Characterization of Echinostoma miyagawai in domestic ducks in Wuhu City / 中国血吸虫病防治杂志

Objective To investigate the morphological characteristics of Echinostoma miyagawai in domestic ducks in Wuhu area, and to explore the feasibility of the cytochrome oxidase subunit-1 (Cox1) gene as a molecular marker for the identification of E. miyagawai. Methods E. miyagawai was isolated from free-ranged domestic ducks in Wuhu area, and the parasites were stained and identified. In addition, the mitochondrial Cox1 gene of E. miyagawai was amplified using a PCR assay, and the amplification product was sequenced and aligned with the GenBank database to yield the homology for the identification of parasite species in combination with morphological findings. Intra-species comparison was done based on the Cox1 gene sequence. Results The prevalence of E. miyagawai infection was 16.67% in domestic ducks in Wuhu area, and the adult E. miyagawai was 6.6 to 13.2 mm in length. The size of the E. miyagawai Cox1 gene was approximately 660 bp, which had a 99.68% homology to the E. miyagawai accessed in GenBank. The morphological findings were in agreement with molecular identification. Conclusion E. miyagawai infection is common in domestic ducks in Wuhu area, and the mitochondrial Cox1 gene is a feasible marker of intra- and inter-species molecular identification of Echinostoma.

Echinostoma macrorchis Metacercariae in Cipangopaludina chinensis malleata from Xiengkhuang Province, Lao PDR and Morphologies of Adults from Experimental Animals

We identified the echinostome metacercariae in Chinese mystery snails, Cipangopaludina chinensis malleata, from Xiengkhuang Province, Lao PDR with morphologies of adult worms recovered. Total 20 snails were examined with artificial digestion method and then the collected metacercariae were orally infected to a mouse and a rat. Adult worms recovered from experimental animals were observed with a light microscope and a SEM. The metacercariae were round, 125×123 μm in average size, with a moderately thick cyst wall, collar spines distributed in the head collar and excretory granules in 2 canals of excretory tube. Adult flukes (3-week-old in a rat) were elongated, ventrally curved and 5.310×1.023 mm in average size. Head collar distinct, bearing 43 collar spines with 5 end group ones on each side. Oral sucker subterminal, prepharynx very short, pharynx well developed, and esophagus relatively short. Cirrus sac well developed, with a saccular seminal vesicle, and ventral sucker very large. Ovary round and on the median line of the body. Testes tandom and elongated. Eggs operculated, elliptical and 90×57 μm in average size. In the SEM observation, the head crown prominent, with 43 collar spines resembled with horns of younger stag. Scale-like tegumental spines were densely distributed on the surface between the head collar and ventral sucker, and their densities were decreased posteriorly. Conclusively, the metacercariae detected in C. chinensis malleata from Lao PDR were identified as those of Echinostoma macrorchis based on the morphological characteristics of adult worms.

Echinostoma ilocanum Infection in Two Residents of Savannakhet Province, Lao PDR

Adult specimens of Echinostoma ilocanum (Garrison, 1908) Odhner, 1911 (Trematoda: Echinostomatidae) were recovered from 2 riparian people who resided along the Mekong River in Savannakhet Province, Lao PDR. In fecal examinations done by the Kato-Katz technique, they revealed echinostome eggs together with eggs of Opisthorchis viverrini (and minute intestinal fluke eggs) and hookworms. To recover the adult flukes, they were treated with praziquantel 30–40 mg/kg in a single dose and purged with magnesium salts. A total of 658 adult fluke specimens were recovered from the 2 people; 456 from case 1 and 202 from case 2. Specimens from case 1 consisted of 335 echinostomes (301 E. ilocanum and 34 species undetermined), 120 O. viverrini, and 1 Haplorchis taichui, and those from case 2 consisted of 36 E. ilocanum, 134 O. viverrini, and 32 H. taichui. Thus, the number of E. ilocanum specimens was 337 in total (average per person, 168.5). From this study, it is suggested that foodborne intestinal flukes and liver flukes are highly prevalent along the Mekong River in Savannakhet Province. The present report describes for the first time human infections with E. ilocanum in Lao PDR.

