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Background: Opisthorchis viverrini (OV) and soil-transmitted helminths (STH) are two of the most common helminths contributing to the Neglected Tropical Disease (NTDs) burden in the Lower Mekong Basin. Although mass drug administration is the cornerstone of control programs to reduce morbidity caused by these infections, this approach has limitations in preventing re-infections. Elimination requires additional measures such as reservoir host treatment, improved hygiene and health education to reinforce MDA's impact. This study aims to examine the impact of a scalable multi-component One Health Helminth Elimination program in the Lower Mekong Basin (HELM) that combines human praziquantel (PZQ) and albendazole (ALB) treatment with a program that includes the "Magic Glasses" and the "Lawa Model" interventions with health promotion at their core., Methods: This study will employ a cluster randomized controlled trial (cRCT) in 18 rural communities (with sub-district or villages as cluster units) across Cambodia, Laos and Thailand. The control arm will receive one round of PZQ/ALB treatment, while in the intervention arm, multi-component HELM program will be implemented, which includes PZQ/ALB treatment together with the Magic Glasses and Lawa Model interventions. OV and STH infections levels will be evaluated in individuals aged 5-75 years at baseline and will be repeated at follow-up (12 months after the HELM intervention), using modified formalin ethyl-acetate concentration technique and quantitative PCR. The primary outcome of the study will be cumulative incidence of human OV and STH infections. Outcomes between the study arms will be compared using generalized linear mixed models, accounting for clustering., Discussion: Evidence from this trial will quantify the impact of a multi-component One Health control strategy in interrupting Ov and STH infections in the Lower Mekong Basin., Trial Registration: Australian New Zealand Clinical Trials Registry (ANZCTR): ACTRN12622000353796. Prospectively registered 28 February 2022., Competing Interests: Declarations. Ethics approval and consent to participate: This study was submitted to and received ethical approval from the Australian National University Human Research Ethics Committee (Protocol number 2022/507), Cambodia Ministry of Health National Ethics Committee for Health Research (NECHR) (209NECHR), the Lao PDR Ministry NECHR (07NCEHR) and Khon Kaen University Ethics Committee for Human Research (Reference No. HE651147). This study is prospectively registered with the Australian New Zealand Clinical Trials Registry (ANZCTR): ACTRN12622000353796. Participation in the survey will be voluntary. Informed consent will be obtained from all participants ≥ 18 years old. Inclusion of participants < 18 years old required consent from a parent or guardian and assent from individuals 5 to 17 years old. The study objectives, intervention components, voluntary participation and the right to withdraw at any time will be verbally elaborated and outlined in the information sheet. Two copies will be signed by the study participants: an original copy and a duplicate. Researchers will keep the original copies of the consents signed by the study participants and leave the duplicate copies with them for their own records. This trial will also involve collection of stool specimens for storage. If the study participants will not allow the storage and use of remaining stool samples for future research, the remaining samples will be disposed immediately after the completion of all the procedures of this study. This study also obtained ethical approval from the Australian National University Animal Ethics Committee (Protocol Number A2022/27) and Institutional Animal Care and Use Committee of Khon Kaen University (Reference No. 660201.2.11/245 (39)). Consent for publication: Not applicable—there are no personal information or identifying images of study participants are presented here or will be presented in reports of the trial results. Competing interests: The authors declare that they have no competing interests., (© 2024. The Author(s).)

Background: Helminths are a major global health issue, impacting health, educational, and socioeconomic outcomes. Infections, often starting in childhood, are linked to anemia, malnutrition, cognitive deficit, and in chronic cases of Opisthorchis viverrini (OV), cholangiocarcinoma. The main control strategy for helminth infection is mass drug administration; however, this does not prevent reinfection. As such, prevention strategies are needed. The "Magic Glasses" is a school-based cartoon health education package that has demonstrated success in improving knowledge, attitudes, and practices (KAP) surrounding soil-transmitted helminths (STH) in China and the Philippines. This study is designed to assess the acceptability and impact of the 2 new versions of the Magic Glasses targeting STH and OV designed for the Lower Mekong audience in Cambodia, Lao People's Democratic Republic (PDR), and Thailand., Objective: The objective of this study is to evaluate the acceptability of the "Magic Glasses Lower Mekong" and "Magic Glasses Opisthorchiasis" education packages among schoolchildren in the Lower Mekong Basin, and the impact of these education packages on students' KAP surrounding STH and OV, respectively., Methods: Schoolchildren will be recruited into a cluster randomized controlled trial with intervention and control arms in rural schools in Cambodia, Lao PDR, and Thailand. Schoolchildren's initial acceptability of the intervention will be evaluated using an adapted questionnaire. Sustained acceptability will be assessed at 9-month follow-up through focus group discussions with students and interviews with teachers. Impact will be evaluated by KAP questionnaires on STH and OV. KAP questionnaires will be administered to children at baseline and at follow-up. Indirect impact on parents' KAP of OV and STH will be assessed through focus group discussions at follow-up., Results: The trial is in progress in Lao PDR and Thailand and is expected to commence in Cambodia in January 2024. The results of the study are expected to be available 18 months from the start of recruitment. We hypothesize that participants enrolled in the intervention arm of the study will have higher KAP scores for STH and OV, compared with the participants in the control arm at follow-up. We expect that students will have initial and sustained acceptability of these intervention packages., Conclusions: This trial will examine the acceptability of the "Magic Glasses Opisthorchiasis" and "Magic Glasses Lower Mekong" interventions and provide evidence on the effectiveness of the "Magic Glasses" on KAP related to OV and STH among schoolchildren in the Lower Mekong Basin. Study results will provide insight on acceptability and impact indicators and inform a scaling up protocol for the "Magic Glasses" education packages in Cambodia, Lao PDR, and Thailand., Trial Registration: Australian New Zealand Clinical Trials Registry ACTRN12623000271606; https://www.anzctr.org.au/Trial/Registration/TrialReview.aspx?id=385315&isReview=true., International Registered Report Identifier (irrid): DERR1-10.2196/55290., (©Suji Y O'Connor, Mary Lorraine Mationg, Matthew J Kelly, Gail M Williams, Archie CA Clements, Banchob Sripa, Somphou Sayasone, Virak Khieu, Kinley Wangdi, Donald E Stewart, Sirikachorn Tangkawattana, Apiporn T Suwannatrai, Vanthanom Savathdy, Visal Khieu, Peter Odermatt, Catherine A Gordon, Sangduan Wannachart, Donald P McManus, Darren J Gray. Originally published in JMIR Research Protocols (https://www.researchprotocols.org), 16.09.2024.)

