Fish-borne Zoonotic Trematodes Research Articles

Immunological and molecular evaluation of zoonotic metacercarial infection in freshwater fish: A cross-sectional analysis

Improperly cooked fish, carrying active metacercariae (MCs), can pose a significant risk for transmitting fish-borne zoonotic trematodes (FBZTs) to human consumers. This study aimed to enhance our understanding of FBZTs by conducting a comprehensive cross-sectional analysis involving various fish species, such as Nile tilapia (Oreochromis niloticus), African catfish (Clarias gariepinus), and red-belly tilapia (Tilapia zillii). These fish specimens were collected from distinct Egyptian governorates, specifically Giza, Kafr al-Shaykh, and Fayoum. The recovered flukes from experimentally infected domestic pigeons were identified as Prohemistomum vivax, Haplorchis pumilio, and Pygidiopsis genata based on morphological features. Furthermore, the identity of the retrieved adult flukes was confirmed using three species-specific primers for PCR amplification and sequencing analysis of the ITS rDNA region and have been deposited in GenBank with the following accession numbers: P. vivax (OR291421.1 and OR291422.1), P. genata (OP099561.1), and H. pumilio (OM439581.1-OP090510.1).Quantitative real-time PCR targeting the immunological genes Tumor Necrosis Factor-alpha (TNF-alpha) and Interleukin-1 (IL-1Β) was employed to compare the cellular immune response between infected with EMCs and uninfected O. niloticus. The results indicated a significant increase in TNF- and IL-1Β levels in FBZTs-infected vs un-infected fishes. Importantly, the presence of adult flukes and EMCs led to substantial histological alterations in both experimentally infected pigeons and naturally infected fish tissues. These changes included the necrosis of fish muscle bundles and a pronounced inflammatory reaction with muscular necrosis in the digestive tracts of experimentally infected pigeons.

Ultra-structure, genetic characterization and Immunological approach of fish borne zoonotic trematodes (Family: Heterophyidae) of a redbelly tilapia

Fish-borne zoonotic trematodes (FBZT) are highly significant zoonotic trematodes that can infect humans by eating raw or undercooked fish harboring active metacercaria. In this investigation, FBZT was found in samples of widely cultivated redbelly tilapia (Tilapia zillii) obtained from the Fayum governorate. Encysted metacercaria (EMC) infection was identified in fish belonging to the heterophyid family morphologically. The prevalence of heterophyid EMC was 30.5%. EMC was identified and implemented in a subsequent study on domestic pigeons (Columba livia domestica) carried out to allow adult flukes of Pygidiopsis (P.) genata; P. summa; and Ascocotyle (A.) pindoramensis species in their small intestine. This study presents the first report that combines ultra-structure, molecular approach of three species of heterophyid flukes, ultra-structure using transmission electron microscope in P. genata, and the study of host immunological responses and associated cytokines during Pygidiopsis species infection of pigeons in Egypt. Using Quantitative Real-time PCR (qRT- PCR), the gene expression levels of six cytokines (IL-1, IL-2, IL-6, IL-10, IFN-γ and TGF-β3) were assessed. The molecular confirmation of P. genata, P. summa, and A. pindoramensis have a registration in the GenBank under accession number MT672308.1, OR083433.1, and OR083431.1, respectively. Throughout the infection, the gut produced cytokines in considerably variable amounts. As a result of the Pygidiopsis species infection in pigeons, our data showed distinctive cytokine alterations, which could aid in figuring out the immunological pathogenesis and host defense mechanism against this infection. This study focused on different types of fish-borne trematodes, particularly the zoonotically important ones.