Characterization of Echinostoma cinetorchis endoribonuclease, RNase H

Echinostoma cinetorchis is an oriental intestinal fluke causing significant pathological damage to the small intestine. The aim of this study was to determine a full-length cDNA sequence of E. cinetorchis endoribonuclease (RNase H; EcRNH) and to elucidate its molecular biological characters. EcRNH consisted of 308 amino acids and showed low similarity to endoribonucleases of other parasites (<40%). EcRNH had an active site centered on a putative DDEED motif instead of DEDD conserved in other species. A recombinant EcRNH produced as a soluble form in Escherichia coli showed enzymatic activity to cleave the 3′-O-P bond of RNA in a DNA-RNA duplex, producing 3′-hydroxyl and 5′-phosphate. These findings may contribute to develop antisense oligonucleotides which could damage echinostomes and other flukes.

Echinostoma macrorchis (Digenea: Echinostomatidae): Metacercariae in Cipangopaludina chinensis malleata Snails and Adults from Experimental Rats in Korea

A survey was performed to know the infection status of echinostome metacercariae in Cipangopaludina chinensis malleata snails from Korea. Total 75 snails collected in 5 localities, i.e., Imsil-gun, Jeollabuk-do, Hwasun-gun and Shinan-gun (Aphae and Jido), Jeollanam-do, and Jinju-si, Gyeongsangnam-do, were examined for metacercariae by the artificial digestion method. Infection rates of metacercariae were 80.0%, 66.7%, 100%, 60.0%, and 73.3%, and their densities were 39, 32, 183, 19, and 30 per snail infected, respectively. The metacercariae were round, 105–118×105–118 μm in size, with a thin cyst wall, collar spines on the head collar, and excretory granules in 2 canals of excretory tube. Adult flukes were elongated, ventrally curved, and 5,167×939 μm in average size. Head collar distinct, bearing 45 collar spines with 5 end groups on each side. Oral sucker subterminal, pharynx well developed, and esophagus somewhat short. Cirrus sac well developed, with a saccular seminal vesicle, and ventral sucker very large. Ovary elliptical and on the median line of the body. Testes tandem and slightly lobed. Eggs operculated, elliptical, and 90–103×55–60 μm in size. By scanning electron microscopy, the head collar was prominent with 45 collar spines resembling horns of younger stags. Scale-like tegumental spines were densely distributed on the body surface between the head collar and ventral sucker. Conclusively, it has been first confirmed that the life cycle of E. macrorchis is indigenously maintained in Korea, and C. chinensis malleata snails are popularly infected with the metacercariae of this echinostome.

Complete Mitochondrial Genome of Echinostoma hortense (Digenea: Echinostomatidae)

Echinostoma hortense (Digenea: Echinostomatidae) is one of the intestinal flukes with medical importance in humans. However, the mitochondrial (mt) genome of this fluke has not been known yet. The present study has determined the complete mt genome sequences of E. hortense and assessed the phylogenetic relationships with other digenean species for which the complete mt genome sequences are available in GenBank using concatenated amino acid sequences inferred from 12 protein-coding genes. The mt genome of E. hortense contained 12 protein-coding genes, 22 transfer RNA genes, 2 ribosomal RNA genes, and 1 non-coding region. The length of the mt genome of E. hortense was 14,994 bp, which was somewhat smaller than those of other trematode species. Phylogenetic analyses based on concatenated nucleotide sequence datasets for all 12 protein-coding genes using maximum parsimony (MP) method showed that E. hortense and Hypoderaeum conoideum gathered together, and they were closer to each other than to Fasciolidae and other echinostomatid trematodes. The availability of the complete mt genome sequences of E. hortense provides important genetic markers for diagnostics, population genetics, and evolutionary studies of digeneans.

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.

Prehistoric Pathoecology as Represented by Parasites of a Mummy from the Peruaçu Valley, Brazil

Paleopathologists have begun exploring the pathoecology of parasitic diseases in relation to diet and environment. We are summarizing the parasitological findings from a mummy in the site of Lapa do Boquete, a Brazilian cave in the state of Minas Gerais. These findings in context of the archaeology of the site provided insights into the pathoecology of disease transmission in cave and rockshelter environments. We are presenting a description of the site followed by the evidence of hookworm, intestinal fluke, and Trypanosoma infection with resulting Chagas disease in the mummy discovered in the cave. These findings are used to reconstruct the transmission ecology of the site.

