Echinostomes and Other Intestinal Trematode Infections.

Intestinal trematodes are among the most common types of parasitic worms. About 76 species belonging to 14 families have been recorded infecting humans. Infection commonly occurs when humans eat raw or undercooked foods that contain the infective metacercariae. These parasites are diverse in regard to their morphology, geographical distribution and life cycle, which make it difficult to study the parasitic diseases that they cause. Many of these intestinal trematodes have been considered as endemic parasites in the past. However, the geographical limits and the population at risk are currently expanding and changing in relation to factors such as growing international markets, improved transportation systems, new eating habits in developed countries and demographic changes. These factors make it necessary to better understand intestinal trematode infections. This chapter describes the main features of human intestinal trematodes in relation to their biology, epidemiology, host-parasite relationships, pathogenicity, clinical aspects, diagnosis, treatment and control.

Effect of Echinostoma caproni on Presumptive Lactic Acid Bacteria Abundance and Salmonella enterica Serovar Typhimurium Colonization in the Mouse Gut.

Co-infections of mammalian hosts with intestinal helminths and bacterial pathogens are common, especially in areas with inadequate sanitation. Interactions between co-infecting species and host microbiota can cause significant changes in host immunity, disease severity, and pathogen transmission, requiring unique treatment for each case. A greater understanding of the influences of parasite-bacteria co-infections will improve diagnosis and therapeutic approaches to control infectious diseases. To study the influence of the trematode parasite Echinostoma caproni on commensal and pathogenic bacteria in the mouse gut, we examined the abundance of intestinal lactic acid bacteria and Salmonella enterica serovar Typhimurium in control mice not exposed to E. caproni (P-) or S. Typhimurium (S-), E. caproni-infected (P+S-), S. Typhimurium-infected (P-S+), and E. caproni-S. Typhimurium co-infected (P+S+) mice, and determined bacterial burdens in the livers and spleens of the P-S+ and P+S+ mice. We also examined a subset of P+S- and P+S+ mice for survival and the relative location of E. caproni in the small intestine. The numbers of presumptive lactic acid bacteria were significantly higher in the P+S+ and P-S+ mice compared to the uninfected mice, and S. Typhimurium colonization in the liver and spleen was significantly reduced in the P+S+ mice compared to the P-S+ mice. Echinostoma caproni were located anteriorly in the intestine of P+S- mice, while in the P+S+ mice, the parasites were distributed more posteriorly. Survival of E. caproni was unaffected in either group. The results of our study suggest that E. caproni facilitates a higher abundance of presumptive lactic acid bacteria in the mouse intestine and reduces colonization of S. Typhimurium in the liver and spleen of the co-infected host.

RNA-seq: The early response of the snail Physella acuta to the digenetic trematode Echinostoma paraensei.

To analyze the response of the snail Physella acuta to Echinostoma paraensei, a compatible digenetic trematode, Illumina RNA-seq data were collected from snails with early infection (5 snails at 2 days post-exposure [DPE]) and established infection (4 snails, 8 DPE), and 7 control (unexposed) snails. A reference transcriptome (325,563 transcripts, including 98% of eukaryotic universal single-copy orthologs; BUSCO) and a draft P. acuta genome (employing available genomic Illumina reads; 799,945 scaffolds, includes 88% BUSCO genes) were assembled to guide RNA-seq analyses. Parasite exposure of P. acuta led to 10,195 differentially expressed (DE) genes at 2 DPE and 8,876 DE genes at 8 DPE with only 18% of up-regulated and 22% of down-regulated sequences shared between these time points. Gene ontology (GO) analysis yielded functional annotation of only 1.2% of DE genes but did not indicate major changes in biological activities of P. acuta between 2 and 8 DPE. Increased insights were achieved by analysis of expression profiles of 460 immune-relevant DE transcripts, identified by BLAST and InterProScan. Physella acuta has expanded gene families that encode immune-relevant domains, including CD109/TEP, GTPase IMAP, Limulus agglutination factor (dermatopontin), FReD (≥82 sequences with fibrinogen-related domains), and transcripts that combine C-type lectin (C-LECT) and C1q domains, novel among metazoa. Notably, P. acuta expressed sequences from these immune gene families at all time points, but the assemblages of unique transcripts from particular immune gene families differed between 2 and 8 DPE. The shift in profiles of DE immune genes, from early exposure to parasite establishment, suggests that compatible P. acuta initially respond to infection but switch to express immune genes that likely are less effective against E. paraensei but counter other types of (opportunistic) pathogens and parasites. We propose that the latter expression profile is part of an extended phenotype of E. paraensei, imposed upon P. acuta through parasite manipulation of the host, following successful parasite establishment in the snail after 2 DPE.

