The Effect of Salinity on Egg Development and Viability of Schistocephalus solidus (Cestoda: Diphyllobothriidea).

Schistocephalus solidus plerocercoids commonly infect three-spined stickleback Gasterosteus aculeatus populations in brackish and freshwaters, but infections are typically absent from marine populations. Here we provide an experimental test of the salinity tolerance of S. solidus eggs, to determine the role of salinity in limiting the distribution of infection in coastal zones. We find that S. solidus eggs, derived from the in vitro culture of 3 different plerocercoids, developed normally in salinities of up to 12.5‰, but above this egg viability dropped rapidly, and no egg hatching was observed at salinities above 20‰. Our results are consistent with the distribution of infections in natural stickleback populations and add resolution to previous descriptive observations on salinity tolerance in S. solidus. They also demonstrate that S. solidus presents a novel disease challenge to marine populations of three-spined sticklebacks entering brackish and freshwater environments.

Egg ultrastructure of the amabiliid cestode Tatria biremis Kowalewski, 1904 (Cyclophyllidea, Amabiliidae), with an emphasis on the oncospheral envelopes.

This is the first report on the ultrastructure of eggs in the cestode family Amabiliidae Braun, 1900. The gravid proglottides of Tatria biremis easily detach from the strobila. Their thick-walled saccate uterus contains numerous rounded or oval eggs measuring about 30-32 µm in diameter. In the early preoncospheral phase, three primary embryonic envelopes are formed around the developing and differentiating embryos, namely: (1) vitelline capsule originating from vitellocyte material; (2) outer envelope formed by two macromeres, and (3) inner envelope originating from a fusion of three mesomeres. Thus, both the outer and inner envelopes of T. biremis eggs are cellular in origin and syncytial in nature. During egg maturation, the three primary embryonic envelopes undergo differentiation into fully formed oncospheral or egg envelopes. Most significant changes were observed in the inner envelope which becomes progressively subdivided into 3 sub-layers: the extra-embryophoral sub-layer, the embryophore, and the intra-embryophoral sub-layer, containing mesomere nuclei. The mature hexacanth is covered by a thin layer of the oncospheral tegument. Within the infective hexacanth larva, five cell types were distinguished: (1) a binucleated subtegumental cell; (2) U-shaped penetration gland; (3) nerve cells; (4) somatic cells representing the myocytons of both somatic and hook musculature, and (5) large germinative cells. Ultrastructural characteristics of T. biremis eggs are compared with those described in representatives of other cestode taxa. Since the functional ultrastructure of cestode egg envelopes is defined by multiple factors such as the type of life cycles, habitats and behaviour of the intermediate hosts, mode of the intermediate host infection, etc., ultrastructural studies of the greater diversity of cestodes are needed to obtain comparative data for fruitful analysis of cyclophyllidean cestode adaptations to their diverse life cycles.

Ultrastructure of the early embryonic stages of Corallobothrium fimbriatum (Cestoda: Proteocephalidea).

Cellular details of early embryogenesis have been studied extensively among cyclophyllidean cestodes, but have been reported for only 2 species of the order Proteocephalidea, both belonging to the genus Proteocephalus. Thus, we performed a detailed ultrastructural analysis of early embryos of a second species, Corallobothrium fimbriatum, including early events in the formation of the embryonic envelopes. Adult worms were collected from the small intestine of brown bullhead catfish, Ameiurus nebulosus, from the St. Lawrence River in North America and processed by standard methods for transmission electron microscopy. The vitelline capsule consists of 2 closely apposed electron-dense membranous layers, separated by a more electron-lucent material. The 2 vitellocytes that accompany each oocyte contain numerous ribosomes, vesicles, and lipid droplets. These fuse to form a vitelline syncytium, which elongates and almost completely encircles the cleaving embryo by the 4-blastomere stage, forming a partial lipid-rich cellular envelope that undergoes apoptosis as cleavage continues. This envelope is later replaced by outer and inner embryonic envelopes. The outer envelope derives from the fusion of the vitelline syncytium with the cytoplasm of macromeres, whereas the inner envelope originates from 3 mesomeres. Simultaneous to the formation of the embryonic envelopes, other blastomeres multiply and differentiate, while some micromeres undergo degeneration or apoptosis. In most respects, ultrastructural features of early C. fimbriatum embryos closely resemble those of previously studied Proteocephalus longicollis, but differ somewhat from those of other orders. This demonstrates that, despite marked ultrastructural heterogeneity within some orders such as the Cyclophyllidea, some embryonic traits distinguish cestode orders from each other.

