Normal and aberrant Mesocestoides tetrathyridia from Crocidura spp. (Soricimorpha) in Corsica and Spain

Tetrathyridia of Mesocestoides sp. were collected from the body cavities of the shrews (Insectivora), Crocidura russula, in Valencia, Spain and Crocidura suaveolens on the Mediterranean island of Corsica, France. Specimens were processed by routine microscopic and histological techniques, including examination with brightfield, phase-contrast, and differential-interference-contrast optics. Most tetrathyridia were clustered together inside host-derived fibrotic capsules, but some occurred free in the body cavity. All specimens examined from both locations had solid hindbodies, i.e., lacking a primary lacuna, thus conforming to the plerocercoid metacestode type; all possessed a single normal tetra-acetabulate scolex. All metacestodes from C. russula in Valencia were normal tetrathyridia. Those from C. suaveolens in Corsica were either normal tetrathyridia or had aberrant deep convolutions of an unusually elongated hindbody. No tetrathyridium from either location or host showed tegumental or excretory duct anomalies such as those reported by several authors from aberrant tetrathyridia and spargana in some other locations. No definitive evidence of asexual proliferation was visible in any of the tetrathyridia, but those with abnormally convoluted hindbodies from a single C. suaveolens in Corsica suggest the potential for asexuality by fission of the hindbody. These results add to our understanding of morphological and developmental variation among metacestodes in this widespread and variable genus.     

Tetrathyridia of Mesocestoides lineatus in Chinese Snakes and Their Adults Recovered from Experimental Animals

Morphological characteristics of Mesocestoides lineatus tetrathyridia collected from Chinese snakes and their adults recovered from experimental animals were studied. The tetrathyridia were detected mainly in the mesentery of 2 snake species, Agkistrodon saxatilis (25%) and Elaphe schrenckii (20%). They were 1.73 by 1.02 mm in average size and had an invaginated scolex with 4 suckers. Adult tapeworms were recovered from 2 hamsters and 1 dog, which were orally infected with 5-10 larvae each. Adults from hamsters were about 32 cm long and those from a dog were about 58 cm long. The scolex was 0.56 mm in average width with 4 suckers of 0.17 by 0.15 mm in average size. Mature proglottids measured 0.29 by 0.91 mm (av.). Ovaries and vitellaria bilobed and located in the posterior portion of proglottids. The cirrus sac was oval-shaped and located median. Testes were follicular, distributed in both lateral fields of proglottids, and 41-52 in number per proglottid. Gravid proglottids were 1.84 by 1.39 mm (av.) with a characteristic paruterine organ. Eggs were 35 by 27 µm in average size with a hexacanth embryo. These morphological characteristics of adult worms were identical with those of M. lineatus reported previously. Therefore, it has been confirmed that the tetrathyridia detected in 2 species of Chinese snakes are the metacestodes of M. lineatus, and 2 snake species, A. saxatilis and E. schrenckii, play the role of intermediate hosts.     

Early post-larval development of the endoparasitic platyhelminth Mesocestoides corti: trypsin provokes reversible tegumental damage leading to serum-induced cell proliferation and growth

Mesocestoides corti is a suitable in vitro model for studying the development of human endoparasitic platyhelminthes. Treatment with trypsin, supplemented with fetal bovine serum (FBS), induces M. corti development from larvae (tetrathyridia) to segmented adult worm; however, the role of this protease and of FBS in post-larval development induction remains unknown. To characterize the participation of trypsin enzymatic activity and of FBS in the induction of tetrathyridia growth and development, both stimuli were added to the larvae either together or sequentially. Additionally, specific inhibition of trypsin activity was also monitored. Finally, the effect of the enzyme on the parasite tegument as well as the proliferative activity and location of proliferating cells after induction of tetrathyridia development were also studied. We conclude that trypsin-induced tetrathyridia development to adult worm is FBS-dependent and that the effect of serum factors is dependent upon a previous trypsin-induced reversible damage to the larva tegument. In dividing and non-dividing tetrathyridia, proliferative activity of cells is mainly located within the apical massif in the anterior region and nerve cords of larvae, respectively. In tetrathyridia stimulated to develop to adult worms, an intense proliferative activity is evident along the nerve cords. Our results suggest that in natural infections the tetrathyridia tegument is temporally made permeable to growth factors by proteolytic enzyme activity in the intestine juice of the definitive host, thus leading to development to adult worms.     