The Road Not Taken: Host Infection Status Influences Parasite Host-Choice.

The manipulation of host organisms by their parasites has captured the attention of ecologists, parasitologists, and the public. However, our knowledge of parasite behavior independent of a host is limited despite the far-reaching implications of parasite behavior. Parasite behaviors can help explain trematode community structure, the aggregation of parasites within host populations, and can potentially be harnessed in biocontrol measures. In this study, we used a simple choice chamber design to examine whether trematode parasites can detect the infection status of a potential host and avoid hosts infected with a competitively dominant species. Our results show that Schistosoma mansoni, a competitively subordinate species, can detect and avoid hosts infected with a competitively dominant parasite, Echinostoma caproni. However, E. caproni, despite showing a significant preference for snails infected with S. mansoni over uninfected snails, showed little ability to detect the infection status of the host or even the host's presence. We propose subordinate species may be under stronger selection to avoid dominant competitors whereas dominant competitors may be more strongly selected to find any suitable host, regardless of infection status. Previous research has focused on parasites distinguishing between 'host' and 'non-host', which does not fully capture the complexity of these interactions. However, the ability of subordinate parasites to determine the infection status of a host results in a consistent evolutionary advantage.

Evaluation of changes in the carbohydrate metabolism of Biomphalaria glabrata Say, 1818 exposed to experimental coinfection by Angiostrongylus cantonensis (Nematoda) and Echinostoma paraensei (Trematoda).

The interaction between intermediate snail hosts and helminths can cause metabolic changes in the former. The snails use their reserves for maintenance of their vital processes, by activating the internal defense system and repairing tissue damage, while also supplying necessary energy for the parasites' development. Our aims were to evaluate the lactate dehydrogenase activity and the glucose concentration in the hemolymph of Biomphalaria glabrata experimentally coinfected by Angiostrongylus cantonensis and Echinostoma paraensei. Besides these aspects, the glycogen content in the digestive gland complex and cephalopedal mass along with histochemical changes in parasitized snails were analyzed. The snails were divided in group A (infected by 1200 L1 of A. cantonensis), group E (infected by 20 E. paraensei miracidia), group A + E (co-infected with A. cantonensis first and after a week by E. paraensei), group E + A (co-infected with E. paraensei first and then by A. cantonensis) and control group (not infected). During four weeks after exposure, samples were collected for biochemical and histochemical analyses. In the infected snails, glucose levels and glycogen content in the digestive gland complex and cephalopedal mass were significantly lower, in contrast with an increase of lactate dehydrogenase activity. These results indicate that the intense energy demand resulting from the presence of parasites causes the host snail to accelerate the anaerobic degradation of carbohydrates to obtain energy, in an attempt to maintain homeostasis. Both parasites were observed in histochemical analysis to cause tissue damages in the snails. So, although the snails were able to sustain the coinfection, several metabolic and tissue changes occurred, mainly in those infected with E. paraensei and then with A. cantonensis.

Sub-chronic exposure to a neonicotinoid does not affect susceptibility of larval leopard frogs to infection by trematode parasites, via either depressed cercarial performance or host immunity.

Little information is available on the effects of neonicotinoid insecticides on vertebrates. Previous work using amphibians found chronic exposure to some neonicotinoids had no detrimental effects on fitness-relevant traits. However, there is some evidence of more subtle effects of neonicotinoids on immune traits and evidence that other pesticides can suppress tadpole immunity resulting in elevated levels of parasitism in the exposed tadpoles. The objective of our study was to assess whether neonicotinoid exposure affected tadpole immunometrics and susceptibility to parasitic helminths. We assessed northern leopard frog tadpole (Lithobates pipiens) levels of parasitism and leukocyte profiles following exposure to environmentally relevant concentrations of clothianidin and free-living infective cercariae of a helminth parasite, an Echinostoma sp. trematode. When comparing tadpoles from controls to either 1 or 100 µg/L clothianidin treatments, we found similar measures of parasitism (i.e. prevalence, abundance and intensity of echinostome cysts) and similar leukocyte profiles. We also confirmed that clothianidin was not lethal for cercariae; however, slight reductions in swimming activity were detected at the lowest exposure concentration of 0.23 µg/L. Our results show that exposure to clothianidin during the larval amphibian stage does not affect leukocyte profiles or susceptibility to parasitism by larval trematodes in northern leopard frogs although other aspects such as length of host exposure require further study.

