The Cytology and Binary Fission of Peranema.By

Virginius E. Brown,Zoological Laboratory, University of California.

With Plates 19-21 and 1 Text-figure.

CONTENTS. PAGE

I N T R O D U C T I O N . . . . . . . . . . 4 0 3

M A T E R I A L A N D T E C H N I Q U E . . . . . . . 4 0 4

M O R P H O L O G Y 4 0 5

M I T O S I S 4 1 0

P r o p h a s e 4 1 0M e t a p h a s e . . . . . . . . . . 4 1 1A n a p h a s e . . . . . . . . . . 4 1 1T e l o p h a s e 4 1 2

D I S C U S S I O N 4 1 2

S t a b o r g a n . . . . . . . . . . 4 1 2K i n e t i c E l e m e n t s 4 1 3C y t o p l a s m i c I n c l u s i o n s . . . . . . . 4 1 5

G E N E R A L S U M M A R Y . . . . . . . . . 4 1 6

R E F E R E N C E S . . . . . . . . . . 4 1 7

E X P L A N A T I O N O F P L A T E S 4 1 8

I N T R O D U C T I O N .

THE Peranemidae are animals of a primitive type and theyshow rather close phylogenetic relationships to the lower pro-tistan forms. P e r a n e m a affords good cytological material.They have a clear cytoplasm and therefore their kinetic elementsare easily determined. Likewise they are easy to culture.Thereby they can be obtained in quantities large enough toallow of good cytological investigation. Also the study of theircytology is of interest, for the conception of the Protozoa asa primitive type has led to various studies which may bring tolight a better interpretation of the riddles of protozoan mor-phology.

404 VIRGINIUS B. BEOWN

The divergence of opinion among those who have written onthe various flagellates belonging to the Euglenoidina has pre-sented many interesting problems. The kinetic elements, thegullet, and the reservoir system of P e r a n e m a are subjects ofcontroversy. The purpose of this paper is to clear up the variousinterpretations that have been made on the morphology andmitosis of this protozoon. It is also my aim to point out thephylogenetic relationships of the neuromotor system.

This work was done in the laboratory of Dr. Eobert C. Ehodesat Emory University. I wish to express to Dr. Rhodes manythanks for his valuable criticisms and general co-operation.Thanks are due also to the University of California for supplyingthe optical equipment used in completing this investigation.

MATERIAL AND TECHNIQUE.

I have successfully cultured P e r a n e m a t r i c h o p h o r u min the following manner: A 200 c.c. culture of E u g l e n ap r o x i m a was centrifuged and the material thus obtained waswashed in distilled water and crushed between two glass slides.To this crushed E u g l e n a I added 100 c.c. of tap-water andallowed the culture to stand for a day before I inoculated withP e r a n e m a t r i c h o p h o r u m . After eight days divisionforms were observed in abundance.

For fixation, the centrifuge method of killing was found to bethe best process as it ensured large quantities of the material.Various fixing reagents were used, but the best for general pur-poses were found to be hot Schaudinn's and strong Plemming'sfixatives. I used the fixatives of Champy and Mann-Kopsch fordemonstrating mitochondria. The most satisfactory nuclearstain was found to be Heidenhein's iron-alum-hematoxylin.Counter stains of eosin or Bordeaux red were used to demonstratethe axial filaments of the flagellum. ' Licht Griin ' was used toshow the spiral striations of the cuticle. The mitochondria werestained with fuchsin and toluidin blue after Champy's fixative,or they were impregnated with Bowen's modification of theMann-Kopsch procedure. Then I stained with safranin to bring

PERANEMA 405

out the nuclear structures. The method of Bowen likewisebrought out the Golgi apparatus.

The plates were sketched by the aid of a camera lucida froma Busch and Lomb binocular microscope with 2 mm. apro-chromatic objective and x 12-5 oculars.

MORPHOLOGY.

The genus P e r a n e m a is characterized by the lack of an eye-spot. It is spindle or cigar-shaped, and it tapers anteriorly toa point. Its broad posterior end is truncate or retuse.

The size of P e r a n e m a t r i c h o p h o r u m is 60 /x to 72 yx inlength and 28 /x to 32 /x in breadth. It has a thin flexible cuticle ofa metabolic nature. A single flagellum occurs, and it possessestwo axial filaments which arise from basal granules situated atthe base of the reservoir. The length of the flagellum variesfrom one-fourth to almost body length.

