:o\er ( Kycleslivmer, '<>! ).

When such a fish community occupied Pond 14, the biota
present gradually changed its <.\\n environmental conditions as

ECOLOGICAL SUCCE>-I<>\ 14!

the former stages had done. The nature of the changes are
evident now when we can compare Ponds ,v and 56 with Ponds
"ja and 76. In ponds ja and ~l> we find water lilies and bladder-
wort in abundance. These are barely pn-eiit in $b and 5f.
Ponds 7a and 76 have much le-- bare bottom than 5 A and $c.

! I ;il I ! • ..| lli.- .||,,u-.;hi I,,

III- 1 uitli \\.ii. •[ lull <ini<l .1

lid I"- ii' '
This J-, pivM-nt in l"a\i.|-al'lr -it uatii >n- \«T\ near tin- water's

edj

I'lir \\A\ c(imiiiiinii\ 0 iii.ulc up ul" tin- -aim- -pcrir- as

the oimmimilN \\illi tin- addition "I th<- Mack liiillln-.id.

~n lack> llu- cluili Mickcr, .UK! ln-n- .is h.i^ liccii n.ucd ihc chuh
sucker lic.ir- tin- -.inn- rt-lati"ii t<» tin- 1 \\ < > |>arts of I'ond 7 as ihc
pcrch does to I'.. n.1 | 5 It is r\idt-iit that it mo\»-d .nil

ol llu- pond from \\liicli a cliamu-l of c\it lias IM-CII opt-n.

The tirst -icp in tlu- transformation of ilu- tish comimmiiv
of I'oiid 14. \\hicli was ecologicall} -imilar to ih.- pn-s.-nt -/>,
into tin- iu-\i l.i icr • . was i In- loss "i -neh ecological i \ pes as

142

YI< IMK

5HELFORD.

dull) sucker and the addition of -nch .1- ilu- black bullhead.
Tin- latter is a well-known "mud-lover" which inhabit^ t In-
oldest Stiller- 'it pond succession. A- the condition^ which
Organisms produced in> d in intensity in the din ciioiis indi-

d, lln < -illy similar to 'In- tadpole cat and the

-pot ted 1 in! Ilu .id disappeared, and with them probably tin- golden
-hiiu-r aUo. \\"iih th< e we have tin- fKh c-onimunii \

which \\as in Pond I i>i lid'orc draining. h i> illustrated by
Pond 5'/ also, in which the plant and animal d'-trilu> ha cted

l-i'. 5. sii.. \\in-j I'.ni'l i i .it moderate !i>\\- \\-ac -t. In contrast wiili i. \v<-
•li.ii it i- clioki-'l with v^'-uiti'Ci an 1 th- margin ni-riipi;-.! l>\- ^Inuli--
bulrushes. etc.

bee; i u -.e it is located al the millet \\here the channel becon
shallower and nammer. l;.co!oLical 1 \pe-.siich as thepickenl,
the black bullhead, and the mud minnow make up this community
in Pond I l. They n-nallv continue until the pond becomes
temporary when ilu v are dt •-iro\-t-d b\ dr\'in.n'.

2. The Fll! !<>•<' of Ilif font!. !'"coloi;y is one of the few biological
sciences in which prediction U po-sible. I shall \cntnre lo predict
the fate, in a state of nature, oi a pond like Pond i^aivasa\ the

ECOLOGICAL SUCCESSION.

lit where the study was first begun, before the draining took
place.

Had Pond 140 not been disturbed by man, it would have
• ontinued to fill with humus. Its center would have o>me to be
occupied by the cat tail-, bulrushes, Proserpinaca, and Equisetum,

all of which invade trom the sides. \Yhen -ueh a Mage wa- reached
ii \\oiild l»e >ubject i.. almost complete drying in extreme droughts

and the li-h \\oiild be eliminated in the order of their ability
10 \\ith-tand draught, and in relation to accidents of distribution
in part> of t : 1 drving t<. various

In ^cpicinber. Ioo>. \\ • able to obtain e\ ideiice on this

point from some of the older pond-. A u'»iip of black bullhead-,

! and mud minnow- were found in the lowest part

•ne of th, ms of p..nd 5*. The pickerel Were dead and

badl\ ed. The bill! land jll-l bcgiimilK

but ihe mudmimio\\-. fully a 1-u-hcl of them, were -till
ali\e ahliough \\ithoul v I hi- lot included -mne of the

large-t indi\ idlial- o.|i<

\\'h« : I '• -nd I ) -h content Would

becom( probl. :ent upon the distribution of

the dilten-iH .;iid shalloue;- parl-

o| the |)ond. and tin - relat* d lo the

the d: I \ j.lence for this i- seen in

Polid I j . v i- the onl\ -111

since a pmbabli 1 df\ili^ of the |)ond.

Indeed. Ue II I he |iredictio|| one -tep farther. \Yllell

-in h a pond become- of tin- -or! the plant- that take hold of
it are i \*< \\ liich till the -oil \\ iih ro.

and form hummock-. I: i'ole to them, -uch pond- be-

rome the breeding place of tl ii-h. During the drought »i

i MO- I found a -chool «.f half ^ro\\ n dog li-h in -uch a pond, which
an-\\ered the description of the la\oHtc breeding place of the
do^ ti-h ,. a 1>\ Kei^hard [*O2 , >uch - thi- \\ill

be folloued b\- the inv.i-ioii of -hrub> and ilu- final de-tructioii
of the pond a- an aquatic habitat.

\\ e ha\e in |ioiid- a p- :\> change in the <-onditioii- and

a p- jive change in the co-logical and |)h\>i< -logical character

of the ti-h comimmitie-. and a succession or evolution of fi^h

144 VICTOR E. SHELFORD.

communities through emigration, death, immigration. ,nid tin-
modification of more* by external -timuli.

\". ECOLOGICAL Succi — n>\ AND Si CCESSION OF SPI « n -.

The difference bet \\een ecological and geological succr—inn
were suggested in my preceding paper. \\V noted that eco-
logical succession deals with tin- succession of ecological types.
We noted also that geological succession N due-, in ihe main, to
the death of a given set of species and the evolution of new ones
throughout geological time. While this is true of the broader
aspects, in the more detailed cases and especially in dealing with
recent post-glacial fossils, the palaeontologist often encounters a
vertical succession of fossils which have been left behind by
the migrations of a succession of species over the locality of fo--
silization (Warming, '09, p. 362, Adams, '05 and '09, Sharff, 07,
esp. Chap. IX. and citations) . Palaeontologists may also encounter
vertical succession of fossils in situations \vhere such succession
as we have been describing has taken place.

i. Vertical Succession of Fossils. — ^Steenstrup ('41) found from
the study of fossils in moors that one kind of vegetation suc-
ceeded another. Various other workers (Andersson, '97) have
found similar arrangements. In the case before us the fossiliza-
tion of species would give a vertical succession of fossils.

Turning again to the diagram, we note that the skeletal parts
of any fish in the earliest stages indicated by hypothetical stage
B might have been preserved as fossils. The accumulations of
humus which lead up to stage I would have covered the fossils
of stage B, the fish of stage B would be present, if at all, as
fossils at the bottom of the pond.

Likewise, the accumulations of humus which led to stage 5
covered those skeletal parts of the fish of stage I, and the fish
of stage I should be found as fossils overlying those of stage B
and underlying those of stage 5. Again, the accumulations of
humus which led up to stage 7 co\ i-n-d fossils which were present
and added at stage 5, and the fish preser\ed .1- fossils from sta-e
5 would lie above all those preserved as fo-^iU lK»m the younj
stages and below those of stage 7. Ko-.-iU from stage 14 and the
intermediate stages would lie at the top of the Aeries and above
those of stage 7.

ECOLOGICAL SUCCE--1"\. 145

However, no >uch arrangement of fo»il- has been found in
the^e pond-. No attempt to ascertain whether or not they are
present ha- been made.

2. Relntion to Clinidtic L'hun^c^. — As ha- l>cen indicated, sued
.-ion is dm- to changes in condition- which make- it impo--il)le
lor a gi\en 'jnmp of or-ani-m- to continue to live at a certain
!•" ality. Accord:n-]\ . -nch a group .-radii, dly di-appear- and
i- .r 'dually -iid .-died l'\ Mip which is adapted to the new

condition

The ch in c'nidition- rdenvd to arc climatic or phv-io-

graphie dial ad other- independent of ph\>iograph\" and

din; Tin- l.i-t type of ch > usually due to the action of

tin isms themselves. The-e »hn-e force- nia\ act separateK-

ther. Tin-! 5 probably the more common. In ca-e

tin- • li lioth the ' -i..n and the

-ii' • "t 5] l-icalit\- might lie pa.rtl\- due !<•

migration of a succession! Narthorsl '70. lid,- \\'ann-

ing, '09, lound that the -il- imderK 'ing moors

areofarcti< 'undra plant-. I: ial dispersal arctic-tundra

plant- and animal- \\ere -n. - .-.nd a— nciali-d

animal-, and I .\ d. . :«lnnii- 1 1 • «1 animal- Adam-.

-

'<itit>n oj //• i ! ' • • a/ -s'/c - essi

1 pointed out thr lact th.it there i- lrei|iietul\ a

hori/oina! -ioiial - •! diltt -rent a.

'ed \\ith c..nlimiou- ph\ -!• 'graphic pr- . -nch a- the

depo-iti.-i id alonj Tlie di.t^ram Fig i illustrates

licith thi' liori/mual and \eriical -erie- ••! c. mdin'i >n- with wln'di

\\ella\el'eelldealil ' 'Ol I 11 f t 1 H f poi II I ed OH I that ll'e

hon/uinal series »\ plain i "in muni tit - nm-t Near a close ( < "1-igical

mMaiH'e or lie pt.K n't ,ill\ identical \\ith the \ertical -. ii< - of

pa-t plant com mini hie- \\ Inch ha\e -ncceeded om- allot her o\ er a

^i\eii loc.tlitN in the older part of the hori/ontal -eric-. lie

pointeil out further that the hori/ontal -eric- may lie taken as

an index of \\hat the vertical Series ha- lieeil. lie \\oiild e\t cpt

the tir-t Si - of plant communities which occupied a locality

at the cl'i-e o| the ic. gi aid \\hich a- ha- ln-eti -tated. an-

•ic plant-. Thc-e arctic plant-, ho\\e\er. nm-t iia\e attd N d

146 VICTOR E. -Ill 1.1 (>R1>.

the soil or pond bottom in much tin- same way as the pimieer
plants of the present climate would affect them. The difference-
resulting from the change of (limale are ihen, those of detail
rather than of principle.

4. The Relation of Ecological Succession to .S'/'rr/V.v of /-'/v//.—
If fish were found fossil in tin- bottom of Pond 14, in the order
which we have indicated, one might conclude at once that it
constituted a proof of ecological su< • > ssion. This seems to have
been the general impression of zoologists who have heard tin-
presentation of these data. The que-tion has been asked, "Do
you find fossil in 14 all the fish which you mention as occurring in
succession there?' My answer to the effect that no such fact
has been discovered seems to have been regarded as constituting
a refutation of the entire statement of ecological succession.

If fish were found fo-sil in the order described above, and the
species and order of species, the same as now found in the hori-
zontal series of ponds, we would have some important data bear-
ing on succession, migration, and other matters of interest to be
discussed presently, but this would yield no crucial ci-idence for
or against ecological succession. Ecological suciv-^ion is based
upon physiology, habits, behavior, mode of life, and the like,
which I have proposed to call mores (opposed to the term form).
Unless the mores of the morphological species found fossil were
the same as the mores of the same morphological species at
present, they would have no weight in the matter, and it would
be impossible to ascertain mores from fossils if Mich fossils were
found.

If the same one or more species were found fossil in each and
all of the vertical stages of a pond like i_|, the evidence would not
refute the proposition of ecological succession because the physio-
logical char.ie'er- of the individual- of a gi\en species living in
the early stages could have been very different from tho-e of
individuals living in later Stages, without the differences beir.L'
shown in the preservable skeletal strueture. Furthermore the
modi liability of animal behavior seems well established. The
same species of plant may remain in a number of dill-Tent pond
stages. Such plants .-how -uiiable functional responses mani-
fested by different gn>\\ th-lOrm- in different stages.

ECOLOGICAL SUCCESSION. 147

The proof of ecological succession must re-t on the result -
experimental \\ork on the animal- of the different stages of -uch
a horizontal series. // matters not from the point of :
ecological succession ichat the forms > lon^ as the arc

'•:rev.t in the different > horizontal ml the

i'li a •'//.'<• ;; :'ed icith the environmental conditions.

When -urli difference- and relation- are found, ecological -IK .

-cniially • -lied. h is ol .i/ed that

within raiher uncertain limit.- the >• ..f a morphological

-p< • rn.iin. in a •:< ner,'.! way. the same throughout it- geo-

Thi- kind of rea-oning mu-t In- pursued \\i;h

i. union. Whi-n applied in detail or to "common" species \\hieh

\\ ide '-.m-iii^, it i- i • ill trustworthy, [f, for example, an

aiination of ihe de;i>-it- in i!n- bottom of a pond, sue!;
I'ond l j. showed the pr« fish \\hirh now habitually

inh.tliit- the you >\ pond -i ion, let it U- a more

ii'Mihern, or tin hose in i he hori/oiita! series

und r cor \\oiild con-tiiiitf -oine e\ ideii( •

logical -i!' -i \\hieh could lie Itirther checked \-\ the -tud\-

o| the modili.'.l'ili". of the mores of that -,prcje- at pre-ent.
HOV • in tin- kind . •! t \ ideiice lie crucial.

I iirthei mori . the i-iiidition- -tich a- \\rre in prim.ie\al I'ond
l | i ould ha\e been i'e. ichrd I -\ a pond like I'ond I in .1 feu hun-
dred- of \. In a pi nd \\itl: I mori MlilaMe ill. ill -and
for the -rouili :llt-, the-e cliail, id lake p!
\\ithin the life time ..f the lualder of Mich a pond Knauthe. '07.

I'- 5;

\ I. i .i NIK \l I >!-« i SSION.
We noted in the precedin. r that physiographic analysis is

a nieih<id. Heieu. \ . d a i lot her met hod \\ hich , \\ h i le

not ph\ -io-r.iphic, i- ,-imilar in principle, -ince it deal- \\iih
-ucce— i"ii over .' ^i\en locality. The cause of thi- succession
i- liiolo-ical and the biological ii, :- proportioiiateK greater.

We ha\e here the -aille attempt to disCOVer -omelhiilL' of the
ph \-iolosJca I character of the or-ani-m as a whole, and to classif}

ini-ni- on the ba-i- of the conlitioii- in which the\- are nm-t
iitarK in ph\ -i"l".^ical e<]iiilibritun. A;j;ain, the mures ol the
li-h of pond Stages in other parts of 'he \\orld are probably Minilar
if the pond- are -iniilar.

148 VICTOR E. -HKLFORD.

However, a !ei.'.i ti male '|i'.e-t ion at tin- point would In- : " \\'h,it
is the value of al! thi- reconstruction and tln.-c complicated
conception- to biolo-ical science and tin- analysis of the organ-
ism?" Indeed, this is a (jiic-iion which we have a-k< d onr-elves
repeatedly while working through the necessary plant data and
all the .-.(-altered unor-.mi/ed literature \\ith which the inve-tiga-
tor in thi- line mu-t deal. The question has always been ans-
wered in a manner satisfactory to us because whatever value the
papers on the ponds of the south end of Lake Michigan and other
similar paper- may possess, i- due to the conception of pond
succession acquired from the plant ecologists. Had I not ac-
quired this knowledge I would have done as others have done-
worked this excellent area over without seeing anything in it.
The problems that have arisen in connection with this work
have given a fresh point of view and a motive for investigation
which has repaid the effort.

In connection with problem- and moiive for their solution, it
should be noted that thi- is one of the most important things
which the plant ecologists regard as of value in the conception of
succession. Cowles, who ha- done more to stimulate work on
succession and the use of physiographic method than any oilier
American ecologist, regards functional response of plants (deter-
mined by watching plants and experimenting on plants) as much
more fundamental than succession. Thi- appears to be ^end-ally
true of plant ecologists. Warming, who has been a .urcat leader
in ecology, emphasizes the physiological side.

\\'e have, then, amonu ihe workers and originator- of the n-e
of this principle, a condition with roped to its place in biolc. \
which is quite different from whal one \\ould expect from .-imply
noting what has been the dominant thin- in their work. The
relation of the historical and ijeneiic side of ecology to more
fundamental physiological and ecological problem-, i- similar to
that of the historical and phylogenetic side of evolution, to the
problems of biology ami ihe motixe- for in\ estimation. \Vheii
Darwin framed the idea <>l evolution into a lo^ic \\hich \\a- not
refutable by academic attack, the fails of biology took shape,
arranged themselves into orderl} relations to < n-h other. Tin-
made possible method- of \\ork \\hich were new. opened up new

ECOLOGICAL SUCCESSION. 149

problem-, tin' attempt at the solution of which has made modern
biology what it is.

If we assume the attitude that nothing can be done in the
organization of natural history materials into a science, we are
do-in- the que-tion to investigation, much as the id. -pecial

creation closed other lines of biological in\ v-ti-ation. Still an
occasional biologist appear- to take this point of view.

A-ide from tin- general questions which we have been di-cu — -
an- many practical applications of pond successsion
dy of behavior .md t<> the economic and quantitative
>idc of l>iol"L;\. The-,- we will di>cu»s in a succeeding pa-

under tin- In ad < if t : ;'>n in pond-.

VII. -M MM \KV.

i. Then- i- s of ponds at the south end of Lake Michigan

arranged in ihe .-rder of their - i- determine<l by the

jili\ .hie history; b\- the relative amount of huinu^ and

ban bolt.iin. and b\ tin- ijualitv' and (|uantit\ <>!' vegetation.

.f li-li are arran^-d in the-r |»oiid> in an i-rderK'

fa -hi 1. 1 1 : i In- order Is related to the age of the ponds.

j. I he p'Hid- • 'I dittt-r'-i in the hi-i'>r\

of c.ldcr |" nub.

4. Tin- horizontal -h cr.iiimunitii-- i- i-.-..|.i-irally
repn-veiitati\e "f tin.- MI, -iv— imi nf ti-li coinmunii ie> \\itliintlie
older puihb.

5. Tin method employed here i- similar to |)h> M'< i^r
anaK-i-; the m..ti\e- ami po-Mble re-ult- are -imilar but
strictK' bi..l. i^ical. because the caus the SU< n are the

iUJMll- tllem>el\

II- . : /

,11.

VIII. ACKNOWLEDGMENTS AND Bir.i.i' »\\< \I-HN .

i. .[<(•' -The writer i- indebted t,. I >r. II. C.

Cowles for assistance with the plant >ide. of the pond problem:
to Mr. \\ . C. All-' '"r assistance in mapping the p<>nd-. The
ideiititicalion ,.f the ti>h i- b\- Dr. S I M k and Mr. S. I

150 -VICTOR !•:. SHELFORD.

Hilclcbrand; some of the coHrt-iin- was done in cooperation with
Dr. Meek and Mr. Hildcbrand. \\'iilumt the a-si>tance of
these gentlemen this paper could n«i h,i\c Uvn \vrium. The
writer is also indebted to Mr. Ellis L. Michael for tin- ivadiiiL;
of the manuscript.

Adams, C. C.

'05 Post-glacial Dispersal of North Amei it an Biota. Hiol. Bull., Vol. IX.. p. 5.}.
'08 Ecological Succession of Birds. The Auk, Vol. XXV.. pp. 109-153

Bi! hy.

'09 Isle Royale. Biological Survey of Michigan (Lansing). < lim.uii- ami
Geological Succession, p. 45, p. 31. Succession of Birds, p. 134. Succession
of Beetles, p. 160. Succession of Mammals, p. 390. Bibliography.
Alden, W. C.

'01 Geological Atlas of the United States. No. 81, Chicago Folio U. S. Geol.

Surv. Maps.
Andersson, G.
'97 Die Geschichte der Vegetation Schwedens. En.ul'-i > Botanical Jahrbiicher

B. 22. p. 433.
Atwood, W. W., and Goldthwait, W. J.

"08 The Physiography of the Evanston-Waukegan Region. Bull. 7, Hlmni-

Geological Survey.
Blatchley, W. S.

'°/7 Geology of Lake and Porter Counties, Indiana. 22d Ann. Rep. Ind. Dcpt.

Geol. and N. Resources, Map.
Chamberlain, T. C., and Salisbury, R. D.

'07 Geology, Vol. III. Henry Holt, New York.
Clements, F. E.

'05 Research Methods in Plant Ecology. Lincoln, Nebr.
Cowles, H. C.

'01 The Plant Societies of the Vicinity of Chicago. Bull. 2, Go.;'.

Chicago. Also Bot. Gaz., Vol. 31, pp. 73-108 and 145-182.
Dachnowski, A.
'08 The Toxic Properties of Bog Water and Bog Soil. Bot. Gaz., Vol. \<>,

p. 130.

Gilbert, G. K.
'85 Topographic features of Lake Shores. 5th Ann. Rep. Dir. U. S. Geol. Surv.,

pp. 69-123.
'98 Recent Earth Movements in the Great Lakes Region. Ann. Rep. I'. S.

Geol. Surv., 1896-7, Part II., pp. 601-647.
Goldthwait, J. W.

'07 Abandoned Shorelines of Eastern Wisconsin. Wise. Ge«>l. and Nat. Hi-'

Surv. Bull., XVII.. Scientific Ser. 5.
Eycleshymer, A. C.

'01 Observations on the Breeding Habits of Anu-iuiii> nelmlixiH. Am. N.it

Nat., Vol. XXXV., pp. 911-18.
Forbes, S. A., and Richardson, R. E.

'08 The Fishes of Illinois. Nat. Hist. Surv. of Illin"i~ (State
Icthyology, Vol. III.

ECOLOGICAL SUCCESSION. 151

Hankinson, T. L.

"97 \Va!nut Lake. Biol. Surv. <.t M i- • Sui 07,

pp. 161-271.
Knauthe, K.

'07 i.lamm.

Leverett, F.

'97 I' •• Features an.l ! • ihr ("hirauo Area, t'h .i.l.

X II Surv. Bull., II.
Meek, S. E. and Hildebrand, S. F.

"10 - Known t within Fil'ty M

3ei ' VI I . . Ni
Reed, H. D. and Wright, A. H.

'09 Tin- \'«-rti-l»ratt-; oi' t!i :i. X. Y Proc. Am. I'liil. >

XI. VI 1 1.. \
Reighard, J.

'03 "I : y Volume, pp.

Salisbury, R. D., and Alden. W. C.

'01 ' , f Cl

Shaler, N. S.

'92 i jili K'-p.. I )ir. U. S. ( .i-ol. Sin \

Shant/, H. L.
'06 A Mesa P Pikes Peak. 1 h<-

Sharff, R. F.

'07 I I >i-ti iliiilion.

Shelford. V. I

'07 !'• in<l

Mull.. Vol. X I V . pp >i ij
'10 I ' iilinn-. III. M. •

II pp
Steenstrup. J. J. S.

'41 .• k iiiul. -~k.i\ • Vednesdam

I. ill. iniini-nliiiKi-iuli- Mi m.i. ik-

nini;iT. h . r i Ahniiuli-Ii

Danske \ I.. IX.. li Warminj

Warming, E.

'09 'i n to the -tiidv 'iiiinun;-

in.

A HETEROCHROMOSOME OF MAI.F oRIC.IX

i\ la -in \oins.

MAVII) HII.T TKXXhX i

The observations of Haltzer ('09) seem in have led to an
immediate conclusion that in cchinoid^ the female is the hetero-
gameticsex while tin- male is homogametic. My observations on

Illpponnc cf1

the „ - cross, taken in conjunction with those of Heff-

1 oxopneustes 9

HIT MO) and Pinney I'll) show that this conclusion cannot be
final. The study of the material mentioned has given con-
vincing evidence that in Jlippouoe cscnloita (Tripneustes esciilen-
///M.a heterochromosome is carried by half of the spermatozoa
and that the dimorphism of somatic chromosome groups in

Uipponoe cf

in straight tertih/ed Inpponoi 'and ,., material must

1 oxopneustes 9

be i orrelated with dimorphism of the spermatozoa.

The heterochromosome in question is hook-shaped in form
// Fi^. i, A, H Fig. 2, B). In a study of fifty- two spindles in

A B C

i. Tuxnfim-H.^lfs 9 X IIif>{x»i»e c? • 'I'lncr 1< >ni;ii ii'linal sections oi
spirull'- in anap!ia-c. In A and /-' twi. chnniK. ;t in i.utliin- ii.r -ak. .-I

\ t, ( lllnlllri-i.llli - -ll'.UIl. lli.uk- ill -iili- \ic\V. XI..S"

which 1 ua> able to reach a definite conclusion a- to tin- pre-ence
or absence of the hook-shaped element , it \va> found to be present

in twenty-eight and absenl in twenty-four instances (Fi^. ^!-

Miss Pinncy ('iu has been abK' to sho\\- the occurrence "t
this element in half of the straight I'eriili/ed llipfxnn' The

152

HETEROCHROMOSOMi: i >| MALE (>RK,IN IN ECHINOIDS. 1 5.;

non-occurrence of such an element in Toxopneustes ey;^ has
been shown by Heffin r '10) and by further Mudyin my labora-
tory with this point in mind. I have been able to determine
that tin- hook— haped chromosome i- pre-ent in hall of the
Toxopneustes eggs which have been fertilized \>\ IIi[>po> - rni.

,'

ABC

!

! i . in irniit vii-w.

By this analysis of the subject it is th< -h«>\\n i lliai tin-

IK -iik--. ha; » d rhr- mn •-. mn- i~ |u-culiar t<> IIif)f)nn«'. J that thi-
chnimii-i.nir i- i arrii-d b\ tin / ermatOZOa, and ; .

that, -inc.- thi- ••!. -nn in i- t«und in half i if the -trai-ht frrtili/i-d

I 'I / ' /

L

I

A TJ P

/v *•

•

NIP In ink- |P!

-and in halt nf tin 'iTtili/cd eggs it nni-t bt- prc-cni in

but half i if the H'rrmat"/'

V , .iiclu-i\i- ^lali-nu-nt can as \ et be made re-ardin^ the
number nf chromosomes in the-, eggs I'he --Id idea that the
xunatic number of chromosomes in echiimdi-rm- i- • -it her ei-htei -n

154 DAVID MILT TEXM:\ i .

or thirty— i\ i- c\ idently incorrect. Piniiev 'i I i h.i- -ho\\n |", ,j-
S- "'" 33- HelTner rio) counted ,y> for To.vopncustcs.
A '11 counted 37 or 38 in anaphase plates in 'J'o.vofmcmirs,
count- which I was al>le to \crif\-. In tin- cross fertilized eggs
ti-mv<l in thi> papc-r tin- counts vary. Fu. i shows .Vi : Fig. 2
>ln'\vs ,^l ; I i^. ;, -ho\\> .^J.

Mi-- I'iniuA ' ' i i i has given as full a <li--u--ion of the >ul>ji-ci
as tin- I, ici- known at this time warrant. For a conclusive
re-time a full knowledge of the development of the LMTIII cell-
in the>e form- now seems alnio-t imperatiw.

VN MAUR Coi.i.i

BIBI.K »GR \1'!IV.
Baltzer, F.

"09 I)i<- ' liiiiiiin-Miin-n v. Mi,)iv-;yl. li\'. 11. I-'cli. i:iinn. Au:li. l". Xcllti il -i li..

Bd. II.
HefiFner, B.

'10 A Study ni Chromosomes of Tox. \'ar. which Slunv Individual Peculiaritic-s

of Form. Biol. Bull., XIX.
Pinney, Edith

'n A Studyof the Chromosomes of Hip. esc. and Moira atr. Biol. Bull. XXI.

HETEROCHROMOSOMES IN THK GUINEA PIG.

X. M -

In \ii-\v of tin- fad lh.it the i:iii' . i- SO much u-ed for

• •rimeiiial breedi in Meiidelian heredity. : icd

di--iraM«- i» secure some further kno\\le<;. the behavior ot

tin- chromatin in ihe ucrm cell-. An attempt ha- t hercf< n\- l>een
maili- in folliiw through the -pern, -. and a l>rie! note

• MI t'he -iil.je.-i \\,i^ puhli-hed in tin- BK>I<" i< \i Hi 1.1.1 n\. \'..l.

XX . \». I. Jaliuarx. I'd I. The material h. <d ti. lie

\i r\ unfavorable, and the re-ult- an- im: ictorj

in !M- desired. A- M. ven a \i-r> full actMtmt of

the transformation of ih« -;irrmaiid- int' I -hall

omit all d !!u- -pern and

tiin- IP inion of the rhromatin in tin'

niirlei and mil- • • 'erina-

tid

MM

\iin-''1!' fixed b) I uietliod- which had

material. It
idem that onl | -'lemming and Mi rniaiin

o-n lie mix i ii-. . ! li xation Hi the chromosomes

ill mi'' -id al-o that th. - nm-t he reino\ed from the

animal. < lit in!" -mall : red to the fixing fluid

\\ilh ^reat rapidit\ i' -1 re-uli-. M<

nu-ihod of n-iiii; ihe lixin^ solul ' pro\ed to In an

im|>ro\emeni on the !d -olution-. Tin >ns, 5^

thick, \\ere -;. lined with I'itlu-r thionin or iron-h.emai"X\ lin,
oral g( ( • In-ill^ u-ed in >om< a p!a-m.. -I tin. Thionin,

'hionin and • ;lu in. -\ n-ult-

cia!l> for ihe chroiuo-onie> in niiti \ * aniline prepara-

tion- \\ere mad' ire-h material and |>ro\cd \alu-

aMc. h ua- found tha - "! the ti--ti- m. d \\ith

needle- in carmine would keep in fairly -on.) ccmditioii in

a ti^lilk -toppered \'ial for several \\eek-.

[.S6

N. M. STEVEN-

SPERMATOGONIA.

As in nihrr vertebrates tin- tc>u-s om>Ut of much coiled
t.ulmle- in \\hich, in mature animal-. -Merman >.u«nua and Sertoli-
cfll- a IT found at the- periphery against the wall of the tulmle:
then coiiK1 lirsi and sccoiul sprrniati H-\ it--^ in \-ari<.ii> stages,
-l>rrmatid>, and unri|)L- s])crmatozoa attached to long processes
r\ u-ndiiv^ from tin- Sertoli-cells toward tin- criitcr of tlic tulmle,
-|)i-niiato/().i licin- found in or near the lumen of the tubule.

I-'H.. i . \u. l.'ii- i .| Sntiili ci-11.
!•"[(.. j. Xiii \<-\i- i)i spermatogonium.
l-n;. 3. Spireme st.t^i-, -]>• i in.it'>-(iniiim.
FK.S. .j anil 5. N'UI lei iinin .1 \niiiit; tc~iis, M--t--t,i.y>' ;nnl
FIG. 6. Fifty--i\ chromosomes <>t .1 -i"1' ni;it"^n]ii.il
prep.

P"!G. 7. Efjuatorial plaii- "i a -pi-i in.it, i^nnial mit(>M~; s'1 'In ....... soincs. All

1 1C i ' j -

h-

Aceto-carm.

HETEROCHROMOSOMES IN THE GUINEA-PIG. 157

In one very young animal only spermatogonia were fi.iiiid. Sertoli-
cells not having been differentiated, and n<> maturation stages
1 1 . i \ i n g yet appeare< 1 .

The nuclei of the Sertoli-cell- ( Fig. i) are clearly different i
from those of the spermatogonia. They are larger and usually
flattened parallel with the wall of the tubule. In the rer. u T of
the nude' i large plasmosome and closely a--ocia!ed with

it one, two or nn>re rhromatin nudeoli, or karvo-omes, which
otldi .tppiMi- to h.'' • ntral vacuole. The retieulimi of the

under u'n with thionin <>r iron-haematoxylin.

Tin- 1 irg< r number of the -permaiovom'al nuclei eoiiiain !
dump- of diroiu.uin of various and tangled thread- into

which the clump lually resolved after a mitosis Fig. 2 .

>hoi-i I mito-i- .ill nf the dump- disappear and a perfect

spiren e i- formed 1 - In a young te-tis nearly aM

"1 the nuclei are Minilar to I _ j. A fe\v are in -pircinc 01
|>rojih. i h ha- pn .\ ed .1 \ er\ dil'licnlt mal

in dei ei mine the number of chromosomes either in the spermato-
ia orspermat [ have never found a case of a perfect !\

ilai plate \\here there was n-> overlapping. After spending

it deal of tin me p"int. I ha\e concluded thai 50

nd 7 is ;i:"l>abl\ llu t number. I i-. (> -!i..\\-.

in outline the cliroiiio-oim-- of a ppipha-e. iv, ,m ,m aceto-carmine

|ireparalion. the cell ha\in^ been much flattened. T\\o 1
pair- can be distinguished, and the oilier- vary con-iderably in
form and Fig. 7 i- the be-t equatorial pl.ite which \\.i-

seen eiiher in .1 nniiie iu'eparalion- or in sections. I hi-

\\.i- taken from Hermann mal. -rial Stained \\ith thionin. It will
ince be -een thai the chromo-oiiie- o\erlap in -udi a manner
a- l.» maki' n curate < ountin;; difficult. T!ie one aberrant i hro-
nio-ome i- not a ch ristic feaiure. ( )ne unu-uall\ lonv pair

of rod- i- conspicuous. There are -everal V's, and the remainder
ha\e the form of -trai-ht <u- -oineuhat cur\e<l rod-. The t \\ <>
Ion- roil- an- always noticeable in an anapha-e, lagging behind
the shorter < hromosomes. The V's are probably ihe same chro-
mosomes \\hich ha\e the -pind!. --liber- attached centrally and
pa-- to the pole- a- V's in the fir-t maturation mito-is. The
heterochromosomes one cannot distinguish at this stage with

158 X. M. STEVICN-.

certainty; tin- smaller one i- one of the -ma'le-t chromosomes,
perhaps y, and .v which seems to have no mate may be i he larger.

I iKsi SPKKM VTOCY1 i-:s.

'I'lu- inn-lei of the young <ermatoeyte- arc r-imilar to those

of tin- spermatogonia Fig. 2.1. clumps of chromatin and line
thread-. Before the clumps have disappeared the chromatin
t lire. ill- are more or less massed at one side of the nucleus (Fij
in a syni/e-i- stage, which resolves it -el I" into an irregular bouquet
e ; .Fig. iji \\ith loop-, much coarser than the threads of the
prc\iov- st.ige. Frequently one dense chromatin ma-- i- seen
half hidden among the bases of the loops. Some of the loop-
look as though formed by telosynapsis. but if this is so, the halves
of the bivalent loops must later become parallel, for the bouquet
stage passes over into a parasynaptic stage (Fig. 10 and Fig. u)
which is followed by a -pitvme stage (Fig. 12) in which then' is
no longer any e\idence of synapsis. In the spireme stage there
always small masses of dense chromatin-like material lying
against tin- nuclear membrane, and staining as deeply with
thionin as the chromosomes in mitosis. A large heterochromo-
some -v is always present , and one or more plasmosdmes can
u>ually be found in sections. In early propha-e- (Figs. i $ and
14) the chromatin appears as though gathering together about
definite centers along the spireme, leaving linin threads between.
Sometime- a longitudinal split is visible in sonic of the larger
( -hrom-ome- i Fig. 14.) In several cases at this stage the two
heterochromosomes (xy) have been clearly separated as in Fi.
13 and 14. hi Fig. 13 a pale plasmosome is also present.

In most first spermatoeyte spindl< - I \\ o chp>mo->ome.-> are
conspicuous (Fig- 15). One of these (.vv) is I he unequal lietero-
chromosome, usually bilobed at the larger end (.Y). The other
(a) is larger than an \ o! i he ot her- and u it lion t doubt is composed
of the two long rods of tin- spermatogonia. As a means of
determining the reduced number of chromosomes, a number of
spindles in the best material were -elected, .md an attempt
was made to draw all of the chromosomes \\hich were divided
between two sections. T \\enty-eight were found in a number
of the clearest cases. The-e are all -ho\\n I'm- the spindle of

HETEROCHROMOSOMES IN THE GUINEA-PIG.

159

8

^-y

13

.

15

14

i-i... p. B..U.|

l'i .111.1 M . I

l'i •; !
i ;. 1 v. tin- 1. -.ir; a,

till-' '.

: :T«m tin- -pin. I!. ••! l;i^. 15.

i 7 I • -i lal pl.itc, 2S > liti'ii,

i6o

N. M. STEVENS.

which Fig. 15 is a part, in Fig. 16. Many of them are • •\idently
parasynaptic pairs attadu-d to ^pindle libers at OIK- end, others
are similar pairs attadn-d in the middle and pulling apart in the

18

19

22

24

23

26

<*»

a

25

27

FIG. 18. The heterochromosome paii • \ y i il

FIG. 19. Metakinesis.

FIG. 20. Anaphase. Heterochromosonn p.m -till muli\iM. 1.

Newly formed spindle; chruiiici-(>nii'~ \\i-ll ailvunrnl in inrtakinc-i-.
Older spindle; metakinesis hardl) l»'_;iin.
FIG. 23. Spindle forming with chromatin in lurm oi a tliic k -piicnic.
FIG. 24. Anaphase; chromosome a separating as .1 pair <>t 1
FIGS. 25-27. Chromosome a in the t'«im <>i a -plit l.aii'l. |ia--in.v; into a
form in metakinesis.

FIG. 21.
FIG. 22.

HETEROCHROMO-MML;-, ix THE GUINEA-PIG. 161

form of crosses. The rods are probably lateral views of cross-
shaped forms. Fig. 17 shows 2S chronio-, mie- in an equatorial
plate. Fig. i >« i- another metaphase in which tin- members of
the heterochromosome pair are ah-' -eparating earlier than

the others. In Fig. 19 tin- separation ot some of the chromo-
some- as rod- ami V- in metakiiic-i- ma\ n. Fi^. Jo i- a
laii- anapha-e in which the heterochromosome pair is still un-
divided. This is unu-ual. but >ln>ws a kind of variation which is
iceable through all of the spermatogenesis. Varia-
tion is nio-t eon-picuon- in the pn>pha-e- of the fir-t maturation
mito-i-. As a rule one find- such earl\ prophasi are sho\\n
in I -"i;;-. i.} and 14, and the ehromoM .mes of the \oim- spindle

\\i-ll on their \\a\ in the process of mctakiii'
bin one -oineiiiiie- hn«!- the ehronio-oine- of an older sj)indle
much K-- I.M- aloii-^ touai'd di\i-ion than in I ig. 21,
and not infrequently I ha\< . ond,-n-ed ^pirenie. as in

I . ell, the nuclear membrane having disappeared,

and a -pindle formil ' - nnisi, I think, be an

\\hiih maturation t.

place at dilieiein linn--, and prob.ibl\- under different stimuli, or
stimuli ot diffen in ini- ' 24 is an inn anap!

-ho\\in. the lar.c chroino-r une pai' as a pair of \ '-

illslead ot ,\o||Id lia\e beell e\|ieeli'd Ifom !

i In tin- pn parations from one animal I repeatedly saw the

largest chromo-oim :io\\n in Figs. J.s. -1' and 27, a -:.lit

band forming ill a later Si Lge Fig. J7 . Ill all ••tiler

mail-rial the la- mosome ha- appea in l-'i^. 15,

\\hich. lo-i-ther \\ith l-i-j. ^4. SUJ lhat the four end- of the

t\\o |,,nw chromo-ome- \\hich lorm the |«.:ir an- usually |o|di-d

I he t\\o lateral hakes of the cro-s sho\\ n in I ij. l~

f • ild i d together would give the form shown in I ig. i .=,-/. and mi- In

then -epaiMle as ii: ' '. The -ame kind of folding ha- very

lil .-i !\ occurred in • hromo-dine- 5 and l<) in the -lioun

in Fig. [6, and po--ibl\ in ..tin - uhere it i- less «-\ident.

' i\D M'l-.KM \ I'" N II-.

Hi-i\\een the t\\o maturation m 'hen- i- a res ge in

\\hich the pa;r- of second spermatocytes may often be found

I 62

N. M. STEVEN-.

connected by spindle fiber- .1- in I'iv. 2*. This timuv sh<>\\-
the larger (.Y) and smaller (vi lieien>ehn>nin-(i]iu- \\itli Himoth
outline, and the remainder <>l tin- rhnmiaiin in the form of
irregular clumps and strands. In sonic cases -ec-md spermato-
cyte spindles have been found associated with firsi -permatnrytes

30

31

33

35

FIG. 28. A pair of second spermatocytes slmwin- tin heterochrom
and y.

FIG. 29. Second spermatocyte metaphase.

FIG. 30. Second spermatocyte equatorial p;

FIG. 31. Second spermatocyte anaphase.

.Pics. 32-35. Four stages in the transformation <>t" tin- spermatid, -li«>\viiii; no
trace of the hetcrochromosomes. All figures 2,000 X i '^ reduced ' _•.

iii mitosis in such away as to indicate that the rest stai^e betwcrn
the two divisions may often be omiited. This is as tar a> the
heterochromosomes can be traced.

In the second maturation spindle I have never been able t<>
count the full number of chromo-i-iiu •.- because <>!" > roudiii^ and
overlapping. In the clearest case- -een in ai rniinc prepa-

HETEROCHROMOSOMES IN THE GUINEA-PIG.

rations, the chrom< re <luml>-l>cll sh.iped. as in l-'i-. 20.

They are, ho\ve\er, rarely so well separatc<l. Fit:. 30 shows 24
out of the 2S which should appear in such a plate, and Fi-. ,^i
an anaphase in which no attempt wa- n, • dr.u all of the

chromosomes. The-e figures though valueless as a dcmonMra-
tion of the number of chronio-omes or of the division and di--
tribniion of terochromosomes, are .J\vn to sho\\- that in

the guinea-pig tin-: no -uch -. -cond synapsis or nninerieal
reduction as ha- la-'-n de-cril>ed l>y C.i: '09, PIO for the

guinea-chicken, the doi '-hieki-n and for man. also l>y Jordan

'M -or the opo—iim. i '-jileir _ in the sections

\\liich -ho\\ that tin- number i- nly not n-dn»-ed <me half,

and pio!,,;M\ n,,i n dih .ill.

iv MA I II

In the younj 5 as in Othei 'inint-diatelx'

follouin- in iln- chroin.uin rs in inv-ular i-liimp^

• :natid stage i- >ho\\ n in
pla-.ino-.niH-. an iciated

\\ilh it in tin ••!! a lai-^c ma^- of ehrom.itin from

\\hieh -tr.un! ihirkt: to ihr niiclr.ir mnn-

liraiif. In a SOmew h f I he enitral elnmp o|

ehroni.ttiii In-, ,ill .: I, and tin- mn leu- i-oiitain- onl\- a

f.iinlK staini ienlnm. In the half .^PIWII >prrmato.

Li-hrd to tin- Sertoli cells, the miel.-n- i- line!\ granular !

I'hi^ licnre \\.'- drawn from paralion -t. lined \\ith

ihioiiin. I'lie iin- \\.i- >taine«! a d« -ep Mile, and the ime1
^ramiK - aUo Mut-. • at the |>ro\imal end. Tin ire-

^i\en a> evidence thai ti rm.itid-, and -j >ermato/( ,.i ,in-

ii"t \i-il>l\ I'imorphie, eaeh of the ti^nre- Ix-jn- t\]>ical for all
of the nnelei of ' A In'ell < SCntS.

->! MM \K\ .

I . The prolialile miml >er of ehroino-. nr.e-, iii the -perm.it o-( mia
of the -uinea-|ii^ i- 5'-. .uid in the >permato< \ tes JN.
2. S\ nap-i-> i- of the para-\ naptic tyt •

; A t\pieal heterochromosome i- pre-ent in the growth stag

of the lirst >,|)ermaP In metakinexj. thi-, i- separated into

a larger and a -mailer component, as in many insect-.

164 N. M.

4. The second >permatocytc> arc vi-ibly dimorphic in tin- n -i
stage, one containing a 1. IT-IT, tin- other a smaller heterochroino-
some.

5. The spermatids and spermatozoa are not visibly dimorphic.

I >IS< i 55 [ON.

One or more unpaired heterochromosomes have been reported
by ('iiiyer ('<»), *IO) for the guinea-chicken, dome-tic chicken
and man. and by Jordan (*Il) for the opossum. This is the
lirM case, so far as I am aware, in which an unequal pair of
heterochromosomes has been found in a vertebrate. Although
it has not been possible to follow the heterochromosomes through
the second maturation mitosis, there is no evidence aiiaiiM the
Mipposition that i-ver>- ( hromo-i mn di\'ides equally in that
mitoM's, ;^ivin^ spermalids one half of which contain a division
product of the larger heterochromosome, the other half one from
the smaller heterochromosome. If it can be shown that the
female guinea-pig has 56 chromosomes, two of which correspond
to the larger heterochromosome of the male, then the fertilization
formula will be like that for similar cases in inseci 3.

Eggs. Spermatozoa. Xy^ote.

\ 2-, +X + 27 +X 54 -f .V .V =9

i\27+X + 27 + y 54+Xl

This is what is to be expected, but, in view of recent develop-
ments in echinoderm spermatogenesis, it is at any time possible
that we may find other conditions; for example,—

Eggs. Sjirrmatozoa.

f 27 + Y + ->7 +A- 54 +x r =cf

'1 27 + y 27H , 54 + 1 1 =9

In this case the female would still be homo/yi;ous Inn
male would have the excess of chroma t in, rcali/ini; in thai
McClung's ('02) original sui;.^esiion in regard to the "accessor)
in the Orthoptera; and the s]ici-mato/(»("ni containing tin- A chro-
mosome would be male-producing instead of female-producing.

It has always seemed to me someuhai improbable that the
small heterochromosome of an ime(|ual pair o\\cs its smaller
size to gradual degeneration, and much more likely that the

HETEROCHROMOSOMES IN THE GUINEA-PIG. 165

unequal pair may, at least in some cases, be composed of an
equal pair of chromosomes with an unpaired heterochromosome
fused with one member of the pair, as McClung ('05) has shown
to be the case in Hesperotettix.

The definite cases of regulation in the number i>f heterochromo-
somes in the male-producing eggs of Aphis and Phylloxera; and
the case ot Rhabditis ni^roi-enosa (Boveri, '11: Schleip, 'u) where
a hermaphrodite with 12 chromosomes produces spermato
with 5 and <> respectively, by rejecting <>ne heterochromosome
in tin- second maturation divi.-ion, naturally suggest that the
in of ih,- impaired h.-tep.clironiosome in the male may be
attributed to a -imilar regulatory process occurring somewhere
in • 'lution ot a hermaphrodite or-anism. and i;ivin^ male

and female-producing -permato/oa. Such a regulation nii-ht
hr-t male- ami hermaphrodites, a not uncommon condi-
tion, bin tion "1" the male reproductive organs in the
hermaphrodite would iv-ult in the u-ual bisexual condition with
ihe . ei|iial iii number-, mile— (];• ratio \\ere ch.m.id
l>\ an environmenl discriminatii in-i one sex, or 1>\ some

ilalion in ihe number of male and feinale-producin- -perma-
tOZOa Ihe une(|iiall\ paired c..ndiiioii of the lieterochromo-
-oine- mulii ea-ily ha\e come aboul b\ In-ion ,,f ih<- heierocliro-
mosomea in boii \\iih anmhi-r [»air of chromosome- in the

COUl ' reduciioii in ihe number of chroino-ome-

characterisdc o| th h i- much more difficult to

ho\\ the change- necessary i" brin^ about I he condition indicated
in the -ecoiid lormula could occur, -inc. \\ . mil- inn either

tor ihe di-appearance of ihre( A' chromosome- lea\ in;< oiil\- the
one A in the male, or !"[• ihe a|)pearanci- of .in A" chromosome

I'axne'- "09, '10 figures for several of the Reduviidae

• other complication^.

Some recenl ob-er\ alimis on other material ha\ e impre— ed up-
on me the probability that the chroinatin units represent in- si \
and other characters may be very -mall indeed, and that in such

cases as that of C«/< 3 M , whereno heterochromosome

differentiation of .my kind ha> been detected, the members of ;i
pair of chromosomes may differ by a sex unit or some other
character unit, and the difference in size come within the probable
error- of most careful observation.

N. M. STEVENS.

\\"'TC it not for a few >t~ condensation of an apparently

equal pair of heterochromosomes in the male germ cells (Lepidop-
tera, Stevens, '06, Pederer, '07, Cook, '10; Anisolnlxi maritinni,
Randolph, '08), and in the female- germ cells (Aphrophora,
Stevens, '06, Rhabditis ni^ron-nosu, Bnveri, 'ii>, one misjit sus-
pect that the condensed condition of the odd chromosome and
the unequally paired heterochromosome of tin- growth stage of
the first spermatocytes might be due to their unpaiied or
unequally paired condition preventing them from joining with
the other bivalent chromosomes to form a spireme, especially
in cases of parasynapsis. In Cuh'.v no such difference is effect-
ive in bringing about a condensed condition of one pair of
chromosomes and in Anopheles, where the difference in size of
the heterochromosomes is slight, condensation is much less
complete than in most cases. In the guinea-pig condensation
is complete from the synapsis stage on.

In the guinea-pig the condensation of the heterochromosome
pair and its behavior in mitosis are as typical as in Tenebrio
and many other coleoptera previously described by the author,
and only the large number of chromosomes of such a consistency
that good fixation in mitosis is difficult to secure, makes it
impossible to give as complete a demonstration of the relation
of the heterochromosomes to sex as in many other forms.

BRYN MA\VR COLLEGE,
May 23, 1911.

LITERATURE CITED.
Boveri, Th.

'n L'ber das verhalten der Geschlcchtschromosomen bei Hermaphroditismus.
Beobachungen an Rhabditis migrovenosa. Yerhandl. d. IMiys.-Mrd.
Gesellschaft zu Wurzburt'. N. F., XLI.
Cook, M. H.

'10 Spcrmatogenesis in Lepidoptera. Proc. Phila. Acad. Sri.. April. 1910.
Dederer, P. H.

'07 Spermatogcnesis in Philosamia cynthia. Hull. Biol., XIII.
Guyer,

'09 The Spermatogenesis of the Domestic Guinea (.Xiimidia mrlra.uii- d'un.).

Anat. Anz., XXXIV., Xo. 20, 21.
'09 The Spermatogenesis of the Domestic Chicken (Gallu< uallu- ilom.). Ibid.,

No. 22, 24.

'10 Accessory Chromosomes in Man. Biol. Hull . XIX.
Jordan, H. E.

'n The Spermatogenesis of the Opossum. Ea-'> m / I ><

HETEROCHROMOSOMK3 IN THE GUINEA-PI* ,. I<>7

mU-r 27, 1910. - XXXIII.. Maivh IO, i'/n. \

in Arch. f. Z<-l!lcir-<:h.
McClung, C. E.

'02 The A rininant? Bi<>l. Bull.. III.

"05 The' ' K "i nnhciptrr;.: Biol. Bull.. IX.

Meves, F.

'99 liber Struktur und Histogenese M ii\v.-iiu-l.

Arch. i. Mikr. An.it.. I. IV.
Payne, F.

'09 I >i-tril>uti.': K tion I

Bull., XVI.

'10 I! :ulti~i>iiii>-.i. Ilii-1.. XVIII.

Randolph, H.

"08 < )n tli' i iiKiritir B I. I'-ull.. X\

Schleip. W.

'11 . '.'111111111 • Rli.ilnl'iiu-iii.i i nkio-

XIX.
Stevens, N. M.

'06 A ( '

R

1 ). II.

'ill'.:" ! • Mull.. XX.

A STUDY OF THE CHROMOSOMES ( )F HIPPONoE
ESCULENTA AND MOIRA ATROPOS.

EDITH IMXXI \

Tlir comparatively recent investigations made <>n fertili/ed
echinoderm ci;i;s with especial regard to the qneMion of chromo-
some individuality had their beginning in an attempt to correlate
the behavior of the chromos, >nics during fertilisation and the
subsequent cleavage stages with the observed facts of parental
dominance in hybrid cultures.

The first observations are recorded by Tennent, '07. He
the equatorial plates of Toxopueustes runt'^iti's, Moini
and Arbacia pitnctnlata in which individual characteristic-
for each species are evident. The contrast between the chromo-
some L^roups in Moini and Arbacia is so apparent that in the

A rbacia cf

equatorial plates of the hybrid the hybrid nature of

Moira 9

the -e-inentaiion nucleus is e.isily reco.u'iii/ed.

Baltzer, '09, in an e\ti-n>i\-e morphological Mudy of the chro-
mosomes of the European forms, Strongylocentrotus Uridiis and
microtuberculatus, was able to identif\- single chronio-

Strongylocen-

s which could be recognized in the hybrid

j he Inn us 9

The main facts which he describe^ ma\- for the sake
o] comparison be brielly stated at this point.

The chromo-omrs of Strongylocentrotus are in general rod-
-haped bodies which \ai~\- in length. The longest are Over three
times | he length of the shorteM. In addition to the>e then- are
in one-half of the e^> i \\ o and in the other half, three hook-
shaped elements. The occasional third hook is smaller than tin-
other two which are equal in sj/r and evidently form a somatic

1 Bovcri in hi- " /.<-V<-\\ stinli.-ii." lint VI., \<>«-. Jena. ra!li-«l aticntion to
Baltzer's observation- uhirh weir licin« ina«lf al iliai lime in Mn\ n i'-, la I >< <i ai • >i y.
Ho publishes oni' fiKun- ni Stt • ••iitrotit* -hd\\-in- a linnk-'-li.i|ii-il c hn iiim-i unc.

He himself in 1890 ha<l iininl tin- m-i uiiciu <• i.| rods "I ililii-n-nt lrni;tli> in certain
echinoderms.

1 68

CHROMOSOMES OF HIPPOXOE AND MOIRA. 169

pair. In all <>1 the eggs tin- -oinati;: number of chronio-omt
iht- -.uiic; /. c., thin y->ix. In accordance with previou- con-
clusion- upon the occurrence of the heterochromo-< .me- in in-cct-.
the dimorphism in the chromosome groups of Strongylocentrotus
i- correlated with the dimorphi-m of ^e\ in cchinodenn>. H<>\\ -

r, Ba!t/er, trom hi- "1 -ervation- on Echinus eggs Ci
lenili/ed with Strongylocentrotus s])erm. to-ether with observa-
tions on nuiltipolar -pindle- in di-pern. < ggs, conclude- ihat the
odd hook i- originally pre-ent in the e.y.u. In-tead of l\\o kind-

perm as, we know. e\i-t in tin- in-rrt-. in ihe i-chinoderm- tlu-rc

tWO kind- • Po QUOti from \\'il-..n. "lO, "il i- ihe

Ifinalc thai i- the lit' ullile the male i- llonio-

in-odui lion in th,-r aiiim.il- mn-i then-Ion-
<-oiiform to iht- lorinu!

:oan (/ /•'(!. l-\-male.
.-. Male."

I - inn nt has obtained evidence which, 1 think, shows thai thi

loMimla^ d<> not ho|<l for all echin 'drnn-. An .n-ioiini of hi-
int<-n--tin^ ob» i\ Ltions a «-l-i-uhi-rr in thi- journal.

In Echinus, \\hirh al-o (oiiiain- lhirt\--i\ « hromo-onu--, ihe

urrence ••! ilmt «; ristic pair- \\ iMi-hnl ; \i/

pair o| lar^r hooks, a -mall hor-t ---hoc or \'--h.tpcd chro-

niii-omi- and a pair o| \»-r\ l»i \ -mall \'--hapt-d un-

paired chromosome is also found in one hall o* t lh \\ Inch

corresponds to the unpaired hook ii • .'rot us.

In a lalrr paper, 'lO, I'.ah/i i dt -• lilu-- fliaraclt-ri-lit elrmt-nl-
in the chromosome groups :nnl<in> and .1 rhm'in

pustulosa. riirimnil" matic chromosomes in ^ 'I'liinns

i- toi i\ -two. All are rod --h aped. T\\o -omalie pair- could ln-u -
c\t-r l>e di-tin;cui-hcil in all of tin . ggs \>\ reason of tlu-ir exi
sive length-. \ ..... hi chrt>mo-ome could I >e demon-trated in thi-
-|n-( it-- dm-, no doiil't. to tin- lack of difterent iat imi in form of the
mi nil M -i- of i he -rou|). . ! -h. '\\eil a numlier of -mall hook-

and I --ha]>ed chromo-oiiie- hut the nnla\ oral >le nature of the
pre\ciited detailed -tud\ . The -omalic mi ml H r i- forl\ . The
c-hromo-ome- in thi- species n--einl>lc tlu»e ol Arbacia fuim'tnldtii
li;^nred l.y Ten in n' . '07. in that the\ a \\ hole Comparatively

short.

EDITH 1'INNEY.

In Kjlo. before the appearance of Balt/er'- later paper. Miss
llciiiHT published observations made in this laboratory upon.
Toxopneustes i'<i>-i<':>ati<s, one of the American species \\hich has
pn>\rd -ii valuable in cross breeding experiments.1 She found
in straight fertili/ed Toxopncit*: ; somatic pair ol long

rod- approximating in length tin- long rods ol" Sphaer echinus.
II ill' of tli. eggs contain t\\o and half tlmv Y-shaped element-.
Tin- impaired Y is undoubtedly the accessory chromosome. The
chromosome number in 'ro.\-<>f»icustes is thirty-six, the same as in
mgylocentrotus .md Echinus. Miss HelTner also examined
Arhddn pituctnlatii but found the eg^s unfavorable for di-taiK-d
c\ tolo-jcal -indy, a fact which forms another point of roemblunce
In tween the American and European specie- di-cribed.

The present paper deals with two additional species of echi-
noid-, Ilipponni' csculcnla (Tripneustes csctilciitns) and Moini
titropos, M\- special object has lieen to determine whether any
or all of the chromosomes exhibit any of the various- expressions
of individuality which have been reported for the other form-
mentioned above.

I am indebted to Dr. Tennent, at whose instance tin- work \\as
undertaken, for tin- material used and direction during the course
of ii- investigation. I wish also to express my gratitude to Dr.
Stevens for her interest and helpful suggestion-.

Hipponoe esculenta.

I'ln- Hif>()nn<H' material was collected and pre-erved at the
Tortu^a- laborator\- of the Carnegie Institution in the summers
of I'jo'i and [i)lo by Dr. Temu-nt. It was pn-parrd in the Usual
manner for study, the onlv -tain u-ed being Heidenhain's iron-
hsematoxylin. ( )b-er\ ation- made on tir-i and second cleavage

-|iindle- form the chief ba-i- of my concln-ioiis.