X-ray microanalysis (EDXMA) of cadmium-exposed eggs of Bothriocephalus acheilognathi (Cestoda: Bothriocephalidea) and the influence of this heavy metal on coracidial hatching and activity.

Over recent years it has been established that pollutants can have a significant impact on host-parasite systems in the aquatic environment, so much so that it has been proposed that parasite fauna may be a useful parameter to monitor water quality. Surprisingly, with perhaps the exception of trematodes and bioaccumulation in adult acanthocephalans, detailed observations on the interaction between helminths, particularly cestodes, and pollutants such as heavy metals, are lacking. In this study, eggs of the carp tapeworm, Bothriocephalus acheilognathi were exposed to a range of cadmium concentrations (0.1, 10, 100 and 10,000 mcirog/L) and coracidial hatching and survival assessed. Results indicated that the egg is highly resistant to heavy metal pollution and hatching occurs even at 10,000 microg/L. In contrast, the activity of the liberated coracidium significantly decreased after 1h exposure to cadmium at 10 and 100 microg/L. Electron microscopic X-ray microanalysis of parasite eggs exposed to 1000 and 10,000 microg/L cadmium revealed that cadmium accumulates on the surface of the egg and does not penetrate detectably into the enclosed coracidium. This means that the parasite eggs may be able to withstand a heavy metal pollutant incident.

[Embryonic development of the cestode Mosgovoyia ctenoides (Anoplocephalidae)]. / Badania rozwoju embrionalnego tasiemca Mosgovoyia ctenoides (Anoplocephalidae).

In this study the cleavage divisions and the ultrastructural analysis of early embryos as well as cellular organisation of infective oncosphere of the anoplocephalid cestode Mosgovoyia ctenoides are described. The early cleavage is unequal and results in the formation of three types of blastomeres: 2 large macromeres containing large electron dense granules, 3 medium-size mesomeres and several small micromeres. In the early stage of oncospheral morphogenesis, formation of three following primary embryonic envelopes takes place: (1) the capsule replaced by thick, rigid outer coat originated form the uterine material secretion, (2) the outer envelope and (3) the inner envelope. The capsule is formed from the vitellocyte material. Two macromeres contribute to the formation of the outer envelope and three mesomeres take part in the formation of the inner envelope. The inner envelope undergoes differentiation into three sublayers: (1) a thick extraembryophoral cytoplasmic layer, (2) an electron-dense embryophore, as a stiff pyriform apparatus, and (3) a thin intraembryophoral cytoplasmic layer containing mesomere nuclei. The oncosphere is located in the extended cupule-like part of the pyriform apparatus. Four egg envelopes surround the mature infective oncosphere of M. ctenoides: (1) a thick outer coat, (2) the outer envelope, (3) the inner envelope with a characteristic pyriform apparatus and (4) the oncospheral membrane. Hook morphogenesis takes place inside six symmetrically arranged oncoblasts, each of which shows a characteristic large nucleus of semi-lunar shape. At the beginning the "hook-forming center" appears in the cytoplasmic part of each oncoblast. It consists of numerous free ribosomes, polyribosomes, mitochondria and Golgi complexes. The hook-forming center is involved in synthesis of a hook primordium, which undergoes differentiation and elongation into the fully developed hook. Mature hook consists of three parts: (1) blade, (2) shank, (3) base, and at the site of its protrusion from the oncosphere, is surrounded by a circular septate junction. Wide bands of hook muscles are attached to the basal and collar parts of the hook. The hook blades project outside the oncospheral body into a large cavity that is delimited by the hook region membrane. In the fully developed oncosphere of M. ctenoides three pairs of oncospheral hooks together with specialized hook muscles form a complex of "hook muscle system", responsible for coordinated hook action. The surface of the infective oncosphere is covered by a thin cytoplasmic layer of oncospheral tegument connected with the so-called "binucleate subtegumental cell", situated deeper in the oncospheral body. Below the cytoplasmic layer are situated wide bands of the somatic musculature responsible for oncospheral body movements. Five major types of oncospheral cells have been distinguished in the infective oncosphere: (1) a binucleate subtegumental cell, (2) a binucleate penetration gland, (3) two nerve cells, (4) numerous somatic cells, and (5) six germinative cells. During development of the oncosphere, changes in the concentration of glycogen and number of lipid droplets were observed. In the early embryos glycogen particles were most abundant in the macromere cytoplasm, whereas in micromeres concentration of glycogen was observed to be lower. In the course of the differentiation of the oncospheral envelopes glycogen was progressively distributed to other parts of the developing embryo. Simultaneously, a great increase in the number of lipid droplets was detected. However, during the preoncospheral phase of development a progressive reduction of lipid droplets was observed. This may indicate that lipids play a role of the energy source for developing oncosphere.

