Mitochondria and Sperm-tail Formation, withparticular reference to Moths, Scorpions, andCentipedes.

By

Vishwa Nath, M.Sc,

Mohindra College, Patiala, India, and Trinity Hall, Cambridge.

(Prom the Zoological Laboratory, Trinity College, Dublin.)

With 3 Text-figures.

INTKODUCTION.

IN a paper on the spermatogenesis of Moths publishedtwo years ago Bowen (3), while agreeing generally with theprevious account given by Gatenby (8), differed in two impor-tant points, namely, the structure of the ' macromitosome ',or mitochondrial' nebenkern ', and its ultimate fate in sperma-teleosis. According to Gatenby the structure of the ' macro-mitosome ' is a ' spireme ', and this breaks up into small fila-ments and directly gives rise to the tail-sheath. Bowen, onthe other hand, considers that the ' macromitosome' is a' plate-work' structure and progressively shrinks duringspermateleosis till it disappears, and the tail-sheath arisesfrom a new ' central substance ' which has nothing to do withthe mitochondrial ' nebenkern '.

The object of the present paper is to offer some originalobservations with regard to the sperm-tail formation in Moths,Scorpions, and Centipedes, and also to review as briefly aspossible the tail formation in the various groups of the animalkingdom.

I have to thank Professor J. Bronte Gatenby, who, aware ofmy interest in gametogenesis, very kindly invited me toexamine his new material and to investigate some of the contro-

644 VISHWA NATH

versial points "with regard to the tail formation in Moths andScorpions. He not only generously placed at my disposal somefresh moth material on which no observations had so far beenmade, but ungrudgingly allowed me to study his old slides ofS m e r i n t h u s popul i and other species on which his impor-tant paper on Lepidopteran spermatogenesis was based, andfurther permitted me to make such use of his other slides asI might wish. I have to thank him also for allowing me toexamine his interesting material illustrating sperrnatogenesisin P a l u d i n a , S a c c o c i r r u s , Cavia, He l ix , andG r a n t i a, which have been very useful to me for the purposesof review. I need hardly say that, without his kind help invarious ways I would not have been able to finish the presentwork so expeditiously.

PREVIOUS WOEK.

During spermateleosis the mitochondria of the spermatid—at least the greater part of them—form some part of thetail of the ripe sperm. It is only in one case (Per ipa tus ,Montgomery, 17) that the mitochondria have been said to besloughed off completely, and this case requires confirmation.

In what we may consider as the primitive sperm the mito-chondria remain clumped together to form a ' nebenkern 'immediately below the nucleus. Such a condition is met within Annelids, e.g. Glycera (Text-fig. A, 1) and Nereis(24 a) ; in Molluscs, e.g. Chi ton (Text-fig. A, 4) ; in Coelen-terates, e.g. Alcyon ium (Text-fig. A, 3) ; in Amphioxus(82), in Nemerteans, in Echinoderms, and in Fishes, e.g.A mi a and Zoarces (22). In his basic work on the germ-cells of Sponges, Gatenby (12) describes a similar arrangement.in Grant ia (Text-fig. A, 14).

From this primitive arrangement of the mitochondria wemay derive the arrangement that prevails in the typical spermof Mammals. In this group the mitochondria descend downthe axial filament for some distance and form the well-known'• middle-piece ' (Meves, 15, Duesberg, 7, Eegaud, 21, Eetzius,

MITOCHONDRIA AND SPERM-TAIL FORMATION 645

22 ; and Gatenby and Woodger, 13). The last workers havestudied the mammalian spermateleosis as illustrated byCavia "with the most modern methods, and have shown(Text-fig. A, 6) that the majority of the mitochondria (M)form the sheath of the ' middle-piece ', while a few (M') aresloughed off with the Golgi apparatus (G) after the latterhas budded off the Golgi bead (G.B) on the ' middle-piece '.I have myself examined some of the material on which thesefindings have been based. An essentially similar mitochondrialsheath of the ' middle-piece ' exists in Eeptiles, e.g. T e s t u d o(Text-fig. A, 10), and in Birds, e.g. Ga l lu s (Text-fig. A, 2).