Secreted and surface proteome and transcriptome of Opisthorchis felineus

IntroductionOpisthorchis felineus, Opisthorchis viverrini, and Clonorchis sinensis are the most medically important species of fish-borne zoonotic trematodes. O. felineus is endemic to the river plains of Western Siberia and Eastern Europe, and it is estimated that more than 1.6 million people could be infected with this parasite. Chronic opisthorchiasis may lead to significant gastrointestinal and hepatobiliary pathology. This study aimed to identify and characterize proteins from the secreted and tegumental proteomes of O. felineus. MethodsAdult flukes were collected from experimentally infected hamsters and cultured in vitro in serum-free media. We extracted proteins from different compartments of the O. felineus secretome, including (i) soluble excretory/secretory (ES) products; (ii) secreted 15K-extracellular vesicles (EVs); and (iii) tegument. ResultsWe also generated a transcriptome using long-read sequencing, and when this was combined with high-resolution mass spectrometry, sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS-PAGE) separation, and protein digestion, we identified 686, 894, 389, 324, and 165 proteins from the ES, 15K-EV, and the three sequentially extracted tegument (TEG) protein fractions, respectively. We conducted in-depth gene ontology and protein family analyses on the identified proteins and discussed comparisons against similar proteome data sets acquired for the Southeast Asian liver fluke O. viverrini and the Chinese liver fluke C. sinensis. DiscussionThe information from this study will form a biologically relevant data set of O. felineus proteins that could be used to develop diagnostic and therapeutic tools to manage the human cost of O. felineus infection and its associated comorbidities.

Prevalence of fish-borne zoonotic trematode infection in Jilin Province, China.

Fish-borne zoonotic trematodes (FZTs) are the most serious food-borne parasites in Asia and have become a burden to public health and a new challenge in food safety. In Jilin Province, China, the prevalence of FZTs in intermediate and definitive hosts has not been extensively explored. In the present study, we investigated the prevalence of FZTs in Jilin Province, China. From July to November 2020, a total of 132 freshwater snails (the first intermediate host of FZTs), 4122 wild freshwater fishes (the second intermediate host of FZTs) and 143 fecal samples from canines, ducks and swine (the definitive host of FZTs) were collected from the Yitong River basin of Jilin Province. FZT species were identified by morphological observation combined with internal transcribed spacer (ITS) sequence analysis. The prevalence of FZTs was then calculated. The results showed that the prevailing species of FZTs in Jilin Province, China, were Clonorchis sinensis, Metorchis orientalis and Echinochasmus japonicus. The total prevalence of FZTs was 29.74% (1226/4122) in fish, the total infection rates were 2.27% (3/132) in snails, 75.00% (21/28) in canines and 37.18% (29/78) in ducks. The coinfection rates of the two trematodes were 13.39% (552/4122) in fish, 35.71% (10/28) in canines and 7.69% (6/78) in ducks. The coinfection rate of the three flukes was 2.60% (107/4122) in fish. Nine of the 12 fish species examined were infected with FZT metacercariae.

Infection rate of Opisthorchis viverrini metacercariae in cyprinoid fish from the markets and its association to human opisthorchiasis in the local community in the Northeast Thailand

The prevalence of Opisthorchis viverrini, a fish-borne zoonotic trematode that can provoke cholangiocarcinoma, is high in the Northeast Thailand. The aim of this study was to determine the prevalence of O. viverrini metacercariae in the cyprinid fish and determine its association of O. viverrini infection among the consumers who regularly buy fish in the markets. A cross-sectional study was conducted in nine provinces covering 20 districts of Thailand, and we examined 778 cyprinoid fish specimens belonging to five species purchased from local markets. Pepsin-HCl digestion method was used to recover O. viverrini metacercariae from fish. In all districts surveyed, O. viverrini metacercariae-positive fish were found with the infection rates ranging from 3.9 to 21.1%. All five fish species studied were positive for O. viverrini metacercariae: Henicorhynchus siamensis (13.7%), Cyclocheilichtys spp. (12.7%), Hampala spp. (8.1%), Systomus spp. (6.9%) and Barbonymus goniatus (5.0%). An average prevalence of O. viverrini infection was 7.1% in the fish consumers surveyed in the markets. The source of fish was determined and our results showed that parasitized fish are sold in markets up to 100 km away from the point of capture, which contributes to the dispersion and maintenance of this helminthiasis. Our results point to the transmission of liver flukes via markets, in spite of many active programs of health education, elimination, prevention and control aimed to reduce O. viverrini infection and subsequent cholangiocarcinoma in the endemic areas of Thailand.