Localization and associated histopathology of asexually proliferative Mesocestoides corti tetrathyridia (cestoda) infecting mouse mammary glands

Female mice of pregnant random-bred, or unmated BALB/c groups were exposed per os to Mesocestoides corti tetrathyridia and necropsied at various intervals postexposure. The right posterior subcutaneous fat pad with two mammary glands was removed from each mouse, stained, mounted whole and examined microscopically for localization of worms. The left fat pad/gland set was processed, sectioned and stained using standard histological techniques. In pregnant mice, tetrathyridia were localized primarily in the fat pad's posteroventral lobe. Unmated mice had few worms in the mammary glands or associated fat pads. The difference in infection levels between the two host groups may result from mouse strain difference or the pregnant condition of one group.     

Mesocestoides corti in the rat: The cellular response in the peritoneal cavity of rats infected with Mesocestoides corti

The tetrathyridium (second larval stage) of Mesocestoides corti elicits an extensive cellular response in the peritoneal cavity of rats which was monitored over a period of 40 days following infection. The total white cell count of female Wistar rats rose in the peritoneal cavity during the first 10 days of infection, then declined slowly. Eosinophilia was characteristic, as it is in most helminth infections. Cellular adherence to the surface of some tetrathyridia was noted. In rats infected with a large number of tetrathyridia, many parasites were found trapped in the mesenteries.     

Mesocestoides corti (syn. vogae, cestoda): characterization of genes encoding cysteine-rich secreted proteins (CRISP)

With the aim of identifying genes involved in development and parasite adaptation in cestodes, four coding sequences were isolated from the cyclophyllidean Mesocestoides corti larval stage (tetrathyridium). Genes showed significant similarity to the cysteine-rich secreted protein (CRISP) encoding genes, a large family that includes stage and tissue-specific genes from diverse organisms, many associated with crucial biological processes. The full-length McCrisp2 cDNA encodes a predicted protein of 202 residues in length, containing 10 cysteines and a putative signal peptide. The expression level of McCrisp2 was estimated by Real-time PCR, relative to GAPDH, showing an increase of 75% in segmented worms compared to tetrathyridia. By in situ hybridization, McCrisp2 expression was localized mainly at the larvae apical region of tetrathyridia and in the proglottids of segmented worms. Taken together our results suggest a possible role for M. corti CRISP proteins as ES products, potentially involved in differentiation processes as proposed for homologs in other organisms.     

Hymenolepis diminuta: The origin of protective antigens

Mice were infected orally with 1,6, or 30 cysticercoids of Hymenolepis diminuta. These were allowed to develop for different periods of time before elimination with anthelminthic, thus exposing the hosts to antigens from the prestrobilate, early strobilate, or fully strobilate worms. Other groups of mice were immunized by intraperitoneal (ip) implantation of a live strobilate worm or by ip implantation of live worms from cysticercoids excysted in vitro. Strong protection against challenge with a surgically transplanted strobilate worm was achieved by prior infection with 6 or 30 worms eliminated as early as Day 3 of infection. By this time these worms would not have strobilated. Conversely, a single worm, strobilating extensively over 16 days, stimulated only weak protection. Parenteral implantation of excysted worms protected mice but parenteral implantation of a strobilate worm had no effect. It is suggested that (i) the tapeworm protective antigens are primarily related to the scolex and/or the germinative region; (ii) the number of worms and the duration of antigenic stimulation in an immunizing infection determine the magnitude of a protective secondary response.     

