Experimental Life History and Biological Characteristics of Fasciola gigantica (Digenea: Fasciolidae)

This study was conducted to investigate the life history, morphology, and maturation of larval stages and adult worms of Fasciola gigantica in experimental mice. Lymnaea auricularia rubiginosa was used as the intermediate host, and Oryza sativa was used for encystment of the metacercariae, while Mus musculus was used as the definitive host for maturation study. Fresh eggs from the gall bladder of water buffaloes fully developed into embryonated ones and hatched out at days 11-12 after incubation at about 29ºC. Free-swimming miracidia rapidly penetrated into the snail host, and gradually developed into the next larval stages; sporocyst, redia, and daughter redia with cercariae. Fully-developed cercariae were separated from the redia and shed from the snails on day 39 post-infection (PI). Free-swimming cercariae were immediately allowed to adhere to rice plants, and capsules were constructed to protect metacercariae on rice plants. Juvenile worms were detected in intestines of mice at days 3 and 6 PI, but they were found in the bile duct from day 9 PI. Juvenile and adult flukes were recovered from 16 mice experimentally infected with metacercariae, with the average recovery rate of 35.8%. Sexually mature adult flukes were recovered from day 42 PI. It could be confirmed that experimentally encysted metacercariae could infect and develop to maturity in the experimental host. The present study reports for the first time the complete life history of F. gigantica by an experimental study in Thailand. The obtained information can be used as a guide for prevention, elimination, and treatment of F. gigantica at environment and in other hosts.     

The effect of host age on Rana temporaria-gorgoderina vitelliloba interactions

Mitchell J. B. 1982. The effect of host age on Rana temporaria-Gorgoderina vitelliloba interactions. International Journal for Parasitology12: 601–604. Two age groups of tadpoles, and newly metamorphosed and adult male Rana temporaria were fed the metacercarial cysts of Gorgoderina vitelliloba. In the younger tadpoles metacercariae died in their cysts. In the older tadpoles excystment took place and juvenile flukes invaded the kidneys, killing the hosts within 72 h. In newly metamorphosed frogs, an immunological response resulted in some of the juvenile flukes in the kidneys being attacked by eosinophils which adhered to and dissolved the tegument, presumably killing the flukes. In contrast, some young frogs were harmed by flukes in their kidneys. Migration away from the kidneys to the bladder took place on about the twelfth day after infection. Juvenile flukes in the kidneys of adult frogs 7 and 14 days after infection, evoked an inflammatory reaction involving polymorphs and lymphocytes. These cells did not appear to damage the parasites.     

In vitro cultivation of Fasciola hepatica metacercariae and of partially developed flukes recovered from mice

Attempts were made to culture the metacercariae of Fasciola hepatica under a wide variety of conditions. Of the media tested, the most successful was NCTC 135 plus 50% heat inactivated chick serum and sheep red blood cells at 37°–38°C. In this medium, somatic development of newly excysted juveniles was similar to that of flukes recovered from the liver of a mouse 11 days post-infection. There was, however, no corresponding development of the genital rudiment. Various supplements, such as liver extract, bile, yeast extract, embryo extract, egg products, monolayer cells and diphasic media were tested, but none enhanced development. The effects of various physical parameters on growth and development in vitro were examined. Cultured metacercariae appeared to be in a state of ‘suspended animation’; when injected intraperitoneally into mice they developed into egg-producing adults. Flukes recovered from the abdomen and liver of mice continued their somatic growth in vitro but their genitalia failed to develop further.     

The early migratory behaviour of young Fasciola hepatica in sensitized mice

Earlier work has shown that when mice are sensitized with irradiated metacercariae the numbers of immature flukes that can be recovered from the peritoneal cavity 2 days after reinfection with normal metacercariae is significantly less than the numbers recovered from non-sensitized control mice. Experiments are now described which investigate the reason for this difference. An inflammatory cellular reaction, most marked in sensitized mice occurs in the intestinal wall but this does not delay the migration of challenge flukes into the peritoneal cavity. No effective protective mechanism operates at the intestinal wall because similar numbers of flukes are present in the livers of sensitized and non-sensitized mice at 12 and 14 days after infection. When livers of sensitized and non-sensitized mice were examined 2 days after infection significantly more flukes had already reached the liver in the sensitized group. This indicates that immature flukes migrate more quickly from the peritoneal cavity in mice previously sensitized with irradiated metacercariae and would account for the difference in the number of flukes recovered from the peritoneal cavity of sensitized and non-sensitized mice at 2 days after infection.     