Epidemiological study of fish‐borne zoonotic trematodes infecting Nile tilapia with first molecular characterization of two heterophyid flukes

Fish-borne zoonotic trematodes (FBZT) are extremely important zoonotically and can infect humans via the consumption of poorly cooked fish containing active metacercariae. In this context, the present study aimed to update the epidemiological information of FBZT among Nile tilapia (Oreochromis niloticus) collected from Giza and Fayoum Governorates. Concerning the molecular characterization and phylogenetic analysis of adult flukes from experimentally infected pigeons and histopathological alterations of both larval and adult trematode flukes. Oreochromis niloticus were found to be infected with different encysted metacercaria (EMC); Prohemistomum, Haplorchis and Pygidiopsis species from wild caught in Giza and farmed fish in Fayoum with the total mean of prevalence that 81.89 ± 6.48, 18.03 ± 2.9 and 34.64 ± 3.42 respectively. Three recovered flukes from experimentally infected domestic pigeons (Columba livia domestica); Prohemistomum vivax, Haplorchis pumilio and Pygidiopsis genata in their small intestine. First molecular characterization and phylogenetic analysis of heterophyid flukes; P. genata and H. pumilio. The sequences obtained in this study were registered in the GenBank with accession numbers (MT672308.1 and MT707975.1) respectively. Moreover, constructing a phylogeny and phylogenetic relationships between two heterophyid species was performed through analytic study of the second internal transcribed spacer nuclear ribosomal genes (rDNA-ITS2). Phylogenetic analysis of P. genata and H. pumilio showed 99.42% nucleotide similarity with that sequence from Israel (AY245710) and 99.71% from Vietnam (EU826636.1). In addition, histopathological alterations of EMC and adult flukes induced necrosis of fish muscle bundles and a severe inflammatory response with muscular necrosis in intestinal tract of infected pigeons.

Prevalence of Clonorchis sinensis infection in fish in South-East Asia: A systematic review and meta-analysis.

Clonorchis sinensis, an important fish-borne zoonotic trematode, is widely distributed in South-East Asia, especially in China. Infections from human and animal reservoir hosts occur due to the consumption of raw or undercooked fish with C. sinensis metacercariae. This study aimed to evaluate the prevalence of C. sinensis metacercariae in fish in South-East Asia via systematic review and meta-analysis. We searched PubMed, ScienceDirect, China National Knowledge Infrastructure, Wanfang and Chongqing VIP databases for studies published between 1976 and 2020 that are related to the prevalence of C. sinensis metacercariae in fish. Studies were screened with keywords based on inclusion and exclusion criteria. Seventy-one eligible articles were identified, covering three countries: China, Korea and Vietnam. The pooled prevalence of C. sinensis metacercariae in fish from South-East Asia was 30.5%, with 35.1% in China, 29.7% in Korea and 8.4% in Vietnam. In subgroup analyses of climate, season, water source and publication date, the highest prevalence was identified in the Dwb climate type (43.3%), summer (70.2%), river (34.5%) and pre-2001 publications (38.9%), respectively. In comparison, the lowest prevalence was found in the Dfa climate type (14.5%), winter (19.5%), lake (8.0%) and post-2001 publications (23.8%). Meta-regression results indicated that country (p=.009), the published time (p=.035) and water source subgroups (p=.003) may be the source of heterogeneity. Overall, our study indicates that a high prevalence of C. sinensis infections occurs in fish in China, Korea and Vietnam, illuminating a significant public health concern in these countries.

Haplorchis pumilio (Trematoda: Heterophyidae) as a new fish-borne zoonotic agent transmitted by Melanoides tuberculata (Mollusca: Thiaridae) in Brazil: A morphological and molecular study

The fishborne zoonotic trematode Haplorchis pumilio (Looss, 1896) is here reported for the first time in Brazil based on morphological, experimental and molecular studies. Pleurolophocercous cercariae emerged from the invasive snail Melanoides tuberculata collected in the municipality of Ceará-Mirim, state of Rio Grande do Norte, Northeast region of Brazil, in November 2018, were used for experimental infection of guppies, Poecilia reticulata. Metacercariae were extracted from the caudal musculature of these fishes and were used to infect mice. Adult parasites recovered in the small intestine of the mice were morphologically identified as H. pumilio. Molecular sequences were obtained for the Brazilian and Peruvian isolates of H. pumilio, and were compared with data available in GenBank. Analyses of fragments of the nuclear genes 28S (1219bp) and ITS-2 (290 bp) revealed 98.48-100% similarity between the South American and Asian isolates of H. pumilio. Moreover, new sequences of the mitochondrial gene cox-1 obtained for the Brazilian (797bp) and Peruvian (646pb) isolates were 100% similar to a Mexican isolate of this species and 97.54% similar to an isolate from Thailand. This finding reveals the potential for occurrence of human haplorchiasis in Brazil, especially because of the increasing popularity of raw fish dishes in the country.