The character of the IIif)()nnot- spindle di-lax-ed somi-v, hat the
solution of the problem at hand. The cytoplasm of the e.^u is
bi-autifully clear and the chromosomes conijtared toother echin-
oderm chromosomes are of -ood size, but during di\i-ion tlu-N-
are so crouded on the spindle that the\ ob-cnre one another.
This may be attributed to several causes. It may be due to the

1 Tcnnont. '07, ' i <m. ' i 06 .ml ' i i .

• IIKOMOSOMES OF HIPPONOE AND MOIKA. I 71

thickness of the chromo-ome- which i> a -pecitie character.
Figs. la, ib and 25 show equatorial plates of Hipponoc and Moiru
re-pectively, and \<y comparing them one sees that the Hipponoc
r<«d- are thicker than those of M«ira. It may 1 >e thai the spindle
radiu.- i- le — in Hipponoc than in other form-. A;.:ain an attrac-
tive force may e\i-t among the element- of one daughter i;n>up
and ihi- nii-ht cause clumph.-

Tht-n- are main reasons why the anapha>e stages are nio-t
lav orable t'or -tndie- of ihi- -ort lmt there i- no doul't that much
< an I"- g iiiH-d from a -unl\- of etjiiatoria! plate-. A- \\ill U- -eeii
from tin- figures n-S-rn-d to, ihe p^lar views of metapha-e Stag* -
are iii.il»U- \<>r determining the relatixe thickiu» of

the chroiii<i-oinc~. although a- 'I'emu-nt. '07, ha- pointed out, the
contrast i enough tn ju-tify tin- i-\clu~i\<- u-e of

i hi- (ha racier as a mean- of distinguishing between the clm>mo-
somes of two in h\ Nrid-. Tl ts of the i ontracted

spireiin- in metaphaS4 have not \«-i lu-i-n inlluenced 1>\ the
>pindle iil'i-r- or the fora \\hii-h produce- .li\i-iou. whatever

thai ina\ be, and th' i uniform thickne--. ^nch

i (|uaioij,il p! iposed i--h iped chromosomes oi

\ai\in. 'i- I'lieN in.i\ IM i. cur\i-d urexwn lieiil at

\ai'iou- angl( In tin- lait-- ii . "iild not l.c deiermined

\\hellier ihr resulting \'-like i hroiil- >-o!in-- uere to lie iddllillid
uilh tin \ '- toimd in tin- di\i-ioii si \< iilu r i- it p"--il>le

to del ermine ihe 1 1 ngth or it i in ; the rod-. ( 'loudinv^ and

«'\ eilapjiiir^ in -ouie parts "I th« jilale pre\eiit- the -eparatinn

ot ihe element- Of theil . mea-lirelllen t . It llo\\ < \ el" t he

cliromo-. une- maintain ci'ii-iaiit -i/e relation-, a- u>- -Imuld • \-
peci fnnu the alum-lam e > •! e\ id.( nee .in thi- point gained from
lorm^ m«'re -uilalile fo-- ii- <!em. 'ii-t rat ion. and since, as I have
>ho\\ n in my li^un -. the thick; i!n r«-d- in thi- Stage i-

- 1. m l . it ^eem- that the n lati 'h- of t he chnuno-. ,nn •- at

(hi- mu-t l-e c.Mi-tant al-o and that in the -ul>-ei|ueni

di\i-ic,n si the thick I the p>d- does e\i<lenil\

\ai\ the l-'iuih relation- Jso. Thi- I c.m-ider .1 ]ioint of

l^reat import. nice since i: i- <>n ju-t -uch a compari-on of ihe
len-th- of the chroino-ome- in di\'i-i.-n Stages thai lialt/er. "lO,
<le|>end- for much of hi- evidence as to the identity of ihe elimi-

! 7 - EDITH PINNEV.

naled chromo-onu - in tin.- hyhrid-. Kilt/er make- IK > com pa rise m
hetwcen tin- equatorial plates of the species thai In- studied.
In hi- figures of aiiaphases of different species the daughter
chromo-omes an- -hown as rods of equal thickness. IK- regards
the apparent variation- in tin- thickness of the rods in seciion-
staincd with Heidenhain's iron-haematoxylin as tin- fault of the
stain and hi- c< >ncli!-ion- as to tin- uniform thickness of the n>d-
are drawn from observations made upon aceto-carmine prepara-
tion-. ( )nc would naturally expect to find the rods which lie
farthest from the surface of the section retaining the stain longest.
The deeper lying rods might then appear thicker hut all of the
rods K'ini; in one optical plane \\otild >how the same width.
All of the rod- in one optical plane, howe\er, an- not of the same
width. It might In- argued that a di-turhance in the section
alter staining or variations in the thickness of the- sections would
produce thi- effecl hut tin- variations in width an- of too frequent
occurrence to he a-rrihed to any such causal factor-.

I am convinced from my own ohservation- that the rods of
one -pindle not only vary in thickness as they are drawn toward
the polo hut they vary irregularly. The explanation is simple.
I hiring division the proximal ends of the two halve- of a rod
are drawn toward opposite poles while' the distal ends are still
united. This latter opposing force results in the lengthening
ot the two daughter chromosome- during their separation. As
soon a- the separation is complete the rods hegin to contract.
Since some rod- are much longer than others the accomplishment
oi their separation and their subsequent contraction would
naturally he delaved and their relative lengths in a late anapha-e
would he more apparent than real.

For illu-tration I ivler to Ralt/er'- later paper, 'lO, Fig-. T, a
and /;. In these he sho\\ - a Strongylocentrotus spindle with only
two rod-shaped chromosomes over eisjit mm. long. The l\\o
-pindle- sho\\n in hi- Fi^-. 4, <i and . and u, n and h from the
same -pecie- each show three rods which exceed eight mm.
On the other hand Fii;-. 5, n and /». sho\\- thirteen and Figs. 6,
a and b, ten chromo-ome- from one spindle \\ Inch are mon- than
ei-ht mm. loi

Again P'igs. 5, a and !>, -ho\\ om- chromosome over 15 mm.

i HROMOSOMES OF HIPPONOE AND MOIRA. 1 73

while the- next lon^e-t measure- only a tritle over 1 1 mm.
If as we \\ould naturally -uppo-e, the two loni;e-t in thi- <
torm a somatic pair, it is demonstrated that tin- force which
causes stretching during divi-ion and contraction after the
daughter clement- have 'ed, may work very irregularly

indeed and thai the chr<>iu< .-< unal length- taken fr< >m the-e

la lb

I '•• // ' . the 5J

•

.HIM1

in a \ > rv ir D 'iin'ari.MUi. Thi-^ i- imi

iiicanl tO impU lliat len^tli • in idi-nl ilica I i. .n i- tu be
elilireK elilliMial. d.

In ihe lal« hree I | I chr'imii-umi^

ap|'iar. ( '• .limn •iu--l aiiiMii^ ih "l--haped i-lenieitl-.

\ \\ ill lie seen !r..ni I . nd 4/1, n.mc ,,t" t!

are ••! i« iu.uk. i1 1. I. ngth. In I u. 4'.' i- ^i\<-n the <ml\ case thai

I I'lUiud ill \\liich a Imu i"«l appeared. Tin- iiiui-ual l«nii; chro-
m"-iniir \\mild ii'»i n-c.|\i- iiM-l! int-i I \\ < • \\ilh the IIK.-I card'ul
t", UHIU. 1 1 - i-i -laii-d i ., c i ; n. in e iu-tilie- the interpret a I i'>n that
it i- an ilhi-l rali. m , ,| the u-iide! ' chromosomes I"

clump l- i-^ri \>-

In IUMM- "t m\ figures "t // •• ill "i the riKJ-, -hi)\\n.

All attempt- in separate the chn>m»-"mal complex into it- unil-
\\ere un-uccc--tul. l"i^-. 2<i and 2:> -Imu . me -pindlc in which
the chroiuo-dinc- \\ere uiiu-ua!l\ \\cll -cparated. Tin- \ariation
in the tlu'ckm -- ..i the rod- ua- repurti-d |i\ I lel'l'iier. 'lO, t'or
7'c.\v'/>;/.'/^.'i-\. I ti'd V' illu-trate the same tact I'or

7//7'/'"""1'- \' 'II-.M \ i- cMivmclv dit'ticult hieve in the

dra\\iu^ of >uch minute olijcct- a- F.chiuoid chromo-oiite-. All
- taken to indicate the true -i/c relation-.

'74

EDITH I'INNEV.

T\vo somatic pairs of Y's occur and these an- shown in 1 ;u~.
5, 6, and 7 to be of two sizes. Their demonstration was not
easy in the late division stai;e^ for as Miss Het'fmT found in
. the arms of the Y's lie parallel. In IlippiDnx- one

2a

2b

h

H

tin/,

3 a

v V v

)

V

\ W

H

3b 4 a 4b

FIGS. 2a and j/'. II i t>t'«>""' • One spindle in anaphase from two sections. l\\"
pairs of \ - .md a Imnk are present.

FlGS. 30 and 36. Hipf''"1" • Saim .. i . j,i and 26. //. tin- lmi,k--h.i|»-d
chromosome.

FIGS. 4aand 46. //;/'/"";( • Lateanaphaa Not all of the rod-shaped chi
somes were drawn in these figures, .\lldia\\inu-ina.i. ,,i ., mavniiH .uinn oi i
diameters.

CHROMOSOMES OF HIPPONOE AND MOIRA. 175

meets with the additional difficulty c.m-ed by the crowding of
the chromosomes. For their study early anaphases proved ii:
favorable. A reconstruction from one of the-e i- -ho\\ n injFig.r6
and the \ "- from one are drawn in Fig. 7. The somatic pairs in
these figures are easily recogni/ed in this stage from their -i/e
from their behavior. In the case of the p.iir*of_lar-i-r

'H

rt'ff <

78

Ml

13 14 15

• •••-

16 17 18a 18b 18c

M I .lIKlpl.

i-i'. ' // : in early anaphase showing the relal

: the \ - md h

I lunik i • -.•Hi-.

1-n. s // . d lic'-ik.

I-l'.^ 9 to I? indusi> Iti' tu"-.nm.-.| i-l. •mi-lit- In- 111 niin- .|il-

I'-i.: -ii|..u.iti\«- Ir.-i|urii, y with wliirli tin-

iliiii-n-nt i-l.-nn ii- lh<- lii u.k i^ tin- m..~! i nii~|ii< unti-. l-'iv;. i^ i-

\\ lii> h . i.iit.iiin-'l n. i lii.uk.

I'..|.ir vii-u- ..i 'l.tiiclui-r \>\ m.l

M • : •

V- one arm . .(' the \' rompleie- it- di\i-i..n first in almo-t e\ . i \
in-tanee. The <iue-tioii a- to whether this i- due to a meeh.inic.il
hindrance in di\i-i<m or to a real diliereiiee in the -i/e of the I\\n
arm- ha- been di-eu— -ed b\- Miss Heffiier. I am inclined to the
opinion that the t\\<> arm- are of different lengths since that

I -I, EDITH PINXKY.

appearance predominates. Fig. 3/< is a typical \ lew. The lai i
pair i- racily identified but one can lie MIIV of only one Miialler \'.
Its mate may be the Mnal! thick element, r, lying iu-\t to on<
tin- larger YV In Fig>. II, 12, 13, 15. 1 6 and 17 some of the
Vs from several otlu-r spindles are shown. Owing to individual
differences in tin- eggs it would hi- impossible to identify the
large or the small pair as such unless the other pair could be
distinguished.

Miss Heffner did not observe any constant difference in size
or behavior which might mark the unpaired Y in Toxopneustes.
The size relation of the Hipponoe Y's is very plain in early ana-
phases as I have already explained.

I ha\e examined sections of Toxopneustes eggs made from the
same material that Miss Heffner used and am able to confirm
her observations upon the appearance and behavior of the Y-
shaped elements. The long rods are also very conspicuous (Figs.
19, 20 and 21). A peculiarity, presumably of the egg, calls for
different degrees of differentiation in staining early and late
anaphases. I could not find on the slides that I examined early
anaphases where the condition of things could be made out satis-
factorily although there were plenty of later stages which were
as clear as one could wish. In these I have observed and give
figures (20 and 21) which show a difference in the size of the Vs
which would indicate that the unpaired V is the smallest, yet I
hcMtate to accept this alone as evidence that such is the case.
Fig. 21 shows two large and one small V in the upper group. One
large V is missing from the lower group. The indications were
that it had been displaced by the knife in sectioning. Miss
Ilellner's I'gs. la, and 3, a and /;, depict two spindles in early
anaphase. From these figures and from what I find in Hipponoe
\\here Mich stages are comparatively abundant and convincing
I feel j u Mi lied in concluding that the unpaired Y or heterochromo-
some in Toxopneustes is the smallest V of the complex.

In Figs. 3, 9 and 10 is shown the- third type of echinodeim
chroni"~<ime, the hook. It corropmids in all aspects to t he-
hook described \>\ Baltzer. In !Iip/>onoc it occurs Miigly and
is not present in all of the c.uv^. ( )f tuent v-nine ci^s in which
its presence or absence' could be determined with certainly seven-

. [Mil i\K .-• 'Mi - ' •! HIPP! >N< '! \V' M' UB \

77

teen contained tin- hook. In late stages it is tin ni«.-t conspicuous
element of the complex and is therefore quite easily detected
when pn-ent; hut when it is not found, one cannot always be
sure thai it i- not hidden union- the other chroinoM >me-. In

• mining the eggs then more of tho>e which lacked the hook
uould lie rejected as po-iiive r\ idence than of ih<.-e in \\hich it

nrred, SO I l>elie\e we are -afe in asserting that half of the

f

• •:*'••*•*
•::••• *

•
• . *

- • •

• •

19a 19b 20a
/.

20 b

22

•:•••.

/

^'ni

21 a

23

24

21b

. !•;.! .ill. I I -A .lll.il-li.i-.- -Iiii\\ini4 t!

!•':• .iii.l 20 i ..ii-pinnm- rlinnn. • ..m a l.n.-

FIGS, a ia ai .' • te .m.i|.l.

I' IGS, .'-• -• i .Hi. I . P .| .|;iii-lit'T |>l;i(. iK.-

III. i- .in. \ ' .li.llllrt.

lertili/ed eggs contain the hook and in half it U al^ent . A- I >r.
I 'linen t, 'l l . this journal, ha- -liou n, i In- diniorphi- in in -oinatic
chroiiioMiiiu- i;rou|)v i.f fertili/eil Ilipponn,- . ggs an In- associated
only \\ith a iliniorphi-in in ihe -perm.

The deci-ion h.i\in- lu-.n reached as to the differential char-
acter o| tin- hoi ik-^h.iped chromosome, the (mention aro-e as to

i;<s EDITH I'INNKN .

the nature of its morphological relation to the other
All such elements of which we have microscopical proof, belong
to t\vo classes, tin- accessory or unpaired chromosomes and tin-
paired hetefochromosomes or idiochromosomes. This does not
take into account those cases like Ciilex and Tlien/xililin where no
differential element is visible in the sex cells. In both of these
classes the germ cells of one sex hick a definite chromatin mass
which is present in the other. The single or unequal pair of
chromosomes of the one is replaced in the other by an equal pair
both members of which correspond in size respectively to the
single clement or to the larger element of the n/ie//i«il pair. In
Hih()ono'c the search for evidence on this point revealed no pair
of chromosomes equal to or exceeding the hook in si/e. This led
to the conclusion that a corresponding pair of hooks does not
exist. It was then thought that if such a pair was present its
members were probably ordinary rods. No rods equal in size
to the combined lengths of the two arms were present in any
of the eggs. The question as to whether the hook was a multiple
chromosome was considered. This probability which I will dis-
cuss later, suggested that perhaps the differential sex element
consisted of only one arm of the hook. No rods equal in si/e
to the long arm of the hook were discovered in any of the eggs.
If such rods were present they might escape detection in main-
instances but it is improbable that they should be so consistently
over-looked in all of the eggs. In Figs. 2a and 2b an attempt
was made to draw all of the chromosomes from one spindle.
The hook and V's are plainly visible. All of the elements shown
could be separated by careful focusing but no rod longer than
the arms of the large V's was observed. It appears highly
probable that the long arm of the hook is unmatcd in somatic
cells containing the hook and that no corresponding pair of Ion-
rods is present in cells which contain no hook. It would be im-
possible from this material to determine \\hether or not the short
arm of the hook was mated. Main short rods approximating
it in length occur but the\ could not be studied mdi\ idnally.
I have failed to determine ihc number ot chromosome's in
these eggs. Figs. i8a, i8b and is, gi\e some of the best polar
views of daughter plates. Fig. is,/ shows thirtx -three bodie-

CHROMOSOMES OF HIPPOXOE AND MOIRA. 171

one of which appears a- though it might coiisi-t of two or more
chromo-omes crowded together. I was not able as were both
Ileffner and Baltzer to identify with certainty the Y's or hook
in such views. As these observers have explained, tin- onlinarx
rod- -(.-en in polar views appear as -mall black dots. Two of tl
in contact arc to In- interpreted as the closely apprised arms of
tht- Y--haped chromosome. Mi-s Hetfner's figures of the polar
viVu- of daughter up'iips of Toxopneitstfs chn >mo-.< mie- an- ver)
r ami her interpretat i« -n of them i- undoubtedly correct.
I', 'I/IT aUo linds tin- V's in jtolar \ie\\- appearing as \\\<> closely
MI dots. In p«l.ii- views ol /fippniioi' I ha\e often oli-iT\e<|
\\hat I al lir-t -ii|ipo-t-.l nii-hi In- ,( \'. Hr-ide- tlu- >mall round
doi~ ?\pic,il of the rnd--h.ipi-il • hroiiio-onu--. one tind- in e\ ery
plaie a number <•! ellijit ie.il I >odi<-- ot twi ce» the size ol the ordinary

do: The-t- one naliiralK inter|)r« i- as ilu- t\\o arm- of a V

\\iih theii .-in surfaces il.nirned lo-rther. In some cases

a MIX (\idnii roii-i rielion ci lincident with their narro\\r-.|
diameter makes -;:• :i an in:' 'lion imp«-iaii\e. Thi- con-

st rit "tii m i- not however evident in the majoriix of cases and v<

often nmre ••! tin-- illiplic.il bodic- appear than are calli-d lor

b\ 'lie mini! -nrd i-lelllelll- ill IHf)f)iUl,' I . I

In .ill cases th( n the <|ii<--tion i- -till open a- to \\liether llir-c are
\ '.id'-haped indi\ idii.il- or even, as i- unite po— ible,

oiil\ one rod \ ii-u ed obliquely. The latter po^ibilit v in add it ion
i" the dillicnhv |ire-enti'i| by untranslatable amorphous ehn>-
matin Imdies in these d.u in.ikr- an .iccnr.ite count

impo^-ibli fhe number counu-d mo-t treiinent 1\- \\a- thirt\-
t\\o but in no • mid I I « -i 1 -nn- that the number counted was

con

M

Thi 'I Moirn i- much better -niled to detailed -tudy

than the I-L;^ of ////'/• I'lu- tendeiicx of the chromosomes

to ero\\d together i- not so pre\alent. .\lthoiii;h my mall-rial
\\a- \ er\ limited and contained comparative!) leu -pindli-s -.,nie
general facts W< • In-red \\hich il ma\- be worth \\hile to

•rd.

5 polar \ iew of the ei|iiatorial plate of the lir-t

Mu-ntation nucleu-. A- alreadx- stated the chromosomes are

IX'

EDITH PINNEY.

nmre slender than t!u>-i- of HippoHor. Fiu. 20 i- a -ection through
a. metakinesis stage. ( )nly a lC\\ of tin- chromosomes are shown.
Three tvpcs appear; the ordinary rods, two mial! YV and t\\o
club--haped elements. In Figs. 2~ and 2N. which are analytical
drawings < it" two early Moiru spindles, the same three types appear.
It could not be determined whether the club-shaped chromosomes

I'M.. 25.
!•!'.. 26.

\1:>irn.
Mnirii.

plate from first sr.ntm-iitat i< MI
anapli

FlG. 27. Moirti. ('liniiiiip-niiii-> limn an r.nl\ anaphase. A", honk?
FlG. 28. Mtiiru. Same as 27, Mai;nifh atinn, i .5110 iliaim ; .

were small hooks or not. Tin- chromo-ome X in each of tin-
figures mentioned above ha^ the appearance ol'ien p|-e-enti-d li\-
the Hipponoe hook. Tin- -mall \"> are iimneroii> Imt their
number is not e\ident. AlthoiiL'h the Moini cliromo-ome^ are
comparatively well separated on the >pindle the fact that the
number is not the same in all of the fignro i> e\ idenci- that e\cn

CHROMOSOMES OF HIPPOXOE AM> MolKA 1M

here mi-takes in drawing are almost inevitable. It may be that
-OUR- -ho\vn are really two chromosomes so close together that
they could not be separated with the magnificat i<m u-ed.
The serie- i- al-o taken from two or more section- and there is
the ever pre-ent | >o-- il ijlit y that some may have been di-placed.
! igs. 290 and 2<)''< -how a Liter anapha-c of which Fi-. ,v> i- a
deiailed drawing. A lio,,k (x i- -ho\\ n to la- pre-cnt. but many
more observations would IK- n - ry to demonstrate its prc-cnce
be\ond a doubt. The club-shaped type of chromosome is nol
in e\ jilciii-e h-re.

A- was to be I rom lateral \ ie\\> of anapha-c- the

only d. ui-hter plate- I found \\civ e-pecially adapted to ail

• i enumeration < .f th< ir element-. The number i- -ho\\n in

Fij ,t</and i vhich are daughter plates of the same spindle,

to I ;\. tlie lar^e-i number \et reported fop the eeliino-

<lerm-. I :. i draun from t \\ •• sections. The charactei

ot the Mn-.-'ti -])indle i- -ueh that I eon-ider the e\idein'e as

to the numb' 'ironio-on. ned from this one plate much

Illoie ( on\ iiu in- th.ui all of the eo 1 11 billed e\ idem e of a like -oft
\\ Inch I obtained I mm // . I hope \» extend the ob-« r\ a-

tion- on thi- species !.•'
The Moira material \\a-coilccted at l-le.mfoi -t . \ . < '.. in 1907.

It- treatment \\a- identical \\ilh that of //;'/'

1 >!-. i SSN »N.

I rom • oin -lu-iou- ed b\ lialt/er. '09, and Temieiil, 'il,

it seems probabU- that in different spC( ies of a limite.l -mup the
hetei'ochrom..-. .me ma\ "ii-inate in dilleretit Both o|

the-e author- u-eil the -auie method oi in\ e-t i-at ion so th.it the
e\ii|enc(- o| ..lie bearing on thi- point po- no peculiar ad-

vantage over that of the dher. Both are equally in-tilled in
their conc!ii-ion-. |M Strongyt '!ns liridit* and I-'.clnnns

niicrdtntn-rcnlutits. it i- the female that i- morphol, .-icallv the
helero-ametic -e\ \\hile the male i- in tin- -ame -en-i- honio-
^ametii'. In the li-lu of i.ur pre-ent kno\\ led-e of the Occurrence
of the .r\ chromo-ome in other groups the-e fact- pre-ent

an intere-tin- aiiomaK .

It as Balt/er found in aid a- Mi-- lletlner

EDITH riNNEV.

found in To.votvit'iistt's the somatic number of chromosome- i> an
even number in both sexes we must conclude that the odd chromo-
some is the eccentric member of an unequal pair. This of course
implies the assumption that in the odd chromosome \\ e an- dealing
with a univalent and not a plurivalent element. The number
of chromosomes in Hiplwnoi'- may or may not be even. In the
five species of sea-urchins already investigated the somatic num-
ber has been reported as even. The conclusions have however

%*

• • •

» ~ *

.%

• •:

31a 31b *9a

'"'I/,

30

FIGS. 290 and 29^. Moira. Anapha-r from two
I-ii.. 30. Moira. Chromosomes from Fix- !>)• -V, hook?
FIGS. 310 and jib. Moira. Polar views of two dauulitn plates from tin-
spindle. r chromosomes. !•'!>;. ,}:'< is from two sections. 1,500 diameters.

been drawn from axtTa^es of main- counts. Since the counts
of single plates \aried an exact statement as to whether the
diploid number of chromosomes is the same and even in both
3, Of differs, beini; e\ en in one and odd in tin- oilier, seems
hardly justifiable.

In comparing daughter plates of /fif>/)oni>i' with those of
'I'<>.\t>f>ncnslfs I found as a rule that ihe chromosomes in '/'t>.\-<>/i-
>ir null's were better separated. K\ en ilu-re tlu- majorit\- of counts
could not be made \\ ith anv assurance of accuracy. Mi-- I let'fner
reported thirt\--six chi-omo-ome- for '/'».\-n/)iii'iistrs. I found four

CHROMOSOMES OF HIPPONOE AND MOIKA. I x ;

plate- which -howed thirtv-ei^lu without any doubt. Tlmv of
them an- -hown in Fi.u-. 22. 2 ;, and 24. I al-o counted thirty-
seven in two very clear daughter plates of the same cell.1 1 made
.: tew rount- \\here thirtx— i\ -eemed to be tlu- number Inn in
tin - I was not -i in- that I had counted correctly. Other

< ount- were made lull they \\ere equally valuele--. Tin- Toxop-
ncns!i's inait-ri.il \\a- too limited to permit definite conelu-ion-
as to i In • art n. il nil nil M -r-. I am t • m\ inred that morr than thirty-
-i\ < lironio-onie- occur in some eggs An .irtii.il determination
i- dilhi ult luit not impo— ible. .\\t-ra-e- in a ra-r of thi> M»rt

\\ hatr\ i r i In- morphological rrl.nion- of the In >ok to the otlu-r
mi-mlii-r- o| the roin|.Ie\. thrir nui-ideralioii lead- u- toa rero^ni-
tioii o| |ni— iMe e\i-teiire of further aiKunalou^ iniidition- in thi>l
Bpecil All unequal pair- o| heterOchrOmOSOmeS Or unpaired

heterochromosomes ol pi- »]'-i rvation find a |>lare in the

M-rie- po-tulati-d |p\ MtAi-n-. ' I I. \\hirli. liei;inninu \\ith ( 'nlfx
and Theo :'•::•:. v. : ichromosome occurs, includes all

«>t iln \aiiou- mn-i|iial pair- and end- \\itli form- ("iitainin^ an
odd ••!• unpaind rliroiiio-onie. \\ r ma\ imagine all of the mein-
bers in thi- series di ii\rd !r"in f'-rm- analo-.iu> to Culcx \>\ ihe

dual dim in in ion and Imal elimination ol one chromosome Iri-m
|i.iir. A — iimini; thai the hook in ////'/""""' i- 'he
nic.|-|)hoi :r.il ei|iii\.ilent "t the odd ehi'omo-ome or the l.u.
meml>ri Ol i inn ;n.tl |>air il follou- that thi- element eannol 1 >e

"iinletl lor in thi- manner. < >n I In- roinrar\ WC mu-i conceive

• •I the i haii^e iri-m the < heterochromosome i\pe io ihe

7/;/'/"1""' 'M"' '" ha\e COme about l.y all ilirrea-e ill the -i/e
of one member of a -omatir jiair. It i- i|iiite a- reasonable l«>
-up|>o-e that the e\olutionary processes \\iihin the nurleii- in-
\ol\e an increase in the amount of rim-matin a- it i- to a — nine
that the\ are a. v unpanied b\ it- elimination. In -up|)ort o|
thi- \ie\\ \\e have the (art that the hook i- the lar^e-t element

in the ////V'"""' complex.

i: \Yil-.-n. in .li ' XII.. T.U. X\"!..

.l.iu.clH' -h"\vini4 .57 >. '"' "'" -"I"'-- ' '"•

i-\|il.iii.iti.-n "I th'- liu'.ir -ii.it tin- nuinl.i-: i-int'-i-

.•-11111; in ihi-i ouuii'i-tion. In carln-i p.ip.-i- li>- rai-.- 1 iln- qu t" \\ln-tlH-r

tin- niunt.iT \\.i

EDITH PINNEV.

I'.altzer has suggested thai tin- hook i> a ] >luri\ alent element
composed of two chromosomes united at their polar ends. Ad-
mit i in- such an interpretation it -till remains probable that
Hipponoe cannot be inclii(k'd in the above series since it was
impossible to rind for the long arm of the hook of some eggs a
corresponding pair of equal rods in others. The long rods of
Toxopiiciistes are so conspicuous in late anaphases that it seems
,i- though the presence of a similar pair in Hipponoe could not
be a matter of dispute. Figs. 19, 20 and 21 show anaphases
of Toxopiu'iistes containing the characteristic long rods. These
Toxopneitxtcs roil- are slightly longer than the long arm of the
hook and the 'groups to which they belong arc perhaps not so
compact as daughter groups in Hipponoe.

With Baltzer's suggestion under consideration the question
.irises as to whether the V's which resemble the hook in all respects
but length of arms are also to be regarded as multiple chromo-
somes. If so the method of grouping in Hipponoe differs from
anything previously described.

The idea that the heterochromosome is associated with the
inheritance of sex is a point of common consideration when
treating of this differential element.

From the experimental evidence as to the inheritance of sex
Castle, '09, has concluded that femaleness depends upon the
presence of some factor wanting in the male, the differential
factor. Two classes of female zygotes are recognized in CastUV
scheme, one, class A in which "femaleness is attained only win n
the differential factor is doubly represented in the individual"
and another, class B, in which femaleness is attained whenever
the dittereiitial factor is present in one only of the conjugating
-ametes which produce the individual." The former for which
Wilson gives the sex formulas; XX female and X :- male,
X in thU case designating the differential factor, is represented
by the insects. Castle says, "Direct cytological evidence of the
existence of class B is not known at present." The formulas,
X< ) •-• female and ()= male einl iod\ ( 'axle's idea.

IlippnntK- falls into class A only upon the assumption that the
1 look is a multiple chromosome, the Ion- arm hein- the differential
element, and that there exists in one-half of the fertilized eggs a

CHROMOSOMES OF HIPPONOE AND MOIKA. 1 s 5

pair of rods the same length a- tin- long arm. There is no obser-
vational basis as yet for either of the-e a— umptions. Neither
dot- our evidence place Hipponoc in class B since Profes-"i
Tennent's hybridization experiment- -how that the hook must
come from the male. If however the hook is not a multiple
chromo-onie the ra-e i- analogous to < la— H, the >e\ formula-
rrvi-r-t-d to indicate that the male i- a hetero/\uou-
and the frmale a pure I ,<. XO = c? ami 00

= 9 . Tin . onclu-ion i-- the -ame if we identify the differential
dement \\iih t In- 1« MI- arm of tlu-hook and a>sumr that none of
tin oiitain a corrf-|iondin- |>air of lorn: ro<l-.

1 If.irly our i !u-ii»n- as t" the eompo-ition ,.f tlu- hook and

it- iiu-anin^ imi-t 1 I until our kno\ has been in-

: \>\ further evidence gained from thi> and otlu-r -pt-t -\< -.
M> i lung's views upon th( -loniif \ahu- of tin- chromo-mnal

• onipli ihat the >tud\ -of other species of the same ^t-nus

\\oiild |>ri-\t- most profit. ilili-. From a eoinpari-on ..f the t \\ o
species "I the _emi- .1 -crilu-il |.\ Tt inient and lialt/i-r

one tin(U -imilaiitie- in the ( hrom. -otnal ;^roiii»> \vhiehlend
nifit an« e to thi- -ir^^e-tion. liroadt-r c\ t< ilo-ieal >tml\' of the

echinoderma is also necessary in order to di- 'he fundamental

prineiplt- \>\ wliieh \\r i an refmi ile ->urh di\er>e plu-nomi-n.:
lho-e |(it-eiiied } <\ and h'.chiniis on the otio

hand and // on the other. The diflieultii-s which attend

tin -tinl\ of e. hinodei in ehronio-onif- make- it dr-iral>lr to
extend tin nunil't i . -f \\orkt-r- in tin- lit-ld.

mm ii 'f.K AI-IIN
Baltzer, F.
'08 i ; Seeigleiern. \Vrh. .1.

.1 /

'098 1 >n- i In hv. u. 1 Ai. li. I. /••!

II.

'ogb i -uiikhi!! • -.ini'lt-n-l- B litivuiiK

ln««iii.itiiiv«-ih.il;ni--' /.. H'l. XXX\'.

'10 I'lx-i .lit- !<••/!. -lui: ii'-n ilnn ('liiniii.itin u. <1. Kntwii kluiiK u. \"rn-r-

l)iiiii;srii-litin. I: i-t.ii.lni. At. Ii. f. Zellforsch., l'>-\ \'.
Boveri, Th.

'07 /rlli-Il-Ul.li'-ll. Mi-It VI .!• :

Castle, W. E.

'09 \ M' -ivlrti.ui Vi. . - Science. N. S Vol. XXIX.. N.I.

1 86 ED TH PINXEY.

Heffner, B.

'10 A Study df Chromosomes of Toxopneustes va:iei;atu- winch Show Individual

Peculiar! ties of Form. Biol. Bull., XIX.
McClung, C. E.

'05 The Chromosome Complex of Orthopteran Spermatocytes. Biol. Bull. I X.
'08 Cytology and Taxonomy. Kan?. I'niv. Bull., Vol. IV., No. 7.
Stevens, N. M.

'n Further Studios on Heterochromosomes in Mosquitoes. Biol. Bull.. Vol.

XX., Xo. 2.
Tennent, D. H.

'07 The Chromosomes in Cross-fertilized Echinoid Eggs. Biol. Bull., XV.
'ioa The Dominance of Maternal and of Paternal Characters in Echinoderm

Hybrids. Arch. f. Entw. Mech., Bd. XXIX.

'iob Echinoderm Hybridization. Publication No. 132 of the Carnegie In-
stitution of Washington, pp. 117 to 151.

'n A Heterochromosome of Male Origin in Echinoids. Biol. Bull. XXI.
Wilson, E. B.

'09 Recent Researches on the Determination and Heredity of Sex. Science,

X. S., Vol. XXIX., Xo. 732.

"10 The Chromosomes in Relation to the Determination of Sex. Science
Progress, No. 16.

Vol. XXL September, ryi i . No. ./.

BIOLOGICAL BULLETIN

A STT'DV OF THi: RELATION < >F TISSUE DTFFER-

ENTIATION TO RATE OF CRnWTll DURING

HI.' IENERATION.1

\V. K. AI.I 1 \.
A' K \i.\\ | | .1.1. Nil \ I-.

Tlii- work was don,- under the directi< >n nf I )r. ("li.i~. /eleny,
\\ho-e .idvire .md (Titin-m h.m- lu-eii in\ .ilu.iMe. I .1111 .il-o
indeliied to I •! II. I'., \\.ini |. ions .unl ciu-<uii

mm I . in Mr. S. I . Prince fur ln-lp u ilii the df,i\\ in--. t.» Mr. \Y.
liuli.iii.i 1 ni\ .1 nihl ystonnt ni.iii-ri.il cllirifin ]\

il .md l«. the lllin.'i- Si. uc l..il><>r.il"i v .,|" \.uur.il
llic ICMII "| \ alu.iMr liicr.iln

11 ECT.

The «il. • inK \\.i~ I., lind \\ hcihcr i he -i (-.iir-i -:

di regeneration occurs 1- it'in- »\- c«iin<-itlcin \\itli ilu-

i|ilcii«>n (•! ii--ur differentiation. If tin- -.nm- ciin-i-l.iiinii

- ili.it was imtc'l 1>\ Minn! [908 iii i.rdiii.in. uiM\\ih, \\(-
c\]'i.t in tiiid di. .\\tli dt.r<,i-iii- .tlu-r ihr

m.iinr tissues li.i\<- IK-I-II \\rll dc\ eloped .md -till more -'. .ifti-r
dillcrcini.ilion i- 'i.ilK 1 1 >iii|>lcic. In the hdp. ECUring

dftiniir information tin- prr-rni riii'iM li.i- lu-t-n m.idr ID secure

d.it.i ill lli.it drlinilr line, ti-in^ t u < > di-tinrt I>IH-- for >tndy.

|-.\n ivi\n \ i 1. A M \ 1 1 KI .\\I\IAI..

'I'm (Mi ...... .in ii \\'(>ivM. Lin:nnt}rilns clafxirctlidiins. l\.u/d.

Mil!, •> inl a ml .\!>'!li<x!s.

The \\ork \\.i- 1't-min in r.irK < >ct( .ln-r. KJIO. At th.it time
then- was not inurh fln.icr of Mi.Urri.il for >urh in\ c-ti-.it ion as .1

i ..nti ilniti«n< iniin : ' .

the .lin-.-tii.n ..1 Il.-niy li. \\'.u.l. N

[88 W. K. ALLKX.

preliminary survey of the local held soon showed. Lack of better
material was therefore the principal reason for choo-ing Linniodri-
lits. Of points distinctly in its favor, tin- mo>t prominent were
accessibility, hardiness, adaptability to laboratory conditions
and rapidity of regeneration. ( )n the other hand, the small si/r,
irrital)ility, quickness and sandy food caused some serious dif-
ficulties. The specimen- n-ed \\ere taken fnun a -andy slouch
margin just north of I'rbana Fair Grounds. All were in good
condition and in a few hours appeared quite at home under
laboratory conditions.

The effort was made to select 120 worms of equal median si/e,
but their activity was so great and their appearance changed so
quickly that the results showed considerable variation, ('.real
care was taken to make exact transverse cuts at the middle, but
here again, activity interfered and there was a con>idcrable
percentage of errors in cutting. For operation each worm \\as
placed on a paraffine block and a quick even cut was made- with
a sharp, thin scalpel. Anterior parts only were retained. Those
of the first five worms bisected were at once stupefied in weak
chloretone solution and killed in Gilson's sublimate mixture (Lee,
1900). All other worms were handled in the same way. The
second five was kept alive I hour after operation. All others
were placed in 10 cm. Petri dishes in which \\as about 4 c.c. of
sterilized native mud with 30 c.c. of tap water. Ten were placed
in each dish except by miscount due to disappearance in the
mud. Two dishes with unoperated worms were kept for check.
One hundred were bisected. \Yater was changed in the Petri
dishes twice daily immediately after taking the temperature of
the water already in the dishes. The highest temperature- re-
corded was 28° C., the lowest 21° C. The greatest duct ii.it ion
in 12 hours was 5°. While it was unfortunate thai laboratory
conditions were not such as to permit more uniform tempera in re,
it is hardly probable that even this variation seriously affected
these hardy worms, none of which died. Li^ht and other con-
ditions were kept as nearly uniform as po-,>ible. 1 )iivci sunlight
was avoided altogether.

As noted before the first five was killed immediaielv after
operation, the second in I hour, third in (> liour^. toiirth in 12

TISSUE DIFFERENTIATION AND RATE OF GROWTH.

I- ,

TABI.K I.

Limnodrilus.

Indivic

rm -f\e Old Length of \\ hole Re-
mber. ! merit in n . Part in •

1st dav

7a .III

1 v» . -f *.

•432

•057

-c .425

.1 I ;

;J

7«

.038

!.iv

7/

.08^

/ ' **T *

7«

L^V/J

7^i

•

7J

.IQO

^'1

8a

.

' 4)

Jtli '

-

sth

; i

Mli da]

•

i

7th iluv

i

i

.1 i

8th .1. i

i

[6

py ••!' tl. in niilliiin ;

\vllii-li (In- yn>\vtli r.it'- I.T tli.- t

'iinlur inlhi'-iiic up. in tl. i in piiH-ni!

190

\\. E. ALL1.N.

TABLE I. — Continued.

Litnnodrilus.

Individual \Vorm

Niin

1 i-n.^tli ..f Sinyli- I >1<1

'i_Mh ..1 \V|],.l,.- R,
i in niiu>..

Qth (lav

I 1C

• 4>6

- 418

nd

•*T J **

•456

/ • tr*r'J
S.OI6

I2b

.380

7.400

I3&

•494

8.132

146

•456

6.840

loth dav

ne

.-342

7 -1OO

I2C

• J'T"
•456

/ "+v-'v
7.068

I2d

•532

(5.928)

13C

.418

7.')80

1 1'

.380

8.626

nth day

1 1/

.418

6.^46

i2e

T

•456

O T1

9.196

I2f

•456

7.6/0

i3d

•494

10.754

I4d

.361

7-752

1 2th day

lie

O.I

o

I2g

y *

9-4

I3«

9-

I3/

9.6

146

9.8

1 5th day

III!

12.7

I2h

10.9

I3S

13.3

13*

ii. S

I4/

18.9

i 8th dav

lit

tO.Q

12*

•* ' • y

17--'

13'

I6.S •

148

t6.3

14/7

13-7

hours and fifth in 24 hours. After that a group of five was taken
each day to the twelfth day, the remainder mi the liftrnith and
eighteenth days. Since the specimens h.id i<> be killed for tissue
study it was necessary to calculate the rate of growth from the
condition in successive groups after killing instead of using
periodic measurements of living worms.

DA i \ .

The following sets of measurement \\ere taken:
I. The total length of the regenerated pai't \\a^ me. inured by
eyepiece micrometer under lo\v |)o\\cr .is ilie ^|iecimeiis la\ in

Day

TISSUE DIFFERENTIATION AND RATH OF GROWTH.

cedar oil in a watch glass. (Table I. and Chart I.) Crooked
specimens caused some inaccuracy here.

Chart 1.

' X /A ' v ^

/ / \/ \\ ; "\\

i i in

III. !
liinki-M '

:

2. At (In- -,mir liiiir .nid under tin- -.tun- o mditioii- an old
in-ill, alum! ti-nlli lV"in lln- lr\rl tit' cill. \\.i- inra-nivd ill
tirdrr l" ;^fi a lu-i- .i|i.iri-nu nl ^t-iu-r.il n-ult-. 1 .iM> I.

M nn-iiifiii- were m.idr at'trr inininiiiiu nu -lidi--. -MHK-
.it'trr -i < limiin- luit nn>-i in (.d.ir oil. In tlii- \\.i\ -unit- til tin-

errors dm- t<> crookedness were drtfi-trd thuu^h tlu- ^fiitT.il

rr-llh i- iim r--riili.ill\ dilk-ifllt. 'I'aMt- II

4. |-'.M-r\ tt-nili -r^inrnt <•!' tin- rand part \\a- int-a-un<l

I'm- Iriuth. I'al.lt- I I

As tin se uu-a-urruu-nt> fi>nn tin- only IMH- for llu- n m-t rut 'lion
of a pt.K-i.ii ('hart I. i<> ^ra]ihifally illu-trali- tin- rale of

192

\V. i •:. AI.LEN.

growth it is worth while to give them a little general discussion.
There are sources of error, which being well recogni/ed, have
been checked upon and guarded against as much as possible.
(i) The measurements were made and the segments counted
without the use of a mechanical stage. Since the specimens
possessed very few good guide marks there may have been slight
errors due to this cause. (2) Only one count was made in
simpler cases although several counts were made in doubtful
cases. (3) The correct measurements may be misleading at
times because of variability in the speed of regeneration due to
lack of uniformity in the bisecting cuts. This is due- largely to

TABLE II.

Limnodrilus.

4th

Day.

5th
Day.

6th
Day.

7th
Day.

8th
Day.

oth
Day.

loth
Day.

i itli

uth
1 lay.

1 >.iy.

i8th
Day.

R .625 .92

2.15

3-o6

5.26

6.94

7.09

8.38

9.29

I4-29

16.43

iRS

.031

.068

•093

.126

.150

.184

•213

•234

.212

.258

.221

loRS

.015

•034

.070

.099

.118

.167

.168

.2OI

.192

•2/3

.246

2oRS

.003

.018

.047

•073

.108

.141

.145 .165

.163 .2 |7

• -' - 7

30/?5

.002

.008

.022

•043

.101

.112

.122

.no

.142

.207

.197

4oRS

.Oil

.O2O

.058

.083

.097

.IO2

•123

.179

.174

SoRS

.008

.014

•037

.048

.068

.072

.092

• 159

.164

6oRS

P

.008

.017

.025

.O2I

.052

.076

.109 .145

•joRS

.008

.Oil

.012

.021

•037

.102

• 1 43

SoRS

.007

.014

.015

.052

• US

goRS .008

.012

01 J

.074

A copy of the averages from which the growth rate for the first and t«-nth
regenerated segments was calculated. A partial revision of the averages for th<-
total regenerated part is also included here. Measurements are in millimci
,R=length of regenerated tissue. iRS, loRS, etc., first, tenth, etc., regi-nrratol
segments.

Explanation of Table II. — Table II. is a copy of the averages, in niilliim i
from which the growth rate for the first and tenth regenerated se.nmcnts \vas cal-
culated. A partly revised list of the averages for the total re{n-iu-i.iti-i| pan is
also included here. A comparison of Tables I. and II., especially cotiMdrimi; thr
measurements of old segments in Table I., will give some indication of one disturl
factor in the polygon, i. e., varying sizes or extension of the w.um- at tin- mm- o)
killing. 7?=total regenerated tissue; i RS, 10 RS, etc., = in-t. until, etc., re-
generated segments.

the smallness and activity of the worms which made it i
to cut precisely across or at the same level in the segments.
Thus some cuts fell on the inter-segmental line, other- .n various
distances within the segment, and several \\rre diagonal. The

TISSUE DIFFERENTIATION AND RATE OF GROWTH.

193

-

\. pin hli.i.

•A

-

present.

-

c

:

-

u

c

lliiln.it ill

1 1 1 1 1 1 1 . 1 1 1 1 1 >

-

—

-
f.

~
.

-

-
.

u

U

-

u

bJ

—

-

0

>.

i

C

-

.

i

D

.

-'

-

f=

-

-

-

-

-
hi

-

_

-

2

s

r

c

=

c

.

_^

•

-r -

—

_g

-

.
_

—

- -

I

~

-

-

a.

- -

—

J

"rt
u

-7

-

5
cd

-

.

is

c c

I

s E

a

c

^

^ -
--

'

:

-

1

--

-
"
*

-
S
~

E k a a a i

.= .= .5 r = s 3

_ _ .~ hi «J «4 «J

s s s s

= E

U

£

E

3

~

—
a

a

fc-

|J

—

~

_

^

^

s

s

•^

__ '

•

•

i

—

E

E

0

0

^
—

^ —

M

4-1

«

3

a

"T

~ ^

E

E

E

*

~

5

<

-

-

•o

_

«

_

B

C

=

r

r

~

-

E

rt

g

2

-
E

ed

E

—

T

T

E

E

E

E

3

—
-

f

/

/

—•

-

c

r

:

-

194 W. E. ALLEN.

last point seems to be especially important as Miiller (1908) and
others have shown that diagonal cuts produce marked differences
in the detail of regeneration. 14) The state of contraction at
time of killing was also probably somewhat variable so that
detailed comparisons would have to be made \\ith caution. The
effort was made to test this factor b\ measuring tin- regenerated
parts with the old segment as a unit. This indicates greatest
speed of regeneration on the tenth day instead of the ninth
as indicated by the other method. (5) The small number of
specimens available gives opportunity for undue prominence in
the showing of individuals. Evident extremes \\ere omitted
from the averages. But, after all, the general effect of most
of these factors would be to diffuse rather than to accentuate
the polygon, and there seems to be ample warrant for saying
that the greatest speed of regeneration is at the ninth or tenth
day, which is near enough for present purposes. In close ap-
proximation to these results we have those from some unpul dished
work by Mr. Frank L. Pinckney, using Limnodrilus in this labor-
tory in the spring of 1910. As he used the method of successive
measurements of living animals, the similarity seems to be
practically conclusive.

In examining the polygons two striking points appear which
are rather difficult to explain. In the first place, there is an
apparent slackening in growth almost to the zero point on the
tenth or eleventh day, the most obvious explanation for which
seems to be in an accidental assemblage of extreme characters.
At any rate there was no change in temperature or other observ-
able laboratory conditions at that time that could possibly ac-
count for it. In the second place there is a rise in tin- later
portion of the polygon. This, however, in the total regeneration
seems to be due to the fact that there are still some 50 or (>o
very young segments, main- of which are just at their maximum,
thus serving to largely balance the decline of the < ilder regenerated
segments, which is not in itself, so very great by that lime. As
the study of tissue differentiation has been made principally on
regenerated segment no. 10, the polygons lOi segments no. i
and no. 10 are shown (Chart I.). The inaMiuum for no. 10
appears here at the ninth da\ Inn is apparently only more or

TISSUE DIFFERENTIATION AND RATE OF GROWTH. 195

less fortuitous!}7 coincident with the general maximum. Coin-
cidence of this sort is hardly to l»e expected in all annelid-. Tin-
abrupt rise for the tenth regenerated Moment after the twelfth

r

day i- e\ idently due to accidental conditions in the gn >u:

For the differentiation study, longitudinal -ection- were u-ed
almo-t e\clu-ivcly. All of these were sagittal or nearly so.
While cro-- -ection- would certainly have given lie tier hi-tological
detail they would have made it very difficult to interpret con-
dition-in gnu-nt-. The number of specimens favor-
able lor sectioning wa- too -mall to allow tin- u-e of l>oth kinds.
'I'lie general feature- of the rate of d i ffen n t iat ion in the rcgenerat-
iiu* p.trt ma\ In- readih -r.i-ped l'\ reference to Table III. The
' rookedni ome »\ the \\orm- and the -mall -i/e of all made
interpretation ol the -ection- rather difticull -ince -o fe\\ -ections
ill each -pci iinen could : a approximately radial. Then'
ua- the turiher dil'licnli \ . \\ith many, of the tine -and in the

ii.hiiu and clouding the -ection even when

it did not tear it. In -pile of the-e di-ad\ am ao •- the character-
IStics "I mo-t ti--ue- -ho\\ed preit\ \\ell their relation- to the
einliiAonic and mature < < mdit i« >n-. Ten ti — ue- and oi-^an- are
laiih ea-il\ di-tin^iii-lu-il in the mid Ixxly region of the normal
\\oiin. Iht '-e \\ill no\\ In- o.n-idered -eriatim.

Matnie cpidrrmi- i- one la\er in thickiie-- \\ith the form of

cell \ a i A iir^ 1 1 < -m tlat to Cuboidal acci ndiir^ to -late ( >\ e\teii-i< .n

li-j. 5 . There i- a \\ ell-<|e\ el. i| H d (iitiile. In regeneration

there i- tir-t an extreme extension ot ( pidennal cell- in the neiiji-

I" 'i hood of the injniA \\ 1 lit-li in a !e\\ houi - results in a covering

-I the i nt surface. Thi- e\ ten -ion i- carried SO far that the eel 1-
aie ijuile thin and aln io- 1 -epa rated Irom each other. Then tollou -
.t period of M-IA rapi«l proliferation in which the cell- are three
or four deep I ig. l These cell- are qui'e -mall, rounded.
irregularU arranged and I!HA ha\e very little c\to|>la-m. Thi-
coiidition .madualK changes until a portion lour days old -Imu-
onl\ a -iir^le !a\ e< . The cell- IA en then are \\ ithollt \\ell-marked
\\all-. their nucU-i are -till prominent and thev lack the ilelinite-
ness SO characteri-'ic of the mature epidermal cell. A cuticle i-
ea-il\' di-tin-ui-hed 1>\- the tilth ila\ and a -ixth da\ <-pidermi-
--entially mature though still -taining a little differently and

196 W. E. ALLEN.

lacking some indefinable quality of the old tissue (cf. Figs. 1-5).

Much the same things may be said about tin- intestinal
epithelium. In thi- ti--ue cilia are well developed on the third
(lav and it is e\idently functional l>y that time but it retain*;
some slight embryonic features to about the eighth or ninth day.

Septa are lir-t distinguishable in the embryonic mass as radial
columns of cells (Fig. i) extending partly across the cu-lomic
space. They rapidly fuse into a continuous membrane reaching
evident maturity by the fifth day (cf. Figs. 1-6).

The longitudinal muscle layer of the body is first noticeable
as a very thin, somewhat scattering layer of long, spindle-shaped
cells just inside the epidermis (Fig. i). It does not change very
definitely except for increase of thickness and compactness. Al-
i hough otherwise well developed by the seventh day at latest,
it is not of typical thickness until later (cf. Figs. 1-5).

The nerve cord is recognizable on the fourth day and almost
mature on the fifth. It is apparently typical by the seventh day.
Note in Fig. 5 the ventral localization of nerve cells in the cord so
characteristic in this group of worms (cf. Figs. 3-5).

The embryonic peritoneum seems recognizable by location on
the third day but is well developed by the fifth (Fig. 5), and in
evident maturity by the seventh (Fig. 4).

The blood vessels were not easily followed. The ventral vessel
is unquestionably well formed by the fourth day. Both were
observed in some specimens of the sixth and both are certainly
mature by the ninth (cf. Figs. 3-5).

The study of chlorogogue was very unsatisfactory. It would
seem that it should show well soon after maturity of the intestinal
wall at least and it is distinguishable on the sixth day and there-
after, but no specimen was found showing it in typical condition
even at the tenth day. It is possible that the sand in the intestine
affected its preservation more than that of the other tissues.

The seta1 are easily followed even in toto mounts. 1 he
embryonic seta sacs are well formed on the tilth day, and the
setai with all their muscles seem to be I'unctional by the seventh.
There is no question of their maturity bv the loih da\ < 1 ig. 4).

Nephridia were rather diltieiill lo identify. An irregular mass
of embryonic nephridial cell*- was first located, mainly by position,

TISSUE DIFFERENTIATION AND RATE OF GROWTH.

on the seventh day. A single loop, probably functional, is
plain on the eighth (Fig. 4), but the organ does not appear to
be mature even on the lenih.

All organs and ii— ne- except the nephridia appear to be func-
tional on the eighth day. They have al-o reached apparent
maturity by the tenth day. -onie before. The data an- not as
romp!' mid be u i-hed but they certainly are very indicative

ot a iorrelation between the maximum rate of growth and the
period ol di-tinct differentiation. Thi- i- e-p.-cially true \\lu-n

\\e con-ider the fact that the tenth regenerated -cement is the one
for uhi.h these data are rer.,rded and that its maximum -peed
rou th fall- on the ninth d.iy.

I KPERIMEN1 II. A\ IMMAHKI \\t\i.\i.. . 1 ;;/'•/ ystonni

T\M-M| i .

M> '.' -</\.

'iid in. ii I'L ton, Ind., probably in the ni<Jn of

Jaim.in -"• •idition at thi- laboratoix

on lebtii.:i\ i. I'hev \\ete kepi in -mall lot- under similar
condition i \ariatioii in uater. until it ua- found that the\

did ver> \\ell ir. i.iji er \\hich ii ua- u-eil regul.irly.

I p in the time «'l haii hin^, ihe main ellun- \»r control \\ere in
the line ••!" char. :i mv belo\\ j, i ( .,

an<l a\ i -id. mi « • •! d:n. • -nnli^ht.

On Febl uai \ -adpolc- . -lei led from a ijtiile

uniform loi and |>la.e.| -in^K in berrv di-he- \\ilh about 50 .
of i.ip wat( : ( Mi the these individual- were about t \\ o

da\- from th noiu- more than Imir nor le-- than one. The

di-he- \\eie jiailK iporation. \\alei \\a-

chan^ed and the di-he- \\a-hcd I \\ o or three time- per ue.k.

I f( -ll \\aler ua- frei;i|elltl\ added betUeell tillK'- if C'\ a] >of, 1 1 ioil

•:H'<| excessive. Ihe temperature record ua- taken twice

dail\ . \o eflforts to feed wen • :ul until l-Vbniar\ 20.

\\'ithin a few da\ - main \\ere feeding readily on live I.innimlrilns
which ua- u-ed i oininiioii-l\ to the end of the experiment.

l'.\ Man Ii ll several death- had occurred but the lai
number of tadjiole . -d to be \\ell .idju-ie I. seventy of tin-

be tier -pecimcn- averaging about J > mm. in length \\ere n-iaim d,

[98

\\ . I . AI.LKN.

TAHLK IV.

.1 »:lily\l(,»ni Tiidpoli-.

Iiulix i.lu.il Number. .Mm. Rcnic>\< Mm. !<'•_:' n- ruled.

I ininc-'li.iti'

70
69

68
67

66
65

64

63
62
61

45
46

16

2

59

29

14
32

17
47
44

28
58
3

33
48
13

43
57
18

27
4
19

49

12

26

56

5
35

6

1'
i i

?.75
6.

5-5
5-5

5-
6-5

7-

7-
7-

6.
7-5
6-5

6.

7-5
6.

7-
5-5
6-75

6.
7-
8-5

6-5
7-
6.

7-
5-5
7-5

7-25
5-2S
6-5

6.5
4-25
6.

6.

7-5
7-

6-75
6.
6.

6.25

7-
7-

.152

•152
.19

•3-S
•38
•35

•45
.76
.91

1.14
died
1.03

..7"
2-43'

1 .(JO

2.05
[.37

2.28
2.05
1.91

2.28
2.13

2. 2O

3-7.S

3-2S
2.25

3-2S
3-
3-75

I -t In nir.

''ih hour

1st dav

2d d.i\ .

3d dav

4th iLiv

Sth dav . ,

otli day1

7tJi dav . . .

8th dav

ijl h dav

I i i h r lav

TISSUE DIFFERENTIATION AND RATE OF GROWTH.

TABLE IV. — Continued.

. \ mhlysloma Tadpole.

Individual Number.

Mm. Removed.

Mm. Regenerated.

1 2th flav

;;

7.C

51

4-

36

7-

4-

i •;! Ii 'lav

21

_ -

i.

9-

4-

-M

7-

4-

I 4 1 1 1 • '. • . ...

;i

^ _,

37

7-

4.25

i s 1 1 1 da j

„

^.

iMli daj

6.

4-5

1 Kill.-'! :it.

llir othel- di-c.irded. ( Ipel'.ltioll U.I- ] U T)'« H'mei 1 nil M.irrl) [6.

T< ii specimens um k< check nnoprrated. Sixtv li.id ap-

pP'xilll.lteK i. II. --I). ill" the t.lil leill"\ I'd. Tile .lll.il open ill'.: .Mid

the lip "I i. til were i.ikm .1- ^nidf in. irk- .mil tin- cllori \\.i- nude
lc (in .'iii--l).ill \\.i\- !M-I\\'«-M "ii i-lini.ilc. Ii \\.i- .il-n iniciidi-d
ili.il tin- nil should l-i- .1 ii-hi .: to thf iiiitiii-lmrd. <»|"

course, there wen errors in !><>th v one <>t tin- iir-t lu-in-

p.irtl\ -ll'iun |iy tin- : lit- ill T.ililr 1\'. 'I'llc cut \\.i-

111. idr \\ith ,i liiit.inic.il razoi nsl .1 M»ik "t I'.n.ittiiic. In

urdi i i 1\ tin- i.i/'T it \\.i- In-Ill .it t-.ich end. tin ,nni- ln-in-

n--tcd mi .1 t.iMc .ind tin- tin i tin- p.MMt'tMic Muck. Sum-

of tin- rimr in judniiic-nt u.i- dm- t" -"inc t.iil- ln-iii;^ inure
.ittciiu.itc .it tip th.in iitln

<>ui , ,t' tin- '»i operated upon. tCur \\en- killed immediately,
three in .me h.mr .ind three in -i\ lic.ur-. The other- \\ere killed
in -ronp- o|' ; mi successive days t'n >in tir-t to -i \tei-nth inclu-i\ e.
( >iie of the three u.i- t.ikeil !'n>in one end o| the t.iMe the Others
l'r« 'in the other in order to ,i\ oid .my po--il>le intlueiK c o|" position.
( 'lil-oif- mixture Le( igOO u.i- n-ed for killing folloued l.y
70 ]>er cent. .ilc.. hoi. i.idi-ed S5 per cent, .ind the n-u.il after
proi edbie. I )elatield'- li.rina t o\\ Ii n I o] lowed h\ picric acid gave

200 W. E. ALLEX.

excellent re-tilt- in staining, the muscles showing especially well.
Two adverse conditions must be kept in mind throughout the
consideration of this experiment. (l) Lack of ability to control
temperature evidently affected the rate of growth seriously as
indicated by Chart II. (2) The old tissue is it>elf so embryonic
in character that it is extremely difficult to make an accurate
measurement of the amount regenerated or an adequate descrip-
tion of its changing features. Some of the vagaries of the
polygon must be attributed to both these causes. General con-
ditioii> \\ere good as is shown by the fact of only one death in
tin- regular series. Drawings are omitted because the requisite
series is too complicated for a paper of this character.