Trematode Metacercariae in Freshwater Fish from Water Systems of Hantangang and Imjingang in Republic of Korea

The infection status of freshwater fish with digenetic trematode metacercariae was examined in water systems of Hantangang and Imjingang (River), the Republic of Korea. A total of 877 (594 from Hantangang and 283 from Imjingang) fishes were examined by the artificial digestion methods. Clonorchis sinensis metacercariae were detected in 7 (1.2%) fishes (in 3 spp.) from Hantangang in Cheorwon-gun, Gangwon-do, and 40 (14.1%) fishes (in 7 spp.) from Munsancheon in Paju-si, Gyeonggi-do. The average densities were 1.9 and 35.6 per fish infected, respectively. Metagonimus spp. metacercariae were detected in 312 (52.5%) and 113 (39.9%) fishes from Hantangang and Imjingang, and their average densities were 47.5 and 9.6 per fish infected, respectively. Centrocestus armatus metacercariae were found in 161 (27.1%) and 70 (24.7%) fishes from Hantangang and Imjingang, and their average number per fish infected was 694 and 82, respectively. Echinostoma spp. metacercariae were detected in 50 (8.4%) and 94 (33.2%) fishes from Hantangang and Imjingang, and their average densities were 9.6 and 23.1 per fish infected, respectively. The infection status of fishes with metacercariae of Stephanoprora spp., Diplostomum spp., Clinostomum complanatum, Metorchis orientalis, and Metorchis taiwanensis were analyzed by surveyed regions. Conclusively, it was confirmed that C. sinensis metacercariae were quite commonly detected in fishes from Munsancheon but rarely from other localities, whereas the metacercariae of other digenetic trematodes were relatively prevalent in fishes from water systems of Hantangang and Imjingang in Korea.

Epidemiology and molecular genotyping of echinostome metacercariae in Filopaludina snails in Lamphun Province, Thailand

OBJECTIVE@#To analyze the prevalence of echinostome metacercariae in Filopaludina dorliaris (F. dorliaris) and Filopaludina martensi martensi (F. martensi martensi) and genotype variation of echinostome metacercariae by using random amplified polymorphic DNA (RAPD) analysis.@*METHODS@#Filopaludina sp. snails were collected from eight localities of Lamphun Province, Northern Thailand and examined for echinostome metacercariae. RAPD-PCR was used to analyze genotype variation of echinostome metacercariae.@*RESULTS@#A total of 3 226 F. dorliaris and F. martensi martensi snails were collected from eight localities. The overall prevalences of echinostome metacercariae in F. dorliaris and F. martensi martensi were 40.89% and 36.27%, while the intensity of infection was 20.37 and 12.04, respectively. The dendrogram constructed base on RAPD profiles, 4 well supported domains were generated; (i) group of metacercariae from Ban Hong, Mae Ta, Meaung, Pa Sang, Toong Hua Chang, and Weang Nong that were clustered in the group of E. revolutum, (ii) Ban Thi, (iii) Lee, and (iv) 3 adults of an out group.@*CONCLUSIONS@#This research demonstrated RAPD profiling has been a useful tool to detect DNA polymorphisms to determine genetic relationship between echinostome metacercariae in Lamphun Province, Northern Thailand.

A Case of Echinostoma cinetorchis (Trematoda: Echinostomatidae) Infection Diagnosed by Colonoscopy

Human cases of echinostomiasis have been sporadically diagnosed by extracting worms in the endoscopy in Korea and Japan. Most of these were caused by Echinostoma hortense infection. However, in the present study, we detected 2 live worms of Echinostoma cinetorchis in the ascending colon of a Korean man (68-year old) admitted to the Gyeongsang National University Hospital with complaint of intermittent right lower quadrant abdominal pain for 5 days. Under colonoscopy, 1 worm was found attached on the edematous and hyperemic mucosal surface of the proximal ascending colon and the other was detected on the mid-ascending colon. Both worms were removed from the mucosal surface with a grasping forceps, and morphologically identified as E. cinetorchis by the characteristic head crown with total 37 collar spines including 5 end-group ones on both sides, disappearance of testes, and eggs of 108x60 microm with abopercular wrinkles. The infection source of this case seems to be the raw frogs eaten 2 months ago. This is the first case of endoscopy-diagnosed E. cinetorchis infection in Korea.