Endocrine and immune responses of larval amphibians to trematode exposure.

In nature, multiple waves of exposure to the same parasite are likely, making it important to understand how initial exposure or infection affects subsequent host infections, including the underlying physiological pathways involved. We tested whether experimental exposure to trematodes (Echinostoma trivolvis or Ribeiroia ondatrae) affected the stress hormone corticosterone (known to influence immunocompetence) in larvae representing five anuran species. We also examined the leukocyte profiles of seven host species after single exposure to R. ondatrae (including four species at multiple time points) and determined if parasite success differed between individuals given one or two challenges. We found strong interspecific variation among anuran species in their corticosterone levels and leukocyte profiles, and fewer R. ondatrae established in tadpoles previously challenged, consistent with defense "priming." However, exposure to either trematode had only weak effects on our measured responses. Tadpoles exposed to E. trivolvis had decreased corticosterone levels relative to controls, whereas those exposed to R. ondatrae exhibited no change. Similarly, R. ondatrae exposure did not lead to appreciable changes in host leukocyte profiles, even after multiple challenges. Prior exposure thus influenced host susceptibility to trematodes, but was not obviously associated with shifts in leukocyte counts or corticosterone, in contrast to work with microparasites.

Biological and histological changes in reproductive patterns of Biomphalaria glabrata experimentally co-infected by Echinostoma paraensei and Angiostrongylus cantonensis.

Investigation of co-infection by different helminths in snails can provide essential information about the biology of parasites co-existence under natural conditions. This study aimed to investigate the reproductive and histological changes of Biomphalaria glabrata experimentally co-infected with Echinostoma paraensei and Angiostrongylus cantonensis. Five groups of snails i.e. uninfected, with single and double infections, either E. paraensei first (E + A) or A. cantonensis first (A + E) were followed; three times a week during four weeks the numbers of egg masses, eggs and hatched snails were counted. Histological analysis of the ovotestis and albumen gland was performed after four weeks. The number of egg masses/snail, eggs/snail and hatched snails showed significative differences comparing the control group to all infected groups, especially in group E + A, with the majority of values of parameters analyzed lower than 50% of those observed for control snails. In addition, the mortality was higher in the group E + A than to control group. The histological analysis showed that presence of both parasites in the reproductive organs was associated with tissue damages.

Reproduction of Echinostoma caproni mother sporocysts (Trematoda).

The localisation and the composition of germinal material in miracidia and mother sporocysts of Echinostoma caproni were studied with the use of histological and electron microscopic methods. Germinal material in miracidia was localised in the posterior body half and was represented by 3-4 undifferentiated cells and 5-7 germinal cells. Taken together, these cells are referred to as the primordium of the germinal mass. In the mother sporocyst, germinal elements also form and develop in the germinal mass, which is located caudally. It comprises undifferentiated cells and germinal cells as well as embryos of various ages (up to the stage of 30-50 blastomeres). Germinal cells divide only by cleavage. New germinal cells are formed only from undifferentiated cells, which can proliferate in the germinal mass and nowhere else. This indicates that the germinal mass is the reproductive organ of E. caproni mother sporocyst.

Nutrient availability modulates both host and parasite life histories in a snail-trematode interaction.

Host nutrient availability can have important life history consequences for hosts and their parasites. The responses of each participant within the association can vary extensively across host-parasite systems. In this paper, we outline the life history responses of the aquatic snail Biomphalaria glabrata and its trematode parasite Echinostoma caproni during host nutrient restriction. The onset of host starvation had rapid and strong effects on snail reproduction, significantly reducing egg output in control snails and eliminating egg production in infected individuals. The combination of E. caproni infection and nutrient restriction also had a dramatic effect on B. glabrata survival, with starved infected snails dying at a faster rate than hosts from any other treatment. In terms of parasite reproduction, host nutrient restriction did not influence the quantity of parasite larvae produced after starvation onset but did influence parasite quality, reducing both larval swimming time and overall longevity. Together these results demonstrate that nutrient restriction can strongly influence both host and parasite life histories, and therefore should be considered in future studies attempting to understand patterns of disease in host populations.