Discovery of Carcinogenic Liver Fluke Metacercariae in Second Intermediate Hosts and Surveillance on Fish-Borne Trematode Metacercariae Infections in Mekong Region of Myanmar.

Countries of lower Mekong regions are highly alarmed by the spread of fish-borne trematode infections, i.e., small liver flukes and minute intestinal flukes especially in Thailand, Lao People’s Democratic Republic (Lao PDR), Vietnam, Cambodia and Myanmar. Moreover; the incidence of cholangiocarcinoma has also been increasing in the endemic area of liver fluke infections. Only a few reports have been published concerning the fish-borne trematodes infections in the central region of Myanmar. However; there is still a lack of information regarding the status of trematodes infections in second intermediate hosts in the Mekong region of Myanmar. Therefore, we conducted surveillance on the distribution of trematode metacercariae in small cyprinoid fishes collected from the Mekong region of Myanmar. A total of 689 fishes (12 different species of cyprinoid fishes) have been collected and examined by pepsin digestion methods. We discovered four species of fish-borne trematode metacercariae infections, i.e., carcinogenic liver fluke, Opisthorchis viverrini; minute intestinal flukes, Haplorchis taichui; Haplorchis pumilio and Haplorchoides sp. in Tachileik, the Mekong Region of Myanmar. The outcome of this study could be a useful index for the fish-borne zoonotic trematode epidemiology in the Mekong area. Besides, the results of our study contribute to filling the gap of information necessary for the control and prevention of fish-borne trematode zoonotic infections in the Mekong region.

Influence of Water Irrigation Schemes and Seasonality on Transmission Dynamics of Opisthorchis viverrini in the Snail Intermediate Host, Bithynia siamensis goniomphalos in Rice Paddy Fields in Northeast Thailand.

Opisthorchis viverrini is a fish-borne zoonotic trematode that causes significant public health problems in Southeast Asia. Its life cycle requires Bithynia snails as the first intermediate hosts, fish, and human and/or carnivore hosts. This study assessed impacts of land use practice for rice cultivation and seasonality on the transmission dynamics of O. viverrini in Bithynia siamensis goniomphalos in rice paddy field habitats. The transmission of O. viverrini cercariae in B. s. goniomphalos was monitored at monthly intervals over a 4-year period from January 2010 to December 2013. From a total of 59,727 snails examined by standard cercarial shedding, the prevalence of O. viverrini was 0.7% (range, 0.0-4.1%). The prevalence of O. viverrini infection in B. s. goniomphalos varied with the amount of rainfall, with peaks of infection occurring in the cool-dry season, that is, after each rainy season. A shift of peak prevalence from cool-dry to hot-dry season observed in 2013 was associated with the increase in preceding water irrigation to support the production of second annual rice crop. Significant positive correlations were found between the prevalence and intensity of cercarial infection and the size of snails. Our results revealed substantial variation between years so that to have a clear understanding of the population dynamics of this complex system, studies should be conducted over an extended period (> 1 year). Results from this study highlight that water irrigation schemes in rice paddy cultivation and seasonality have a significant effect on the prevalence of O. viverrini in B. s. goniomphalos.