Effect of cobalt-60 irradiation on the infectivity of Paragonimus westermani metacercariae.

A study was made to observe the effect of cobalt-60 irradiation on the viability of Paragonimus westermani metacercariae in Sinopotamon chekiangense crabs. The crabs were collected in mountain regions of the Zhejiang Province of China in which paragonimiasis is endemic. Adult cats and albino mice were infected with metacercariae irradiated at different doses. Dissection of the host animals was conducted 90 or 30 days, respectively, after infection for recovery of lung flukes. Anti-metacercariae antibody in infected mice was measured by enzyme-linked immunosorbent assay (ELISA). Results showed that metacercariae were unable to grow into adult worms in cats after exposure to gamma irradiation at a dose of 0.10 kGray. However, a small number of metacercariae exposed to a dose of 2.0 kGray excysted and survived in 1 mouse for 30 days. No worm was recovered from mice when the metacercariae were irradiated at a dose of 2.5 kGray. Seropositive results by ELISA were obtained when the mice were infected with metacercariae irradiated at doses ranging from 2.0 to 3.5 kGray.     

Mucosal Immune Responses of Mice Experimentally Infected with Pygidiopsis summa (Trematoda: Heterophyidae)

Mucosal immune responses against Pygidiopsis summa (Trematoda: Heterophyidae) infection were studied in ICR mice. Experimental groups consisted of group 1 (uninfected controls), group 2 (infection with 200 metacercariae), and group 3 (immunosuppression with Depo-Medrol and infection with 200 metacercariae). Worms were recovered in the small intestine at days 1, 3, 5, and 7 post-infection (PI). Intestinal intraepithelial lymphocytes (IEL), mast cells, and goblet cells were counted in intestinal tissue sections stained with Giemsa, astra-blue, and periodic acid-Schiff, respectively. Mucosal IgA levels were measured by ELISA. Expulsion of P. summa from the mouse intestine began to occur from days 3-5 PI which sustained until day 7 PI. The worm expulsion was positively correlated with proliferation of IEL, mast cells, goblet cells, and increase of IgA, although in the case of mast cells significant increase was seen only at day 7 PI. Immunosuppression suppressed all these immune effectors and inhibited worm reduction in the intestine until day 7 PI. The results suggested that various immune effectors which include IEL, goblet cells, mast cells, and IgA play roles in regulating the intestinal mucosal immunity of ICR mice against P. summa infection.     

The ultrastructure of the tegument and the digestive caeca of in vitro cultured metacercariae of Fasciola hepatica

Davies C. 1978. The ultrastructure of the tegument and digestive caeca of in vitro cultured metacercariae of Fasciola hepatica. International Journal for Parasitology8: 197–206. The ultrastructure of the tegument and digestive caeca of metacercariae of Fasciola hepatica grown in vitro in two different media is described and compared with the development of these two systems during maturation in vivo. Although the tegument of metacercariae grown in Medium RC showed no development, that of flukes cultured in Medium CS began to produce T-1 and T-2 granules typical of the liver phase of development in vivo. The gastrodermal cells showed some degree of conversion to an adult-like morphology in vitro with the production of typical secretory granules, a limited amount of orientation of the GER and the development of junctional complexes with adjacent parenchyma cells—this was particularly evident in flukes from Medium CS. The growth achieved in each of the culture media is correlated to the degree of development of the tegument and the digestive caeca.     

On the viability of Fasciola gigantica metacercariae ingested by Lymnaea auricularia

The effect of ingestion by Lymnaea auricularia on the viability and infectivity of Fasciola gigantica metacercariae was studied. The cyst wall was unaffected by the snail's digestive processes. Two rabbits, each infected with 50 ingested metacercariae, died at 83 and 87 days post-infection. Eight and 10 immature flukes were recovered from the livers, indicating that the metacercariae had remained infective after passing through the intestine of the snail.     