An ultrastructural study of embryonic envelope formation in the anoplocephalid cestode Mosgovoyia ctenoides (Railliet, 1890) Beveridge, 1978.

In this study the ultrastructural aspects of egg envelope formation in the anoplocephalid cestode Mosgovoyia ctenoides are described. In the early stage of oncospheral morphogenesis, formation of three following primary embryonic envelopes takes place: (1) the capsule, (2) the outer envelope, and (3) the inner envelope. The capsule is formed from the vitellocyte material. Two macromeres contribute to the formation of the outer envelope and three mesomeres take part in the formation of the inner envelope. The three primary envelopes undergo further differentiation and transformation into the secondary envelopes, the so-called oncospheral or egg envelopes. In the advanced preoncospheral phase, the inner envelope undergoes differentiation into three sublayers: (1) a thick extra-embryophoral cytoplasmic layer; (2) an electron-dense embryophore, as a stiff pyriform apparatus; and (3) a thin intra-embryophoral cytoplasmic layer containing mesomere nuclei. The oncosphere is located in the extended cupule-like part of the pyriform apparatus. The two embryophoral horns elongate and fuse, thus forming a rigid cone. Four egg envelopes surround the mature infective oncosphere of M. ctenoides: (1) a thick capsule; (2) the outer envelope; (3) the inner envelope with a characteristic embryophore, in the form of the pyriform apparatus; and (4) the oncospheral membrane. The differentiation and ultrastructure of the egg envelopes of M. ctenoides are compared, in particular to those described in other anoplocephalids, and in general to the oncospheres of other cestode species.

Vitellocytes and vitellogenesis in cestodes in relation to embryonic development, egg production and life cycle.

Vitellocytes have two important functions in cestode embryogenesis: (1) formation of hard egg-shell (e.g. Pseudophyllidea) or a delicate capsule (e.g. Cyclophyllidea), and (2) supplying nutritive reserves for the developing embryos. During evolution any of these two functions can be reduced or intensified in different taxa depending on the type of their embryonic development, degree of ovoviviparity and life cycles. Within the Cestoda, there are three monozoic taxa with only one set of genital organs: Amphilinidea, Gyrocotylidea and Caryophyllidea. In these monozoic taxa and some polyzoic groups with well developed vitellaria (e.g. Pseudophyllidea, Trypanorhyncha) a single oocyte [=germocyte] and a large number of vitellocytes (up to 30) are enclosed within a thick, hardened egg-shell, forming a type of eggs typical for the basic pattern of Neodermata. Only one type of egg-shell enclosures, the so-called 'heterogeneous shell-globule vesicle' is common for the above mentioned cestode taxa. Each membrane-bounded vesicle of mature vitellocytes contains numerous electron-dense shell globules embedded in a translucent matrix. In free-living Neoophora and Monogenea there are two types of vesicles with dense granules; the second is considered to be proteinaceous reserve material. Within the Cestoda, the numbers of vitellocytes per germocyte are reduced in those taxa forming eggs of the 'Cyclophyllidean-type' (e.g. Cyclophyllidea, Tetraphyllidea, Pseudophyllidea). This is particularly evident in Cyclophyllidea; for example, in vitellocytes of Hymenolepis diminuta (Hymenolepididae) there are numerous vitelline granules of homogeneously electron-dense material; in Catenotaenia pusilla (Catenotaeniidae) there are three large, homogenous vitelline vesicles, while in Inermicapsifer madagascariensis (Anoplocephalidae) there is only one large vitelline vesicle, containing homogeneously electron-dense material, which occupies most of the vitelline cell volume. In this respect the Tetraphyllidea and Proteocephalidea, in forming eggs that lack a hard egg-shell, hold an intermediate position. A comparison of interrelationships which exist among types of vitellocytes, vitellogenesis, types of embryonic development, ovoviviparity and life cycles indicates parallelisms and analogies in adaptation to the parasitic way of life in different groups of cestodes. Knowledge on cestode vitellogenesis may also have an important applied aspect. Vitellocytes, due to their high metabolic rate, represent a very sensitive target for analysing effect of anthelminthic drugs upon the egg formation (ovicidal effects); rapid degeneration of vitellocytes is usually accompanied by a cessation of egg production.