Background: Despite the national campaigns in 1997 to control Opisthorchis viverrini (OV), a significant risk factor for cholangiocarcinoma (CCA) was accounted for in the northeast of Thailand, which is also considered the highest-risk area for CCA as per the latest national-wide survey in 2019. Social innovations were employed to address OV and CCA issues in this area, but they have yet to be thoroughly investigated. Investigating social innovations in areas where issues can already be solved is a particular approach for expanding on a broad basis., Objective: This study aimed to search for social innovations used to handle OV and CCA in northeast of Thailand., Methods: Researchers coordinated with the Office of Disease Prevention and Control Region-10 (1 of 12 regional offices under the Department of Disease Control, Ministry of Public Health) to specify the model areas for taking OV and CCA. In addition, researchers cooperated with local coordination units for data collection. The data were coded and classified for content analysis, and triangulation was performed for comparative analysis. Finally, a meeting was conducted to summarize the social innovations discovered in each study site area., Results: Our findings have been presented according to the epidemiologic triad: agent (risk factors), host (individuals/communities), and environment. The researchers and experts addressed and proposed social innovations for dealing with OV and CCA, such as the "3 Health" (Health Behaviours, Health Hygiene, and Environmental Health), in order to achieve a CCA-free society., Conclusions: This is the first study to highlight the significance of social innovations in managing OV and CCA. The findings will be utilized to further create strategies for addressing these public health issues in this and other regions in a sustainable and successful manner.

Opisthorchis viverrini is the etiological agent of the disease opisthorchiasis and related cholangiocarcinoma (CCA). It infects fish-eating mammals and more than 10 million people in Southeast Asia suffered from opisthorchiasis with a high fatality rate. The only effective drug against this parasite is Praziquantel, which has significant side effects. Due to the lack of appropriate treatment options and the high death rate, there is a dire need to develop novel therapies against this pathogen. In this study, we designed a multi-epitope chimeric vaccine design against O. viverrini by using immunoinformatics approaches. Non-allergenic and immunogenic MHC-1, MHC-2, and B cell epitopes of three candidate proteins thioredoxin peroxidase ( Ov-TPx-1 ), cathepsin F1 ( Ov-CF-1 ) and calreticulin ( Ov-CALR ) of O. viverrini , were predicted to construct a potent multiepitope vaccine. The coverage of the HLA-alleles of these selected epitopes was determined globally. Four vaccine constructs made by different adjuvants and linkers were evaluated in the context of their physicochemical properties, antigenicity, and allergenicity. Protein-protein docking and MD simulation found that vaccines 3 was more stable and had a higher binding affinity for TLR2 and TLR4 immune receptors. In-silico restriction cloning of vaccine model led to the formation of plasmid constructs for expression in a suitable host. Finally, the immune simulation showed strong immunological reactions to the engineered vaccine. These findings suggest that the final vaccine construct has the potential to be validated by in vivo and in vitro experiments to confirm its efficacy against the CCA causing O. viverrini ., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Shah, Sitara, Sarfraz, Shehroz, Wara, Perveen, Ullah, Zaman, Nishan, Ahmed, Ullah, Ali and Ojha.)

Opisthorchis viverrini infection is a pressing health issue in rural Southeast Asia and is associated with the risk of cholangiocarcinoma. Despite control efforts, high infection rates persist, including evidence of reinfection post-treatment. This study aimed to address this public health concern through an integrated One-Health approach in endemic areas in rural Thailand over a 3-year period. The study included data from 3600 participants from Udon Thani Province, Thailand, during the years 2020 to 2022 and involved integrated epidemiological data collection and risk factor analysis to understand the impact of various interventions on disease transmission in the community. The efficacy of interventions was assessed by monitoring the incidence of O. viverrini reinfection in 2021 and 2022. In 2020, 218 cases of O. viverrini infection (6.0%) were identified. Significant risk factors included proximity to water bodies and consumption of raw fish. Variables contributing to infection risk among participants (P < 0.001) were education level, engagement in traditional ceremonies, poor sanitation, absence of ducks in nearby water bodies, self-medication for parasitic conditions, and multiple infections within a household. Dogs, cats, and cyprinoid fish showed prevalence rates of 5.4%, 6.3%, and 11.5%, respectively. Geographic analysis revealed clusters of infected households around water bodies. Interventions, including in-depth interviews, focus-group discussions, health education, anthelminthic treatment, and biological control using local free-range ducks, were implemented, resulting in no human reinfections in the second year and a minimal 0.3% prevalence rate in the third year. This study offers valuable insights into the dynamic changes in infection prevalence, making a significant contribution to effective disease control and community health promotion. This integrated One-Health approach proved to be an effective strategy for the prevention and control of opisthorchiasis., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Opisthorchis viverrini is a parasitic liver fluke contracted by consumption of raw fish, which affects over 10 million people in Southeast Asia despite sustained control efforts. Chronic infections are a risk factor for the often fatal bile duct cancer, cholangiocarcinoma. Previous modeling predicted rapid elimination of O. viverrini following yearly mass drug administration (MDA) campaigns. However, field data collected in affected populations shows persistence of infection, including heavy worm burden, after many years of repeated interventions. A plausible explanation for this observation is systematic adherence of individuals in health campaigns, such as MDA and education, with some individuals consistently missing treatment. We developed an agent-based model of O. viverrini which allows us to introduce various heterogeneities including systematic adherence to MDA and education campaigns at the individual level. We validate the agent-based model by comparing it to a previously published population-based model. We estimate the degree of systematic adherence to MDA and education campaigns indirectly, using epidemiological data collected in Lao PDR before and after 5 years of repeated MDA, education and sanitation improvement campaigns. We predict the impact of interventions deployed singly and in combination, with and without the estimated systematic adherence. We show how systematic adherence can substantially increase the time required to achieve reductions in worm burden. However, we predict that yearly MDA campaigns alone can result in a strong reduction of moderate and heavy worm burden, even under systematic adherence. We predict latrines and education campaigns to be particularly important for the reduction in overall prevalence, and therefore, ultimately, elimination. Our findings show how systematic adherence can explain the observed persistence of worm burden; while emphasizing the benefit of interventions for the entire population, even under systematic adherence. At the same time, the results highlight the substantial opportunity to further reduce worm burden if patterns of systematic adherence can be overcome., Competing Interests: The authors have declared that no competing interests exist., (Copyright: © 2024 Kamber et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Opisthorchis viverrini infection is endemic in the lower Mekong subregion. The liver is an organ that worms are drawn to and cause damage. However, the immune-related susceptibility in the liver is poorly understood. In this study, we investigated T helper (Th) cell responses in the liver of BALB/c mice and golden Syrian hamsters during 2-28 days post-infection (DPI). We found that Th cell responses were distinct between mice and hamsters in terms of dynamics and polarization. Mice exhibited the early induction of Th1, Th2, Th17, and regulatory T (Treg) cells responses after the presence of O. viverrini worms at 2 DPI. In hamsters, the late induction of Th1/Th17, downregulation of Th2/Treg responses and early elevation of suppressive cytokine interleukin (IL)-10 were found together with swift reduction of Th cell numbers. Interestingly, expressions of IL-4 (Th2 functional cytokine) and Foxp3 (Treg lineage) were completely different between mice and hamsters which elevated in mice but suppressed in hamsters. These results suggest that early induction and well-regulation are related to host resistance. In contrast, late induction of Th cell response might allow immature worms to develop in the host. Our findings provide a greater understanding in Th cell response-related susceptibility in O. viverrini infection which would be targeting immunity for the development of immune-based intervention such as vaccine., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Cholangiocarcinoma (CCA), caused mainly by Opisthorchis viverrini (OV) infection, is a public health issue. Health literacy can play a significant role in preventing OV and CCA and adopting preventive behaviors. Therefore, this study aimed to evaluate, summarize, and synthesize the current evidence on health literacy programs for preventing OV and CCA.A systematic literature search, with Thai and English languages, was performed using electronic databases through PubMed, Google Scholar, ThaiJo, ThaiLis, and Embase to identify studies examining health literacy programs to prevent OV and CCA. We followed PRISMA 2020 guidelines. In addition, we used the RevMan software to perform a meta-analysis to analyze effect sizes using a fixed-effects model and measures of heterogeneity using Cochran's Q and I 2 . This meta-analysis included seven studies that met the criteria. The results showed that the people who received a program had an increased health literacy overall and in each aspect with a statistically significant (p < 0.001). So, health literacy programs can assist people in understanding their health and gaining access to health information and services. Additionally, the effect of programs (communication abilities, self-management, media and information literacy, and decision-making in practice) can help prevent OV and CCA. As a result, multi-disciplinary healthcare teams are crucial to developing preventive programs to prevent OV and CCA. Further studies need to be done and applied to these programs to modify behavior to avoid other diseases., (© 2023. The Author(s) under exclusive licence to American Association for Cancer Education.)