The genus is likewise characterized by a ' crawling' method oflocomotion. Here the contour of the animal assumes variousprotean forms by means of a series of wave-like contractions andcontortions of a fugacious character which run along the cuti-cular surface. These are clearly apparent when the animaldoubles on itself by an abrupt retrogressive motion, which issimilar to that described by Mast, 1922. The flagellum is usuallykept straight forward and rigid, except the tip, which is bent intothe form of a hook. This bent portion vibrates in a continuouswhirl. The point of the flagellum when in motion describes anellipse. P e r a n e m a g r a n u l o s a often swims by whirling theentire flagellum around and hurls itself backward.

The vacuole system is complex ; it consists of a large flask-shaped vesicle (= Hauptvacuole of Klebs, 1863) ; also a smallcontractile vacuole is situated at the base of this structure, whichI prefer to call the reservoir after Ehodes, 1926 ; Baker, 1926 ;Calkins, 1926. At each systole the contents of this adjacentcontractile vacuole (= Neben vacuole of Klebs, 1863) is emptiedinto the main reservoir. A slight contraction of this main reser-voir seems to aid the flagellate to reach the substratum ; sowe suggest that its function is not only an organelle for the

406 VIBGINIUS B. BROWN

storage of waste materials, &c, but also that it is of a hydrostaticnature.

I believe that the old family Peranemidae (Stein) should stand,because all of the group, with the exception of E u g l e n o p s i s ,

TEXT-FIG. 1.

neck of reservoir

cytostomzrodorganor Siaborgan

ayiaL FiLamentcentrobLepharoplastreservoircontractile vacuoLeGoLgi bodies

endosome

mitochondrium

FLac/eLLum

Food vacuole

siriations oncuticLe

I- GoLgi network

A diagrammatic sketch of Peranema tr ichophorum.(Note that the crescent-shaped Golgi bodies shown around the reservoir

are not present when the Golgi network occurs.)

possess a rod-organ, or ' Staborgan '. Hall and Powell were un-fortunately not informed of this fact. The rod-organ of P e 1 a t o-ruonas is small and the lateral rods are quite short. Also

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Scy tomonas , T rop idoscyphus , Mar sup iogas t e r ,and Dinema have well-developed rod-organs. In Urceolusthe rod-organ is spread out into an urn-like structure ; whereasthe rod-organs of Anisonema and E n t o s i p h o n extendposteriorad, as an internal' siphon ', almost the entire length ofthe body. I see no reason, therefore, to split up an old familyof Peranemidae into several families which have one or moreflagella. Shaeffer (1916) stated that the Peranemidae are holozoicand ingest solid organic matter which is composed of pieces ofplant or animal tissues. I wish to add that they exhibit greatselectivity with relation to their food.

Pe ranema t r i c h o p h o r u m lives almost entirely on deadEuglena p rox ima , and has been observed to engulf E n t o -s iphon and Chi lomonas . The animal rarely if ever attacksimmotile forms, but it is content to tear encysted or deadEuglena to bits. On the other hand, Pe ranema g ranu l i -fera eats living Zoochlore l la . This type of food selectionis not uncommon among protozoa; for example, Didin iumfeeds almost entirely upon Pa ramec ium (Calkins, 1926). Itis quite reasonable to believe that Pe ranema is almost entirelyholozoic, because a culture will not live unless it is inoculated withsome Euglenoid. For this reason as well as the one men-tioned above, I do not think it advisable to place Pe ranemain the same family with the saprozoic Astasidae of Calkins (1926),but I prefer to accept the classification of Doflein and place thegenus in the old family Peranemidae (Stein).

Wager (1900) was the first to describe the insertion of aEuglenoid flagellum. He stated that the flagellum of Euglenav i r id i s bifurcated on entering the main vacuole (= reservoir),and that each of these strands was anchored at the base of thisvesicle to a basal granule. Later writers (Rhodes, 1926 ; Baker,1926; Eatcliffe, 1928) have suggested that these strands becalled axial filaments.

Hartman and Chagas (1909) believed that the flagellum ofPeranema was single and that it was anchored at the base ofthe main vacuole (= reservoir) by a basal granule.

They also stated that another short flagellum arose within

408 VIRGINIUS B. BROWN

this vesicle from a basal granule adjacent to the other one andtraversed the vesicle to become anchored to another granule onits upper surface.

Hall and Powell (1928) believed that the flagellum remainedsingle after entering the reservoir and that a new flagellumstarted to grow out of the reservoir early in mitosis.