Echinostoma revolutum: Freshwater Snails as the Second Intermediate Hosts in Chiang Mai,Thailand

The occurrence of 37-collar spined echinostome metacercariae in freshwater snails was investigated in 6 districts of Chiang Mai Province, Thailand, from October 2011 to April 2012. A total of 2,914 snails that belong to 12 species were examined, and 7 snail species (Clea helena, Eyriesia eyriesi, Bithynia funiculata, Bithynia siamensis siamensis, Filopaludina doliaris, Filopaludina sumatrensis polygramma, and Filopaludina martensi martensi) were found infected with echinostome metacercariae. The prevalence of metacercariae was the highest in Filopaludina spp. (38.5-58.7%) followed by B. funiculata (44.0%), E. eyriesi (12.5%), B. siamensis siamensis (8.2%), and C. helena (5.1%). Metacercariae were experimentally fed to hamsters and domestic chicks, and adult flukes were recovered from both hosts at days 15 and 20 post-infection. The adult flukes were identified based on morphological features, morphometrics, host-parasite relationships, and geographical distribution. They were compatible to Echinostoma revolutum or Echinostoma jurini, with only minor differences. As the adults were recovered from both hamsters and chicks, our specimens were more compatible to E. revolutum rather than E. jurini (reported only from mammals). This is the first report for metacercariae of E. revolutum in the snail host, C. helena, and also confirmed that Filopaludina spp., E. eryresi, and Bithynia spp. act as the second intermediate hosts of E. revolutum under natural conditions, which are indigenously distributed in Chiang Mai province.     

Serological and vaccination studies with bloodstream and culture forms of Trypanosoma musculi

Khazindar S. H. and Dusanic D. G. 1982. Serological and vaccination studies with blood-stream and culture forms for Trypanosoma musculi. International Journal for Parasitology12: 257–264. Trypanosoma musculi bloodstream forms (BSF) were collected from immunosuppressed infected mice and extracted with phosphate buffered saline (PBS). ML-15, O'Daly's and LMC media, each containing 5% fetal calf serum (FCS) and a dialysate medium were investigated to identify the medium providing the optimal growth of T. musculi culture forms (CF). Because of the ease of preparation, ML-15 containing 5% FCS was selected and the culture forms were harvested when the parasites attained concentrations of at least 1 × 10⁷ trypanosomes/ml. Cellular antigens present in PBS extracts of the BSF and CF parasites were analyzed with rabbit antisera by crossed immunoelectrophoresis and tandem crossed immuno-electrophoresis. Absorptions of rabbit antisera with CF, BSF, media and normal mouse blood extracts were performed on immunoabsorbent affinity columns prior to crossed immunoelectrophoresis to further study the unique and shared antigens of the parasites. A minimum of 13 antigens were shared by these trypanosomes. Four antigens appeared to be unique to BSF and a single antigen to CF. In immunization studies, two groups of C3H/Anf mice were immunized with the equivalent of 1 × 10⁸ frozen-thawed BSF or CF/injection. Two groups of five animals injected with PBS or uninoculated medium and one untreated group served as controls. Animals in each group received 6 injections administered at 3-day intervals. Three days following the last injection, all animals were challenged with 1 × 10⁴ BSF. Hemacytometer counts were performed every 4 days until no parasites were seen in wet blood preparations of the untreated group. None of the animals inoculated with BSF homogenate displayed parasitemias, while animals inoculated with CF homogenate were found to be infected. Parasitemias in mice immunized with CF were lower than those of the control mice.     

Scanning electron microscopic study of Opisthorchis viverrini tegument and its alterations induced by amoscanate

Scanning electron microscopic study of Opisthorchis viverrini tegument and its alterations induced by amoscanate. International Journal for Parasitology16: 19–26. When examined by scanning electron microscopy, the surface of adult Opisthorchis viverrini is covered with short microvilli that are closely packed together. Microvilli are more numerous and are taller on the ventral surface. Distributed among microvilli are two types of papillae, each one with a dome-shaped base (approx. 3 μm in diameter) with a projecting cilium in one case but not the other. Papillae are scattered in groups over the surface but are especially numerous around the suckers and laterally.When the flukes were treated with a potent schistosomicidal agent, amoscanate (C 9333-Go/CGP 4540), the tegument was damaged. Lesions that occurred on the flukes recovered from infected hamsters 1, 9, 30 and 90 days after treatment were compared with those which occurred when the flukes were exposed to the agent in vitro. Total disruption of the basic structure of the tegument was noted within 2 h of in vitro incubation with 1% amoscanate; the damage was more severe after 24 h of treatment. Flukes obtained from hamsters 24 h after treatment for 4 consecutive days with a total of 40 mg also showed tegumental lesions, including pronounced swellings into large bulbs that eventually ruptured and sloughed. However, complete regeneration of the tegument was noted within 30 days after treatment.     