DATA .

These descriptions are somewhat indefinite because the facts
themselves with few exceptions lack clear definition.

P.v tlu- end of the first hour the epidermis had stretched over
and very nearly covered the cut surface. In six hours it showed
rapid proliferation, with cells closely and irregularly massed and
three or four deep. This condition remained much the same in
general appearance except for outward extension of the entire
mass at the end of the first day. In two days the older regen-
erated epidermis was indistinguishable from the neighboring
primary epidermis, though still proliferating at the tip. This
general condition remains unchanged to the tenth day which is
as far as the study of sections could be carried with any pretence
of general accuracy.

The parenchyma showed some sign of reorganization in one
da\ and there was a considerable apparent difference on the
second day, but thereafter there was no appreciable difference
from the ordinary conditions.

Bloodvessels were not in evidence in the regenerated tissue
until the second day when some were seen which \\ere e\ idently
functional almost to the distal extremity of the new tissue. Alter
the MO md day a fairly large vessel is uniformly found just
beyond the distal end of the ner\ e n>rd. and some capillaries
were identified (|iiite near the tip in almost all specimens. Many
of tllCM were easily ol>MT\able in the live specimen-.

TISSUE DIFFERENTIA!!' >N AND RATE OF GROWTH. 2OI

The fir-t -i-n of extension of the nerve o-nl <OU-JM- in a com-
paratively loose mass of nerve cell- or nuclei at the extreme end
on the fir-t day. These have -ome appearance of miration.
Little difference appear- on the -econd day although a distinct
lengthening of the cord has taken place. The third day shows
rlv a half millimeter of regenerated n<-r\e cord hut the end
-hou- no very detinite -i-n of rapid growth. After that the
condition c.mnot be distinguished from the ordinary.

The noioc hord -hou- some reneual in one day and a -li^hl

literalion on the 3( < "iid after which it appear- normal except

for a \er\ -mall area .it the end \\liich retain- the proliferating

charai ter. I hi- i- true till ahout the fifth day \\hen the di-tal

OIK- h. ill to t hit •c-ii. urth- of the notochi.rd is markedly different

in .!]•! " I'Mii the re-t. The I'ell- are -mailer and n.

-lion^K -tained. c\cn tl- -pla-m taking tin- ha-!nat"\\ lin

-lain -li^hiK ' I -tlldy of the >lide indicate- that thi-

di-ial p.n i oi i!i hord i-inatran-ition.il- dil'leivn-

li.tiion and that it ha- ap; l'\ ,i -imultane. m- change in the

neu cells dc-tined t<- l-iriu the di-tal not.ichord. It i- inien -li
to note that uh> ys tlu- noiochord had

hehind tin i : it quickly became «.-tc|-minal \\ilh and

:idcd -liv;htl\ l'e\ond the ner\ « i . .rd li\ the ti^hth or
ninth da\ . lli: iiotiichord de\ elopmnit i- \

-lion-! int ident \\itli the period • test

•dini atiou.

The In-l -ivii o| nui-cle de\ elopnient i- found on the -ixth
da\ ulien in i ell ma--e- ai- ni/alile a- aiila^eii i .| the

neu mu-cle ti--ue I di-lini;iii-hal>li- thn>u;Ji al-oin

one-half the extent .i| lieu tissue. < >ll the -eXelllll d.l\ a leU

pi-'ximal >trand- ol uiu-cle -ho\\ fair de\ elopmi-nt . And on ihe
eighth da\ -mne are found throughout tin- |>ro\imal third. In
nine da\- a leu \\ell de\el..ped lil.er- are di-t in^ui-haMe half
u.i\ out in tlu-jieu ti — lie an<l on tlu- tenth day it i- extremely
ditlicult to ideiitil\ the 1 mrder line lielueen the |iroximal

neraled mu-cle and the old tisSUi

While mo-t of tin- tissues -tudied reach tl inal State ot

de\elopment too -0011 to -ho\\ an\ connection \\ith or inllueiu.-
upon tin- lii^he-t -peeil of iv-eiiera t ion iCharl II.', it should he

202

W. E. ALLEN.

remembered .that all except the muscle are so generalized in thru
original condition that they require little change before reaching
the original limit of differentiation. On the other hand, the
muscle is rather highly specialized and if it be taken as a standard
.1 rather marked correlation is noted. It is just becoming dis-
tinguishable at the sixth day when growth has its greatest speed
according to this polygon, and it gradually takes on maturity
until it is well developed by the ninth day which is evidently

Chart 11.

Temperature.

15°

Cfrowth Incite.

Sections

Toto

B

10

12 13

IS 16

(.'hurt II. — Tempi-ratlin- i° C. t<> i JM inrh rise. Growth rate - M.I nun. to
!0 inch rise.

somewhat past the time- of most rapid extension. A -lance at
the chart indicates the probability of greatest speed having
been -ho\\ n at about the seventh or eighth day if the temperature
had been under control.

Chart II. is a polygon constructed from micrometer measure-
ments under the compound microscope, of the tissues in toto

TISSUE DIFFERENTIATION AND RATE OF GROWTH.

203

-

9 I

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-

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— —

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r - -i

f E

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7.

f

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1

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E

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-

:

-

u

—

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_:

— '.

— i

as

0

c

C

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=

M

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'E _;

S —

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X

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M

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Ci

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i.

-. _!

- -

^

Z

Z.

—

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"3

. E
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=

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- - -

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P

B

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-

X.

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d

C

w

~n

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73

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—

=

E

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T

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^T

E
T

7

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X

Z

w

-

...

Z\

1

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^«
-

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u

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—

s

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-. 1

^ I

1 2

~ r

-

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X

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H

204 W. E. ALLEN.

and after sectioning. The solid line indicates the former and
the broken line the latter. Temperature fluctuations are re-
corded in the upper part. It was very difficult both with the
toto examination and the section examination to distinguish the
exact plane of cut, and of course, a very slight error in identifi-
cation of this level would make a marked difference in the poly-
gon. This difficulty rapidly became worse as the regenerated
tissue grew older and measurements taken by any method after
the tenth day have a purely suggestive value since they have no
basis in strict accuracy. A set of measurements was taken
with dividers, using a binocular dissecting microscope, but as
their results are not essentially different they are not plotted
on the chart. Table V. is self explanatory.

SUMMARY.

It seems that the results of these two studies are sufficiently
clear to warrant assumptions as follows though further data
are very desirable.

1. When adult tissue is removed the resultant regenerative
growth reaches its greatest speed just preceding, or at least not
later than, the time when the major somatic tissues become
typically differentiated.

2. When immature tissue is removed the resultant regenerative
growth rate shows no relation of any sort to the differentiation
of the more generalized somatic tissues. There is, however, a
marked correlation with such a highly specialized tissue as muscle,
the differentiation of which is coincident with the period of
maximum growth rate. And the peculiar development of the
notochord also strongly supports this view.

Of the literature available, only that which was found dis-
tinctly useful is listed here. Krecker's list is good and a more
general bibliography, especially concerning annelids, can be ob-
tained from his paper.

PAPERS CITED.
Durbin, M. L.

'09 An Analysis of the Rate of Regeneration throughout the Regenerative

Process. Journ. Exp. Zo51., 3, 397—420.
Krecker, F. H.

'10 Some Phenomena of Regeneration in Limnodrilus and Related Forms.
Zeitschr. wiss. Zool.. 95, 383-450.

TISSUE DIFFERENTIATION AND RATE OF GRO\YIH. 2O5

Lee, A. B.

'oo The Microtomist's \'ade Mecum. London.
Minot, C. S.

'08 Age, Growth and Death. NY\v York.
Morgan, T. H.

'oi Ki-LM-n. -ration. N"f\v York.
Morgulis, S.

'07 Observations and Experiments in Lumbriculus. Journ. Exp. ZoOl., 4.

'n !<• 'In Water Content in Regeneration. Journ. Exp. Zool., 10.

321-3 :
Miiller, C.

'08 !<• K IK- an I.umbricuhi aus und TuhilYx rivulorum.

An li. i '. i • j6, 2OO-J77.

ABBREVIATIONS

R m =11111

niatui- :<•. rd.

RS ^ m. nr =iu-phridium.

b - i ~*cl. f>e - inn.

C -clil": •'ili-ratniK in.;

e " rpidi-rn 5 »!ii-ptiilii ><r M-ptal anl.

ss -s«'ta sac.

206 \V. E. ALLEN.

EXPLANATION OF PLATE I.

All figures are from camera drawings and partly diagrammatic. All have the
same magnification (375 diam.)- None are intended to represent cell detail.
All show only most conspicuous features of the ventral side in or near the tenth
regenerated segment.

FIG. i. Third day. Septa almost formed. Muscles just distinguishable.
Proliferation of epidermis and embryonic cells just inside muscle layer.

FIG. 2. Fourth day. Septa formed. Epidermal proliferation less. Cells
massing to form nerve cord inside the muscle layer.

FIG. 3. Fifth day. Nerve cord, bloodvessel and peritoneum formed. No
proliferation.

FIG. 4. Ninth Day. Typical ventral localization of cells in ner%-e cord.

FIG. 5. Old, mature segment. Note larger size, the thin epidermis and thick
muscle layer.

FIG. 6. Eighth day. Optical section. Immature nephridium. Mature seta
sac and muscles.

8IOLO&CAL BUUETIN. VOL. XXP

PLATE I

1»»»»*\

u

Fig. 5

THE SMALLEST POLYODOX.

THOMAS BARB01R

An article which appeared in a recent BUM ( M ,i< .\i. Hi i i I n\
h>- hanforth. entitled a "74 mm. I'olvodon"1- reminded me th.it
sonic yeai I had seen -ome -mall -pecimens of Polyodon

in the -tudv collection of Fi-he- in the Mu-eum of ( "omparat i\ e
Zoology. \ n •• xamination < >f the rn.in.-ri.il pre-erved th'-re re-
V'-aled tli' 'tuple-, tin- smallest hitherto recorded, as well

afl other- which an- anion- the -nialle-t known. Tin- three im-
portant example- are .V> mm.. .^7 mm. and 05 mm. lon^. The
la-i iii-lix idual \\a- -ome\\ h.n drii-d and ua- ol.viou-ly -imilar
in form an<l development to the spreimeii de-cribed and figured
1>\ I >aiiforili. The -nialle-t ha- been l>adl\ -o| teiied am 1 -\\ olh-n
li\- immei-ion in \\eak aleuhol. Thi> individual \\a- collected
in Ark.ui-a- l-\ Mi < .. Stollev and canu- to tin- museum hi-lore
0. 1 he other t\\o -mall -|ieiini.-n- a- \\ell a- three more.
which are 80 mm. ,85 mm. and 03 mm.. < a me from near St. 1 .oui-,

MO., and \\eie Collected in September. 1 x- J. 1-v a I >i . l;.a -till. ill.

The\ \\ere iii the private collection of Professor l.oni- Agassiz
which vva- aci|iiired by llar\'ard 1'niver-itv a- the foundation
lor the Mii-eimi < :npaiati\'e Zool". '. lhe-e live example-
have been kept in -troiu clear alcohol and are well preserved.
Tin- example of \~ mm. then is the important -pecimen in that
it i- the Miialle-t known which is reasonably well pre-erved.

1'ioic— or ('. II. 1 igenmann, who ha- seen thi- -pecimen. -n^-

•ed that it wa- probably about three month- old. in which
Case the c^ lav in- would lake place in July. If thi- wen- not
really tin it i- 'jiiite po--ible that I )anforth'- individual

whiih was about ;, in. he- Ion- was not the -wift-^rown vdimu of
the year but a -peeimen which had lived over one winter. < '.row th
mav be verv -low or alnio-t arre-ted during the cold month- in
the latitude of St. l.oiii-. It. a- i- quite po--ible, the .^7 mm.
\OUHL; wa- reallv much \-oim_uer than three month-, then this

1 HIOI.. Hi i i ... XX

207

208 THOMAS BARBOUR.

rate of growth would seem reasonable enough. Emphasis is laid
on this point luvunse great search has been made for the young
during the months of spring and early summer. This search
has been fruitless, while these inch-long fishes were got in Sep-
tember.

The drawings which have been made by Mr. E. X. Fischer
with special regard to the proportions, show three aspects, dorsal,
lateral, and ventral of the 37 mm. example, as well as of one 80
mm. long; two aspects of a fish 130 mm. long and a lateral view
of a three-foot adult. These figures serve to show the remarkable
features of the young as will as the changes in form which the
species undergoes during growth, better than would a lengthy
verbal description.

As may be readily seen from the drawings the most interesting
characters to be observed in the young are the short sturgeon-
like snout which has not yet become in the least spatulate; the
extremely heterocercal form of the tail; the relatively long bar-
bels and the less developed fins throughout. Unfortunately the
notch near the outer extremity of the caudal fin is not shown n
the very young because of the frayed margin of the tail fin.
This character is marked in the slightly larger examples and,
as is shown in the lateral view of the 80 mm. specimens, the notch
marks the point where the cartilag nous fin rays cease to be
developed. This character would seem to suggest that seen in
the caudal fins of certain sharks wrhere there is a similar notch.
The whole general form of this, the smallest known selacho-
stomous ganoid is certainly most strikingly shark-like as well as
reminiscent of the adult forms of other ganoid genera.

210 THOMAS I3ARBOUR.

EXPLANATION OF PLATE I.

37 mm. Polyodon, X 3. Fig lA, lateral view; Fig. iB, ventral view; Fig
iC, dorsal view.

BIOLOGICAL BULLETIN, VOL. XXI.

PLATE I

I.I

E. N. FISCHER DEL.

212 THOMAS BARBOUR.

EXPLANATION OF PLATE II.

80 mm. Polyodon, X 2. Fig. 2.4, lateral view; Fig. 2B, ventral view; Fig.
2C, dorsal view.

/

\

'

.

214 THOMAS B ARBOUR.

EXPLANATION OF PLATE III.

Figs. 3/1 and 3.B, 130 mm. Polyodon, lateral and ventral vie\\s. XiJs- Fig.
4, 3 ft. individual from Boston market, 1/6 natural size.

,

•

.-.

-

z

NOTES ON TWO CROSSES BETWEEN DIFFERENT

RACES OF P!C,K« INS.

T. H. M< )R«-AX.

The- pn-M-nt note i- inu-mk-d to rro.nl tin- iv-ult- of a U-w

crosses with pi-con-. Through a -.i.-rir- of acridem^ tin- rxprri-

inriii cam.- to an untiim-ly t-nd. Tlu- hn-i-din^ <• irriol t" tin-

at-ratidii ha- ^i\cn \»<> t\-\\ indixiduaU lo warrant any

'•rali/;iti«»iix. luit ciiou-h I ' llu- di-iral >ilit\' <>f IIMH-

fxtfinlcd \\ork aldii^ tin- -anu- linr. A- it \\ill takr t\\n years

ID iair\ llir en thi '"iid -riHTanoii it sri-ins

\\«irth \\liilr t<> n-rord the fa. ir ol.iaim-d.

CROSS I' l I \\ I I \ \ \Ylll 1 l I \\ I \ll \\D "SWALL< >\\ ."

\ • -. It- fantail I i. I \\ as pain-d to a male

I-K.. I.

-.\\allo\\ Fig. -1 One of tin- chi« f featun ;he breed of

fantaiU i- the large nuinlii-r of t.iil feather-; in the bird u-ed
hete ,,J tCatlu'r- \\ere pre-i'iit. I \\i-hed to find out how this

21 S

2l6

T. H. MORC.AN.

character would beha\e \\lien brought into connection with the
normal numUr u ot" tail feather*- possessed by tin- "-wallow."

2.

The First Generation —Seven young were reared from this cross.
The tails of these hybrids had respectively 17, 12, 13, 15, 14, 13,
13 feathers. The result ^hows an intermediate condition be-

FIG. 3.

tween the parent types, with a distinct approach to the normal.
The tail of the hybrid was not carried in the erect position as-

TWO CROSSES BETWEEN DIFFERENT RACES OF PIGEONS. 217

sumed by the fantail nor was it flat as in the other type, hut
formed a broad rather flat wedge like that of the fantail at rest,
but not as high. It is shown in 1 i_ ;.

The FI Generation. — Only four birds were reared in thi- genera-
don Fach had only twelve feather- in it- tail. It may appear
cither thai there is a further loss of tail feather- in thi- generation
or that the normal number, t\\elye, repre-ents the dominant
type, and that tin- number of indi\ idu.il> i- too -mall for the re

• type i" reappear. Farmer number- mu-t be obtained to
tie thi- point, for. ii -eem- not improbable that F-j birds with
mon- I. -at In T- than twelve.uv expected, even although the original
number .^2 may -eldom or m-\rr reappear.

Olln-r Churn. . [t is not the object of thi- note to examine
in dd ail the inherit a IK e ol Other « ha racier- of the-e two breed-,
but the l'oll"\\ in- point- may be mentioned. The tantail ha- a
plain In. id Fig. i. . /. c.. n •; tin- >wallow ha- a nv-t . The

I bird- all ; rest. S 'lie of the ]•'.. bird- have a crest,

other- lac k it . ( »ne bird I ha I \va- killed i- IMI recorded for ere-t.

The -wallow ha- leather- on the tar-u- and tOCS Fk- J;, the

fantail lacks thes I . i fhe FI birds have feathers on tarsus

and IOCS, bill le-- developed than are tllo-e of the father. Two
ol the I . bird- lack leather- <p|i the tai-n- and loe-, one bi d
re-einble- the FI livbiid. and mie ha- well leathered lai-n- and
toes. In the la-t case the feather- are a- lon^, and a- \\ell
developed. .1- ill llle -\\allo\\ .

The tantail ua- pun- uhile, the -wallow ha- a dark top to
the head, dark v\ iiu-. and the feat her- of the feet are liijll bluish.
The \\ing- an grayish blue and broun v\ith two brown bai'-
The tail. baek. nei k and veiitra' -urfaee are white.
The l-'i bird- ate pure white. Two of the-e bird- >howed two
or three slat Bathers near the outer ed-e of the

winj leiallv in be -een oiilv" b>' bru>hing back the while

leathei- or when the wii j Mended. Two ol the F-_- binl-

aie |nnc white, twii \\ei-e -plutehed, one with red, and one with
brown. ( >nlv a -mall number of the-e colored leather- were
pie-ent. but thev weie -cattered unequally over the body and
showed no icndencv to be grouped on the winu- and he, id alone.

'wiiii.1 t ion jury !>>• tat - tin- In-. i< I \v.i- t"" iiuu Ii catrii in 14 i\ -.- , t-i tain rt-cor^ls tdr
thi- »K -i.

2l8 T. H. MORGAN.

Summary .--The FZ generation was inte mediate n number of
tail feathers, feathers on legs, and color, although in regard to
the last point the bi ds were practically 1 ke the white parent,
a few feathers alone placing hem 'n the intermediate class.
The crest \vu> dominant. The F2 generation showed segregation
practically complete for the feathers on the legs, it showed segre-
gation of white color to some extent, the recessive type appearing
only to a very limited degree. The number of tail feathers
showed a return to the normal, but the number of individuals is
too small to lay much emphasis on the point. Yet it is significant
that in all other directions the F2 birds sufficed to show the in-
fluence of both parents.

CROSS BETWKKN A TURBIT AND A STARLING.

The FI hybrids of those birds were given to me by Mr. E. B.
Southwick, who kept both parent stocks. The turbits (Fig. 4)
were not very high-bred strains, but showed the main character-
istics of their class. Their color is red or blue. The head is
uniformly colored and has a cres : the- wing bars are dark; along
the middle line of the breast there is a series of reversed feathers
(Fig. 4). The starling (Fig. 5) was a black bird with a white top
to the head and white wing bars. On the breast there is a large
light crescent with a, metallic tinge. There are no reversed
feathers on the breast.

The two hybrids (Fi) were red birds (Fig. 6). The crest was
well developed. The top of the head was white (like that of the
starling), but in one case not so sharply delimited. The wing
bars were dark.

From these two hybrids eight, Fj birds were reared. Five
have crest and two lack the crest, five are red and three blue,
all but one have darker bars on wing. T is one has a dark bar
on most of the feather^ of the wings but a fe\v feathers ha\ e \\ hii<
bars.

Summitry. — The most interesting result of this combination is
the failure of the reversed feathers on the breast to reappear in
the second generation. The result i> like that of the number of
tail feathers in the fantail-s\vallo\\ cross, but here also the
numbers are too small to make any conclusions possible, however

TWO CROSSES BETWEEN DIFFERENT RACES OF PIGEONS. 2IQ

significant the absence of rever-ed feathers may appear. The
iJiite top of the head and the creast appear to be dominant;
while the ichitc winy bars are recessive.

5I( >NS.

A- -tated ,il.o\r the immlitTol' individual- reared i- too -mall
to \\.in-ant Ctensivi /ation-. hill il i- apparent

that uhile . erlain clia: evidence of mono-hybrid in-

h. Titan. .-. three characters d«. not fnrni-h as ^.'od evidence.

'llu-c three charactei 'he nninl'ei ..t tail hMther> in the

i.iil. the (ol.-i | .ittein .'I tl . -\\allo\\. and the re\er-ed lnvaM-

feathera < •!' the tnrl.it. It i- prol)vil.li- that cat h of tlu-e char-

acters has nol been formi ' i • a -in-l<- -tep. l.nt l.\ .1 s< rie- of

-tep-.1 It tin- ! HOWS thai tin' condition- pre-ent

ill the tu< ii\el\ ran not I >e fe| n'e-nited l.\ a pair of

M. ndeli.m chara. ters. 1 here are then t \\ <• iiit.TpnMat ioii-: first,
it the characters Mendi-li/e there mn-t In- more than t\\o factors
in\ol\ed; -e. ond the ih. - in (jiM-tioii ma\' come under

-.-nil- other kind of inherit. inn- in \\hirh Miupli- Mi-ndeliai:

ion do,-- not occur, or p, — il.lv the re-nlt- may lie partlx Men-

delian, parlK not. A large nnml.erof !•", otl-prin- will I .e neces-

-ar\ In tore the-e point- ran l>e -ettled, l.nt enough IKI- been

loimd, I think, to -h»\\ at lea-t in cases like the-e when a t \ pe

t ^, in, T. H ' atal ZoOlogy," page 171. New York. 1907-

220

T. H. MORGAN.

has been formed either under domestication, or in Nature, not

by one mutation. il >iep alone, hut bv a M-ries of succe^ive steps

FIG.

along the same line, that the crossing with the original type or
with a related type does not give Mendelian inheritance for one

FIG. <>.

pair of characters. What we should expert under such circuin-
stances \\ e do not know a- vet for \\ e are now dialing with a

TWO CROSSES BETWEEN DIFFERENT RACES OF PIGEONS. 221

series of progressive or orthogenetic variations and not with a
n-tro^n— ive mutation.

There is another result that seems to me to be especially
significant. The white on ihe top of the head of the starling is
dominant to the plain (colored) head of tin- turbit . Inn the white
win^ bars of the starling are recessive to the dark wing bars of
tin- turliit. I'nless these are two different kinds of white, one
dominant ' inhibitor of color) and the other recessive ( absence of
color: then-Milt is difficult to explain, unles> as I have suggested *
tin gation in mosaic types i- not germinal but ontogenetic.

1 " 'I In- Intlm-: ;ity aiv! : :r"ninciu in I Vlri mining the- ('".it

' ' . X X I.. Jul> . \<ii i .

THE (ENOCYTES OF PLATYPHYLAX DESIGXATUS

WALKER.

WILSON P. GEE.

INTRODUCTION.

The oenocytes as unique cells of the body of all of the orders
of insects which have been investigated, except the Aptera, have
received much attention within the last couple of decades, but
with relatively little more than mere conjecture as to their defi-
nite function. \Yith the idea of throwing added light on the
nature of the activities of these cells, the investigations here dis-
cussed were made on the larvae, pupae, and adults of Platyphy-
Id.v desif.'jnttiis, a caddis-fly, which occurs quite abundantly in
fresh-water springs in the vicinity of Madison, Wis.

The results here presen;ed were derived from a study of the
structure of the cenocytes in several periods of the life-history
of this insect, and by injection experiments. Several stains
\\ere tried, Heidenhain's iron-haematoxylin proving the best for
detailed work. Delafield's haematoxylin wrh orange G or eosin
\\ere the stains used in determining the location and number of
the cenocytes. The cytoplasm of the oenocytes takes up eosin
very readily, as it does also orange G. The substances used in
injections were methylene blue as an intra-vitam stain and
sulphindigotate of sodium to test the excretory nature of the
cells.

The papers of Perez (1903), Rossig (1904), and \\Vi-senberg
(1907), presenting a rather complete review of the work done on
cenocytes, it will here be necessary only to briefly mention some
of the chief contributors to the subject.

According to Ro^ig (1904), the lux to observe tin- u-nocytes
special organs was M. Kabrc (1856).

Landois (1865) distinguished them from the fat-bodv, and
gave to them the name " Respirationszellen " 1> icause of the rela-
tion he obser\ed them to bear to the tracheae.

(ENOCVTES OF PLATYPHYI.AX DESIGNATUS. 223

llaberlandt 1^72) observed them in the -silk-worm, where
they were described by him as an organ of unknown function.

Graber (1873) separated the oenocytes sharply from the
pericardial cells, and called them "ein.ne-pren^te Zellen." He
-aid regarding their function: "Ohm- Xweifel halien wir es hier
mil ei-niarti.uen rin/rlli-ru 1 >rii-en y.u thun, iil>er dereii -eeivt
ullerdini;- nicht da> < ".erinu-tr liekannt i-t."

The name " < leiiorythen " ua- ;^i\ni to the-e cell- by \\ielo-
\\iei-ki [886 "ii account of their n -rmblamv in rolor, in the
larva of Chinni< white \\ine. Tlu-y \\ere ob-erved by

him lo be mrtamrrieally n-peated under the hypodermic of the
alMlnmiii.il segments and arran-o-d in -mall ur"iip> -iirrotindin-
or ne.ir t" tl, M.ii.i. Thi- ua- tin- nio-t r\ieti-ive \\ork on

the OBI • hi- time- for \\'ielo\\ iei-ki wa- -in i'e--ful

in ideiitif\ inv; tlii-m in the 1 )iptera, ( 'oleojurra. 1 1\ iiH-no|it.-i .1

and Lepidoptera. He in. hide- tin- . \\\.- elemt-nt-

of the M'.od ti — u«- .lll'i . "... d.i-- -ic alle

\ oil drill -if lllllv' Ix-ndrn Mrdilllll ^»-\\i--r ^lol'lr all f Hell Illell .

yeit\\ri-r .mt-pri hrrn re-|>. \rrarlieiten und ir^«'iid \\rlehe I m-
>at/prodnkte .in da--rll»e zui i'«-n und dadtuvh .ml die in

den I l.uipti;r\\< : ai-mii- \nr-ieh ^rlirndrii .\--imila-

tioii-- und 1 )r-.i — imilatioii-.p- einrn Kiullu— an-iil irii."

K"\\.ile\ -k\ [887 .it tributril to thr u-lloe\ ; ^l.illdlllar

function.

\Vr-i -n ls'il J in hi- \\ork on the -ilk-worm gives them the
name "< . llulr ^l.mdnl.iri hypostigmatische," and find- their loe.i-
tion e\( ln-i\r|\ under anc| behind the -ti^m.ita of the abdominal
-eminent-. A- e\ ideiicrd b\ thr number of \a.-u..|r- iii iln-ir
i \ lopla-ni during the lar\al - lie attributr- to them a -lan-

dtilar lunetion. At the time of the -pinnini; of tin in, t \v o

or three d.i\- before pupation, a -ee. md irnitcyir -nirr.it ion ap-
pear- in bro.id |.: :i tin- \nitral part of the third, fourth, and
the lifth abdominal -e-meiit-. Thr-r attain a -M/e of (.»^ in
the -i\-da\ |.npa. and br-in at thi- t'me to di\idr bv amito-j-.
The lar\al .enoc\ tr- prr-i-t to the end of the life of the individual,
- mueh as i.V'M m "'/l'-

Grabei [891 places the cenocytes with the blood ti--ue and
i;i\e- their origin a- .•etodermal. Hi-. howr\.-r. eoii^iders them

224 WILSON P. GEE.

as being metamorphosed into the fat-body, and also as giving
rise to the Hood corpuscles.

\Yheeler (1892) in an extensive paper on the blood tissue of
insects deals with the oenocytes at considerable length. He ar-
rives at the following conclusions:

1. "The oenocytes are derixcd by delaniination from the ecto-
derm just caudad to the tracheal involutions They are also
metameric organs.

2. 'They are limited to the eight trachigerous abdominal
segments.

3. 'They appear to be restricted to the Pterygota, in all the
members of which group they probably occur.

4. 'They give rise neither to the fat-body nor to the blood
but represent organs sui generis.

5. "Alter their differentiation from the primitive ectoderm,
they never divide, but gradually increase in size."

\\ itli regard to their function, the only suggestion that Wheeler
makes is in his discussion of their vacuolate structure in certain
of the larva? of the Trichoptera. Here he says: "One is re-
minded of certain gland cells which store up vacuoles of a specific
substance in their cytoplasm, preparatory to secretion."

Heymons (1895) in the embryogenesis of Forjicuhi, points out
that the oenocytes are found in the eleventh abdominal segment
as well as in the others. The eleventh segment bears no stigma,
and thus they cannot bear any relation to the stigma as such.

Pantel (1898) classes the oenocytes as an organ because of
their metameric repetition, and treats to a slight extent of their
cytological changes in different periods of the insect's life. He
hesitate^, lumever, to commit himself as to their function, but
rather inclines toward the excretory phases of their probable
act i\ il ies.

Berlesc (1*99), in his work on tin- ants, 7 \i /uiionui erratic nnt
;md I'hcidole pallid/ilu, finds that, in the beginning of mei.i-
morphosis, the larval u-nocytes are multiplied, give up their
lixed position, and, becoming amu-boid, are scaiterrd about
among the fat-bodies. In his work on Politics 'jalllca and on the
honey-bee, he ImiU a persistence of the o-nocytcs, \\itli no special
second generation. Their function he considers as excretory,

CENOCYTES OF PLATYPHYLAX DESIGNATl -

serving during the periods of moulting and pupation, when the
Malpighian tubes art- functionless.

An-la- 1900) describes the oenocytes as more or lc>- anneboid,
and with n<> -pecial distribution in the Hymenoptera, except
their existence in the abdomen alone. Except for a relative
increase in volume, they remain throughout the life-cycle, in
both di-tribution and structure, very similar to what they are
in the e.irK larxal stages. He sucue-t- that they perhap- -ecrete
about them-el\e- ferment-. an<l con-ider- them a- glandular cells

.mating from the hypodermi- and functioning as organs of
internal SC( ret ion pcrhap- for di— • >ciati\ c purpo-r-. Tin- pnul-
in i- <i| theii ; ion may serve for the general nutrition of

tin- ti--iie- of tin- bodv or even tor the cytol\-i- of th<>-c larval
cell- de-tined to di-apprar.

KO-I hc\ niko\ [900 . in hi- \\ ork mi the tat -bod v and < i-m >c\ tes
ol the hotn-\ -be.-, hnd- that nio-t ot the lar^e CEDOCytes "| the
lar\a • li-int • - rate in the pupal -omr of them continuin-.

ho\\e\i i. into the bei;inniii^ of the ima^inal stage. In a \oim-
])ti|i.i, a second 0 .'-neration, to \\hich he gives the name

" Imaginaloenocyten,1 arises from the h\ podermi-, and tip

OenO( \ l« - an lab i di-tribllted aiilon- the fat-bodie-. 'l'he-e are
onl\ one tilth a- 1 .:•. the lar\al cenOCytCS and are toiined

01 1 K in the pupal st . Ib observed inelu-ion- iii -onie of tin-

cell-, and on t hi- ba-i- came to t he « oiiclu-ioii that I hex are to be
loii-idend .1- excn in-, uithout " Au-fiilinm^-^an.

\\hi(h serve as Storage place- tor > product- ol excretion.

Vane) 1902 says < • : \3iSimulia, \e Culex, \eChironomus,

durant la | eriode iivmpliale ie in \oj- aiicun chani;i-ment dan-
celluli llou,\er, he ob-et \ e- iii .^iniiilinni that the num-

ber of Oenocytes appears tO have increa-ed at the end of pupation.
He conclude-: "ChCZ le- /'.'/'/•'• Ie- OenOCytCS He -llbi— ellt
aiicune hi-lo|\-e durant la ii\m]tlio

1'en / [903 states relative to tlu- di-i overy of l\o-ehe\ niko\ ,

(oiiiernin^ the origin of a -econd oenocyte generation, that this
opinit ni -eem- to In erri >iu-i >u-.

!<<"• — i- 10041 t!iid> that, in the ('\nipid.e. the CEttOCytes are
limited to tin- abdomen. The lar\al oenocytes n-.ich a length
as great as l.^oM and attain a proportionate length one tilth that

226 WILSON P. GEE.

of the body. These larval oenocytes degenerate in the pupal
stage. The imaginal oenocytes originate at about the same time
through mitotic division of the hypodermal cells on the ventral
side of the abdominal segments. They are described as very
much smaller than the larval oenocytes, never equalling them
in size.

\Veissenberg (1907) finds in a well grown larva of Torymus
ni'^ricornis, over one hundred oenocytes, presenting no definite
arrangement, and scattered about among the fat-bodies. They
are in no way limited to the abdomen, nor do they necessarily
have the definite tracheal attachments observed by Wielowiejski
and others. The principal degeneration of the larval cenocytes
occurs in the stage which he calls the "gelben Puppe." The
larvae shortly before pupation contain the imaginal cenocytes in
the form of special cenocyte imaginal plates, which are laid in
the niches of the dorsal hypodermis in the fifth to eleventh
abdominal segments.

With regard to the function of these cells he says: "Schon
oben habe i h mich gegen die Auffassung der Zelleinschliisse als
Exc ete ausgesprochen, mochte aber hier nicht das fur Urate
characterische Bild der braunen, radiar gestreiften, runden Krys-
tallkorper darbieten, sondern an den frischen Zellen homogen und
farblos erscheinen. '

ORIGINAL OBSERVATIONS.

In the larva.' of Platyphylax designatus, the cenocytes are limited
to the abdomen. A careful search through serial sections of
the thorax of several individuals in different larval stages showrd
no traces of cenocytes in this portion of the body. They are in
no way limited, however, to certain abdominal segments bin
occur in all, the larger number being present in segments two to
seven. The definite tracheal attachments observed by many of
tli • \\orkers on this subject have not been seen in this insect.
This ob-.erv.ii ion agrees essentially with that of Weissenberg
(i<)<>7) in Torynms ni^rirnrnis: "Von einer Befestigung an Tra-
cheencapillaren, habe ich nicht> bemerkt."

The eenocytes occur both dorsally and ventrally, by far the
gre.tter number occupying the hitler position. The- larger <IMIO-

CENOCYTES OF PLATVI'HYLAX DESIGNATUS. 22~

cytes, so conspicuous in the early larval stages, occur veiurally
in pair-. one on each side of every abdominal segment. The
smaller oenocytes of the earlier, as well a- the later larval stages,
are found principally in gn>up> of from two to five, the larger
ip- appearing uion- ventrad. In -e\ rral specimens they were
observed near to or even grouped around a t radical tube to which
they, however had no connection. The location of all of the
• i-ii" is distinctly pleural.

The distribution of the o-noi \tt-s does not char. ' ntiallv

\\ith the iin rease in -i/e of the larva-, but there i- a le-s marked
tendeiK \ for them to occur in Croups. \\'ith the increase in
si/e ot the lat-bod\ more of them become partially or compleielv
-m rounded by this structure. The-e ch more marked

in the late larval and in the pupal si but \\e frequently MIN!

in the :- m.inv as ti\e oenOCytCS, and this

lendem \ to grouping i- observed even in the ima^inal sta

\ i aivlul < ounl ot tl;, • a lar\ a eijit millimeters in

length totals about one hundred and t\\eitt\-ti\ •. the-e bein-
ap|iro\imat«-l\ ei|ii.ill\ 'itioncil lie! \\een tin H'Jit and left

sitle- of the bod-. I • 'her estimates it i- '..mid that this

number i- about iv; I h. n seems

to be no \\ide tin- tuation in numbers in dillerent |>etiods nf the
li|e-|iisio|-\ , though the ima^inal stages appeal to ^}\<>\\ a d'
in niimbi • ;he lar\al, due perha|>- to disinuvrat ion. The

loiulusioii that the numbei is con-taut lor all the larva- of the
-an • i I belie\e. hardlx ju-l iliable, but there is no \\ide

\aiiatioii in tin I. In no case. ha\e I s,-,-n evidences of

di\ is'i .11 eitlu-r b\ mito-i- . .r a mi to- is in an\ ol the stages si udied.
nor an in in numb»-r in later larval, pupal or imaijnal

• .\ ei that in the earlier lar\ al perioiU.

\\hile the u-iio. .ppear practically constant in number

lhroui;hoiit the >tai;es examined, there is to be noted a \ ery
matked increase in tlu-ir - th- lar\a advam< .routh.

The lollouin^ tabli- tcprcsent- the >i/,- ,,f an average oencx

derived from measiireineiii - of several ol these celU in the

indicated.

The lame-i sj/i- «,f tin- oenocytes is reached in the pupal stage;
the greatest lenuih of the nucleus, proportionate to ihat of the

228 -\YILSOX P. GEE.

Length of Larva. ' Length of Cell. M .ength of Nucleus.

4.5 mm. i8.4M 10.4/1

6 mm. 24 fj. 12 n

12 mm. 40 // 20. 6/1

22 mm. 77 n 43-5M

pupa 103. IM 63.5/1

c;uly imago 94-3M 62.9/11

cell is found in the imaginal stage. There is to be noted, how-
ever, a decrease in the total length of the oenocytes of this latter
stage, this being due, beyond doubt, to degeneration, manifest
signs of which are present.

There are two sizes of oenocytes in the earlier larval stages.
One of the larger type, from a larva six millimeters in length, is
shown in the accompanying plate (Fig. 3). As already noted,
these large oenocytes are present in each of the abdominal seg-
ments, ventral in position, one occurring on each side. These are
many times larger than the oenocytes of the smaller type, and are
pressed rather closely against the hypodermis. In the earlier
stages of larval life, the greater comparative size makes these
oenocytes very conspicuous, while in the older larva this is not
so marked; in fact the smaller cenocytes in the oldest larvae
attain a size equal to or even greater than the formerly larger
ones. Careful examination of serial sections of these larger ceno-
cytes showed no traces of a duct leading from them.

The smaller cenocytes are more numerous than the larger. In
the early larval stages, they are, as noted above, much smaller
than the other type, attaining, in the old larvae, an equal or
greater size. Their distribution in the body is, however, very
much more irregular.

In the earlier larval periods the cenocytes appear much more
rounded in form than in the earlier stages. The nucleus shows
the same tendency. The cytoplasm presents a finely granular,
homogeneous appearance, and takes up rather evenly the orange
G and eosin. In these stages, however, the cytoplasm appear-
rather more dense than in the succeeding periods.

Wheeler says with regard to the ceimcyies of an unidcniilM-d
phryganeid larva on which he worked that the nuclei contain no
nudeole. In a 4.5 mm. stage of Pl<tty/>/iyI<i.\- (IcsiatHilm;, stained
with safranin, a nucleole is seen to stand otn \ ery dearly (Fig. i).

CEXOCYTES OF PLATYPHYLAX DESIGXATUS. 22Q

This apparently disappear- in later stages. The hulk of the
chromatin appear- in small, rounded granules connected to form
a skein. One easily observes several larger ma--e- of chromatin
-landing out rather conspicuously in the nuclei of the ienoc\
of all the larval -tages. The nuclear membrane appear- -harply
defined in all of th. .1 the imasjnal.

Be-ide- ,i relative increase in the size of the cell, there appear-
little change in the character ,,f the oenocytes up to the la-t larval
Ai this period the nucleus b< 'riionately much

uid mure irregular in outline. Th' 'la-m -ho\\ -. be-

en the nuclear membrane and cell boundary, an inner and
an outer /one <,r l.i\er. The more central of ihe-e i \\ o I.e.
-hou- th' ontain more and larger vacuoles, and

to ha\e ,i d' die outer, -mailer /one, on the

con md -mal' None »f tlie-e

ir, but -pile m. iv di-tingui-h in the l.i

b\ i ..!' ful :••' USinj . main mule-, which \\ould -eem

to ind: .f more than a hoi .nteiit.

Til' . . YVheeler

.iiu! oilier-, all o| whom iir them as indicatixe of

ler (1892 \ aciK.le- are

bin and an- not Thi- condition

the o-n mid ill a number of |.: nd I beliexe.

represents .1 normal i I )nc i^ reminded

'- \\ hi. -tic -lib-tain e

in iher .n."

In the |nipal si find- that the nucleu- ha- a more ir-

:lar outline, and th.it tin ia-m contain- comparatively

tf\\ vacuoles. It i- in th: e that the oenocytes attain their

l.n. .\\tli. 1'hi- .upleil with the appearance- ,.|

dvity in the late larval ! ma\ perhap- be taken as indication-;

of a great -i\it\ of the-e cell- during the-, i \\ «»

period-,.

In ihe imago, jn-I after enu imw marked

- of degeneration. The boimdar\ of the cell- ha- become

indi-tinct and fra\ed, and the outer portion of the cviopla-m

•.lined rather deeply \\ith the h.emato\\ lin Fig. ~ . At this

-tage. the nuclear membrane i- not di-tingui-hable, and but few

230 WILSON P. GEE.

vacuoles are to be noted in the cytoplasm. The chromatin has
gathered into irregular masses along the sides of the adjacent
cytoplasm (position of former nuclear membrane) and the linin
appears in masses which have separated from the chromatin
material. In an imago two or three days after emergence, but
which had been kept enclosed in a small dish, the degeneration
of the nucleus is seen to have proceeded further. The cell does
not, however, present an essentially different appearance to that
shown in the imago two or three hours after emergence. This
third-day imago was the latest stage of which material \\ as ax-ail-
able. It would be interesting to determine from older imagoes
whether the oenocytes entirely degenerate and are taken up for
the purpose of general nutrition of the tissues, thus affording a
reserve food supply in addition to their secretory function. This
latter view would, however, be hard to understand considering
the short life of these insects, and also the fact that egg-laying
goes on shortly after emergence from the pupal case.

Various functions have been attributed to the cenocytes.
Landois (1865) gave to them the name "Respirationszellen" and
assigned to them a place in the respiratory system. Graber
(1873), in his early work, speaks of them as unicellular glands, the
secretion of which was of unknown function; later, howi-vrr, he
holds that they are metamorphosed into the fat-body, and give
rise to blood corpuscles. Berlese (1899) considers the cenocytes
as excretory in function, as does also Koschevnikov (1900).

Anglas (1900) says: "Ce fait permet d'affirmer qu'ils s6cretent
autour d'eux des ferments. Aussi les considerons-nous cominc
des cellules glandularies (nees de l'hypoderme) et jouant le role
de glandes a secretion interne, mais de glandes dissoci£es." Ver-
so n (1891-2) considers them as secretory in nature as do also
Rossig (1904) and Weissenberg (1907).

Holmgren (1900) in Apion flavipes, experimenting by injecting
small quantities of powdered methylene blue and alizarin-cyanin
into the body cavity, came to the conclusion that they were
excretory in function. He found that the cenocytes took up the
color in much the same manner as the Malpi.uhian tulirs, tin-
nucleus of the cenocyte showing signs of staining somewhat rarlirr
than the nuclei of the latter organs. Three hours after injection,

(ENOCYTES OF PLATYPHVLAX DESIGNATE.

he found that the methylene blue had passed into the lumen
of the Malpi^hian tubes in sufficient quamitie- to odor them;
the oenocytes at the end of an hour and a half had lost their
color.

Following out this line of inve-ti-ation. a do/en or more -peei-
men- of Platyphylax Jcsi'^nalns were injected with an aqueous
solution of methylene lilne and the pan- di— ccied and remo\ t d
to a jJv<iTine mount. It v. a found that immediately afu-r
injection the oen< and the spinning-glands, SO prominent in

tlii- form, had both taken up the -tain the tenoeyte- t<( a much
1. - marki-d degree than the latter. In onlv OIK- in-tance. \\a-
it found that the Malpi-Jiian tube- had taken up the methvleiie

blue, and in this case the color was seen in but a -in-le tube. The
• iii' ;o«,k ii]> the -tain more in the micK-n- than in the , \ tO-

|'la-in. th- latter. houe\er, beini.; distinctly tinned. The Iar\.e
killed a halt hour alter injeition shou ed the tEnOCytCS more
decjiK : lined, but no . • ion was ob-er\ ed in t he Malpii;hian

tllbe-. l.alA.e killed niie hour .ll'tlT illieclioii -ho\vcd that the

Mal|.ichian tube- had be^un to take up the blue color, but that
the 01 and spinning-glands at tin- time \\ere becoming le--

inten-e in ilu-ir i o[( ,rat ion. The-r n--ult- \\ould lead one to
rather dilleieiit ( mu lu-ion- than tho-i- reached b\ llolm^nn.
It \\oulil be interred from the -imilarity of the reaction ot the
<i-ii' and Spinning-glands as detailed abo\e, that the former

h ot a d tteretu nature and inien^itx to that
ot the vpiiiiiiiie.-ij.md-.

I hi- \ie\\ i- further !u-neil b\ injection experiment ^

with sulphindigotate of sodium, l-'iirth 190 l)iee\'

|oii-i In- I'.edt lining der M a! i <\-^ hi'-chen ^chlauche oftenbart sil h
am h in un/\\eidt: \\'ei-e dun-h ihr \ermo-cn ,lir AII--

-cheidiin^ VOn 1 arb-totfen /u bewerkstelligen." >chimller I -
found b\ injection ot a -olutioii ot sulphindigotate oi -odium into
the bod\ cavity ot < '. yllotnlf'.i that the Malpii;hian tube-
excr< I'd tlu- -ub-taiiie \\ithin twenty-four hour-.

An ai|iicoii- -olution ot sulphindigotate of -odium \\a- injected
into the abdomen of a do/en or more ]ar\.e of Flnfyphylax ./r>.';--
tiatits 15 2O mm. in length, the in-ertion of the hypodermic -vrinue
1'einv. \ entral and 1 1 but tin- -olution ( •\i--ndin.u thoroughly

232 WILSON P. GEE.

into all the space between the hypodermis and the digestive
tract. Specimens killed within an hour after injection and
mounted as in the former experiment, showed a slight accumula-
tion of the solution in the lumen of the Malpighian tubes. This
was even more marked two and a half hours and four hour- after
injection, a diffuse blue appearance being then very apparent
throughout the cells of most of the tubes. The oenocyles, how-
ever, showed no signs of coloration at any of the periods observed
nor any evidences of excreting the substance injected.

Just what the function of these peculiar cells may be it is
difficult to say. In no case have I observed in the cytoplasm of
any of the oenocytes what could be interpreted as accumulations
of solid products of excretion. Their behavior toward methylene
blue, and sulphindigotate of sodium would seem to show that
they are secretory rather than excretory in function. The nature
of their secretion is difficult and practically impossible to deter-
mine. Measurements show their greatest size in Platyphylax
designatus to be in the pupal stage, and evidences from a purely
structural basis seem to point towards the greatest functional
activity during the late larval and the pupal stages. Their in-
timate relation to the fat-bodies seems vastly more significant
than the occasional grouping about a tracheal tube. Their rela-
tive increase in size, while of course proportionate to the increase
in size of the body, seems to bear a direct relation to the size
and development of the fat-bodies. It is also found that their
identity is retained throughout the period of metamorphosis.
Can it be that their function is the secretion of a substance or
enzyme which is of aid to the fat-body in its constructive \vork
or is it that their function is, as Anglas (1900) suggests, that of
internal glands of a dissociative nature, their secretion serving to
aid in the elaboration of products for general nutrition of the tis-
sues or for the disintegration of larval cells destined to disappear?
These are very pertinent questions, but ones to which a definite
answer cannot as yet be given. It seems safe to conclude that.
they are internal, ductless, secretory glands, whose function may,
in the light of recent progress in the field of internal secretion.be
for a long time a matter of conjecture.

These investigations were taken up at the suggestion of Prof.

CENOCYTES OF PLATYPHYLAX DESIGNATUS.

\V. S. Mar-hall, to whom I am indebted for many su.^e-tions
in the direction of the work.

/• ^LOGICAL LABORATORY.

I.K-I iv iii- WISCONSIN.

RKM-KKM 1 -
Anglas, J.

'oo Ol servatione -ur k-- nii'tamor -le la Guf-pe ct de 1'Al'oillo.

Hull ranee et Bd.uiqm-. XXXIX'.

Berlese, Antonio

'99 Ossei ilurante la nin in- iti

kiv. I 8.

Fabre, M.

'56 I • 1'in-ti: n-. Ann. S . n.it.

V I.

von Furth, Otto
'03 \ •

Graber, V.

'73! . Ai< h. inikro-;. Ai.

! : • i-

'95 Die 1

Hnini. ran, "•
'02 i

A: II.

Koschevnikov. G. A.
'oo I

III
KI'-A • .A.

'87 I- A nLluiiK •'• i M u-i idi 11.

' \ .
Landois. I .

'65 I • ' . X\'.

Pantcl, J.

'98 I i I Iii i\i- >n 1 i num. 1 .1 i • Hi.

Perez. Ch.

'03 ( '..iitiitur I'... 11. Soc. 1 n Ki 1-

Kii|i.< . \X\I\
Rossii . H.

'04 YI-M \\i-Kln-n • nlaivi-n u'-lit tin \<< 17. /ur HiMuiiK

i|i-I I'! X X.

Schindler. E.

'78 1'.' M.llj'l. '-.trll. /ritsrhr.

\\ i
Vaney, C.

'02 ("initiil'iitii'ii .'i r.'tu.ii- ill-- l.n\' - nn'-iai: let Dipt

Ann. lui\ 1 yon nouv. ser.)i Sc. Mnl.. Fasc. IX.

234 WILSON P. GEE.

Verson, E.

'91 Cellule glandular! ipostigmatische nel Bonil/yx inori. Boll. Soc. entomol.
Ital.. Anno XXIII.

'92 Altre cellule glandulari (epigastriche) di origine postlarvale. Richerche

Anat. Staz. Bacol. Padova, \'II.
Wheeler, W. M.

'92 Concerning the blood-tissue of insects. Psyche, VI.
Weissenberg, R.

'07 Onocyten von Torymus nigricornis Boh. Zool. Jahrb., XXIII. (Anat.).
Wielowiejski, H. R.

"86 Uber das Blutgewebe der Insekten. Zeitschr. wiss. Zool., XLIII.

EXPLANATION OF PLATE.

All figures drawn with a camera-lucida. X 630.

FIG. i. CEnocyte from a 4.5 mm. larva.

FIG. 2. One of the smaller a-nocytes from a 6 mm. larva.

FIG. 3. One of the larger cenocytes from a 6 mm. larva.

FIG. 4. CEnocyte from a 12 mm. larva.

FIG. 5. CEnocyte of late larval period showing vacuoles in cytoplasm.

FIG. 6. CEnocyte of pupal stage.

Kin. 7. CEnocyte from newly emerged imago, showing signs of degeneration.

BIOLOGICAL BULLETIN, VOL. XXI

PLATE I

• »

•
..

'•
'• \

•

.

•

7

WILSON P

THE MATING HABITS OF F( H K SPECIES
OF II IK BRACHYURA.

1 E. < HIDESTER.

KK 1'NIYIKMIY.

i. Impi.liii-ti'>n ...... ......................

................

3. M.ii .......

4- '
-S- I-

I . INI K1"!'! « 1 1« .v.

In I lie i < .111 -.1 activity of the rra\ ti>h

\\liich h.i >ied tnv .mention for tin- pa-t three years, I

me in- 'I in tlic mating hal.it- of \i i hn >\ » >da.

Durinj tin- MHHI: ; i'>. uliilc .uiin : iciii] inr.iry

istanl in I In.- \\i.«.il- Hull- l.il.i'iMi«.r\ ••!' tin- Miin-.iu o l-'ish-
ciir-, I li.nl iln- "|>pi>rtiinif, rve iln- in.iiiiii.' h.il>ii

fur .-liiiHiil \\illi tin-in t<>

(IftiTiiiinc it |io--il'lr their in. in: lin.itii.n.

In tin- p. MM i I >h.ill l.rirtK «!• ~. ril.t- tin- iv-ult- ol.i.iiin'tl .intl
iniil.ii \\ork iloin- |.\ \.ui<.u- in\ tors on ill'- otln-r

Arilm •;•< "l.i.

A. knou Ic.l-iiu-iit i- ill\- in. uli- to tin- Hon. Geo. M.

lioui i-. LJ. S. ( oninii--i"in-r "I li-linii S, l"i tin- opportunity to
\\oik in tin- \\ooiU Hole l.il.i.r.it. .r\ : .uul to |'ro|'i---or J. I'.
roller o| (I. nk I ni\i-r-it\ for n-.idinv: -ind iTitici-in.u thi> papi-r.

' )|;-l RVATIONS \\l> K.M'l KIMI-.N 1-.

Tin- species observed W< .•«////>-, tin- Mm- cral»;

C'/;;/Cfr irronitiis. tin- common rock cral.; (,irci)ins ma;>iiis, the
i;ivt-n crab; /'A;/ vc;/ V(-///r.v oi'i'llntnx. tin- lad\ « Tali.

\ i|»--. i ipiion of tin- differences between tin- male and the

female; the \ ir-in female; tin . .\ igerOUS female and tin- o\ igeroUS
female about to la\ , \\ill apjiK" in general for all of ilie -pi ,

236 F. E. CHIDESTER.

In the male the abdomen is narrow and the intromittent organs
or verges are the first pair of abdominal appendages. The paired
penes are formed by the everted posterior parts of the efferent
ducts. The penes transmit spermatophores down the groove
lit the verges into the seminal receptacles of the female.

In the- virgin female- the abdomen is much like that of the male,
except that the telson is narrower. Hay (15) discovered the
presence of a pair of hooks which project from the body and fit
into a pocket on either side of the abdomen, holding the abdomen
tightly flexed against the ventral surface of the carapace. The
swimmerets are very small n the virgin female. Crabs moult
several times during the summer, according to Herrick (16).

At the moulting the broad abdomen of the ovigerous female is
liberated from its narrow quarters in the old virginal shell, and
the female is ready for impregnation. The abdomen of the ovig-
erous female is held flexed only by muscular contraction.

A few days after impregnation, the female may be seen to have
two spermatophore plugs projecting slightly from the vaginae
and effectually closing the entrance. These plugs are probably
secreted by the glands of the male. Andrews has found that the
spermatophores of the crayfish are held in the annulus ventralis
of the female by plugs secreted by the glands of the male (2).
It is not known by me just when the plugs are forced out from
the vagina' of the female crab, but probably they are retained
until just before the laying. This would be the case if the crabs
are to be classed at all with the crayfish in which fertilization is
external, and in which the annulus ventralis acts as a temporary

sperm receptacle.

SPECIMENS STUDIED.

I • males.
M.iles. Virgin. Ovin<-r.m-. With Eggs.

Blue crabs 3 3

Rock crabs 3 2 7 i

Green crabs 2 2 2

Lady crabs 2 . 2 2

Iii the above table the virgin females are those that \\ere
brought into the laboratory and moulted there.

Distinction between the ovigerous females showing spermato-
phore plugs and tho>e not showing them will be made as tin-
experiments are d

M \I ENG HAB1 I- ' >F Mil BRA( HI OF \. 237

The crabs were kept in a sea water aquarium 2 X -1.' X 4
feet. In the case of the blue crabs, observations and experiment-
were made only on those confined in this tank. The other species
\\ere -mailer and individuals were from time to time remo\ed
io circular bell jars, <> in. high and 9 in. in diameter, with about
o in. of sea water in them.

Blue Crabs. — The female about to moult is seixed by a male
and carried about by him for a day or two until she i- ready to
-lied lu-r -hell. I hiring the pre-nioulting period the female is in
a normal upright po-it ion, being held by the -« < olid, third and
fourth pair of ambulatory appendage- of the male. It i- Mated
by I la\ l - that the male appear- able to distinguish the female
which i- about to moult. I \\a- not able to te-t thi- in the blue
crab-, but Iroiu tl, t\atioii- on the beha\ ior of the rock

era'1 < n irab- and lady crab-. I am inclined toquc-tioii the

Matrmrni and to 1" 'hat the male i- not able to diM ingui-h

the female about to nioul pt fr. .m the fart that -he i- less

:ii. I ha\e noti-d this -luggi-Onie— and apparent \\eakne--

in the i ra\ ii-ln everal days before moulting 7 .

\\licn ti < :le i- in the pi Mioultiirj. -he i- left foi- a

time b\ the mali I M n he approach- - her and turn- her over
so that her head i- beiic.ith hi- abdomen. I ua- unable tO
the lii-l ap|H"a< he- "I the male to tin -helled

lemale. but -cparated t \\ <• pair- man\ times and \\atchcd the

readjustments.

\\ hen the temale ha- been turned o that her \entral -urla< e i-
ap|)o-ed to that of the male, ami her hi ad i- -iuiated beneath hi-
abdoinen. both remain i|ir< - ral minuti-- nn!e-- di-iurbed.

Alter a |.i iii"l ..I Irom thirt\ -ecoiid- to ten minute-, the male
tap- the temale li'JnK \\ith hi !ir-t and second ambulatory
appendage- and turn> her -lov, 1\ around underneath him. The
male is nut, how \er, the only active party. No -ooiier ha- the
female been an-u-ed f .mi her -emi-h\ pilot ic Mate \<\ the mo\ e-
ment and tactile Mimuli of tlu' male, then -he be^in- to u-e her
ambulatoiA a|ipendaui - in turning herself. In order to deter-
mine it an external Mimulu- would c.m-e an i arly adui-tmeiit, I
tapjied lightK on the edge of the thorai it iv J< m of (he l\-m.ilef
b\ mean- of the tip of a -mall seeker. This \\ a- tried oil tl.

