The Embryology of Temnocephala novae-zealandiae Haswell.
[Received by the Editor, December 20, 1946; issued separately, September, 1947.]
Detailed studies of the embryology of the Temnocephalida have been undertaken by Haswell (1909) and Fernando (1934). Haswell described mainly the development of Temnocephala fasciata, an Australian species and did some work on four other species, including T. novae-zealandiae. He stated that he was preparing material for a fuller study of the New Zealand species, but the results of his study do not seem to have been published. He acknowledges that his account of the embryology of the group is not complete, and it was with a view to elaborate on Haswell's account and to endeavour to gain a complete picture of the development of the group, that Fernando undertook the study of the embryology of Caridinicola indica, and published his account in 1934. These two published works, together give a full study of the development of typical members of the Temnocephalida. The object of the present study was to find out how the development of the New Zealand species compared with the known facts.
This paper is a condensation of that submitted as a thesis for the degree of M.Sc., University of New Zealand. The writer wishes to acknowledge the help and criticism given him during the course of the work by Professor H. B. Kirk, Professor L. R. Richardson, and Dr. H. B. Fell, all of Victoria University College, Wellington.
Materials and Methods.
Temnocephala novae-zealandiae adults and eggs were obtained from fresh water crayfish, Paranephrops neozelanicus, inhabiting a stream near Wellington. Both the adults and the eggs occur in large numbers on the host. The adults are found on all parts of the body but the majority live on the great chelipeds. They are very active, especially when disturbed, and are difficult to dislodge from the host. The eggs are cemented firmly to the surface of the crayfish, and most of them are laid on the proximal podomeres of the great chelipeds. Some eggs are found at the base of the antennae and antennules and around the regions of the mouth. T. novae-zealandiae breeds throughout the year, and eggs can be obtained in quantity at any season. Eggs of known age were obtained by keeping several crayfish, which previously had all eggs removed from their appendages and had a large number of adult Temnocephala present, in breeding tanks, and by examining them every day for newly laid eggs.
All specimens were kept in Durham tubes, and, in the preliminary work of dehydration, softening egg membranes, etc., the liquids were changed in the one tube by means of a pipette. This was preferable to transferring eggs to a fresh liquid in another tube. Eggs were fixed in Schaudinn's Fixative (Lee, 1928). Good fixation was obtained after the eggs had been in the fixative for half an hour. They were then preserved in 70 per cent. alcohol and used as required. Adults were fixed with Zenker's fluid (Lee, 1928). They were placed on a glass slide, covered with hot fixative, flattened under a glass cover-slip, and left for 30 minutes. They were then washed in water for several hours and preserved in 70 per cent. alcohol with iodine.
Imbedding. The egg membrane is thick and tough, and made section cutting a difficult process. Haswell's (1909) method of softening the membranes by placing the eggs in an aqueous solution of sodium hypochlorite was tried. It was found, however, that even when the eggs were passed through this solution, the membrane still remained comparatively tough and offered too much resistance on the microtome to make the method an absolutely sure one of obtaining a good and complete series of sections from all eggs. A modified imbedding procedure was finally adopted, and is described herewith.
It was discovered that the egg membrane could be removed with comparative ease from the eggs in the 70 per cent. alcohol preservative. The membrane was teased off the embryo by needles under a low power lens, and in most cases the embryo was liberated unharmed. The embryos thus obtained were imbedded in paraffin first in a watch glass and then oriented in another bath of paraffin.
The purpose of the first paraffin bath was to facilitate orientation of the embryos into suitable positions for section cutting, and since the majority in this block were lying with their longitudinal axes parallel to the surface of the paraffin, they could be extracted from it together with a small mass of paraffin and placed in the second block in the desired positions.
Adults were imbedded in paraffin and the procedure given by Carleton (1938) was followed.
The sections of both embryos and adults were stained with Heidenhain's iron Haematoxylin and Eosin. These stains proved entirely satisfactory and produced very good differentiation.
The majority of the sections were cut at 6 μ, a thickness which appeared to be the most satisfactory to enable the details of individual cells to be observed.
Diagrams of sections were made with the aid of a camera lucida, and reconstruction diagrams were done by the graphic method of Pusey (1939).