Seasonal prevalence of zoonotic trematode parasites in commercial catfish species in the Ho Chi Minh City area

An epidemiological study of fishborne zoonotic trematodes (FZT) infecting three major catfish species, hybrid catfish (Clarias sp.), river catfish (Pangasianodon hypopththalmus) and black-spotted catfish (Pangasius larnaudii), produced locally in Ho Chi Minh City, Vietnam, was conducted during 2017-2018. The impact of seasonal changes associated with the wet and dry seasons was assessed. The only species of FZT found was Haplorchis pumilio, a new host record for black-spotted catfish. Farm prevalence of H. pumilio was highest for black-spotted catfish (20.0 %) and hybrid catfish (16.7 %), and lowest for farms producing river catfish (5.0 %). The FZT prevalence in hybrid catfish (N = 300 per season) was higher in the wet season (2.0 %) than in the dry season (0.3 %); however, it was not significantly different (P > 0.05). For all three catfish species (N = 550 per season), seasonal prevalence was significantly higher (P < 0.05) in the wet season than in the dry season (1.5 % versus 0.2 %). These results demonstrate that, overall, most aquaculture farmers in this area employed management practices that were effective in lowering the risk of parasite infection in grow-out catfish compared with prevalence data for catfish reported from the Mekong Delta. However, the basis for the seasonal effect is not clear. The increase in snail intermediate host populations and pond contamination with parasites eggs from surface run-off are risk factors, generally, but the significantly higher impact on hybrid catfish production requires further investigation.

Comparative analyses of complete ribosomal DNA sequences of Clonorchis sinensis and Metorchis orientalis: IGS sequences may provide a novel genetic marker for intraspecific variation

Both Clonorchis sinensis and Metorchis orientalis are the fish–borne zoonotic trematodes, and have a wide distribution of southeastern Asia, especially in China. Due to the similar morphology, life cycle, and parasitic positions are difficult to differentiate between both metacercariae. In the present study, the complete rDNA sequences of five C. sinensis and five M. orientalis were obtained and compared for the first time. And the IGS rDNA sequences were tested as a genetic marker. The results showed complete rDNA lengths of C. sinensis were range from 8049 bp to 8391 bp, including 1991 bp, 1116 bp, 3854 bp, and 1088–1430 bp belonging to 18S, ITS, 28S and IGS, respectively. And the complete rDNA lengths of M. orientalis were range from 7881 bp to 9355 bp, including 1991 bp, 1077 bp, 3856 bp, and 957–2431 bp belonging to 18S, ITS, 28S and IGS, respectively. Comparative analyses reveal length difference main in IGS, which has higher intraspecific and interspecific variations than other ribosomal regions. Forty four repeat (forward and inverted) sequences were found in the complete rDNAs of C. sinensis and M. orientalis. The phylogenetic analyses showed that the sequences of ITS1, ITS2, 18S and 28S could be used as different level genetic markers. In IGS phylogenetic tree, Opisthorchiidae, Paramphistomidae, Dicrocoeliidae, and Schistosomatidae formed monophyletic groups, and the same length sequences were clustered together in the same species. These findings of the present study provide the new molecular data for studying the complete rDNA of C. sinensis and M. orientalis, and indicate IGS sequences may used as a novel genetic marker for studying intraspecific variation in trematodes.

Prevalence of trematode in wild fish from fishing in Ho Chi Minh City

A research on the prevalence of fishborne zoonotic trematodes (FZT), metacercariae stage, in wild fish from fishing in Nhieu Loc – Thi Nghe, Tau Hu – Ben Nghe and Thay Cai – An Ha canals was conducted in Ho Chi Minh City in the period of May 2017 and April 2018. The results showed that no metacercariae were found in fish in Nhieu Loc – Thi Nghe and Tau Hu – Ben Nghe canals. However, wild fish in Thay Cai – An Ha canal were infected with Haplorchis pumilio and Centrocestus formosanus. Metacecariae were infected in silver barb (80%), climbing perch (11.7%), snakehead (7.5%), and walking catfish (1.7%). The FZT prevalence in the wet season is significantly higher than in the dry season (P

Metacercaria Infection Status of Fishborne Zoonotic Trematodes, Except for Clonorchis sinensis in Fish from the Heilongjiang Province, China.