238 F. E. CHIDESTEK.

Micce-sive occasions and in r.n-h case tin- female began to move
herself into the copulating position by walking around the legs
of the male, plating her legs between and against his. In one
of these cases, tin- f finale moved so rapidly that the male may
have thought ^he was attempting to escape, for he beat upon her
carapace in the attempt to quiet her.

It is extremely in ten- Mini; to see such activity on the part of
the female crab, when one has as a contrast the passivity of the
female crayfish.

\Yhen tli^ female crab has turned about two thirds of the way
around, adjustment of the abdomens must be made. As above
stated, the abdomen of the ovigerous female is not bound by
hooks to thi- body and is held flexed by muscular contraction.
During the preliminary maneuvers, the abdomen of the female
i- Ill-Id slightly extended, and the male mn>t push this partially
extended obstacle out still farther and interpose his own telson
between it and the body of tin- female. In the case of the blue
crabs this is not difficult. The verges of the male are inserted
in the seminal receptacles of the female, and copulation takes
place. The male carries the female about, grasping her with the
tips of his ambulatory appendages, and holding her for hours,
sometimes days. \Yhile the act of copulation is going on, the
female is exceedingly quiet. The male is very pugnacious.
Bachelor male-, were placed in the vicinity of a mated pair of
blue crabs and were beaten away by the vigorous defense of the
male in copula.

Tin- two male blue crabs under observation were tested as to
their recognition of females, hard and soft, of the three other
specie-. In no case did a male attempt to copulate with an alien
female.

kork L'rahs, (irccn Crabs and Lady Cnihs. — These species are
all smaller than the blue crabs, and are not materially different
in appearance from each other. In the case of the blue crabs,
I did not succeed in getting a female with ^permaiophotv plugs.
In the case of the three -pecies under present consideration, I
did secure M\er,tl o\ igeroiis females that had been ferlili/ed
before they were brought into the laboratory, and also M-cured a
rock crab \\ ith egg-.

MATING BABITS OF Till BRACHYURA. 239

The method of capture of the female by the male is tin- same
in these crabs as it is in the blue crabs. The ability of the blue
crabs to distinguish by siidit. however, seems to be greater. In
handling the members of the different species, one is -truck imme-
diately by the remarkable visual powers of the blur crab. He is
alert to moving objects and knows when a body comes \\ithin
striking <li-iance.

Males of all three species were tested by removing the soft
female- and replacing them by hard males. They attempted in
all cases io copulate with the hard male-. Then iu the case of
the rO< k < rab, two male- wen 1 by placing \\ith them. hard.

ovigerous female-, in which the -permatophore pin-- \\ere not
protruding. In tin -lit- male- -ucceedcd in in-ertin;.; the

verges and actual copulation took pla-

In th- M- male- \\ith whom vvciv placed other male-, the

approach of ih< 'iil.'tin^ male to ihe -tr.m-er male

\\a- made in a -imilar manner to hi- approach to the lo-i parliu r.
\\hen ill-- stl d violently, the <>ther male attempted

to beat him into pa--ivitv iu-t a- he -iicce--fullv did \\ith the
female-, hard and fusl 1 ha\e | >rev iou-ly held in the

•:-h. I no\\ held that the ability of the male crab
to di-i iiu'ui-h tl llv a matter «\ inabilitx to hold and

copulate \\ilh the male iin li\ idual. It -eem- that discrimination
i- tactual and kin.e-ih«

Am-. n^ tin femali - captun-d for me \\ere fniir ovigerous lock

i rabs, t\\ 'i ci'ab- and t\\o o\ i-erou- lad\ crab-,

all of uliich had evidently been feriili/ed >ome time belmv, lor

lhe\ were -m - • --tully projected from a second teriili/alion.

about ;, mm. around the aperture- of the -eminal recejiiacle-.

the -hell Ua- -oft. Ill the o|>enin-- of the Vaginae \\ere -ecu I \\ o

\i-h vermicular plugs. These were pulled out, the operation

evidently cau-iii'., di-comfort to the female. The plii1^- \\en-

curved in such a manner thai tlu-\- appeared to have been moulded

in the va-ina-. At the internal end- were the -permatoph'
eni|itied and Mat. In order I rinine if the-e -permatophofe

plu^- were formed before they were vi-iblc from tin- out-ide,
I di— ecieil a female \v hi ch had been killed tWO days after copula-
tion, and found that the plugs were present, but that the hanK
outer 1'in 1- \\ ere m 't \ n'< >\ rudin^.

240 F. E. CHIDESTER.

Experiments were made \\itli males of the three species under
consideration by placing with them females of the same species
known to be fcrtili/ed and to have protruding plugs. In no case
was a in. ilc successful in his attempts to copulate. In the case
of one rock crab, the female was beaten into submission and the
male succeeded in touching the plugs. The discomfort caused
-4ie -li violent struggles on the part of the female that she was
released.

The female rock crab with eggs was placed with a male1 of the
same Aperies, but his efforts to overcome her resistance were
futile. In neither the fertilized and spermatophore plug-bearing
female, or the egg-bearing female is a true passive state produced
by the beating of the chelae and ambulatory appendages of the
male.

In the case of a large male and a small female, the adjustment
of the abdomens is very difficult. In the case of one large male
it was noted that the adjustment was not possible in the normal
position, and that he turned upon his back so that the female
floated above him and her abdomen was extended to a greater
degree. Then he slowly drew her into position, and having in-
serted his verges, assumed the normal position above her. In
all of the species studied, the female is not passive in the later
stages of adjustment, but uses her ambulatory appendages to
assist in the turning of her body. I should have liked to place
a soft-shelled male with a male that had just been separated from
.1 -.oft female, but was not able to do so.

3. MATIN* ; HABITS OF OTHER ARTHROPODA.
('rn.^tiK'i'a. — Herrick did not study the mating habits of the
lobster i in i but noted the fact that there were external seminal
pouches in the female. He states that the copulation is inde-
pendent of the moulting of the female. I lerrick mentions the dis-
covery by Milne-Edwards of a female ('inner (xi^ints which was
fcrtili/ed and in which \\cre \\liat M ilue-Kdwards at first look
to be the wands of the male, broken off after copulation as in the
insects. Later, Milne-Edwards said in reference to this observa-
tion that it seemed probable that the stoppers found in the
copulating poncho of t he female were "of the nature of spermato-
phores rather than a fragment of tin penis."

MATING HABITS OF THE BRACHYUK\. 24!

Holmes (17) found that in Amphipods the ovipo-ition occurs
after the moulting of the female. The males capture the females
before the moult and leave them for a few hours, resuming
possession of them after the moult. He found that tin- method of
recognition i- neither by sight nor smell. He removed antemve
and blackened the eyes, and -till the males gra-ped the females.
Holme- conclude- that the male ha- the instinct to seize and
carry another individual of tin- -ame -pecies, while the female has
ill-- in-tinct i<. lie <|uiet \\hen -< i/ed. He found that dead indi-
vidual- \\ere not carried and explained this by the lack of an
isional mo\.-ineni causing a -iniu:Je on the part of the male
lo reiain hi- hold. Holme- ha- mentioned a!-o the v, ork of
I • du on Arti-min. in which the latter found that the male-
ot Artemia cla-p the female- bv mean- of peculiarly -haped
antennae, and the two s > im about to- ether for several days.

I'ohni ||.,|m.-,. 17 noted that the male- of coprpod- while
mating, -uim about on the back- of the female- much as in the
. \mpliipoda.

Dearborn 9), Andrews 2, 3), Chidester 6, 7) and Pearse (26),

\\orkin- uith cra\ li-h, ha\e -ho\\ n ih.it munition i- largely
tacltial. . \ndir\\- di.-i nnulu- \ eiit rali- in < 'tis,

the Amei ii an ira\li-h foiim' of the Kockic-. Thi- -i ;

to retain the spermatophores until the female lav- her eggs. In
the rra\li-h, I'ertili/alion of the ci^- i- .\trrnal. Chide-ter

lonnd o that males grasp other males and attempt t<> copulate

\\itli them. I'ear-e found that male- attempt to copulate with
dead female- of the -ami- and different Species 26 , Audi'

holds that the male recognizes the temali a- a female because

-he d«r- noi resist, hencr tlu- dead or bound male i- taken for a

female. llr-a\- ; I - ttius possible that from the crayfish

-tand|>oint the miK difleivncr bi-tui-ni the sCXCS i- a diften-nce
in beha\ ior ,md not a difference in form, and moreover a d ff ere nee
i\e-l b\ mn-( lr and touch sense and not in effect upon any
other -i-ii • M-."

It -rrm- to the \\riirr. that although in the cra\ ti-li and the
lob-ter, he male- do not capture the female- and watch o\ er
them while the\- moult, the process i- analo-.m-. for the freshly
moulted female i- -non captured and Icrtili/ed. In the ca-e of

242 F. E. CHIDESTER.

the crayfish which have annuli, the subsequent copulations are
futile on account of the presence of the hard vermicular plug
deposited in the false pouches by the male. Again, in the lobster
and in the European crayfish, As (icns, although there are no
pouches, the spermatophores would naturally adhere to the
freshly moulted surface better than to a surface covered with
accretions.

The term "recognition" has been used by Pearse and others
without a proper appreciation of the fact that we have absolutely
no right to assume a true r cognition in the Arthropoda. An-
drews has brought out this fact in his statement regarding the
crayfish, Cambarus affinis. He says (3): "Sex 'recognition' ex-
ists apparently, only in the sense that the male may carry out
all the stages of conjugation if a female happens to be seized,
but not if a male is seized."

"Recognition," it seems to the writer, should be relegated to
disuse in speaking of the lower animals. The term "discrimina-
tion " may be safely used. Arthropoda may use vision, "contact-
odor," kinaesthetic or other senses in discrimination without really
"recognizing" sex.

Insecta. — Fer6, in a study of cockchafers (n) found that sexual
coupling failed to take place when the antennae were removed.
He also found that males which had just coupled with females
proved sexually attractive to other males. It is not just certain
that Fer£ was correct in supposing that the males were attractive
to other males, because the former retained the odor femina. He
found similarly in Bombyx that males sometimes proved attrac-
tive to other males (12).

Mayer, working with moths (21), found that the male moths
find the females by smell (Forel's contact odor sense?) rather
than by sight. Males flew to the abdomens of female^ which
were placed near the detached winged bodies. Males \\ould
pair with a female minus wings, but would not approach a male
with female wings. In the moths, there would seem to be a
distance-odor perception.

Berlese observed that in flies, there was only a slight activity
of the external organs of the male, and a great activity of the
female. The male mounted the female, but the female \\a> thru

MATING HABITS OF THE BRACHYURA. 243

the active party, introducing her ovipositor into the genital
atrium of the male ( 5

Maeterlinck's popular description of the life of the bee, states
that the male honey bee captures the female in her nuptial
flight ami leaves part of his copulatory appendages in her seminal
receptacles (20). The factor of natural selection is active in the
mating of the bee, for the strong males are the ones to capture
the females.

1 ielde iv. \Vlni-lrr .vp and others mention the mating of
ant 5i / 'imination i- 1 >y -mell. Forel calk it "contact-
odor."

Mit< hdl JJ . worked with mosquitws, finding ih.it tluj sexes
di-< riminati- I-.K li other by the -<>n-. The female tlic< into a
(1<. ml of dancing male-. It i- po—ible that di-criminat ion is
risnal at lir-l. rather than auditory a- Mi — Mitrhe'l belic\e-.

In conlmement, the female -tand- quietly >n the floor of her
e \\ilh tlie male U-neath and clin-im; with hi- body ]).iralk'l
to Inr-. Anv di-iiirb.im e which cau-e- the female to mo\c,
\\hen the p.iii aie •; Ull.it i llg, catl-e> the male to seize her

1 he mating i- ivpeaird e\'er\' hour or ohriier, \\hen the
male i- (\riinl IP\ a nioM-nient of the temale. Tin- ma\ 1 u-
luoii^hi alion! K\ a tap on the cai;e. When nm online. 1, one
male unite- \\ith -c\eral t«-inale-. A -in^le fert ili/at ion la-t-
foi five layings. Mi— Mitchell conclude- that tin- male con-
tinue- to approach the female l.v mean- of the olfadory -timuli
through the -en-ili\e hair- on the antenna-. 1 >i-tant locali/ation

B according to her idea, by sound ; ili-crimination of the quiescent

lemalein-ar.it hand i-l>y-mell. Moulting a- o, ( nrrin- l.efoi.-
fertili/atioii i- not meiitioiu d.

Mockard, \\orkini; \\ith thr \\ alkin.^--lick. Af>lof>us Mnycri
mini that male- mated with arliticial female- made by
fa-teniim ihe alidomeii- of tVmale- to -tick- and supporting the
-tick- on u iie 1< j

.\nnliniilii. Montgomery, 1903 23), and Porter, 1906 (30),
ha\e -ummari/ed prett\ fully the in\ estimation made on the
matins' habit- of -|.ider-. Both these writer- ha\e contril mled
much original in\ e-t Ration. I -hall not attempt to give all of
the literature, i vial i\ e to t he mating habits of spiders, but content
m\-el! \\ith a brie! -uinmary of the more important work.

244 F> E> CHIDESTKK.

The Pcckhams ha\ r taken up in detail tin.- courtship of spiders,
and hold that 127) In the spiders, Darwin's theory of sexual
selection is upheld.

Emerton (10) finds some variation in the ferocity of th female
spiders. He states that in Liuyphia and Tlicridioii, the male
and female live peaceably together in the same web and the
male is not attacked by the female during his approaches.

In A galena the male s the stronger of the two. He takes the
female in his mandibles and lays her on one side and inserts one
of the palpi; then he turns her so that she lies on the other side
with the head in the opposite direction and inserts the other
palpus. The female lies as though dead. Emerton holds that
the method used in all of the spiders is for the male to discharge
the seminal liquor on a little web spun for the purpose, and to
dip the palpi in it; then he approaches the female and inserts
the palpal organs into her epigynum. One palpal organ at a
time is replaced by the other. Emerton believes that few males
are eaten by females.

Montgomery has made most extensive studies of the mat ng
habits of spiders (23. 24). He holds that in recognition, touch
is the most important and sight is next in importance. He dis-
credits conscious sexual selection on the part of the female.

Porter agrees with Montgomery in his statement that the
female chooses that male which "first and most surely announces
by his movements that he is a male." Porter found that in the
Ar^iope (30) the male waits unc 1 the female has moulted, and
then successfully mates with her. He found that in some cases
the male left the web after approaching the female who was not
ready to moult and finding her hostile. After the copulation the
f male wraps the male just as she does any prey, and 1 aves him
hanging in the web with her old skin. Frequently there are as
many as five male spiders on one web.

Petrunkcvitch holds that sight is the only sense of sex recogni-
tion used in the hunting spiders. After sex has been recognized,
courtship begins and touch is the chid mran^ by which tin- male
excites the female and tests her willingness to accept him (29).
Petrunkevitch found that, in /tyv/crn, the male approached the
female whenever she br-.m to

MATING HABITS OF THE BRACHYURA. 245

Xuttall and Merriam have made a -tudy of the copulation of
ticks (25). They find that there are three distinct movements
in copulation. The male introduces his mouth pan- into the
sexual orifice of the female, dilating it and probably excitin- the
female; then he apposes hi- <>\\n -e\ual aperture to that of the
female and expels the spermatophore, whose neck adheres to the
tip of his hypostome and is then pushed into the vagina; the
t movement on the part of the male is to rein traduce his
mouth pan- and rupture the tip of the -pcrmalophoiv.

4. ( )ON< i.i -!• >NS.

1. In the four species of Brachyura -tudied. -e\ di-criniination
i- tactual.

2. In tin I'.: chyura, tVnili/ation i- effected \\hen the female
ha- n •« enlly nunilted.

3- Mali-- of die four species -tudied attempt to mate with
male- and lertili/ed female- of the -anie -pecie- but do not
allem|.l to male \\ith indi\idiial- of a different species.

4. In tin- !'•• ichyura, the female i- not entirely pa--i\e in the
mo\emeiit- \\hich preeeile copulation; -he re-pond- to tactu.il
-limiili v:i\ en by the male.

5. Inihe( in-- is a whole, copulation occurs at any time,

of the moulting of the I. -male. It i- highly probable
thai moulting i- a necessarj toiidition for -ucce— fill fertili/a-

tion.

<>. In the Crustacea the female i- u-ually pa— i\e during th-
ai l of copulation.

7. s« A discrimination in the Cni-tacea i- kin.e-tln-'ic and
tactual.

8. In the ( " ru-lac. a, the male- -em-rally die - i alter fertili/a-

liou. A -mall proportion of male- li\e for -ome time.

9. In the In-ecta. -e\ di-crimination is b\ -mell; |-'ore|'- "con-
ta» t-o.|or sense" i> the fully de-cribin^ u rm for a lar^e decree of

thi- discrimination.

10. Moulting ha- not been -ho\\ n to be a nece— ary forerunner

of Fertilization in the In-ecta.

1 I. The male- oh -ome -pi cies of In-ecta die immediately after
copulation.

246 F. E. CHIDESTER.

12. In some species of the Arachnida, particularly the spiders,
the females devour the males after copulation.

13. Moulting on the part of the female before fertilization is
not the rule in spiders. It occurs in some genera.

14. In the ticks, the mouth parts of the male are used to
moi>u-n and stimulate the sexual aperture of the female and
thus prepare the way for the injection of the spermatophore.

15. Sexual selection on the part of the female has not been
definitely established in the Arthropoda. It appears that the
successful male is the one who is strong enough or wary enough
to mate with the female, or who has learned soonest by his
senses the location of the female.

1 6. Since chance is a great factor in the presence of the male
at the proper time, we must consider that the successful male
is the one who first demonstrates his maleness to the female.
Though strength is a great factor, opportuneness of proximity
appears to be a greater one.

5. BIBLIOGRAPHY.

1. Andrews, E. A.

'95 Conjugation in an American Crayfish. Am. Nat., Vol. 29, pp. 867-873.

2. Andrews, E. A.

'04 Breeding Habits of Crayfish. Am. Nat., Vol. 38, pp. 165-206.

3. Andrews, E. A.

"10 Conjugation in the Crayfish Cambarus affinis. J. Exp. Zool., Vol. 9,
pp. 235-264.

4. Bell, J. C.

'06 Reactions of the Crayfish. Harvard Ps. Studies, Vol. 2, pp. 615-644.

5. Berlese, A.

'01 Copulation of the House Fly. Rev. Patol. Vegetale, 9, pp. 345-356.

6. Chidester, F. E.

'08 Notes on the Daily Life and Food of Cambarus bartonius bartoni. Am.
Nat., Vol. 42, pp. 710-716.

7. Chidester, F. E.

'n The Biology of the Crayfish. Am. Nat.
8 Darwin, C.

'86 The Descent of Man, and Selection in Relation to Sex.
9. Dearborn, G. N.

'oo Notes on the Individual Psycho- physiology of the Crayfish. Am. J.
Physiol., Vol. 3, pp. 404-433.

10. Emerton, J. H.

'78 The Structure and Habits of Spiders. Boston. B. \VhiuM.-n.

11. Fere, C.

'98 Experiences relatives aux rappmts li<>m<>~<'.\uels chez les hammetons.
C. R. Soc. de Biol., T. 5, pp. 549-55 i .

MATING HABITS OF THE BRACHYURA. 247

12. Fere, C.

'98 Experiences relatives a 1'instinct sexuel chez le bombyx du murier. C. R.
Soc. de Biol., T. 5, pp. 845-* ;;.

13. Fielde, A. M.

'05 The Progressive Odor of Ants. Biol. Bull., Vol. 8.

14. Forel, A.

'08 The Senses of Insects. Tr. by Macleod Vearsley. Mrthiu-n and Co.;
London, 1908.

15. Hay, W. P.

'05 The Life 11 • ! the Blue Crab (Callinectes sapidus). Bull. 1". S. F. C.,

>5. PP- 399-.;

16. Herrick, F. H.

'95 'I lie Ann ii. .,i; ' Bull. U. S. F. C., 1895. pp. 1-25 j.

17. Holmes, S. J.

'03 Sex Ki-< 'ignition um«mx Amph.; Biol. Bull.. Vol. 5. p; -12.

18. Holmes, S. J., and Homuth, E. S.

'10 I acll in • Ui,>l. Bull.. \".il. is. pp. 155-160.

19. Hewlett.

'07 ' : .

20. Maeterlinck, M.

'02 The Life of thi Be D I Mea & I \. \".. H;M.-. pp. i - .\\ -.

21. Mayer, A

*oo < »n tin- Mating In-tin, t in V nd Mag. Nat. I Ii~t.. Vol. 5,

Pi

22. Mitchell. E. <-

'07 .NT P. Putl

23. Montgomery. T. H

'03 Mii'li'-- "ii tli> II !il> tlii>-i- i.: the Malini; l'i-ii«iil.

I'' ;o.

24. Montgomery, T. II.

'10 I Sexual < - of

AT. in. i : . ! - V \-i 1 11. |.|i. ! 5 I -J -- .

25. Nuttall, G. H., and Merriam, G.

1 1 i ! n < >inr : ! \'. .1. 4,

PP

26. Pearse, A. S.
'09 Observal

•:. Am. Na( . Vol. |3, |'|>. 7 :

27. Peckham, G. W. and E. G.

'89 Ol • exual Selection in Spiders of the Family Attidae. i

Aiki-i> ami ' Miluau',.' • WU., 1889.

28. Peckham, G. W. and E. G.

'05 S 'itary \\a-p-. Il'.ii-Ju.iii and Mifflin, 1905, pp. 1-311.

29. Petrunkevitch, A.

'10 r.niit^hip in I ' Biol. Bull.. Vol. 19. pp. 127-129.

30. Porter, J. P.

'06 Tin- lla'.it-. In-tin. -t- an. I M.-ntal Pnwn -< nf SpMi-t -. Genera Argiope and

ICpi-iia. Am. J. m" 1'- . \'..l. i 7. pp. .;<>'.

31. Romanes, G. H.

'83 M Mlution in Animals. D. Appletmi and Co., 1883, pp. 1-520.

I?4'S F. E. CHIDESTER.

32. Stockard, C R.

'08 Haliits. Reactions and Matin.u Instincts of the "Walking Stick," Aplopus
Mayi-ri. Papers fr. Torttigas Lab., Vol. 2, pp. 43-59.

33. Washburn, M. F.

'08 The Animal Mind. Ma. -inillan. N. V., 1908.
34 Wheeler, W. M.

'10 Ants, their Structure, Development and Behavior. Henry Holt & Co..

N. V.. lc;l ii.

Vol. XXI. October, 1911. No. 5.

BIOLOGICAL BULLETIN

EXPERIMHXTS ON PATTERN-VISION OF THE

HONEY BEE.

• II. TCRXKR.

I vi K'Hir. ii, ,\.

In my |M|M-r mi ihr "Color \'i-i"ii « >f tin- H..IK-V Brr," pul>-
li-hrd in the BIIII..,.,I< \i. BULLETIN for < >rt.>u-r. 1910, some

<-\prrimriit- were «!«•-. -rilx-.l which, I think, dnn. >n-l ralr roii-
< ln-i\rlv ill. it I •!•(•- c.m di-t riminalr l>n\vrrn colors. hi .\H\cni-

IHT of ihai same year, J. II. l.o\rll' puMi-hrd the results of some

researches ><>iidii<trd in .1 IILUIIHT (jiiiir diUfn'iu from my
mclliod; Inn uliiili Kd him to form lln- -,mic n mclu-ion.

On pagi 277 of inv p.iprr is ihc follouin- p.ir.i^r.ipli : " Ho\\
miniiii- are the drt.iiU that l.f«-, observe I .un not pn-p.irrd to

say; Inn th.u iln-\ do observe details is indicated by the following

observations I In thr boxes u-rd in tin- i-\prriiin-iii- of s,.|-i,.^
l\ . iln- lr.i\- o| (he IMIM-, n-i d \\i-n- riitcrt-d, from ilu- purtici .

l>\ i in MI is oi an eccentric opening. In mo~i of the boxes this door-

u.i\ \\.i- K i I. iii-nl. ir. iii otluT- it u.i- i in nl. ir. \\"hrn .1 !.<•.• ln~t

approached one of these boxes, she had to search for the entrance.

. \lici .1 tcu \\-\\\~. -In- \\onlil Kind on tin- porliro dirccth in front
ot thr entrance, .ind, in -omr cases, -lir uould llv into ihr ir.i\-
\\ilhont r\cn p.ni-iii;< on thr portico. - lor .iliout ,m hour

3 li.nl ln-rn rollrrlin^ honry Iroin -omr of thr rrd .irtil.i<
h srrmril ih.it iir.nK all of ihr 1 iri's ill that part of thr held were
rollcrtiii- tioni tho-r artifacts. Almi^ >idr of one of thr rrd
!>o\r- I jilarrd a rrd 1 n>\ on ihr ^i<lr^ and top of \\hich I had pa -led
of \\ hit.- paprr. Thi- gave a 1 >o\ \\ith rrd front and >poll«-d
Into thi- l>o\ I plarr.l -oino honey. The bees that ap-

1 I. n veil. J. H., " I 1 1 , it tin- Hoin-y B.-c; Can BI-I-- I )i-tiiiL'iii~li ( , ,i,,;

.\tn<-r. \ilt.. \'nl. XI. IV.. Ni'V.. l';l". pp.

-49

250 C. H. TURNKK.

preached this box from the front alwav- entered immediately;
the majority of tln»e that approached tin- sides pau-ed a nionu'iit ,
then went to the nearest red box." This experiment indicated
that bees could di-criminatc between patterns. The purpose of
the researches to be described in thi.-. article was the establish-
ment of a definite experimental answer to that <]iieMion.

The- method decided upon wa- to permit a few bees to learn
that honey could be collected easier from artifacts of a certain
color pattern than it could be from ordinary ilo\\ers; and, after
that had been thoroughly learned, to see if those bees could
select artifacts of that color-pattern from those of a different
color-pattern; 1st, when the artifacts to be Delected contained
honey and the others did not; 2d, when not only the artifacts
to be selected but some of the others also contained honey; .vl,
when none of the artifacts contained honey.

The artifacts used were constructed out of cardboard and were
of three kiixU: circular discs about forty millimeters in diameter.

FIG. i. A few of the artifacts. From left to right: front view of a transversely

striped box, side view of a transversely strip<>>l box, front \ir\\ of a longitudinally
striped box, side view of a. longitudinally striped box.

cornucopias made in the manner described on page 260 of tin-
article published last year, and special experiment boxes made
in the manner described on pages 271 and 272 of that article.
The artifacts used this year were smaller than those used last.
The cornucopias, excluding the lip, were fifty millimeters long
and thirty millimeters in diameter at the largest part. The
boxes, excluding the lip, were- sixty millimeters long, forty milli-
meters wide and twenty millimeters high. The pattern that
distinguished any artifact was placed upon both tin- inside and
the outside. The following colors and color patterns were used:
plain red, plain green, alternate red and green longitudinal stripe^,
alternate red and green tran-\erM- stripes, alternate black and

PATTERN-VISION OF THE HONEY BEE. 251

white longitudinal stipes, mottled red and green. With the ex-
ception of the red of some of the plain red artifacts, the red and
green used on all of the artifacts was of the same hue. The
strip'-- were about five millimeters wide; the spots on the mottled
artifact-, varied from two to five millimeter* in length; they were
irregular in shape and arranger 1 in a haphazard manner.

Fl'.. -'. A IfV. -I ill' :

•;i"ttlr'l ; rii'linally ?iripr«l oitimoipia.

i-iid v i.-u ..| ,i loi

In order to h.i\ c tin- bet - \\orking under normal field ci UK lit ion-,
an abandoned in, ,1 !.,\\ was -elected as the -ite of tin- e\| lerimeiit
plot. M..-I ol 'in- t'n-ld \\a- rank with Melilolns ulna Lam.; but
hen- .nid then- were patches of almo-l barren ground. The
maioriiN D| tin experiment^ \\ere performed in one of ii
partb. barn n spots, in \\iiieh .1 feu scattered mint plaiil> and a
le\s | ilaiil - ol t he \\ hite 5WI

i i«'\ in i in l-.xi'i K[\II \ rs.

\ I . I July I } 17.]

The purpose of thi- preliminar\ series of i-xperimeiit - ua- to
permit the l-ee- to Irani that IKMU \ < mild In- ol.tained much
mole ca-il\ I rom artilael- marked \\it!i alternate red and -n-en
lon-itudinal line- than it could lie fnnn the llo\\t-r> of the nieliloi
or the mint from \\hich the\ \\ere fora^ii

I \ri ICIMI \i i. l:<:rly in the tiftcnioou of July 14, tico ili^ .
nnirkci! icitlt ffiirallcl red ami and .ell supplied icifli

honey, were attached to the brand. lilotns bush from ichich

'•re collect in^ honey.

Although the di-c- remained on that plant all of the alternoon,
no response was made to them \<\~ any of the be

I \ri ivi\ii \i J. Three discs, marked icith parallel red and
n stripes. .ttaehed to melilotns lici^s and a lar^e drop of

252 C. H. TURNER.

honey placed on each. These were placed in position at eight
o'clock on the morning of the fifteenth of July.

These discs were watched until two o'clock in the afternoon;
but not a bee made any response to them.

EXPERIMENT 3. At about eight A.M.. July 77, 1911, two disc*,
marked with red stripes on a green back-ground were attached to the
tops of upright sticks placed in the midst of a mint weed from which
several bees were collecting honey. By means of a string the bra nches
of the plant were drawn together in such a manner as to have several
blossoms near each of the discs. Honey was placed on the discs
and on some of the blossoms near by. Accidentally some honey
fell upon some of the leaves.

After waiting, restlessly, in the hot sun, for nearly two hours,
I was rewarded for my patience by seeing a bee alight on one
of the discs, and, while hanging suspended from its edge, sip the-
honey. When it had finished and before it started for the hive,
the bee made a short flight of orientation. A few minutes later
a bee alighted on the top of disc number two and began to sip
the honey. Before departing for the hive, it made a flight of
orientation. From this time to the close of the experiment,
thirty-five visits were made to the discs, twenty to disc one and
fifteen to disc two. Often bees were on both discs at the same
time. On one occasion, three bees were on disc two and two on
disc one. On another occasion, two bees were on each disc.
Soon after the bees began to collect from the discs I noticed
that several bees were collecting honey that had dropped on the
leaves. To check them, wherever I saw honey on a leaf I
covered it with dust. By this means I hoped to cause those
bees to notice the honey on my artifacts. The habit of col-
lecting from leaves had become fixed and those bees spent the
remainder of the afternoon searching the leaves for honey. Oc-
casionally they would find a new supply and coHect therefrom
until I covered it with dust. By the close of the afternoon, about
six bees were collecting from the artifacts and four hunting for
honey among the leaves.

EXPERIMENT 4. About two hours before departing for home,
disc two was placed in a new position and a cornucopia marked
with alternate red and green longitudinal lines was placed when-

PATTERN-VISION OF THE HONEY BEE. 253

it had been. Disc one was left in its original position. All of
these artifacts were well supplied with honey.

lp to the close of the experiment, eleven visits were made to
the artifacts; four to disc number one. three to disc number two,
and four to the cornucopia.

Exi'Kimn-.NT 5. Half an hour before departing for home, disc
nuniher o>: in a new position and a second cornucopia,

resembling nuniher one in its color pattern, was placed in the place
r<->:dfri'il I'niiint l>y (lie removal of disc nuniher one. (Cornucopia
nuniher one and disc nuniher tu >i the positions occupied

in experiment j. All of the artifacts .'ell supplied with honey.

Tin' bee- made tuehe visits i<> the-e artifact: four to disc
mi 1 1 ilier din-. i\\n to di-i- uuinlier \\\«. tlm > • nuco|iia nu ml " r

our, .UK! ilm-e to < "rnueopia nuinlM-r two.

K\perimeiit number thrrc \\.i- bejun .it about ei^hl A.M..
experiment number li\i- < |o-ed ,n fi\e I'M. Throughout the
\\hole o| ili. n interval tin- artifari- \\tTt- o|>M-r\ed e<>m inu<>u-ly .

< >n |i-a\in- |i.r home ,tll iif the artifact- \\ere \\ell -upplied
\\ iili li«>iic\ ,nid Ict'i in tin- lirld.

I In- bees ' "IN-' 'in-; li'i'in tli"-r .irtil.ici- \\crc <>nlv .1 -in. ill |n-r
cent. oi those visiting ih.it i .in "t the field : Inn the\ lu-h.i\ < d -<•
unlike I IK- "I In -r- ili.n I i mild recoyni/f on.- .1- -mm .i> it arri\ < <1
\siiliin .1 \.nd "I the artifacts. Tin- bee- ih.it h.ul imt lie^uu i"

i -n!l( « I h"lle\ |"n mi ni\ .'.nil.irt- nr holle\ --Ille.ued le.ixe-, oil

hin:: 'he e\|.t -i imeM.il plot, \\ould >iui]>l\ \\.uider from llouer
i" ll"\\ei. p. iii-iii- .i mmni-iit .it -<mie and -inipK Ll.inrin- ,n
othrr-. < )u the oilier hand. ln-i> thai \\en- colle. i in- hone\
Inmi m\ arlit'ai I- »r h"iu | red leaves, "ii leaehin- the plot,

\\nuld de-ciilii- a numliei- «if i-imiple\. irregular, mimlier eighl
curves, n<> tuo "I u hii-h -« eim-d io 1-e in tin- -a me plane and \\ hirh
became gradualh -h'-rtei- and shorter until the a nil .nt m- hone\ -

smeared leaf \\a- fmillil 1>\ llle bee. T le Inokin^ On, il \\a-

allllo-t imp"— ible to re-i-I the belief that tll»~e beCS WCrC -eekin:;
-oiiH- deliniie thin-. ..r thii

//. IJuly [8, I'M I.]

» Mi reachini: the field, at about eiijit thirt\' A.M.. there \\a- no
hone\ in any of the artifaet-: but the bee- were nio\in- in and

254 C. H. TUKM'K.

out of the cornucopias as though they \\vrr hunting for some-
thing.

Exi'i-.KiMKNT 6. One of I lie cornucopias used in the last experi-
ment was replaced by an experiment box marked with alternate red
and green longitudinal stripes. The other cornucopia was left in
place and called striped cornucopia number one. Three groups of
three short stakes each were erected near the plant containing the
two stakes mentioned above. The Croups were about a foot apart
and the stakes in each group separated by an interval of about two
inches. The Croups iscrc numbered from two to four, two being tin-
nearest to the two stakes and four being the most remote. The stakes
were arranged in a section of a curve. On the top of one of the stakes
in cadi group a cornucopia marked with alternate red and green
stripes was placed, while a similar plain red and plain green cor mi-
ned each of the other stakes. In no two groups was tin'
arrangement of the cornucopias the same. In the striped box and
in the striped cornucopias, honey was placed. The other artifacts
e empty. The position of the artifacts remained the same
throughout the experiment.

This experiment began at about nine A.M . and extended until
noon. Except during the time consumed in taking a photograph,
the artifacts were watched continuously. Not a single visit was
made to either the red or the green cornucopias: but forty-seven
entrances were made into the cornucopias that were marked
with alternate red and green longitudinal stripes. These visits
were distributed as follows: striped box number one, nine; striped
cornucopia number one, twenty-one; striped cornucopia number
two, eight; striped cornucopia number three, seven; striped cornu-
copia number four, two. Tints all of the striped artifacts were
visited. Sometimes a bee would enter one artifact and then
leave and enter another and sometimes yel another. It usually
sipped honey in the last visited artifact only. In some cases the
cause of leaving tin- first artifact was the presence of sonic
bee; in most cases, however., I could not discover the cause for
departing.

EXPERIMENT 7. .-1 fourth group of three stakes was added to tin-
above group and called group Jive: it was arranged about half way,
in a straight line, between croup two and group four. All of the

PATTERN- VISION OF THE HONEY BEE.

cornucopias were replaced by experiment boxes of the same color
patterns. At irregular intervals, the arrangement of the boxes in
each group was altered. All of the boxes marked :.•//// alternate
n and red longitudinal stripes contained honey, the other arti-
facts were empty.

Thi- experiment extended from noon to five P.M. Kxrept
\\hile tak-'nu a photograph, the artifact- v.ere \\atched cominu-
ou-lv. hiirin- that tinn- not a -.in^le entrance \va- made into
either tin- green or the red boxes; but eighty-nine entrance-
\\ere made into the -triped boxes; and each -uch lio\ was \i-ited
eral times. The visits were distributed as follo\\^: -tripe.!
\<n\ numlier one, ei-ln : -triped IK>X number t\\<>. lifieen; -triped
box number three, i\\e!\e; -triped box number four, seven;

:ped box number fl\ e. I U ellt y-one.

line of the cornui u-ed in thi- experiment leaked and

lormed. on the -lightly inclined -take, a line of honey. ^e\eral
be - disi overed this honey and began to carry it to the hi \e. The

line of hoiic\ was «n the -ide of the stake that ua- turned auay
from me and I did not knou that the bee- had formed the habit
of collec tin^ lione\ from the -ide of the -take until experiment
-even \\a- half over. I -a\\ bee- living to that -ide , ,f the -tak( .

bill I Ml|ipo-ed tlle\ Uete SOfTlC ot tile I " V- \\llo had fonilelh

formed ihe hab: - honey from -meaivd leaves, and,

-ince ! hail -ecu to it ill at I here \\ a- no hoin \ onanj "I the lea \ e-,
I di<l not i;i\e them much attention until I noticed that bee- \\eiv

!ie<|iiei)tly ll\in- Iroin that |i'a<e to the hi\e. A n a- I

ivali/ed that the\ had tormeil that habit. I co\ ered the hotie\-
\\ith road du-t, hopin- to induce the bee- to attend to m\ arti-
tact-. In-iead of entering the artifact-, the\ lontimied to ex-
amine the -take; -ionally one \\ould tl\' to another -take and
examine it. I hi- ua- not because thev had not noticed the
artifacts, but because the artifacts had acquired no meaniir.1 to
them. Thev \\oiild frei|tieiitl\ ap|)ro.ich one ( ,f the ! con-
taining honey and ho\er betore it momentarily and then return
to searching the -take-. To them the -take- had come to mean
" hotie\ -bearei - " a nd they -eenied to it -e t he art it act - and neigh-
boring plant- as reference points.

Mo-t .•!' the b. -e- . . .n tinned to de-cribe tho-.' complicated mini-

256 C. H. TURNER.

her eight curves, whenever they arrived 'at the experiment plot;
but the number of curves described were perceptibly fewer and
the lengths much shorter, and some of the bees seemed to fly
practically direct to tin- artifacts.

On departing for home, all of the artifacts, except three striped
boxes, were removed. Those boxes were well supplied with
honey-
July nineteenth, I arrived at the field at 9:30 A.M. and found
the boxes empty and the bees living into and out of them as
though hunting for something. It began to rain soon after my
arrival, which made it impossible to continue my experiments
that day. On leaving, all of the artifacts were removed from
the field.

Scries III. [July 20, 1911.]

The morning was quite cloudy and when I arrived in the field,
at about half past seven, the bees were not foraging.

EXPERIMENT 8. Three groups of three experiment hoxes each
were arranged in the manner described above. In each group there
was one box marked with alternate red and green longitudinal stripes,
one green box and one red box. The order of the boxes was different
in each group. When the experiment was about half over, the ar-
rangement of the boxes in each group was changed. All of the hoxes
marked with alternate red and green longitudinal stripes contained
honey; the others were empty. The experiment was an hour long.

Twenty-five entrances were made into the interior of the striped
boxes; but not a single entrance into either the red or the green
boxes. The visits were distributed as follows: striped box num-
ber one, ten; striped box number two, eight ; striped box number
three, seven. Only once did a bee, on arriving at the plot, make
the irregular number eight curves mentioned above. < >n arriving
at the plot, the bee would either lly directly to one of the striped
boxes, or else fly along, examining box alter box, until it reached
a striped box. On leaving, the bees did not al\\a\s make flights
of orientation. Often a bee would arrive at tin- hind end of one
of the experiment boxes that contained honey and attempt to
find an entrance; then it would sidle around to the front end ,md
enter.

EXPERIMENT <). Three groups of hoxes were arranged as in

PATTERX-VISTOX OF THE HONEY BEE. 2 57

experiment ei'^ht; but honey was placed, not only in the striped box,
but also in red box number one and green box number two. When
the experiment teas about half over, red box number one was placed
where striped box number one had been and striped box number one
placed on the stake wade vacant by the removal of red box number
one; and green box number two was placed where striped box number
two had been and that striped bo.x placed on the stake made rat-ant bv
lhe removal of <^reen box number two. The time occupied was about
toil' hour.

Twentv-two entrances were nude into the striped boxes, lull"
before .iinl hall" .ificr ihc change \\.i- made in ihr arrangement
of the boxes; lull not a -hide \i-it \\a- made into the interior
(•I ciihcr tin- n-d or \\\- . although one box of each of

iho-e color- ( out. lined hoiu-\ . Thr \i-it- \\riv di-tributed as
|o|l<i\\-: -triped box number one. thirte. n: -triped l>o\ nnmber
t\\o. lour: ^iriped |,o\ niinilier three, li\t-. ( >nr bee Imxered
belon- ihc hind end o» red box number one t'or -t -\eral M-COIK|»
• ind ihen lieu to -iriped I >> >\ mimlier one. Another bee alighted
on the I ron t end o| red bo\ number om- ,md then departed. I'
olten paused moniciitariK' before all of the boxes; but the\ never
enterecl an\ that \\ere not marke-l \\ith alternate red and i;iven
Ion-it udinal -tri:

I xri Ki\n \ i io. / nrniii'.'.i-il a\ in

:n any of the boxes ami

none hail ever , > duration of the experiment

abon!

Fourteen entrances were made into the >tri|K-d bo\c~ ; bm not

a -iinje \i-it \\.i~ made to the interioi" of either the green <>r the
red box< I he \i-it- \\ere distributed as follou-: -triped box

number one. -i\; -nipe<l box number t\\o. three; >iripeil box
number three, four.

I KPERIMEN1 II. / iirtifu. >ved from

the field; thus /<•.. '>i!y one '^roup of boxes, all of which ;

c»:f>ty and non, • ich had ever tontained ho>>ey. The time

allo, -out ten minutes.

Ten entrain v- \\ere made into the -triped bo.x, but not a ^inije

visit was made to the interior of either of the other boxes. \\ hen

I terminated the experiment, about -ix bee- were lio\rrin;j about
the Mriped box. examinin- it carefully.

C. H. TURM-.R.

Series I V. [July 21, i<)i i.]

Exi'KKiMKNT 12. Two sets of free boxes iccre arranged on
stakes. I)> each group then- was one of each of the following arti-
f acts: box marked with alternate red (ind green longitudinal strides,
box marked with alternate black and white longitudinal stripes,
box mottled id III green and red, a plain red box, and a plain
green box. The arrangement of the boxes in each >^ronp was dif-
ferent, and was changed tii'ice during the experiment. The box
marked with alternate red and green longitudinal stripes was sup-
plied with honey; the others were empty. The time consumed was
about one and a half liours.

Sixty-three entrances were made into the boxes marked with
alirrnair n-d and grec'n longitudinal stripes and one into the
boxed mottled with green and red; no visits were made into the
interiors of any of the other boxes. Bees frequently hovered
before other boxes and twice bees alighted on the ponieo of the
box marked with alternate black and white- longitudinal stripes,
but did not enter. The visits to the artifacts marked with
alternate red and green longitudinal stripes were distributed
forty-two to box one and twenty-one to box two.

Exi'i.KiMKNT 13. One set of six boxes was arranged on stakes.
To the kinds of boxes used in experiment 12, I added a box marked
with alternate red and green longitudinal stripes. The box marked
with longitudinal red and green stripes was supplied with honey; the
others were empty. Starting at the left, the arrangement of the boxes
as follows: plain green, marked with alternate red am! green
longitudinal stripes, mottled with green and red, marked with alter-
nate black and white longitudinal stripes, plain red, marked with
alternate red and green transverse stripes. The time consumed
was about half an hour.

Nineteen entrances were made into the box marked with
alternate red and green Ion-it udinal stripes; but no visits \\cre
made lo the interior of any other box.

l.xri KiMt'.xi 14. The same number ami kinds of boxes were
used as in experiment thirteen; but the mottled red and green box
placed where the box containing honey had been and the box contain-
ing the honey icus placet! on the slake rendered vacant by the remortil
of the mottled box. The lime consumed was about half an hour.

PATT] RN-VISU '\ ' •!•' ! HI- I!i iNl- \ HI I . 259

Twenty entrance- were made into tin- 1>"\ marked with ulu-r-
naic red and green longitudinal stripes and one im<> the box that
was mottled with red and green. No \i-it- \\--re made into the
interior of any other box.

Kxi'i KIMI.N i 15. The same number and kinds of
used us in experiment fourteen; but the box marked icith alternate
red mi/I '^reen transrersc stripes isas plaeed ichcre the box containing
the honey had been, and the box containing the honey icas placed
li'here the box marked icith alternate red and ^reeii transverse stripes
IKK! been. The time consumed :cas about three-quarters of an hour.

Thirtv--i\ entranee- \\ere made int«> tin- box marked with
alternate red and ^iveii 1« uiuit ndinal -tripe-. HX into the box
marked \\ith alternate red and -iven transverse Stripes, and two
into the bo\ m.irki-d \\itli alternate blark and \\hite longitudinal
>tri| \o \i-it- uere made into the interior ,,(' ,tii\ other bo\.

1 In- bees that selected the \\ron- bo\e- did not enter them as
I'p'inptlv a- did tlm-e that entered the ri^ht box. In eaeh case

the bee llo\ ereil b< |.,|- I lie ell I IM Hi 'e ( | II i t e a \\ Ilile bel'ofe entering.

All o| these mi-tak. made during the !ir-t ten minute- of

Illellt .

I •. \ri KIMI \ I i<>. / . •//// alter>: d and

itudin,: marked i.ith alternate red

find •>! stakes, in a line, icitli

all ' '.-. l-'or the 'irst half

5 alternated: during the
latter halt'. ///, : ndinal stripe in the middle.

: ndinal stripes contained honey, the other* .
•!y. Th( tin :tt half an hour.

The pnrp" this expeiiment \\a- to a< en-tom the bee- to

i^noiiiu the tran-\ i r-el\ -trijied bo\e-. Thirt \ -t hive entrances
\\ere made into the longitudinally -trit>ed bo\e- ,md none into
an\ • 'l the • 'tin :

I \ri KIMI \i 17. /• ••.'• number and kinds of in-
used as in experimei. >: and they . -I in the follo:cin^
order: box marked icith alternate black and ichitc longitudinal stripes,
box marked icilh alternate red and <^rccn transrerse stripes, box
marked icith alternate red and ^reen longitudinal stripes, plain
>i box, box )>: ;.'//// red and <^rccn. The box marked icith

26O C. H. TURNER.

longitudinal red and green stripes contained honey; the others teere
empty. The lime consumed teas about fifteen minutes.

Eleven t-nt ranees were made into the box marked with alternate
red and green longitudinal lines; no entrances were made into
any of the other boxe-.

Exi'i KIMI:\T 18. The same kind and number of boxes teere used
as in the above experiment, hut they tee re arranged in the following
order: box marked teith alternate black and teliite longitudinal
stripes, box marked teith alternate red and green transverse stripes,
plain green box, box marked teith alternate red and green longitudinal
stripes, box mottled teith red and green, plain red box. Xone of the
boxes contained honey or ever had contained honey. The time con-
sumed teas about fifteen minutes.

Thin v entrances were made into the box marked with alternate
nil and green longitudinal stripes, but no visits were made into
the interior of any other box.

1 \IM KIMKXT if). An empty box marked teith alternate red and
green longitudinal stripes and teliich never had contained honey
teas placed on the palm of my hand and by its side teas placed an
empty box marked teith one of the other patterns. The entrances
of both boxes teere made to face the same teay. All other artifacts
teere removed from the field and my hand held near the stakes upon
tehich artifacts had rested. At the end of about tteo minutes the
position of the boxes teas altered. This maneuver teas repeated
teith each kind of box used in experiment eighteen.

At least a hundred entrances were made into the box marked
with red and green longitudinal stripes; but not a bee entered
any of the other boxes. One bee acted a.s though it was going
io enter the box marked with alternate red and green transverse
stripes, but it did not do so. Towards the end of the experiment
t lie bees entered the box marked with alternate red and green
longitudinal stripes M> rapidly that it was almost impossible to
keep an accurate record. To one looking on, it seemed as ihoiigh
the bees were looking for something which they expected to find
in those boxe-.

When I left for home the bees \\en- searching over and around
the barren stakes for something that \\.is not.

PATTERN'- VISION OF THE HONEY BE1 . 26 I

INTERPRETATION OF THE EXPERIMENTS.

These experiments furnish additional evidence, if such is
needed, that Plateau is mistaken when he claims that odor is
the sole factor that leads bees to the honey of Hovers: for my
di-< -, marked with red and green stripes and well supplied with
honey, remained exposed to bees for fully two day- before a
-ingle bee collected honey from them [Ex. 1-5].

These experiment- -trengthen the impression, gained from
previous work with hymenopterous insects, that such insects
can lie induced to form new a— ociations that will influence- their
behavior, it" only the experiment is so planned as to lead the
in-e( i to accidentally make the a--< .nation. So far a- my experi-
eno . ii i- almo-t impo--ible to coerce them into forming

nev. iation-. and an abundance of time i- ohm ivquiied

for i he !.. rmation of chance associations. Lack of response to a
-liimilu- does not mean that that -timulu- ha- not been noticed
by tlie in-ect-; but that, to them, it ha- not vet acquired a

meaning.

In induci: -, in the held, to re-pond to new food-object-,

the ne\\ associations are formed much more quickly when the
1 1 i- •'! t" \ie\\ than \\heii it i- hidden. The\ ma\ In-

induced to form -uch associations, however, even under the latter

condition-,. "l"he-e -lateiiiein- are -upported by the fact that
la-t year, when I conducted a lengthy series of experiment- \\iih
di-c- an<l cornucopia-, artifact- which did not hide the honey
In. m \ie\\. I soon had more bee- re-ponding to the artifact- than

[ could keep an accurate record <.i : whereas this year, when I laid

a-ide my di-c- and cornucopia- .1- -oon a- about half a do/en
bees had made the a--ociation, at no time wen- more bee- at
\\ork than could be counted. At the clo-e of the series about
twice a- many bee- \\ere collecting from the artifact- as at the

lining.

\\lien a bee had di-co\ered one of my honey-producing arti-
lact- ami collected therefrom, it would make a flight of orientation
and then lly home. < h\ returning to the field it would, by mean-
ol a -eric- ot ever narrow inc. complex, figure eight curves,
search out the experiment plot, and then, by a closer scrutiny
ot the surrounding obji-cts, locate the artifact desired. [Last

C. H. TURNIvR.

paragraph of Series I.] After a bee had made several trips to
tin- plot, it would rly directly to it without making the compli-
cated searching movements. This striking behavior furnished a
sure- means ol telling when a new bee had formed the association.
I changed the arrangement of the individual artifacts so often
th.it tlu- bees usually were forced to search for the honey-produc-
ing box. After tin- asocial ion had been well established, the
bees ii-ually departed tor home without making a careful flight
of orientation. It,ho\\c\er, I had made a marked change in the
position of the artitact since the last visit of the bee, then a care-
ful Might of orientation was always made. Xo one who had
watched these pronounced searching movements could doubt that
bee- are largely guided by memory pictures of the environment.

After a bee had learned, by experience, that artifacts bearing a
certain color pattern contained a more copious supply of easily
obtained honey than ordinary flowers, 'it would select artifacts
bearing that color pattern from those marked in a different way.
This was true: (i) when several of the artifacts to be selected
\\ere scattered among a number of plain artifacts of the color-
used in making the color-pattern [Ex. 6-1 1] ; (2) when the artifact
to be selected was scattered among several other artifacts, some of
which were plain and some of which were marked with patterns
unlike that of the artifact to be chosen [Ex. 12-15, 17, 1 8]; (3) when
tin- only difference between the artifacts was that one was marked
with transverse and the other with longitudinal stripes [Ex. id):
i when the artifact to be selected contained the honey and
the others did not [Ex. 8, 12-17]; (5) when honey was to be found
not only in the artifact to be selected, but in some of the other
artifacts also [Ex. 9]; (6) when none of the artifacts contained
honey |Ex. to, II, 18, 19].

All of the facts mentioned in the above paragraph are recorded
in the follou in- table.

Although this table shows conclusively that bees can recognize
color patterns it does not tell the whole story. It does not show
that t\\o of tin- mi-takes \\ere made in selecting a box mottled
with red and green for one marked with longitudinal red and green
stripes; that two mistakes cons'sted in selecting a box marked
with black and \\hite longitudinal stripes lor one marked with

PATTERN-VISION OF THE H'>M;Y BKK.

263

I.ATED RESPONSES OF HONEY BEES TOWARDS PATTI K\^.

The bee to select an arti!.-i< t marked with alternati

tudinal •.iri|n-s wh'-n .i-vi:i.tu-.l with

j

—

-

:

. -
-

Plain 11 'I .tii'l plain j-n-i-n ariita' <[iial numl".-r "1 tin-

thr'-c kind- Iji-inn n-i-'l: wln-n tl.i- uriiiavi in l>i- -i-lt-rit-d

-.liiit-i! Imni-y ami tin- ntln-t- did nnl ................ if.i ir.i o IOO.O

Plain n-d and plain ^in-n ariit.ut-. an i-qua! miml»-:

n: I In- tlin-i- kind- Ix-ilii; u-rd ; u hen tin- ,11 t it art - in In- -i-li H trd

1 -niin- nl i -a. h nl tin- nthi-i 1 ind- inniaini-d hniic\ . 33 IOO.O

Plain : jilain x'( '•» -<i n <-<|iial iHMiil»T >•] <

kind-; iinin- nl tin- ait: .taiii'-d ' . .1 i uo.O

Arlilait- ni.ni.i-d uilli ahi-i • ; and HM-<-» Iran--.

nnl liniu-y. lln-

• illicr -. ui-ii- ••nij • . , c, 100.0

• a, plain n-d.

umiilcd n-d and gri i \\itli a • Mark and

• idinal -ti i| •

li'd llnll' , 1

( l||,- n| c-.ii ll n| I 1|. |. ,||, IV

nmltli-d |i-d ai .nid

\vh • ite red

ii-i t,'d

< >IH- of each of tl Q, plain i<-d.

inntlli-d n-d .u .,nd

wliiii- Inni'.itiid; \\illi a! nid

M.-d

Imiii -.

An\ in t!i«-

di.-J a ci|iial liuiiilii-i

Mllli'd !, , U ........... 0

•l.il

n-d .mil v;ict-ii longitudinal -tripi--. .nid lli.it llir ullu-r -i\ mi-l.ikf-

consisted in -r|( t i in- .1 |m\ ni.u-kcd u iih ml .nnl green transverse

-~lii|ic- t"i "in- m.iiknl \\illi ii-'l .'.ml -n-ni Imi^ii udin.il --iri
And il d"(-- ii"I -ln>\\ lh.it tin bees SOOn li-.inu-d I" ,i\"id in.ikini;
r\ i-n ! In -r nii-!,iki

Tin -IT \\ ,1- nun h in tin- lu-h.i\ iiir "t tlu--i- bees I" indii .He tli.n .
in thi-ir al>ilil\ I" di^i iiii;iii-li niinuii drl.iil-. tlu-y an- m-.ir
-i-hlrd. ( )n lA.imininv; tin- .util.nl-. ritln-r \\ln-n -i-.n-rliin^ for
.in .init.n i nr in making ii- lli-ht nt" orientation, tin- lu-c ,ilu,i\-
lio\rn-d uitliin about a rcni inn-HT of tin- ol,jivi examined. Tin-
lii'-t !M.\ a lu-i- tvarlird, oil arri\ inv; at the ]»|o|. \\.t- u-ualK'
not ilu' doircd our. and, after a momentary examination, it
\\< mid |>a— on to tin- next .

264 C. H. TURNER.

Evidently bees can distinguish between color-patterns, and
this is of value to them in recognizing plants that yield honey.
I K-IKV, since insects can distinguish colors and the fine details
of color pattern, there is nothing about the visual powers of bees
that militates again-i the theory that the colors and the color
of Mowers are adaptations to insect visitors.

SI-MMKR UK. H SCHOOL.
ST. Lovis, Mo.,
July 22, in.

PHOTOTACTIC REACTIONS AND THEIR REVERSAL
IN THE MAY-FLY NYMPHS HEPTAGKMA
INTEKIM'NCTATA (SAY

J. E. WODSEDALEK.

REACTION-, n> LH.IIT.

\\"ln-n nymphs of the may-fly Hcptu^cnia interpunctota are
placed in a Ion- :Ja — di-h i.f water near a window, they im-
mediately -uim away from the light, and upon reaching the end
of tin- di-h they repeatedly collide \\ith and cla\\ against the
invisible ob-iructioii. The-e nio\emeiit- may l»e kept up, \vith
interval- <if rest, for a uhole da\\ \\"heii (|tiiet. the n\ni]ih- do
imi alua\- face a \\.iy IPUU the li:cht, nor \\hen they -\\iin do
the\ alu.i iii a -trai^ht liiu- aua\ from it. (Mini if there

are lar^e dark ol»;e(t^ in the room, lhe\- \\ill > \\ini a^ain^t the
-ide o| the di-h in the direction o| the olijci-t^.

A- ihe lieha\ior o| die nxnipli-, i- often much allertcd |i\-
olijcct^ iii tin- nei'Jiltorhood. nio-t of the experiment-, on the
reactions to li;^ht \\en- pcrtormed in a dark room in a lar

evenly colored box constructed of white cardboard, thu^ >lnn t in^

o|| all oliject^ and unde-iiaMi li'Jit. The liv.lit u-liall\- emplo\ed
\\a^ deri\ed liom an onlinar\ -ivterii laiulle pourr incainle-ceiit
lamp attached t<. a uire and movable about at \\ill. Alnio-t
\\iihoiit , Aceptioii, the n\ni])h- -\\.im a\\a\ from the li-ht. » '• -
ca-ioiiall\ . -oine -pn inieii- alter repeatedly clauin^ a^ain-l the
transparent barrier and not -e( uriii;^ a hold \\ould fall down on
their back- uith their le.i^ >tickinv; up, remaining in that po>iti<m
Munetime^ for hour-,. If a -tone \\a- placed in the end of the
ba-in neare-t to the li'Jit, the n\ niph would attach it-elf and
remain there for days. When tin- li.^ht \\ a- brought near the
-tone and m-.\ ed about, the- iii-ec'l would -i-ek the -haded area.
but liijn i-\en much more inieii-e would >eldom lorce it to de-ert
the -tone entirely.