The Reproductive Organs.
Fyse (1943) has given a full description of the reproductive organs of Temnocephala novae-zealandiae. Prom material examined in the present study the germarium is seen to be an ovoid body situated dorsally on the right of the genital complex. In a normal sized adult it is 71 μ wide and 78.5 μ long. The ova at all stages of
development can be seen in the germarium. The ovum nearest the germiduct is the mature one, and is 50 μ wide. The remaining ova are wedged in the ovary and are correspondingly smaller. The nucleus of the mature ovum is relatively large, and is a conspicuous object. It is oval, and is 28.6 μ long and 21.4 μ wide. The nucleolus has a diameter of 10.7 μ. (Plate 49, Fig. 1.)
The testes are four in number, one pair being situated on either side of the body. Sperm is seen in the receptaculum seminis, the seminal ducts and occasionally in the receptaculum resorbiens. A spermatozoon is 7 μ long, with a head of approximately 1 μ diameter. (Plate 49, Fig. 1.) The vitelline glands are described below.
A ripe ovum leaves the germarium and passes into the ootype through the germiduct. There it is fertilized, yolk material and shell material added, and it passes into the uterus, genital atrium, and finally out through the common genital pore. The completed egg thus consists of a zygote surrounded by yolk material and all enclosed in an egg membrane.
In describing the nature of the yolk portion of the egg Haswell (1893 and 1900) did not clearly indicate the existence of yolk cells, but later (1909) he distinguishes yolk cells as forming the greater part of the mass of the fully formed egg, and he describes their subsequent coalescence to form a syncytium. Fernando (1934) also identified initially polyhedral yolk cells which later fuse to form a syncytium.
The yolk cells are formed by the vitellarium and Benham (1901) considers that this structure in the Temnocephaloidia (and many Turbellaria) is a modified anterior portion of an original female gonad which in the course of evolution has become divided into a gremarium and a vitellarium.
From the sections examined in the present study the vitelline glands are seen to be extensive structures in the form of irregular lobate follicles lying in the parenchyma (Plate 49, Fig. 5) on the dorsal and lateral sides of the intestine. In some mature animals they are also found in the parenchyma on the ventral side of the intestine. A section through one of the follicles of the gland shows that the yolk material is in the form of irregular cells, average diameter 25 μ, each containing a comparatively large nucleus and cytoplasm that is heavily laden with yolk granules (Plate 49, Fig. 4). The nuclei of these cells measures 10 μ in diameter, and the yolk granules have an average diameter of 1.6 μ. There is a single vitelline duct on each side (Plate 49, Fig. 5). These ducts unite to open into the ootype about the region of the germiducts (Plate 49, Fig. 3). Yolk material in the form of complete cells is produced in the vitelline follicles and passes down the vitelline ducts to the ootype (Plate 49, fig.g 2). There the yolk cells enter into the formation of an egg by surrounding the fertilized ovum. The yolk cells are being secreted by the vitelline glands continuously, and when an egg has been completed and lies in a portion of the ootype, the yolk cells formed at this time are surplus and enter the receptaculum resorbiens to be
ultimately absorbed into the intestine. The yolk cells which enter into the formation of an egg are similar to those cells described in the follicles of the vitelline glands. They are polyhedral in shape and have an average diameter of 30 μ Each cell has a large nucleus, average diameter 12 μ, in which the chromatin material is very extensive and conspicuous (Plate 50, Fig. 6). Yolk granules are very numerous in the cytoplasm of these cells, and they have an average diameter of 2 μ.
In appears, therefore, that in Temnocephala novae-zealandiae the condition described by Benham (as stated above) holds good. The female gonad has become differentiated into germarium and vitellarium, and when an egg cell leaves the germarium it is surrounded by vitelline cells which supply nourishment for the developing embryo. These yolk cells soon lose their individuality and fuse to form a syncytium. There is no evidence that the cells of the vitelline glands break down and that secreted material passes down the vitelline ducts and helps in the completion of an egg.
The completed egg when laid is cemented to the surface of the crayfish by material secreted by cement glands which surround the genital atrium. The egg is pear-shaped, the long axis measures 450 μ, the broadest diameter is 350 μ, it has a small filament at its distal extremity, and is attached to a crayfish by a short stalk. (Plate 50, Fig. 9.)