In the third arrangement the mitochondria descend downalmost the whole length of the axial filament and form thetail-sheath. In doing so they may either first pass through theprimitive condition as described for G l y c e r a , &c, in whichthey are grouped immediately below the nucleus, or they maydirectly descend down the axial filament. This arrangementmay be illustrated by S a e c o c i r r u s , P a l u d i n a , Li m a x ,and Scorpions.

In S a e c o c i r r u s (Gatenby, 11) the numerous small mito-chondria of the spermatid run together to form large spheresat the base of the nucleus, as shown in Text-fig. A, 9. Laterthese spheres are pulled down the axial filament and give riseto the tail-sheath (Text-fig. A, 8).

In the typical spermatid of P a l u d i n a (Gatenby, 10) eachof the few rod-shaped mitochondria of the spermatocyte con-denses to form a spherical body (Text-fig. A, 5). They thenelongate along the axial filament and form the tail-sheath(Text-fig. A, 7).

In the Pulmonates Li max and H e l i x (Gatenby, -9)probably a few of the mitochondria directly form the tail-sheath, whereas the majority are always sloughed off (Text-fig. A, 11, 12, and 13).

In an Arizonan scorpion, C e n t r u m s (Wilson, 24 b), eachspermatid contains two spherical, homogeneous mitosomes,each one of which is one-eighth of the spermatocyte chondrio-some ring which is formed by the union of numerous small

646 VISHWA NATH

mitochondria. The two mitosomes of the spermatid becomeattenuated along the axial filament and form the spiral tail-sheath. In O p i s t h a c a n t h u s (Wilson, 2ib) the numberof the mitochondria in the spermatid varies from five to seven,which become closely grouped together at the base of thenucleus like the primitive arrangement that prevails in Gly-c e r a , &c. Later they are carried backwards along the axialfilament and form the tail-sheath.

In P a l a m n a e u s b e n g a l e n s i s (Gatenby and Bhatta-charya, 14) the number of mitochondria in the spermatid variesfrom four to eleven. They become grouped together at thebase of the nucleus (Text-fig. B, 11) and later form the tail-sheath as in O p i s t h a c a n t h u s (Text-fig. B, 7 and 8).

The only apparent case of the complete sloughing off of themitochondrial material is the well-known case of P e r i p a t u s(Montgomery). In this form a mitosome is formed by theunion of the mitochondria and this is bodily discharged, whilethe tail-sheath is a new structure altogether. This caserequires confirmation, and is interesting owing to the phylo-genetic importance of P e r i p a t u s . The complete dischargeof the mitochondrial material in P e r i p a t u s which is con-sidered, on all hands, to be a primitive animal, conflicts withthe evidence obtained from a considerable number of otherso-called primitive forms in which the mitochondria form atleast a part of the ripe sperm.

The formation of the tail-sheath in insects seems to be themost specialized in the whole animal kingdom. Meves (16)and Gatenby (8) working on Lepidoptera, Duesberg (see Agar)working on B l a t t a g e r m a n i c a , and Charlton (5) workingon L e p i s m a d o m e s t i c a , all claimed that the mito-chondrial ' nebenkern ' directly forms the tail-sheath.

Gatenby (1917) was the first worker on Lepidopteranspermatogenesis to employ modern technique. Before thepublication of his paper our knowledge of Lepidopteran or anyinsect spermatogenesis was in a confused state, as all theprevious workers, e.g. Meves (16), Cook (6), Platner (20), andMunsen (18), had employed fixatives containing acetic acid.

MITOCHONDRIA AND SPERM-TAIL FORMATION 047

It is only recently, therefore, that clear light has been thrownon the questions associated with the structure of the mito-chondrial vesicles; their behaviour during meiosis; theirultimate union to form the ' niacromitosorne ' ; the formationof the acrosome, the ' micromitosome ', and the centrosomes.

TEXT-FIG. A.