Spontaneous sexual differentiation of Mesocestoides corti tetrathyridia in vitro

Barrett N. J., Smyth J. D. and Ong S. J. 1982. Spontaneous sexual differentiation of Mesocestoides corti tetrathyridia in vitro. International Journal for Parasitology12: 315–322. Tetrathyridia of Mesocestoides corti, from the body cavity of mice, maintained in the laboratory by intraperitoneal infection, were used for in vitro culture. In an initial experiment, after 50 days asexual multiplication in vitro one tetrathyridium spontaneously segmented and developed into a sexually mature adult. Further experiments were carried out in an attempt to determine the conditions favouring segmentation and sexual differentiation. A combination of 5 or 10 ml liquid medium S1OE.H (basically composed of CMRL 1066 and foetal calf serum with supplements) changed every 3 days, in a Leighton tube (19 × 105 mm), rotated at 38°C and gassed with 10 or 20% CO₂, containing between 100 and 200 tetrathyridia, has proved to be most suitable so far. Numerous adult worms with normal male and female genitalia have been obtained in this system. However, segmentation is sporadic, rather than consistent and only a few shelled eggs with hooked oncospheres have so far been obtained, suggesting that impregnation and fertilization in vitro is not fully comparable with that in vivo.     

Inequalities in size and intensity-dependent growth in a mermithid nematode parasitic in beach hoppers

Inequality in body sizes is a common feature in populations of helminth parasites, with potential consequences for egg production and population genetics. Inequalities in body lengths and the effects of intraspecific competition on worm length were studied in a species of mermithid nematode parasitic in the crustacean Talorchestia quoyana (Amphipoda: Talitridae). The majority of the 753 worms recovered were relatively small, and an analysis using a Lorenz curve and Gini coefficient suggested that there were no marked inequalities in body lengths among the worms. Total worm length in the 356 infected amphipods (i.e. the sum of the lengths of all the worms in a host) increased steadily as a function of the number of worms per amphipod, whereas the length of the longest worm per amphipod peaked in amphipods harbouring intermediate numbers of worms. This last result was not significantly accounted for by the observed increase in host size with increasing intensity of infection, but resulted from a correlation between worm length and host size. As the number of worms per amphipod increased, the relative sizes of the second-, third-, and fourth-longest worms per host increased markedly. This means that relative inequalities in sizes become less pronounced, i.e. subordinate worms get closer in size to the longest worm, as the number of worms per host increases. The main consequence of this phenomenon is that worm sizes in the mermithid population are more homogeneous than they would be if intraspecific competition had stronger effects on worm growth.     

Preventing overexploitation in a mutualism: partner regulation in the crayfish–branchiobdellid symbiosis

For a symbiosis to be a mutualism, benefits received must exceed costs incurred for both partners. Partners can prevent costly overexploitation through behaviors that moderate interactions with the other symbiont. In a symbiosis between crayfish and branchiobdellidan annelids, the worms can increase crayfish survival and growth by removing fouling material from the gills. However, overexploitation by the worms is possible and results in damage to host gills. We used behavioral observations to assess the degree to which two species of crayfish (Cambarus chasmodactylus and Orconectes cristavarius) use grooming to moderate their interaction with branchiobdellids. We found that grooming could effectively reduce worm numbers, and the proportion of total grooming directed at worms differed between crayfish species and as a function of worm number. O. cristavarius increased grooming in response to the addition of a single worm, while C. chasmodactylus only increased grooming in response to ten worms. These differences in the number of worms that trigger grooming behavior reflect differences between crayfish species in field settings. We also assessed whether antibacterial compounds in circulating crayfish hemolymph could limit bacterial gill fouling. O. cristavarius hemolymph inhibited some test bacteria more effectively than C. chasmodactylus did. Differences in the antibacterial properties of crayfish hemolymph may therefore help explain differences in both worm-directed grooming and worm loads in the field. We conclude that crayfish can use grooming to reduce worm numbers, which could lower the potential for gill damage, and that the level of grooming varies between crayfish species.     