The zygote was not observed in the present study, but from measurements of the mature ovum in the germarium, it would have a diameter of 50 μ or more.
The first division of the zygote forming a two-celled embryo was not observed. However, from subsequent stages, the nature of this division was clearly seen. The fertilized egg divides unevenly into two cells, a large macromere and a small micromere. The macromere soon divides and gives rise to a macromere smaller than the original macromere and with a diameter 43.3 μ, and a small micromere smaller than the first formed micromere and with a diameter of 20 μ.
The three-celled stage is seen in Plate 50, Fig. 7. Here are shown the single large macromere and the two micromeres. The largest micromere results from the first division of the zygote and has a diameter of 33 μ. In the prepared slides showing this stage, no normal nuclei were observed. They were represented by a series of small, clear vesicles each of which contained a small mass of chromatin material. Haswell (1909) noted this nuclear state, but Fernando (1934) did not record it in Caridinicola indica.
Following on this three-celled stage it appears that the macromere now divides evenly, producing two macromeres, and the micromeres divide evenly producing four micromeres.
Repeated divisions similar to these now occur, and the number of cells increases rapidly. The two types of cells are clearly discernible in all sections. In serial sections of an embryo of seven cells the macromeres measure 30 μ diameter and micromeres 15 μ. (Plate 50,
fig. 8.) At a later stage they become smaller still and have diameters of 13.3 μ and 6.5 μ respectively.
At this stage Haswell (1909) describes three sets of cells differing from one another in size and character of nuclei. Fernando (1934) describes two sets of cells but states that the smaller cells (micromeres) vary somewhat in size. From my material in Temnocephala novae-zealandiae it appears that there are only two distinct types of cells as described above.
These cells constitute the blastoderm. When it is fully formed it is an ovoid mass of cells situated near the centre of the yolk mass. (Plate 51, Fig. 11).
The cell membranes are clearly discernible in the early blastula stage (Plate 50, Fig. 6) but when the blastoderm is almost formed the boundaries tend to disappear and a syncytium is formed. (Plate 51, Fig. 11.) The cells therefore must be described from their nuclei. In Plate 51, Fig. 11, there are large nuclei of average diameter 8.0 μ, representing macromeres and hence called macronuclei, and small nuclei of average diameter 5.0 μ representing micromeres and hence called micronuclei. Most of the macronuclei lie towards the proximal end of the blastoderm, which is the future posterior end of the embryo. The division between these two masses of cells is by no means distinct and sharp, for several large macronuclei are found in the mass of micronuclei.
Haswell (1909) states that no germinal layers are recognisable in the blastoderm, but Fernando (1934) whose description of the blastoderm corresponds fairly well with Haswell's, names two germinal layers on the evidence afforded by the fate of parts of the blastoderm. He considers that the mass of macromeres (macronuclei) is the hypoblast and the mass of micromeres (micronuclei) is the epiblast. In the present case it is seen that the archenteron, parts of the alimentary canal, and probably the reproductive organs and excretory system arise from the mass of macromeres. All these organs are usually regarded as being of typical hypoblastic origin. Also, the epidermis, muscles of the body wall, pharynx, and nervous system, all usually regarded as of epiblastic origin, arise from the mass of micronuclei. Thus there are clearly two germinal layers present, represented by the regions of different sized cells, and they can be called the hypoblast and epiblast respectively with confidence. (Plate 51. fig. 12.) The arrangement of the hypoblast and epiblast in the blastoderm is, however, far from a normal one, and in Temnocephala it is to be regarded as a condition very much modified from that of a normal embryonic development, due to the presence of the large mass of yolk cells and their arrangement about the embryo.
During the development of the blastoderm the surrounding yolk cells are undergoing changes. They gradually lose their individuality; the cell walls disappear and a yolk syncytium is formed. This formation is begun at the periphery of the egg, and when completed there is a mass of cytoplasm surrounding the embryo, very heavily laden with yolk particles and containing numerous large nuclei. It appears that the time when this yolk syncytium is formed varies in different individuals of this species.