When a piece of white cardboard i- placed on the water in a
small ija-- di-h. the nymph- -ooii attach ihem-el\e- to the lowi-r

265

266 J. E. WODSEDALEK.

side of it. By illuminating that side of tin- paper they soon
crawl upon tin- top which is shaded. Then, when the strong
light is again thrown from above, they move below, and in that
manner can be made to nn>\e back and forth repeatedly, though
not with Mieh precision after the experiment is kept up for some
time. The nymphs respond to light most readily when occupying
the upper sjdi- of tin- paper, and the best results are obtained
\\hen white paper is used. Ouite similar results are obtained
b\ using gray or black paper, but in such cases the nymphs lake
a much longer time to move trom one surface to another.

I have performed quite a number of series of experiments with
different groups of nymphs using two lights, differing in intensity,
one at each end of a glass basin twenty-four inches long. I
noticed that by placing the nymphs in the1 water half way between
the two lights they would often continue going in the direction
in which they first started, regardless of the intensity of the light,
though many of them manifested much promptitude in going
towards the end of the dish from whence came the light of less
intensity, even when there was a difference of only a few candle
powers. The excitability apparent in many specimens at the
instant of being placed in the water, which is largely responsible
for their reckless behavior was almost totally reduced by placing
the nymphs in a narrow tunnel made of glass strips, which was
placed in the middle of the large basin at right angles to the rays
of the two lights. The tunnel was sufficiently wide to enable
the specimens to go forward, but not wide enough to enable
them to turn around in it. Then the nymphs were put in at
one end and allowed to pass through two or three1 inches of the
tunnel, thus getting the full force of the two lights at the same
time, and also a chance to resume a more normal attitude, Uy
the time the end of the tunnel was reached, which was in line
with the two lights, it was only a mat ter of choice which direction
was to be taken. Sometimes the nymphs would manifest no
choice, but remain in the middle ot the dish for several minutes,
especially when the two lights differed little in intensity. Hut,
generally the insects exinced considerable precision in choosing
the end of the basin from which came the light of less intensity.
The results obtained in some o| the experiments are given in the
following tables.

PHOTOTACTIC REACTIONS IN THK MAY-FLY.

267

TABLF I.

:'-S.

Turn> I Hward
80 c.p.

Indifferent.

Turns Toward
16

1 "tal.

I

O

O

IO

IO

2

0

0

10

IO

3

0

10

IO

4

O

0

IO

IO

5

0

O

10

I"

'•

0

[0

in

i

o

60

TABLI II.

i

-5

i

i

'1

i i

1

-5

1

TABLI III.

i

i

in

i

f, HI

I..

in

in

i

in

i

'

In tnrlhcr lAprrinifiit- .in .ipp.ir.it u- essentiall) th<- -.inn- as
ih.it UM-d 1>\ Strasl :u hi- uork on rrnti.\t<i u.i- ciii|il<'\ .-d.

It con-i-trtl <•!' .1 |>ri-m i.l' .1 -«ihilii>M <>l Imli.i ink |)l.uc<l over .1
L.III; glass tniU^ll tin- sides .unl lM,|t(,ni of \\llicll \\riv IM
over \\ith \\hiii- ]I.I|HT I ' n spe< inn n- \\rn- ]il.u'i-il iii tin- r
of tin- trough uml.T tin- thin portion of tin- pri-ni, .uul uhni
I'.iirU i|iiirt tin- li^ht \\.i- ri'lln tnl ] M T] .mi i icul.irly through tin-
pri-in i;i\iii^ .t perl .i<l.ition in tin.- intniMtv of li-ht troin

one mil of the troii'Ji t«. tin- otluT. Th rev <>f tin- specimens
immrili.itrly -\\.ini to\\.ir«l tin- «1. irk i-ml of tin- trough ; four otln-r»
fo|lo\\cil \\ithin h.ill .i niinntr; and the rfin.iinin- three collected
into .1 -ioup n-in. lining th.it \\.tv for ,i lV\v ininutr> \\lx-n tlu-y

J. E. WODSEDAL1 K.

were separated, and within another minute also left for the dark
area. Sometimes a specimen would return to the less obscure
end of the dish and not finding anything favorable to its thigmo-
tactic inclination, it went back whence it came from. Thi> ex-
periment was repeated several times, always with similar results.

The light was next permitted to enter the trough obliquely,
the-^hicker end of the prism being next to the light. Again five
of the individuals made for the darker end of the trough almost
immediately, this time going against the rays of the light. About
half a minute later two other specimens followed and within
three minutes all had deserted the light area. Thi^ experiment
was repeated many times with ten different sets, and always in
less than one minute a large majority of the specimens were in
the dark portion of the trough. There were almost always a
few slow specimens which required a longer time, from one to
ten minutes and even much longer, to be moved. If left in the
trough, almost invariably at the end of a few hours the nymphs,
with some exceptions, would gather into a group in the obscure
region and remain there indefinitely. If the prism and light
were reversed, leaving the entangled colony undisturbed in the
more illuminated end, the colony would slowly break up and with-
in several hours again form in the more obscure territory.

Another apparatus was so arranged that the length of the
prism could be increased or lessened by tipping the trough holding
the solution, at various angles, and thus augmenting or diminish-
ing the contrast between the two ends of the trough. It \\a-
observed that the proclivity of most nymphs to choose one end
of the dish in preference to the other varied directly in proportion
to the contrast in the illumination of the two extreme regions
of the trough. Here again, we find that some individuals react
far more readily than others.

It was also observed that some nymphs under ordinary li^ht
manifest no signs of agitation when in a uniformly lighted area,
yet when an area in their neighborhood becomes slightly shad< d
they soon aggregate there.

CONTROL OF PHOTOTACTIC RF..V riONS HY ( "in \IKALS.
It was found in the experimental \\ork on the reactions to
light that II. intcrpunctata nymph-- are practically all negatively

PHOTOTACTIC REACTIONS IX THE MAY-FLY. 269

phototactic, but that the different individuals vary in the in-
tensity of their negative n -ponse. Also, out of about five or
six hundred specimens several were quite indifferent, and a tew
were even \\cakly positive. This obvious variability in response
among the different individuals, in an identical external environ-
ment, is apparently due to the variability in tin- internal con-
ditions of the various specimens.

1.0(1,' says that in all probability light produces chemical
change- in the eyr <>r -kin of the animal-, and that these changes
are re-pon-ible for tin- heliotropic react i« >n-. In a brief pre-
liminary communication2 on the control of heliotnipic reactions
in fre-h water crustaceans by chemical-. he says that specimens
ol (inninmrns (nilcx, which are naturally negatively heliotropic
can be made po-iii\r b\ -pecilir chemical -ub-tanc. •-. 1 •". -r ex-
ample, if carbon di"\ide i- allowed to bubble through tin- water,
or it ihe animal- are thrown into \\aier previously charged \\ith
i arlion dioxide. ihe\ ai on, ,• become po-itively phot, iiaclic. The
-a me i. nib- were obtained \\ith othei" a. id-, a- hydrochloric,
oxalic and aCCUC, but boracic acid produced no -ncll ettecl.
AIIIOII- the salts, lie found that onlv tli.»-e of ammonium could
be ( 'impaled in their ell. < i- \\ ith tli id-.

Ja< k-on .\pei-imeiitc.l \\ith another amphipod, //:.:.'. lid
knit kfr'tnx kcri, \\ hich i- al-o nev;at ivel\- phoiotactic. and obtained
re-ult- similar t.> tho-e ••! I ..>« 1,, except, that Ilynlcll.i" were made
p..-ili\e b\ ! id. Tartaric acid, ho\\e\er, |iro«luci-d no

change in their reaction. \Yith the -ab- and alkali.-- it \\a-
a-aill found that there -eenied to be no relation between the
cla-- "I > liemical u-ed and the reaction-.

Ma-t' found that A '•: larva' \\hich are iKirmally |>o-iti\e,

become negative in -olutioii- coiitainiiu1 various narcotics, at id-.
alkalie- or neutral -alt-, and In- claims that the sense of the re-
action in the-e form- -eem- to be due to the general State of the

'Loeb, J., "Compa ' :i aini t'lUHpaial: . :.'il.,ny."

1910,

1 ,„•!,. J. • Heliotropi. I<« I • ins,

l.v Chemicals, especiall; • uy »i t'.iliiornia I'ut.li. ati-n- . "Physi-

l'. i. i'.
i. II 11. 1 . 1 h> < ontrol ol I'i R tions in ll\<il,-llu l>y

'• •'- 20 ^

' •

2~O J. E. WODSEDAL1 K.

organism a- a \\lmle, rather ihan to the specific effect of acid or
other solution- on tin- chemical state of some postulated Mib-
stance.

My experiments on the effects of chemicals on photoiaxis in
//. inter punctata nymphs were performed in a dark room and the
source of light was an incandescent bulb of seventy-five candle
power. In each experiment luenty fresh specimen- decidedly
negative in ilieir reaction to light were put in a long and narn>\\
glass basin, three and a half bv t \u-nty-six inches, of about 2,000
c.c. capacity half filled \yith distilled water. The concentration
of the solution was then slowly and gradually increased by adding
constant small amounts of the given chemical at about tin or
twelve. minute intervals. A careful record was kept of tin- num-
ber of insects positive and the number negative at the different
concentrations throughout the experiment, h was noticed that
as the concentration of the solution was increased there was a
gradual increase in the number that became positive. For ex-
ample, in the experiment with hydrochloric acid at .01 per cent.
solution, of the twenty specimens four became positive; at .02
per cent, eight were positive; at .03 per cent, fifteen were positive:
and at .04 nineteen were positive. It was the same with the
other chemicals employed. As a rule, specimens that once be-
came positive remained positive throughout the experiment, and
usually the insects that seemed to be the least annoyed by t In-
light in fresh water were the first to manifest their positive photo-
taxis in the solution. In general the specimens that evim ed
their dislike for the light most obviously in fresh water were the
ones that made the most frantic efforts to get at the light when
they once became positive. Ho\\e\er. the specimens as a rule
were very persistent in their normal reaction, remaining negative
until death occurred.

Of the different classes of chemicals emplo\cd the acids were
the most effective in reversing the photoiaxis of //. inter punctata
nymphs. A large majority, and in >onn- cases all ol tin- twenty
specimens became positive in tin- acid solutions. \Yhen ( '()o \\.i--
bubbled through the water, again, a large percentage of tin- speci-
mens became positive. The concentrations sufficient for this
effect with the different acids u-ed are, -hydrochloric .04 per

PHOTOTACTIC RELATIONS IX THE MAY-FLY. -~I

cent., tartaric .04 per cent., nitric .03 per cent., sulphuric .03
per cent., boracic .05 per cent., and acetic .07 per cent. Similar
n -ults were obtained with the salts, although the number of
-prrimrn> which became positive- \va- xuiu-uhat Miialler. Ho\\-
ever, all tin- >alt- employed H'frrtrd a re-version, and the con-
centrations necessary to do thi- are as follow-: pota--ium iodide
.3 PIT cent., pota— iiim chloride- .3 prr ci-nt., ammonium bromide
.3 prr cent., -odium chloride .3 per cent.. >odium Milphite .06
PIT cent., and -odium -ul|>hai>- .08 prr cent. 'I'lu- alkalic- were
less effective, only about hall" of tin- -prrimrn- becoming po-iiivc
in tln-r -oluiinn-. In general all of tin- -prrimrn- U-ramr quite
inarti\r, and uhni -pi-i iiiu-n- \\hich faili-d to manifest a po-iii\«-
n->|ion-i- urn- plan-d in ihr^riid of tin- ba>in nraiv-t ID thr li^lil,
tin \ -rldom mailr an\ rl'fort to -ct au Ml tlirrr of thr

alkali.-, -odinin h\<lro\idr. |)ota--ium h\dro\idr, and ammonium
li\dio\idr, uhi(h were rmplo\r<l. produii-d -imilar rr-ull- at

about the same concentration, which was ,09 per cent. Alcohol.

too. at a ( OIK ( iniaiioii of about .8 prr cent, produced a iv\rr-al
ol thr ivarl!..|) in a lar^r perCCnl 1 hr -prcilllrll-,.

Ill lllrii Mallir.ll rll\ innimrllt tllr-r inseCtS li\r on lllr Ulldrr
side ol Stones and r\ni in thr a.|iiaria I lia\r m\ i tin-in on

llir llppri- surl \\ in n a i n tain amount of an\ of thr above

mmiioiird rhrmir.iU ua- addrd to thr \\aliT ill thr ai|liaria.
thr n\ n 1 1 ill- \\ould -o. .n ( T.IU 1 up on to|i. 1 e acids, a^ain. \\rrr
pan i< ulail\ rllrcii\r iii tin- change, and although ordinarx da\ -
liljll Ua- -lllticirilt lo brill;^ lllr ll\lll|>h- UJ). lllr I>-lllt-> brraillr
nioi, ob\ ion- uhrn llir aquaria u rrr taken in the dark room and
a -lion- liijit ua- -u-prlidtd OVCf thrill.

It ;^i\r- nir ]>lra-urr here \« ackllouln|-r m\ indebt I'd lle-^ to
Trof. s. |. llolmr-. for hi- hrl|> .ind kindly critici-m-.

I vi • -u \ l»kV.
I M\ i k-i iv ..i \\ i-. . INSIN.

A PRELIMINARY NOTE OX THE RELATION OF NOR-
MAL LIVING CELLS TO THE EXISTING THEORIES OF
THE HISTOGENESIS OF CONNECTIVE TISSUE.

JEREMIAH S. FERGUSON, M.S., M.D.,

ASSISTANT PROFESSOR OF HISTOLOGY, CORNELL UNIVERSITY
MEDICAL COLLEGE, NEW YORK CITY.

The connective issue of the adult is in the embryo derived
from the mesenchyme. The same is true of certain other
tissues, endothelium, muscle, and cartilage, bone and dentine,
if these last be not included within the scope of the term con-
nective tissue. It is in this narrower sense that the term con-
nective tissue is herein used.

In considering the origin of connective tissue one has to
take into account two distinct elements, cells and fibers. Of
the cells there are at various stages in the embryonic mesenchyme
three distinct types: (a) the wandering cells, viz., leucocytes,
which are more directly concerned with the processes of hse-
matopoiesis; (b) those cells which give rise to the true wandering
connective tissue cells of the mature tissues, the mast cells,
plasma cells, etc., \vhose history is more or less closely concerned
with that of the embryonic leucocytes; (c) those typical con-
nective tissue cells which are related to the fibers and which
are commonly known as the "fixed" connective tissue cells.
It is without doubt only the last type which is concerned with
the origin of the connective tissue fibers, hence the present note
deals only with this type of cell.

At least two types of fibers have also to be considered as
arising in the connective tissue mesenchyme, the elastic and the
collaginous. We are here concerned only with the latter type,
for the reason that the elastic fibers arise at a later period; thus
the origin of the connective tissue libcr> concerns primarily
the collaginous, or so-called "white fillers." It is unnecessary
to distinguish at the early stage under consider.it ion between

such varieties of collaginous fibers as "reticulum" and " librog-

272

HISTOGENESIS OF CONNECTIVE TISSUE. -75

Ha," for the former undoubtedly arises in exactly the same
manner as the ordinary "white fibers," and of the histogenesis
of the latter little or nothing is known if, indeed, it can be con-
sidered as an entity distinct from the other connective tissue

tiliers.

The numerous theories which have been proposed to account
for the origin of the connective tissue fibers may be reduced to
three chief divisions:

i. The embryonic connective ti--ue or mesenchymal cells
!.(•((. me directly Iran-formed by elongation into connective
ti.—ue lib. •

J. Tin- tibi r- arise in the .unmix! -ub-tance betueen the
cell- eitlit-r by i rail-formation of that Around -ub-t.mce or a-
.1 -' < n lion from ;ln- .idiai < in i <-\\>.

v Tin- liber- ari-e \\ithin the cell- either a- di-tinct fib
- 'lu.mn. i- -mule- \\hich fn-e to form liber- iS|)iiler.

l6j I.axini, [909 . as an epicellular protopla-mic libroii- layr

I \ iff, [889 , or as an ectoplasm about the cell • I Ian-en, i -

or forming the -\ iicytiuin Mall, i

The ihe<,ry of dire, t trail-format inn by elongation of cell-
into hber- max \\ell be abandoned ami, except il be con-trued
aliuiLi ihe line- of the \aiioii- theori--- o| int racellular origin,
it i- \\nrth\- only of pa--iir^ notice. . \nion- other tiling- the

existence oi an overwhelming num!»er of fiber- r<-lati\e to the

Illllllbl I of i ell- ple-ellt ill Cop llei 'I i \ e ti--lle at'Vlle- ajaifl-t

dire, i transformation, and mod.-ni mii-ro-coi»ical meihoil- arc
not able to detect tin- pi uch t rail-format imi either in

normal growing ti— lie- or in the p ..I uomul re|».iii. I'or

Several decade- the re-lilt- of ob.-er\ at ion- ha\e been ill -llpport
of eiiher the iniei < ellnlar or t In- int ra< vllnlar -ronp of theorie-.

To Hellle I^}I we OWe ihe theor\- of the intercellular

lacellnlai origin of coimei li\e ti— lie fiber-, the liber- bein-

-uplio-ei| to ari-e Irom the intercellular ground -ub-taiice. not

directly from the cell-. Latterly thi- the..ry ha- received but

little support mile-- it be from the <>b-ervation- ,,\ Merkel

[895 . uho found that in the umbilical cord liber- appear lir-t

to ari-e in location- relatively remote from the cell-. Later

the cell- \\andered into the-e tibroti- area-, and at a -till later

274 JERKM1A1I S. FKRCU -MX.

period the fibers had so increased in numbers that they neces-
-arily laid in relation to the cells.

The weak point in Merkel's deduction appears to be the dis-
regard of the locomotive properties of mesenchymal or youn^
connective ti-Mie cells. If these cells, except for the incidents
of mitosis, are stationary, then Merkel's observations would
render conclusive evidence to support the theory of the extra-
cellular origin of libers. But that such cells possess at least a
limited power of motion simulating the anm-boid character
has long been known. An adequate theory must account for
the activity of these cells. Merkel's theory presupposes them
to be stationary till fibers have appeared and only later to
acquire amoeboid characters.

In -e.irching for a tissue which would offer opportunity for
the study of the activities of connective tissue cells in the living

SW1PM. Mf, S.50I 5.51 5.5Z

£68

6.00 „

6-/0

6.07^7 6.08 6.09 ^4

oJ L , VS^-A ^v/

FIG. I. < Hitlincs of a stc-llate conned iv ti^^uc ci-11 in tlu- caudal fin of a I-'uiiiln-
/HV rinl)i >•(> 4.5 mm. long (one clay alter hatching). The figures record the moment
of completion of each drawing, the whole ol>~er \-ati on extending over a period of j.j
minutes. The fish was afterward returned to water and swam for two days I"
li'-iny killed I'll tiirther histoli .yical use. The cell observed corresponds in appear-
ance with that -hown liy \\'. 1- leiiimin.- i \r,h. I. Aunt., 1897, Fig. 9, Plate \"I.)
in the me-entery of a Snlnni:itiil?r lai \ a. It slmws active amoeboid motion and
ash e change- oi iiiim. It- locomotion was not recorded, it was (|itite limited.
X 800, camera lucida.

animal under normal conditions, 1 have found that the median
tin o| voinr; lish embr\-o-> allonh a subject tor stud\' \\ hich throws

on Merkel's observations.

HISTOGENESIS OF CONNECTIVE TISSUE. -, ;

In a Fund id us embryo of 6 mm. total k-n^th — se\ -oral day- after
hatching- — I find in the median fin between the double layer
of cutaneou- epithelium tin- tir-t -;-n- . .f tin- dermal fin-ray-.
R!ood-ve— el- have not entered, and between the tin-ra\- are
fr\v if any connective ti — ue fibers, certainly HOIK- in the distal
portion of tlu- fin. nvadiirj the ba-e of the fin is a ma-- of m« --
• in hymal cell-, mostly of the numd cell type. s< al i«T'-d through
the fin are, here and there. i-«»lated c< mnect i\ e ti.— ue cell- in
very limited number-, di-tribtited over an area repre-i nt iii^
the proximal one half or two third- of the tin. and forming a -on
of -kinni-h line, as it urn-, in advance of the army of round cell-.
I Stellate cells e.m often be di-linctly seen in the li\ in-

animal, l.aier. coniiecii\e ti--ne cell- in abundance \\ander into
the tm and fiber- appear. Rut it cannot be -aid that the tilur-
appear in ad\ame of ihc cell-, for the "-kirmi-h line" pivc.de-

the appearance ot tiber-. t! dvance • eiu becoming later

intermingled \\ith tho-e of the -ub-e(|iH'nt in\a-ion.

The ad\allie cell-, like all other -tell. lie connective ti — lie

cell- in the tin- o| einbr\o ti-ln -, I tind to be actively anxeboid,

capa!'!' "iil\ o| motion bn: me < \ient of loioinotion.

>iii(f ihe-e till-, are !ia\ellinu :hrou-h an area in which liber-

oiil\ ju-t beginning to appear, the location of early fibers al

point - idati\el\ di-tant from the c»-ll in "fixed" li — ue al once

lo-e- ii ificance and thus robs the theorj of the extracellular

origin of fibei -iipl»on- M \ ob-er\ at imi-

ma\ be readilx \etitied on an\ li\iirc, free-swimming l-'innhilns

embryo within the fir-i week after hatching

1 inni tive li--ue liber- ma\ be -ti|)po-ed to ari-e a- a |)rodin i
of se< retioti of the me-ench\ ma! i ell-, the extracellular deposition
of fiber- tliu- occnrrini; under the intliience of the cell-. Thi-
i- bin a -liijit deviation from the -trict ext r.n ellular origin |>re-
supposed by the theon ot Ibnle and Kolliker. Such -i-cntimi
mn-t appear either in fluid form or a- granule-. In either case
it i- difficult to conceive why the -ecretion -hoiild take on a
linear form it the cell- remain Stationary, and -till more difficult
if the cell- are in motion, mile-- that motion be extrcmelv -lo\\
and in one direction only.

Rv ob-er\ aiion of the tins of c-mbr\'o fi-h. chieth' h"undulus,
I ha\e been able to ol that the -pindle cell- ot embrvom'c

2/6 JERI.MIAH S. FERGUSON.

connective tissue possess locomotion and move largely in one
direction, but their motion and locomotion is extremely active,
and so far as I can observe in the living animal they leave behind
no trail of secretion, certainly no observable granules: still more
important, the fibers appear to have been formed in great numbers
prior to the appearance of the type of spindle cells in any con-
siderable proportion relative to the other types of cells, round
and stellate, already present in the primitive connective tissue.
Hence I cannot conceive of the spindle cells as producing fibers
by direct secretion.

But it is at the time of the predominance of stellate cells that
the fibers make their first appearance in the fish's fin, as is
likewise the case in the tissues studied by other observers.
These stellate cells rapidly change their form, throwing out and
retracting their processes in rapid succession, as can be seen in
any living, free-swimming /'"nmlnlKs embryo under 25 mm. total
length. Fig. I shows the outline of such an early connective
tissue cell in the caudal fin, its changes of form at intervals for a
period of twenty-four minutes being accurately depicted with the
aid of the camera lucida. It is impossible to both watch and
draw all the changes in form occurring during this period, but
Mifficient are shown to demonstrate very active amceboid motion.
By observing these cells in relation to a relatively fixed object,
e. g., a chromatophore, or a joint in an adjacent fin-ray, I have
been able to detect in them considerable locomotion. 1 find
that, unlike the spindle cells, the stellate cells appear to travel
equally well in all directions. Neither following their locomotion
nor in trail of a retracted process do I find any indication of
secreted granules, nor of anything other than the most delicate,
preformed fibers. One would suspect that it new fibers were
being formed by a process of secretion they would appear within
i \\enty-foiir minutes after such a cell or its process had occupied
a gi\en position in which liber should reasonably be expected
to be formed. Though I ha\ e repeated the experiment many
times I have failed to ol»MT\e such evidence of secretion, and
the-e re-nlt-, do not appear to be con^i-tent with a theory of
dire> i -eiTetion of libers by the primitive connective tissue cells.

The intr.ii t llnlar theories of the origin of connecti\e li^-ue
fibers may be divided into I wo classes, those in which the libers

HISTOGENESIS OF CONNECTIVE TISSl I .

are presumed to arise in the peripheral portion of the cell, as
proposed by Lwoff (1889), or in a modified peripheral portion of
the cellular or syncytial protoplasm, ectoplasm or exoplasm.
as proposed by Hansen (1899) and afterward- by Mall [902 .
and those in which the fibers are presumed to have a direct
intracellular origin as observed by Spuler (1896) and later by
Livini (1^09).

Ill' "circurncellular" origin of fibers proposed by Lwoff,
since it mu-t take into account the -eparation of fiber- from the
cell- as udl as their origin from int racellular granules, doc- not
fundamentally differ from tin- later . riopl.t-mic modilicat i« »n-

c

I-i... .-. \ ..f tin- l.» 'iiiii.tii'n i -I a -pim!!.- i-niim-riivi- t: !. a. for

a 1" riodof 10 iiiiiiut. !:r.iiiKiti-iih. il -rin.v; ..n the- fin-rays <>n ciih.-r

'•i-iiii; taken a- the- n-lativ.-ly lixcd p ,int-;. The cell \va- t'<uml to have covered

•ut ^".u. : i/j in every I-' -x.ni,!-. u ,/, o«iiii.-i-tiv.- i.

C'-H- • .1 i tun : cell, tin- Otl I; Ch', Ch", tw ilir..matophores;

Old, fll.l.itlu-lilllli ill til.x..

of Ilaii-ni and Mall. Thc-c- Liter author-, ba-inv their inves-
tigation on "fixed" material, pre-uppo-e a constant but gradual
recession of the endoplusinic area with coincident changes in

2/8 JEREMIAH S. FERGUSON.

the exoplasm in order to account for the transference of fibers
from a position within the cell, in the- ectoplasm, to an extra-
cellular position. This hypothec- i- entirely harmonious with
i lu- appearances to be observed in "fixed" tissue with the methods
used. Ho\\e\vr, it is open to two objections, viz., the obser-
vations of Spuler, confirmed in a casual and not wholly satis-
factory way by Livini, that fibers actually lie within the cells,
and not always at the periphery, as Hansen's and Mall's theories
presuppose, but even in direct contact with the nucleus; and,
secondly, if the cells are in active amoeboid motion, with loco-
motion as well, as is certainly the case in the fins of the embryo
Fiunliilns, then one can hardly conceive that the more or less
fixed syncytium described by Mall, whose observations can be
readily verified by any of the usual histological methods applied
to the tissue used, is sufficiently stationary to permit the retrac-
tion or shrinkage of endoplasm, thus leaving the endophismic
fibrils outside the bounds of the cellular protoplasm, for according
to Mall's theory true cells are lost, being replaced by nucleated
endoplasmic areas anastomosing with their neighbors to form a
continuous syncytial net.

1 have many times observed the fins of Fiiuditlns embryos
between the lengths of 4.5 mm. (at the time of hatching) and
25 mm., the pectoral and caudal fins being usually selected, and
in no single instance have I observed either a stellate or a spindle
cell which did not exhibit changes of form and locomotion, the
latter being sometimes very limited, at other times, as shown
in Fig. 2, quite extensive. Hence the ultimate theory of con-
nective tissue histogenesis, it would seem, must take into account
the relative activity of the mesodennic cells. So far as I ap-
preciate them, none of the theories thus far advanced fully
meet all of the ol»erved conditions. Further studies of both
living and "fixed " tissue may further elucidate the true conditions
involved in the histogenic process.

The fins of living fish embryos offer for the development of
connective ti-^ue a most desirable subject for study, for in the
subcutaneous tissue betueen the dermal fin-rays one finds a
definite connei li\e ik-ue area, in which, with the exception of
the >ome\\ hat related chromatophores, there is no other structure

HISTOGENESIS OF CONNECTIVE TISSUE. -" -

to confu-e the picture. This area is bordered on either side by
the dermal fin-ray, beneath which an efferent blood-vt
descends, and alongside of which the return or afferent \.
ascend-- the tin; it i- covered only by the pavement cell- <.)f the
dermal epithelium, who-e outline- are -harply detined, lyiii£ at
a more -uperticial focal plain- than the connecti\e ti — ue. The
chromaiophore- are ea-ilv reco-ui/ed both by their color, black
to yellow, and by their peculiar branching form. -<> that e\ en
iho-e re|,iti\ ely or almost entirely de\oid of color can be ea-ilv
ilitfereniiated from ihe -mailer colorle— connective ti--ue cell-.
Moivo\er, th,- chroma tophores are prone to lie in immediate
relation to the tin-ray- and to the blood-\ v— el-.

In com lu-ion I de-ire to einpha-i/e the opinion that in the

-tlldv of hi-to-i-lie-i- of conilecti\e li — lie \\ e Illll-t take into

con-ideratioii the (..ndition- -urroimdiiiK the living cell- and
dra\\ i OIK In-ion- troin "ti\e«l" ti--ne only in the Irjit of our
knou 1. t the li\ i:

I turni-li (oin In-' 'ill- li\iiu cell- mu.-t be ob-er\ed

under noriii.il condition- and not merely under tho-e mo-t un-
u-iial. or e\en pathological, conditions \\hich -urroimd 'lie ob-
servation <-ither in the me-enterv -tudieil by

Meinmiiu -mil other-, open l» the ciitiii-in of the pi'e-i-nce of
inllainmatoi \ change-, or in tin ti--ne culture- b\ the more
nt method (i II ii-oii. ( arrel and liurrou-. \\hich i- -ur-
roimdetl b\ the i the inlerpretalioii of te-nlt- ari-in^

umlei einireh ne\\ -i i rroii ml i i i'j - and condition- as \et but little
under-tood. Both in the nie-entei\ and in li — tie culture-
movements of the conne<ti\e ti — lie cell- and even locomotion
have been observed. I no\\ add to the-e former re-ull- the record
of i he ob-ei \ a I ion of motion and locomotion in cell- under wholly
normal . ondition-. in animal- \\hich umleruent no operation
other than in -onie in-tance- tin- admini-t rat ion of chloretone,
and \\hich remaiiu-d ali\e. active and normal for -oinc time -ul>-
M'ljuent to the period of observation. 1 or example, the par-
ticular ti-h \\hich furni-hed the >ubject for I-'I'L;. I \\a- retnnu-d
to -ea \\aierand remained acti\el\ -\\immiiiL; tor i \\ o da\-; he
could ha\ e been kept much I- inger.

l-'or the ojiportunit\ of pur-uinv; thi- -tud\ 1 am indebted
to the Marine Hiolo-ical Laboratory at Wood- Hole. M ISS.

THE METHOD OF CELL DIVISION IN MONIEZIA.

C. M. CHILD.

Since the appearance of Richards's paper on the method of
cell division in Miuiiezia (Richards, 'n) Dr. Richards, in answer
to a request of mine, has very kindly permitted me to examine
some of his slides of early stages in the development of the ovary
and of cleavage stages, with the privilege of publishing the re-
sults. The present paper consists of a brief statement of the
results of my examination of these slides, together with some
consideration of recent criticisms of my observations on amitosis.
I take this opportunity of acknowledging my indebtedness to
Dr. Richards and also of acknowledging an error of my own
which though it does not directly concern the chief point at
issue certainly requires correction. Dr. Richards is undoubtedly
correct in his statement that the early cleavages in Moniezia
are mitotic, either entirely, or to a much greater degree than my
earlier statements would indicate. With this acknowledgment
of my error the chief point of disagreement between Richards's
positive observations, so far as they go, and my own is removed.
The difference between us seems to me to rest no\v on Richards's
failure to recognize, or to interpret as I have done, what the
material actually shows.

The whole problem of amitosis seems to me somewhat un-
profitable at present as a subject for investigation on a purely ob-
servational basis. Experimentation is necessary be- fort- we can
reach any certain conclusions as to the significance of either mitosis
or amitosis in the life of the cell. Moreover, cytological thought
is at present dominated by morphological hypotheses, which are
themselves based on direct observation rather than on experi-
ment. So long as this remains the case all observations which do
not accord with the current hypotheses will be explained a\\,i\
by one course of 'reasoning or another and brought into agreement
with ihesc hypotheses. So long then as our conclusions are
b;i~<-(| Dimply upon observation in fixed material and without any

280

METHOD OF CELL DIVISION IN MONIEZIA. 28 1

attempt to control conditions in that material there will always
be room for endless discussion and controversy. It seems to me
highly improbable that cytology will make any very great real
advances in thi- direction. But when cytolo^i-t- -hall -eriously
dt vote their energies to the problem of the physiology of the cell,
a field of inve-ti.LMtion that ha- been entered only here and there,
and to the control of cellular phenomena, then I believe we shall
n-al pi . together with the <:on-i-nment to oblivion of

-oine ot the hypothe-e- now nio-t fondly cheri-hed.

A- regards Mnniczin, however. I have at p re-em nothing to
:it further observations, or rather my own interpre-
tation ot \\hat 1 h.: 'i in Richard-'- material a- a^ain-t his
interpret. iiion of uhal he ha- -ecu. li. -\.nid the prr-ent paper
I do not propo-r to enter into .my further con-ideration of the
problem of ainiio-i- until I .an d<> so "ii a different ba-i-. as

[ hope to be able to do ue time in the future.

Tim- far I ha\e .,nly attempted to defend mv own observa-
tions and i "in lii-ion-, .i-.iin-t uh -:ied to me unmerited
ci-iii<i-m and -.kepiici-m ba-ed on in-ullicient <la!a. Richard-'-
paper i.n Moni( :i<i i-, hov. d n-in\ e-t i^at ion of the Mib-
and as MI. h I e.m "til\ \\elcoine it and accord it the coii-
-ider.itioii \\ Inch i- due to .my piece of -, i.-n title \\ork. alt hou^h
I am eom|»elled to Criticize it in \arioti- point-. The pre-ent
pa|»ei include- a -tatemeiit of the re-ult- of my observations on
Richard-'- -lide her with some di-cu— i<>n of hi- argument-
and coiiclu-ioii- an 1 !•• to critici-m- olfen-d b\- other
authors and to tin- \ alidit\ of rytolo-ieal theor\ .

The .-lide- contain exactlv the -aim- picture- a- my own.
Tin re i- no i|iie-tion In - of fixation or of -t. lining.

It amito-i- OCCUrs in m\ malerial then it OCCUrs in Richard-'s
in the -ame region- of the body and the -am. - "t develop-

ment.

I. Tin I'KiMi'kiuv OF mi-: OVARIES AND DICTS.

I igs. i x are > imei : drawing- from the primordia of the

oxarie- in Richard-'- -lide-. The\- were drawn from -everal

slide- \\ith different method- of fixation and -taining. In the

ires 1 have attempted to reproduce what I have seen as

C. M. CHILD.

exactly as possible, though this is a somewhat difficult matter
as regards the (.'.\tru nuclear phenomena. Godlewski ('09) has
recently objected to my earlier figures on the ground that they
were too highly schematized. This objection seems to me to be
based largely on unfamiliarity with the material. In the early
stages of the gonad visible cytoplasmic boundaries are often
entirely lacking and the question as to whether cytoplasmic
division follows nuclear division has no point, since it is quite
impossible in most cases to distinguish cells. In later stages
the germ cells do become distinct, but in those stages amitosis
does not occur or occurs but rarely. In my earlier work I
attempted to reproduce the essential features of the nuclei as
exactly as possible. The only schematization consisted in not
attempting to indicate an intranuclear achromatic structure
except where such a structure was clearly visible and in leaving
out various extranuclear details which did not bear upon the
problem under consideration. In many of the nuclei the kary-
osome alone is stained and the rest of the nucleus appears almost
or quite homogeneous; in other cases a more or less distinct
structure of strands or an apparent reticulum does appear. In
cases where I could not see such a structure distinctly I did not
attempt to represent anything in the nucleus except the karyo-
some. It seems at least probable that the apparent achromatic
nuclear structure is in many cases simply an artifact due to the
action of the fixative. In the figures of the present paper I have
represented a vague structure in all nuclei, but for most of the
nuclei this must be regarded as little more than an attempt to
show that something is there.

Richards's statement (p. 135) that the parenchyma is not a
syncytium seems to me to require qualification. It may well
be that it is not a continuous s\ncytium throughout the whole
proglottid in later stages, but there cannot be the slightest
doubt that several or many nuclei are often found imbedded in a
more or less continuous mass of cytoplasm. This is true of the
early stages of the gonads as well as of other parts. Where the
nuclei are far apart they are usually, as Richards states, sur-
rounded by more or less definite and distinct cytoplasmic masses,
but this is certainly not true- in caM-> \\ here they lie close together.

MKTHOD OF CELL DIVISION IN MONIEZIA.

Fig. I shows two nuclei in close contact, with the surfaces of
contact flattened. If each nucleus surrounds itself with cyto-
plasm it is difficult to understand how a condition like Fi£. I
can arise except l>y division. Moreover, this is not the usual
result of mitotic divi-ion in Moniezia, nuclei which ari-e by
mito-i- arc -eparatcd by an appreciable distance when they form
and are tonally of equal -i/e, which the two nuclei in the figure
are not.

I . J sho which I am unable to interpret except as a

din-ci division. The nio-t careful focu-in^ -ho\\ - that the nuclei

3

£fej

5

. .

7

8

are in ihe >ame plane and that the iin-inbr.ine -eparatin^ them
i- ii"i a -!iand of "linin" but a n-al membrane. Moreover, thi->
membrane i- appareiilly thinner and more delicate than other
pi'rtit'ii- of the nuclear membrane.

In 1 g ; two nuclei .ire -ho\\ n which -eem to be just com-
pleting di\i-ion. much a- in l-'i^. I.

1 i^. 4 shou-, t\\.. \et\ di>tinctl\' hemispherical nuclei with
llatteiied surfaces parallel and -t-parated by a Miiall amount ••!'

284 C. M. CHILD.

cytoplasm which stains less deeply than other parts. They
appear as if division had recently occurred and cytoplasm had
begun to form on the adjoining surfaces. If it were not for
certain preconceived ideas concerning the method of cell division,
I think no one would hesitate to regard such a case as a very
strong indication — I do not say proof — of direct division.

In Fig. 5 the apparent division of a small nucleus is shown.
The membrane between the two parts is more delicate than the
rest. The position of the two "nucleoli" is important. They
are close to the separating membrane and directly opposite each
other. This position is of very frequent occurrence in such
cases, far too frequent I believe, to be the result of chance. It
suggests very strongly that division of the nucleoli may occur
in many cases.

Fig. 6 is a case of apparent "endogenous" division. Richards
(p. 14) was unable to find anything of this kind in the material.
I found a number of such cases in his slides.

Figs. 7 and 8 show cases in which the two halves of a nucleus
are slightly different in color, a phenomenon of not infrequent
occurrence, which Richards also failed to observe. I do not
know, of course, that such a condition indicates division, but
I regard it as highly suggestive in that it certainly indicates some
difference in condition in the two parts of the nuclei and a certain
degree of independence of each other.

Figures of this sort might be multiplied indefinitely from
Richards's slides, but there is no reason for giving others here.
My chief object is to show that exactly the same pictures appear
in his material as in my own. I am perfectly well au.uc t hat-
none of these figures and likewise none of my earlier figures
constitute a real demonstration of the occurrence of amitosis,
for such a demonstration is impossible in fixed material.

Numerous similar pictures were found in the early >tav,i-s of
the genital ducts, the vitellaria and the parenchyma during the
stages when increase in the number of nuclei is occurring in the>e
regions. There is, however, no reason for giving figures from
these organs, for on the basis of current cytological theory tin-
point of chief importance is whether amitosis occurs in the
germ cells. If its occurrence is admitted for tln-M- cells, most

METHOD OF CELL DIVISION IX MONIEZIA. 2S;

cytologists would, I think, be willing to admit at once that
similar phenomena in somatic cells doubtless indicate its occur-
rence there.

\\ 'r may now consider the value of such pictures as evidence
and tin; force of some of the criticisms made- by Richards and
others.

As regards the first point, I have observed picture- of this

kind only in region- where the nuclei are increa-in^r in number.

'Ie\\-ki '09 h.i- -u^e-ted that I may have mi-taken nuclear

lu-ioii- tor divi-ion-. It would be -trance indeed it" nuclear

lu.-ion ^hould occur only or chiefly in regions where the nuclei

undoubtedly iticr. ;n number in some way.

•ii.it the tuo nuclei in a cell repre-eiu

the paternal and maternal chroinatin refer- primarily to the
aiii]ihiltian cell- and perh. ip- he him-clf would n«>t rc-ard it as
ad" '11111 t'<ir t! nces in .17 ' Mainly

then- i- no evidence of t: ratii>n of the two chroinatin- in

milotic di\i-ion in M< en in . A- iv-ard- the

amphibian i "-1I- \\hich I ti^un-d in an earlier paper '.Child,
|o and l l ii -lu>uld I- •••<! that t ': !i.-<l

\\ere n-.i cleava . but mo-ll\- lar\al , in \\hich I

(i-uld IIIK! ii' ': I took especial < iicrriiiiu'. this

point, '•p.iratinii of paternal and maternal cliromatin in

mitoiic di\i-i«ni. I d» imt believe that thi- -iiu^e-tii)ii \\ill
"lint for the phenomena even in .1 and it certainly

\\ ill ii">t in .I/.-

A- i< ^ard- the po— ible in ot the nuclei by migration in

parti"iilai regions, «. ,., the prinmrdia of the •o'liad-. '\ichanls
him-elf admit- 'hat the i-\ idence tor migration U not

strong, M' even if migration di cur in the formation

of the -..nad-, there niu-t ne\ ei t lule— be a coii-ider.tble ])riniar\-
increa-e in the parcnchymal mu l.-i befori' tin- ouiad- are formed.
Kichanl- pp. [38 9 find- the aciual increase in parenchym.il
nuclei to be .'.bout equal to the increase iii \olume of the pro-lot-
lid-. Such an increa-e i-. I belie\e, entireK' inade(|iiat"- to ac-
count on the b.i-i- of migration for tin- very lar^e number of
nuclei which appear in ^miad formation.

t 'oncerniii;c the occurrence of mito-i- Richard-'- actual ob-

286 C. M. CHILD.

serrations are essentially identical with my own; he finds it but
rarely, in fact he has not figured anywhere a case showing so
many mitoses as my Fig. 8 (Child, '070), in which seven cases
of mitosis are shown. His argument concerning the ratio of
resting to dividing cells in sections (pp. 139-40) concerns matters
of which we really know almost nothing one way or the other;
moreover, I am fully convinced that in many cases besides
Moniezia where the infrequently observed mitoses have been
supposed to account for all nuclear increase, numerous amitotic
divisions actually occur. But besides all this, my conclusions
concerning Moniezia are not based primarily on the frequency
or infrequency of mitosis, but upon the actual observation of
what I cannot regard as anything but amitosis. The infre-
quency or apparent absence of mitosis in many regions of Mo-
niezia was what first led me to undertake a further investigation
of the material and the remarkable infrequency of mitosis ac-
cording to all observers in the neck region, the developing gonads
and the genital ducts still seems to me to be highly suggestive,
especially when we add to this fact the very frequent occurrence
of what seems by all criteria at present available to be direct
division.

Concerning periodicity in mitosis (Richards, 'n, p. 140)
the only positive observation which suggests anything of the sort
is mine, which was cited in an earlier paper (Child, '10). This
was a case of scolex, neck region and a few proglottids in which
most of the nuclei were dividing mitotically. I have no means
of knowing whether this is a normal occurrence, but if it is, it
seems probable that this specimen is a young scolex, for the nuclei
throughout the scolex are dividing as well as the others and these
nuclei certainly do not take part in the formation of proglottids
in later stages. I have considerable evidence for various forms
which seems to me to indicate that many organisms or parts
may start a period of nuclear multiplication by mitosis, while
in later stages amitosis becomes increasingly frequent and it
this scolex is normal it may be a case of that kind. It is quite
possible, however, that it may be- the result of certain special
conditions of which we know nothing and not a normal stage of
development in Moniezia at all.

METHOD OF CELL DIVISION IX MONIK7I A. -V

It is certainly most strange, if mitosis occurs periodically
or in waves as Richards suggests, that neither Richards nor
Young nor I — except in tin- case above mentioned — have hap-
pened to ob-er\e any of these waves of mitosis in the numerous
animal- t hat we ha\ e -ectioned. All the evidence which we have
obtained indicates a remarkable infrequency and a scattered
occurrence of mito-i-.

lint tin- argument ba-ed on mit<>-is is not direct evidence
lor or auain-t the occurrence of amit<>-i-. \\'heii \\ e tind the
picture- \\hich I have figured occurring frequently and confined
to re- ion- in which the nuclei are knoun pn-iii\el\ to be incr«

in number in -ome \\.iv. the direct e\ ideiice for the occur-
r< -in e "I amito-i- seems to me to ha\e far greater value than any
indin-c t e\ idence l.a-cd on the treojueiuA or infre(|iienc\- of

mitosi

•Mi pagi i . lianl- that in th<- primordium of

theo\ar\ and duct- the re-jon .if unclear increase is at thepe-

ripherv and that it i- here that mito-i- i- chielly found.

Wit h the latter statement I jree, bul I imd it difticuh to mxler-

-land the v:i"'ind- I-T hi- ("iichi-ioii that unclear increase i-
• niilined to the pel i| iliei-\ . ( CttainU' tlie iiM'i'ea-e in the number
<il mil lei at llie peiipheiA i- n.it \i • it, \\hile in the central

ion "I the pi inioidinin the nuclei are \ ei \ niimeioii-, cm \\ded
i li.-e together. M"ie. .\ ei , these central nuclei are in earl\ -t.:
\et\ much -mailer than the peripheral. I nle-- there ha- be, n
e\ten-i\e mi^iaiiiiii from ihc periphery to\\ard the center it i>
impossible to accoimt for the i^tvai number of nuclei then except
b\ pM'lileiation. Ki< hard- ha- failed to lind amitn-i-, in thi-
region, but 1 ha\e found nnmeroi: — ( if \\hat I regard as

amito-i- in hi- -liile-. I l< maintain-, that the central nuclei
are be^innin^ ditleieiil iat ion (p. [50). I have been unable to
Observe an\ \i-ible dilleientiation in this region at the-e -t.:
and be-i<le- the-e ct.i\\«led nuclei are on the average much
-mailer than anv nuclei of the \\hole |)rimonlium after the vi-iMe
difli'fentiation ha- be-nn. In later st •.\lieu. as I believe.

|)iolil\'ration become- le-- rapid, the-e nuclei -ladiialh enlarge
a-ain and became more \\idely -eparati'd and then visible dif-
ferentiation begin-. For the-e reasons I am unable to agree with

288 C. M. CHILD.

Richards that the region of greatest proliferation is the periph-
ery of the primordium. Such a conclusion fails, it seems to
me, to take account < >f all the observed facts. The great number,
the very small sixe, the crowded condition of the nuclei, the
relatively small amount or cytoplasm and the very frequent
occurrence of apparent amitoses force me to maintain my original
position, that the central region of the primordium is a region
of more rapid proliferation than the peripheral. In connection
with this point I have evidence from various species that in
regions of more rapid proliferation amitosis is more frequent,
while in regions of slower proliferation mitosis may be the chief
or only method of division.

On p. 150 Richards states that "the method of cell multi-
plication in the female sex ducts of Moniezia cannot at the present
time be positively stated." Here evidently the author has lost
faith to some extent in his own argument, for the conditions in
the ducts are not essentially different, so far as observation
shows, from those in the primordia of the gonads themselves.

Of the possibility of observational error (Richards, 'n, pp.
140, 141, etc.) no one can be more keenly aware than myself.
I can only state again that I believe that I have used every pre-
caution possible in direct observation of fixed material. I have
often spent hours on a single case with the most careful focusing,
changes of illumination with the aid of diaphragm and mirror
and the alternating use of artificial light and sky light with and
without color screen. I'ntil Richards can show that he- has
taken at least equal precautions before concluding that direct
division is absent, he lays himself open to the charge of error
or superficial observation.

As regards the difficulty of observation in the small and
often crowded nuclei of cestodes, a point which Richards em-
phasized in his earlier paper (Richards, '09), it is of course true
that the nuclei are small and often crowded, but if one devotes
sufficient time and care to the matter it is possible to find nuclei
which show apparent direct division with almost diagrammatic
clearness and I have seen many such, both in his material ami
in my own. In such eases I have not been able to convince
myself by any means which I could devise that the appearance

METHOD OF CELL DIVISION IX MOXIEZIA. 289

of direct division was a deception or an artifact, or that it lay
within the limits of observational error, as Richards asserts
for all apparent cases of the sort observed by him.

It would l»c of interest to kno\v how far Richard- i- familiar
with the appearance of amito-i- in forms and ti--ue- \\here it is
•jencrallv admitted to occur. I have spent considerable time
familiarizing myself with such material and I find many cases of
apparently dividing nuclei in .Uo/nYcM which -eem tn me to be
id i -ntii -a I in .i|)|iearance with « l» - «\ di\ i-ion in such ti — -ue-.

And linalK . K'i.'hard- admit-- finding mniienm- " paired
nuclei" and nuclei l.etueeii \\hich no lavcr of cytoplasm could
he di-iin^iii-hed: it i- of intere-t incidentally to note that hi-
statement- OM thi- point concerning the duct- and vitell.iria
an- much less caution- than tho-c concerning the <>\arian pri-
mordium it-elf. And although lioih he and I have oli-cr\ed
inito-i-. in tin- duct-, he i- \\illin^ to .idmit concrrnini; them that
then- is "no certain evidence I'or amito-i-^ and th.it for miio-i- i-,
jierhap-. in-ultn icnt to :nt for the Drouth \\liich ha- taken

pi.. : And « on, ruling the \itellaria he -a\-. "if

mito-i- i- not <lratl\ pto\,-,| as the -uUiciriit CaUS€ o| ( ,-H niill-
ti|>li< ation, amitOSlS i- c.-itainU lc iceinin^ tlu-

o\ . ilium piinioidiuni hi- -talemeiit- arc much more guarded.
I I- ic the influence of < \ io|( ,-j. .il In polhe-c- appear-. Mo-i
author- are \\ illi' dmit the occurrence of a mito-i- in -oinai it-

cell-, though thc\ \\oiilt! 1 as rank heiv-v the a--ertioii

that it OCClirs in the i^ern; cell-. ( Vrt.iinly Mmi!i-~i,i allonl-
no lia-i- lor -nth a di-lim lion ami • [ trd- main" other forms
\\e have not the -lijjite-t e\ii|ence ihat the verm cell- in earl\
Stages an ditlcicnt ill an\ \\a\ from -omatic cells. The mo-t
that ol.-erxaiioii , ,m n-ll u- on tin- point i- that in -oine forms
tlu-\- appear earlv. in other- late.

\- to \\hat con-titute- e\itlence for amito-i- Kichanl- does
not -eem to !>»• eniireK clear. A- regard- the \itellaria lie says

"Inn for amito-i. there i- only iu-^ati\e evidence" i|>. 147 .

\et iii the -ame paragraph he admit- that "the arrangement
tit nuclei in [i.iii- i-. pcrhap-, more in e\ idence here and in<lented
nuclei are -omeuhat more numeroii-." These oli-er\ei| facts
reprc-cnt condition- \\hich we should expect to tmd pre-eiit it

29O C. M. CHILD.

amii<>-i- incurred and therefore constitute not negative but
positive e\ idence of considerable value. They do not, of course,
constitute a demonstration, but, as I have pointed out abo\e,
an actual demonstration or proof is impossible' in fixed material.
To my own mind, all the facts concerning which Richards and
I agree, together \\ith the cases which I regard as nuclear division,
but which Richards has failed to find, constitute a practical.
though not a logical demonstration of the occurrence of amitosis
in Moiiiczia. On p. 127 Richards asks "whether the failure to
find evidence of a certain process, using proper methods and
exercising due diligence, is not positive evidence of the lack
of that proces- .J ' Such a question can of course only be an-
swered in the affirmative, but this does not constitute a refutation
of my statement that Richards had presented only negative
evidence for the occurrence of amitosis; it is of course merely the
same statement in other words. And we are all aware that pos-
itive evidence of the absence of something must be of the strong-
est character before it can be regarded as refuting direct posit i\ e
evidence of the presence of the same thing. Richards is unable
to show with anything approaching certainty that mitosis is
the only method of cell division in the ovarian primorciium.
I have presented positive evidence that amitosis occurs in ad-
dition to mitosis, though my evidence does not amount to an
actual demonstration since that is impossible. 1 believe, how-
ever, that the evidence for the occurrence of amitosis in the
(•\.irian primordia of Moniezia is almost if not quite as conclusive
as that tor its occurrence anywhere else in other species.

II. Tin; CLEAVAGE STAGES.

As regards the cleavage of .\foniezia, my examination of Dr.
kichards's slide-, enble^ me, as mentioned above, to correct
the error into which I fell in maintaining that "cases of mitosis
.iii r.irely seen after the first cleavage, but amilosis is ot frequent
occurrence' '< 'hild. 'ojM. Richards's O|>MT\ ations as stated
in his paper led me to believe that I must ha\e been in error
on this point and the examination of his slides removed any
doubt which might ha\e remained.

There is no question but that tin- earlier cleavages beyond the

MKIHMD OF CELL DIVISION IN MONIEZIA. 29!

first were almost entirely absent from the proglottids on which
most of my study of cleavage was made and that my Figs. 21-26
(Child, '076) are in reality sections of considerably later cleavage
stages than I had supposed.

The passage of the eggs into the uterus in Mnniczia undoubtedly
occurs periodically and as the eggs are fertilized in the course <>f
ihis passage it follows that in any Dingle proglottid containing
( li-avage stages not all the stages will be present. In the- pro-
glottids on which I based my study of cleavage it happened (hat
hi--i cleavages \\<-n- numerous. /. c., a period of pa— -age of eggs
from the ovary to the uterus had occurred recently. Hut as a re-

ex.unination ot the-e -lide- after my study of Rit h.ir.U'- material,
shous, tin stages bctueen the tir-t and occasional second cleav-
nd much later Stages were almo-t entirely absent. M\
ol" supposed early ( leavages are in realitv sections « ,\ t
containing a < < m-ii leral >Ie numl'er of nncK-i, but the plane of
-'' lion happen- to pass SO .1- I" show only or chielly the L
nuclei. Thai I could mistake ihe-e f. .r early cleavages \\a- due
to the tact thai I \\.t- much more interested in the condition
ot i he inn lei than in i he course »i i leavage, and m\ observations
on ihi- latter poim \\tic .ml\- incidental and so led to a wrong
com lu-ii.n. Tim- m\ <\\»v on thi- point i- explained, but I do
not • illcr i he explanation as an excuse.

!• unhcrmoic. Richards i- ciiiiivl- ct in }\\~, a— -eriion that

liming iln- earlier cleavages tin- blastonieres possess di-tinct
boiimlai ii - and tl,> IS not a s\ ncytial ma--.

< >n the other hand m\ examination of Richard-'- material
has enabled me to <li-co\er \\hat -eetiis to me a \ cry important
point that he ha- tailed to note. In his material the early
cleavages are, as he says, mitotic and the bla-toinerc- distinct.
The ( \ to] .la sin in these M. ;m- rat her dee] il\ and uniforml\-

and i> not \acuolaled to an\ great extent. At certain Staj
ho\\e\er. \\hich ma\' \ar\ in ditlerent e-;^> to some extent, but
at \\hich tlh consists of a considerable number of blas-

tomeres it und. a marked change in appearance. The

cvtoplasin lose> its t ini;ibili t y to a lar^e extent and become-,
highly \ai-uol.ited and the di-tinct boundaries of the blastomeres
disappear. Apparently thoe boundaries disappear gradually,

2Q2 C. M. CHILD.

for in many cases some blastomeres ivm.iin more or less distinct
while others have completely disappeared and in Mill other cases
the whole mass of the egg appears to he a syncytium. In my
earlier studies of the cleavage 1 interpreted these stages as stages
in the appear, nice rather than the disappearance of the cell
membranes.

In these and later stages I find, both in Richards's material
and in my own, the most striking evidence for the occurrence
of amitosis that I have seen anywhere in Moniezia. Here the
nuclei are large and they are not crowded, consequently they
can be studied with less difficulty and a larger number of good
cases can be readily found.

I give here a few figures drawn from Richards's slides to sup-
plement my earlier figures.

In Fig. 9 five nuclei, one of them "double," are shown. No
cell boundaries are visible and the cytoplasm is highly vacuolated
and stains only slightly The two parts of the double nucleus
are in immediate contact and the relation of the nucleoli to the
dividing membrane and to each other which was mentioned
above is seen.

Fig. 10 shows four nuclei, one of which appears to be dividing
or to have recently divided. The two parts of this nucleus are
of different size and are closely apposed.

Fig. ii is an interesting case: the blastomere containing the
double nucleus still retains its dense un vacuolated cytoplasm
and to some extent its distinctness, though no actual cell mem-
brane can be seen. The two parts of the nucleus which it con-
tains are in immediate contact and flattened against each other.
It is difficult to understand how they could attain such a position
as the result of mitotic cleavage like that of the earlier stages.
Moreover, in those earlier stages neither Richards nor 1 ha\e
found any evidence of the separation of paternal and maternal
chro matin.

In Fig. 12 is shown a case of apparent division which seemed
to me, as I examined it, particularly convincing. Here the t\\o
parts of the nucleus are of different si/e and a ne\\ nuclear mem-
brane appears to be in process of formation U-t\\een them. Its
formation apparently began on one side and on that side the

METHOD OF CELL DIVISION IN MONIEZIA.

293

I

~~"

V

V

\-^— ^

II

10

16

294 C. M. CHILD.

nuclei have separated, \\hile on die other side it is still very
delicate and the nuclei an- Mill closely connected. The dense
mass in the upper p.irt <>t" the figure is the cytoplasm of a large
blastomere which has not yei .undergone the change in appear-
ance; its nucleus is in another section.

Fig. 13 shows a section without any cell boundaries and with
\aciiolated cytoplasm in which one small nucleus is apparently
dividing.

In Fig. 14 a constricted nucleus is shown. The cytoplasm
of a large dense blastomere is also seen in the figure.

Fig. 15 shows another case which is to me very convincing.
The double nucleus is in a mass of cytoplasm which is denser
than the rest and still rather distinctly marked off from it.
Evidently this represents a blastomere. Here, as in Fig. 12,
the division of the nucleus apparently began from one side, for
the membrane is much more distinct on the side where separation
is most advanced and becomes more and more delicate toward tin-
other side.

Fig. 1 6 represents a case of almost diagrammatic clearness in
which a large nucleus is apparently dividing from one side.
Other nuclei are in other sections of the egg.

Figures of this sort might be multiplied indefinitely from
Richanls's material as \\ell as from my own. There is, I think,
but little chance of observational error here, for the nuclei are
relatively large and not crowded and in many cases almost
diagrammatically distinct. If pictures of this sort are of any
\alue as a criterion of the occurrence of amitosis, then the
e\ idence is certainly very strong for these cleavage stages.