To investigate the metacercarial infections of fishborne zoonotic trematodes (FZT), a total of 6815 freshwater fish (in representing 13 species of 5 families) were collected from Songhua river (n = 2636), Nenjiang river (n = 1935), Mudanjiang river (n = 301), and other lakes or ponds (n = 1943) in 36 representative regions in Heilongjiang Province, China, from August 2012 to December 2015. Metacercariae of four FZT species, that is, Clonorchis sinensis, Metorchis orientalis, Isthmiophora hortensis, and Metagonimus yokogawai, metacercariae were detected in the examination by the artificial digestion method. As the partial data for C. sinensis were previously reported, the remaining three FZT species are to be treated in this study. The overall prevalence of M. orientalis, I. hortensis, and M. yokogawai, metacercariae was 10.54%, 0.28%, and 1.35%, respectively. Metacercariae of M. orientalis were detected in seven fish species, that is, Pseudorasbora parva, Hemiculter leucisculus, Saurogobio dabryi, Rhynchocypris lagowskii, Carassius auratus, Rhodeus ocellatus and Perccottus glehnii. Their prevalences were the highest in false dace, P. parva (26.81%), and in fish from Songhua river (17.94%). Metacercariae of I. hortensis were detected in only one fish species, Misgurnus anguillicaudatus, from Nenjiang river only. Metacercariae of M. yokogawai were detected in three fish species, that is, P. parva, H. leucisculus and S. dabryi. Their prevalences were the highest in sharpbelly, H. leucisculus (6.05%), and in fish from Mudanjiang river (5.65%). This study first demonstrated the existence of M. orientalis, I. hortensis, and M. yokogawai in freshwater fish from Heilongjiang Province, posing a major public health concern. Eight fish species, namely M. anguillicaudatus, P. parva, H. leucisculus, S. dabryi, R. lagowskii, C. auratus, R. ocellatus, and P. glehnii, cannot be eaten raw. Moreover, the findings of this study not only extended the second intermediate host range of FZT, but also improve the information of the distribution of FZT in China.

Comparative Risk of Liver and Intestinal Fluke Infection from Either Wild-Caught or Cultured Fish in Vietnam.

A survey for fish-borne zoonotic trematode (FZT) metacercariae in fish from the Northern Mountain Region of Vietnam was conducted from May to August 2014. The major objective was to evaluate the impact of water body types on the prevalence of FZT. A total of 846 fish (31 species) were collected from three water body types, including aquaculture ponds, rivers, and reservoirs. A pepsin digestion method was used for the recovery of metacercariae from fish. Overall, 29 of the 31 fish species were infected with FZTs. Prevalence was 56.8% in river fish, 46.7% in pond fish, and 29.3% in reservoir fish. The prevalence of Clonorchis sinensis metacercariae was 10.6% in reservoir fish, and 2.6% in river fish; fish from ponds were uninfected. The infection intensity of C. sinensis was 29.7 metacercariae/reservoir fish and 2.1 metacercariae/river fish. The prevalence of C. sinensis was highest (25.9%) in Toxabramis houdemeri. The intestinal flukes Haplorchis pumilio, Haplorchis taichui, Haplorchis yokogawai, Centrocestus formosanus and Procerovum varium were recovered from 29 fish species, with an overall prevalence of 46.8% and a mean infection intensity of 23.6 metacercariae. The results indicate that a C. sinensis sylvatic cycle involving wild fish species is important in the epidemiology of liver fluke and that consumption of raw or inadequately prepared wild-caught fish is a the major risk factor for human Clonorchiasis in this region. Both wild-caught and cultured fish present a high risk for infection with intestinal flukes. The habitat requirements of the snail host may be the important determining factor in favoring a sylvatic cycle of C. sinensis. This information is relevant to the development of effective prevention and control strategies, and emphasizes the priority that must be given to education of the local communities on the risks of consuming raw or improperly prepared fish dishes.

The role of rice fields, fish ponds and water canals for transmission of fish-borne zoonotic trematodes in aquaculture ponds in Nam Dinh Province, Vietnam.