Snail species diversity impacts the infection patterns of Echinostoma spp.: Examples from field collected data.

Rapid losses of biodiversity due to the changing landscape have spurred increased interest in the role of species diversity and disease risk. A leading hypothesis for the importance of biodiversity in disease reduction is the dilution effect, which suggests that increasing species diversity within a system decreases the risk of disease among the organisms inhabiting it. The role of species diversity in trematode infection was investigated using field studies from sites across the U.S. to examine the impact of snail diversity in the infection dynamics of both first and second intermediate larval stages of Echinostoma spp. parasites. The prevalence of Echinostoma spp. sporocysts/rediae infection was not affected by increases in snail diversity, but significant negative correlations in metacercariae prevalence and intensity with snail diversity were observed. Additionally, varying effectiveness of the diluting hosts was found, i.e., snail species that were incompatible first intermediate hosts for Echinostoma spp. were more successful at diluting the echinostome parasites in the focal species, while H. trivolvis, a snail species that can harbor the first intermediate larval stages, amplified infection. These findings have important implications not only on the role of species diversity in reducing disease risk, but the success of the parasites in completing their life cycles and maintaining their abundance within an aquatic system.

The distribution of echinostome parasites in ponds and implications for larval anuran survival.

Parasites can influence host population dynamics, community composition and evolution. Prediction of these effects, however, requires an understanding of the influence of ecological context on parasite distributions and the consequences of infection for host fitness. We address these issues with an amphibian - trematode (Digenea: Echinostomatidae) host-parasite system. We initially performed a field survey of trematode infection in first (snail) and second (larval green frog, Rana clamitans) intermediate hosts over 5 years across a landscape of 23 ponds in southeastern Michigan. We then combined this study with a tadpole enclosure experiment in eight ponds. We found echinostomes in all ponds during the survey, although infection levels in both snails and amphibians differed across ponds and years. Echinostome prevalence (proportion of hosts infected) in snails also changed seasonally depending on host species, and abundance (parasites per host) in tadpoles depended on host size and prevalence in snails. The enclosure experiment demonstrated that infection varied at sites within ponds, and tadpole survival was lower in enclosures with higher echinostome abundance. The observed effects enhance our ability to predict when and where host-parasite interactions will occur and the potential fitness consequences of infection, with implications for population and community dynamics, evolution and conservation.

Host food resource supplementation increases echinostome infection in larval anurans.

Host-parasite interactions are often influenced by environmental factors through multiple mechanisms. For example, changes in host food resources may affect multiple host traits (e.g., body size, behavior, immunocompetence), which may increase or decrease infection levels and the impact of parasites on host fitness. We often lack an understanding of which traits are most important for parasite transmission and fitness effects, posing challenges to predicting consequences of changing environmental conditions (e.g., eutrophication). Here, I examined the effects of food resources and host traits experimentally in a larval frog (Rana clamitans Latreille, 1801)-trematode parasite (Echinostoma revolutum Looss, 1899) system. I hypothesized that higher food resources reduce parasite infection and parasite effects on host growth and survival, due to increased host investment in parasite defenses, which I tested in a laboratory experiment. Contrary to my hypothesis, the results indicated that increased food levels enhanced infection in hosts, while the effect of parasites on survival did not depend on host food resources. A potential explanation for the positive effect of food level on infection was size-dependent infection rates (i.e., higher food levels increased infection through increased host growth), which is supported by a positive relationship between host body size and infection. These findings emphasize the complex relationship between host food resources and parasitism and the importance of environmental context and host traits (i.e., body size) in mediating interactions with parasites. The results also have relevance for conservation in light of rising anthropogenic impacts on aquatic systems and recent amphibian declines.

In vivo and in vitro effects of the herbicide Roundup(®) on developmental stages of the trematode Echinostoma paraensei.