Differentiation and ultrastructure of oncospheral and uterine envelopes in the nematotaeniid cestode, Nematotaenia dispar (Goeze, 1782).

The oncospheral envelopes of infective eggs in Nematotaenia dispar include the outer envelope with 2 sublayers, the inner envelope with a fibrillar embryophore and 2 cytoplasmic sublayers, and the oncospheral membrane. They differentiate from 3 primary embryonic envelopes, capsule, outer and inner envelope. The uterine envelopes are formed around the early embryos by processes of uterine epithelial cells, which surround the capsules. They degenerate rapidly in later stages; however, some structural components of the uterine envelopes were still visible in gravid proglottids as flattened perikarya with pyknotic, lobate nuclei, residual membranous structures and cellular debris situated usually between eggs. The following ultrastructural features of oncospheral envelopes differentiation appear to be characteristic for N. dispar: (1) lack of the outer capsule or shell in the fully mature eggs; (2) bi-layered structure of the outer envelope and tri-layered structure of the inner envelope; (3) absence of hook region membrane resulting probably from its early disintegration; (4) presence of small vesicles or "pits" incorporated into the inner envelope plasma membrane; (5) presence of densely packed microtubules in the external layer of the inner envelope; (6) changes in number of mitochondria and free ribosomes in the external and internal layers of inner envelope during egg maturation; and (7) probable "passage" of mitochondria and free ribosomes through the embryophoral pores in the developing eggs.

Ultrastructure of oncospheral hook formation in the nematotaeniid cestode, Nematotaenia dispar (Goeze, 1782).

Ultrastructural characteristics of oncospheral hook morphogenesis in the nematotaeniid cestode, Nematotaenia dispar, are described. The primordia of embryonic hooks appear in the advanced phase of the pre-oncosphere in 6 specialised hook-forming cells or oncoblasts. Each hook primordium, situated near an invaginated part of the nucleus, is surrounded by numerous free ribosomes, mitochondria and extended Golgi regions. Simultaneously with the hook primordium elongation and transformation into a blade, handle and base, the hook material differentiates into an electron-dense cortex and a less dense, inner, crystal-like core. The exit of the blade of the mature hook, protruding from the oncosphere, is surrounded by a circular, septate desmosome and 2 rigid, dense rings on either side. The pattern of oncospheral hook morphogenesis in N. dispar is compared with that of 2 previously examined cyclophyllidean cestodes, Inermicapsifer madagascariensis and Catenotaenia pusilla.

[The life cycle and fine morphology of the embryophores in Microsomacanthus paraparvula (Cestoda: Hymenolepididae), a parasite of diving ducks in Chukotka]. / Zhiznennyi tsikl i tonkaia morfologiia zarodyshevykh obolochek Microsomacanthus paraparvula (Cestoda: Hymenolepididae) parazita nyrkovykh utok Chukotki.

It was found out, that the cestode Microsomacanthus paraparvula Regel, 1994 being a common parasite of diving ducks in Chukotka uses a caddisfly Grensia praeteria (Trichoptera) as an intermediate host in its life cycle. Mature fragments of the cestode have been collected from droppings of the experimentally infected nestling of the kittiwake Rissa tridactyla (non-specific host) and used for the fine morphology study of embryonic shells and for an infection of intermediate hosts.

Embryos and embryonic envelopes in eggs of two cyclophyllidean cestodes.