Immunological damage to Hymenolepis diminuta following a challenge infection in C57 mice

Immunological damage to Hymenolepis diminuta following a challenge infection in C57 mice. International journal for Parasitology17: 795–803. Ultra-structural observations were made on the scpleces of Hymenolepis diminuta from 10-worm 2° infections in C57 mice to determine whether the immune stress causing reduced establishment, stunting of growth and early rejection was accompanied by significant damage to the worm fine structure. In almost all of the worms recovered there was extensive damage to tissues throughout the scolex, even in those examined 24 h post-secondary-infection (p-2°-i). Many of the muscle, tegumental and parenchyma cells were completely disrupted and dispersed, leaving only isolated strands of cytoplasm and cell debris. A number of excretory ducts were breached, and large and apparently empty spaces were evident throughout the scolex tissues. There was an increase in lipid accumulation in the tegumental syncytium, underlying cytons and in the suckers. Extensive vacuolation also occurred in the sucker tegument. Although some scoleces showed certain regions to be apparently undamaged, close examination revealed a fine structure consistent with enhanced metabolic activity. The structural abnormalities observed are discussed in relation to host immunity.     

Hymenolepis diminuta: ultrastructural abnormalities in worms from C57 mice

Ultrastructural studies, including stereological analyses of micrographs, have been made of five-worm primary infections of Hymenolepis diminuta from C57 mice to determine whether the immune destrobilation/rejection process was accompanied by significant changes in the fine structure of the scolex tegument. Destrobilation/rejection of worms occurred from Days 9-12 after infection. For the first 5 days after infection, the scolex tegument showed no detectable differences in ultrastructure compared with that of "control" worms from either Wistar rats or immunosuppressed C57 mice. By Day 6, large lipid deposits were observed in the tegument and associated musculature of worms from untreated C57 mice. Further, worms recovered from Days 6-8 after infection also showed increased activity of the Golgi apparatus, GER, and mitochondria of the tegument, resulting in increased numbers of discoidal secretory bodies. Concomitant with destrobilation/rejection from Day 9 was a drop in the number of secretory bodies, an increase in autophagic activity throughout the tegument, and a blistering of the tegument surface plasma membrane. The possible functional significance of the results is discussed in relation to host immunity.     

Effects of nickel, zinc, and lead-contaminated soil on burrowing rate and coelomocytes of the earthworm, Allolobophora chlorotica

We have shown previously that stubby worms Allolobophora chlorotica are sensitive to environmental stress, including metal-polluted soil. In order to discern the mechanisms of this sensitivity, adult (clitellate) Al. chlorotica were exposed in the laboratory to soil samples soaked with water (control) or Ni (1 and 2 mg/kg), Zn (1.25 and 2.5 g/kg) or Pb (5 and 10 g/kg) chlorides. Worms avoided contact with metal contaminants by prolonging burrowing time in metal-soaked samples, especially in the case of lead. Higher concentrations of the investigated metals were lethal for worms. During a 3 week exposure to lower metal concentrations, nickel and lead readily accumulated in the bodies of worms while zinc was efficiently regulated. However, body weights and numbers of non-invasively retrieved free coelomocytes (consisting of amoebocytes and riboflavin-loaded eleocytes) were significantly lower only in zinc-exposed worms. We assume that zinc regulation in worm bodies is more energy-demanding than nickel or lead bioaccumulation, thus this might be responsible for inhibition of the body gain and diminution of immunocompetent cells in zinc-exposed earthworms. Alternatively, missing free coelomocytes may actually be involved in Zn trafficking and removal through nephridia and/or in the formation of multicellular brown bodies, since metal can unbalance host/bacteria relationships.     

The first report of peritoneal tetrathyridiosis in squirrel monkey (Saimiri sciureus)

This report describes a case of peritoneal larval cestodiasis caused by tetrathyridia of Mesocestoides sp. in an adult female squirrel monkey. The monkey had lived in a zoological garden in Japan and had a clinical history of wasting. At necropsy, numerous whitish oval masses were found in the liver and peritoneal cavity. These masses contained larval cestodes. Morphological observation and molecular analyses of the mitochondrial 12S ribosomal RNA gene and cytochrome c oxidase subunit 1 gene sequences allowed us to identify the larva as the tetrathyridium of Mesocestoides sp. This is the first report of Mesocestoides larvae in a squirrel monkey in Japan.     