A

1, Ripe sperm of Glycera (after Retzius). 2, Ripe sperm ofGal lus (after Retzius). 3, Ripe sperm of A lcyon ium (afterRetzius). 4, Ripe sperm of Ch i ton (after Retzius). 5 and 7,Maturing sperms of P a l u d i n a (after Gatenby). 6, Maturingsperm of Cavia (after Gatenby and Woodger). 8 and 9, Matur-ing sperms of Saccoc i r rus (after Gatenby). 10, Ripe sperm ofT e s t u d o (after Retzius). 11, 12, and 13, Maturing sperms ofLi max (after Gatenby). 14, Sperm of G r a n t i a (afterGatenby). A, acrosome; A.F, axial filament; O, Golgiapparatus ; M, mitochondria ; N, nucleus ; X, bodies believedto form a part of the sperm-tail; Y, yolk-spheres.

So far as t a i l f o r m a t i o n in Moths is concerned Gatenby's' spireme ' structure of the ' macromitosonie ' and its directtransformation into the tail-sheath has been contradicted byBowen (3), who favours the ' plate-work ' structure of the' macromitosome ' and describes a progressive dwindling of thelatter and the appearance of a new ' central substance ' whichforms the tail-sheath.

648 VISHWA NATH

Bowen (3) has brought the Lepidopteran spermatogenesis inline with his previous work on Hemiptera (2), and maintainstha t ' a further comparison with the Orthoptera and Coleopterawhich I have studied gives unmistakable indication of thefundamental similarity in the processes of sperm formationin all insects '. The essence of Bowen's work is that the tail-sheath is not directly formed from the mitochondrial ' neben-kern' which gradually disappears, but is a new structurealtogether.

In a recent paper (4) Bowen has worked out the tail formationin Coleoptera and Aptera. His figures of L e p i s m a clearlyshow the gradual condensation of the mitochondrial ' neben-kern ' and the simultaneous appearance of the sheath-formingsubstance or Bowen's ' central substance '. In Coleoptera hedoes not figure any new ' central substance ', but says (p. 354) :' The usual condensation phenomena are apparently passedthrough by the " nebenkern ", but on so small a scale that I havenot attempted to study them.'

OBSERVATIONS.

Lep idop te ra .—The slides of lepidopterous testes uponwhich the following observations are based were prepared frominsects bred by Dr. Heslop Harrison in connexion with his workon the genetics of Lepidoptera. Pupae were sent to ProfessorGatenby and prepared by his method (Flemniing-without-acetic acid, iron haematoxylin). This material has never beendescribed before, and I must thank both Dr. Harrison andDr. Gatenby for allowing me to examine it.

Unless otherwise stated the following remarks are based ona cross between O p o r a b i a d i l u t a t a a71 and 0 . a u t u m -n a t a ? . The cells in this form are very large as compared withS n i e r i n t h u s and P i e r i s .

My account starts from that point in spermateleosis atwhich the mitochondria begin to coalesce to form the mito-chondrial ' nebenkern '. For a proper understanding of theextraordinary changes which the ' nebenkern ' undergoes it

MITOCHONDRIA AND SPERM-TAIL FORMATION 649

is necessary to recall the structure of the mitochondria beforethey begin to run together. In optical sections each mito-chondrion has a chromophilic rim surrounding central chromo-phobic material. The chromophilic material forms really acomplete thin envelope round the chromophobic material;but in sections, owing to the effects of curvature, it appearsin the form of a rim and cannot be seen in surface view.

Text-fig. C, 5, shows a transverse section through the ' neben-kern ' at a stage when the mitochondrial vesicles have startedrunning together. The central stippled area is chromophilicand, if properly differentiated, is distinctly alveolar as shown inthe figure. Lying on the periphery of this chromophilic areain an irregular fashion are alveoli of different sizes which,except for the chromophilic rims surrounding them, are per-fectly chromophilic. As differentiation of the ' nebenkern 'proceeds further, the chromophilic rims surrounding the peri-pheral chromophobic alveoli are progressively withdrawn tillwe get a large chromophobic cavity surrounding the centralchromophilic area (Text-fig. C, 6 and 7). At the same time thenumber of alveoli in the central chromophilic area is progres-sively reduced till only a few ' lines ' can be seen traversing thisarea (Text-fig. C, 6 and 7). We have now reached a stage inwhich the ' nebenkern' possesses a prominent peripheralchromophobic cavity in the centre of which lies a chromophilicmass. In properly differentiated sections the central chromo-philic area always shows a number of ' lines ' traversing it.If, however, the stain is not properly extracted the centralchromophilic mass appears structureless.