Doflein figures P e r a n e m a with a single flagellum which hastwo axial filaments similar to those described by Wager (1900)for Eug lena v i r i d i s .

It is possible that Hartman and Chagas (1909), as well asHall and Powell (1928), misinterpreted the second axialfilament as a new flagellum of the daughter individual. I findthat P e r a n e m a has a flagellum which is formed by the unionof two axial filaments which arise from basal granules situatedat the side of the reservoir.

P e r a n e m a does not possess the granule which has beendescribed to occur on one of the axial filaments by Wager,1900 ; Baker, 1926 ; and Eatcliffe, 1927 ; however, in all otherrespects its flagellum is similar to that of E u g l e n a . It ispossible that this granule is absent in P e r a n e m a because itdoes not possess an eye-spot. Wager (1900) believed this granulewas associated with the eye-spot of Eug lena and that itaided in orienting the animal to light. Since this granuleis absent in P e r a n e m a it is possible that this statement iscorrect.

The occurrence of a cytostome has been detected by Carter,Clarepede, and James-Clark, but Klebs (1883) gave the firstdescription of its organization. He believed that the mouthopening was ventrally placed and that it was separated fromthe ' Hauptvacuole ' or reservoir. However, he failed to considerthe pharynx as a tubular structure, and he believed it was com-posed of two adjacent rods on the ventral side of the cuticle.These he termed the ' Stabapparat '.

Ehodes (1926) has shown that the reservoir of H e t e r o n e m aacus is a separate structure from the ' staborgan ' (Stabapparatof Klebs), and that it consists of three rods. The outer lip of thecytostome is bounded by a falcate rod, which he terms ' trichite '.

PERANEMA 409

He stated that the tube which leads from the mouth is boundedon each side by lateral rods.

I agree with Klebs (1883) and Bhodes (1926) that the' staborgan ' and the gullet of P e r a n e m a are not connected inany way with the reservoir, but that the cytostome is a separateopening on the ventral side of the body. The cytostome iscapable of great distension and its outer lip is reinforced witha heavy falcate rod. The inner or proximal' lip ' is protoplasmic.These rods of the ' staborgan ' probably act as a support tostrengthen the sides of the gullet and to prevent the prey fromtearing its own cuticle. The third rod (= falcate rod or trichiteof Bhodes, 1926) seems to act not only as a support to the lowerlip, but also as a trigger or valve which prevents the cytostomefrom opening except when the protozoan is feeding. It also aidsin holding its prey so that it can be pinched off by the aid of theother two rods which border the gullet (=Cytoesophagus ofEhodes).

I agree with Ehodes, 1926, that the rod-organ is displacedduring the early prophase and that it later disintegrates in thecytoplasm. New rod-organs are formed in the early anaphase.The new cytostomes are formed by an inpitting of the cuticle.At the base of this pit a granule is formed and out from it growtwo rows of granules. These condense to form the lateral rodsof the gullet. (I prefer to use this term because it has been usedextensively in protozoology; however, it should not be confusedwith any part of the reservoir.) During the telophase the thirdor the falcate rod forms by an outgrowth from the top of themedial lateral rod (fig. 11, PI. 20 ; fig. 12, PI. 21).

The mitochondria of P e r a n e m a vary from a small roundform to a large disc-shaped type with a clear centre. The smallspherical forms range from | ^ to \ fj., whereas the disc-shapedtypes vary from \p to lfi in width and are £ju to J/M in breadth.They are usually grouped around the nucleus and the base ofthe reservoir. When the protozoan divides most of the mito-chondria move anteriorly along with the nucleus and groupthemselves about the reservoir. The ovoid mitochondria seemto be capable of becoming the disc-shaped types by growth.

410 VIRGINIUS E. BROWN

These disc-shaped types show a differential staining with thetoluidin blue and fuchsin method. The core is magenta, whereasthe cortex is dark blue. The small mitochondria often showconnexion^ or form dumb-bell structures. These are in allprobability division stages. The mitochondria are straw-colouredwhen stained by the Mann-Kopsch method, whereas the Golgiapparatus is black.

The Golgi apparatus of P e r a n e m a is a network of long inter-woven fibres which are concentrated in the posterior portion ofthe animal (fig. 15, PI. 21). This network is not as dense in thelater division stages of P e r a n e m a as it is during the earlyprophase. Neither the reservoir nor the contractile vacuole takesan osmic impregnation. However, the Golgi apparatus formsspheres around the reservoir during the early prophase.

MITOSIS.