Resistance of mice immunized with irradiated and lyophilized stages of Trypanosoma cruzi to infections with metacyclics

Okanla E. O., Stumpf J. L. &; Dusanic D. G. 1982. Resistance of mice immunized with irradiated and lyophilized stages of Trypanosoma cruzi to infections with metacyclics. International Journal for Parasitology12: 251–256. BALB/c mice were immunized with either irradiated or lyophilized metacyclic, epimastigote or bloodstream forms of Trypanosoma cruzi in three weekly injections of 1 × 10⁸ trypanosomes/injection. The lyophilized trypanosomes were emulsified in equal quantities of Freund's complete adjuvant. Two weeks following the final immunization, the mice were challenged subcutaneously with metacyclics obtained from either culture or the vector Triatoma infestans. The mice challenged with metacyclics from culture included groups of mice immunized with each of the three stages, while those challenged with metacyclics from the T. infestans included mice immunized with the epimastigotes or metacyclics. Mice immunized with the irradiated epimastigotes, metacyclics and blood-stream form trypomastigote challenged with metacyclics from culture exhibited reduced parasitemias compared to mice of the control groups. Parasitemias were lowest in those mice immunized with irradiated metacyclics. The parasitemias terminated in the immunized mice before those of the control animals. No protection was detected in the mice inoculated with lyophilized trypanosomes and challenged with culture metacyclics. Groups of mice injected with either irradiated or lyophilized epimastigotes or metacyclics and challenged with metacyclics from T. infestans exhibited resistance both by reduction of the parasitemias and the duration of the parasitemias when compared to the infected control animals. This study demonstrated the comparative effectiveness in mice of irradiated and lyophilized vaccines produced from either metacyclics, epimastigotes or bloodstream forms when challenged with metacyclics obtained from culture and the vector.     

Molecular Description of Macroorchis spinulosus (Digenea: Nanophyetidae) Based on ITS1 Sequences

We performed a molecular genetic study on the sequences of 18S ribosomal RNA (ITS1 region) gene in 4-day-old adult worms of Macroorchis spinulosus recovered in mice experimentally infected with metacercariae from crayfish in Jeollanam-do Province, Korea. The metacercariae were round, 180 μm in average diameter, encysted with 2 layers of thick walls, but the stylet on the oral sucker was not clearly seen. The adult flukes were oval shape, and 760-820 μm long and 320-450 μm wide, with anterolateral location of 2 large testes. The phylogenetic tree based on ITS1 sequences of 6 M. spinulosus samples showed their distinguished position from other trematode species in GenBank. The most closely resembled group was Paragonimus spp. which also take crayfish or crabs as the second intermediate host. The present study is the first molecular characterization of M. spinulosus and provided a basis for further phylogenetic studies to compare with other trematode fauna in Korea.     

Identification of Fasciola hepatica recombinant 15-kDa fatty acid-binding protein T-cell epitopes that protect against experimental fascioliasis in rabbits and mice

Vaccination with fatty acid-binding proteins (FABPs) from Fasciola hepatica has been shown to confer significant levels of protection against challenge infection in mice, rabbits, and sheep. A recombinant 15-kDa FABP (rFh15) has been purified and also shown to be an immunoprotective molecule. From the rFh15 molecule sequence 2, 12- and 10-mer putative T-cell epitopes were identified, the first an Fh15Ta of amino acid sequence IKMVSSLKTKIT, and the second an Fh15Tb of amino acid sequence VKAVTTLLKA. The synthesized oligonucleotides were cloned individually into a pGEX-2TK expression vector. The overexpressed fusion protein was affinity purified using glutathione S-transferase (GST) by competitive elution with excess reduced glutathione. These GST fusion proteins were emulsified in Freund adjuvant for rabbit immunizations or further purified as peptides after digestion with thrombin. The purified 12- and 10-mer peptides were either emulsified in Freund adjuvant for immunizations in rabbits or used in an adjuvant-adaptation (ADAD) system, followed by challenge infection with F. hepatica metacercariae in mice and rabbits. In vaccinated-challenged rabbits, the highest levels of protection were found in those treated with GST-epitopes (Fh15Ta 48.2% and Fh15Tb 59.1% reduction, respectively), as compared to GST-immunized controls. Moreover, those immunized with Fh15Ta had higher (84%) numbers of immature flukes as compared with Fh15Tb (41%) or GST alone (64%). The rabbits immunized with the putative T-cell epitopes in adjuvant had a 13% reduction in flukes in those with Fh15Ta and also were highest with immature flukes (46%). In vaccinated mice challenged with a lethal number of metacercariae, both CD-1 and BALB/c mice treated with complete ADAD-GST-Ta had the highest (40%) survival rates of all groups by 47 days postinfection. Thus the Fh15Ta and Fh15Tb polypeptide epitopes warrant further study as a potential vaccine against F. hepatica. Antibody isotype studies in mice revealed a mixed Thl/Th2 response to vaccination.     