( )ne question, ho\u-\er. must be raised here, vi/., whether
these are normal stages in development or degenerating embryos.
To settle this point absolutely tin- study of later stagi-s i- neces-
sary and thus far 1 have not been fortunate enough to obtain
such si.iges. But in many proglottids practically every embryo
shows this syncytial condition and if this indicates degeneration,
then all, or almost all. the embryos of those proglottids are
degenerating. Ii seems probable therefore that \\e ha\e to do
here with normal stages in ihe development.

METHOD OF CELL DIVISION IN MONIEZIA. 295

III. CONCLUSION.

My examination of Richards's material has only confirmed
me in my conclusion that direct division plays an important
part in the developmental cycle of Moniczia. in the germ cell-
as well as in the soma. I believe I ha\t- at least shown clearly
enough that this material contains something besides negative
c\ idence fur amitosis.

We are, I think, ii"t yet acquainted with the nucleus as a
dynamic system. We know something of ii morphologically
and almo-t all our hypothe-e- .uid theories c< mcernitiL; il are
ba-ed on the morphological data obtained from lixed material.

I'm e\cn \\hen we <oii-ider only the re-ult- of direct obser-
vation ii is difficult to under-taiid ho\\ cytologists can continue

as many do in ignore or minimi/e the importance of the rapidlv
iin rea-inv; number of o|.-er\ation- mi tin- • 'ivmTeiici- nf direct
di\i-ion. ( Vrtainlv tin- \\ork of recent years on proto/oa, to
mention only tin-, ^roiip. ha- brought to li-ht many apparent
facts \\ hich are not in agreement \\ iih current cytological theory.
\\heie there i- SO much ^nmke it \\<nild -eein that there mu^t be
Collie tin .

Moreover, uhen \\. -d the micleii- as a (Kn.unic s\-tem

rather than a UK >rph"l< njcal ^trudure the nece^-it\- for a^>
the indi\ idualil\ <>r contimiii\ of the chromo-ome-
eiitireK. The reappearance o| a <letmite and coil-taut number
ol chroino-i.iiH-, in successive i-ell -eiirratioii-- i- e--ein ially the
>ame problem a^ the reappearance <.f ti\e tinker- on the hand
in successive -.< in ration^ of man, <>r any other case "f the in-
heritano oi organs or pan- in definite con-taut number yel ue
do not regard it .1- necessary to a--ume that the-e fin^ei- .it-
part- maintain their indi\ idualit \ at all Stages of the life cycle.
Instead of bein- tile ba-i^ of heredity the chronio-ome i< it -elf
a problem in hereditx. The theory ••! -o-ealled chr« nno-ome
indi\ idualit\ i- a \\holly nime<(--ai\ hvpothe-i- and a hea\ \
burden for cytolog) I" c.inx. -ince it i- imt in accord \\ith (he
fact'- and require- -upplemcntarv h\pothe-e- at every ]>oint.

There i- e\ t r\ reason to belie\e that the nucleii-. like other
dynamic -\-lem-. \\ ill beha\ e differeiilK under difh-rent internal
and external condition-. What omceixable rea-oii i- there

296 C. M. CHILD.

to doubt that it can under certain conditions undergo fission or
fragmentation and still retain the capacity t<> become a "whole"
again;' \\V know that the capacity for regulation is a very gen-
eral characteristic of living things: are we justified in asserting
without the strongest experimental evidence that the nucleus
is wholly without this capacity?

If it were not for the fact that it makes no essential difference
for the phenomena of life, including those of heredity, whether
the chromosomes are continuous individuals or not this hypothesis
would doubtless have been abandoned long ago. But since it is
in a sense outside the field of scientific investigation, at least
so long as present methods of cell study are in vogue, the chro-
mosome theory has developed into a wonderful inverted pyramid
of hypotheses in the construction of which the chromosome
appears as a veritable dens c.\ muchina.

The great need of cytology is a substitution of the experimental
for the descriptive method: more experimentation and control
and less inference from observation, a dynamic rather than a
static point of view, rigorous proof rather than loose speculation;
with such changes we may hope to gain some knowledge of the
cell. The disadvantages of the present method are sufficiently
manifest in the present paper.

II' I.L ZOOLOGICAL LABORATORY,
rxivKRsiTV OF CHICAGO,
July, ion.

BIBLIOGRAPHY.
Boveri, Th.

'07 Zellen-Studien, H. 6.
Child, C. M.

'073 Studies on the Relation between Amitosis and Mitosis. I. Development

of the Ovaries and Oogenesis in Moniezia. Biol. Bull., Vol. XII.
'o7b Studies, etc. III. Maturation, Fertilization and Cleavage in Moniezia.

Biol. Bull., V..1. XIII.
'070 Amitu-i> as a Factor in Normal and Regulatory Growth. Anal. Anz.,

Bd. XXX.

'10 The On iim-nrr of Amitosis in Moniezia. Biol. Bull., Vol. XVIII.
Godlewski, E., Jr.
'09 Da Vererbung i>mM<-m im Lichte der Entwicklungsmechanik betrachtet.

Voiti. 11. Ant-. 11. Entwickelungsmechanik, H. IX.
Richards, A.

'09 On the Method of Cell Division in T;rnia. Biol. Bull., Vol. XVII.

'n The Method of Cell Division in the Development of the Female Sex Organs

of Moni.via. Bi,.l. Bull., Vol. XX.

DROSOPHILA AMPELOPHILA LOEW BRED IX THE
DARK FOR SIXTV-XIXE GEXERATIOXS.1

FERXAXDUS PAVXE.

In a short note fBiuL. BULL., '10) I stated that I had been
breeding Drosophila in the dark for forty-mar .genera t ions or
nion- than two j that the darkne-s had produced no visible

Ct either in the color of the body or in the structure of the
: but th.it thi-iv -eemed to be a difference in their reactions
to light, the Ones bred in the dark react in- more ^lowly than tl
bred ill the livjit. The e\ idence on which tin- la-t statement
was based \\a- \\ It \\.i-, obtained by placing a number of

Ilie- in a clean \ ial and then iv\ol\ -\\\± \\ 50 as to brin^ lir-t one
end and then the other toward tin- li-ht. In tliis way it was
demonstrated thn the the- bred in the dark were noticeably
much -louer in theii ions than tho-,,- bred in the li^lit.

These observations as far as they went \\ irrect, bul as we

-•hall 866 they uei'e not extensh ' an I hence led loan

i i roneoua 'ttion.

I ha\e iust linished :i -nt in which each lly was te

indi\ idualK'. I'hi- inieiit ua- ba-ed upon four Aeries of

llie-, I .000 in each ;i ,. I , dlOSC bred ill the

dark for sixty-nine generations jeries no. 2), th

bi'ed ill the dark lor -i\tv-loiir ^em-rat ion-, and then placed in
tin- liijit tor -i\ -t iieratioii-; third -erie- no. 3), tho-e bred otil\-
in the li:<ht: and fourth . new --train from series

m>. .1 bred in the dark for live o-iieration^.

The apparatus for testing each ll\- coiiMMed of a \\"eUba. h
lamp, .t h. reen three inche^ thick and a ^la<s tube one inch

in-ide diameter and nine and three eighths inches luii;<. The
a|)paratn- \\a- arranged a-> -hown in the diagram l;i-. l), the
end of the tube toward the lii;ht ln-in^ ten and three fourths
inches from it. Tin- Ua— • tube \\a- perfectl>" clean on the inside
and wrapped with black cloth on the out-ide. The llie^ \\

1 Cuntriluitimi limn th ical Lah- .tati-ry < >!" In liana triivrr-ity, No. 120.

2 -•

FERNANDUS PAYNE.

introduced into this tube by means of a shell vial three and one
half inches in length and large enough to slick' over the end of
the glas> tube. The end and about one half inch of the side of
this vial were also covered with black cloth so when the vial was
slipped over the tube, the tube admitted no light except at the
end toward the lamp. By means of a stop watch I obtained tin-
time it took each fly to travel the distance of nine and three
eighths inches toward the light. Someiimes the flies did not
react and in such cases they were counted out at the end of one
minute. Records of these flies were also kept. This limit
wa> placed at one minute, as experience showed that if they did
not react within this time they might not react for five or ten

B

FIG. I. Arrangement of the apparatus. A, the shell vial, three and one half
nches in length; B, the glass tube, nine and three eighths inehc< IHUL;; C, the heat
screen, three inches thick; D, the light, ten and three fourths inches from the end
of the glass tube. The parts colored black wen? wrapped with black cloth.

minutes, or even longer. Extreme care was used to have ex-
ternal conditions as nearly uniform as possible. The temperature
of the room at the time of testing varied between 74 and 78
degrees Fahr., but I could not see that a variation within this
range made any difference. All flies were changed to perfectly
clean vials and allowed to Mand from ten to thirty minutes before
testing, so they might become accustomed to a clean surface
such as the inner surface of the tube into \\ Inch they \\eiv intro-
duced. The flies bred in the dark \\ere placed in the light during
this ten to thirty mimito in onlrr to overcome any temporary
effect of the darkness. All tlirs \\ere tested at approximately
the same age, from six lo twenty-lour hours alter hatching.
They were fed on ban. ma which wa^ penniiied \o reach about
the same degree of fermentation before using.

DROSOPHILA BRED IX THE DARK. 299

\Yith these precautions it was found that offspring of different
parents within the same strain, those bred in the light as xvell
as in the dark, vary considerably in their reaction-. Further,
this difference does not seem to be due to inheritance from the
parents. To tesi thi- I selected five Hie--, each <>i which failed
to pass through the tube in five minutes. Tin- next day, I
retested the-e -aim- fi\v flies and again they failed to react.
This sluggishness \va- not due to poor development, a- tlu-v looked
Strong and vigorous. These five (lie- \\etv bred and their off-
-pring tested. The avi Mine of tin,-.- \\hich passed through

the tube and the percentage which fail<-d was approximated
tin- same as tor tin- MTU--, indicating that -omelhing other than
inheritance i- ih< cause "I" thi- differem v in reaction. 1 h.i\e
taken t\\o pair-, brother- and sisters, f-d them on tin- -ame food,
kept tin in -ide by -ide in tin- -aim- room and yet tlu-ir offspring
-li'iucd a decided din. in their reaction- to light. This

fact has caused m< >t -leal of trouble and led to the eiTO-

neouscon< eption -tated in the pre\ ion- note. In fat t I can obtain

the -ame re-nli between the offsprii lifferenl parents in the

same Strain \\hidi I formerlx -tated \\a-obtained b\ c, .mp.iring
Ilie- bred in the dark \\ith tho-r bred in tin- light. Often this
dillen i -I that it i- noticeable before the llie- are

ted, thi a-t beJDg more ea-il\ e\ciiei| b\- handling.

I • OD!\ possibl thi- dilieren.-e \\hich 1 have to £

i- liie food, a- it max '.in. nt degrees of lennentalioD

after it i- placed in the xial. Another thing which lead- me to
beliexe thai the |o»d max be the uiiderlx ing caii-e i- thai the
la-t the- taken Irom anx one xial are u-u.illx more -higgi-h than
the lit.-t one I .en \\ith thi- differi'iici- in the reaction- of

tile Offspring Oi «litlel« III parent-. I belieXe the reacticiD- of I.O.KI

individuals trom manx dillereni jiarent- gives .1 close approxi-
mation to the average for the -train.

The lollou ing table gives the axerage t ime of t he the- in pa — itrj

i.cx- i- in

I thi- I 'ite.

i : -'5. i per i-t-iit.

l : . lid-. Ml.

Sen. } No. 3 i 'iuls. nt.

n-l-. r cent.

3OO FERXANDUS PAYXK.

through the tube and the percentage which failed to pass through
in one minute.

This table shows that the average time of the flies bred in the
dark for sixty-nine generations (series no. i) is least, 13.9 seconds,
and the average of those bred only in the light (series no. 3)
is greatest, 17.62 seconds. This is a difference of 2.72 seconds
in favor of the flies bred in the dark. Likewise the percent-
age of flies which failed to pass through the lulu* in one minute
is less (25.1 per cent.) in series no. I than in series no. 3 (29.5
per cent.). Series nos. 2 and 4 were run as controls and the aver-
age time is intermediate between the two extremes, while the
percentage which failed to go through the tube is less than in
series nos. I and 3. However, I do not believe the difference
between any of the series great enough to be of any special sig-
nificance and the only conclusion which can be drawn is that the
darkness operating through a period of sixty-nine generations
has produced no visible effect in the reactions -of Drosopliila
to the light of a Welsbach burner. It should be pointed out,
however, that the only cave condition present in the experiment
is darkness; that it is possible and indeed probable that factors
other than darkness (constant temperature and moisture) may
play some part in the changes which have taken place in cave
animals. If this be true the real test comes not in rearing animals
in the darkness but in a true cave environment.

That the constancy of the environment may be a potent factor
in the production of degeneration in cave animals is strengthened
by the recent unpublished experiments of Tower. In a letter,
August 2, 1911, Professor Tower makes the following state-
ment:1 "It has been my experience that there is hardly any-
thing so injurious to breeding stock as a constant environment.
It will produce degeneration, reduced activity, and not infre-
quently will result in actual elimination of the race." Al-o
in the experiments of Calkins and others on pmto/o.i, it seems
that the constancy of the food supply and the consiuncx of the
chemical make up of the medium in which they lived, brought
about, at least a shorter cycle than occurs in a food Minnlv am!

1 1 wish to thank Professor Tower for the permission to use tliN -Md-incnt before
it has appeared in print.

DROSOPHILA BRED IN THE DARK. 3OI

medium which are changed at intervals as shown by Woodruff's
experiment (Archiv fur Protistenkunde, 'u).

In caves conditions are practically uniform. Also in dark
corners under stones and logs, conditions are more'nearly uni-
form than in the open. Since it seems almost certain that forms
which now inhabit caves once- lived outside under stones and in
dark corners and that they had varied in the direction of cave
animals before they entered cavi -, is it not possible that the
more nearly constant environment of the dark corners under
stone- and lo-s ha- st.irted the degenerative change-; which have
been carried to their present condition in the more constant
en\ ironiiu-ni of tin- ra\ • Po be -lire .ill changes in rave animals

not degenerative. The tactile sense, for example, becomes

highly developed, but no doubt a sec >nd (\ui-viti\e fart..r eir
here.

SOME PARASITES OF SIMCLITM LARY.K AND THEIR
EFFECTS OX THE DEVELOPMENT OF THE HOST.1

E. H. STRICKLAND,
CARNEGIE SCHOLAR IN ECONOMIC ENTOMOLOGY.

During the spring of the present year (1911) while studying
entomology at the Bu^rv Institution of Harvard University,
I made numerous collections of Simulium larva1, which arc
extremely abundant in the neighboring streams, with the in-
tention of studying the development of the imaginal discs which
are unusually well defined in this genus of diptera. As, however,
I found that a large percentage of these interesting larvae were
heavily parasitized by two very different organisms, namely a
worm and a protozoon, I turned my attention rather more
directly to these and their effects on their hosts, during the feu-
weeks intervening between my first discovery of the parasites
and the pupation of the insects.

Before giving any details I wish to take this opportunity to
offer my sincere thanks to Professor Wheeler, who by his kind
suggestions and advise enabled me to bring together the fol-
lowing facts, which, though very incomplete in form, do not
appear to have been recorded before. My thanks are also due to
Professor Johannsen for naming the species of Simnlinni larvae
and to Professor T. H. Montgomery who identified the worm
parasite as a species of Mermis.

LIFE HISTORY AND STRUCTURE OF SIMULIUM LAUV.E.

A brief summary of the structure, and mode of life, of the
Simulium larva? may not be out of place here.

If during the months of March to May one examines the rocks
or vegetation in any swiftly flowing stream in the neighborhood
of Forest Hills, Mass., especially where ii-> bed causes a small
cascade, one will, in all probability, observe a large, dark,
gelatinous mass where the current is >\\ittest and the water

'Contributions from the Entomological Laboratory <>\ the lln—i \ Institution,

Harvard University, No. 45.)

302

SOME PARASITES OF SIMULIUM L \R\VE. 3°3

shallowest. On closer examination the mass \vill be seen to
consist of numerous curiously shaped larvae standing upright
on the rock, to which the hind end of the abdomen is firmly
attached. These larva*, the largest known North American
species of which measure when mature -ome 12 mm., have the
following characters: The soft-skinned body is more or less
cylindrical, though the posterior thin! i- -omcwhat swollen.
The segments are poorly defined but the first three behind the
he.id. namely, the thoracic segment-, are usually distinctly
marked off from tin- following abdominal -eminent-.

The prothoracic segment bear- a -inije leg (Plate I.. Fig. l)

which i- apparently t\\ «-\< >inted : tin- distal joint i- -mail and

retractile and terminate- in a -inker which i- armed \\ith numer-

on- hook-. \o other -e-meiit bear- any trace «.f le^-, with the

piion ol I'M- a|»ical abdominal -e-ment when- the pair of anal

prolei;- of -onir oilier lar\.e i- represente 1 by a powerful, armed

di-k like -inker, by mean- of which the larva firmly attaches

ii-'ll i" rocks or \ew.-iation \\hen at re-t. The -kin i- u-ually

: vjveiii-li i^ray color in immature larva-, but gives place to a

reddi-h bn>\\ n as t he\ inatn:

The head i- -nb-c\ lindrical and i- n-uallv of a <larker color
than the re-t of the bod\ . It- chitinoii- inie-unieiit i- much
den-er and less ela-tic than that co\i-rin^ the remainder of the
boi|\. Tin- re-ult-. durii - >uth. in an e< d\ -i- of the heail
cap-ule alone, bein- n< \ more trei|in-ntl\ than that of the

• -r.il cntii iilar io\erin^ "1 the body. The lieu head cap-nle
e\po-ed alter -mil an e< d\ -i- i- |»erfec!ly pi 14 men tie— in Sininlimn
liirlifx-), llioii-h dark 5] "in form on it- \erte\ I'late I.. I

2, a. i> and , . lolloucd b\ a -r.idn.il inlii-cation of the \\liole
-nrlace. Indixidiial- \\ere observed in which the almo-t black-
head c.tp-llle \\.i- half reillo\fd. e\po-ill<^ the new pi^melllle--
cap-ule. entiivlv inde]iendeiiil\ of the bod\" cuticle, the ecdysis

of \\hich \\a- never observed, except at pupation.

( )n either -ide of the head are two black eye spots and on the
anterior border are two lateral tan-like organs borne mi elongated
pedicel-, which are u-e.l by the larva in procuring food. Th
fans con-i^t of numeroii- cur\ed rake- 'Plate I., 1 i-. 3) bearing
on one -ide Ion- -tiff" cilia which, when the fan is expanded,

3O4 E. H. STRICKLAND.

stretch from one rake to the next, thus forming a very fine
strainer which allows water to pass through it, but retains any
small organisms, such as the diatoms, on which the larva subsists.
By means of a curious flicking motion the^e fan- can be closed
and their contents brushed into the mouth orifice. Here are
situated two large glands (Plate IV., Figs. 9 and 10 (d)) on the
dorsal side- of the pharynx, whose function appears never to have
been determined. They are covered with an apparently porous
membrane which is clothed with short stout hairs. It would
seem that they secrete some sticky material onto these hairs
which removes the particles of food from the rakes when the
latter are brought in contact with them.

It is usually stated that the fans are used to set up currents
in the water and thus sweep food toward the mouth. My
observations, however, lead me to believe that they act as a
"strainer" and this is supported by the fact that, living as these
larvae do, in the swiftest currents, such movements would be
useless.

The salivary glands are very large and secrete powerful silken
threads which are used by the larvae as anchor lines to hold them
in an upright position no matter how strongly the current of
water. may flow.

Although the larvae appear to be very sluggish, they can move
about actively on the rocks by a lopping motion similar to that
of the geometrid larva. It is interesting to note the care with
which these larvae, when thus moving about, assure themselves
that the adhesive disc of the prothoracic leg is firmly attached
before relaxing the anal disc and vice versa. Respiration is
accomplished by means of retractile finger-like blood gills,
normally three in number, which are situated on the dorsal
surface of the last abdominal segment just above the anus.
These gills, which can be distended at will by the insect, by means
of blood pressure, are apparently inadequate for use in any but
rapidly flowing water which contains plenty of oxygen, for if the
larva? be placed in a jar of still water very few will survive for
more than eight hours, though life may be prolonged for two or
three days by placing them in small numbers in I'etri dishes
containing only sufficient water to just cover them.

SOME PARASITES OF SIMULIUM LARV.E.

The form and size of the imaginal discs, or histobla-t-. will
here be described in detail as they are of especial interest in
connection with the parasites.

The thoracic hi-toblasts are twelve in number and aiv very
large and conspicuous in normal larva? as they approach lull
growth i Plate I., Fig. _j .

The six discs to be found on each side of the thorax are the
following:

1. Adult prothoracic leg histoblast — situated at the base of
tin larval prothoracic leg.

2. Adult mesothoracic leg histoblast — -ituated ventro-laterally
on tin- me.-oihora< i'- -e-ment.

.v Adult metathoracic leg histoblasl situated ventro-laterally
on the metathorai j, . m .

4. Adult \\iiu hi-lobla-t - -imated dor-o-laterally oil the
mesothoracic segment Tin- in th»- inatun- lar\a i- by far the
largest di-c and it -• •« m COmes in contact with the nie-othoracic
ICL; <li

5. Adult halteric hi-N-ltla-t This in the early -ta^es is
alino-a ,i~ large as tin- u in; disc, but it- 'vM'iiu th ami de\ elopment

are verj -\<>\\ an-1 ii soon disappears under the rapidly expanding

\\ ini; di-f.

6. I'upal iv-pirati>ry tutt hi-t< >l >la-t . Tlii- i- -it n.ttfd mi the
pmtliorax and in ii- VOUl ha- the ap|»earanee ••!' bein^
quite lioiimli'^i'ii- \\ith a pn >t In -raeie \\in.u. It, h«>\\e\er. -n«>n
begins t«> lake on .1 ddinin- t'..rni, and the r<miparati\ ely
traeheal tube- ran In- BCCn ^i"\\in- and leii-t helling

the traii-pai'ent cuticle till tlle\ berime e. tiled tip a- indicated in
I'late I., Fig. 4.

It -liquid be noted that tin- hi-tobla-t i- imt in the true sense
of the \\onl an "inia^inal" di-c since the re-pirator\- tilanieiiis
an- e\clu-i\ely pupal Organs, and ha\ e no e(|iii\al'-nt -tructnre
in any adult. Their -imilariu lo a pr«»i honn ic \\ in- de-titute
of the \\ ini; meinbrane is of intere-t . A le\\ day.- before pupation
tin- "|>upal" hi-!obla-t becin- to darken. ri-nieiiiatioii com-
mence- at the apex of the folded tube- and slowly work- back-
ward- to\\ard tin- ba-e \\hen the di-c lake- on the appearance
shown in Plate lib. I . Later, immediately prior to pn-

E. H. STRICKLAND.

inn. the cuticle over this disc ruptures and liberates the fully
formed and now functioning filaments.

The growth of these various histoblasts causes the thorax
to >well considerably, thus giving the body the appearance of
being const rioted in the middle.

Internally there is comparatively little tissue. The abdomen
contains the alimentary tract, and the much elongated salivary
glands which lie normally in a ventro-lateral position with regard
to the alimentary tract. The sexual organs are small and not
ea-ily found even in serial sections. The remaining portions of
the body cavity are filled with blood plasma in which is suspended
a quantity of fat body, mainly collected near the apex of the.
abdomen and causing this region to become slightly swollen.
As the larva- mature this fatty tissue very materially increases,
and when dissected out, is of a stringy nature.

METHODS.

Most of the larvae were killed as soon as they were brought
into the laboratory, but a few of the more heavily parasitized
ones were kept, alive in running water by covering the mouth of
a jar with fine net I ing and introducing a piece of rubber pipe into
the jar, through which tap water was run. In this manner
-pecimens were kept alive for several days.

The following killing fluids were found to be the most satis-
factory among several tried:

1. JI<>! Water. — Water was just brought to the boil when the
lar\;r were immersed and allowed to cool in the water. This
method was most unsatisfactory from a histological point of
view, but it had the advantage of leaving the skin as transparent
as in its normal condition. It was also possible to dissect larva-
thus killed.

2. Ci il son's Fluid. — This was used hot as described above and
gave good n-Milis, but had the disadvantage of making staining
with the haematoxylins difficult.

;v Kalile's Flu id, consisting of 30 parts water, 15 parts 96
per cent, alcohol, 6 parts formalin (40 per cent.), I part glacial
acetic acid. This fluid has been recommended by W. Kalile
(1908) and proved to be superior to Ciilson's fluid both for

SOME PARASITES OF SIMULIUM LARV.K. 3OJ

fixing and staining, and in the end \vas exclusively used. It
was also used hot.

The chief advantage of both of these fluids was that. besides
giving excellent histological preparation, they cau-ed tin- hi^to-
blasts to turn milky white so that their position and form could
be readily observed immediately after the specimen- \\erc killed.

Staining. — Heidenhain's iron haematoxylin combined with
orange G was almost entirely used as it gave the U>t diftVr-
entiation, though -tamiim \\a-ratherslow. Replacing orange G
with i-o-iii (crated staining but differentiation \\a-le—

Mi KMI- SP. I'AK \-i n/iNi, SIMII.H M L\K\ i.
\ number of larx.e were placed in a jar of water and left
over niijii. The following morning I was -urpri-ed to
-e\eral white worm- mo\in- about at the bottom of the jar.
h was c\ident that the-e had o.me out of tin- SiniitHnm
larvae, SO I went OUt to a ripple W lien- the lar\ a- were particularly
abundant and examined the colonie- on & Mia! Stones. I then
noticed the large size o| man\- indixiduaU. and on examining
these I loiniil that manv of them liad a \\orni coiled up \\ilhin
the abdomen I'latc I . I - When a <|iiaiititv of material

\\a-luon-lii i in o i IK- laboratory it \\.t- found that the-e para-^iti/' d
larxa- were much more -In — j>h than their lieabhv companion-,
\\hicli iapidl\ explored the iar in \\hiih tlp-\ were « oiiliiu-d.
\\ith their peculiar loopi: ;. The para-iii/ed indi\ idual-,

h<i\\e\(i. seemed mm h Ic-- concerned as to their new -nnouiid-

iirc-. and maii\ o| them \\eic ~oon motionle--. -laiulin^ tipri^ln
on the boiiom and -ide-, of tin- jar with tlu-ir rakes expanded,
i«ad\ to caich aii\ lood which mi-lit lloat tlieir way. Mere
i- a po--ible explanalion of their larger size. <)\\in- (,, their
curiou> form of feeiliir^ it i- e\i.|ent that the more >hi^i-h an
in.li\idual i- the more f ..... 1 it can obtain, and -.liould it .^row
bin a little larger than it> companion-, it will reach abo\ e their
innumerable oiit-t retched rakes, SO that its food Mipplv will be
\ ei \ matciially inrrea-et 1.

It mu>t not ho\\e\er be taken for granted that all |iara-ili/e<]
larxa' \\erelaixcr than their lu-alth\- companion-, for -o me re-
mained i|iiite -mall, \\heiva-, main' of the larger lar\a- -lio\\e,| no

3O8 E. H. STRICKLAND.

signs of worm parasites. As a gnu-nil rule, however, ilu- largest
larva? were found to contain one or more worms coiled up within
the abdomen. One would naturally expect this rule not to
be YITV constant, if, as conjectured, it is simply due to a more
>luv;:J-!i u-mprrainent and increased appetite on the part of the
parasitized larva?.

A case of a Mermis parasitizing ants was described by Professor
\Yheeler ('07) and here also he noticed a great increase in size
of the host due to a greatly increased appetite during the larval

A number of tin- worms were dissected out and sent to Professor
T. ]]. Montgomery who pronounced them to belong to an
undetermined species of Mermis.

RETARDATION OF DEVELOPMENT OF THE HISTOBLASTS DUE TO

MERMIS.

On making a closer examination of the parasitized larvae a
far more remarkable effect than that of increased size was noticed,
for it was found that the presence of Mermis parasites, no matter
in what numbers, has a direct effect on the development of the
histoblasts. In a normal larva of about 10-10.5 nim. which is
the maximum length, and is attained immediately prior to the
blackening of the respiratory filament histoblasts, the latter are
quite large and owing to their white color readily visible to the
naked eye; especially when the larva has been killed in Kahle's
or Gilson's fluid (Plate II., Fig. 6). If a parasitized larva of
the same, or greater, size be examined no trace of the histoblasts
can be discovered with the naked eye, and can only be detected
with difficulty under a dissecting lens. Under the low power
of a compound microscope, however, they are seen to be repre-
sented by small white traces of the organs which should at this
time be far advanced in development (Plate II., Fig. 7). A
do-i- examination fails to reveal any differentiation of these
in led histoblasts into the component parts of the adult,
or pupal, organ.

The parasitized larva rarely develops beyond this stage, though
I have observed specimens which were turning reddish brown,
and contracting slightly as a healthy larva would, shortly before

SOME PARASITES OF SIMULIUM LARV.E. 309

maturation. In some cases, even, the rudimentary respiratory
filament histoblast begins to darken in color.

I examined numerous specimens from the same Miv.im at
Forest Hills, and, with slight variations in intensity, they all
showed this nearly complete suppression of the imaginal di-c-.
Some weeks later I chanced to pass a small stream at Norwood,
about .-even miles from Forest Hills, and seeing Simulium lar\ ,e
pre-ent in small numbers, I took nine specimens from the rock-
in «.nler to see whether the species w.t- identical with that common
in our >treams at Fore-l Hill-. They proved to belong to tin-
same specie-, namely, Simulium hirlipt-s, but I was much sur-
prised io find that hen- al-o ihe .l/,7/;;/x para-iu \\a- much in
evidence for o| tin- nine -pecinien- taken. -i\ contained the \\orm.
Tli' •- |)O\M \erin th»--i- -pecinien- were- not -. ;• marked a- in

tho-r found nearer home, lor tour of the -i\ had turned bn>\\ n
and althoii-h the hi-inbla-t- \\eiv much -mailer than is normally
the case ihe\ \\« r- readily visible to the naked eye. rndoubtedK ,

houe\er. malui.ition u.i- impo— ible. t if even should the lar\ a
manage to |iii]iate it \\oiild -ooti ilie for \\aiil of oxygen, -illCC
the ie-piratorv hlanu nl- \\ere but half formed.

In .1 third I'M.iliiv. the Slonx brook Reservation near Fop

Hills, a different sp< larva was found. l'r»: loh.mn-en

-.t. Me- that thi- laiA.i i- quite neu to him and mav be the tin-
d( Drilled lar\al Stage of ':t»:. uhich OCCtirs I Vei [iient ly

in thi- . but as I found only adult- of hirtipt* it wa> im-

p"--ible i i ill rm thi- supposition. The-e lar\ a- u ere. ho\\ e\ er,

al-o f..imd t" -utter Inun .Ucr;;;/\ para-iti-m, but in a much

le-- tAlellt than tll.>-eo| .S . /'..'•. The el'lecl- nil tile '

were, however, as one \\mild expect, identic. il \\-it.i tho-e on

hirtipcs, though the increa-ed -i/e \\.i- no) SO exideiit.

A large number ol latx.e \\ere measured, cut «'p(-n and the
\\ollll- remoxed. It i- ill tere-l i IIU here to Hole th. 1 e\-en ill
li\ed specimen- \\here tin- lluid had cati-ed the skin t" become

opaque, the presence or absence of worm- eon'd in alJ ca-es be

determined \\ith certaint\ by a .lance at the hi-toblanl -. The
\\onn. it should be noted, did not cause the abdomen to -well
up or Income di-torted to any appreciable <!•

A I. \\ -elected results -ho\\ n by the exam ina i ion are appended :

E. H. STRICKLAND.

1

V,. ..!'

1 '-ii^th.

i ••!"! .'t l..ir\;i. Histoblasts.

\\ 't m>.

S e "t W..ims.

I

10.75 mm.

Gray.

Minute.

12

A.V. .75 i cm.

2

1 1. 50 mm.

( '.ra\'.

Minute.

I

2.75 cm.

3

n.o mm.

i tray.

Minute. i

3.00 cm.

4

ii.o mm.

Gray.

Minuti'. 3

1.3. 1.5. 1.5 cm.

5

10.5 mm.

Brownish.

Miimif. I

3.00 cm.

6

9.0 mm.

Gray.

(Jiiitt- small.

3

.9, 1.3, 1.5 cm.

7

8.5 mm.

Brownish.

Large.

0

8

10.5 mm.

Gray. Large. o

9

8.0 mm.

Brownish. Large. o

10

7.0 mm.

BrcnvnNli. Large.

P

I !

.v.S mm.

Gray. Large.

0

12

n.o mm.

Gray. I Large.

1)

From this table it will IK- seen that when there is but one worm
in ,1 hn-i ii .it Min- a length of about 3 cm. (luring its parasitic
lil'c; that is, roughly eibout three times the length of the host itself,
since the average length of a parasitized larva is 10.5-1 1 mm.
It will al-o be -cen that in one case as many as twelve worms
\\ere removed from a single host, and that in this case they all
remained small, owing doubtless to insufficient food supply. In
other instances, however, where several worms were present in
one host ii \\a- noticed that one had attained almost to the normal
size of 3 cm. wherea- the remaining worm or worms were less
than I cm. in length.

The larval measurements showed that parasitized larva- a\ erage
about i i mm. (Plate III., Fig. 8</ ) whereas mature, and therefore
-' niie\vhat contracted, larva-, a\ crage some 8-8.5 mm. (Plate III.,
Fig. 86), though a few are abnormally large.

Li !••!•: HISTORY OF TIIK WMKM.

As yet nothing is known of this except during the latter pan
of tin- life in its host. In all probabilit\ tin- eggs or young
worm-, are caught as they float down stream by the outspread
larval rake- and are M\ept into tin- mouth, passing into tin- ali-
mentar} .ract through the walls of which they bore- their way
till they enter the body cavity. It should however be noted
that no Bears or hypertrophy of the wall of the alimentary
tract \\cie noticrd in -.everal serial section'- made of parasiti/ed
larvae. Tin- alleniati\r hvpothi--is i- that the larval \\orm- bore
into their hosts through the thin cuticular covering of the ab-

SOME PARASITES OF SIMULIUM LAKV.1 .

domen. That thev -hould be present in the S'nnitlium e^> i-
hardly possible, and this hypothesis can be with safety rejected.
Either of the former hypotheses offers a po--ible explanation
for the variation in development "t" the histoblasts of para-iti/ed
larva- i.ikcn from different streams, for in the case ot those taken
,n Norwood where the di-e- were of a moderate si/e. the worms
may not have entered the body cavity till the di-e- were -ome-
\\hat dc\ eloped Then ai^ain when- OIK- worm ha- -ro\\n to
nearly the normal -i/e of ^ cm. \\lu-rea- other \\orm- in tin- -aim-
hosl ha\e n-m. lined -mall, it i- probable that the lar^»- \\orm
ciilcre<l the body cavity -onic time In-fore the other-.

I- i;^ urc ( i, I Mate [V., which is a drawing of a median longitudinal

ion of .1 p.u.i-ili/ed larva, >ho\\- the ventral position of the
\\orm \\itli re^anl to the aliment. iry tract, though it will 1 M-

i that the me~eiitcroii lia- lieni -oineuhal pn-hed to i.iie -ide
l.\ the coiled parasite. It \\ill al-o In- -een 1>\ comp.irin^ I u. <i.
IM.ilt l\ '.. u ii h 1 • ii-. I I , I Male I I I .. I hat tin- para-iti/e;l lar\ a ha-
much le-- fat IM,(|\ than thai of a liealtlu individual a! aliotll

the same rom th.

Another e||. 'lie ho-t i- that apparently the -e\nal oi-an-

do noi develop in paia-iti/ed l.n \ .e. The-c arc- n-.t easilj seen

ill a lieallh\ Simi4 rva, as ihe\ are \er\ -mall, lull lhe\

can n-nallv lie found in . ies "I -eciion-; in sections <»l

|i.na-ili/ed l.n \ e I ha\e l.een eiitin-K imalile to locate them.

< Mh ei u i-e the parasite appear- in ha\ e im el led on the internal

in- ot the ho-.t. \\ith the excejition of di-]ilacin- the -pinning

-land-, the I. ;iortion- < .1 \\hich normalK lie in the po-itimi

later occiljiied \>\ the para-ite. Alt hotU'h the\ m.iy lie -o di--

plai'ed a- to lie d.ii-.ul of llie alimeiilaiA tract ' IMale 1\ . I

their functioning •- in no \\a\ impaired, since these larvae -pin
.mile as man\ -ilken threads as do those which are uninfested.

The .\/<T;;//> prokiMy teed- <lirectl\- on the Mood pla-ma and
fat t\ ii— ne of the ho-t.

A- the lar\a reache- maturity the contained \\orm |.e«nne-
\er\- ri--tle--. and if a li\in- lar\a la- placed under tin- lo\\ pouer
of a micro-cope in a cell -lide the mo\emeiit- of th-- \\orm can he
ea-il\- ol>-cr\cd. I hie thn- \\atched lor aliout half an hour \\ a-
to explore the h\ |iodermi- of the ho-i , e\ idem 1\ atlemptin.u

312 E. H. STRICKLAND.

to find a place of exit. The head was kept constantly moving
over the intern, il surface of the abdominal hypodermis and was
even sometime- thrust far into the thoracic region. The move-
ments appeared to cause the host great pain, especially when the
thoracic region was visited. Finally after the entire worm had
been several times t\\ isted round in tin- body the head was pressed
against the junction of the third and fourth abdominal segments
and a hole was quickly made through which the worm slowly
emerged. The worm, measuring 3 cm., took in all 27 minutes
to disengage itself after puncturing the skin of its host. At first
the operation was very slow and the constant writhing and turn-
ing of the host impeded rather than aided the movements of the
parasite. When about 50 mm. were exposed the head was twisted
around the body and the remainder of the worm was more
rapidly forced out, finally becoming detached from the host in a
tightly knotted mass which soon straightened out. The larva
meanwhile rapidly shrank, and in about half an hour was dead.
The worm apparently can leave the abdomen at almost any
point, though the thin junction between two segments is usually
chosen. It is probable that the death of most parasitized larxa-
is directly due to the escape of the worm, which in all obser\ ed
cases occurred some time before maturity.

On raising leaves in the bed of a stream a little below a large
colony of Sim it! in ni larv;e it was found that they had under
them several of these white worms. This year most of the worms
had CM -aped by the beginning of May, and ha\e since been lost
sight of, though while examining a stone microscopically for a
second brood of Sininliiini eggs, on May 29, one or two minute
worms were seen, which may have, been young Mt'rniix. Xo
Sinniliitm eggs could be observed. At a still later date, August
3, a full grown, though dead, Mermis was found under a stone
in the bed of the .stream.

PKK< EN i A.GE OF I. AKY.K l\l I SI I-.D.

Several leaves co\en-d with larv.r were taken from a stream
in Forest Hills for the purpose of determining the percentage of
parasitism.

These larva?, which repiv-eiiied \\\(- species Sininliiini hirli/>fs.

SOME PARASITES OF SIMULIUM LARV.E. 313

gave the following results: Xumber of larvae present 174; number
of parasitized larv;e 41. This means that in this particular
locality about 23.5 per cent, of the larvae would be unable to
mature on account of tin- Mermis parasite.

The species living in the Stony Brook Reservation was found
to be para-it i/ed only to the extent of some 3-4 per cent. A few
6". hirtipes were also present in the same stream and were para-
-i i i/ed only to the same extent, so it is probable that thi< smaller
attack wa- not dm- to tin- host's belonging to a different species.

CAUSI "i IHE RETARDATION «\' mi; HISTOBLX-I-.
In order to arrive at some definite conclusion as to \\liv the
hi-tobla-t - ol ,1 parasitized larva should not develop normally,
it will In- o| advantage to review briellv xuiu- of tin- theorie-
which have been advanced to explain the normal development
ol ihe-e <\\~< -,, and at tlie same time to consider the fact-, which
ha\e In- n brought to li-ht by the study of several cases n<>\\ on
• id in u liich the di-.cs have developed with abnormal rapidity.
tlm- prodiK in.; larvae \\hich p - character- that normally do
pot make their appearance until the pupal or adult condition is
reached. This abnormal condition ha- been termed "pro-
thetely" bv Kolbe (1903). l For the abnormally retarded con-
dition as produced in the Sinuilinnt larvae by the pre-ence of
M,->n:is I should like to surest the word " metal heteK ."

The earlie-i re|)ort of prothetely was made by Ilexnion-
'96 . In tip larv;p of Tencbrio nwlit<» \. . \\en- found which,

ulien matuic. |'ro\rd to be abnormally developed as follo\\-:

I The meso- and meta-thorax possessed expanded lateral por-
tion- ol the termite-, \\hich resembled the wing-pads oi the pupa,

though the\ \\eie not folded under the body as in the latter,
but \\ere direcit d I'.n kuards.

2. The antenna- had additional incompletely segmented joints,
ihu- a|)pn>achin- tin- i i -jointed adult condition.

,v The abdominal termite- \vere modi tied SO that they resembled
the termite- of the |)U|)al abdomen.

The>e lar\,e wen- rai-ed in the meaUvorm cultures of the

' \lpotf(iv. tn run before, .ml ^'\'->5. th.-
\\>Tat>(tv, to run Ix-liiml. .in-1 -f\os. tin-

314 E- H. STRICKLAND.

Berlin Zoological Institute, but no statement is made as to
whether conditions during development were quite normal. Hey-
mons concludes his paper by suggesting that the abnormal de-
velopment described above is due entirely to an accelerated
development of the histoblasts, but makes no suggestions as to
the cause of the acceleration.

In 1903 Kolbe reported and figured an interesting case of
prothetely in the larva of DetidroHmus pini L. He received the
larva when in its fourth moult, at which stage it had the following
characters:

1. The larval antenna? were replaced by elongate antenna?
showing simple primary division into about seven segments.

2. The larval thoracic legs were replaced by three pairs of
jointed legs possessing all the adult parts, namely, coxae, tro-
chanters, femora, tibiae and tarsi.

3. The mouth parts were modified.

Kolbe points out that all these organs were in an immature
adult, or true pupal form, thus showing, it would seem, that the
development was quite normal, but simply accelerated. \Yinne-
guth succeeded in breeding from a similar larva an iidult which
hatched as a small male that w^as apparently quite normal except
for its dwarf size. These abnormal larva? were produced from an
artificially hatched generation kept indoors and from parents
which did not hibernate.

Hagen ('72) gives an account of silkworm larva? obtaining
wings before pupation, which condition was accompanied by
other abnormalities as follows: The head was small and had two
small facetted eyes, and the thorax became modified but the
abdomen remained in the normal condition of a larva in the
fourth moult. The fore wings were long and narrow and rather
more gray than usual, while the hind wings were short and nar-
row. As this anomaly occurred frequently and was therefore
liable to be of economic importance, its causes were investigated
by Majoli who came to the conclusion that it was due to the
larva? being kept at a temperature above the normal.

A case similar to that of Heymons was described by Riley
('08) from another coleopterous larva, Dendr aides canadensis,
which \v.i> bred by a student at Cornell I'niversity.

SOME PARASITES OF SIMULIUM LARV.E.

It will be seen from these cases that in every instance they
occurred in artificially reared larvae, and it is probable that the
prothetely was due to an increased temperature, perhaps with
the aid of abundant food, in some way hurrying on the develop-
ment of the histoblasts.

This fact suggests that the histoblast- may be cau-ed to
develop, though at a much slower rate than the larval organs until
pupation, by some enzyme secreted by the in-ect and that they
can develop only as t'a-t a- thi- -timulant is formed. Ii- -upply
tlm- acts .1- a regulator. An increa-ed temperature i- in nio-t
•d\ antag< «u^ to en/yme action and in thi- case it i- prob-
able tli.it it either causes more of the en/yme to be -ecreied or
-tiinnlate- the action of the amount already available.

I ). v 5 , after numerous experiment- <m retardation and
a« i eleration of development and pupation arrived at the tol-
li. \\iiu i "in lii-ion-: Development and pupation are caii-ed by
enzymes \\hich are not very evident in the early lar\.il

The\ , lio\\e\ei. increase \\ith the growth ot tin- larva until ii-
|)upation. ,n \\hich period they are at their maximum -tivnvjh;
tin \ then begin to dimini-h. till at the end of the pupal period
their action entirely ceases. He also state- that the en/vine
action can be hindered by the presence of another b«»l\ •. and that
by obtain! IK' an en/yme of an increased strength before pupation
de\ el"pnii-ii! can be abnormally accelerated.

I 1 i- exident that the cells of the hiMobla-t- .in- can- d to
dexelop b\ a different stimulus from that \\hich causes the de-
\ elopineiii of the larval organs, for in the former case d«-\ elopmeiit
i- \ei\ -lo\\ during the larval stage, \\lu-n the latter organs are
de\elopin. rapidly, and it is only when the-e ha\« ir.irhed
their limit of de\elopment at pupation, that the adult organs

n in de\ (lop \\ ith an\ rapidity.

Thi> suj - that there may be in the insecl body t u o sets

of eii/> me- \\hich one may term "larval" cn/yme- and "ima:;-
inal " enzymes. The-i- are -ulficiently different so that condition-,
which cause ;he ac. -el. -ration or retardation of one of them n< ed
not ne(e--aril\ h.i\e an\ effect on tin- oilier. It thi- be SO, one
ha- a probable explanation of prolhetely and al-o. a- I -hall
attempt to -how in the l'ollo\\in;c paragraph-, ol metal heteU.

3l6 E. H. STRICKLAND.

At first sight one would suppose that the retardation in develoj >
ment of the histoblasts in parasitized Simiilium larva? is simply
due' to a lack of proper nourishment for thr>e <>r-,m^, MIHV the
lar\-a, besides having to supply the requirements of its own
developing larval tissues, has also to supply the demands of its
fast-growing parasite. This may be true to a certain extent,
but later observations, when another parasite is also present,
indicate that there must be some other more potent factor \vhch
accounts for this inhibition. This second parasite is a Spotozoon
which, owing to the vast numbers in which it occurs in a single
host, is far more bulky than the worm and must, one would
imagine, make far greater demands on the resources of its host.
In this case, however, the histoblasts are usually unaffected in
size, though in many cases they are distinctly smaller than normal.
Two individuals, however, were seen in which the histoblasts
were minute. On dissection it was found that, in each case, a
small worm measuring only some 7 mm. was living embedded in
the mass of spores, and it was evident that this minute worm was
responsible for the retarded condition of the discs, even though
it had evidently absorbed very little nourishment. One must,
therefore, in all probability look to some toxin secreted by the
worm as the cause of the inhibition of development in the discs.

The researches of Verson and Bolle, as quoted by Fischer
('06), proved in the case of lepidopterous larva? that in their
early stages their body fluid is alkaline and that this alkalinity
decreases as the larva matures. This would suggest that an
alkaline condition encourages the growth <>1 larval tissues whereas
acidity, or the absence of alkalinity, permits of the development
of the adult organs. Hence the histoblasts would develop but
slowly till the larva? are nearing maturity when the decreased
alkalinity of the blood allows the "adult" enzymes to stimulate
the cells of these discs to rapid division. I have been unable
to find any account of the excretions of Mermis or of closely
related Nemathelminthes, but should they be proved to have an
alkaline reaction the probability of the above contention would be
very greatly strengthened, for here we should have a case of the
alkalinity of the system being maintained in maturing larva?, and
thus preventing the normal, though slow, development <>t the

SOME PARASITES OF SIMULIUM LARV.E.

adult organs, without affecting the larval organs to any great
extent, except in so far as they are kept well supplied with
alkaline fluids and'are thus capable of developing to their utmo-t
extent. This would account for the somewhat larger size of
parasitized individuals.

It may he. however, that the Mcrmis doe- not actually secrete
an alkaline -ill -lance, hut brin-- ahi'in an increased alkalinity
in the hodv of it- h-.-i by ah-orhini; whatever arid- are lonned
in it. The prohahility of this being the true condition i- in-
1 hy the fact that closely related \\orm- li\ e in acid media.
Midi as \int-var or sour paste (An'^nillnln nccti Chrb-. . \\liich
uould point to the fact that the worm require-, and ah-orhs,
acids 'liriir^ its development. In either case the eliect on the
ho-t \\ould he the same in that there is a reduction in the acii\ iiv
of the arid- \\hich ap[)ears to be essential to tin- d.-\rlopineiii ,-t
the hi-iohla-i- Whether this. is the true explanaiion or not,
it i- it-nail) that the presence of the worm does ha\e a direct
inliihiioiA .11.. t upon the development of the ima^inal organs,
hni does not ha\e a similar effect on the lar\al tissm

A -uppre--ioti of pupal and adult organ- \\ill natnrallv he o|
advantage to the parasite for two reasons:

1. \onii-hment is not required for building up the-e ti— ue-
and then lore more \\ill he available in the \»»\\ cavity o|
the ho-t.

2. The maturity of the host will be deferred ilui- ^ivin;^ the
para-ite a longer life, should it require extra time for develop-
ment. In mo-t of the observed cases, ho\\e\er. the \\orm killed
it- ho-i, h\ emerging before or at about the -ame time that the
uninle-ted latxa- \\ere pupating.

A -imilar though le-- marked case of metal heieK due to para-
^iti-m h\ Mt-ntii* ha- been described and h.^iind h\ Mia/ek 'OM
in the queen- of a Kniopean ant (Lasius alienus I. In ihi- case the
para-iti/ed lar\a- matured and produced adult ant- \\hirh were
noiniil in all external characters with tl:< piioii of a -real

reduction in the -i/e of the wings. Through the kindne-s of
I'roU-— or Wheeler 1 ha\e been able to examine some -imilarly
para-ili/cd -|)ecimen- of a closely relat.-d American SpCClCS
/.,;.v//(.v ;/<v)?//^cn in order to see whetlu-r the de\ <-lo|unent of the

31$ E. H. STRICKLAND.

legs had been in any way affected but a careful comparison of the
measurements of the legs of the parasitized ants with those of
healthy specimens failed to reveal any inhibition of theirdevelop-
ment on account of the Mermis, although the wings, which in
normal ants measured some 10-11 mm. in length were reduced
in the parasitized individuals to 6-6.5 nun.

In the case of Sinnilium larvae, as before stated, the develop-
ment of the legs also is inhibited by the presence of Mermis,
though comparative measurements of the wings and leg histo-
blasts in healthy and parasitized larvae show that whereas in the
former case the wing histoblast covers about four times the area
covered by that of the mesothoracic leg, in parasitized larvae
these histoblasts bear a relation to each other in size of about
2.5 : i, showing that the wing histoblast suffered a greater inhibi-
tion in development than did that of the leg.

A further interesting case of Mermis parasitizing ant larvae
was described by Wheeler ('10), in which case the worker larvae
of Pheidole commutata were parasitized, and resulted in the
adults of such larvae not only possessing all the normal " worker '
characters perfectly developed, but owing to excess of feeding on
account of their constant hunger these " worker " larvae developed,
when mature, characters such as the ocelli which arc normally
only found in the sexual ants.

A SPOROZOON PARASITE OF SIMULIUM LARVAE.

While dissecting out worms from a batch of larva taken on
March 30, I chanced to cut open one larva which from the
whiteness of the abdomen I took to contain a worm, but was
much surprised to find that the body cavity was closely packed
with a white substance which had the appearance of cotton wool.
A little of this substance, however, when smeared on a slide was
seen to be composed of countless organisms as illustrated in
Plate V., Fig. 14. My first impression, very naturally, was that
these were spermatozoa, which they resemble very closely in
general outline. It was, however, very difficult to imagine to
what possible organism these spermatozoa could brlong. The
larva itself could surely not produce them; but it not tin- larva
what then? The only explanation seemed to IK' that they \\nv

SOME PARASITES OF SIMULIUM LARV.l ,

*

formed by one of tin- worms, which could not then be Mcrmis
but must be a new form closely related to Allanioncnm or Sphfc-
rnlaria, in the females of which the uterus becomes protruded
and is finally many time- the -ize of the original worm.

I naturally visited the >t ream in order to obtain more specimens
suffering from this disease, only to find that a quantity of oil
which had been flowing down tlu--tn-am for someday- pa-t had
succeeded in killing off all the larva,- in that neighborhood. It
wa.s then-fore necessary to find a place further up the -m-am,
aborr the contamination, where the larva- were li\inu. About
1 1. ilf a mile'- \\alk led to such a place, -i mated under a -tone arch,
in which the larva- were present literally in thoii-md-. They
forim-d greal masses covering the whole -nrface of the rock-. An
•ninaiion of a few rocks soon showed that the .\[<rniis \\ a- \ cry
plentiful lii-re, but it was some time before I found -pecimen-
\\ith 'In- ctiriou- white abdomens for which I \\a- -earchin^. I
(haii-i-d. h"\M\er, to pull up a piece of water weed that ua-
llo.it in^ in a -u if 1 1\ running swirl and here I found quite a number
of individual-, -miie showing immense abdominal swelling- dm-
to the parasite. On examining a few small specimens I found thai
the para-iie was more numerous among them than annui- the
lai-.i i indi\ iduals; a closer examination of the rock* showed that
the-e -mall parasitized individuals were quite commonly scattered
amon- the larger healthy ones. A quantity of material \\a->
taken back to the laboratory to be studied on the -anie line- as
that adopted for the .UVrw/s parasite.

K\u KNAI. EFFECTS ON THE L.\RV\.

The iir-t effect IK 't iced in badly infested larva- i- the immen-ely
distended abdomen. In some cases as in that illn-trated in
I'lati- \ .. I Ig. u. i he a]«i-x of this region of the body was thn-e
to four time- it- normal size. Unlike the .\fcnnis, thi> ])ara<it«-
is not contin.-d to the \entral portion of the bod\- but entirely
surrounds the caudal extremity of the me-eiiteron. and -mall
deiached colonies were not infrequent in the thoracic re-ion.
I'he Malpi^hian tubule-, however, can usually be seen on the.
surface of the ma— and are plainly vi-ible through the tightly
Stretched skin, \\hidi is (|uiie transparent in place- and appears
to be on the point of bur-ling.

E. H. STRICKLAND. .

The length of the larvae next claims attention. No larva con-
taining this parasite was found to be exceptionally large ; whereas,
as before mentioned, many were extremely small, measuring some
2-75 or 3-5 mm- at a time when all normal larvae measured some
9 mm. or more. This may possibly require a similar, though
opposite, explanation to that suggested on page 281 to account
for the large size of the individuals affected by the Mermis.

In confinement these small specimens were extremely restless,
they were continually twisting about and moving over the sides
and bottom of the jar with a peculiar jerky movement. The
larger, and more heavily parasitized, larvae on the other hand
were more sluggish though nol more so than the healthy larvae.

The effect of these parasites on the histoblasts is very hard to
state with any certainty. Fig. 13 is an illustration, made with
a camera lucida, of the histoblasts of a nearly mature individual.
In this case the parasite was confined to the abdomen. It will
be seen that the discs are but half developed (compare with Fig.
6). In other cases, however, the development of the discs was
not, so far as could be seen, affected in any way till pigmentation
of the respiratory filaments commenced. Then it was noticed that
the entire histoblast only turned a slate gray color instead of
blackening at tin- apex of the filaments and finally over the
complete disc. In other cases, however, the discs entirely
blackened in a perfectly normal manner.

\Yhen colonies of parasites are located in the thorax, as is not
infrequently the case, the histoblasts are materially decreased
in size, while in the very small specimens development appeared
to have been arrested. This is hardly surprising. My general
conclusions were that the parasite affected the histoblasts but
little, though the larvae rarely advanced so far toward maturity
that the respiratory filaments became pigmented. It should be
noted that the power of spinning silk was in no way affected by
the presence of this parasite.

NATURE OF THE PARASITE.

While visiting other localities I made numerous observations
on the larvae to be found in the streams, and was much surprised
to find that species of the parasite were extremely common and

SOME PARASITES OF SIMULIUM LARY.V. 321

that in almost every brook visited some of these conspicuously-
distorted individuals were to be found. On making micro^cop-
ical examinations I observed that the organism- taken from dif-
ferent localities varied very much in form and apparently
represented different species. A list of these different form- and
the'r hosts is appended:

1. Ovoid bodie- bearing a "flagellum "-like organ varying in
length from about that of the "body" to three lime- it- K-ngth
(Plate V., Fig. 14 . Ho-t : ShnnUinn hirtipcs. Habitat: Kotv-r
Hill- and Him- Hills, Mass.

2. Bi-annulated ovoid bodie- bearim; a " Ma^ellum," which i-
rn-ver much longer than the "body" (Plate V., Kiu. 15). H<>-t :
Simtt/tttm species undescribed. Habitat: Stony Un>ok Reserva-

t'on. Mass.

.v <>\()jd bodies ha\'ing the "flagellum" replaced by a trail —
pan in flattened disc (Plate \'., Fig. !<• Host: Sinndium
species undescribed. Habitat: Stony Brook K< •-« t \ ation. Ma--.

4. simple o\oid bodies destitute of all appendage- Mate \ '..
17). Ho-i : Simnlium species undcscriUd. Habitat : Stou\
Bi I. l\- < : \ation and Blue Hills, Mass.

Tht !.. Mowing characters were common to all of tin-. <>i-.m-
i - 1 11 - :

1. Thr\ were all similar in size.

2. I'mlrr i he highest power of the mi> ipe thc\ had a faint
oli\ • i-h tinge.

\\hcii in a IM -h condition absolutely n» internal <li tail- \\ .
\i-iblc an<l in -| >n imens which had been killed, fivd and -tainrd
verj little more could be seen. In many of the first or "sperma-
to/oid" t\|>«- a tlaikriied central area iudicati-d the presence
of a verj large iimlcii-, while a smaller dark -pot jn-t bcfi-rr the
ba-e of the lla'^flltini might be taken for the mieleulu-.

Many of the " -i m pie ovoid " type, although but -lightly -tain-
ing, -Imueil a m'eat -hrinkage of the internal -ub-taiiee: oiher-
\\i-e nothing imiM be -een in them. The -peeimeii- \\ith a
flattened di-e. and ih.>-e bearing t\\o annuli and a lla^elluni,
al-n -ln>\\ed praetieallv no differentiation. Melh\l green and
\arioii- h.emato\\ lin i < -mbination- \\ere tried without success.

He-idi - tlu-M- \ari(.u- fi irms, no two of which \\ire ob-cr\ed

322 E. H. STRICKLAND.

to occur in the same individual, another form of cell (Plate Y.,
Fig. 1 8) was found in much smaller numbers, but possibly in some
way connected with them, for it was seen in association with
each form, but was not found in healthy larva?. This cell, which
varied much in diameter from but little more than the numerous
"spores" to three or four times the length of their longest axis,
was apparently globular in form, and in a fresh state was very
transparent and could only be discerned with difficulty. The
following characters, however, were seen. The substance of the
cell was finely granular, and often contained a number of large
transparent globules. When fixed and stained the only dif-
ferentiation was that of a large dark mass, apparently the nucleus
(Plate V., Figs. 18, a and b). In some cases these cells seemed to
be dividing (Plate V., Fig. 18, c}.