In Plate 50, Fig. 6, which represents an early blastula stage, the polyhedral yolk cells are clearly visible. In Plate 50. fig. 10, a stage that is almost at the same stage of development, the yolk portion of the egg does not show the same degree of cellular structure. In Plate 51, Fig. 12, there are shown traces of the individual yolk cells in the yolk tissue immediately surrounding the embryo which is at a late blastula stage, whereas in Plate 51, Fig. 11, showing an earlier embryonic stage no trace of yolk cells is visible.
Organ Rudiment Formation.
The blastula stage is succeeded by a period in which the organ rudiments make their appearance as partially segregated masses of cells of either the epiblast or hypoblast. At first, what might be conveniently called a “gastrula” is formed. This is characterised by the formation of a cavity—the Endocoele. The endocoele, first given its name by Haswell (1909) is presumably homologous with the archenteric cavity since it is the first cavity formed in the embryo when it is composed of two germinal layers, and it is formed in the group of cells considered to be the hypoblast. It develops in the mass of the hypoblast first as a small space surrounded by the macronuclei. (Plate 51, Fig. 12.) As development proceeds the lumen enlarges (Plate 52, Fig. 13) and the macronuclei immediately surrounding it form a well-defined syncytial epithelium. At its maximum development the endocoele extends for approximately one-third of the length of the egg. (Plate 52, Fig. 14.)
The endocoele and its surrounding cells being comparatively large in mass, tend to override the mass of micromeres which is situated anteriorally in the blastoderm. It is when the endocoele is almost fully formed that the masses of macromeres and micromeres start to be clearly differentiated into organ rudiments. Comparatively large, well defined regions of hypoblast lie one on each side of the endocoele (Plate 52, Fig. 13), also one on the mid-ventral region. The lateral masses probably give rise to the testes, prostate glands and excretory apparatus and the ventral mass probably differentiates into the female reproductive apparatus. The epiblastic mass partially segregates into three regions; a pharyngeal rudiment on the antero-ventral aspect of the endocoele (Plate 52, Fig. 14, and Plate 53, Fig. 15); a nervous system rudiment anterior to the endocoele (Plate 52, Fig. 14 and Plate 53, Fig. 15); a region ventral to the endocoele which gives rise to external epithelium and muscles of the body wall. (Plate 52, Fig. 13).
Growth of Embryo.
Once the organ rudiments have made their appearance, further differentiation occurs in them. The endocoele becomes greatly reduced until its cavity is represented by a mere slit. The pharynx meanwhile has enlarged considerably and develops into a very conspicuous organ. It lies close to the antero-ventral surface of the embryo but does not yet communicate with the exterior. The nervous system is well formed by the time the embryo has hatched. While the elaboration of the nervous system and pharynx is going on there is a marked growth of the cells of the two endodermal bands and the
medium ventral ectoderinal mass towards the posterior end of the body where they enter into the formation of the reproductive organs, ventral sucker and posterior portions of the body wall. This posterior mass of cells is of such extent as to give the embryo an appearance of having two centres of development—an anterior and a posterior region. The central mass where once the large endocoele was present is now a mass of yolk tissue destined to become contained in the lumen of the intestine and to be ultimately absorbed by it as food. On the ventral side of the embryo there is a thin, narrow connection between the two regions of development, consisting of the epithelium of the body wall and several layers of cells of epiblastic origin destined to become incorporated into the body wall as muscle tissue or to be converted into parenchyma tissue. (Plate 53, Fig. 15.) The six anterior tentacles make their appearance at an early stage, and as these grow in length they are forced to bend over either the ventral or dorsal surface of the embryo. At the stage when the first small lumen of the pharynx appears and when the endocoele is still relatively large the epidermal epithelium is formed. It apppears first on the ventral surface of the embryo but soon forms a complete covering to the egg. The embryo immediately prior to hatching has a well developed canal in its large, bulbous pharynx, a short oesophagus and an intestine as yet without a cavity of any size owing to the presence of the yolk mass; a nervous system that has a well defined brain and a nerve net together with a pair of eyes; six tentacles, and a well-formed ventral sucker; a body wall consisting of n single-celled layer epithelium with underlying muscular layers; a well-developed and functioning excretory system, and a reproductive system that is only partly mature.