Intestinal Parasites among Wild Rodents in Northern Gangwon-do, Korea

To determine geographical patterns of natural parasite infections among wild rodents, a total of 46 wild rodents from 3 different localities in northern Gangwon-do (Province), Korea were examined for intestinal parasite infections. Along with nematodes such as hookworms and Syphacia spp., Plagiorchis muris (2 specimens) (Trematoda) were collected from striped field mice, Apodemus agrarius. In a Korean wood mouse, Apodemus peninsulae, the overall nematode infections were similar to A. agrarius, but an adult worm of Echinostoma hortense (Trematoda) was collected. In addition, 2 species of cestodes, i.e., Hymenolepis nana and Hymenolepis diminuta, were collected from A. agrarius. Through this survey, A. agrarius and A. peninsule were confirmed as the natural definite hosts for zoonotic intestinal helminths, i.e., P. muris, E. hortense, H. nana, and H. diminuta, in northern Gangwon-do, Korea. Considering increased leisure activities around these areas, seasonal and further comprehensive surveys on wild rodents seem to be needed to prevent zoonotic parasite infections.

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.

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)x113-125 (120) microm, 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 3.9-6.3x0.7-1.1 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 88-95x56-60 microm. 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.

Zoonotic Echinostome Infections in Free-Grazing Ducks in Thailand

Free-grazing ducks play a major role in the rural economy of Eastern Asia in the form of egg and meat production. In Thailand, the geographical location, tropical climate conditions and wetland areas of the country are suitable for their husbandry. These environmental factors also favor growth, multiplication, development, survival, and spread of duck parasites. In this study, a total of 90 free-grazing ducks from northern, central, and northeastern regions of Thailand were examined for intestinal helminth parasites, with special emphasis on zoonotic echinostomes. Of these, 51 (56.7%) were infected by one or more species of zoonotic echinostomes, Echinostoma revolutum, Echinoparyphium recurvatum, and Hypoderaeum conoideum. Echinostomes found were identified using morphological criteria when possible. ITS2 sequences were used to identify juvenile and incomplete worms. The prevalence of infection was relatively high in each region, namely, north, central, and northeast region was 63.2%, 54.5%, and 55.3%, respectively. The intensity of infection ranged up to 49 worms/infected duck. Free-grazing ducks clearly play an important role in the life cycle maintenance, spread, and transmission of these medically important echinostomes in Thailand.

Echinostome Flukes Receovered from Humans in Khammouane Province, Lao PDR

Echinostome eggs were detected in 22 (1.1%) of 2,074 people residing in riparian villages along the Mekong River in Khammouane Province, Lao PDR. In order to recover the adult flukes, 9 persons were treated with praziquantel and purged with magnesium salts. A total of 55 echinostome specimens (4 species) were recovered from the 9 persons. Echinostoma revolutum (8 specimens) was recovered from 3 persons, Artyfechinostomum malayanum (8 specimens) was from 2 persons, Echinochasmus japonicus (33 specimens) was from 7 persons, and Euparyphium sp. (6 specimens) was from 1 person. In Lao PDR, only human infections with E. japonicus were previously known. Therefore, the present study describes human infections with E. revolutum, A. malayanum, and Euparyphium sp. for the first time in Lao PDR. These results indicate that the surveyed villages of Khammouane Province, Lao PDR are low-grade endemic foci of echinostomiasis.

Echinostoma revolutum: Metacercariae in Filopaludina Snails from Nam Dinh Province, Vietnam, and Adults from Experimental Hamsters

We detected metacercariae of Echinostoma revolutum in Filopaludina sp. snails purchased from a local market in Nam Dinh Province for the first time in Vietnam. Adult flukes were harvested from experimentally infected hamsters at days 14 and 17 post-infection. The metacercariae were round, 170-190 microm (n=15) in diameter, with a cyst wall thickness of about 12 microm. A total of 37 collar spines were arranged around the head collar, and large excretory granules were seen in 2 canals of the excretory bladder. The 14-day old adult flukes were elongated, ventrally curved, and 5.0-7.2x0.8-1.3 mm (n=20). The head collar had a total of 37 collar spines arranged in 2 alternating rows, including 5 corner spines on each side. The cirrus sac contained a saccular seminal vesicle, a prostatic gland, and an unarmed cirrus. Two tandem testes were smooth or slightly lobed. Eggs were ovoid to elliptical, 110-118x70-75 microm. These morphological characters were similar to those of E. revolutum and E. jurini. We tentatively identified it as E. revolutum because the validity of E. jurini remains to be elucidated. The taxonomic relationship of E. revolutum and E. jurini is discussed.