In the face of all these diverse types of cell, which live in
precisely the same way, it is evident that the forms first found
are not spermatozoa, and further the fact that the flagellum is
replaced in one "species" by a flattened disc eliminates the
possibility of their being Flagellates. It is therefore probable that
one must look to the Microsporidia among the Sporozoa as the
group to which these bodies belong. It is further noticed that
many of the forms are very similar to those met with in the
genus Ghigea to which the well known pebrine disease (G. bomby-
cis) of silk-worms belongs. One must therefore conclude that
it is a pebrine-like disease, which is killing off a high percentage
of the Simuliid larvie in the neighborhood of Boston, though on
account of the vast difference in structure and mode of life of
the two hosts the life history of the parasite in the SimuUum
larva1 is very unlike that described by Pasteur ('70) and Stempell
('09) in their work on the disease in silk-worms.

LIFE HISTORY OF THE PARASITE IN ITS HOST.
As in the case of Mermis, I know very little about this, since
I did not discover ii (ill the final stages of the larva were being
approached, and in every instance but one ii was apparently in
exactly the same condition, namely, the "spores" were all
formed and were simply awaiting the death of their host and its
resulting decomposition to escape into the water.

SOME PARASITES OF SIMULIUM LARV.E.

The one exception, however, was that of a larva found on
April 17. It was one of the first discovered, in the abdomen of
which could be seen, with a dissecting lens, large white bodies
floating in the blood plasma. On dis-rrtion the-e bodic- proved
to be rounded masses of flagellate spores, t few of which had
become di-'-niMjed and were apparently movin- about by their
own impetus in the blood plasma. A-, ho\\(-\ vr. I ha\ e ne\ er «ni
any -ub-e<|nem occasion ob-erved MIC:; a movement amon- the-e
spon - I .1111 inclined t» believe thai thi- was -implv a Bn>\\ man
movement, \\hieh I ha\e di-tinctlv re. o-m'/ed in later examina-
tion-,. Tliouuh I -••arehed carefully for other lar\ a- \\ ith para-ile-
in a similar -lage I was unsuccessful.

In -.-rial -e< lions of parasitized larva- similar effects to th.
exhibiied \\hen Mcrmis is present, are noticed in that the fat-
bo.lv i^ much redmvd (Plate I\".. Fig. IO) and the -exiial organs

i onld not be detected, whereas the spinni nd- and mu-cu-

la i tin- a|i]«ai-«-nil\- remained unaffected. The pata-ite pi-obabk-
enters ihe ho-t through the alimentary tract for in all cases of
infected larvae i I \\.is found that the mesenteron was di-ioi-te<| in

one or more places showing distinct hypertrophv as if tin- celU had
be< M !'.nll>- initati-d but had healed over again often pcrmitiin-
the -pores to pass through into the body cavity Plate IV., I
io. , In one case a small colony of "spores" \\ as found in-ide the
alimeniaiA tract . but it is <|iiite possible that the-.- h.,d been
n;l\ taken in with tin- water, as parasiti/e.l !ar\.e \\ere
constantly dxini; in the colony, and an\- material, tloatin^ in
the water -uch a- liberated "spores," would be caiijit by the
i ( phalic I, in- oi still li\ing larva?.

In all recorded cases of Microsporidia parasites it ha- been
noticed that the "-pores" live, at least during their early -t.i.
in-i<le -ome l.od\ ti--ue of the host. An example of thi- j~, \,t
be !'< mud in pel >rine < .f -ilk-worm larva.1, in which case I he -pinnin-
gland- are the main seat of attack. It mn-t be borne in mind,
houe\ei, that an attack upon these organs in the larva now
under consideration \\oiild, in all probability, iv-nlt in the early
death of the host, for it is only by means of the much -tren-t hen<-d
silk thread- that the lar\ a i- enabled to maintain in rapid strcain-
the perpendicular po-ition \\hich is essential to obtaining food.

3^4 E. H. STRICKLAND.

while at the same time the larva has to depend to a large extent
on these threads for retaining any foothold on tin- rocks, for when
moving its position, the adhesive disc of the proleg frequently
loses its grip and the larva is \vashed clear of the rock. Very
rarely, however, the anchoring threads, which are always present,
break so that the larva is able slowly to draw itself once more to
its support. Sections of the Simulium larva disclose the fact that
there are very few other tissues in the body. The muscular
system is much reduced, and the only tissues available for attack
without rapidly killing the host are the fat-body and the sexual
organs, and I am under the impression that it is the latter which
are usually the original seat of attack. In frontal -sections of a
very young larva taken during March, only one testis could be
found, but on the other side of the body a small mass of minute
cells, taken at the time for small cenocytes, was situated a little
back of the normal position of the missing testis. The cells are
too small to show any structures but it is possible that this is a
diseased testis to which the spores made their way as soon as
they entered the body cavity. Later sections also show traces
of a very thin membrane investing the mass of "spores."

I kept a large number of diseased larvae in running water,
hoping to see some of them pupate, but in every case they died,
and soon liberated the spores, whereas many of the healthy
larva1, which were approaching maturity, pupated in captivity.
It is thus evident that heavily parasitized larvae never pupate, but
die in the stream, liberating their countless spores in the water.
\Yhat happens to these spores I have been unable to ascertain.
Larvae were allowed to die in distilled water and the liberated
organisms were examined at intervals, but though they strongly
resisted decomposition they never showed signs of movement or
altered condition. Sections of pupse and adults obtained from
badly infested localities failed to reveal any cases of the disease
being carried beyond the larval stage. It is therefore probable
that the disease is not hereditary as is the case with pebrine,
and as before stated it was seen that the presence of this parasite,
in every case examined, apparently resulted in castration of t In-
best. It is, however, possible that among the vast numbers of
larva' a few were only slightly parasitized and did not have tlu-ir

SOME PARASITES OF SIMULIUM LARXVE.

sexual organs entirely destroyed. Such individuals should be
capable of pupation and might in that way carry the disease over
as in the case of pebrine. An examination of several hundreds of
larvae did not reveal one in which such a condition was possible.
XVere there overlapping generations of larvae the maintenance
of the parasite could be more easily under-tood. but the spring
brood pupated and hatched during the first half of May. and
Him- an- noi at ilu- time of writing any signs of more eggs bein-
depo-it.-d on the rocks. It thus seems that there imi-t be a
•ndary hosl in \\hich this parasite passes tin- summer.1

Tin-: PERCENTAGE OF INFESTED I.AKV i

The number of infested larvae varied to a great «-\tent in
ditlt ifin streams. In the part of the stream in Foiv-t Hills
\\li.-n- the para-ite-, were first noticed, les- than i per reni. of
the lar\.e \\ t <-iti/ed, half a mile further up the Mivam a

little under 40 PIT cent, were infested, while in the Stony Brook
l\t -i-r\ aiion \\lnTi- two types of "spores" \\nv pn -nit among
tin- par s-iii/i-d larva', between 70 and 80 per cent, of ihe in-
di\idual- \\ere to be seen with the immensely di-ti-ndrd and
\\hiiened abdomens which proclaimed tln-m to be -nth ••

11 ihi- I'at.il disease.

h \\ill ilui> be seen that should this di^. Aether \\iih the

previously di--i ribcd Mcrmis parasite, pro\e to .-\i-t a- abun-
daniK in other localities as it has during the pa-t -prinu in the
\itiiiii\ oi Bo-ion it must be of considerable im|)ortaiife in the
natural .ontrol of the black Hies which an- -neh an amioyaiiee
to man aiid bea-t, especially in more tn.pifal regions, and it' the
Mippo-i-d secondary host of the Sporoxoon doe-, not \>\-»\ e to be a

li-h or animal "t an\ \ alue it should be po>-ible to inter t -tr<-am>
\\lu-re simnliid lar\a- breed, and diminish their number- verj
lai.t-K \\itliout the danger of poisoning the li^h b\ apphin^
oil or other Mil»Miires to the water.

SUMMARY.

.^iniidinni lar\a-. which are found in \ a>t number- in small
rapid streams around Boston, are seen to feed by standing per-

1 I II.IM- >ino- July i\ imtiri-il isolated specimens of a different species of larva
in OIK- >>! tin- >ti«Miii- u In n- tin- p.irasite abounded in the sjinng brood, but have
not tomi'l any in uhii.li tlu-ro \\ ^ of the para-itc In-iiiK present.

326 E. H. STRICKLAND.

pendicularly on rocks to which they are attached by a strong anal
adhesive disc, and kept in position by silken threads secreted by
the salivary glands. While thus anchored they spread out a pair
of cephalic fans which act as strainers and collect small particles of
food from the water. The head capsule is moulted independently
of the body cuticle and exposes a new capsule which is at first
white with a few dark spots on the vertex, but which rapidly
becomes uniformly darkened all over. The thorax bears un-
usually well defined and large histoblasts of the imaginal wings,
halteres and legs, and also on either side a histoblast of the
pupal respiratory filaments, which by turning black when the
larva is mature becomes very conspicuous at this stage of growth.
The larvae are infested by two parasites, namely a Mermis and a
Sporozoon, both of which live in the body cavity.

The Mermis does not affect the larval development to any
extent, except by slightly increasing its size, but it inhibits the
development of the histoblasts to such an extent that pupation
becomes impossible.

The embryo worms are probably caught by the cephalic fans
of the larvae and pass into the alimentary tract, through the
walls of which they" bore and live in the body cavity of the
host till the latter matures. They then rupture the abdominal
cuticle and pass into the water where they live a free life under
stones in the bed of the stream. The number of worms con-
tained by a single larva is usually only one, but as many as
twelve have been found. A single worm measures 3 cm., which
is about three times the length of tin- ho>l. In some streams
25 per cent, of the larvae were infested with this parasite. Para-
sitized larvae never pupate, bin are killed by the worms when they
escape.

The retardation in the development of the histoblasts is the
opposite condition to that met with in prothetely which is usually
caused by keeping larvae at an abnormally high temeprature.
This probably results in an increased supply of the cnzyjnes
which cause these histoblasts to develop. The Mermis ap-
parently excretes some substance which lessens the supply or
action of these enzymes and leads to metathetely.

The Sporozoon parasite occurs in several forms in different

SOME PARASITES OF SIMULIUM LARY/E.

localities. All these forms, however, live in the same way and
appear to be related to the pebrine disease of Lepidoptera. Tin-
body, especially near the apex of the abdomen, becomes much
distorted and swollen on account of its interior beinu closely
packed with a white wooly material which on di—ection is seen
to consist of count less "spores" of minute si/e. Such parasitized
larvae are usually rather smaller than healthy individuals, but
tin- hi-.tol.la~t- do not appear to be much affected. The para-ite
apparently enter-, the 1,. >dy cavity in the -ame manner as that
dt--cril>ed in ill'- case of Mcrmis. K\ idemv of thi- i- seen in a
h\ pi-rirophied condition of part- of tin- mesenteric \\all. From
h.-n- it seems to pass to one or both of the -exnal organs \\hich
.ire d'--t ro\ i-< 1 .mi 1 IM-. i niie the nuclei tor the great ma-- ot -pore-.
\\hich eventually fills the abdomen. The para-iti/ed larvae in
tin- al-o \\i-re never observed to pupate but died \\lu-n

mature. The -pores are liberated by a rupture of the abdominal
\\all -oi.ii .itter the death of the host and pa-- into the \\ater.
after \\hicli stage ih'-y have not been -ecu. l"p to >o j.i-r cent.
of the laiA.e iii -oine -iream- were found to contain lar^e ma-
o| tin- p.ira-ite but no cases of slightly para-iti/eil lar\.e were
ob-er\ei|. There has been no -econd brood of Simulium larva-
tin- year, -M> it would seem that if the par.i-iie i- t.. ap|u-ar next
\ear then- must be a sccondar\- h<i>t in \\hich the -umniei i-

ed.

POSTSCRIPT.

The foregoing account of the Spon./oid ]>ara-iti- o| Simulium
hirti(>f> \\a- \ cry kindly reviewed for me by Professor G. V

Calkin-. ..I I "olumbia I'niversity. 1-roin mounted -|.ecimen- of

the -p. -it him he confirmed my opinion thai ihe-e represent

some species oi M\ \<-|)oridea, and drew my at tent ion to a paper
by I.oiii- I ; . \\hich 1 had o\cilo,,ked because of it- not

beiii- catalogued under Simulium in the cards of the " Concilium

Bibliographicum " in the C'ambrid^e librar\ . In thi- papt-r a
ne\\ -P.-MC- of Cilii'^fti parasitizing the lar\ae of the luin.|»ean
Sininlinni onuitnni i- <K--i ribeil i :ritins. The note- on thi>

-|>ecic- are in brief a- follow-: The abdominal r.-i.,n of the
in!\--ted lar\a i- greatl) di.-ti-ndi-il ami contain- lar^c mass<
a free |).ira-ite in the form of opaque. milk\' white, irregular -

E. H. STRICKLAND.

Some larvae contain but one mass whereas others contain two
to four. The muscles are unaltered, but the fat body is much
reduced. The alimentary canal always appears to be contorted.
In only one case had the Microsporidea failed to sporulate and
they then formed a swelling on the external intestinal surface.
The sacs contain countless ovoid, refractive spores, which, when
treated with iodine, show a filament 15-20 times as long as their
longest diameter. Spores of two sizes are present, the smaller
measuring 4-5/11, the larger 8/j.. In a subsequent note written
in collaboration with Hagenmueller ('08) Leger states that the
spores are sometimes present in a polysporic and sometimes in
an octosporic arrangement. These authors also refer to a simi-
lar parasite in the larvae of Tipula gigantea.

From the foregoing notes it will be seen that the disease which
occurs in S. hirtipes is very similar in its main features to that
described by Leger, and I do not hesitate to regard the organism
responsible for its occurrence as a closely related form. Pro-
fessor Calkins is inclined to consider the various forms I have
described as belonging to a single species. For this I would
propose the name Glugea polymorpha sp. nov. Future investi-
gation, however, will quite possibly show that the various forms
occurring in different localities are not all representatives of the
same species, since numerous dissections of diseased larvae
showed that certain types of spores were peculiar to different
localities even though present in two different species of Simidiiim
larvae.

SOME PARASITES OF SIMULIUM LARY.E. 329

BIBLIOGRAPHY.
Dewitz.

'05 Untersuchungen iiber die Venvandlung der Insectenlarven. II. Archiv

ftlr Anatomic und Physiologic, 1905, pp. 389-415.
Fischer.

'06 Uber die Ursachen der Disposition und iiber Friihsymptome der Raupen-
krankheiten. Biologisches Centralblatt. Bd. XXVI.. Xr. 13, 14, 15, 16.
Hagen, H. A.

'72 Stettin, ent. Ztg., pp. 392-393-
Heymons, R.

'96 Sitzungsber. d. Ges. nat. Fr. Berlin.
Kahle, W.

'08 Die I'aedogene-i- .l.-r ri-ri.lmnyiden. Zoologica. Heft ?=;.
Kolbe, H. J.

'03 l"b«T ••!!(• Kntwickeluni; I'r.«tln-t'-!ie . Allgem. Zeitsch. t'ur Knt.

Bull., - [i. 28.

Leger, L.

'97 (,l: < K

Leger, L. and Hagenmuller.

'08 K .iiiiin.itix .1'. < K. \-~.

Mr.'izck, Al.

'08 M unky. 111. •. \ 1. IV.,

pp. i vj-i.\'>. -1 '
Pasteur.

'70
Riley, Wm. A.

'08 1 i. : ni.il A; • I Inlmi),

'
Stcmpcll. W.

'09 i i.. i t., XVI., p]

Wheeler, W. M.

'07 '!!-• r -A lib .in A'-i'iMir.: ! Singular Abnormalltlea

diii- 1" I'.ii.i-i:i-:ii. li'ill. Am. N.it. Hi-t., \'nl. XXIII.. pp. i .;;,

jil. I \ I.

'10 I "ln-r Ki;. 'ion in ! Journ.

\ III., p,

330 E. H. STRICKLAND.

EXPLANATION OF PLATE I.

FIG. i. Prothoracic leg of a Simuliid larva showing the histoblast (a) at an
early stage of development.

FIG. 2, a, band c. Types of spotting on the head of a recently moulted larva
of Simiiliiim hirtipes.

FIG. 3. A single rake from the cephalic fans.

FIG. 4. Profile of thorax of a half-matured larva showing the histoblasts.
a, respiratory organ histoblast (pupal organ); b, wing histoblast (adult organ);
c, halterer histoblast (adult organ) ; d, e and /, pro-, meso- and metathoracic leg
histoblasts (adult organs).

FIG. 5. Ventral view of a Simidium larva with Mermis parasites in situ.

BIOLOGICAL BULLETIN, VOL. XXI

PLATE I

t. H. STRICKLAND

332 Ft. H. STRICKLAND.

EXPLANATION OF PLATE II.

FIG. 6. Histoblasts of a healthy full-grown larva measuring 10.5 mm.

FIG. 7. Histoblasts of a larva parasitized by Mermis measuring 10.5 mm.

BIOLOGICAL BULLETIN, VOL. XXI.

PLATE II

0

0

0

E. H. STRICKLAND.

334 E. H. STRICKLAND.

EXPLANATION OF PLATE III.

FIG. 8a. Average size full-grown parasitized larva of Simulium hirtipes meas-
uring II mm.

FIG. 8b. Average size mature larva of Simulium hirtipes, measuring 8.5 mm.
showing darkened pupal histbblast, a.

FIG. ii. Median sagittal section of a half-grown healthy larva to show the
alimentary tract, b, the normal position of the spinning glands, which extend
backwards from the pharynx on either side latero-ventrally to the alimentary
canal, doubling back on themselves at about the point b,' where one has been cut
in cross section; c, the normal quantity of fat body which increases still more as
maturity approaches; d, one of the pharyngeal glands. (This is not seen in an
exact median section, as the two glands are narrowly separated medially.)

BIOLOGICAL BUILETIN, VOl . XXI

-t.O

E. H. STRICKLAND.

336 E. H STRICKLAND.

EXPLANATION OF PLATE IV.

FIG. 9. Median sagittal section of a larva parasitized with Mermis sp. to
show the somewhat displaced alimentary tract, and a, the coiled up worm; b, the
displaced spinning gland, a short longitudinal section of which has been cut; c,
the much reduced fat-body; d, one of the pharyngeal glands.

FIG. 10. Median sagittal section of a larva parasitized with a sporozoon to
show a, the mass of spores, a few of which are shown enlarged; b, the spinning gland;
c, the much reduced fat body; d, one of the pharyngeal glands, and e, the some-
what distorted wall of the alimentary tract.

BIOLOGICAL BULLETIN, VOL. XXI

PLATr IV

10

I.

.0-4

l

I -.a

E. H STRICKLAiD

E. H. STRICKLAND.

EXPLANATION OF PLATE V.

FIG. 12. Dorsal view of a Simttliitm larva with Sporozoid parasites in situ.
Compare with Plate I., Fig. 5, which is normal in shape.

FIG. 13. Histoblasts of a larva parasitized with the Sporo:oa measuring 9 mm.
Compare with Plate II., Fig. 6.

FIG. 14, .4, B and C. Simple "flagellate" type of parasite, X4,ooo.

FIG. 15, A and B. Annulate "flagellate" type of parasite, X4,ooo.

FIG. 16. Type of parasite with "flagellum" replaced by a flattened disc,
A showing surface view of disc, X4,ooo; B showing side view of disc, X4,ooo.

FIG. i", .4, B, C and D. Simple ovoid type of parasite, X4,ooo.

FIG. 18, .4 , B and C. Other bodies found in small numbers among the "spores,"
X4.0OO. Many are larger in proportion than those illustrated. C shows one
apparently dividing.

BIOLOGICAL BULLETIN, VOL. XXI

12

E. H STRICKLAND

\\>l. XXI. November, iyn. Xo. 6

mOLOGICAL BULLETIN

THE PERSONAL EQUATION" IX BREEDING EXPERI-
MENTS INVOLVING CERTAIN CHARACTERS OF

MAIZE.1

RAYMOND PKARL.

Iii the summer of 1908 some experiment-, in the i TO-- -breeding
iin types of maize were begun 1>\ the \\ riter .UK I hi- former
colleague, I )r. Erank M. Surface. Tin- pve-ent paper ha- tn do
\\ith a part of the results obtained by crossing a \\hitr -\\eet
\atieiy (c? parent) with a yellow dent \arirtv (9 parent'.
Hoth \anVties used were "pure," in tin- -en-e that each bred
true to the general type to which it belonged. The hi-tor\ of the
sweet variety used has been detailed in another place' and in . d
not be repeated here. The important tiling tn In- iMinl at thi-
time is that in its whole historv thi- -ui.-t ((.in n-<d in tin-
cross breeding experiments had never been known t<> prudinr
anv but su'eet (sugary) kernels of an exceptional de^n-e o|
whiteness*

The dent corn used in the experiment^ ua- al~o of knoun
lii-toi'\ . A discussion of its hi.-t'>r\. and of the characteristics
of the corn has been given elsewhere.4 The essential point tobe
noted here is that during a long period d years ii ha- ne\ er
ced anything except starchy kernel- ot a d(i p
when ri|)C.
I'.ipi-rs from the Biological Laboi. nluir.il l-;\|i.-iinnm

St.lt I. ill. \. i. 2Q.

I ''iil. R.. and Surface, F. M., "I Corn." Me.

•it. Stat. Ann. Report fur tgio, ;

A pure chalky white or. put in tin- mlit-i uay. tli.- (.ntiro .ili-ciici- »\ yellow
color, i- .in absolute essential of a lii.yh Ki.nii- . .1" mm fnun tin- P.I. k. r'~ -tamlpnint.
Ilu irn here under discussion is regarded by expert ;inn-

ally tiii<- >tiair, for their purpose.

1 iv.ul. K.. "The Mendelian Inhcritan. c ••!' Ortain Invi-ilil«.- ("ln-n.
chr. f. Ahst.- u. \'crcrb.-'.< • I'. \ 1 ;ii.

34O RAYMOND PEARL.

The general results which follow the crossing «\ a yellow dent
(9 ) with a white sweet (cf) maize are well known, Yellowness
of endosperm is dominant over "whiteness" of endosperm, and
"starchiness" over "sweetness." Consequently the Ft kernrk
are externally indistinguishable (in fact as well as in theory)
from those of the pure yellow dent parent. These FI kernel-
planted give rise to plants bearing ears of which each should
have four distinct kinds of F* kernels which ought, by theory,
to occur in the simple dihybrid ratio, 9 yellow dent , 3 white dent,
3 yellow sweet, I white sweet.

The present experiments1 entirely confirm in all essential re-
spects this general Mendelian result. Certain novel points
arose, however, in the course of the work, which led to the present
investigation. These points may now be considered.

A large quantity of ears bearing F2 kernels was raised. These
ears were well matured. This was indicated both by their
appearance and by the way the seed from them germinated.
One of the assistants in the laboratory, Miss Maynie R. Curtis,
undertook the sorting and counting of these p2 kernels on an
extensive scale. In this work the following situation immediately
developed and was called to the writer's attention. While in
general the F2 kernels fell without any doubt or difficulty into
the four classes or categories, yellow starchy, white starchy,
yellow sweet and white sweet, yet there were a number of kernels
on each ear that were extremely difficult of classification. These
kernels were, in short, intermediate in respect to their external
visible somatic characters, and might, in the individual case,
be put with equal propriety into either of two classes. Into
which class such an intermediate kernel would actually be put
plainly depended upon the personal bias of the observer, rather
than upon any peculiarity of the kernel itself. This result ap-
peared to be of enough interest and potential significance to
warrant a more extended and thorough investigation of the
matter. The present paper deals with tin- results of such a
study.

1 A detailed description of the conditions and manner of these experiments has
been given elsewhere (Pearl, he. cit.) and need not be repeated.

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 54 '

-I.VII-.MENT OF PROBLEMS AND PLAN OF INVESTIGATION.
The problems with which this work is concerned may be
.-ummarily stated as follo\\ - :

1. To what extent is the personal equation of the observer a
lificant factor in the Mendelian ratios described for simple
• •riments with cross-bred maize? In other word-, h«>\v closely

Mould the different individuals of a group of competent biological
ob-ervers agree in their classification ami count of the same V»
in. nrrial from a maize cross involving -uch rclati'.ely -imple and
e.i-il\ judged unit characters as color ot" endo-perm, or chemico
phy-ical character of endosperm (starchy »r swe<

2. I )oes somatic "intermediateness" in mai/e imply garnet ic
"intermediateness"? In other word-, do F_. kernel* \\hich are
inn rmediute snmatically give rise to any diffen-nt •-on of progeny
\vheii pi. uitcd than do kernels which belo, \\\ and indubi-
i.iliU n> one or another of the well-defined :>imcti(- d '">\ \ .'.
If ihe\ are true "blends" in the (ialtoniaii . the\ \\ould

ainly be expected so to do. If, ho\\ever, they merely re;
sent a phenomenon essentially like the incomplete <>r p.mial
(somatic) dominance so frequently oh>«-r\ei| in Mendelian work,
it would be expected that their progeny would dill'er in im essen-
tial particular from that obtained from ~»ni,uii .i!l\ nun-inter-
mediate kernels having the same gametic c<m-titution.

To test these questions the following plan wa- di \ i-ed : I'mir
«.ii- ln-.iring p2 kernels were taken quite at random from a lot
ot about two bushels of such ears, which in turn was a random
-ample of a whole crop which included a much larger number of
bu-liels. Each of these four ears was given an arbitrary number
and was separately shelled, great care bein- taken to see that
im kernel* were lost. All the kernels from each ear were pre-
. i -d t' Aether in a bag (or box). The -hell in- was done in t h--
\\riter\ laboratory in the presence ol -e\eral \\orker-. so thai
there can be no question whatsoever, that all of the kernel- in
each one of the four parcels originally a< \\ upon the -ame ear.

Three of the ears so dealt \\ith No uid 101 \\eienormal

in everj re-pect. Kar No. 11 was -li^htl\- abnormal in the
re-pert that a fungus had attacked -ome of the urain-. ^i\in^
them a -li.uht pinki>h tinge in addition to their o\\ n propi-r

342 RAYMOND PEARL.

rcolor. This was especially noticeable in the case of the "white "
sweet grains of the ear, because in a mature, dry sweet corn
kernel " white " means merely the absence of any color (yellow or
other). The grain is translucent and "not colored." Any
extraneous color such as that arising from a fungus attack will
be the more evident. The same considerations apply to the
white starchy kernels, except that here the starch of the endo-
sperm gives the grain a positive white color.

The kernels from each of the four ears having been separately
shelled and preserved as described, fifteen persons (including the
writer) were asked to sort the kernels of each ear into the four
categories, yellow starchy, white starchy, yellow sweet and white
sweet, and then count and record (on blanks provided for the
purpose) the number of each sort found. Four small vials con-
taining typical kernels of each sort were given to each observer as
comparison samples. The only instructions given the observers
were :

1. To sort and count the material independently.

2. To open and handle only one parcel of seed at a time.

3. Not to lose a kernel.

4. To count correctly, i. e., to make sure that the total numbers
of kernels, counted tallied with the total numbers in the parcel,
which numbers were set down on the blank for each ear.

Especial pains was taken to insure that no observer (with the ex-
ception of Nos. VI., \ II., \ III. and XI.) should know, in advance
of his count, the nature of the experiments which gave origin tot he
material, or the expected Mendelian ratio between the several
classes of kernels. No observer1 was, of course, allowed to see
the results of the counts by others until after his own had been
completed. In short every effort was made to insure in all
possible ways that the counts tabled should be the unprejudiced,
unbiased, independent and purely objective statements of the
opinions of a group of competent biological observers as to the
proper classification of the F2 kernels from these four ears of maize.

\Ye may next consider the observers who took part in this
work. At the outstart tin- writer wishes to express his indebted-

1 With the single exception of No. XI., and in this case it was some months
later that his own counts were made.

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 545

340

310
295

YELLOW STARCHY

65

45
105

YELLOW SWEET

75

50
80

WHITE STARCHY

I I I I I

50

20

WHITE SWEET

I II III IV V VI VII VIII IX X XI XII XIII XIV XV
FIG. i. -i showing tin- ouint of the ditt'-rriit olwervi : the

lour classes »i knn.-l- tor ear No. 8.

344

RAYMOND PEARL.

ness to all of those who cooperated in the investigation, and his
appreciation of the painstaking interest and care given to the
sorting and counting by all. Table I. gives the name, academic
degree and official position of each of the cooperating indi-
viduals. For convenience of reference in the paper each observer
has been assigned a Roman numeral.

TABLE I.

LIST OF OBSERVERS COOPERATING IN THE PRESENT STUDY.

No.

Name.

Academic
Degree.

Official Position.

I.

W. J. Morse.

M.S.

Plant pathologist, Maine Experiment

Station.

II.

C. E. Lewis.

Ph.D.

Associate plant pathologist, Maine Ex-

periment Station.

III.

G. E. Simmons.

M.S.

Professor of agronomy. University of

Maine.

IV.

M. E. Sherwin.

M.S.

Assistant professor of agronomy, North

Carolina College of Agriculture.

V.

Wallace Craig.

Ph.D.

Professor of philosophy. University of

Maine.

VI. Raymond Pearl.

Ph.D.

Biologist, Maine Experiment Station.

VII. Frank M. Surface.

Ph.D.

Biologist, Kentucky Experiment Station.1

VIII.

Maynie R. Curtis.

M.A.

Assistant in biology, Maine Experiment

Station.

IX.

Lottie E. McPheters. Computer, Maine Experiment Station.

X.

Frank Pearl.

Farmer and practical corn breeder.

XI.

W. Johannsen.

M.D.

Professor of plant physiology, University

of Copenhagen.

XII.

P. Boysen Jensen.

Ph.D. Instructor in plant physiology, 1 niviT-ity

of Copenhagen.

XIII. Jenny Hempel.

M.Sc.

Assistant in plant physiology. University

of Copenhagen.

XIV. Gerda Dohlmann.

•

Assistant in plant physiology, University

of Copenhagen.

XV. 1 Oilman A. Drew.

Ph.D.

I'rotc — r oi Uinlcigy, 1 niversity ol Maim

Certain points regarding this list of cooperators need to be
discussed. In the first place it is obvious that any one of them
(with the possible exception of X.) might in the ordinary course
of his work carry out a Mendelian experiment with maize, either
independently or in cooperation with someone else. It this were
done and the results published they would certainly be accepted
by the biological public as a precise and true statement of the
facts regarding the material which was in the experimenter's
hands. That is, if any worker in this list published a statement
that a Mendelian experiment which he had conducted with

1 At the time this work was done: Associate Biologist, Maine Experiment
Station.

PERSONAL EQUATION IN BREEDING EXPERIMENTS.

mai/f l«-d to a ratio of, for example-. 7,vi : 24.} : 252 : (>o this
stati-rnrnt would not be doubted or questioned.

In the second place it is worth while to consider the training,
or lines, of work with which the-r 15 observers have had to do.
<M-ix(Xos. I., II.. XL. XII.. XIII.. XIV. ilu- training and work
ha- been primarily botanical. Four of tlu--e the I>ani-h croup.

255

YELLOW STARCHY

225

70
55

85

70

25
10

YKLLOW SWEET

WHITE STARCHY

WHITE SWEET
I I I I

I II III IV V VI VII VIII IX X XI XII XIII XIV XV

:.im showing the count ,of tl;
f"iii rncls for car No. 9.

Nos. \l to Xl\'. inclusive) ha\«- had particularly to do \\iih
tin- data of experimental plant breeding, in c< .nnrrtion \\ith the
brilliant and fundamental re-ran -In •- <>l I'n ilV--or JohaniiM-n.
The training and special field »t \\<>rk of ti\e \»-. \".. \'L. \ II .

\ III. and \\".) of the ob-rr\iT- ha- bn-n : . < M i'

ii\e thvi'i- Nos. VI. . VII. and \ III. ha\ c had experience with

the data and method- o| investigation in e\])erimfiital

;•

RAYMOND PEARL.

TABLE II.

SHOWING THE CLASSIFICATION OF THE KERNELS OF EAR No. 8 BY THE DIFFERENT

OBSERVERS.

Mendelian Expec-

299.25

99-75

99-75

33-*5

399.00

133.00

tation.

I.

352

IO2

52

26

404

128

II.

322

49

82

79

404

128

III.

298

75

108

51

406

126

IV.

332

IOI

71

28

403

129

V.

305

101

86

40

391

141

VI.

3i3

IOO

90

29

403

129

VII.

308

86

95

43

403

129

VIII.

3ii

IOI

92

28

403

129

IX.

327

IOI

78

26

405

127

X.

308

92

95

37

403

129

XL

3"

97

92

32

403

129

XII.

313

99

9i

29

404

128

XIII.

308

97

95

32

403

129

XIV.

3"

104

9i

25

403

129

XV.

333

97

73

29

406

126

Totals.

4.753

1,402

1,291

534

6,044

1-936

Means.

316.87.'

93-47

86.67

35.60 402.93

129.07

Another of the five (No. V.) adds to the special training of the
zoologist that of the philosopher and psychologist, which by
traditional standards, at least, ought to aid in the development
of a discriminative judgment. The training of two of the ob-
servers (Nos. III. and IV.) has been agricultural. Further, both
of these men belong by birth, early life and education to the
"corn belt" section of the country, and arc thoroughly and
intimately familiar with maize. They have had experience in
corn judging, which demands the appreciation of very small
differences in ear characters. Observer No. X., while not a
scientific student of breeding, has had successful practical expe-
rience in corn breeding, and is a careful observer. Observer
No. IX. has been specially trainrd in biometric work in the
writer's laboratory and has had considerable experience in meas-
uring, sorting small variations out of mixed material, and similar

work.

RESULTS.

I.

The results of the counts of the four ears by the different
observers are set forth in Tables II. to V. inclusive. Each oi

I'l.RSONAL EQUATION IN BREEDING KXPF.RIMENTS. 347

250

240

65

45
90

"JO
45

I- -1 • — 1-_ l-^t^

r rfa

YELLOW STARCHY

I 1 I I 1

YELLOW SWEET

WHITE STARCHY

WHITE SWEET

15

X XI XII XIII XIV XV

I- 1'.. ; I>i.i--ram showing the count <>f tl. ..! the-

loin . I.i-M's of kernels for car No. 10.

tlie-e taMes is arranged as follows: Column- are v;i\eii for the
lour dillereiit classes of kernels, \vllo\\ -i.inln. \ello\\ -\\eet,
\\ liite -iarch\ and white sweet. Al-o column- .in- ij\ m tor toi.il
^l.iirhx mil toi.il >weel. The first rou o|' r.u h i.iMt- -Imu- tlir
Mi n<li-!i.m c\|)i-ci.[!ion for each clas-. Tin- lollouin- line- -Imu
t!u- <li-tril>uiiou of the kernels as reporlcil li\ r.ich of tin- tifircn
i il i-(-r\ '

I In- data for the color classes gi\i-n in tin •-(• i.iMc- an- -ho\\ n
-i.iphii all\ in I igs. I 104 inclll>i\c. < »m- of tli:--r diagram- i-
di-\oii-d to r.ich of the four rar- u-rd in tin- -tud\ . Ivach tivuiT
ui\c- the (iloitiug of each ol>-rr\ »-r'- count of the four cla--rs of
krrnr!-. The Mendt-lian rxpcctaiion i> plotted in each ca-e as
a dotted -trai^ht line and the mean of the re-u]t> of the different
oli>er\er- as a >trai^ht line ol da>li'

l-'rom the-e table- .md diagram- \\ e note tin- fol!o\\in^ ]ioint-:

348

RAYMOND PEARL

TABLE II!.

SHOWING THE CLASSIFICATION OF THE KERNELS OF EAR No. 9 BY THE DIFFERENT

OBSERVERS.

Classes of Kernels.

Observer

Yellow

Yellow

White

White 1

Starchy.

Sweet.

Starchy.

Sweet.

Starchy.

Sweet.

Mendelian Expec-
tation.

237-33

79.11

79.11

26.37 .

316.44

io5.4S

I. 234 71

89

28

323 99

II. 247 69

76

30

323 .

99

III. 230 62

93 37 323

99

IV.

249 75 74

24 323 99

V.

227

8 1 84

30 311

III

VI.

242

79 81

20 323

99

VII.

240

75

82 25 322 100

VIII.

241

78

82 21 323

99

IX.

242

80

79 21 321

IOI

X.

238

82

84

18 322

IOO

XI.

245

75

78

24 323

99

XII.

243

77

80

22

323

99

XIII.

244

75

79

24

323

99

XIV.

246

77

79

22

323

99

XV.

246

75

77

24

< '/>

99

Totals.

3.614

1.131

1.215

370

4.829

1.501

Means.

240.93 75.40

81.00

24.67

i-M.g.5

100.07

1. For no one of the ears is there entire agreement among
all the observers as to the number of kernels falling in any one
of the color classes. There is entire agreement among the ob-
servers as to the total number of starchy and sweet kernels in
the case of two ears (Nos. 10 and 1 1), leaving out of account the
loss of one starchy kernel from ear No. 10 between the time
when this ear was counted by observers X. and XI. In the
case of the other two ears (Nos. 8 and 9) there is some disagree-
ment as to the number of starchy and sweet kernels. In no
case, however, is the disagreement in regard to these characters
so marked as that in respect to color characters.

2. The relative amount of divergence among the observers in
regard to the distribution of the kernels in color classes is strik-
ingly different for different cars. Ear No. 9 plainly bore kernels
which were relatively easy to classify. The same was true of
ear No. 10. On the other hand the kernels of ears 8 and II
offered many difficulties in classification. But in the case of
ear No. i i the difficulty was largely confined to the sweet kernel-,
there being close agreement between all the observers but one

PERSONAL EQUATION IN BREEDING EXPERIMENTS.

TABLE IV.

SHOWING THE CLASSIFICATION OF THE KERNELS OF EAR No. 10 BY THE DIFFERENT

OBSERVERS.

eet.

ite

White

-

hy.

M- i ;.ec-

243-00

81.00

81.00

27.00

..-X

108.00

I.

251

51

45

II.

251

53

336

96

III.

248

51

336

96

IV.

260

65

76

31

336

96

V.

245

65

31

336

96

VI.

251

75

85

21

96

VII.

246

61

90

96

\ III.

247

75

21

IX

249

70

247

74

250

73

96

XII

247

71

96

XIII

247

76

88

xr

74

X

78

I.OI2

M. '

249.00

67.47

1

.

\". i) with regard to the yellow and \\hitc -t.uvhy kernel- of
tlii- ear.

.V The cause of the discrepancies 1>« lut.-n tin- i-ount- <>t" the

• r.il ol)servcrs is obvious from the data. It \\ill IM -< en at .1

vj.im c 1 1 mil the diagrams that generally \\ IH-II .111 observer's « omit

of tin- yt'lloic starchy kernels of an f.ir. for example. <li-\i.iti-(l

I'n. in tin- me.m in excess, this same observi mnl of tlir ,.•////«•

-t.m h\ kernels deviated from the inc. in in del'r.i. ,iiul l>y ,tn

.imoiint approximately corresponding tn the pn-iii\e de\ i.ition

in the other rase. In other words, certain kernel-, either starchy

\\eet . \\ hich \\ere called "yellow " \>\ one observer were • .died

"\\ hite" l>y another. This bring^ out in .1 -trikin^ \\.iv \\ li.u was

o|i\ioii- to e.u h observer who handled (hi- m.ii/e, namely, that

there were on each ear a number of both Diarchy and -ueet kerneh

\\hii-h \\ere inierniediate in re-|u-et to eolor. The distribution

-nch kernel- into the Mendel!, m categories <le|>eiid- ujion the

1 Bi-turi-ll tin- tillH' \S 1> ' •'lIHi'.l i

XI. . hv krinel was 1..-: < •:i---|in>iuly the • .111 <.t" all

-t.it. li\ .in.! all - -mall. : X \\'. in. 1

th.in |..r t!i«- .-tlirr ol

350

RAYMOND PEARL.

YELLOW STARCHY

YELLOW SWEET

WHITE STARCHY

WHITE SWEET

10

I II III IV V VI VII VIII IX X XI XII XIII XIV

FIG. 4. Diagram showing the count of the different observers of each of the
four classes of kernels for ear No. n.

personal "equation" or bias of each individual observer. As a
matter of fact it was possible (and this was done) to make a
perfectly graded series of either starchy or sweet kernels from a
single ear which ranged from pure white at one end to pure deep

PERSONAL I.OUATION IN BREEDING EXPEKIM I-NTS. 35

: 210

30

XII VI XIII XI VII VIII IX X V XIV IV III II
PlC. S ' '•'">-' l'": total deviations in all tl.

TAIILK \ '.
SHOWING nu. < i s OF THB KBRNBLS OF EAR No. 11 \.\ i:u

l.KVERS.

• w <>w

!iv Swccl. : . ti\

^ 218.87 76.39

I

II.
Ill
I\
\ .
\ I
\ II
\ III

IX

XI
XII
XIII.
Xl\

•.M-.

392

87

331

53

67

313

75

323

66

76

224

72

82

87

221

64

223

84

221

78

222

79

1,0

7'

7

7»

77

I'

77

76

76

75

75

7i

21

7 7

M

4.1

'-•

o-unt of this ear l>\ N JC\ :>• lti'!''.l in t!.

35 2 RAYMOND PEARL.

yellow at the other end, with each intermediate step practicallx
as small as one cared to make it. An attempt was made to
obtain photographs of such series of kernels which would demon-
strate the fact of this gradation pictorially, but the photographic
resources at command were not equal to the task and it had to
be abandoned.

4. The data presented fully demonstrate, I think, the interest-
ing fact that if each of these fifteen competent, and with one
exception (No. X.), specially trained observers had independently
undertaken an investigation of Mendelian inheritance in maize,
and all used the same seed, of at least the two strains here em-
ployed, grown their crops in the same place, and even studied
identically the same progeny ears, no two would have fully agreed
in the numerical values of the F2 ratios.

II.

Let us now consider the question as to whether these deviations
due to personal equation are of sufficient magnitude to be prac-
tically significant. The whole of the remainder of this paper
will be devoted to a discussion, from different standpoints, of
the quantitative aspects of the recorded classifications of the
several observers. All these data will bear upon this general
point. To answer the question specifically raised in this section
it will only be necessary to show the range of the variation
exhibited in the counts made. Table VI. gives for the four ears
and the four classes of kernels on each ear (a) the mean numbers
of kernels found by averaging the counts of all observers, (6)
the minimum and the maximum recorded number of kernels, (c)
the total range of variation shown in the records, and (d) the
percentage which this range is of the mean of the same class.

It is evident from this table that the personal element is one
of real significance. When two careful observers can diflVr in
their count of the same set of objects by as much as one and a
half times the actual number of the objects counted the factor
which leads to this difference is certainly not to be neglected-

An examination of the standard deviations and coefficients
of variation of the counts leads to the same result. These
constants are shown in Table \ II. It should be said in this

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 35

TABLE VI.

THE RANGE OF VARIATION EXHIBITED BY ALL OBSERVERS IN THE
SEVERAL CLASSIFICATI-

1

Class.

in.

;Ill.

int.

Rai

Y'-H hy.

316.87

298

54

17-0

'•t.

93-47

55

8

WI !iy. •

86.67

108

56

Whit'- -wet.

35-6o

25

54

151.7

Yd' hy.

240-93

9.1

l»\v sweet.

75-40

62

20

9

\\'liit'- -tarchy.

81.00

74

93

Whit-- sweet.

24.67

18

ig

:

[0

Y«-II»w starchy.

249.00

245

15

6.0

[0

low sweet.

67.47

51

;s

to

Wl. hy.

86.67

76

15

17-3

[0

\\ ' '-I.

28.53

18

1 I

hy.

228.36

221

71

31-1

I i

••t.

76.00

53

u

1 I

Whin- -tarchy.

70.64

7

71

n-

1 I

Win!- -'A.-^t.

32.00

14

41

i

M> an.

connection th.u f-»r the particular sort of proMnn hcic dt.ilt
\\iih ii \\<mld ,i!i]M-.ir that the nu'thcxl of i-\|iiv--in- tin- <lr.

• it \ .u i.iliilii v u 1 1 id i i- usrcl in TaMt- \'I. i /. e., tin- .il •-. ilut«- \ .ilui-

• if i he ranyc and ii- rd.nion U) tin- mean i- ]»r<>l>.il»ly nf n

n .il \.ihn- than an- the conventional con-i.mi- '^i\iii in T.il.K-
\ I!. In the privsent instance it is the run.'- of v.m.nion
ilu (\nt-ine amounts by which different <>|IMT\«T- dittcr in iln-ir
(oiini^i \\hifh is the thing of primar\- intnc-i .uid ]>i,uii«.il

nificance.

I .tMr \1I. gi\-es the standard deviation .ind < m UK icni <>t
\.iii.iii"n for the counts of each class of krrnd-.

TAHI.I. \ !l.

mi. IE AND RELATIVE \'\KIVIMN IN n ,11:

\ 1 I | \ss |- V ll|.' ;

'iy. 'chy.

'S

-44

4.24

12.90

i ;

g

(>

-09

19.00

[0

i n

9.08

i

..

17

17

i .,

!i'>t usi-d in any d«-taili-'l i-i'inpai i~'Hi« it h.i^ nut
tlu.nv.lit • !. ul.it.- )>[..l..iM. All tlr .tiy .l.it.i .1:

h.in.l. h« \\r\ct. il aiiyon. in.ik" t!. It m-.-.l ..n'

n •mi-nil', -n-.l th.r i .in.l P.. 11 = 15. an.l |..r i-ar I I . n = i ).

354 RAYMOND PEARL.

This table brings out several points which need discussion.
These are:

1. The amount of variation, both absolute and relative, in
the counts is shown by the measures here used to be very large
for some ears and classes of kernels. For no ear, taken as a
whole, can the variation fairly be considered negligible. Thus
the conclusion previously reached by another method is con-
firmed.

2. The amount of variation in the sorting and counting is
distinctly different for the different ears. From the values of
the constants it would appear that ear No. n presented the
greatest difficulty in respect to the classification of starchy
kernels. In respect to sweet kernels ears No. 8 takes rank as
offering the greatest difficulties. The starchy kernels of ear No.
10 were the easiest to classify of all starchy kernels. In the case
of sweet kernels ear No. 9 had fewer intermediates (i. e., was
easier to classify) than any other ear.

3. Relatively there was closest agreement among the observers
in respect to yellow starchy kernels, and least agreement in
respect to white sweet kernels. This table illustrates the fact
which was evident to the observers themselves, that there were
marked differences in the ease with which the kernels of different
ears and different classes could be sorted.

Now while it has been shown that the fifteen observers do
not agree in their classification and counts, and that the dif-
ferences are too large to be neglected, it may fairly be asked if
the same result would appear if the group of observers participat-
ing were not merely scientifically trained and familiar with maize,
but in addition had had a considerable amount of actual expe-
rience in the detailed study of variation and inheritance in plants.
In other words, is not that special familiarity with the object
which comes with the active prosecution of research in a particu-
lar field worth something in reducing the magnitude of one's per-
sonal error or "equation"? To get some light on this point Table
VIII. has been prepared. This is made up in exactly the same
way as Table VI., except that only observers VI., VI I ., VI 1 1 ., IX.,
XI., XII., XIII. and XIV. are included. These eight observers,
comprising the staffs of Professor Johannsen's and the \\riirr'-

PERSONAL EQUATION IN BREEDING EXPERIMENTS.

laboratories have certainly had more extended experience in the
din (i and immediate <tudy of plant breeding and of variation
in plants (involved in all breeding investigation' than have the
oilier observers of the original fifteen. Lists of published pa;
could be cited in proof of thi- were it ncce— ary, but the tart is
obvious. Will this group of workers on problem- of variation
and inheritance show a similar d of variability in their

nits to that brought out in Table \ l..J

TABLE VIM.

IMG THE RANGE OF VARIATI - VI. :•> IX

INCLUSIVE. AND XL TU XI\'.

Clou.

Mean.

8

• >w starchy.

312.88

SWeet.

98.13

18

8

Whiti- -tarchy.

90.50

8

\\ .-i.

30.50

9

liy.

242.88

0

9

'•t.

77.00

5

9

\N 1 liy.

79-75

77

5

9

\\'liiti- sweet.

22.38

4

10

• iw starchy.

248.38

^

10

Yellow sweet.

-1.88

6l

15

IO

White starchy.

87.13

85

5

10

White sweet.

24.13

20

1 1

: .w starchy.

224.50

]

1 1

Yellow sweet.

81.50

IS

1 1

While starchy.

74-50

7

1 1

White sweet.

26.50

IS

li ii from comparison o| thi- table \\ith Table \ I. iliat

the amount of variation in the -<>,iin- an-1 c-niniiir^ i- di-tincil\
leil-iied in llu- group of student^ o! \.iriatioii. \\lnna- lln-
average jierccntage of range in mean ; I ible \ I., ii i-

bui i«).o, or onl\- approximate 1 '.hird a- much, for Table

\III. Tluis it appears that in tlii- ia-i as \\ould be

expected on general ground-. 1 experience »v pr.nii.-e

in a particular line -n-.illv re<luce> the per-on.il ei|ii.u ion .
It nui>t be said, ho\\e\er, that e\en \\ith the -nuip of ob-
servers included in TabK \ 111., the dil'lVrence- are to-. I.
in be neglected. When the ran-e <if variation amongsl ilil'ten-nt
observers of the >ame thiivj amount- on the a\< • approxi-

356 RAYMOND PEARL.

mately one fifth (19.6 per cent.) of the mean value of the thing
counted it indicates a source of error not lightly to be dismissed.

III.

It is desirable next to examine somewhat more closely into the
nature and distribution of the discrepancies among the observers.
A point of particular interest is to determine to what extent
the counts indicate a definite and persistent bias on the part of
an observer. There may be great variation in the counts of
several observers of the same set of things and yet each observer's
judgments may be distributed quite at random about the mean.
In order to get more light on this and some other matters
Table IX. has been prepared. This table gives in successive
columns for the four kernel classes, first, the mean deviation
from the mean, all deviations being taken together without refer-
ence to sign (i. e., the mean total deviation), and second, the
mean net deviation from the mean, got by taking the algebraic
sum of the deviations. All four ears are used in getting these
mean deviations. An example will make clear the method of
obtaining the values given in this table. An examination of
Tables II. to V. inclusive shows the following set of deviations
from the means in the counts of yellow sweet kernels by observer
Xo. V.

-i- Deviations from Mean. —Deviations from Mean.

7.53 (ear 8) 2.47 (ear 10)

5.60 (ear 9) 10.00 (ear 1 1)

13.13 == sum of -f deviations 12.47 :: sum of - deviations.

— = 6.40 = mean total deviation from mean.
4

13.13 - • 12.47

+ 0.165 -- mean net deviation from mean.

4

The last column of the table gives the total deviation from
the mean of each observer, all cars being taken together and the
deviations summed without regard to sign.

It is strikingly evident from the mean net deviations in this
table that each observer was "a law unto himself." \< nl\
everv one of the fifteen evidently had a different system ot si >rting.

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 357

TABLE IX.

SHOWING TIII-: MEAN DEVIATION FROM THE MEA i. \\i> NET) AND TOTAL

IATIONS OF THE COUNTS OF ALL OBSERVERS

ny.

Yellow Sweet.

iy.

:= —

lee

d

3 = =

3 = =

—

.

- - d

~ - ~

z ~ 5

/ I

|Sq

**

_

*" i" ' ~~

"

r

=

=

. i-

.-. i

£

- - -

-

I.

+ 2.3

IO.IO

- 0.34

26.995

+ 0.14

II.

22.09

— 22.09

4-68

+ I.

III.

- 9

14-34

-14-34

IO.26 ;

+ 1

IV.

+ 7

2.86

+ 0.93

9-93

- 6.

—

V.

- 8

6.40

+ 0.17

3-34

+ 3-

+ :

VI.

— i

4.42

+ 4-42

2-59

+ r-

- 1

'

VII.

- 4

4-59

- 4-59

4-26

+ 4-

+ :

VIII.

- 3

5-92

+ 5-92

3-26

+ 3-

— '

'

IX

+ 2

6.42

+ 6.42

2.84

— 2.

49

— •

X.

— 4

6.65

— 0.085

5-26

+ 5-

6.65

-

— i

4-37

+ 4-17

3.84

+ i.

51

- '

+ 3-17

3-51

+ 3-

OI

- •

XIII.

+ 1.67

4-51

+ 3-

51

-

XIV.

+ 9-17

3-16

— 0.

25

-

'

XV

+ 4

6.67

- s-

1 1

-

<>! ilu-i- (lilii-i'i-in c-s are very intcn -iin^. I'm-
No. I. had a hi^li "<•' <l«-\ i.iiimi on starchy krnu-U. u-ndin.

•iinatc tin- yellows and practically zero n« i dr\i.itii>n "ii
-\\cri k. rnels; XI\*. is exactly the op . h.i\iir^ verj -in. ill

net dt-xi.iiions on the starchy and tending -tnnnjv i<> over-
estimate the yellows among the sweet k(iiu-U No. II. li.id tlu-
iciidi-iKA to underestimate the yellow sweet>, and c< n-rc-ixuidin^ly
to ov< i' -lim.iic tin- white sweets. No. III. oni-i-M-niK umi
f-iini.iu-d ratht-r lu-a\-ily all yellows and o\ --n •-n'ni.iti-d all u hiic-.
No. IV. did ! I >' the opposite. No. \'. >h«i\\ - a \»-r\ irraiic

se1 «•! net di-ii-ilnitii>ns, o\vin^ to his idi<i-\ m IM-\ respecting the
di-t rimination i n starchiness and n«in--t.in-hiii' Tlic

n -ult i- that \\hilr he underestimated tin- vll<>\\ -tan h\ ki-nn-1-,
In- «>\ »-i\-- tiinat i-d all I he ol In -r rla-- No. \ I. -••nicu hat ninlcr-

estimated the yellow starchy, but »\ n-c-tiniatrd tin- \ clln\\ sweet.
No. X shows an lAiraonlinarily Miiall net dc\ialion <ui tin-
>\\rd kcnii-l-. Inn di-tinctly un«li-n>tiniatfd the \cllc\\ -larrhy.
In yrnt-ral the table shows in a striking way, that the indi\ i«luality
i>t" tlu- i.li-i-i-\i-r i- . -v to IK- nrkiMu-d \\ith in work of thi-

sort.

RAYMOND PEARL.

It is of some interest to examine the trend of the total devia-
tions given in the last column. The data are shown
in Fig. 5, arranged in order from the smallest to tin
deviation.

This diagram illustrates a point frequently overlooked. It is
commonly argued that the more independent judgments one ob-
tains regarding any point the more accurate will the average
result be. We are apt to say that if ten men measure a stick
the average of their measurements will necessarily be nearer to
the true dimension than if but three men measure and their
average be taken. But it is plainly evident from Fig. 5 and
Tables VI. and YIII. that the inclusion of observers I., II., III.
added nothing to the accuracy of the mean. The point which is
forgotten in assuming that greater numbers necessarily mean
greater accuracy is apparent if we examine the equation for the
probable error of a mean which is

P.E.J, •• .67449

V n

The probable .error, to be sure, varies inversely with n, but
it also varies directly with a, the standard deviation. And, what
is here of primary importance, the standard deviation tends to
increase as n increases. Whether the probable error shall be
smaller or not as the number of observations is increased depends
upon what has happened in the meantime to the standard devia-
tion. When n is small, as in the case here under discussion, the
effect on the standard deviation of taking n + I observations as
compared with n may greatly outweigh its effect in the denomina-
tor of the probable error fraction.

IV.

The next point to be considered is the relative constancy of
the same observer's error. If each of the fifteen observers had
made a second count of all the ears at some considerable interval
of time after the first, how closely would the recounts tally with
the original counts? Such an experiment really tests, of course,
the stability or constancy of an observer's judgment . It indicates
the degree to which his standard of sorting is absolute, and to
what extent it fluctuates.

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 359

It was not feasible to ask all of the original fifteen observers to
go to the labor of recounting these ears. Second counts made
after a relatively long lapse of time are, however, available from
three observers (namely, VI., VIII. and IX.) for all four ears.
\\hile this gives only comparatively meager data, -till some
point-- of interest appear. The-e d.iui are given in Tables X.,
XI. and XII. It should be -aid that the recounting was done
in the same way as the original count. In each case the observer
had no access to the original data while the second count was
in progress. No one of the three had any remembrance of what
hi- »r her) original counts were. The writer has not been able
to di-cover any factor which would make these recount- any-
thing other than what they were intended to be. namely, really
independent determinations of the same mail-rial by the same
observers after a long lapse of time.

It will be remembered (cf. p. ,}_).<) snf>ni\ that one kernel I'n.m
ear No. 10 was lost in the course of th' nal counting, h is

theieion- obvious that all the recounts of thi- ear mu-t of m .
HI y be one kernel smaller than the tir-t count-.

TAUI.I-: X.
• INAL AND SECOND COUNTS - 8 TO u i .\». VI.

I • Is.

Count.

Yellow
Starchy.

Yellow .
Sweet.

rial.

int.

313
3«

IOO
100

yo

|

.I.mu.iry J i , K;in.
[pll.

— I

';. I 'I i.;ilial.
• ;. Ki-i-'llllt.

2.»2

79
76

81

J.tnu.irv 1 1 . K;IO.
AniMHt II. ivi I.

ice.

~~ 2

-3

+ 2

nal.

!<• '.ml.

251

75
73

_l

JaiiiKiry Ji, IQIO.
AII^II-'. ii. [911.

-7

— 2

I I, ( >iii;iii.il.
II, Krr.illllt.
I )ilt<-I. I1CC.

223

76

75

7"
7''

0

.1. mil. u v 21, i'
• IO, i<;i i .

O

— i

The data in the-e table- indicate that, -o far at lea-t a- tlie-e
thrc-e observers are con<vrne.d. the judgment of the indi\idiial
i- reasonably constant. This i-, plain it' the total deviation of

3 6o

RAYMOND PEARL.

TABLE XI.

ORIGINAL AND SECOND COUNTS OF EARS 8 TO n BY OBSERVER Xo. VIII.

Kar and Count.

Clasees ol

Kernels.

Date.

Yellow
Starchy.

Yellow
Sweet.

White
Starchy.

White
Sweet.

8, Original.
8. Recount.

311
309

— 2

241
243

101
93

-8

78
79

92
94

28
36

January 20
August 12,

January 20
August 12,

.1910.
1911.

Difference.

9, Original.
9, Recount.

+ 2

82
80

+8

21
2O

, 1910.
1911.

Difference.

10, Original.
10, Recount.

+ 2

247
246

+ i

75
73

— 2

89
89

— I

21
23

January 20
August 12,

, 1910.

! • .. ! [ .

Difference.

ii, Original,
ii. Recount.

— I

224
223

— 2
82

81

0

75
76

+2

26
27

January 20
August 12,

, I9IO.
I9II.

Difference.

— I

- i

+ 1

+ 1

TABLE XII.

ORIGINAL AND SECOND COUNTS OF EARS 8 TO n BY OBSERVER No. IX.

Ear and Count.

Classes of Kernels.

Date.

Yellow
Starchy.

Yellow
Sweet.