The next changes that the ' nebenkern' undergoes arecharacterized by a progressive dwindling and ultimate disap-pearance of the central chromophilic mass and a simultaneousappearance of a new substance in the peripheral chromophobiccavity. Text-fig. C, 8, 9, 10, and 11 show these changes. AtM is the chromophilic mass and at S.S is shown a new substancethat appears in the chromophobic cavity and ultimately formsthe tail-sheath. In Text-fig. A, 10, the chromophilic substanceis reduced to a minute circular area, and in Text-fig. A, 11,

650 VISHWA NATH

which is a transverse section of a very old sperm, it has com-pletely disappeared.

We may now study these changes in longitudinal sectionsrepresented in Text-fig. C, 1, 2, 3, .4. In Text-fig. C, 1, at M,is the ' nebenkern '. The central chromophilic mass is stippled,and this is surrounded by a chromophobic cavity traversed bynumerous ' strands '. In Text-fig. C, 2 (there are of coursemany stages between Text-fig. 0, 1, and Text-fig. C, 2) the' strands ' traversing the chromophobic cavity have completelydisappeared, and at the same time the sheath-forming substance{8.8) has put in its appearance. In later stages the chromo-phobic cavity containing the new sheath substance is progres-sively drawn out, and simultaneously with it the chromophilicmass becoming smaller and. smaller ultimately disappears(Text-fig. 0, 3 and 4).

When it first appears in the chromophobic cavity the sheathsubstance is in the form, of small vesicles which do not stainreadily, and are therefore in sharp contrast to the centralchromophilic mass (Text-fig. C, 2). Gradually, however, thesevesicles run together and form irregular strands which usuallyrun parallel to the longitudinal axis of the cell and begin tostain readily. With the progressive spinning out (?) of thechromophobic cavity containing the sheath substance, andthe simultaneous dwindling and ultimate disappearance of thecentral chromophilic mass, the cytoplasm of the cell becomesless and less prominent and ultimately disappears (compareText-fig. C, 10 with Text-fig. C, 11).

Another point which is worthy of note is that the ' neben-kern ' is a bipartite structure (Text-fig. A, 7 and 8). In thesefigures the partition passes through the middle of the chromo-philic mass, but this is not always the case. In Text-fig. C, 9,the chromophilic mass is unequally divided, while in Text-fig. C, 10, it lies entirely in one-half of the chromophobic cavity.

The same series of changes which have been described abovefor the ' nebenkern' of O p o r a b i a also characterize the'nebenkern' of S m e r i n t h u s p o p u l i (Text-fig. C, 12)and P i e r i s r a p a e (Text-fig. C, 13).

MITOCHONDRIA AND SPERM-TAIL FORMATION 651

The various other structures in the cell—the nucleus, theacrosome, the acroblasts, the Golgi apparatus, the centrosome,and the axial filament—have been incidentally shown in thefigures, but since they have been fully discussed by Gatenbyand Bowen a description of them is quite unnecessaryhere.

From the above description it will be clear that the chromo-

TEXT-FIG. B.

1, Spermatocyte of P a l a m n a e u s fu lv ipes m a d r a s p a l e n s i s(original). 2, 3, 4, 5, and 6, Spermatids and maturing sperms ofthe same (original). 7, 8, and 11, Maturing sperms of P . ben-ga lens i s (after Gatenby and Bhattacharya). 9 and 10,Maturing sperms of L i t h o b i u a f o r f i c a t u s (original).A, acrosome ; a, acroblasts ; A.F, axial filament; Cy, cyto-plasm ; O, Golgi apparatus ; Ml, mitochondria ; N, nucleus.

philic area of the ' nebenkern ' is central, and the chromophobiccavity in which the new sheath-forming substance develops isperipheral. Bowen has named this new substance as the' central substance ', which appears to be an unsuitable name.I prefer to call this new substance the ' sheath substance 'because it ultimately forms the sheath of the axial filament.