It is known that Opisthorchis viverrini (OV) is the most significant risk factor for the development of cholangiocarcinoma (CCA); hence, it is also known as carcinogenic parasite. Effective control and elimination of OV infection should significantly reduce O. viverrini-related CCA. This chapter includes details of the three recently developed innovative tools, namely the Isan cohort database software, an OV-RDT for screening of O. viverrini, and an ultrasound telecommunication system. Past and current control programs, i.e., education, medication, and sanitation were discussed and stressed the need for a comprehensive control program which encompasses primary, secondary, and tertiary patient care programs for confirmation and management of suspected CCA cases. The approach of mathematical modeling for control of OV and CCA was also briefly described. Additionally, we highlighted the current progress toward control of OV and CCA in Thailand and potential for expansion into nearby countries in Southeast Asia., (© 2023. The Author(s), under exclusive license to Springer Nature Switzerland AG.)

Host mucins have crucial physical roles in preventing the parasitic establishment and maturation, and also in expelling the invading parasites. However, some parasites utilize mucinase enzymes to facilitate the infection. Recently, we have identified a mucinase enzyme of the liver fluke Opisthorchis viverrini , Ov -M60-like-1, which exhibits metallopeptidase activity against bovine submaxillary mucin substrate. Here, we aimed to study the localization of this enzyme in O. viverrini and the bile duct of hamsters using immunohistochemistry and functional analysis by mucin digestion in hamsters and mice tissues. The results showed that Ov -M60-like-1 was detected strongly in the tegument, tegumental cells, vitelline glands and mature eggs with miracidium. Expression in the gut, ovary and testis of the parasite was moderate while parenchyma showed slight colour intensity. In addition, the mucinase was also detected in the host biliary epithelial cells and goblet cells surrounding the worm. The mucinase assay revealed that the Ov -M60-like-1 could digest neutral mucin in the parenchyma, testis and seminal receptacle, but not the mucin in the tegument, tegumental cells and vitelline glands of the worm. The enzyme can also digest mucin in the cholangiocytes and modified the mixture type in the bile duct goblet cells of the infected hamsters, a susceptible host. In contrast, the enzyme was unable to digest neutral, acid and mixture mucin in the bile duct of the mice, a non-susceptible host. These findings indicate that Ov -M60-like-1 may have functions in both housekeeping tasks and host–parasite interactions, especially in modification of host susceptibility.

Fasciola spp., Opisthorchis spp. and Clonorchis sinensis are common liver flukes that can cause a variety of diseases, mainly cholangiocarcinoma induced by clonorchiasis and liver damage and associated pathology induced by fascioliasis. Because these trematodes are parasites of humans and domestic animals, they have greatly affected the economy of agricultural industries and public health worldwide. Due to the emergence of drug resistance and the living habits of flukes, among other reasons, a possibility of reinfection remains even when antiparasitic drugs are used. Therefore, developing a safe, efficient and cost-effective vaccine against trematodes is an important goal. Here, we briefly describe the progress in the development of vaccines against liver flukes. Related innovations may provide effective protection against these helminths and the diseases that they cause., (Copyright © 2021 The Authors. Published by Elsevier Inc. All rights reserved.)

Transmission of the carcinogenic liver fluke Opisthorchis viverrini is ongoing across Southeast Asia. Endemic countries within the region are in different stages of achieving control. However, evidence on which interventions are the most effective for reducing parasite transmission, and the resulting liver cancer, is currently lacking. Quantitative modelling can be used to evaluate different control measures against O. viverrini and assist the design of clinical trials. In this article we evaluate the epidemiological parameters that underpin models of O. viverrini and the data necessary for their estimation, with the aim of developing evidence-based strategies for parasite control at a national or regional level., Competing Interests: Declaration of Interests No interests are declared., (Copyright © 2021 The Authors. Published by Elsevier Ltd.. All rights reserved.)

Background: The human liver fluke Opisthorchis viverrini is a food-borne trematode that causes hepatobiliary disease in humans throughout Southeast Asia. People become infected by consuming raw or undercooked fish containing metacercariae. Development of a vaccine to prevent or minimize pathology would decrease the risk of severe morbidity, including the development of bile duct cancer., Methods: We produced an oral vaccine based on recombinant Bacillus subtilis spores expressing the large extracellular loop (LEL) of O. viverrini tetraspanin-2 (Ov-TSP-2), a protein that is abundant on the surface of O. viverrini secreted extracellular vesicles (EVs). Recombinant spores expressing Ov-TSP-2-LEL were orally administered to hamsters prior to challenge infection with O. viverrini metacercariae., Results: Vaccinated hamsters generated serum IgG as well as bile IgG and IgA responses to Ov-TSP-2-LEL, and serum IgG from vaccinated hamsters blocked the uptake of fluke EVs by a human bile duct epithelial cell line. Vaccinated hamsters had 56% reductions in both adult flukes and fecal eggs compared to the control group., Conclusions: These findings indicate that oral vaccination of hamsters with recombinant B. subtilis spores expressing Ov-TSP-2-LEL is efficacious at reducing infection intensity and could form the basis of a vaccine for control of carcinogenic liver fluke infection in humans., (© The Author(s) 2020. Published by Oxford University Press for the Infectious Diseases Society of America. All rights reserved. For permissions, e-mail: journals.permissions@oup.com.)