Because it is enclosed in a comparatively firm egg membrane the mature embryo is contorted to a greater or lesser degree. The six tentacles in all cases are bent back over one body surface, and in some cases the anterior portion of the embryo is also bent back. This bending is very marked in a few cases, but in the majority of cases it is not excessive.
The embryo hatches as a juvenile. The egg membrane dehisces and the animal escapes from its shell and attaches itself to the crayfish. Considerable growth takes place after it is hatched, and in time the adult may grow to have a length of approximately 5 mm.
Elaboration of the Alimentary Canal.
Pharynx. On the immediate antero-ventral aspect of the endocoele a section of the mass of epiblastic cells (micronuclei of the blastoderm) becomes partly isolated from the remaining ectodermal cells to form the pharyngeal rudiment. A lumen soon appears in this mass, and it is surrounded at first by a well marked epithelium of a single layer of cells. The remaining cells of the rudiment surround this epithelium. (Plate 52, Fig. 14.)
The lumen, which is the first cavity of the pharynx, then enlarges. The epithelium is well marked and the whole mass of the rudiment becomes a well-differentiated mass whose identity is always clearly defined from the rest of the embryo. The lumen is first
formed at the time of greatest development of the endocoele. As the lumen of the pharynx grows the endocoele gradually diminishes in size, and its cavity shrinks away until it is a very small vestige. (Plate 53, Fig. 15.).
There seems to be no very-marked bending of the embryo at this stage as was described by Fernando (1934) for the species Caridinicola indica such that the endocoele was divided into anterior and posterior cavities. In several sections it was seen that there was an apparent bending of the embryo and consequent partial obliteration of the cavity of the endocoele, but this was due to the collapse of the cavity during section cutting.
The endocoele does not entirely disappear, for in its reduced state it forms the oesophagus. At first there is no connection between the pharynx and endocoele, but later the two cavities unite. This occurs when the pharynx has a relatively large lumen.
As the development of the pharynx proceeds, it proximates to the ventral anterior surface of the embryo (Plate 53, Fig. 15) so close in fact that later there is only the narrowest septum left between the cavity of the pharynx and the exterior. (Plate 54, Fig. 17). This septum is retained until after the embryo is hatched or until just prior to hatching. The epithelium lining the cavity of the pharynx forms a, comparatively tough layer by its gradual shrinkage and hardening. (Plate 54, Fig. 19.) All traces of cellular structure of this layer have disappeared by the time the embryo hatches. The outer layer of cells surrounds the pharyngeal cells and also forms an epithelium. The mass of cells between the two epithelial layers is gradually differentiated into muscle tissues. This differentiation is not completed by the time the embryo is hatched, but most of the pharynx at this stage consists of large muscle masses.
At a stage when the muscular tissues of the pharynx are being: formed there appears the first rudiment of the pharyngeal vesicle. The septum is still present and so there is no direct connection between the exterior and the vesicle. The pharynx, however, has direct communication with the vesicle, and it protrudes into its cavity to a greater or less degree. (Plate 54, Fig. 19.) When adulthood is reached the pharynx is capable of being extruded through the vesicle to the exterior. In some mature embryos the pharynx is not closely opposed to the exterior as at the earlier stages of its development but, because of folding and bending of the embryo, it appears to be situated well in the middle of the anterior region of the embryo. (Plate 54, Fig. 19.) The true mouth is regarded as being the opening of the pharynx into the vesicle.
Oesophagus. The remnant of the endocoele forms the oesophagus. As stated above, the endocoele is reduced to a mere vestige. This vestige, which retains a small cavity, lies close to the posterior end of the now-large pharyngeal mass. Ultimately the cavity of the endocoele is united with that of the pharynx, and so forms an oesophagus. (Plate 54, Fig. 18.) The oesophagus leads back into the yolk mass which will ultimately form the bulk of the intestine.