White

Starchy.

White
Sweet.

8, Original.
8, Recount.

327
314

101
98

78
89

26
31

January 21
August ii.

, 1910.
1911.

Difference.

9, Original.
9, Recount.

Difference.

10, Original.
10, Recount.

-13

242
240

-3

80
76

+ 11

79
83

+5

21
23

January 21
August ii,

, 1910.
1911.

— 2

249
246

-4
70

+4

87
89

+ 2

+ 2

26
25

January 21
August ii,

, 1910.
1911.

Difference. —3 +i

— I

II, Original.
ii, Recount.

Difference.

228

222

8?
88

71
77

21
20

January 21, 1910.
August 12, 1911.

-6

+1

-i

recounts from original counts be considered. In the case of ob-
server No. VI. a total of sixteen kernels were differently classified
in the recount from what they were originally. Since the whole
number of kernels involved in the experiment was 1,792, iliU
means a discrepancy of less than one per cent, between two
independent sortings more than a year apart. Such an error is

PERSONAL EQUATION IN BREEDING EXPERIMENTS. 3^ r

certainly negligible. Observer Xo. VII I. cla--ified eighteen
kerne]-, .ill told, differently in the recount than in the original.
Thi i i- only about one per cent, of the total kernels handled,

and cannot be regarded as a significant error. In both of these
case- VI. and YIII.) the discrepancies had to do entirely with
the color classification. With observer IX. this was not the
case. < >n both ear 8 and ear 9 she classed two kernel- as -\veet
in tin recount which she had originally called starchy. Alto-
gether this observer < lassificd thirty-five kernels differently in the
r< (oiini from \\li.it -In- did in the original. This ho\\e\vr repre-
sents .1 P lati\e error of a little less than two per cent. V> very
i stress ' ould be laid upon such an error.

I roin i In- i.iM. s it will be noted that there was a marked and
nearly uniform tendency on the part of all three observers to
underestimate tin \ellows (both starchy and sweet) and to over-
« stimate the \\hin •-. in the recounts as compared with the orig-
inal-. It seems probable that the cause of this lie-, in part at
lea-l. in a fading of the yellow color during the lime -ince the
• inn- \\eiv made. Thus it may be that kernel- which were
plainly vellou \\lien lir-t c, united are now white or very nearly
so. A further fact which would indicate that fading had occurred
i- found in ihe menial impressions of the observer-. All three
found the material di-tinctly more diflicult to classil\- \\hen re-
counted than whi'ii ori^inalK counted. One feels certain that
a part, ai le.i-i. of ihi- i- due t<> a change in the man-rial it-elf.

l\eco-ni/in- tully the nieauerness of the material, the fact-
so far as the) ;^o seem to indicate clearly that tin- -ame < >\ »sen er
i- likcl\ to « !a— ity the same material in about th. -ame \\a\-
ever) lime. It a particular kind of bias is sho\\ n in one count
it \\ill appear i--(iiiially unaltered in succes>i\e tri.il-. Thi- i-
I u i '1 i.il >1\ UK ire 1 1 IK- < •!' i 'I i-ervers especially experienced in dealing
\\ iih the data of \ a riat ion than in the case of tlm-e \\ it In mt -nch
experii nee, though figures are lacking to demon-irate thi^.

V.

\\ . c< urn- next to the consideration of the -ecotid ijm-lion pro-
at the beginning (p. 341). Thi- u a-: " I >o( •- -, unat it • 'inter-
--' in mai/e im])l\ gametic ' intermedia teness '? In

362 RAYMOND PEARL.

other words, do ¥2 kernels which are intermediate somatically
give rise to any different sort of progeny when planted than do
kernels which belong clearly and indubitably to one or another
of the well defined gametic classes in F«?" To answer this ques-
tion carefully controlled plantings of somatically intermediate
kernels were made in 1910. Series of starchy and of sweet
kernels were formed ranging in each case from pure white -at
one end to pure deep yellow at the other end. Then rows were
planted as follows: (i) pure white, (2) deep yellow, (3) the lightest
yellow to be found (== somatic intermediates), (4) the yellowest
whites to be found ( := somatic intermediates). The kernels
in classes (3) and (4) were such as would be classified with
the yellows by some observers and with the whites by others.
The rows included about twenty plants each and were made
in duplicate, and in some instances triplicate for both starchy
and sweet series. In each row a varying number of ears were
self-fertilized (i. e., pollinated by hand with pollen borne on the
same plant). Owing to the numerous vicissitudes incident
to hand-pollination, together with pressure of other work, as
large a number of good ears as would be desirable was not ob-
tained. Some of the possible gametic combinations were not
represented at all in the progeny ears. This part of the investi-
gation is, in consequence, not complete. It seems desirable,
however, to present briefly the general result shown by the fifty
odd ears at hand.

This result was that there was no discernible difference what-
ever between the progeny of groups (i) and (2) as a class, and
that of groups (3) and (4) as a class. In (3) and (4) some of the
kernels planted were of course heterozygotes and some were
homozygotes. The same was true, however, of the kernels of
(i) and (2). In each case a typical Mendelian result was ob-
tained, and ihis result could have been predicted in every case
(with the exception to be noted presently) had the gametic con-
stitution of the kernel been known when it was planted. It
could not have been predicted from the somatic appearance ol
the kernel.

The only behavior of an exceptional character observed in
these selfed ears was that in certain of the white sweet kerm K,

IM KSONAL EQUATION IN BREEDING EXPERIMENTS. 363

which \\< re homozygous recessives in respect to absence of yellow-
ness aiul starchiness, selfing brought out a latent red.1 The
thn-r ears of this type which were obtained all came from kernel-
< la-sified in the planting as pure white (group (i)). Xo such
ears were obtained from selfed sweet kernel- in group (4). The
toi.il number of homozygous, non-yellow sweet ears obtained
was too small, however, to make it at all certain that .-imilar
n-d • ars might not, with larger number-, be obtained from unmp
I ' ' rnels.

It Is planned to get further data on thi- portion of the investi-
gation. using for planting the kernel- of ears s. 9, 10 and 11
\\liich formed the material for the personal equation part of the
\\ork. It can be said at this time that tin- experiment- \\ith
'id maize so far conducted furni-h no evidence thai
-omatic " intermediateness" comx.' metic intenncdiaieii'

I !:<• progen} "I a deep yellow kernel -dl'ed i- not \ i-ibly dilleivnt
In. m that of a lii;ht yellow kernel -elfed, pro\ idi-d both are • •!
the -aim tic constitution. The tv-ult nl thi- experiment

pn-ci-eK \\ith Darbishi re's2 extensive stud) <.! « •--. ntially

the same problem with peas. Indeed hi- final < •< >m -hi-i. >n
( it., p. 71) applies here without change of \\onlii ' That in the
attempt to predict the result i\eii mating the somatic char-

a< it i not only of the parents and of the ancestors of the indi-
\ idual- mated, but of the individuals thi -in -i -1\ e-. max be nitin 1\

It-tt oni of account ; and that the expectation ba-i-d »n a tin
of the contents of the germ cells of the two indi\ idual- i- fulfil led

I MSCUSSIOX AM) Si \IM.\K\.

l\e-nlt- MM h as are set forth in thi- paper \\mild crriainK ha\ e
been at oin- time proclaimed by some as furni-hiir^ a refutation
of Mi ndi-li-m. In fact one of the carlie-t i i itii i-m- « .f M.-ndi-liaii
\\ork was mainly devoted to calling attention t" the existence of
Mich somatic intermediates between Mendelian categories in the

1 A mill. H i' suit !i i I'Vrmly IM-I-IL i ..n, K. A. Ri-jit. An.

we., \'I.. 233-237. i'J' i-
•' 1 ).n l.i-lim . \ D., "An I mental] -muition <>f tin- Fheory of Ai

•iiluitinii- in I h-n-.lity." ]'' *> !'.. \ 1. Si, pp ' i ;•;. 1909.

3 \\YMoii. \\ . I- . K.. " Mi'ii '• 1 - I \ltrriiutivi- Inli'-r itaiK-i- in I'

trikii. \'«\. I . |>|>. J-'N -'5|. I'l.iti-- I. .ui'l II.. -

364 RAYMOND PEARL.

case of peas as are here shown to exist in maize. That such
variation, provided it be really somatic or fluctuational, is, how-
ever, of no real importance in relation to the cardinal facts of
Mendclian inheritance has been shown by all experimentalists
who have devoted attention to the matter. Bateson1 (loc. cit.,
pp. 240-244) gives an illuminating discussion of the whole matter,
with special reference to the phenomena in peas. East and
Hayes2 discuss the same point with reference to maize and sh<>\\
that somatic intermediates behave in inheritance in accord with
their gametic constitution rather than their somatic appearance.
Certainly the time is past when facts such as are set forth in the
present paper can be adduced in criticism of basic Mendelian
principles.

The essential point brought out by this study is, it seems to me,
that the well known general fact that every datum of science
is a function (in the mathematical sense) of two variables, namely,
the observer and the thing observed, is once more emphasized
by a particular case.

A thorough investigation which brings out essentially this
same point, though conducted on a different class of material
and with a somewhat different object in view, has been made
by Yule.3

It will be freely admitted by everyone as an abstract proposi-
tion that the personal idiosyncracy of the observer constitutes a
source of error in all scientific observing. Yet how often does
the biologist not working on strictly quantitative problems make
any effort either to eliminate or determine the magnitude of this
source of error in his case and in a specific instance? Anyone
who has not experimented for himself on the matter can hardly
realize how important, on the one hand, and how difficult on tin-
other hand, it is to attain to any considerable degree of real
objectivity in results. While the "exact" sciences are some-
what better off in this regard than biology, they are after all not
greatly so. There has, to be sure, been a great deal of work d< >IH-

1 Bateson, W., "Mendel's Principles of Heredity." 2cl Edit., Cambridge, 1909.

2 East, E. M., and Hayes, H. K., "Inheritance in Maize." Conn. Agr. K\pt.
Stat., Bulletin 167, 1911.

8 Yule, G. U., "On the Influence of Bias and Personal Ki|ii;itinii in Statist!'
Ill-defined Qualities." Jour. Anthropol. Insl., Vol. XXXVI.. pp. .i-'.S J«i, 1906.

EQUATION IN BREEDING 1 XI'I KIMI.XTS. 365

on the theory of errors of observation, particularly as related
to a-ironomy, phy-ic-. and like subject-, yet so late as H)O2
iV.ir-on1 d'-mon-trated in a mo-t convincing manner that much
of the then <-urrently accepted theory wa- \\ron-. and that all
of it quite ..\erlooked a factor which miijit be exceedingly
important, namely, correlation of judgments.

The pre-em -tudy i- by no mean- a complete in\ e-i i^at ion of
tin- proltlem of per-on.il equation in Mendelian work. (Correla-
tion of error- mi-lit t., U- -tndi«-d. and certain other matter- ELS
well. But the pre-eiit material i- -tati-ticallv entirely inade-
quate f»r the di-cu--ion of these point-, and it dor- not -rein
• olli-n more extensive data, -in* • in\ •< >\\ e- too

it a tn on the time and good nature of bu-y worker-.

I null. -r tlr material here presented brings out clearly the pri-
maril\ « — -iiiial ]>oints. It shows that in .1 M'lidelian ratio tin-

il eijuaiion of the observer mark- a -our. rror \\hich

in iln- il mai/e i- of considerable magnitude. Tin- >ource

r quit- hadows in magnitude, in thi- error

due to random -amplin.. \'et it is the latter alone \\liich i-

ordinariK considered I .lelianwc '!'!)«• probaM. « -nor

i Mendi liali latio a- i oiuiinmly c.ilculated tell- one the pmb-

aliilii\ that the sample < omite<l i^ l( tme representation "I tin-
population from \\ hich it \\ a- dra\\ n. It tell- one no; Inn-
\\ hale\. i -about the unconscious bia- of t lie . -oiuiter a- a l'a< tor
in prodiiein- : he result set down.

l'.\ \\a\ o| -uminary it may be >aid that in thi- paper i\ idem e
i- pn-eiited \\hich shows that:

i. The ob-er\ed \:2 Mendelian ratio- determined from the
same foul • n - "I mai/e b\- fifteen competent ol , all dilti r

I'n mi on. aiioth-

j. The failure of all observers to agree in their di-tribution of
kiin.l- into -t \eral categories n--uh- from t\\o causes, \i/..
the e\i-tenre of -..mat i( all\- inn-rmei liate kernel-, and (6) the

onal bias or idiosyncracj "i iheob-er\er.

3. The magnitude of the difference- betueen the -e\eral
ob>ei-\er- i- -ucli a- to denum-irate that tin- per-oiial equation

'IV.u-'in ; ;!n- M.iili' mi-lit, with

S|M . ial !•' Phil. I A, Vol

pp.

RAYMOND PEARL.

is a factor which cannot safely be neglected in work of this
character.

4. The observers who have had most experience in the appre-
ciation and measurement of variation have the smallest personal
equations on the class of material and the problem here treated.

5. There is no evidence that the progeny of somatically inter-
mediate kernels is different, in any respect whatsoever, from the
progeny of distinctly non-intermediate kernels of the same
gametic constitution.

DIFFERENTIATION OF THE HTMAX CELLS OF

SERTOLI.

I II' (MAS II. M"\ K.' 'MHKY. JR.

-ITY OF I. VAN! A.

This study is based on the examination «.f the testis "I a

ro aliout 40 year- of ,i-c. piv^TM-d in Xenker's fluid \\hile
Mill \\arm after hi- e\eeiitioi). The fixation \\.i- not as excellent
.1- niiijit In- desired, cytop'.a-mic detail- ln-iu- not always pre-
served, 1'Ut the preservation "f tin- nuclei \\a- mi tin- \\ln>!e MTV

d, and of -pindle fiii'ii'' Mcni. A considerable \ariety ot

>taiuiii'j IIH thods were cinploytd, of \\hieh the nio-t Iruitful
|ii.i\t-il id In- 1 Icidnihain'- iron li.rmaio\\ line, \\ith \ arion>

•'i- of r\tr.i< lion, follout-d |i\- .ili'oholi.- eosin. 1'araltiin-

ions were made of 5/1 and Sju.

I op tlie '^ift of tlli'- malt-rial I am indrlit d to tin- kindiic-- of

I >i . . \ddi-on, of the I 'ni\ i-r-ii >• of Pennsylvania.

i. GENERAL < M M.IM «>i mi PROCESS.

'I'hf tf\t diagram exhiliii- tin- i hi' I iv-n!'- oliiaint-d. Tin-

antepenultimate spermatogonia t-oniain eaeh a r«Mi (R.) \\itliin

ihc ( \ lop!.i-m. This doe- not di\ idt- in inilo^i-. i «\\^ i|iifiitly

iu-t half of their daughter cells, the penultimate spermatogonia,

foiiit- to i-oiiiain each a rod. \\hilt- half of iln-in laek it. In iln-
di\i-:on of thf-c pcnultiniate spermatogonia thf rod ddi- not
di\idt- Inn lu-foiDf- di-ti-iliutfd to oiu- nuartfr of the ultiinatf
-|i- i inaloLoiiia. Each of ever} llm-f ultimate -ptTmato-onia

product-^ 1>\ di\ ision two primarv spermatocytes, and ihf-t- (<•!!-

\\ h ifh I n -Ion- to the true .germinal c\ clt- laek the rod f mi rely. But

ii four ih ultimate spermatogonium presen es the rod. and this

cell without further division enlarges and becomes a cell of Sertoli.

In t hi- cell of S'l'ioli a ]>rimar\ n>d let (r. j) buds oi'f from the rod :
then the rod disappears, while the prim ili-t di\idf> inio

t\\o -econd.ii \ rodlei- r.2 ind the latter persist in the Sertoli cell

throughout it- hi-lory.

367

368

THOMAS H. MONTGOMERY.

The line of the Sertoli cell is therefore <h lennined In- the
presence of the rod; one Sertoli cell is produced to every three

Antepenultimate
Spermatogonium

Penultimate
Spermatogonia

Ultimate
Spermatogonia

Sertoli
cell

ultimate Spermatogonia that lack the rod, or one Sertoli cell
to every twenty-five spermatids.

2. THE ANTEPENULTIMATE SPERMATOGONIA (Fics. 1-8, PL. I.)-
These are the largest germ cells in the adult testis, and like
the other generations of Spermatogonia arc situated at the pe-
riphery of the seminiferous tubules. Frequently their nuclei are
of irregular shape, as shown in Fig. I. Within the nuclei ( Fig. 3)
are two kinds of nuclcolar structures: acidophilic plasmosomes
and basophilic bodies; it would take a detailed Mudy to deter-
mine whether the latter are chromatoid nuclei >H or modified
chromosomes (allosomes). In their cell bodie- ;nv tound ehro-

THE HUMAN CELLS OF SERTOLI. 369

malic rod-. never more than one to a cell, various forms of which

dra\\n in Figs. 2-8, that of Fig. 5 being the largest found.

The rods are homogeneous in appearance, dense, and they -tain

with ha-ir -lain- but u.-ually not as intensely as in the later

Denial generations. Such rod- are u-ually in contact

with tin- nuclear -urface, hut not always, and do not occupy

constant po-iiion- with regard to the pole- of the cell. Inar-

acteristic of the antepenultimate spcrmatogonia i* 'he relatively

-in, ill -i/e of • >d- and their frequently twi-ted form.

Thirty of the-e ceil- were carefully examined, and tuvntv-
thn-e i»f them showed each one n,,|. ( )f the remaining -e\ en,
fi\e were ii"i \\holl\- within the plane of the section, so that their
rod- mav have been present in the < i portion-. It i-, prob-

able thai each < ell of this g< •]]• -rat i«>n comes t<> contain a rod, and
thai the rod- are first produced in thi- generation; no -perm,

ia o| e.irlier ^mcrations \\ere pre-ent, houexer. consequently
there can I"- no -urety of the latter point.

What the nietliod of origin of th I- mav lie could not

In- determine<l. They are quite di-timi from the idio/ome. at
least when fully formed, as shown in Fig. 2. In our • i

no rod was found lull an irregular granular mass which i-

po--ilil\ .1 precursor of the rod; if this be so, the roil mi-hi I.e

con-ideieil to lie produced l>\ the glomeration of granules

at lir-! • red in the cell body. Hut the -tate of fixation of

the c\ to|ila-m was not sufficiently reliable to allo\\ of an\ -ali--
factorj d« ii i inination of this matter. There i- no e\ idence that
the rod- are directly produced from the nucleii-. I heir origin
is thus unexplained, though their -ul >-ei iiieiit hi-tor\ i- perfect 1\-

( leal .

\o mitoses of these cells were found, but there can be no doubt

that the rod- do not become divided for ju-t half of the cell- of
the next generation contain rod-.

;v Till I'l \i LTIMATE M'l KM \ \l\ FlGS. -i !<• ,

l'ort\ -nine cell- of this generation \\en- examined, care being
taken to -tu<l\ only those that lay entirely within the plane of
the section, and of the-e tuenty-foiir exhibited each a rod \\hile
t\\entv-fi\e -houed no rods. ANo -ix cases \\ere found of tuo

3/O THOMAS H. MONTGOMKRY.

nuclei in one cell body, indicating nuclear di\ IMOII without .cyto-
plasmic division of antepenultimate spermatogonia: in .ill MX
of these cases only one rod was present, and an example i-> -ho\\ n
in Fig. 9. There can thus be no doubt that hall" tin- penultimate
spermatogonia contain rods and half do not.

Characteristic appearances of the rods are illustrated in Figs.
9-11, PI. I., 14^16, PI. II. They differ on the average from those
of the preceding generation in being usually larger, straighter,
and more deeply-staining with haematoxyline, which indicates
they have been undergoing growth changes.1 Not infrequently
they are curved around the nuclear surface (Fig. 14) and tin-
length of a rod may equal the diameter of a nucleus. Conse-
quently they are in this stage very prominent constituents of the
cell bodies and easily differentiated by safranine or haema-
toxyline.

Mitoses of these cells were not frequent, but two clear cases
(PI. II., Figs. 12, 13) were found, showing that the rod (R.)
passes undivided into one of the daughter cells, and this is fully
borne out by a study of their distribution in cells of the following
generation. Fig. 18 shows the end result of such a mitosis in a
case where the cell body had not divided and here there is but
one rod. \Yhat is the nature of the scattered globules shown in
Figs. 12 and 13 is doubtful; they may be discharged nucleolar
material.

4. Tiii': ULTIMATE SPERMATOGONIA (Fios. 17-24, PL. II.).

These are the smallest of the spermatogonia and tin- most
numerous in the testis studied. One quarter of them contain
each a rod; three quarters lack rods. One hundred and forty-
two of these cells were studied, at stages before any of them had
enlarged into Sertoli cells, the precaution being taken to include
only cells lying wholly within the section; of these twenty-fix r
showed each one rod, and one hundred and seventeen showed no
rods. This ratio is somewhat less than I : 3, which is readily
explained on the ground that some of the spermatogonia \\ith
rods had already become Sertoli cells and then-lore were not
included in the count. A very important and clear case i- that

'The condition of the pair of rodlcts (r. 2) in Fig. 10 will l><- .\plainrd luti-r.

THE IITMAN CELLS OF SERTOLI. 37 T

of Fig. 17; this shows four nuclei, the granddaughters of the
nucleus of an antepenultimate spermatogonium. while there has-
been no division of the cell liody. and it will he seen there is
but one rod to the four nuclei. The evidence is then decishe
that one quarter of the cells for this generation contain each
one rod.1 In these cell- the rods arc on tin- average more- ma--i\ e
than in preceding grin-rations (Fig-. 17 24), and while usually
iiion- or less curved an- never t\vi-ted. (Juite frei|uently one end
of i he rod is bent off at an .ingle Figs. 17. -1-1 . In these cells
al-o the rod- an- nio-t den-e ,md acquire their maximum -tain,
staining fully as inten-elv as the basichromatin ; they generally
hut not aluays touch the nuclear .-urfat v.

5. DIFFERENTIATION AND HISTORY "i un SERTOLI CELLS

(Fit.-. j5 50).

All thr ultimate SpermatOgonia that contain n>d> hecoine cells
of ^i-rioli, and ihosi- only. Nothing like either rod- or rodlets
Were found in any of the SDermatOCytes or -permatid- The
N-iioli cell- hrconir e-| le. i. il I \- marked h\ their great

25, I'l. [I., shows the beginning ol -uch ^n>\\ ih,
(ell (A) growing out heMmd it- sister uliimate -|iennaio-.uiia
I H ,ind ( . I , | and .i-. I'l. N I .. -lm\v ^ertuli cell- in a later

growth 'agr in their eiitiret\. and 1 19, 41-46 exhihit

portions of them in still later -i 1 !n se • elU hec.mie icl.i-

ti\i 1\ enormous as shown in I i. I'l. \'., \\liich n-pre-eni- a

pi-rtii'ii of a transection of tlu- \\all »t a -emiuilerou- iiihule; in
thi> ligure tlu- shaded portion represents the hmlie^ nf the Sertoli
cell-, \\hich have grown far into the lumen of the tuhtile \«
emhi.n e i he -per ma lids. Thi- ^icat growth i- due mainl\ to the
formation of \'acuoles within the e\i«>pl.i-m. and in the li-mv-

onl\ the larger of the vacuoles are shown, not tin- gn-at numher

<>1 minut. . These vacuoles are drop- ,,)' a noii — laiiiin-

llu id. like that contaim-d \\ ithin the ca\ it\ of the tuhule; only in
rare in-tances are an\- concretion-, found in the \ at uole>. ( )ne
end, the ha-al, of eac'h Sertoli cell remain- adherent to the lihrous
\\all of the tuhule and in the figures lim - are draun to denote

1 Spi-iinat<«;:i'iii.i \\ itli two. tlin-c or l<.nr nucli-i in a -ini;!.- o-ll bmly arc- unusually

In .|in-iu aii'l in >u. 1, • Ut l( i an i:.-i[iii-nily »\ (jiiitc um-qu.il v«ili;-

THOMAS H. MONTGOMERY.

the inner border of the tubule; the other end, the distal, is the
•one that grows out and forms branches ramifying around the
spermatocytes and spermatids. In the later history of the
Sertoli cells large spaces are found within them, as shown in
Fig. 50, which are cavities in which germ cells had been situated
before their transformation into spermatozoa. Boundaries be-
tween the Sertoli cells become indistinguishable, so that these
cells come to constitute a syncytial cytoplasmic net of extreme] v
vacuolar structure (Fig. 50). In the basal portions of the Sertoli
cells parallel bundles of fibrils may be seen at certain stages

(Fig. 43).

The nuclear changes are also characteristic, and represent a
gradual transformation of the structure of the resting nucleus
of an ultimate spermatogonium. The reticulum changes first
into microsomal masses (Fig. 25, PI. II.). Then takes place a
flowing of these masses together (Figs. 34-39, 41-43, PI. III., IV.)
until all the basichromatic substance of the nucleus becomes
concentrated into a mass or karyosphere, and the particles re-
maining without the mass are oxychromatic. Figs. 44 and 45,
PL Y., represent the result of this process. Then follow stages
of dissolution of the karyosphere into minute granules, all of
which become gradually oxyphilic (Figs. 46 and 48), Fig. 49
representing a degenerate nucleus at the close of the cell's cycle.
During all these stages the nuclei become very irregular, with
deep indentations and Jobations at their margins and grooves
passing along their lengths. This irregularity of form and the
central karyosphere are diagnostics by which these nuclei may
be readily distinguished from those of neighboring germ cells.
Further, the nuclei do not remain at the basal end of the cell,
as they do in certain other mammals, but move out beyond the
level of the spermatogonia (Fig. 50).

After passing through the series of changes just described the
Sertoli cells degenerate, for there is no evidence that they go
through a second cycle. This is proven by the later stages of
these nuclei (Figs. 47-49) which gradually become wholly achro-
matic and then disappear from view. Their vacuolar substance
must at that time mingle with the fluid of the tubule. Fig. 25
(PI. II.) is interesting in this regard, for it exhibits a yom i^ Serioli

THE HI-MAX CELLS OF SERTOLI.

cell (A) pushing out before it an old and degenerate one (D).

A Sertoli cell i- therefore produced to every twenty-four -perma-
tid-, and alter the latter have metamorphosed into spermatozoa
and the-.- spermatozoa have become discharged from the tubule-.
that Sertoli cell de-enerates. Formation of Sertoli cells must
then continue through life as \«\\- as formation of germ cell-
continues.

\Ve now pa-- u» the hi-tory of tin- rod in the Sertoli cells, that
n markable body which differentiate- them from the functional
uerm cell-. Tin- i- at lir-i a -imple md. and may remain such
n in the bcjinniii'j enlargement of the- cell Fig. 25, 1'!. II.).
I)iil tin- rod di\ide-. and in in before the ^ertoli cell

IDS it- Drouth. - ; it- divi-ion are rarely found, and

tin- only on.-- oli-erved are illustrated in FU-. ?<> JO, I'l. III.
It \\ill l>e -eeii that in the case- of I'u-. jo j^ ;lu- rod i- under-
[ual longitudinal cleavage, a more -lender and -hortcr
rod .tb-lriciinv, from a portion of tin- larg< r on< ; this -mailer rod
may lie < ailed the primary rodlet. 1'erhap- one rea-oii \\liy
tli« are SO M-ldom found i- lu-cau-e tin- di\i-ioii tan 1 ie

•i i learly only \\ hen the rod lie> at a jiarticular an^le of \ i-ion.
\\ 'helher I imply an nmiMiallv ln-nt rod, or one

that i- in process of di\i-ioii. i- hard to determine, for it \\ a-
an i-ol.iied ca-e. The condition immediately follo\\ inu thi- di\ i-
>ioii i- -lio\\n in Fig. 31, \\ith an uniisiialK Ion- piiniar\ rodl.-t
(r. 1 1 i -01 1 1 pie 1 1 -I \- separated from the rod R.) ; this is a cell body
of the \oliuiie of that of an antepenultimate spermatogonium,
\\heie .n i "i<lini;l\- cytOplasmic di\i-ion h.id not otcurred, and
\\here the original inn leu- had di\ided \\hile onl\ one of it-
daughter nuclei had di\ided a^ain. .\ of rod and primarx'

rod let together at an unu-ually lat' i- n-pre-ented in I

P. ri. iv.

In four fifth- of tii -, in M out of KJO cell- examined, the

large rod conipleteK- di-appear- before tin- Serti>li cell Starts in
it- -routh and in -uch ca-e- only the primarx r..dlci i- to I n:
seen I igS. \2 J3, I'l. III. . Ju-t SO soon a- tin- cell enla:
a ]>aii- o| secondary rodlet- are seen in-lead of the primar\-
rodlet I ig. ; } . and uithout dotilit the-.- are produced by equal
longitudinal cleavage ol tin- primary rodlet. for they are alway- of

374 THOMAS II. MONTGOMERY.

the same length and lie close together. In their later stages
(Figs. 43, 46, 47) these secondary rodlets undergo some increase
in thickening, and in all cases these rodlets persist within the
Sertoli cell until the end of its cycle; they also probably degenerate
there, for no signs of them were found within the germ cells or
free in the fluid of the seminiferous tubule.

But in one fifth of the cases, 19 out of 100 cells examined,
the original rod continues visible for a shorter or longer period
after the secondary rodlets have been produced, as shown in
Figs. 35-30, 42, 45; and in Fig. 41 is drawn an unusual case of
late persistence of the rod and primary rodlet together. The
rod may persist for a while as a single dense body (Fig. 38).
Fig. 30 shows a case of such a single rod that has segregated into
chromatic and achromatic parts, a rare condition. But as a rule
it divides longitudinally as exhibited in Figs. 35-37, 41, 42, 45;
this division begins and is most prominent near the middle region
of the rod, when its ends may be still undivided, but cases were
found where the rod had completely divided into two secondary
rods (Figs. 35, 40, Fig. 40 being a rod from a cell of about the
stage of the cell shown in Fig. 30). In the instances where the
rod persists after the secondary rodlets have been produced, it
never stains quite as deeply as the latter, and gradually becomes
less and less chromatic until it can no longer be seen; no rod
was observed in any cell after the karyosphere of the nucleus had
disintegrated.

There is accordingly considerable individual variation in the
behavior of the rod after it has abstricted the primary rodlet;
in four fifths of the cells it then promptly disappears, in one fifth
it persists for a variable period, but never until the end of the
cycle of the Sertoli cell, and then undergoes a second longitudinal
division which is this time an equal division. The rod when it
persists generally remains in the basal portion of the cell body.
The secondary rodlets arc at first usually in contact with the
surface of the nucleus, either basal or distal, while they are later
found near the distal pole of the nucleus and usually sep.ir.iied
from it. Whether the disappearing rod contributes substance
to the formation of the fibers in the cytoplasm (Fig. 43) could
not be determined. It is also difficult to decide whether the

THE HUMAN CELLS OF SERTOLI. J] ;

roils and secondary rodlets are or arc not always enclosed in
vacuoles.

< e.;,,in aberrant cases need mention. In a single instance a
pair of secondary rod lets were found together xxith a rot! in a
penultimate spermatogonium FL. io),a precocious case of rodlet
formation; what the constricted acidophilic body in the cyto-
plasm of this cell mav be. ! do not know. Then in each ot t\\o
• i rather late Sertoli cell-, in-tead of tin- general case of one
pair of -econdary rodlet-. two pair- \\ere found Figs. 44, 48);
the-e might have been produced from an unu-ually long primary
t, -IK h as tin- "in- -ho\\n in Fig. .>!. by the occurrence of a
.1- x'.ell as a longitudinal divi-ion.

6. DlSCl --!M\ AMI CON< I.I SIGN.

Il i- triiK -nrprising that no i Ix m >ii'Ji account ha- \ et !

n ..f tin- hum. in cells of S-rtoli; indeed, the -tndie- made so
far are rather hi-tol. i^ical ihan e\ t> •!• rj.ic.il.

\tleiition in t! il- was tir-l draxxn bv srrt"li [865), \\ho

(ailed them "cellule r.i mi In at i . " M.i-t \\riter- -iuce his time
haxe given them hi- name; but the term "follirle cell" ...iiied

bv Valette St. George i- fre«nieinl\- employed. as \\ell a- the

t e| 111 "foot cell " ( J. I'".. S. Mii'H'e , \\ llile \ . K bile I '7 1 e|l)p|o\ e(|

t lie name -permatoblast , and I 'end.: : that of vegetative cell.

The name lullidc cell is generally u-ed for the cell- comp«i-iir^
the >permaii-iA>ts of iilMitebrate- and |.»\\er xertebrate-, ami
thai "1 ^ei'ioli cell for the physiologically corre-pniident cell-, nt
mamnialiam test

A- t" the -i netic relation- n| the Sertoli Cell- In the -erill cells

proper the \\ritei- I. ill into -rmip-, \\hich \\'.tlde\er '06 ha-
de-ign.ited as the dtiali-t- and the moni.-t-. The tir-l of il,

iid the t\\o kind- of cell- as of entirely diltCrent origin, the
-permaii'voni.i pi m ••eiliug from primordial germ cell- and the
cell- of ^ertoli from other element-. A- dna!i-t- are to be da--ed
\\ata-e. I', .:.|eleben, lieiida. \\'.llde\ er and Sieph.m. \Vata-e
'92) and Manleleben '07 . -m-ider t!:e ^ertoli cell- to be inier-ti-
tial ie-ii- cell- that haxe \\andered into ihe -eminiferoii- tubule-;
but liardeleben'- li-nre- are (jiiite indeci-i\e, and \\'aia-e. in his
x erx brief account of little over a page, drcxv hi- coiiclu-ion- from

376 THOMAS H. MONTGOMERY.

the similarity in color of the cells of Sertoli and the- interstitial
cells after staining with cyanine, chromotrop and erythrosine.
Though Bardeleben thus holds the two kinds of cells to be of
different origin, he nevertheless thinks that the Sertoli cells give
rise to "a rudimentary second form of spermatozomes." Benda
('94, '98), and Waldeyer relying upon him, considers the cell
of Sertoli to arise from the indifferent cylindrical peritoneal cells.
The dualists generally hold that a differentiated Sertoli cell re-
mains functionally active during the life of the individual and
does not regenerate more than the distal portion of its cell body;
and they are also of the opinion that the cells of Sertoli proliferate
themselves by division — amitotically, according to Bardeleben
and Stephan, or mitotically according to Benda. On the other
hand Prenant ('87), Schoenfeld ('01), Regaud ('99) and Bugnion
('06) consider both cells of Sertoli and spermatogonia to be de-
rived from one kind of cells, by a process of division of labor;
and this is in agreement with the results of most writers who
have studied the origin of the follicular cells of the ovaries and
testes of invertebrates — the follicular or nurse cell being generally
regarded as a modified germ cell. Yet Regaud and Stephan
('02) hold that the fully formed Sertoli cells proliferate germ
cells as well as nourish them; while Bugnion believes "the
primordial spermatogonium gives place to a plurinucleate plate
(part of the parietal synctium) which contains in a common cyto-
plasm spermatic nuclei and sertolian nuclei," after which the
germ cells delimit themselves from the syncytium that remains
as a Sertoli cell.

My conclusions differ practically in their entirety from those
of the writers mentioned. In tin- human testis the cells of Sertoli
are of common origin with the germ cells, one out of every four
ultimate spermatogonia becoming a Sertoli cell. Sertoli cells arc
thus not differentiated from the germ cells merely in early fatal
history, but so long as ultimate spermatogonia continue to be
produced. A Sertoli cell of man once differentiated does not,
so far as I have observed, divide again, and consequently does
not give rise to germ cells; further, a Sertoli cell die- completely
after the spermatozoa that are associated with it depart Irom
its surface, and it does not persist to nourish a second ^em-ration of
spermatozoa. There being one Sertoli cell to every three detini-

THE HUMAN CELLS OF SERTOLI.

live ultimate spermatogonia there is necessarily one to every
twenty-four spermatozoa; accord ingly. in man the number of
-permaio/oa, spermatic bundle, associated with one Sertoli cell
cannot be "8 or 16" as Bugnion states.

But the point oi the greatest interest with regard to the dif-
ferentiation of the human Sertoli cell. i- thai it is determined by
the inclusion of a peculiar cytoplasmic rod, thi- rod tir-t .iri-inu
in the antepenultimate -pennatogonia. No such "Sertoli cell
determinant" ha- been made km>\\ n in any other object. In the
e of the dili« n •niiati-in of tin- oo-onia from the nur-e cell- in
the ovary <>i the beetle />v//.v</fv, SO \\ell de-cribed by ('liardina
rO] , and corroborateil by I >ebai-ieii\ '»> i . there is a remarkable
met hani-m of differentiation of the nur-e cells: here the cell-; that

are io become oocytes receive a cast-ofl n-ticular pan of the

inn It ii-, \\hile the cell-, \\hich lack thi- extruded mass become

nur-e cell- I \\ill be seen that thi- i- an entirely-different
pro. ess from that described In me for man. for in man the Sertoli
(ell- are tho-e that contain the differential in:; body.

The de\e|opment of the human Sertoli cell is dearly a very

beautiful case of -omatic differentiation. In fact, one may re-
gard the m u It ice] hi lar oivaiii-m a- ha\ in^ I \\ o period-; of -omaiic

diiii n niiation: the first \\hen tin- ti— ue cell- become differen-
tiated from the germ cells, and the second, when in the early mass

n ( elK the primordial ^onad. the Sertoli cell- become dif-
ferentiated from the germ cell- proper. For the Sertoli cell-
ma\ piopeiK be classed as the soma of the testis.

Nothing like the rod that differentiate- the human Sertoli cells
from the other ultimate spermatogonia -< em- to be known in any

other case of somatic differentiation. In the classical case <>f

. di-co\ered b>' lio\eri, pm-pecti\e bod\ cell- ca-t off into
the c\ topla-m the end- of their chn>mo-ome-. In cn|»e|)od- and
insects, acconlin- to I lacker and >il\ e-lri n>pecti\ ely. a nucleolu-
or a mass "I nucleolar substance thro\\n out from the -erminal
V( icle of tin 'ine> ultimateK to lie in cell- of the germinal

cycle. The origin of the rod of the human S.-rtoh cell- I c«.iild
not determine, be\ olid that it i- lir-t apparent in the c\ topla-m
of the antepenultimate -permatogonia, and that it probably
form- then- during the re-i -tage of the cell-. It conn-, to
dc\elop in all antepenultimate -permatogonia, theref-ire. before

3/8 THOMAS H. MONTGOMERY.

the distinction of Scrtoli cells and germ cells: it becomes trans-
mitted without division to one quarter of tin- ultimate spermato-
gonia, and that quarter transforms into Sertoli cells. Under
these conditions, on account of the precision of the process, this
rod must be regarded as a Sertoli determinant, and as a cyl<>-
plasmic and not a nuclear determinant. Whether the rod, or its
substance, emanated fn the first place from the nucleus, can be
determined only by some fortunate observer who has more and
better fixed material than was in my hands. But there is no
reason to regard it as mitochondrial, as a chondriosome, because
granular mitochondria have been described in mammalian Sertoli
cells by Benda and others; in my material no mitochondria were
seen in the spermatocytes and spermatids, they were evidently
dissolved by the action of the fluid of Zenker, and it is therefore
probable they were dissolved also out of the spermatogonia.

The rod that comes to determine the Sertoli cells increases in
size while in the cytoplasm, becoming most voluminous in the
ultimate spermatogonia; outside of the nucleus, also, occurs its
process of abstriction of the primary rodlet and the division of
the latter into the secondary rodlets. It is therefore clearly an
extranuclear determinant of the Sertoli cell; and this as yet
unique process of somatic differentiation seems to be controlled
by an extranuclear body.

It has not been my intention to decide upon the function of the
Sertoli cells. They increase greatly in size to produce a syncytial
mass loaded with intracellular droplets, probably of fatty nature;
they envelope closely the rapidly growing spermatocytes and
for this reason they are generally supposed, and probably cor-
rectly so, to nourish this generation of germ cells. The fluid
within the seminiferous tubules contains, so far as I have ob-
served, neither erythrocytes nor leucocytes, therefore is probably
derived from the droplets of the Sertoli cells and not from the
blood scrum. The spermatids at the commencement of their
histogcnesis lose their first connection with the Sertoli cells, while
the nearly mature spermatozoa exhibit their heads buried in tin-
substance of the Sertoli cells; the latter is then a second orient. i-
linii of the germ cells to the Sertoli cells, one that cannot subserve
nutrition, for the developing spermatozoa do not inrre.i-e in
size, but which is rather, as Loisel ('07) has shown, the r\piv— ion

THE HUMAN CELLS OF SERTOLI. 379

of some chemico-tactile response. It may then be the Sertoli
cells fulfill three functions: to nourish tin- spermatocytes, to
furni>h the fluid within the seminiferous tubules and to attract
the spermatozoa into oriented bundle-.

It is certain that much more study i- needed of the Sertoli rell>,
both from the standpoint of somatic differentiation as \\ell a-
that of the physiology of the germ cells them>el\ < •-.

I.ITKRATrRK LIST.
Bardeleben, K. v.

'97 I: • n Ili-tologie des Hodens inn I /ur Spi-nna: bciin Mi-n-

-. li'-n. Arc h. Anal. 1'liy-. An.it. Alith. Su;

'98 Weitere !• :m Mi-n-ch.-n. Ji-n.i. /<-it.. si-

Benda, C.

'94 Anat"iuii- ILiii'llnK-h <ler Hani- uml

I dp

'98 I eber 'li.- S|» -MI. • ten ini'l h..lierer Invort'-l'iati-n.

i . An li. An.it. I'liy-;. Al«th.
BuRnion, E., et Popoff, N.

'06 la i] nili. atidi; Hibl. \ i\ I'

Debaisieux, P.

'09 I .-- ,1. !,i,-.'
Ebner. V. v.

'7- i ber den Bau der £

llll<l l.l-llli \li-II-.ln-Il. Ki'llclt I'!:'.-, lli-t.

Giardina, A.

'01 Originedel • 'lul<- nutiiri n.-l I < '. n.a. Mmuiix In.

. 18.
Loisel, G.

'07 ( ••ntiiliut

Mi in :iil>iyiil. I' i

Prenant. A.

'87 1 i . '•' l'.iii.->.

Regaud. C.

'99 SIM !.i in. • Anal i '

Schoenferd. H.

'oi I B taui. an.

Sertoli.

'65 I vir i • Mi ]i.n ti' iil.n i cellule ramil i -ainifere

<lc!l i. II Mori;av:iii.

Stephan, P.

'02 1 .'. \ ..Iiitii>n (!«• la eelli!' Sen - < . K I1 vj.

Waldeyer, W.

'06 \i\- • len. Hi-itwi'.:'- Haivlhiah <li-r I-!ut\v. \\'irl'i-ltii:i<-, I.

Jei
Watase, S.

'92 11,. i1::. •: ol the Sell ill's • • 11. .\in--i. Nat..

THOMAS H. MONTGOMERY.

EXPLANATION OF PLATES I-V

All figures have been drawn by the author with the camera lucida at the level
of the base of the microscope, and reduced one third in size in reproduction. Fig. 50
was drawn with Zeiss obj. C, ocular 12, all the others with the apochromatic im-
mersion objective 1.5 mm., ocular 12.

The following abbreviations have been employed:

Id., idiozome.

R., rod.

r. i, primary rodlet.

r. 2, secondary rodlets.

S.C., Sertoli cells.

Sp.G., ultimate spermatogonia.

PLATE I.

FIGS. 1-3. Entire antepenultimate spermatogonia.
FIGS. 4-8. Rods of antepenultimate spermatogonia.
FIG. 9. A binucleate penultimate spermatogonium.
FIGS. 10, ii. Penultimate spermatogonia.

BIOLOGICAL BULLETIN, VCL. XXI.

PLATE I.

o 4

p

No

x-~

7

. >— «.

A'

-7?

A'

T. M. MONTOOMFBT

382 THOMAS H. MONTGOMERY.

PLATE II.

FIGS. 12, 13. Penultimate spermatogonia in divis-'on.

FIGS. 14-16. Rods of penultimate spermatogonia.

FIG. 17. Quadrinucleate ultimate spermatogonium.

FIG. 1 8. Binucleate ultimate spermatogonium.

FIGS. 19, 20'. Ultimate spermatogonia.

FIGS. 21-24. Rods of ultimate spermatogonia.

FIG. 25. An incipient Sertoli cell (A) next to two ultimate spermatogonia
(B, C), and to a degenerate Sertoli cell (£>). The inner margin of the wall of the
seminiferous tubule is at the left.

BIOLOGICAL BULLETIN, VOL.

PLATE II

21

T. M. MONTGOMtRY

3 §4 THOMAS H. MONTGOMERY.

PLATE III.

FIGS. 26-30. Primary rodlet abstricting from the rod. early Sertoli cells.
FIG. 31. Trinucleate ultimate spermatogonium.
FIGS 32, 33. Early Sertoli cells with primary rodlets.
FIGS. 34-36. Secondary stages of Sertoli cells.

BIOLOGICAL BULLETIN, VOL. XXI.

PLATE i::

33

\

29

28

r i

r i

R

t

//

r J

386 THOMAS H. MONTGOMERY.

PLATE IV.

Succeeding stages of Sertoli cells arranged in the order of the nuclear changes
Fig. 40 exhibits a dividing rod of a stage similar to tha of 41.

BIOLOGICAL BULLETIN, VOL. XXI.

PLATE IV.

— R

^ )

_ / I

r J

r 2&--

r / -

/

T. H. MONTGOMERY

388 THOMAS H. MONTGOMERY.

PLATE V.

FIGS. 44-47. Later stages of Sertoli cells, Figs, 44 and 47 being oblique trans-
sections.

FIGS. 48, 49. Degenerate nuclei of late Sertoli cells.

FIG. 50. Portion of a section of a seminiferous tubule. Uppermost is the wall
of the tubule, and next to it a layer of ultimate spermatogonia. The syncytium
of the Sertoli cells is expressed by dark shading, and thrir nuclei an- ili-.iin^ui-liahlt1
by their angular form and central chromatic body. The other cells shown are
chiefly early spermatids.

BIOLOGICAL BULLETIN, .OL. XXI.

PLATE v.

II

R -'
\

.

\

«•-•«.

•. - »

f

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T. M. MONTGOMERY

ON THE CAUSE OF AITOTOMY IX TTBULARIA.

OSCAR RIDDLE.

In the cour-e of studies on the oxidizing and reducing; poxxer-
of ilir xariou- ti--ue- and body re-ion- of h\ dromedu-a1 the
phenomenon of .uUotoniy \\a- ob-erx ed to occur so -enerallx-
and SO rapidly in Tidndar'm as to invite attention to it- cause.
That Tnhulariti \- capable of aiiioiomy /. e., of the -elf-dix i-ion
of it- bodx ha- Ion- been known, the process haxin- been ob-
-erxed b\ (liard. Loeb, Drie-ch. Morgan, and other-. Two in-
vestigators ha\e definitely -on-lu in determine thi of the
phenomenon. The conrln-ion- »\ the-e two worker- seem, ho\\ -
i-\er. not to be in accord. < "lodleu.-ki1 maintain- that degenera-
tion of i he h\ dram 1 1 precedes and condition-, the autotomy, so

that il i- a <l>>. •• lie rdtcd ludranth that i- -e\etvd from the body.

Mo -\\ho seems unfortunately to ha\e o\erlooked ('.odiew-
-ki'- pajier \\ritin.u in this journal -tate- that antotoiny ma\r
Mr \\itlioui an initi.il de-eiu-r.it ion in the ludranth, if one
i an ind-e of ihi- liom hi-iolo-i( al examination.

M \ o\\ n experience with autotomizing 'I't-.-ndnria indicate- thai

\\lu-ii thi- ]n is effected \er\ -lo\\l\ and -^radnalK', .1- i-

orilinarilx the « ne can c.rlainK -ometinu- lind. as did

< ,oillc\\ ^ki, that before the actual -e|iaration of ihe lixdranth

the latter ha- undergone -iderable degeneration. ( h\ the

other hand. I ha\e had main aiitoi. >mi/ed Indranth- to !i\e in
a|>]i.irentl\ p«-rfect condition for three and lour day >. < .odle\\ -ki
hiin-el! noie- one Mich hydranth \\hich li\ed lor t\\o day-,. It
i- quite easy, tOO, to confirm Mor-e's -tatement that a ri-c- in
temperatiiu- tax or-, the occurrence of autotomy. Neverthel<
the peculiar conditions under which I \\a- able to obserx e the
aiitotomx conxinced nu- that neither of the aboxe mentioned
Mil)po-ed causes, nor xet both combined, i- the immediate or

' .i«!li-\v-ki. 1" . "/in Ki-niUin- del K'-v;iiLitii'r. i I :i>> iiltiriii

•:u»l." l\.-:i\' ArchtV, \'-l. I S, I')04.

'Morse, M the II>ai.nuh -i 'I'u'-ulnria."

Hei i i i IN. \',.l. if.. 1909.

389

390 OSCAR RIDDLE.

adequate cause of autotomy. I therefore gave a little speci.il
attention to this subject, the results of which may be summarized
here.

The particular experience which seemed to contravene the
proposed causes as being tin- actual immediate cau>e \vas the
following: If normal healthy indi\ iduak of T. mesembryanthemum
be held by the lower stem or stolon and drawn through any one
of a variety of solutions — sodium tellurite, sodium selenite, etc.—
complete autotomy may occur in less than one minute! Clearly
degeneration is not the cause of the autotomy in these cases.
Other members of the colony taken from their moorings and
placed in vessels supplied with fresh sea water remained for
days without autotomy. When some of these wen- similarly
drawn through pure sea water they remained intact without
autotomy. It was found, moreover, that the autotomy likewise
occurred even when the animal was dipped into a solution of
NaoTeOa, NaSeO3, etc., of lower temperature than that from
which it had just been removed. Here, too, the autotomy was
rapidly and decisively effected. In these cases the autotomy
plainly could not have been caused by a rise in temperature; the
temperature change actually being in the opposite direction. In
many cases the animals were removed from water at 16° C.
and drawn through a solution at 13° C.

In order to study the changes occurring in the rapidly air
totomizing animals these were examined with a Zeis^ binocular
while being drawn through the solutions. By this means it was
found: (i) that as soon as the animal touches the solution there
follows a very strong contraction of tentacles, hypostome, peri-
stome, etc.; (2) that the "neck" region becomes extremely con-
tracted and narrow, and apparently so much weakened as to be
unable longer to support the weight of the hydranth; or rat la r
too weak to sustain the slight pull on the hydranth as it is being
drawn through or lifted from the solution. The appearance hen-
is such as to indicate that the contraction of the circular libers
of this region is of sufficient force, not only to close completely
the central channel, but also to separate and cn»\\d out. many
entodcrm cells, and likewise to weaken their o\\ u adhe>i<>ii and
that of the other ectoderm cells to each other. .1 ri^onms con-

ON THE CAUSE OF AUTOTOMV IN TUBULARIA. 39 I

traction in response to stimulation therefore seems to be the ef
muse of antotnmy.

It ha- lieen po— iMe in a few instances to ;^et a beautiful demon-
:iion of tin- -tn-n-ih of the contraction in the "neck" re-ion.
It .1 i iilmlarian In- found with the gastro- vascular cavity of the
hydranth well expanded and full, it fan sometime- 1 >e induced—
I)V prii -knitf the peri-toim — to contract tin- peri-tome lir-t . and
thu- retain the \\ hole of the tin id of the cavity. In this condition
the hvdranth -omewhat re-eml>!e- a ruM>er hall: ihe channel to
the ont-ide In-ill- c!o-ed 1 ,y the c. MI i meted peri-tome, and the
po-ti-rior < 01 mm i.t tion with tin- ca\ ity of the -tern or l>od\ U-in-
interrupted 1>\ the above-mentioned coniraction of the "neck"
on It iiou . \\iih an appropriate Mum instrument, piv— ure
l><- liroii-lii lo IPIMI- upon tin- ^phere. and the u hole |nc,, .-cdin-
earrii-d out iindii the I >inn< ular. one <MII \\ at ch e\ er\ t hin- and
gauge \\ith one's "\MI mu-cle- the -tren-th of the coniraciion
in the peri-tonn- and "ii<-ck." In the in-tame- \\hen- I ha\e
< arrii-d »m ihi- experiment / //<KT n< >n iihlc tn force the

<>/><•>! />i\' ni tin- t humid in the nc< k re-jinn. Some part of the
h\ pOStOme \\all i- t he tir-t to break.

It is, too, this ven strong contraction thai carrie- the caenosarc

ot the ;/, <>n (|iiite a\\a\ from the peri-arc see I ig. o).

l'io|iaM\- tin 'ii that the point <.) the aut«ilom\ i- al\\.i\-

!elinitel\ loc.ili/ed in tin- "neik" re-ion as ua- tir-t re<
ni/e<l \>\ i .iard —is that tin- remainder of the -lender portion of
the animal i- i o\ ered uith a chitiiioii- peri-arc \\hich i- r.ither
impermeaMe and hi'^hK pn>tecii\e a^ain-i -timuli.

In "normal" cases ihi>-e in \\hich the process • <\ aniotmiu'

xtended over a period o| several hour- or ,1 «|,i\ or t \\ o — it
i- \\ell knoun that a \ei\ complete histolysis of the cell- iii the
"neck" re-ioll OCCUrS. I'llele i- gOOd e\ idelice lloU e\ er that

ill the-i cases, i"". the hi-iol\-i- i- preceded l>y a rather strong
or 1>\ a prolonged ci>ntra< lion of thi^ region. In \ er\ \\eak -olu-
tion- of tellurium and -eleiiium -alt-, ot acid- and alkali-, and
e\eii |i\ \\atchtnl mechanical stimulation of the animal into
coiilimioll-l\ coiltrai led -late. I ha\e lieen aMe to effect I he

Giard, \ . "I.'Aui"t"iiii.' il.in • aiiinmlf." R ';//'/•/«.•. p.

392 OSCAR RIDDLE.

autotomy of healthy hydramhs more gradually in period- ex-
tended to one to four hours, and \» watch the course of the
process in a single individual during thi> time. From these
observations I may here record one or two point- \\hi.-h -eem to
throw some light on the reason for the histolysis just mentioned.

I cite the case of a tubularian which \\.i- kept mechanically
stimulated by touching or pricking tin- hydrant h with a dissecting
needle and in which the process of autotomy had advanced at
such a rate as readily to separate at the end of four hours when
lifted from the sea-water. With the beginning of contraction
in this animal the circulatory current in the "neck" region was
stopped; indeed the circulatory-nutritive fluids were quite ex-
pelled and excluded from approximately two millimeters of this
region; the entodermic walls of the tube here being completely
and tightly apposed. The closure at this point also largely
stopped for a time the circulation through the stem. The fluids,
however, were as before in contact with the walls of the gastro-
vascular cavity everywhere except in the much contracted neck
region. That is to say, in the contracted animal the normal
nutritive fluids were in contact with all the structures with which
they are normally in contact except at one point — the "neck"
region; it is always at this latter point that histolysis and autotomy
later occur.

In a little less than an hour it was found that the dissepi-
ment which divides the gastro-vascular cavity of the stem into
two channels — one for the anterior, the other for the posterior
flow of the circulatory fluids — had been broken at a point a little
below the contracted "neck," and that the usual circulation of
fluids was again established within the stem. Soon however
there accumulated at the point immediately below the neck a
quantity of the red pigment and other debris from the circulation;
thus the channel became so firmly plugged that even a relaxa-
tion of the contracted neck region could not now effect a ree>tal>-
lishment of circulation in this "neck" region.

It is my opinion, furthermore, that the reestablish ment of this
circulation after a few hours of contraction might be prexciiied
or at least greatly hindered by another circumstance if for
any cause the debris just mentioned should fail io collect and

ON THE CAUSE OF AUTOTOMY IX TUBULARIA.

393

act as an effective block. I refer to the accumulation of a
gelatinous mass between the pi-ri-arc and coenosarc which begins
to be secreted by the coenosarc soon after it pulls away from its
contact with the perisarc. The secretion is perhaps of the nature

// Y /).—

^ c.c.

.

•

<„/.< .

!<• pi. -entin- tin- oni'litim; i> in a tubularian. Such om-

ilitioii< .in- present in J':<!':iLiriii which have licen k.-pt -tiinulatoil tn/m one to
several li.-ins. //\ .' h\.li.mt!i; B.D.=break in .li-epinient; C.C. =c<rnosarc
"i linily \\.ill; 1>. --deliii- leit 1-v eiii ulalini; current; (./'. -.nelatinnti- n

:vity of body; /'. |ifii
-ti.'ii.^ly o>tur.. .1 ="ncck."

of iiKilcrial for .1 iir\\ pn i-.ii'r. .uul as noU-d 1>\' < lodlcu -ki it
i- mi r« mi act \\ ith \\atcr. The accumulatii >n and hardening

394 OSCAR RIDDLE.

of this mass would probably make a reopening of the closed
channel very difficult or quite impossible.

\Ve see then that the contraction of the contractile parts of
Tnbnlaria acts differentially upon its various organs. Such con-
traction does not rob the hydranth of its contained fluids, nor
of its ability to circulate these fluids. The same is true for the
stem region, except that there the actual movement of the fluid> i-
in abeyance for a very short time. The contraction which occurs
in the neck region, however, brings about far different relations
between the contracting area and the nutritive medium. Here
there results, not only a cessation of the circulation of fluids, but
a complete loss of contact with these fluids following the complete
closure of the channel; whilst finally the breaking of the dissepi-
ment immediately below the neck region, and the subsequent
plugging of the end of the connecting channel with pigment and
debris, preclude the possibility that such contracted region may
again regain its circulation together with the food it brings. This
area then necessarily disintegrates; and the break — the autot-
omy — necessarily occurs at this the weakest point.

Our conclusion is that autotomy in Tubularia is the result of
the contraction of the animal; similar but weaker contractions
being common and central features in the behavior of the animal.
If the contraction be either too strong, or too much prolonged,
autotomy will follow. That is to say, if a very slight strain be
put upon the "neck" region while its circular fibers are in a state
of extreme contraction separation results at once. It the con-
traction be not so strong, but considerably prolonged, readjust-
ments are effected in the circulation which prevent the ingress of
food to the contracted "neck" region. Degeneration now occurs
in this region and the break — the autotomy — follows at this same
point. Then- is, then, no great mystery attached to "ramputa-
tion spontanee " ; not even a complex organic correlation to direct
a watchful and sacrificial neck in severing an offending head
from an unoffending body.

The present work has been done at the Zoological Si.it ion at
Naples while occupying a table supplied by the Carnegie Institu-
tion of Washington. To the president of the Oirnrgir Institu-
tion, and to the director and assistants of the Zoological Station,

OX THE CAUSE OF AUTOTOMY IN TUBUI ARIA. 395

I am much indebted. My best thanks are due to Professor
Paul Mayer, of the Zoological Station, for special apparatus and
for very main- kindnesses. For other support of this work I am
further indebted to the Laboratory of Experimental Therapeutics,
The University of Chicago.

NAPLES,

March, 1911.

MBL WHOI LIBKAJn

UH 17JS I