The freshwater snail Bithynia siamensis goniomphalos serves as the first intermediate host of liver fluke Opisthorchis viverrini, a foodborne parasite, of which human infection has persisted in Southeast Asia for decades. The snail Filopaludina martensi martensi has been proposed as a biological control agent against B.s. goniomphalos, but knowledge on the snail ecology and population dynamics between the two species remains rudimentary. This study investigated selected abiotic and biotic factors influencing the distribution and abundance of B.s. goniomphalos and F.m. martensi. Water quality, soils, and snails were collected from 34 localities in Northeast Thailand. Soil properties and snail soft tissue elemental concentrations were analyzed. Experiments were performed to examine interspecific competition. Statistical analysis was conducted to explore the associations between water and soil properties and soft tissue elemental concentrations. The results showed that B.s. goniomphalos had the highest mean dominance in streams and red-yellow podzolic soils, while F.m. martensi snails preferred ponds and latosol soils. Negative correlation in species abundances was found between the two species. Interspecific competition was detected, with B.s. goniomphalos growth rates hampered by the presence of F.m. martensi. Despite the possibility of using F.m. martensi to control B.s. goniomphalos, B.s. goniomphalos exhibited a greater adaptability to different water and soil properties, suggesting that the species could colonize a wide range of environmental conditions. This study provides further insights into the ecology of the two snail species, underscoring the importance of considering abiotic factors when assessing the possible biological control agent to control O. viverrini transmission.

To clarify the reinfection profile associated with risk factors of opisthorchiasis, we conducted an epidemiological study on the chemotherapeutic effects on reinfection with O. viverrini in the endemic areas of Northeastern Thailand for 3 years. A total of 3,674 fecal samples were collected from participants in villages of 5 provinces. They were examined microscopically using a modified technique of formalin ethyl-acetate concentration. Egg-positive residents were reexamined year (2018) by year (2019) after treatment with a single dose (40 mg/kg) of praziquantel. Health education was provided to the participants yearly. The egg-positive rate of O. viverrini was 14.3%, and was highest (22.2%) in the 20-30 year-old group in 2017. The egg positive rate was 15.3% in dogs and 11.4% cats. Human reinfection rate was 15.5% and 6.3% in next 2 years, and was highest (23.2%) among the fishermen. Relative risk factors of reinfection were significantly higher for males, over 40-year-old age, or working as fishermen or farmers, and eating uncooked fish within the preceding year. A significant difference resulting from a health education program was observed in the third year. Therefore, health education and sustainable surveillance for opisthorchiasis should be maintained to decrease the risk of reinfection.

Liver fluke infection caused by Opisthorchis viverrini is recognized as a potential risk factor for cholangiocarcinoma (CCA). The National Strategic Plan to Control Liver Fluke Infection and Cholangiocarcinoma has implemented microscopic-based stool examination screening. However, eggs of O. viverrini and minute intestinal flukes (MIFs) are nearly morphologically similar and could result in inaccurate O. viverrini diagnosis. Stool specimens were collected from eight districts of Chiang Mai Province in northern Thailand. Opisthorchis -like eggs were identified with the Kato-Katz technique and differentiated for O. viverrini and MIFs using molecular study by PCR and PCR-restriction fragment length polymorphism targeting the internal transcribed spacer 2 (ITS2) gene. Prevalence of Opisthorchis -like eggs was 5.9% from a total of 9,570 specimens. From PCR assays, all liver flukes were O. viverrini and all MIFs were Haplorchis taichui. The distribution of species was H. taichui (38.2%), O. viverrini (10.5%), coinfection of H. taichui and O. viverrini (37.2%), and 14.1% were negative from PCR. Totally, H. taichui was found in 75.4% of infections from Opisthorchis -like specimens. ITS2 nucleotide sequencing analysis showed a single variant of O. viverrini with no variation and two variants of H. taichui . This study first revealed the genetic background of Opisthorchis- like eggs in northern Thailand. Minute intestinal flukes are occasionally misdiagnosed as O. viverrini leading to misinterpretation and overestimation of the burden of O. viverrini infection. Molecular diagnosis such as PCR could effectively discriminate species of Opisthorchis -like eggs and help shape the robustness of epidemiological data to control liver fluke infection and raise awareness of other risk factors for CCA.

Opisthorchiasis is a serious public health problem in East Asia and Europe. The pathology involves hepatobiliary abnormalities such as cholangitis, choledocholithiasis and tissue fibrosis that can develop into cholangiocarcinoma. Prevention of infection is difficult as multiple social and behavioral factors are involved, thus, progress on a prophylactic vaccine against opisthorchiasis is urgently needed. Opisthorchis viverrini tetraspanin-2 (Ov-TSP-2) was previously described as a potential vaccine candidate conferring partial protection against O. viverrini infections in hamsters. In this study, we generated a recombinant chimeric form of the large extracellular loop of Ov-TSP-2 and O. viverrini leucine aminopeptidase, designated rOv-TSP-2-LAP. Hamsters were vaccinated with 100 and 200 µg of rOv-TSP-2-LAP formulated with alum-CpG adjuvant via intraperitoneal injection and evaluated the level of protection against O. viverrini infection. Our results demonstrated that the number of worms recovered from hamsters vaccinated with either 100 or 200 µg of rOv-TSP-2-LAP were significantly reduced by 27% compared to the adjuvant control group. Furthermore, the average length of worms recovered from animals vaccinated with 200 μg of rOv-TSP-2-LAP was significantly shorter than those from the control adjuvant group. Immunized hamsters showed significantly increased serum levels of anti-rOv-TSP-2 IgG and IgG1 compared to adjuvant control group, suggesting that rOv-TSP-2-LAP vaccination induces a mixed Th1/Th2 immune response in hamsters. Therefore, the development of a suitable vaccine against opisthorchiasis requires further work involving new vaccine technologies to improve immunogenicity and protective efficacy., Competing Interests: Declaration of Competing Interest The authors confirm that they have no conflicts of interest in relation to this work., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Infection with the liver fluke Opisthorchis viverrini is the major causative factor inducing cholangiocarcinoma in the Mekong region of Southeast Asia. Northeast Thailand has the highest incidence of this cancer worldwide leading to about 20,000 deaths every year. Infection with the liver fluke comes from eating raw or undercooked fish, a tradition in this area that can potentially be countered by education programs at school level. Here we develop a school-based health education model, based on protection motivation theory (PMT), including module design, learning materials, student activities, and capacity building amongst teachers. This education program was applied and tested in primary school to pupils (9-13 years) in Khon Kaen province, northeast Thailand. Using a randomized control trial, four schools served as intervention groups ( n = 118 pupils) and another four acted as controls ( n = 113 pupils). Based on PMT constructs, we found that the pupils in the intervention group had significantly greater knowledge and perceived the severity, vulnerability, response efficacy, and self-efficacy parameters concerning the dangers of eating raw fish and of developing cholangiocarcinoma than those in the control schools ( p < 0.05). All of the PMT constructs measured were significantly intercorrelated with each other ( p < 0.001). At the same time, some background knowledge, from community-based education programs, was present in the control schools. The result from this initial study suggests that PMT can be used to predict protective attitude as well as behavior changes in evaluating the consequence of school health intervention programs.