Intestine. The intestine is uncompleted at the time of hatching, but some progress in its differentiation is apparent. In a freshly hatched individual there is present a single layer of cells representing the intestinal wall. Incorporated in this wall are the first elements of the connective-tissue septa which project into the cavity of the intestine. (Plate 53, Fig. 16.) The wall is formed from cells of the endocoelie remnant and so is hypoblastic in origin. The lumen of the intestine is practically full of yolk until some time after the animal has hatched, but a small lumen may make its appearance in some mature embryos. This is formed by the gradual absorption and breakdown of the yolk cells in the intestine, at first near the oesophagus. (Plate 53, Fig. 16.)
In the mature adult the intestine has its lumen completed and its wall is divided up into irregular inwardly projecting septa.
The Body Wall and Associated Structures.
Both Haswell (1909) and Fernando (1934) describe the formation of the body wall as taking place by a process of migration of cells from blastoderm to exterior. A similar condition was observed in Temnocephala novae-zealandiae.
Cells that bear the same description as cells from the epiblastic region of the blastoderm are seen in various positions in the surrounding yolk material. (Plate 52, Fig. 14.) These cells are first noticed when the pharynx is first being formed and its lumen is as yet small. Later they apparently reach the outer surface of the yolk material, where they flatten out and coalesce with their neighbours to form a thin syncytial epithelium. (Plate 53, Fig. 15.) By a continuation of the process of migration an epithelium that surrounds the whole egg mass is gradually built up.
Beneath the epithelium the other elements of the body wall are formed. In the fully formed embryo the basement membrane and the muscular layers are clearly visible. (Plate 54, Fig. 19.)
The ventral sucker is formed from the epiblastic cells that migrate to the posterior region of the egg. (Plate 53, Fig. 15.) The sucker is developed quite early and consists of an epithelium similar to the epithelium surrounding the rest of the embryo and very extensive muscular tissues.
About the same time as the pharynx is being formed the buds of the tentacles come into being, and the comparatively long tentacles are produced. They bend usually along the ventral surface of the embryo. (Plate 53, Fig. 16.) Bach tentacle has a well-formed epithelium from the time the bud has first formed.
ThE Nervous System.
This system is of epiblastic origin, and is the first one to show any marked differentiation. Before the pharyngeal rudiment has developed a lumen, some cells of the nervous system rudiment have become chunsred into true nervous tissue and exhibit the characteristic fibrillated structure of the “brain” of Temnocephala. The “brain,” or mass of nervous tissue that is situated anteriorally in the embryo develops considerably and soon shows a bilobed struc-
ture. This grows to a comparatively large size, and from it are derived the main longitudinal nerve trunks, the nerves to the tentacles and the pair of eyes. (Plate 54, Fig. 18.)
The Excretory System.
Both Haswell (1909) and Fernando (1934) describe the earliest stages of the excretory system as being connected with the cells surrounding the endocoele. The system can be regarded therefore as being of hypoblastic origin. The earliest stages were not observed in this study. As development proceeds the two excretory rudiments shift from their position near the endocoele and come to lie close to the exterior. (Plate 54, Fig. 19.) Large terminal sacs are formed, and as each sac contains two nuclei they are regarded as being formed from two original hypoblastic cells. In each sac there is a canal which opens to the exterior and, in the confines of the sac, bends sharply back on itself. (Plate 54, Fig. 18.) This canal is the end of the intricate system of excretory canals which commence in the flame cells of the parenchyma, and is, by nature of its formation (Haswell, 1909), intracellular.
The Reproductive System.
In the series of sections examined few details of the reproductive system were seen. The male reproductive system develops before the female system, but whereas the male system may be functional shortly after the individual has hatched, the female system is not mature until some considerable time after hatching. The first rudiments of both systems were not observed, but because in the embryo there is a mass of hypoblastic cells (derived from the posterior elongation of the lateral and medium ventral hypoblastic bands) situated posteriorally to the intestinal yolk mass and, because in the adult animal the reproductive organs are situated posteriorally to the intestine, it is considered that the reproductive organs develop from these cells and are thus hypoblastic in origin.
The vitelline glands are in the process of formation before the embryo hatches. In a mature embryo the first few small vitelline follicles are situated near the dorsal body wall, posterior to the intestine. (Plate 53, Fig. 16.) The remaining parts of the female genital apparatus do not show clear differentiation of structure in the mature embryo since even at this stage these parts are still in a rudimentary state.