Prophase.—During the early prophase the nucleus ofP e r a n e m a t r i c h o p h o r u m migrates anteriorly to the baseof the reservoir. The nucleus at this time is of a vesicular nature ;its centre contains a large dark staining body of chromatin.This body may be single or fragmented (Hall, 1926). Thisendosome (= Binnenkorper) is filled with vacuoles of varioussizes. No centriole occurs within their centre, as some writershave believed. Therefore I agree with Hall and Powell (1928)that these vacuoles are of no significance. In the early prophasethe dispersed chromatin comes together to form very thinchromomeres. These chromomeres are formed from 'spherules'which are connected together to form loops. Within thesechromatic structures one can easily discern basophilic granulesof variable sizes. During the growth of chromatin these loopsthicken and the nucleus moves anteriorly and comes to lie onthe edge or the base of the reservoir. By continued anteriormigration it comes in contact with the largest basal granule(fig. 4, PI. 19). This granule or centroblepharoplast seems to pullthe nucleus upward and strands can be traced from it into theendosome. The endosome begins at once to elongate and thenucleus swings around on its axis until the other portion of

PEKANEMA 411

the elongated endosome is connected by similar lines to the otherbasal granule. The chromatin loops then thicken and arrangethemselves parallel to the elongated endosome. At this periodthe longitudinal splitting of the metaphase occurs. The chromo-somes separate from only one end to form V-shaped structures(fig. 5, PL 19).

At this stage a polarity is established between the oppositecentroblepharoplasts at each end of the nucleus. Also fibres arenoticed connecting each centroblepharoplast with the end of theelongated endosome (fig. 6, PL 19). The nucleus then pushesup against the reservoir. This not only distorts the vesicle butalso it pushes the rod-organ out of position (figs. 5, 6, PL 19).The rod-organ is thus cast out into the plasma of the animal,where it disintegrates before the end of the anaphase (figs. 5,6, PL 19, and fig. 7, PL 20). This disintegration of the rod-organhas been described in H e t e r o n e m a acus by Rhodes, 1926.

During the prophase the basal granules divide twice to formnew basal granules ; out from these grow axial filaments. Eachof these filaments unites with one of the old axial filaments toproduce a new flagellum. A part of the nagelluni thus persiststo form one of the newflagellar filaments of a daughter individual.The chromosome count at this time has been estimated to bethirty-two.

Metaphase.—The longitudinal splitting of the chromosomestakes place during the prophase when the chromatin is arrangedin a group of granular chromosomes. These pull apart to formV-shaped chromosomes (fig. 4, PL 19). These V-shaped struc-tures widen and contract into chromosomes which come to lieparallel to the endosome (fig. 5, PL 19). Such a situation isusually termed the equatorial plate. The nuclear membraneremains intact.

Anaphase.—The endosome continues to elongate and oftenvacuolates or splits longitudinally (figs. 8, 9, 10, PL 20). Thechromosomes during the early anaphase separate or pull apartfrom the last end which splits during the metaphase, and therebyan appearance of a pseudo-transverse splitting is noticed. Thechromosomes become more granular and group themselves at

412 VIRGINIUS E. BROWN

the ends of the endosonie ; the nucleus then shows a constrictionin the central portion (figs. 8, 9, 10, PI. 20). The attachmentbetween the endosome and the centroblepharoplasts persists.

During the anaphase the new cytostomes are formed byinvagination of the anterior cuticular surface. On the edge ofthis newly formed cytostome a group of four granules grows insize and later they collect into two distinct rod-like structures(fig. 9, PI. 20). These form the lateral rods of the rod-organ.The daughter reservoirs pull farther apart, and a partition beginsto form between them. This division of the daughter reservoirsis produced by an outgrowth from the base of the old reservoir.

Telophase.—The telophase begins by a constriction of theanterior end of the animal. This deepens and the animal beginsto split from the anterior end so that the animal divides bybinary fission. The continued constriction of the nucleus causesthe central portion to break apart. The chromosomes formgranular loops around the endosome (fig. 12, PI. 21). The thirdrod of the staborgan forms by an outgrowth from the top ofthe median rod. This connexion persists after the rod is formed;the same condition is found in H e t e r o n e m a (Bhodes, 1926).

Division is completed by the daughter individuals pullingapart (fig. 13, PI. 21). This action is very violent and the entireprotoplasm is kept in motion. The flagella grow out, and theyare kept in violent motion until the division is completed.

DISCUSSION.