Membranes and envelopes around the embryos in eggs of Cotugnia digonopora and Raillietina (R.) echinobothrida have been studied in detail. This is the first study giving details of the egg-shell of a Cotugnia species. There are 3 membranes and an equal number of envelopes around the embryos in eggs of C. digonopora , while there are 2 membranes and only one envelope in case of eggs of R. echinobothrida . Noteworthy is the presence of a membrane in eggs of C. digonopora , termed "middle capsule", between and separating the 2 envelopes viz. the outer and inner. The membranes and envelope around the embryos in eggs of R. echinobothrida have been found to be identical with those of R. galeritae .--The 3 pairs of embryonic hooks of the 2 cyclophyllideans studied are collared --a feature of specific importance.

Ovoviviparity in platyhelminth life-cycles.

The encapsulated embryos of platyhelminths may be retained and complete their development in utero in a range of circumstances. However, hatching within the parent (the criterion of ovoviviparity) is relatively rare and larvae generally emerge only after deposition. Viviparity is characterized by the nutritional dependency of the unencapsulated larva upon the parent, but in several cases larvae retained within a shell also receive parental nutrients during intra-uterine development. Uptake of exogenous nutrients via shell pores occurs in Schistosoma mansoni but the eggs, which gain all the advantages of intra-uterine retention, are supported by host nutrients. Intra-uterine larval development avoids the hazards of development in the external environment and eliminates the time delay between oviposition and infection. Deposition of immediately infective offspring may be concentrated in time and space to exploit periods of host vulnerability. The control and precision of transmission is illustrated by examples in which the opportunity for invasion is restricted because of either host behaviour or environmental instability. This strategy has been an important factor in the evolution of polystomatid monogeneans, and its effectiveness is demonstrated by comparison of the life-cycles of Polystoma integerrimum and Pseudodiplorchis americanus. Ovoviviparity also increases reproductive potential in some polystomatids by extending the period of multiplication and by increasing established populations through internal re-infection. In Eupolystoma alluaudi, the capacity for ovoviviparity is programmed into larval development and this regulates population growth within individual hosts.

Studies on the embryogenesis of the tapeworm Cittotaenia variabilis (Stiles, 1895) using transmission and scanning electron microscopy.

The embryogenesis of the tapeworm Cittotaenia variabilis from the oocyte to the oncosphere is followed using scanning electron microscopy (SEM) in conjunction with ethanol cryofracture. Transmission electron micrographs and histochemistry are used to corroborate the inferences drawn from the SEM studies. Evidence is given for oocyte granules, the uterine epithelium, and the outer envelope as sources of components for the outer capsule. The contributions and senescence of the inner envelope and its relationship to the embryophore are given.

Histochemistry and fine structure of the bladder tegument of a larval Multiceps endothoracicus.

Certain differences were found in the histochemistry and fine structure of an active bladder tegument of an infective larva of M. endothoracicus and a regressively changing bladder of an aging larva of this species. The bladder tegument of an aging larva contained an accumulation of acid mucosubstances and phospholipids, that of a younger larva neutral mucosubstances, and it reacted less strongly than the former to tyrosine, cystine and tryptophane. Evidence was obtained with the scanning (SEM) and transmission (TEM) microscope for regressive changes in the bladder of an aging larva: its microtriches were more slender, less tightly packed and fibrously interconnected, and there were spherical formations adhering to the microthrix border. Sometimes, the vacuolation of rod-shaped bodies was so much advanced that these bodies looked like vacuoles arising to the surface of the distal cytoplasm. Another sign of bladder regression was the formation of vacuoles in the cytoplasm of subtegumental cells with contents of a granular to crystalline structure.

[Development of the neural apparatus of Triaenophorus nodulosus (Cestoidea, Pseudophyllidae) during ontogenesis]. / Razvitie nervnogo apparata Triaenophorus nodulosus (Cestoidea, Pseudophyllidea) v ontogeneze.