The condensation stages of the chromophilic area are accom-NO. 276 X X

652 VISHWA NATH

panied by the appearance of the ' sheath substance ' in thechromophobic cavity. In later stages the small droplets ofthe ' sheath substance ' unite to form irregular strands orfilaments, which Charlton (5) working on L e p i s m a , inagreement with Gatenby (8) working on S m e r i n t h u s ,interpreted as having been directly derived from the chromo-philic substance. Bowen, on the other hand, working onC u l l o s a m i a (?) and P y g a e r a interprets this new ' sheathsubstance' (his ' central substance') as a new substancealtogether. After a careful study of the slides of O p o r a b i a ,S m e r i n t h u s , and P i e r i s I am convinced that the ' sheathsubstance ' is not d i r e c t l y derived from the chromophilicsubstance, which never fragments but undergoes a progressivedwindling and ultimately disappears. On the other hand, Iam quite prepared to grant that as the chromophilic substanceis undergoing condensation, its substance may somehow betransformed into the ' sheath substance ' which is simultane-ously developing in the chromophobic cavity. The i n d i r e c torigin of the ' sheath substance ' from the chromophilic sub-stance is perhaps likely because it is difficult to imagine ofanother source from which it could develop.

We may now consider the structure of the ' nebenkern '.According to Gatenby the structure of the ' nebenkern ' isa ' spireme ' which is formed by the flowing together of themitochondria, ' forming at first elongated structures, thenloops, and finally filaments, the latter joining up gradually toform a tangled anastomosing figure '.

Bowen, on the other hand, regards the ' nebenkern ' as a' plate-work ' structure. He considers that it results from themitochondria running together into bigger alveoli which arearranged like one soap bubble within another, with more or lessirregular septa running across the chromophobic contents ofthese alveoli. His main objections to the ' spireme ' theory are,firstly, that if the structure of the chromophilic area is a' spireme ' it is very surprising that we can never see the cutends of the threads in any section. Secondly, the progressivedwindling of the chromophilic area is best understood on the

MITOCHONDRIA AND SPERM-TAIL FORMATION 653

' plate-work ' theory by the simple withdrawal of the chromo-philic septa or plates.

I agree with Bowen that it is impossible to see the cut endsof the threads of the ' spireme '. I further agree with himthat the condensation phenomena of the chromophilic areacannot be understood on the ' spireme ' theory. In the laststage of condensation the chromophilic area appears quite likean ' ovoid shell' (Text-fig. C, 4,10, and 13), and this appearancecan be better understood on the ' plate-work ' theory by sup-posing that the outer shells have disappeared and their chromo-phobic contents have merged into the peripheral chromo-phobic cavity. On the ' spireme ' theory it is difficult tounderstand how we can arrive at what I may term the singleshell stage.

Another objection to the ' spireme' theory is that themeshes of the chromophilic ' threads ' should appear quitecolourless. On the other hand; when the finer adjustment of themicroscope is used one very clearly sees chromophilic platesjoining up the chromophilic ' threads '. As Bowen has rightlypointed out, the chromophilic threads are simply due to anoptical illusion resulting from the effects of curvature.

In all justice to the pioneer work of Gatenby, however, Imust say that appearances are sometimes very deceptive,and one may sometimes be tempted to regard the structureof the chromophilic area as that of a ' spireme '. This is due tothe fact that if the osmic acid is not sufficiently removed fromthe material by washing (as it was not in Dr. Gatenby's firstpreparations), the staining capacity of the chromophilic areaof the ' nebenkern ' is considerably impaired, consequently onecan see the plates only at the points of curvature where theyappear in the form of ' threads ', but when seen in surface viewthey cannot be made out at all and the ' meshes ' of the'threads ' thus appear quite colourless and the whole structuremay wrongly be interpreted as that of a ' spireme '.

Scorpions.—The species of scorpion, P . f u lv ipesm a d r a s p a t e n s i s , on which the following observations arebased, was collected from Madras (India). The material was

x x 2

654 VISHWA NATH

prepared according to Gatenby's method (Plemming-without-acetic and iron haernatoxylin). Sections were also chromed•on the slide and stained in Benda's alizarin and crystal violet.

It is not my intention to give a full account of the mito-chondria, the Golgi apparatus, and the acrosome. For thatthe reader may refer to the paper on the spermatogenesis ofP . b e n g a l e n s i s by Gatenby and Bhattacharya (14). Ishall only mention certain points of difference with regard to thestructure and behaviour of the mitochondria in the two species(see ' Nature ', July 12, 1924).