R o d - o r g a n or 'S t abo rgan ' .—The Staborgan of P e r a -n e m a consists of the rods which I have described above. Thereis no evidence that these rod-organelles are parabasals, as Calkins(1926) and Hall (1926) believed.

In fact P e r a n e m a seemstouse them in feeding, and they areused just as a ciliate uses its trichites. The falcate rod opensand closes the cytostome when the animal feeds. Hall (1926)believed the parabasal to lie adjacent to the gullet and that theparabasal body doubled during division. Hall and Powell (1927)stated that they believed the parabasal body to be the equivalentof the 'Staborgan'. Later (1928) they ignore this statement

PERANEMA 418

(Hall, 1926) and refer to it as the pharyngeal rod apparatus.I object to the use of this term since the cytostome is not con-nected with the reservoir, hence it cannot be of a pharyngealnature. I suggest that the term 'Staborgan' or rod-organ (= rodorganelle) be applied to this structure since this term is found inall of the literature (Doflein, 1912; Ehodes, 1926). Also I havenot been able to find a parabasal in P e r a n e m a , and I see noreason to believe that the 'Staborgan' is either analogous orhomologous to the parabasal body. Let us consider this questionfrom a phylogenetic view-point. In the primitive protist,E u g 1 e n a, we find a homology between the parabasal body andthe kinetic complex (Baker, 1926). This kinetic complex is derivedfrom the endosome. Now in a higher form like P e r a n e m a itis reasonable to believe that such a body is either lost during theevolution of this protozoon or that it still remains within theendosome. Obviously the endosome is part of the kinetic mass ;I do not believe that P e r a n e m a has any homologue of theparabasal body, but I do believe that the endosome is thekinetic reserve mass.

Hall and Powell (1928) state that P e r a n e m a has only one' pharyngeal' rod element in the early stages of binary fission.This they believe suggests that one of the two original rodspasses to each of the daughter organisms. I have not noticedanything like this and I suggest that Hall and Powell studiedthe 'Staborgan' from a lateral view so that one would be abovethe other, and unless care is used this structure may appearas only one. The 'Staborgan' does not split during binary fission,but the old 'Staborgan' disintegrates and new ones form fromgranules on either side of the new daughter cytostomes. Thesegranules are possibly of mitochondrial origin.

K i n e t i c Elements.—Theblepharoplast-rhizoplast-centro-some complex is of a primitive type and has been described tooccur in many of the lower flagellates. Likewise we find sucha complex in most of the Polymastigotes and the Hypermasti-gotes. Associated with this centroblepharoplast-rhizoplast-centrosome complex, in the Polymastigotes and the Hyper-mastigotes, is a dark staining paradesmose. This structure

NO. 291 E 6

414 VIRGINIUS E. BROWN

connects the daughter centrosomes during fission (Kofoid andSwezy, 1915). I do not find such a paradesmose in P e r a n e m a .The kinetic elements of P e r a n e m a are of a type like that ofO c h r o m o n a s (Doflein, 1912). Here we have the centro-hlepharoplast, but no paradesmose occurs. During mitosiscentroblepharoplasts are connected to the ends of the endosomeby numerous spindle-fibres. No permanent rhizoplast exists.In fact I see no reason to believe that the ' blepharoplast-rhizoplast-centrosome ' complex occurs in P e r a n e m a ; thou-sands of granules occur around the nucleus, but these do notfunction as a centrosome.

Although the paradesmose is known to occur in the Hyper-mastigotes and in the Polymastigotes, I suggest that it does notoccur in any of the Euglenoid flagellates, because it has notbeen demonstrated by any of the recent workers on the Euglenoidgroup (Baker, 1926 ; Katcliffe, 1928).

After a due consideration of the behaviour of the centro-blepharoplast of P e r a n e m a during mitosis, I have decided thatthere is a relationship between the centroblepharoplast and theendosome. In other words, the action of the centrosome, orbetter, the centroblepharoplast, does not initiate mitosis alone ;but that this kinetic force is an interaction of both the centro-blepharoplast and the endosome as well as intranuclear physio-logical forces. If such a kinetic mass as the endosome is chargedby a type of ' mitokinetism ' or any type of electrostatic force,and if that force is associated with other forces which are ofa physiological nature, there will be balance between all theforces which may initiate mitosis. Now if this balance is upsetand a change in polarity occurs and the endosome is elongated,then this structure will split (fig. 8, PI. 20). In all probabilitythe same interacting forces cause the chromosomes to splitlongitudinally and to ' flow ' apart. If the endosome is the centreof this kinetic force, then it is quite reasonable to expect thata division centre (or even a kinetic reserve complex) is given offin some of the lower protistan forms. Otherwise it would behard to establish a phylogenetic line between P e r a n e m a andthe lower flagellates. Such division centres or kinetic reserve

PERANBMA 415

masses have been described by various workers (Kater, 1925 ;Baker, 1926).