BackgroundFish-borne zoonotic trematodes (FZT), such as Clonorchis sinensis, Opistorchis viverini (Opisthorchiidae) and intestinal trematodes of the family Heterophyidae, constitute a public health hazard in Vietnam and infections with these trematodes has been linked to consumption of raw or undercooked fish from aquaculture. The FZT transmission pathways, however, are more complicated than just the presence of intermediate snail hosts in aquaculture ponds as ponds may exchange water with surrounding habitats such as rice fields and irrigation canals and thereby these surrounding habitats may be a source of snails and cercariae and contribute to FZT infection in cultured fish.MethodsThis is a longitudinal descriptive study on selected farms (n = 30) in Nam Dinh Province which is endemic for FZT. At each farm, we sampled one pond, a small irrigation canal used to supply the pond with water, and a nearby rice field. At each of these three sites, we estimated the density of the FZT intermediate snail hosts and determined their trematode infection status. Comparative analysis was performed for the prevalence and density of FZT infections in fish and snails.ResultsSpecies of the Thiaridae, and most notably Melanoides tuberculata, the most important host species for FZT belonging to the Heterophyidae, were particularly abundant in ponds and small canals, i.e. M. tuberculata was found in 27 ponds and 13 small canals. Bithynia fuchsiana, a potential host for both Heterophyidae and Opisthorchiidae, was rarely found in fish ponds but common in rice fields. A total of 12 types of cercariae were found in the snails and pleurolophocercous cercariae, primarily FZT, constituted about 40 % of all cercarial infections. The fish species cultured were mainly carp species and Haplorchis pumilio was the dominating trematode species infecting fish. Clonorchis spp. were not recorded in any of the ponds. FZT transmission to fish was intense during the summer period (May-June to November) but less intense during the winter months (December-January) partly because cercarial emergence ceases due to the low temperature.ConclusionOur findings highlight the complexity of FZT transmission within aquaculture farm settings and suggest that efforts to control these infections must take a holistic approach using interventions against all stages of the transmission cycle.

Two-year intervention trial to control of fish-borne zoonotic trematodes in giant gourami (Osphronemus goramy) and striped catfish (Pangasianodon hypophthalmus) in nursery ponds in the Mekong Delta, Vietnam

Fish-borne zoonotic trematode parasites (FZT) pose a food safety and public health problem in Vietnam. The transmission cycle is complex as domestic animals, especially dogs, cats, fish-eating birds and pigs together with humans serve as reservoir hosts and contribute to FZT egg contamination of aquaculture ponds and the environment. This intervention trial was conducted to determine the effectiveness of various on-farm interventions, including reduction in FZT egg contamination through treatment of infected people and domestic animals, reduction in snail density through mud removal from aquaculture ponds prior to fish stocking, and various other measures in reducing FZT infection in juvenile striped catfish (Pangasianodon hypophthalmus) and giant gourami (Osphronemus goramy). Interventions were implemented on 5 farms for each fish species during production cycles in 2009 and 2010 while 5 similar farms for each species served as control. For both fish species, both prevalence and intensity of infection did not differ significantly between intervention and non-intervention farms prior to the interventions. The interventions significantly reduced both prevalence and intensity of FZT infection in the juvenile fish compared to control ponds. For giant gourami, odds of infection in intervention ponds was 0.13 (95% CL: 0.09–0.20; p<0.001) of that in non-intervention ponds after the 2009 trial and 0.07 (0.03–0.14; p<0.001) after the 2010 trial. For striped catfish, these figures were 0.17 (0.08–0.35; p<0.001) after the 2009 trial while after the 2010 trial all ponds with interventions were free from infection. Metacercariae intensity (no. of metacercariae/fish) in giant gourami from intervention ponds was 0.16 (0.11–0.23; p<0.001) of that in fish from non-intervention ponds after the 2009 trial and 0.07 (0.04–0.15; p<0.001) after the 2010 trial; for striped catfish these figures were 0.18 (0.09–0.36; p<0.001) and 0.00 (confidence limits not estimated), respectively. The aquaculture farm pond intervention approaches taken in this trial have the potential to reduce or eliminate FZT infections in fish and may be implemented across the entire region if adjusted to local conditions and fish species.

Infection Status of Zoonotic Trematode Metacercariae in Fishes from Vientiane Municipality and Champasak Province in Lao PDR.