The anatomy of the nervous apparatus of Triaenophorus nodulosus at all stages of its life cycle was studied by means of Zherebtsov's hystochemical method. Judging by the cholinesterase activity the mass of nerve cells is situated in the oncospheres of the coracidium. At the procercoid stage the ortogonal nervous system arises with three pairs of the longitudinal nervous trunk. In plerocercoids the number of longitudinal trunks increases up to 7 pairs, the rough nervous plexus and inner plexus develop too, In mature, cestodes only secondary changes take place associated with the development of the genital system. Both in larvae and adults all longitudinal trunks are situated at the same level, on the border of the cortical and medullary parenchyma, and only the inner nervous plexus first described by the authors from cestodes passes through the medullary parenchyma. The arrangement of all elements of the nervous apparatus at the same level corresponds from the authors point of view to the most primitive state of the nervous system in the order Pseudophyllidea.

[Cleavage and gastrulation in Microsomacanthus paramicrosoma (Cyclophyllidea, Cestoda)]. / Droblenie i gastruliatsiia u Microsomacanthus paramicrosoma (Cyclophyllidea, Cestoda)

On the basis of hystological studies a description of fission and gastrulation in Microsomacanthus paramicrosoma (gasowska, 1931) is given. Eggs lacking morphological characters of polarity and symmetry display features of duet and spiral fission characteristic of lower groups of Turbellaria. Serial sections (5 mcm thick) have shown that the fission ends in the formation of coeloblastula consisting of 26 +/- 3 cells. Gastrulation is expressed in immigration into blastocoel of two micromeres which later degenerate. These micromeres are homologous to entomesodermal micromeres of other Platodes. Blastocoel disappears as a result of local reproduction of ectodermal cells that is regarded as an anlage of ectomesenchyma. Ectoderm and mesenchyma are not morphologically separated from each other.

[Embryonic development of three Cestoda from the genus Acanthobothrium (Tetraphyllidea, Onchobothriidae) (author's transl)]. / Développement embryonnaire de trois cestodes du genre Acanthobothrium (Tetraphyllidea, Onchobothriidae)

The embryonic development from the egg to the oncosphere is examined in three Cestoda: Acanthobothrium coronatum (Rud., 1819), Acanthobothrium filicolle, Zschokke, 1888 and Acanthobothrium zschokkei Baer, 1948 (Tetraphyllidea, Onchobothriidae). The three ontogeneses have in common the following data: -- Two vitelline cells pass with the zygote into the ootype where a thin shell is formed out of a material which comes from the vitelline cells. -- At first the cleavage is equal, then it becomes unequal resulting in the formation of four types of blastomeres: macromere, secondary macromere, mesomere and micromere. -- The preoncospheral phase is characterized first by the blastomere multiplication and later by their decreasing number and differentiation. -- The embryonic envelopes are formed within the shell. The vitelline layer includes the cytoplasm, a vitelline nucleus and possibly the secondary macromere, the nucleus of which always lies against the outer membrane of this envelope. The syncytial embryophore develops from mesomeres coming from the embryo. -- The oncosphere is limited by its owm membrane whose posterior region seems to double in order to form a kind of cap bending over the six hook tips. The final number of embryonic hexacantha cells is relatively low.

A comparative study of the two alternative larval forms of Hymenolepis nana, the dwarf tapeworm, with special reference to the process of excystment.

Studies of the cysticercoids of Hymenolepis nana from insects and from mouse villi revealed important differences in cyst structure and function. The insect form resists low pH unless treated with bile salts which render the cyst permeable and reduce infectivity to mice. Bile salts are not essential for scolex activation. Activation is inhibited by pH 2.5 and under and by 1% succinic acid up to pH 4.0. The importance of scolex immobility and energy conservation in relation to cyst impermeability is discussed. The villus cysticercoid has no special insulating layer. It is vulnerable to low pH and cannot infect mice orally. Bile salts are without effect and excystment occurs unaided by external agents. The structural differences between the two forms revealed by the electron microscope may be attributed to changes in the relative rates of development of the various tissues.

The fine structure of the cysticercoid of Hymenolepis diminuta. III. The scolex.

Transmission electron microscopy of the scolex of the 8-day-old Hymenolepis diminuta cysticercoid demonstrates its resemblance to the scolex of the adult. A syncytial tegument composed of external and internal layers is connected by cytoplasmic extensions. Fully developed microtriches are present. Furthermore, a basement membrane, muscle layers, and medullary region containing flame cells, nerve tissue, and other cell bodies are observed. Of particular interest is the presence of discrete sensory endings whose function is discussed.