Opisthorchiasis affects millions of people in Southeast Asia and has been strongly associated with bile duct cancer. Current strategic control approaches such as chemotherapy and health education are not sustainable, and a prophylactic vaccine would be a major advance in the prevention of the disease. Tetraspanins are transmembrane proteins previously described as potential vaccine candidates for other helminth infections and are also found in the membranes of the tegument and extracellular vesicles of O. viverrini. Here, we investigated the potential of a recombinant protein encoding for the large extracellular loop of O. viverrini tetraspanin-2 (rOv-LEL-TSP-2) in a hamster vaccination model. Hamsters were vaccinated with 50 and 100 μg of rOv-LEL-TSP-2 produced from Pichia pastoris yeast combined with alum CpG adjuvant via the intraperitoneal route. The number of worms recovered from hamsters vaccinated with rOv-LEL-TSP-2 was significantly reduced compared to adjuvant control groups. Fecal egg output was also significantly reduced in vaccinated animals, and the average length of worms recovered from vaccinated animals was significantly shorter than that of the control group. Vaccinated animals showed significantly increased levels of anti-rOv-TSP-2 IgG in the sera after three immunizations, as well as increased levels of several T helper type 1 cytokines in the spleen including IFN-γ and IL-6 but not the Th2/regulatory cytokines IL-4 or IL-10. These results suggest that rOv-TSP-2 could be a potential vaccine against opisthorchiasis and warrants further exploration.

Opisthorchis viverrini infection, opisthorchiasis, is a food-borne trematodiasis that is the main cause of cholangiocarcinoma, a bile duct cancer, in the Lower Mekong sub-region of Lao PDR, Cambodia, Vietnam, and Thailand. Despite extensive research on opisthorchiasis, the eradication of this disease has yet to be achieved. One of the major reasons for this failure is due to the multi-host life cycle of the parasite, which requires complex medical and public health interventions to eradicate. Another reason is due to a lack of knowledge of not only the interactions between the parasite and the human immune system, but also the interactions between the parasite and its various hosts during its complicated life cycle. Recent advances in various high-throughput omics technologies has allowed for the identification of key biomolecules crucial to the processes of parasitic transmission, and the identification of novel drug and/or vaccine targets. In this paper, omics studies dealing with O. viverrini host-parasite biology will be reviewed. In particular, there will be a focus on the strategies O. viverrini uses to trigger, evade, and manipulate the host's defense systems. Recently-identified biological molecules with potential as targets for interventions will also be reviewed. The results obtained from these omics approaches to analyzing O. viverrini and host interactions will be of great importance in the future when developing effective and sustainable medical and public health models for the prevention and control of opisthorchiasis and opisthorchiasis-induced CCA., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Background: The liver fluke Opisthorchis viverrini infects several million people in Southeast Asia. Adult flukes live in the bile ducts of humans, where they cause hepatobiliary pathology, including cholangiocarcinoma. Here, we investigated the potential of extracellular vesicles (EVs) secreted by the fluke and defined recombinant proteins derived from EVs to generate protective immunity in a hamster vaccination-challenge model., Methodology/principal Findings: EVs isolated from the excretory-secretory products of O. viverrini and two recombinant EV surface proteins encoding the large extracellular loops (LEL) of Ov-TSP-2 (rOv-TSP-2) and Ov-TSP-3 (rOv-TSP-3) were adjuvanted and used to vaccinate hamsters intraperitoneally followed by challenge infection with O. viverrini metacercariae. The number of adult flukes recovered from hamsters immunized with EVs, rOv-TSP-2, rOv-TSP-3 and rOv-TSP-2+rOv-TSP-3 were significantly reduced compared to control animals vaccinated with adjuvant alone. The number of eggs per gram feces was also significantly reduced in hamsters vaccinated with rOv-TSP-2 compared to controls, but no significant differences were found in the other groups. The average length of worms recovered from hamsters vaccinated with EVs, rOv-TSP-2 and rOv-TSP-3 was significantly shorter than that of worms recovered from the control group. Anti-EV IgG levels in serum and bile were significantly higher in hamsters vaccinated with EVs compared to control hamsters both pre- and post-challenge. In addition, levels of anti-rOv-TSP antibodies in the serum and bile were significantly higher than control hamsters both pre- and post-challenge. Finally, antibodies against rOv-TSP-2 and rOv-TSP-3 blocked uptake of EVs by human primary cholangiocyte in vitro, providing a plausible mechanism by which these vaccines exert partial efficacy and reduce the intensity of O. viverrini infection., Conclusion/significance: Liver fluke EVs and recombinant tetraspanins derived from the EV surface when administered to hamsters induce antibody responses that block EV uptake by target bile duct cells and exert partial efficacy and against O. viverrini challenge., Competing Interests: The authors have declared that no competing interests exist.

Clonorchis sinensis, Opisthorchis viverrini, and O. felineus are liver flukes of human and animal pathogens occurring across much of Europe and Asia. Nevertheless, they are often underestimated compared to other, better known neglected diseases in spite of the fact that many millions of people are infected and hundreds of millions are at risk. This is possibly because of the chronic nature of the infection and disease and that it takes several decades prior to a life-threatening pathology to develop. Several studies in the past decade have provided more information on the molecular biology of the liver flukes which clearly lead to better understanding of parasite biology, systematics, and population genetics. Clonorchiasis and opisthorchiasis are characterized by a chronic infection that induces hepatobiliary inflammation, especially periductal fibrosis, which can be detected by ultrasonography. These chronic inflammations eventually lead to cholangiocarcinoma (CCA), a usually fatal bile duct cancer that develops in some infected individuals. In Thailand alone, opisthorchiasis-associated CCA kills up to 20,000 people every year and is therefore of substantial public health importance. Its socioeconomic impacts on impoverished families and communities are considerable. To reduce hepatobiliary morbidity and CCA, the primary intervention measures focus on control and elimination of the liver fluke. Accurate diagnosis of liver fluke infections in both human and other mammalian, snail and fish intermediate hosts, are important for achieving these goals. While the short-term goal of liver fluke control can be achieved by praziquantel chemotherapy, a comprehensive health education package targeting school children is believed to be more beneficial for a long-term goal/solution. It is recommended that a transdisciplinary research or multisectoral control approach including one health and/or eco health intervention strategy should be applied to combat the liver flukes, and hence contribute to reduction of cholangiocarcinoma in endemic areas.