The male apparatus, however, is more advanced, for the prostate glands are formed and the first phases of spermatogenesis are taking place before the embryo has hatched.
This study shows that the development of Temnocephala novaezealandiae follows closely that already described for T. fasciata by Haswell (1909) and Caridinicola indica by Fernando (1934).
The egg consists of a fertilized ovum surrounded by yolk cells and all enclosed in a membrane. The paired vitelline glands may be regarded as representing the anterior parts of a pair of original
ovaries which in the course of the evolution of this species (and other members of the sub-order. Tempocephalida) have become differentiated into a vitellarium and a germarium as suggested by Benham (1901). The yolk cells are thus in one sense ova which are well supplied with nutrient matter and upon which the developing embryo feeds.
The blastoderm resulting from the segmentation of the zygote is composed of cells of two sizes. The larger cells are described as the hypoblast and the smaller as the epiblast. Fernando (1934) described only these two germinal layers in Caridinicola indica and his findings are paralleled in Temnocephala novae-zealandiae. From either of these two germinal layers all the organs and structures of the adult can be traced.
The endocoele is regarded as being homologous with the archenteric cavity for reasons similar to those suggested by Fernando (1934)—viz.: it is formed in the hypoblastic cells; it forms that portion of the alimentary canal (the oesophagus and intestine) which is posterior to the epiblastic portion of the canal (the pharynx); it is persistent throughout the life of the animal. The term “endocoele” was suggested by Haswell (1909), who did not regard it as homologous with the archenteron, and this name is retained for convenience and because of the nature of its formation, which differs from that hitherto commonly attributed to an archenteron. There is no process of invagination of cells of the blastoderm. Rather, a small lumen appears in the mass of certain cells of the blastoderm, and this lumen grows until it attains a comparatively large size. It is suggested that this mode of formation of the archenteron is a result of the peculiar nature of the egg wherein an ovum is surrounded by a relatively large mass of yolk material. Although the endocoele is persistent throughout the life of the animal as the oesophagus it shows much degeneration from the gastrula stage wherein it is a large cavity extending through approximately one-third of the egg. From this stage, which can be regarded as the climax in the development of the endocoele, it is reduced to a mere fraction of its original size. In this study there was no indication that the condition stated by Fernando (1934), when describing the reduction of the endocoele, takes place. There was no evidence that the endocoele is divided into two portions by the bending of the embryo and that the posterior portion disappears. Rather it appears that, in the species under consideration, the endocoele is reduced in size without being divided into two portions and that the vestige is situated close to the posterior edge of the now large pharynx.
By the time the endocoele is reduced, the embryo has developed considerably and most of its organs are well on the way to completion. A mass of yolk replaces the endocoele, and there appears to be two primary centres of development in the embryo—an anterior and a posterior one. Haswell (1909) described this state, but when we regard the central mass of yolk as being a kind of elementary form of the intestine whose differentiation is not proceeding as fast as the rest of the embryo the idea of having two separate foci of development does not describe the position so well. The central yolk
mass ultimately becomes incorporated into the intestine, and in fact almost completely fills its cavitiy for some considerable time after the animal is hatched.
Why the endocoele should attain such large proportions is a matter for conjecture Haswell (1909) suggests that it may act as a water reservoir to serve the embryo during dry spells when its host may be forced to spend much time out of water either in the mud of dried-up pools or under rocks. Such conditions do not hold in New Zealand, and if the large endocoele has been developed for this function, it appears that in Temnocephala novae-zealandiae it is a state that has been developed during the course of evolution to meet the condition in some other age and that has been retained by the species to the present day.
It is worth noting that as the endocoele is degenerating to become a mere vestige of its former state the pharynx is enlarging, and in time comes to have a cavity that is almost as large as that of the endocoele of the gastrula stage; also the pharynx does not communicate with the exterior until the animal hatches, and so its cavity may be able to serve as a water reservoir in the same way as is suggested for the endocoele.
Temnocephala novae-zealandiae breeds throughout the year.
The egg consists of a zygote surrounded by yolk cells, and all are enclosed in a tough chitinous membrane.
The yolk cells are secreted by extensive vitelline glands which are considered to be modified anterior portions of an ovary that has become divided into a vitellarium and a germarium during the course of evolution.