I have recently seen the Dublin preparations of P . ben -g a l e n s i s and can strongly support the statement of thewriters that ' the mitochondria form the sperm-tail directly '.In Text-fig. B, 7, 8, and 11, are drawn three stages in sperma-teleosis in P . b e n g a l e n s i s . It will be seen that the mito-chondria, at first spherical (Text-fig. B, 11), later apply them-selves to the base of the nucleus and become ' leaf-like ' (Text-fig. B, 7). These leaf-like bodies become more and moreattenuated and ultimately form the tail-sheath (Text-fig. 8).

A comparison of the mitochondria of P . f u l v i p e sm a d r a s p a t e n s i s with those of P . b e n g a l e n s i s revealedcertain minor differences. In my preparations of P . fu lv ipesm a d r a s p a t e n s i s the mitochondria in late spermateleosisappear absolutely hollow like a ring (Text-fig. B, 4, 5, and 6).In the sperrnatocytes as well as in the young spermatids theyare not so hollow (Text-fig. B, 1, 2, and 3). It would thusappear that during spermateleosis the chromophilic lipoidmembrane of the mitochondrion containing a central chromo-phobic material becomes progressively thinner and thinnertill we reach a stage when in optical sections the very thinlipoid membrane cannot be seen in surface view and becomesvisible only at the points of curvature. Hence each mito-chondrion appears like a ring—a hollow surrounded by a rim.The mitochondria at this stage are very similar to those ofinsects, e.g. Lepidoptera (Gatenby). In P . b e n g a l e n s i sthe lipoid membrane of each mitochondrion appears to be muchthicker, and can be seen not only at the points of curvature

MITOCHONDRIA AND SPERM-TAIL FORMATION 655

but also in surface view ; hence they never appear ring-like.These differences may be due to slight differences in technique,although the fixative and the stain used in both the specieswere the same.

Another minor difference is with regard to the relative time

TEXT-FIG. C.

1, 2, 3, and 4, Longitudinal sections of the maturing sperm ofO p o r a b i a d i l u t a t a x a u t u m n a t a , showing the progres-sive dwindling of the mitochondrial ' nebenkern ' (M) and thesimultaneous appearance of a new sheath-forming substance(S.S) (original). 5, 6, 7, 8, 9, 10, and 11, Transverse sections ofthe same showing the same phenomenon. 7 shows a somewhatoblique section (original). .12, Longitudinal section of thematuring sperm of S. popu l i showing the same phenomenon(original). 13, Longitudinal section of the maturing sperm ofP i e r i s r a p a e showing the same phenomenon (original).A, acrosome, a, acroblasts ; A.F, axial filament; Cy, cytoplasm ;O, Golgi apparatus; M, mitochondrial' nebenkern'; N, nucleus ;S.S, sheath substance.

of elongation of the nucleus and the state of development ofthe mitochondrial vesicles. This is shown clearly by comparingText-fig. B, 6 (P. fulvipes madraspatensis) with Text-fig. B, 7 (P. bengalensis).

As to the behaviour of the mitochondria in P. fulvipes

656 VISHWA NATH

m a d r a s p a t e n s i s : after the stage represented in Text-fig. B, 6, each ring-like mitochondrion breaks down into smallpieces of mitochondrial material which are seen on the axialfilament at various intervals, and the whole appearance of thecyst is such that, were it not for the evidence obtained from thestudy of other species of scorpions, one would hesitate to saythat the mitochondria form the tail-sheath.

Professor Gatenby is of the opinion that the bladder-likemitochondria of P . f u l v i p e s m a d r a s p a t e n s i s aredifficult objects to manipulate and liable to bursting. Afterhaving personally examined his preparations of P . ben-g a l e n s i s in which each mitochondrion becomes attenuatedalong the axial filament, I think that the fragmentation of themitochondria in P . f u lv ipes m a d r a s p a t e n s i s isartificial. In this connexion the figures of E u s c o r p i u sc e r p a t h i c u s by Sokolom (23) may be studied with advan-tage. This worker figures large vesicular mitochondria in theyoung spermatids, but in later stages he actually figures smallpieces of mitochondrial material on the axial filament. Probablythis also is due to the bursting of the elongating mitochondria.