Echinostoma macrorchis in Lao PDR: Metacercariae in Cipangopaludina Snails and Adults from Experimentally Infected Animals

The echinostome metacercariae encysted in Cipangopaludina sp. snails that were purchased from a market in Vientiane Municipality, Lao PDR, were identified as Echinostoma macrorchis (Digenea: Echinostomatidae) through recovery of adult flukes after experimental infection to rats and a cat. The metacercariae were round, 113-128 (121)×113-125 (120) µm, having a thick cyst wall, a head collar armed with collar spines, and excretory granules. The adult flukes recovered from the rats and cat at day 14 and 30 post-infection, respectively, were elongated, ventrally curved, and 3.9-6.3×0.7-1.1 mm in size. The head collar was distinct, bearing 43-45 collar spines with 5 angle spines on each side. Two testes were large (as the name implies), tandem, and slightly constricted at the middle, with irregular margins. Eggs were operculated, ovoid to elliptical, and 88-95×56-60 µm. In scanning electron microscopy, the head collar was prominent, with 43-45 collar spines. Scale-like tegumental spines were densely distributed on the ventral surface between the oral and ventral suckers. Sensory papillae were distributed mainly on the tegument around the 2 suckers. It is confirmed that E. macrorchis is distributed in Lao PDR using Cipangopaludina sp. snails as the second intermediate host.     

Echinostoma mekongi: Discovery of Its Metacercarial Stage in Snails,Filopaludina martensi cambodjensis,in Pursat Province,Cambodia

Echinostoma mekongi was reported as a new species in 2020 based on specimens collected from humans in Kratie and Takeo Province, Cambodia. In the present study, its metacercarial stage has been discovered in Filopaludina martensi cambodjensis snails purchased from a local market nearby the Tonle Sap Lake, Pursat Province, Cambodia. The metacercariae were fed orally to an experimental hamster, and adult flukes were recovered at day 20 post-infection. They were morphologically examined using light and scanning electron microscopes and molecularly analyzed by sequencing of their mitochondrial cox1 and nad1 genes. A total of 115 metacercariae (1–8 per snail) were detected in 60 (60.0%) out of 100 Filopaludina snails examined. The metacercariae were round, 174 μm in average diameter (163–190 μm in range), having a thin cyst wall, a head collar armed with 37 collar spines, and characteristic excretory granules. The adult flukes were elongated, ventrally curved, 7.3 (6.4–8.2)×1.4 (1.1–1.7) mm in size, and equipped with 37 collar spines on the head collar (dorsal spines in 2 alternating rows), being consistent with E. mekongi. In phylogenetic analyses, the adult flukes showed 99.0–100% homology based on cox1 sequences and 98.9–99.7% homology based on nad1 sequences with E. mekongi. The results evidenced that F. martensi cambodjensis snails act as the second intermediate host of E. mekongi, and hamsters can be used as a suitable experimental definitive host. As local people favor to eat undercooked snails, these snails seem to be an important source of human infection with E. mekongi in Cambodia.     

Fasciola hepatica: Simple,large-scale, in vitro excystment of metacercariae and subsequent isolation of juvenile liver flukes

A simple method has been developed for the in vitro excystment of metacercariae of Fasciola hepatica, and for the isolation of large numbers of juvenile liver flukes free from intact metacercariae and from cyst-wall material. In this method, the outer cyst wall was removed by gently grinding the metacercariae between small glass plates. The metacercariae were activated by incubation for 1 hr under $\text{60\%}\text{CO}{}_2\text{40\%}\text{N}{}_2$ and excysted by the addition of 10% sterilized sheep bile or an equivalent amount of taurocholic acid. Excystment was accomplished in an experimental apparatus allowing the newly excysted juveniles to escape from the bile-containing excystment medium into a medium with low bile content. The yield of isolated liver flukes was 60–80%; their protein content was about 125 ng. Both bile and taurocholic acid, though necessary for excystment, were detrimental to the survival of the juvenile liver flukes. The presence of bile in the host intestine may be a stimulus for juveniles to leave the gut and enter the abdominal cavity.     