Early segmentation results in the formation of a blastoderm showing two germinal layers; an epiblast and hypoblast.
The hypoblastic cavity is called the endocoele and is regarded as being homologous with the archenteron.
The epiblast gives rise to the pharynx, nervous system and body wall including tentacles and ventral sucker.
The hypoblast gives rise to the oesophagus and intestine, and it is considered to give rise to the excretory system and male and female reproductive organs.
The endocoele is persistent throughout the life of the adult as the cavity of the oesophagus.
There is no marked bending of the embryo at the post-gastrula stage such that the endocoele is divided into two cavities.
The alimentary canal consists of an epiblastic portion—the pharynx and pharyngeal vesicle, and a hypoblastic portion—the oesophagus and intestine. These two portions unite.
The body wall is built up by cells which migrate from the embryonic tissue in the centre of the mass of yolk cells to the periphery of the egg, and there coalesce.
There is no larval stage, the animal hatches from the egg as a miniature adult.
The adult does not reach sexual maturity until some time after hatching.
The embryonic development of Temnocephala novae-zealandiae is comparable to that previously described for T. fasciata and Caridinicola indica.
Benham, W. B., 1901. A. Treatise on Zoology, Part IV. The Platyhelmia, Mesozoa and Nemertine. (Ed. B. Ray Lankester), London.
Carlton, H. M., 1938. Histological Technique. London.
Fernando, Wilfred, 1934. Studies on the Temnocephaloidea, II, The Embryology of Caridinicola indica. Proc. Zool. Soc., p. 827.
Fyke, M. C., 1943. The Anatomy and Systematic Position of Temnocephala novae-zealandiae Haswell. Trans. Roy. Soc. N.Z., 72, 253.
Haswell, W. A., 1893. A Monograph of the Temnoeephaleae. Proc. Linn. Soc. N.S.W., Macleay Memorial Volume, p. 93.
—— 1900. Supplement to a Monograph of the Temnocephaleae. Proc. Linn. Soc. N.S.W., 25, 430.
—— 1909. The Development of the Temnocephaleae. Part I. Quart. J. Micr. Sci., 54, N.S., 415.
Lee, Bolles, 1928. The Microtomist's Vade-Mecum. 9th Edition. (Ed. J. B. Gatenby). London.
Pusey, H. K., 1939. Methods of Reconstruction from Microscope Sections. J. Roy. Micr. Soc., 59, 232.
Key to Abbreviations used in Diagrams.
Ec.R.—Rudiment of body wall.
Ep.C.—Migrating epiblastic cells.
Fl.—Fold in embryo.
M.—Muscles of body wiill.
Ma.—Macromere or macronucleus.
Mi.—Micromere or micronucleus.
N.R.—Nervous system rudiment.
O.N.—Nucleus of ovum.
T.Ex.—Terminal excretory sac.
Y.N.—Yolk cell nucleus.
Fig. 1.—Section through the female genital complex of an adult Temnocephala novae-zealandiae.
Fig. 2.—Section through one of the lateral vitelline ducts (see Fig. 5), showing two yolk cells in the process of passing down the duct to the ootype.
Fig. 3.—T.S. through the lateral vitelline ducts where they unite and enter the ootype.
Fig. 4.—Section through one follicle of the vitelline glands.
Fig. 5.—Reconstruction of portion of the vitelline gland, showing the two lateral vitelline ducts uniting in the female genital complex.
Fig. 6.—L.S. through an egg showing an early blastoderm and its relative position in the mass of yolk cells.
Fig. 7.—Portion of a L.S. of an egg showing the three-called stage.
Fig. 8.—Portion of a L.S. of an egg showing a seven-called blastoderm.
Fig. 9.—Egg of Temnocephala novae-zealandiae.
Fig. 10.—Portion of a T.S. of an egg showing an early blastoderm.
Fig. 17.—Portion of a L.S. of an embryo showing the pharynx and the thin septum that separates its cavity from the exterior.
Fig. 18.—Portion of a L.S. of an embrvo showing the pharvnx in connection with the endocoele forming the oesophagus.
Fig. 19.—T.S. through the anterior end of an embryo just prior to hatching.