The infection status of fishborne zoonotic trematode (FZT) metacercariae was investigated in fishes from 2 localities of Lao PDR. Total 157 freshwater fishes (17 species) were collected in local markets of Vientiane Municipality and Champasak Province in December 2010 and July 2011, and each fish was examined by the artificial digestion method. Total 6 species of FZT metacercariae, i.e., Opisthorchis viverrini, Haplorchis taichui, Haplorchis yokogawai, Haplorchis pumilio, Centrocestus formosanus, and Procerovum varium, were detected in fishes from Vientiane Municipality. The metacercariae of O. viverrini were detected in 50 (49.5%) out of 101 fishes (6 species), and their average number was 154 per fish infected. The remaining 5 species of heterophyid metacercariae were detected in 36.8%, 65.8%, 9.4%, 23.9%, and 5.1% fishes examined, and their average densities were 12, 1,038, 4, 15, and 13 per infected fish, respectively. In fishes from Champasak Province, 3 species of FZT metacercariae, i.e., O. viverrini, H. taichui, and H. yokogawai, were detected. Only 2 O. viverrini metacercariae were found in only 1 Barbonymus schwanefeldi. The metacercariae of H. taichui and H. yokogawai were detected in 60.0% and 50.0% of fishes examined, and their average densities were 47 and 28 per fish infected. By the present study, it has been confirmed that several species of FZT metacercariae are prevalent in fishes from Vientiane Municipality, with P. varium being a new member of FZT in Lao PDR. In comparison, FZT metacercariae are less prevalent in fishes from Champasak Province.

Effect of pond water depth on snail populations and fish-borne zoonotic trematode transmission in juvenile giant gourami (Osphronemus goramy) aquaculture nurseries

Infection with fish-borne zoonotic trematodes (FZT) is an important public health problem in many parts of Southeast Asia. People become infected with FZT when eating raw or undercooked fish that contain the infective stage (metacercariae) of FZT. The parasites require specific freshwater snails as first intermediate host and a variety of fish species, both wild caught and cultured, as second intermediate host. Aquaculture production has grown almost exponentially in SE Asia and in order to produce fish free from FZT metacercariae, it is important to mitigate factors promoting transmission to fish. Here we report results from a cross-sectional study to look at the association between pond depth and infection with FZT in giant gourami nursery ponds. Density of intermediate host snails was positively associated with pond depth (count ratio associated with a 1m increase in pond depth was 10.4 (95% C.L.: 1.61–67.1, p<0.5)) and this may partly explain the higher prevalence and intensity of FZT infection in juvenile fish. High fry stocking density (>200frym−3) was associated with lower host snail density (count ratio=0.15) than low stocking density (<100frym3). Ponds stocked with 100–200frym−3 had snail counts 0.76 (95% C.L.: 0.33–1.75, p n.s.) of those in ponds stocked with fry density of <100frym−3. Since density of intermediate snail hosts was associated with FZT transmission to fish, effort should be taken to reduce snail density prior to stocking the fry, but focus should also be on habitats surrounding ponds as transmission may occur through cercariae produced outside ponds and carried into ponds with water pumped into ponds.

Seasonal transmission of Opisthorchis viverrini sensu lato and a lecithodendriid trematode species in Bithynia siamensis goniomphalos snails in northeast Thailand.

Seasonal changes play roles in the transmission success of fish-borne zoonotic trematodes (FZT). This study examined the seasonal transmission patterns of Opisthorchis viverrini sensu lato (s.l.) and a virgulate cercaria (family Lecithodendriidae) in the snail intermediate host, Bithynia siamensis goniomphalos in northeast Thailand. Snail samples were collected monthly during the rainy, cool, and hot seasons during 2012-2013 to determine the prevalence and intensity of larval trematode infections. The prevalence of O. viverrini s.l. varied significantly with season, being 0.31%, 1.05%, and 0.37% in the rainy, cool, and hot seasons, respectively (P < 0.05). Similarly, the prevalence of virgulate cercariae was 3.11%, 6.80%, and 1.64% in the rainy, cool, and hot seasons, respectively (P < 0.05). The intensity of larval trematode infections also varied between seasons and peaked in the hot season (P < 0.05) in both species. The snails infected with O. viverrini s.l. were significantly smaller (P < 0.05) and those infected with virgulate cercariae were significantly larger (P < 0.05) than uninfected snails. Seasonal variation and the different sizes of B. s. goniomphalos parasitized by O. viverrini s.l. and virgulate trematodes indicate complex host-parasite interactions with important implications for the epidemiology of O. viverrini s.l.