Opisthorchis viverrini infection is widely prevalent in Southeast Asia. In Cambodia information on this helminth infection is scare. Recent reports suggest that O. viverrini is an emerging public health problem. We aimed to synthesize all information in relation to the infection, epidemiology, and morbidity of O. viverrini in Cambodia; from published as well as thus far unpublished sources. First reports on O. viverrini date back to 1995. In 2006 an O. viverrini initiative was launched by the national helminth control program. Since then O. viverrini has been reported in all - except two - provinces. Villages with high prevalences (>20%) were found in provinces from Preah Vihear to Takeo. The infection has a highly focal distribution. In many villages no infections were detected. O. viverrini infection was also reported in cats, dogs and intermediate hosts. No report on morbidity associated with O. viverrini was found. The current evidence suggests that O. viverrini infection remains underreported in Cambodia. It is likely that the transmission will further increase in the future with potentially serious consequences for human health., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

Human infection with the Southeast Asian liver fluke Opisthorchis viverrini and liver fluke-associated cholangiocarcinoma cause significant disease burden in Southeast Asia. While there has been considerable work to understand liver fluke pathology and to reduce infection prevalence, there remains a limited understanding of the environmental determinants of parasite transmission dynamics to inform treatment and control programs. A particular setting where targeted control efforts have taken place is the Lawa Lake complex in northeast Thailand. Here, we describe the recent history of host infections, as well as the hydrologic characteristics of this floodplain ecosystem that influence the extent of snail habitat and fish mobility and the transport of human waste and parasite cercariae. Using mathematical modeling, we outline a framework for reconstructing environmental transmission of O. viverrini over the course of the Lawa Project control program from its inception in 2008 until 2016, using locally acquired but fragmentary longitudinal infection data for both humans and environmental hosts. The role of water flow in facilitating movement between snail, fish, human, and reservoir hosts is a particular focus with respect to its relevant scales and its impact on success of interventions. In this setting, we argue that an understanding of the key environmental drivers of disease transmission processes is central to the effectiveness of any environmental intervention., (Copyright © 2018 Elsevier B.V. All rights reserved.)

In Italy, food-borne outbreaks of opisthorchiasis have occurred since 2003, all of them originating from some lakes in Central Italy where the only European liver fluke, Opisthorchis felineus (Rivolta, 1884) is endemic. The aim of the present study is to investigate the factors that may facilitate the life cycle of O. felineus in the area of Bolsena Lake, the knowledge of the local population about opisthorchiasis, and preventative healthcare measures. We performed a descriptive observational study through the submission of questionnaires to three target populations: fishermen who fish exclusively in Bolsena Lake, restaurateurs whose businesses are close to the lake and general population living in the province of Viterbo. Results showed hazardous behaviours and eating habits in the three target populations, as well as a poor knowledge of opisthorchiasis and preventative healthcare measures.

We adapt a population-based model of Opisthorchis viverrini transmission dynamics to determine the effectiveness of three different interventions. The model includes the definitive hosts, humans; the reservoir hosts, dogs and cats; and the intermediate hosts, snails and fish. We consider the interventions: education campaigns to reduce the consumption of raw or undercooked fish, improved sanitation and treatment through mass drug administration. We fit model parameters to a data set from two islands in southern Lao PDR. We calculate the control reproduction number, simulate different scenarios and optimise the interventions with optimal control. We look at the potential of the interventions to eliminate transmission within 20 years. The model shows that education and improved sanitation need a very high coverage to fulfil the goal of elimination, whereas annual drug distribution at medium coverage is sufficient. The best solution is a combination of drug distribution at a medium level of coverage and as high as possible coverage of education and improved sanitation., (Copyright © 2018. Published by Elsevier Inc.)

Although any fish-eating mammals could be potential definitive hosts of Opisthorchis viverrini, only a few, especially cats and dogs, are actually known reservoir hosts for this parasite. Both animals usually get infected via consuming raw or undercooked contaminated fish, fish dishes or food remains from households. The infected animals sustain parasite egg spread via open environment defecation. Cats are the most important reservoir with higher prevalence rates of O. viverrini infection than dogs in endemic areas. Usually Opisthorchis-infected animals do not exhibit apparent clinical symptoms or specific abnormalities in laboratory examinations. Pathological findings in these animal reservoirs are basically similar to those seen in humans and experimental animals, namely periductal inflammation, biliary hyperplasia and periductal fibrosis. However, O. viverrini-associated cholangiocarcinoma has not yet been reported in the reservoir animals at present. Praziquantel is a treatment of choice not only for humans but also for animal reservoirs. Integrated control of opisthorchiasis in animal reservoirs is based on holistic approaches such as EcoHealth/One Health concepts. In fact integrated control of opisthorchiasis in humans in ecosystem has also proved successful, for example, the Lawa model for opisthorchiasis control in the endemic area of Khon Kaen, Thailand. Other feral and wild animals in endemic areas might also be potential reservoirs, and this requires more investigation. In addition, genetic diversity and evolution of the flukes might also influence zoonotic capability., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Opisthorchiasis is a neglected tropical disease, caused by infection with the fish-borne trematode Opisthorchis viverrini sensu lato that afflicts more than 10million people in Southeast Asia, including Thailand, Lao PDR, Vietnam and Cambodia. The disease is characterized by a chronic infection that induces hepatobiliary inflammation, especially periductal fibrosis, which can be detected by ultrasonography. This chronic inflammation eventually leads to cholangiocarcinoma (CCA), a usually fatal bile duct cancer that develops in approximately 1% of O. viverrini-infected individuals. In Thailand alone, CCA kills up to 20,000 people every year and is therefore of substantial public health importance. Its socioeconomic impacts on impoverished families and communities are considerable. To reduce O. viverrini-associated morbidity and CCA, the primary intervention measures focus on opisthorchiasis control and elimination. Accurate diagnoses of O. viverrini infection, in both mammalian, snail and fish intermediate hosts, are important for achieving these goals. Despite extensive efforts over several decades to find sensitive and specific diagnostics for opisthorchiasis, a simple and robust diagnostic method is still required. Here we review earlier and current developments in the search for new diagnostics for opisthorchiasis, with practical applications in the research laboratory, the clinic and the field. Of the methods currently available, the urine antigen assay shows considerable potential for the diagnosis and screening of opisthorchiasis. Nevertheless, these new assays require validation, determination of their cost-effectiveness when applied for mass screening in an endemic setting in support of policy decisions for national public health programs aimed at the control and elimination of opisthorchiasis., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The Lawa model is a successful integrative and sustainable means of controlling opisthorchiasis in Thailand. The model integrates the EcoHealth and One Health holistic approaches with systems thinking to target the interruption of Opisthorchis viverrini transmission. Using the six principles of EcoHealth and emphasizing the three domains of One Health (human-animal-ecosystem), the program targets each step of the parasite life cycle, thus maximizing the chances of interrupting the life cycle of the parasite. The main drivers of success are the village health volunteers and health-promoting hospitals, together with the support of the government and academia. The success of the model has led to continuous expansion, adoption nationally and internationally and application to the control of other neglected tropical diseases as well as noncommunicable diseases., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

The liver fluke Opisthorchis viverrini (Ov) is endemic in Southeast Asia where more than 10 million people are estimated to be infected. The infection is associated with several hepatobiliary diseases, including cholangiocarcinoma (CCA). Northeast Thailand is a hotspot for Ov transmission, and, despite extensive public health prevention campaigns led by the government, the prevalence of Ov infection is still high. High infection rates result from cultural and ecological complexities where wet-rice agrarian habitats, centuries-old raw-food culture, and the parasite's complex biology combine to create an ideal transmission arena. Here we review the state of our knowledge regarding the social-ecological determinants underlying Ov transmission. We also describe an integrative research rationale for liver fluke control better aligned with sustainable health development., (Copyright © 2017 Elsevier Ltd. All rights reserved.)