Echinostoma ilocanum Infection in Oddar Meanchey Province, Cambodia

Fecal examinations using the Kato Katz technique were performed on a total of 1,287 villagers (945 students and 342 general inhabitants) of Oddar Meanchey Province, Cambodia in May 2007 and November 2009. The overall intestinal helminth egg positive rate was 23.9%, and the most prevalent helminth species was hookworms (21.6%). Other helminth eggs detected included echinostomes (1.0%), Enterobius vermicularis (0.8%), small trematode eggs (0.7%), which may include Opisthorchis viverrini and Haplorchis spp., and Hymenolepis nana (0.4%). In order to recover adult echinostomes, we treated 2 patients with 10-15 mg/kg praziquantel and purged. Total 14 adult echinostomes, 1 and 13 worms from each patient, were collected. The echinostomes characteristically had 49-51 collar spines and 2 round or slightly lobated testes. They were identified as Echinostoma ilocanum (Garrison, 1908) Odhner, 1911. So far as literature are concerned, this is the first record on the discovery of human E. ilocanum infection in Cambodia.

The effect of early infection with Echinostoma paraensei on the interaction of Schistosoma mansoni with Biomphalaria glabrata and Biomphalaria tenagophila

Infection caused by the trematode Echinostoma paraensei has been shown to interfere in the natural resistance to infection by Schistosoma mansoni. Biomphalaria glabrata is susceptible to infection, while Taim isolate Biomphalaria tenagophila is resistant to infection by S. mansoni. These two snail species were assessed for infection with E. paraensei two days after exposure to S. mansoni miracidia. The number of B. tenagophila and B. glabrata infected with E. paraensei was lower in co-infected group, suggesting an antagonistic relationship. B. glabrata showed an increase in its susceptibility to S. mansoni, whereas B. tenagophila maintained its refractoriness to S. mansoni infection. Weekly comparisons made between the E. paraensei cercariae released from B. tenagophila and B. glabrata mono-infected snails revealed no quantitative differences. In contrast, S. mansoni cercariae released were higher in the B. glabrata co-infected group. Mortality rates were significantly greater in both species pertaining to co-infected group and unexpected mortalities were also observed in B. tenagophila exposed only to S. mansoni miracidia. Our study revealed that the B. tenagophila Taim isolate is susceptible to E. paraensei infection, although infection did not alter its resistance to S. mansoni infection.

Echinostoma hortense and Heterophyid Metacercariae Encysted in Yellowfin Goby, Acanthogobius flavimanus, from Shinan-gun and Muan-gun (Jeollanam-do), Korea

Fishborne trematode (FBT) metacercariae were investigated in yellowfin goby, Acanthogobius flavimanus, collected from Shinan-gun and Muan-gun, Jeollanam-do (province), Korea. All collected fishes were examined using the artificial digestion method. In all of 15 gobies from Aphae-myeon in Shinan-gun, metacercariae of Stictodora spp. (334 metacercariae/fish), Heterophyes nocens (153/fish), and Heterophyopsis continua (20/fish) were detected. In 2 of 14 gobies from Jido-myeon in Shinan-gun, 8 Echinostoma hortense metacercariae in total were detected. In 15 gobies from Haeje-myeon in Muan-gun, the metacercariae of H. continua were found in 100%, Stictodora spp. in 86.7%, and H. nocens in 6.7% of fish examined. The average numbers of metacercariae per infected fish were 23.3 (H. continua), 416.0 (Stictodora spp.), and 2.0 (H. nocens), respectively. The metacercariae of E. hortense found in gobies were elliptical, with 150 x 138 micrometer in average size, and had 27 collar spines on the head crown. The above results suggest that yellowfin gobies from 2 localities may be the potential infection sources of FBT. Moreover, it is proved for the first time that the yellowfin goby, A. flavimanus, acts as a second intermediate host for E. hortense.