Maternal transmission of asexually proliferative Mesocestoides corti tetrathyridia (Cestoda) in mice

Asexually proliferative Mesocestoides corti tetrathyridia were studied to test the hypothesis of in utero transmission in mice and define more clearly the path of transmammary transmission. In utero transmission was not observed in 132 fetuses (22 litters) taken by caesarean section from infected mothers. However, 19 of these mothers had tetrathyridia in their mammary glands at the time of operation, nine had worms in the uterine lumen, and one had a single worm in the maternal blood space of a placenta. No tetrathyridia were found in amniotic cavities. No infection was found in 32 young (7 litters) examined immediately after birth to infected mothers, but before nursing. No infection was found in 30 young (5 litters) removed from infected mothers before nursing and raised by uninfected fosters. Of 29 uninfected young (5 litters) allowed to nurse on infected mothers, 18 became infected. Whole mounts and sections of infected mammary glands showed proliferating tetrathyridia free in larger milk ducts and free and encapsulated in mammary parenchyma. These data suggest that maternal transmission of M. corti tetrathyridia in mice occurs primarily or perhaps exclusively by the transmammary route.     

Trichinella spiralis: mediation of the intestinal component of protective immunity in the rat by multiple, phase-specific, antiparasitic responses.

Rats infected orally with Trichinella spiralis developed an immunity that was induced by and expressed against separate phases of the parasite's enteral life cycle. Infectious muscle larvae generated an immune response (rapid expulsion) that was directed against the very early intestinal infection and resulted in the expulsion of worms within 24 hr. This response eliminated more than 95% of worms in an oral challenge inoculum. Developing larvae (preadults) also induced an immune response that was expressed against adult worms. The effect on adults was dependent upon continuous exposure of worms to the immune environment throughout their enteral larval development. Immunity induced by preadult T. spiralis was not expressed against adult worms transferred from nonimmune rats. While adult worms were resistant to the immunity engendered by preadults they induced an efficient immunity that was autospecific. Both “preadult” and “adult” immunities were expressed in depression of worm fecundity as well as in the expulsion of adults from the gut. However, the two reactions differed in respect to their kinetics and their efficiency against various worm burdens. Preadult immunity was directed mainly against fecundity whereas adult immunity favored worm expulsion. All responses (rapid expulsion, preadult and adult immunity, and antifecundity) acted synergistically to produce sterile immunity against challenge infections of up to 5000 muscle larvae. These findings indicate that the host protective response to T. spiralis is a complex, multifactorial process that operates sequentially and synergistically to protect the host against reinfection.     

Formation and excretion of calcareous bodies by the metacestode (Tetrathyridium) of Mesocestoides vogae

The function of calcareous bodies, commonly found in the parenchyma of cestodes and trematodes, is relatively poorly understood. The present histochemical ultrastructural study of the proliferative tetrathyridia of Mesocestoides vogae revealed that calcareous corpuscles begin to form as organic (lipid-protein) masses that coalesce in parenchymal (calcareogenic) cells. Concentric accretion of organic and inorganic crystalline material then leads to the formation of typical refractile calcareous bodies. The precise composition of such bodies, determined by x-ray diffraction, revealed that their major inorganic constituent is indeed calcium, with significant amounts of phosphorus, silicon, and zinc as well. Emission of calcareous bodies through the tegument was observed by scanning electron microscopy, explaining their accumulation in the intracapsular spaces around worms embedded in liver tissue of the host. Following their emission, the crystalline substance of corpuscles dissolves, leaving only nonrefractile, membrane-bound cytoplasmic debris. These observations do not preclude the possibility that calcareous bodies may play some role as buffers or reservoirs of inorganic ions. However, it is difficult to accept such a function for unencapsulated worms in the coelom or intestinal lumen; we suggest that excretion is a more likely role in such sites.     