L i t h o b i u s for f ica tus .—A full account of the spermato-genesis of L i t h o b i u s f o r f i c a t u s will be found in anotherplace. Here it is my intention -to show that in Chilopods,a group which has not yet been studied by the more recentmethods of cytology, the mitochondria form the sperm-tail.Text-fig. B, 9, shows an egg-like spermatid attenuating intoa sperm. The mitochondria are at M. In later stages when thenucleus moves forward, all the mitochondria come into the tailregion and form the tail-sheath directly. Text-fig. B, 10, showsa transverse section of a fairly old sperm. At A .F is the axialfilament and the mitochondria at M are forming a sheathround it.

MITOCHONDRIA AND SPERM-TAIL FORMATION 657

SUMMARY.

E e v i e w .

1. In Sponges, e.g. Grantia (Gatenby), some Annelids,e.g. Glycera (Eetzius), Coelentrates, e.g. Alcyonium(Eetzius), some Molluscs, e.g. Chiton (Eetzius), Am-phioxus (Eetzius), Nemerteans, Echinoderms, and Fishes,e.g. A mi a (Eetzius), the unchanged mitochondria remainclumped together to form a ' nebenkern ' at the base of thenucleus of the ripe sperm.

2. In Mammals (Meves, Eegaud, Duesberg, Eetzius, andGatenby and Woodger) the mitochondria form the sheath ofthe ' middle-piece'. An essentially similar condition prevailsin Birds, e.g. Gallus (Eetzius), and Eeptiles, e.g. Testudo(Eetzius).

8. In Saccocirrus, Paludina, Lirnax (Gatenby),and Scorpions (Wilson, Gatenby and Bhattacharya, and Nath)the mitochondria form the sheath of almost the whole lengthof the axial filament.

4. In the highly specialized case of Insects, e.g. Aptera,Orthoptera, Lepidoptera, Coleoptera, and Hemiptera (Bowen),the mitochondrial ' nebenkern ' undergoes a progressive con-densation, and the sheath of the axial filament is a new struc-ture altogether.

Original Observations.

1. In Oporabia, Smerinthus, and Pieris the mito-chondrial ' nebenkern ' does not directly form the tail-sheath,but undergoes a progressive dwindling and ultimately disap-pears. The sheath-forming substance arises as a new substancein the chromophobic cavity round the ' nebenkern '.

2. The structure of the Lepidopteran mitochondrial' neben-kern ' is alveolar or ' plate work ' and not a ' spireme '.

3. In the centipedes (Lithobius forficatus) the mito-chondria form the tail-sheath directly.

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LIST OP LITERATURE.

1. Agar, W. E.—' Cytology with particular reference to the MetazoanNucleus', 1920.

2. Bowen, Robert H. (1922 6).—" Studies on Insect spermatogenesis.III. On the structure of the ' nebenkern' in the insect spermatidand the origin of ' nebenkern ' patterns ", ' Biol. Bull.', vol. 42>1922.

3. " Studies. V. On the Formation of the Sperm in Lepidoptera ",' Quart. Journ. Micr. Sci.', vol. 66, Part IV, 1922.

4. " Studies. VI. Notes on the formation of the sperm in Coleo-ptera and Aptera, with a general discussion of flagellate sperms ",' Journ. Morph.', vol. 39, 1924.

5. Charlton, H. H.—" The spermatogenesis of Lepisma domestica ",ibid., vol. 35, 1922.

6. Cook, M. H.—" Spermatogenesis in Lepidoptera ", ' Proc. Acad. Nat.Sci. Philadelphia ', vol. 62, 1910.

7. Duesberg, J.—" Nouvelles recherches sur l'appareil mitochondrial descellules seminales " , ' Arch. f. Zellf.', Bd. 6, 1911.

8. Gatenby, J. Bronte.—" The cytoplasmic Inclusions of Germ-cells.Part I. Lepidoptera " , ' Quart. Journ. Micr. Sci.', vol. 62, 1917.

9. Ditto, Part III, ibid., vol. 63, 1919.10. Ditto, Part IV, ibid.11. Ditto, Part X, ibid., vol. 66, 1922.12. "The Germ-cells, Fertilization, and early Development of

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