The C y t o p l a s m i c Inclusions .—Hall (1928) describescertain cytoplasmic inclusions which he believes are mitochon-dria and Golgi elements. The mitochondria he states are smallelongated structures in P e r a n e m a and they lie in spiral rows.Likewise he noted numerous spherical inclusions similar in struc-ture to the ' Golgi' elements of higher animals. I find that themitochondria are rarely in spiral rows and that they assume thisposition for only a short while during the early prophase. It ispossible that Hall (1928) has confused the Golgi network withthe mitochondria, because at this time the Golgi apparatus isa network of long fibres. These fibres of the Golgi network aregrouped in spirals round the nucleus and round the base of thereservoir (fig. 15, PL 21). When P e r a n e m a is not in division,the Golgi apparatus is a network of long fibres arranged ina tangled mass. During the early prophase these fibres condenseinto a tangled network which lies in the posterior portion of theanimal. This mass breaks up into small irregular bodies whichgroup themselves round the nucleus during the metaphase. Butthe typical network of long fibres form again during the ana-phase and persist as such during the interkinetic phase. Themitochondria are spherical and disc-shaped (fig. 14, PI. 21).The large disc-shaped types are grouped heavily round thenucleus and the base of the reservoir. They lie deep within thecytoplasm, whereas the spherical forms are more superficial andoften take dumb-bell or rod-like shapes. The spherical formsare almost evenly distributed throughout the cytoplasm, whereasthe large disc-types are grouped together. These disc-shapedtypes quite often show a clear centre. Earely these disc-shaped types form ' roulettes '.

In the same slides with the P e r a n e m a material I foundE u g l e n a p r o x i m a and E u g l e n a g r a c i l i s . The mito-chondria are similar in shape and distribution to those I havedescribed in P e r a n e m a . This leads me to believe that themitochondria of E u g l e n a described by Causey (1926) are onlya part of the chromatophores and possibly the pyrenoids of this

416 VIRGINIUS E. BROWN

protist; however, I shall discuss the behaviour of the cyto-plasmic inclusions of E u g l e n a during binary fission inanother paper.

GENERAL SUMMARY.

1. P e r a n e m a t r i c h o p h o r u m is holozoic in nature andselective in its food, but not predaceous. It feeds usually ondead and encysted E u g l e n a p r o x i m a , E u g l e n a g rac i l i s ,and rarely upon C h i l o m o n a s and E n t o s i p h o n .

2. The 'Staborgan' or rod-organ is not connected with thereservoir, but it opens into the cytostome which lies ventrallyto this vesicle. Therefore the term gullet should not be appliedto the neck of the reservoir.

3. The chromosome count of P e r a n e m a t r i c h o p h o r u mis estimated to be thirty-two in number.

4. The 'Staborgan' is thrown out of its position during mitosisand it disintegrates in the cytoplasm. New rod-organs grow outfrom granules which form at the base of the new daughtercytostomes. These granules may be of mitochondrial origin.

5. A centroblepharoplast is described. No paradesmose ispresent.

6. A theory is suggested which supposes that an interactionbetween the centroblepharoplast and the endosome occurs.The centroblepharoplast acts as a kinetic attraction spherewhich carries the nucleus anteriorly in order that the blepharo-plasts can function as extra-nuclear division centres ; therebya co-ordinated interaction is brought about between both intra-nuclear kinetic elements and all of its cellular components. Sucha reaction or interrelation of parts is necessary to initiatecellular division.

7. The mitochondria of P e r a n e m a were found to bespherical; these may grow into large disc-shaped types withclear centres. The latter have a tendency to group themselvesround the nucleus and the reservoir.

8. The Golgi apparatus was found to be a network of longfibres. These Golgi bodies seem to be concentrated in theposterior end and round the reservoir. Neither the contractile

PEEANBMA 417

vacuole nor the reservoir was impregnated by osmic acidmethods.

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EXPLANATION OF PLATES 19-21.Pigs. 1-15 inclusive are of Peranema tr ichophorum.All figures were drawn with the aid of an Abbe camera

lucida. Figs. 2-4, 6-13, from preparations fixed in Schaudinn's