What are the evolutionary constraints on larval growth in a trophically transmitted parasite?

For organisms with a complex life cycle, a large larval size is generally beneficial, but it may come at the expense of prolonged development. Individuals that grow fast may avoid this tradeoff and switch habitats at both a larger size and younger age. A fast growth rate itself can be costly, however, as it requires greater resource intake. For parasites, fast larval growth is assumed to increase the likelihood of host death before transmission to the next host occurs. Using the tapeworm Schistocephalus solidus in its copepod first intermediate host, I investigated potential constraints in the parasite’s larval life history. Fast-growing parasites developed infectivity earlier, indicating there is no functional tradeoff between size and developmental time. There was significant growth variation among full-sib worm families, but fast-growing sibships were not characterized by lower host survival or more predation-risky host behavior. Parental investment also had little effect on larval growth rates. The commonly assumed constraints on larval growth and development were not observed in this system, so it remains unclear what prevents worms from exploiting their intermediate hosts more aggressively.     

Co-expulsion of Ascaridia galli and Heterakis gallinarum by chickens

Worm expulsion is known to occur in mammalian hosts exposed to mono-species helminth infections, whilst this phenomenon is poorly described in avian hosts. Mono-species infections, however, are rather rare under natural circumstances. Therefore, we quantified the extent and duration of worm expulsion by chickens experimentally infected with both Ascaridia galli and Heterakis gallinarum, and investigated the accompanying humoral and cell-mediated host immune responses in association with population dynamics of the worms. Results demonstrated the strong co-expulsion of the two ascarid species in three phases. The expulsion patterns were characterized by non-linear alterations separated by species-specific time thresholds. Ascaridia galli burden decreased at a daily expulsion rate (e) of 4.3 worms up to a threshold of 30.5 days p.i., followed by a much lower second expulsion rate (e = 0.46), which resulted in almost, but not entirely, complete expulsion. Heterakis gallinarum was able to induce reinfection within the experimental period (9 weeks). First generation H. gallinarum worms were expelled at a daily rate of e = 0.8 worms until 36.4 days p.i., and thereafter almost no expulsion occurred. Data on both humoral and tissue-specific cellular immune responses collectively indicated that antibody production in chickens with multispecies ascarid infections is triggered by Th2 polarisation. Local Th2 immune responses and mucin-regulating genes are associated with the regulation of worm expulsion. In conclusion, the chicken host is able to eliminate the vast majority of both A. galli and H. gallinarum in three distinct phases. Worm expulsion was strongly associated with the developmental stages of the worms, where the elimination of juvenile stages was specifically targeted. A very small percentage of worms was nevertheless able to survive, reach maturity and induce reinfection if given sufficient time to complete their life cycle. Both humoral and local immune responses were associated with worm expulsion.     

Tongue worm (subclass: Pentastomida) infection and treatment in two domesticated reptiles – A case report

Tongue worms (Subclass: Pentastomida) are endoparasites found in carnivorous reptiles, fish, amphibians, birds, and mammals. Several pentastomids cause pentastomiasis, a zoonotic disease. We encountered tongue worm infection in two reptiles imported into Japan: a Wahlberg's velvet gecko (Homopholis wahlbergii) bred in Germany and a wild-caught green tree python (Morelia viridis) from the Republic of Indonesia. The infecting tongue worms were identified as Raillietiella sp. and Armillifer sp. in the gecko and python, respectively, based on morphological and molecular analyses. Gecko is a newly recorded definitive host harboring Raillietiella tongue worm. Live tongue worms were first detected in domestic reptiles in Japan, and this indicates a risk that these tongue worms could be spread to other animals, as well as humans. Therefore, the diagnosis and treatment of infected pets are of importance. In the case of the two infected reptiles, the availability of fecal egg examination for diagnosis was shown. In addition, the effectiveness of treatment with 200 μg/kg/day ivermectin orally once weekly was demonstrated, without no side-effects and no re-detection of eggs after the cessation of treatment.