A comparative ultrastructural study of in vivo and in vitro derived adults of Microphallus similis

Davtes C. 1980. A comparative ultrastructural study of in vivo and in vitro derived adults of Microphallus similis. International Journal for Parasitology10: 217–266. The ultrastructure of in vitro cultured adults of Microphallus similis was examined by TEM and SEM and compared to that of metacercariae and in vivo grown adults. In cultured flukes the most conspicuous abnormalities were observed in secretory cells, especially those of the tegument, digestive caecum, forebody glands and vitellaria. In the tegumental cells and the forebody gland cells, the secretory granules appeared to lose some of their contents within the cell bodies suggesting that there may be some defect in the transport and/or packaging of secretory granules in vitro. In the vitellaria of cultured flukes some of the granules lost their characteristic appearance, becoming ragged in outline and very electrondense. The premature tanning of the vitelline secretions within the vitellaria is correlated with abnormal egg production in vitro. The caecal cells of cultured flukes differed from those of the metacercaria and the normal adult in several important respects which suggested that their function was probably impaired.     

Prevalence of Zoonotic Trematode Metacercariae in Freshwater Fish from Gangwon-do,Korea

The infection status of zoonotic trematode metacercariae was investigated in a total of 2,293 freshwater fish collected from 11 rivers or streams in 9 administrative regions of Gangwon-do, Korea for 5 years (2009-2013). All fish were collected by netting methods and examined using the artificial digestion methods. Clonorchis sinensis metacercariae were detected in 4 fish species, i.e., Pungtungia herzi, Squalidus japonicus coreanus, Acheilognathus rhombeus, and Ladislabia taczanowskii, from only Hantangang in Cheorwon-gun. Metagonimus spp. metacercariae were found in 1,154 (50.3%) fish and their average number per infected fish was 55.8. Among the positive fish species, especially Tribolodon hakonensis from Namdaecheon in Yangyang-gun and Plecoglossus altivelis from Osipcheon in Samcheok-si were most heavily infected. Centrocestus armatus metacercariae were detected in 611 (26.7%) fish and the average metacercarial burden per infected fish was 1,032. Two chub species, Zacco platypus and Zacco temminckii were highly and heavily infected with C. armatus metacercariae in almost all regions surveyed. Echinostoma spp. metacercariae were also found in 24 fish from a few localities, but their numbers per fish infected were very low. From the above results, it is confirmed that the metacercariae of intestinal flukes, especially Metagonimus spp. and C. armatus, were heavily infected, while C. sinensis metacercariae were rarely found in fish from Gangwon-do, Korea.     

Intestinal histopathology and in situ postures of Gymnophalloides seoi in experimentally infected mice

The intestinal histopathology and in situ postures of Gymnophalloides seoi (Digenea: Gymnophallidae) were studied using C3H/HeN and C57BL/6 mice as experimental hosts; the effects of immunosuppression were also observed. The metacercariae isolated from naturally infected oysters, 300 or 1,000 in number, were infected orally to each mouse, and the mice were killed at days 3-21 post-infection (PI). In immunocompetent (IC) mice, only a small number of flukes were found in the mucosa of the duodenum and jejunum during days 3-7 PI, with their large oral suckers pinching and sucking the root of villi. The intestinal mucosa showed mild villous atrophy, crypt hyperplasia, and inflammations in the villous stroma and crypt, with remarkable goblet cell hyperplasia. These mucosal changes were almost restored after days 14-21 PI. In immunosuppressed (IS) mice, displacement as well as complete loss of villi adjacent to the flukes was frequently encountered, otherwise the histopathology was generally mild, with minimal goblet cell hyperplasia. In these mice, numerous flukes were found, and it seemed that they were actively moving and rotating in situ. Several flukes were found to have invaded into the submucosa, almost facing the serosa. These results indicate that in IC mice the intestinal histopathology caused by G. seoi is generally mild, and the flukes do not penetrate beyond the mucosa, however, in IS mice, the flukes can cause severe destruction of neighboring villi, and some of them invade into the submucosa.