At the beginning of the new millennium, helminth infections continue to be prevalent, particularly among impoverished populations. This study attempts to create the first health informatics model of helminthiasis in Thailand. The authors investigate how a health informatics model could be used to predict the control and eradication in a national control campaign. Fish-borne helminthiasis caused by Opisthorchis viverrini remains a major public health problem in many parts of South-East Asia, including Thailand, Lao PDR, Vietnam and Cambodia. The epicentre of this disease is located in north-east Thailand, where high prevalence coexists with a high incidence of cholangiocarcinoma (CHCA). The current report was conducted to determine a mathematical model of surveillance for helminthiasis while also using a geographic information system. The fish-borne helminthiasis model or the predicted equation was Y1 = 3.028 + 0.020 (elevation) - 2.098 (clay). For soil-transmitted helminthiasis, the mathematical model or the predicted equation was Y2 = -1.559 + 0.005 (rainfall) + 0.004 (elevation) - 2.198 (clay). The Ministry of Public Health has concluded that mass treatment for helminthiasis in the Thai population, targeting high-risk individuals, may be a cost-effective way to allocate limited funds. This type of approach, as well as further study on the correlation of clinical symptoms with environmental and geographic information, may offer a novel strategy to the helminth crisis.

Opisthorchiasis caused by the carcinogenic liver fluke Opisthorchis viverrini is a major foodborne parasitic zoonotic disease in Thailand and neighboring Mekong countries. The infection is associated with several hepatobiliary diseases including cholangiocarcinoma (CCA). The rates of CCA in regions where the parasite is endemic are unprecedented. Extensive research on various aspects of opisthorchiasis has been conducted in recent decades. However, the current status of O. viverrini infection in the country is approaching 85% prevalence in certain endemic areas even after more than 30years of control programs in Thailand. The complex life cycle of the fluke, which involves several hosts/environments, makes it difficult to control by conventional methods. Therefore, a new control strategy using the EcoHealth/One Health approach named the "Lawa model" was introduced into the liver fluke endemic Lawa Lake region in Khon Kaen Province. This program has been underway for over ten years. The program includes treatment with anthelmintic drugs, novel intensive health education methods both in the communities and in schools, ecosystem monitoring and active community participation. We developed the "Liver fluke-free school program" as a part of the Lawa model with successful results. All key stakeholders were empowered to obtain competency in their control activities for the sustainability of the program in the community. Nowadays, the infection rate in the 12 villages surrounding the lake has declined to less than 10% from an average of 60% at the start. The Cyprinid fish species now show less than 1% prevalence of infection compared with a maximum of 70% during the baseline survey. No infected Bithynia snails in the lake were found during recent field studies. Of the lessons learned from the Lawa model, the importance of community participation is one practical and essential component. The key to the success of the project is multi-stakeholder participation with the active local Health Promoting Hospitals and the village health volunteers. The idea of the Lawa model is on the national agenda against liver fluke infection and CCA and is being scaled up to work in larger areas in Thailand. Internationally, the "Lawa model" is one of two programs that are showcased as successful control programs for helminths by the WHO Neglected Zoonotic Diseases (NZD4). Several media outlets have broadcast news reports about the program., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Opisthorchiasis caused by Opisthorchis viverrini is a major food-borne zoonosis in Greater Mekong sub-region. Even though campaigns discouraging the consumption of raw fish have been launched to public, the disease still remains highly endemic. The unsuccessful eradication of the disease is probably because of the persistence of the parasite in animal reservoir hosts, particularly felids. Praziquantel (PZQ) is the drug of choice for morbidity control of opisthorchiasis in humans and animals. However, there is no specific study on its dosage regimen for feline opisthorchiasis. Thus, the effective treatment dose of PZQ, as well as its adverse effects, was evaluated in O. viverrini infected cats. Twenty-eight infected male and female cats from the endemic area of Khon Kaen and Maha Sarakham Provinces, Thailand were enrolled in this study. Physical, hematological, blood chemical and urine examinations were analyzed, as indicators of health status, on the day before and 30days after treatment. Intensity of the infections was determined by the formalin-ethyl acetate sedimentation technique. Cats were equally allotted into the low infection group of 14 cats with egg count per gram of feces (EPG) <300 and the high infection group of 14 cats with EPG higher than 300. Cats in each group were equally divided into two subgroups of 7 cats; thus, there were two low infection subgroups (L1 and L2 subgroups) and two high infection subgroups (H1 and H2 subgroups). A single dose of 25mg/kg PZQ was orally administered to each cat in the L1 and H1 subgroups and a single oral dose of 40mg/kg PZQ was administered to the L2 and H2 subgroups. Complete clearance of O. viverrini eggs was found in all cats in the L1, L2 and H2 subgroups; thus, the cure rate (CR) and egg reduction rate (ERR) were 100%. However, partial clearance was observed in two cats with high EPG (1502 and 1518) in the H1 subgroup, which received 25mg/kg PZQ. Regards, CR and ERR for these two animals was 71.4 and 99.5%. No significant difference among the 4 subgroups was seen. Almost all hematological, blood chemical and urinalysis data were within normal ranges, except for the eosinophilia and an increase of alanine aminotransferase (ALT). Hookworm infection seen in all cats would cause eosinophilia. As for drug safety, there was no side effect observed in any cats. In conclusion, this study suggested that 40mg/kg PZQ is a highly effective and safe dosage for the treatment of feline reservoir hosts of human opisthorchiasis., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Background: Foodborne trematodiasis (FBT) is a significant global health problem, with the liver flukes Opisthorchis viverrini, O. felineus, and Clonorchis sinensis contributing to half of the global burden of FBT. North-eastern Thailand where O. viverrini is endemic and un-cooked fish dishes remain an integral part of the food culture has the highest reported incidence of opisthorchiasis, including associated cholangiocarcinoma. Both food sharing and eating practices are potentially important factors in FTB, suggesting an important role for the social ecology of disease transmission in these rural communities., Methods: Two rural Thai-Lao villages that were part of a 12-village project in Northeastern Thailand were selected for detailed investigation of O. viverrini infection risk associated with sharing of raw fish dishes among households. The project included screening individuals for infection and cholangiocarcinoma, a household questionnaire, and offering treatment options for positive individuals. Social network mapping was used to construct raw fish dish-sharing networks and create a proxy variable capturing variability in the degree of food sharing (DFS), measured as the number of different households with which each household shared fish dishes. Measures of associations between DFS, O. viverrini infection, the frequency of raw fish consumption, and the number of raw fish dishes consumed were generated using binary logistic regression, proportional odds ordinal logistic regression, and Poisson regression., Results: The results showed that the probability that a household has members infected with O. viverrini increased by ~7% (P < 0.01) for each additional household included in its network. Moreover, the frequency and number of types of raw fish dishes consumed increased significantly as the DFS increased. Of the two villages, that with the highest infection prevalence (48% versus 34.6%) had significantly higher social connectivity overall (P < 0.001)., Conclusions: Our findings suggest that the social ecology of human settlements may be key to understanding the transmission dynamics of some FBT. In the case of O. viverrini in Thai-Lao communities, for which food sharing is a traditional practice supporting social cohesion, food sharing network mapping should be incorporated into community-based interventions. These should encourage fish dish preparation methods that minimize infection risk by targeting households with high DFS values.