The exposure of wildlife and humans to toxic residues of Roundup(®) through agricultural practices or the food chain has been reported since the herbicide was found contaminating rivers. Glyphosate, N-(phosphonomethyl)glycine acid, is a nonselective post-emergent herbicide and is formulated as an isopropylamine salt with the surfactant taloamine polyethoxylate (POEA) representing the commercial formulation of Roundup(®). There is little knowledge about the effects of the herbicide on helminth parasites, particularly those whose life cycle is related to water bodies. Here we investigated the effects of the Roundup(®) on the food-borne trematode Echinostoma paraensei in experimental conditions using different developmental stages (eggs, miracidia, cercariae, metacercariae, newly excysted larvae (NEL), helminths at seven days and helminths at fourteen days). Three different herbicide concentrations were tested based on concentrations typically applied in the field: 225, 450 and 900 mg/L. Specimens were analyzed in vitro for hatching miracidia, mortality and excystment rate of metacercariae and in vivo for parasitic load and egg production. There was a significant difference in the hatching miracidia rate only for the newly embryonated eggs. The mortality of specimens and excystment rate of metacercariae were concentration-dependent. There was a significant difference in the miracidia mortality with respect to concentration until 56.3 mg/L. The same effect was observed for cercariae, and mortality was observed from 15 min onwards at concentrations of 225-900 mg/L. At low concentrations, mortality was detected after 30 min. The effects of the herbicide concentration on NEL and on helminths at seven and fourteen days showed a significant difference after 24 h. There was no significant difference in parasitic load and egg production after infection of rodents with exposed metacercariae. All developmental stages of the trematode E. paraensei were affected by Roundup(®) exposure under experimental conditions. These results suggest that dynamics of transmission of the trematode could be affected in the natural environments. The study also reinforces the usefulness of this trematode as a good model organism to test pesticides regarding human and environmental health.

Animal Models for Echinostoma malayanum Infection: Worm Recovery and Some Pathology.

Echinostomes are intestinal trematodes that infect a wide range of vertebrate hosts, including humans, in their adult stage and also parasitize numerous invertebrate and cold-blooded vertebrate hosts in their larval stages. The purpose of this study was to compare Echinostoma malayanum parasite growth, including worm recovery, body size of adult worms, eggs per worm, eggs per gram of feces, and pathological changes in the small intestine of experimental animals. In this study, 6-8-week-old male hamsters, rats, mice, and gerbils were infected with echinostome metacercariae and then sacrificed at day 60 post-infection. The small intestine and feces of each infected animal were collected and then processed for analysis. The results showed that worm recovery, eggs per worm, and eggs per gram of feces from all infected hamsters were higher compared with infected rats and mice. However, in infected gerbils, no parasites were observed in the small intestine, and there were no parasite eggs in the feces. The volume of eggs per gram of feces and eggs per worm were related to parasite size. The results of histopathological changes in the small intestine of infected groups showed abnormal villi and goblet cells, as evidenced by short villi and an increase in the number and size of goblet cells compared with the normal control group.

An update on human echinostomiasis.

Echinostomiasis, caused by trematodes belonging to the family Echinostomatidae, is an important intestinal foodborne parasitic disease. Humans become infected after ingestion of raw or insufficiently cooked molluscs, fish, crustaceans and amphibians, thus, understanding eating habits is essential to determine the distribution of the disease. Despite the public health impact of echinostomiasis, it has been neglected for years. Traditionally, echinostomiasis has been considered as a minor disease confined to low-income areas, mainly in Asia. However, the geographical boundaries and the population at risk are currently expanding and changing in relation to factors such as growing international markets, improved transportation systems, new eating habits in developed countries and demographic changes. These factors make it necessary to improve our understanding of intestinal trematode infections. Herein, we review the main features of human echinostomiasis in relation to their biology, epidemiology, host-parasite relationships, pathogenicity, clinical aspects, diagnosis, treatment and control.

Effect of Infection Duration on Habitat Selection and Morphology of Adult Echinostoma caproni (Digenea: Echinostomatidae) in ICR Mice.

The course of infection of Echinostoma caproni was followed in female ICR mice, a permissive laboratory host, from infection to natural termination. Twenty-one mice were infected with 20 metacercariae via oral intubation and housed 3 per cage. Three mice from a randomly selected cage were necropsied at 1 mo intervals. A second group of 15 mice was infected approximately 1 yr later to replace mice negative at necropsy in the first group. Mice in the second group were examined weekly for the presence of eggs in the feces. Mice negative for eggs on consecutive days were killed and necropsied. The location of individual worms and worm clusters were located in 20 segments of the small intestine. Adult worms were killed and fixed in hot formalin, stained, and prepared as whole mounts. Standard measurements were taken using a compound microscope fitted with an ocular micrometer. The infection spontaneously resolved in 10 mice from 7 to 32 wk PI, indicating the host response is highly variable and extending the maximum recorded length of E. caproni infections in ICR mice to 31 wk. A moribund worm was found in the feces of an animal that continued to pass eggs for an additional 2 mo indicating individual variation in worm responses. Worms located preferentially in the ileum (segments 11-13) during the first 3 mo of the infection but shifted to the jejunum (segments 8-9) during weeks 4-6. Morphologically, worms of different ages clustered together in multivariate space, with substantial overlap between the 3- and 4-mo-old infrapopulations and between the 5- and 6-mo-old infrapopulations. Muscular structures increased in size throughout the experiment, while the gonads increased in size for the first 3 mo and then declined during the last 3 mo. The relationship between E. caproni and ICR mice is more nuanced than previously reported. The reduction in gonad size and the shift from the ileum to the jejunum in the last 3 mo likely are related. These changes might be attributable to a localized immune response by mice to E. caproni that results in the ileum becoming less hospitable and a resultant relocation of the worms to a less favorable location in the jejunum.

Auto-infection by Echinostoma spp. cercariae in Helisoma anceps.

Auto-infection is a life history strategy used by many parasitic organisms, including digenetic trematodes. The process of autoinfection most frequently involves the transfer of a life cycle stage of the parasite from one site to another inside the same host, usually accompanied by morphological transformation. Moreover, among trematodes, the stage being transferred may also move from one host to another in completing the life cycle, i.e., an indirect cycle. Echinostoma spp. parasites offer the opportunity to study auto-infection because they utilize gastropods as both first and second intermediate hosts. Rejection of a null model predicting independent infection of first and second intermediate larval stages coupled with the presence of rediae being the best predictor of metacercariae prevalence and intensity suggests that auto-infection by Echinostoma spp. cercariae is occurring in their molluscan hosts. Shell length was also found to be a significant predictor of metacercariae intensity in the snails hosts, but this is most likely attributed to larger snails being more commonly infected with Echinostoma spp. rediae as opposed to an increased likelihood of cercariae infection. Auto-infection as a life history strategy increases transmission success of the parasite, but may also have negative consequences for the parasite that necessitate auto-infection coupled with the release of cercariae to maximize transmission success and host survival.

Differential alterations in the small intestine epithelial cell turnover during acute and chronic infection with Echinostoma caproni (Trematoda).

BACKGROUND: The intestinal epithelium plays a multifactorial role in mucosal defense. In this sense, augmented epithelial cell turnover appears as a potential effector mechanism for the rejection of intestinal-dwelling helminths. METHODS: A BrdU pulse-chase experiment was conducted to investigate the infection-induced alterations on epithelial cell kinetics in hosts of high (mouse) and low (rat) compatibility with the intestinal trematode Echinostoma caproni. RESULTS: High levels of crypt-cell proliferation and tissue hyperplasia were observed in the ileum of infected mice, coinciding with the establishment of chronic infections. In contrast, the cell migration rate was about two times higher in the ileum of infected rats compared with controls, with no changes in tissue structure, indicating that an accelerated cell turnover is associated with worm expulsion. CONCLUSION: Our results indicate that E. caproni infection induces a rapid renewal of the intestinal epithelium in the low compatible host that may impair the establishment of proper, stable host-parasite interactions, facilitating worm clearance.

A re-assessment of species diversity within the 'revolutum' group of Echinostoma Rudolphi, 1809 (Digenea: Echinostomatidae) in Europe.

Species of Echinostoma Rudolphi, 1809 (Digenea: Echinostomatidae) belonging to the 'revolutum' species complex were re-examined based on material gathered in an extensive sampling programme in eight countries in Europe. The morphology of the life-cycle stages was studied in naturally and experimentally infected snail and bird hosts. A review, with an updated synonymy, is presented for six European species, including one new to science, i.e. Echinostoma revolutum (Frölich, 1802) (sensu stricto) (type-species), E. bolschewense (Kotova, 1939), E. miyagawai Ishii, 1932, E. nasincovae n. sp., E. paraulum Dietz, 1909 and Echinostoma sp. IG), and keys to the identification of their cercariae and adults are provided.