The International Agency for Research on Cancer (IARC) has classified two liver flukes as carcinogenic to humans (Group 1): Opisthorchis viverrini in 1994 and Clonorchis sinensis in 2009. This review is focused on O. viverrini, the most studied of these two trematodes, which infects nearly 10 million people in Southeast Asia. The life cycle involves two intermediate hosts living in fresh water: a snail of the genus Bithynia and a ciprinid fish. The definitive hosts (human, cat, dog) become infected by ingesting raw fish containing metacercariae, the infective stage of the parasite. Adult flukes attach to the epithelium of the bile ducts where they feed for as long as 10 to 30 years, resulting in chronic inflammation, epithelial hyperplasia, periductal fibrosis and formation of granuloma. For a long asymptomatic, the distomatosis is revealed by a chronic cholangitis when the parasite load becomes high. Complications can occur with time: gallstones, cholangitis, liver abscess, pancreatitis and, after a few decades, cholangiocarcinoma (CCA). The epidemiological correlation between the prevalence of O. viverrini infection and the incidence of CCA has been demonstrated in the northeast of Thailand. Specifically, the Khon Kaen province has the highest incidence rate in the world. The CCA can develop asymptomatically for a long time, especially in intrahepatic locations. It is often discovered at a late stage, unresectable. Its prognosis is dreadful with a survival rate less than 5% at 5 years. The phenomenon of carcinogenesis induced by O. viverrini is multifactorial. It has been specially studied using experimental infection on the Syrian golden hamster. Three intricated mechanisms are involved: (i) the direct damage caused by adult worms on the bile duct epithelium, (ii) the immunopathologic processes related to chronic inflammation (oxidative stress) and (iii) the mitogenic and anti-apoptotic effects of the proteins secreted by the parasite. Exogenous cofactors are also involved, such as nitrosamines in fish-based dishes undercoocked or fermented, very popular in these endemic regions. Despite the effectiveness of praziquantel to successfully cure this distomatose, opisthorchiasis persists endemic in areas where the incidence of CCA tends to progress. Mass deworming campaigns are ineffective due to the frequency of reinfection in the exposed population. Repeating alternatively cures and reinfections may promote carcinogenesis. The failure of prevention programs reflects the difficulty of changing the traditional habits of consuming raw or fermented fish. Pending a vaccine prophylaxis, control strategies are based on integrated measures involving the treatment of reservoir hosts, sanitation and efforts of continuing information and education to deter the consumption of uncooked fish and to improve the sanitation in rural areas.

Opisthorchiasis caused by Opisthorchis viverrini infection, is a serious public health problem in Lao PDR, Cambodia, Vietnam and Thailand. Cyprinid fish are required for the transmission of O. viverrini metacercariae to fish eating-mammal hosts including humans. The transmission of infective metacerariae is an important event, potentially resulting in the development of cholangiocarcinoma in cases of chronic opisthorchiasis. For a better understanding of the development of O. viverrini metacercariae in the second intermediate host, Barbonymus gonionotus (Cyprinidae), the immune response of infected B. gonionotus through fibrous tissue formation and superoxide dismutase (SOD) activity were investigated. The immunosuppressant (prednisolone) was administered to B. gonionotus infected with O. viverrini cercariae to study infectivity and development of the metacercariae. Additionally, the effects of prednisolone were studied by the investigation of fibrous tissue formation around the metacercariae, as well as SOD activity. In our study, prednisolone administered prior to infection in B. gonionotus had higher metacercarial recovery when compared to the non-administered control group. On the other hand, the administration of prednisolone after the O. viverrini infection allowed metacercariae to develop to the infective stage. Energy consumption from encystation to the infective stage required 443.9-463.7degree-days. The results show that immunosuppression decreases fibrous tissue and SOD activity in infected fish, and also affects the ability of O. viverrini metacercariae to develop to the infective stage. The immune response to metacercarial development of O. viverrini in cyprinid fish should be further studied to gain a greater understanding of this process, and hence, a better understanding of the mechanism(s) involved in metacercarial wall formation., (Copyright © 2016 Elsevier B.V. All rights reserved.)

Contamination of a popular fermented fish dish, pla-som, by Opisthorchis viverrini metacercariae (OVMC) is a possible cause of carcinogenic liver fluke infection in Thailand. Affected individuals are at risk of bile duct cancer, which is a major health problem for people in the Greater Mekong Subregion. In order to investigate concerns about food safety, we studied the effects of fermentation time and low temperature on the viability and infectivity of OVMC during the pla-som production process. Pla-som was prepared at room temperature for up to 1 week in duplicate experiments using cyprinid freshwater fish obtained from an O. viverrini-endemic area. OVMC were then isolated and identified under a stereomicroscope. Complete and viable OVMC were found on days 1-4 of fermentation, while their morphology was degenerated thereafter. After OVMC were fed to hamsters, the percentage of the worm recovery after 1 to 2 months of infection was 52%, 44.7%, 11.3% and 1% for days 1, 2, 3 and 4, respectively. In order to measure the effect of low temperature on OVMC, fish were kept in a refrigerator (4 °C) for up to five days and then subsequently fermented for three days. In fish stored in a refrigerator for 1 and 2 days, viable OVMC were clearly observed and were able to infect hamsters, a worm-recovery percentage of 3.3% and 12.7%, respectively. By contrast, in pla-som prepared from fish stored for 3 to 5 days, OVMC were degenerated and could not infect the host. In conclusion, pla-som fermentation for more than four days and refrigerating fish for three days before pla-som processing can prevent O. viverrini infection. This study may increase awareness of fermented-fish dish preparation to prevent liver fluke infection., (Copyright © 2015 Elsevier B.V. All rights reserved.)

A quasi-experimental study was conducted to develop a health education modification program based on self-efficacy and motivation regarding liver flukes and cholangiocarcinoma development in Keang Sanam Nang district, Nakhon Ratchasima province, Thailand. A total of 36 individuals were invited to participate in the program and were screened for population at risk of liver fluke infection and cholangiocarcinoma using SUT-OV-001 and SUT-CCA-001. Development of health education modification program regarding liver fluke and cholangiocarcinoma prevention included 3 steps: (1) preparation, (2) health education program, and (3) follow-up and evaluation. The study was implemented for 10 weeks. Pre-and-post-test knowledge was measured with questionnaires, Kuder-Richardson-20: KR-20 = 0.718,and Cronbach's Alpha Coefficient = 0.724 and 0.716 for perception and outcome expectation questionnaires. Paired and independent t-tests were applied for data analysis. The majority of the participants were female (55.6%), aged between ≤50 and 60 years old (36.1%), married (86.1%), education level of primary school (63.9%), agricultural occupation (80.6%), and income <4,000 Baht (44.4%). The results revealed that after the health education program, the experimental group had a mean score of knowledge, perception, and outcome expectation regarding liver fluke and cholangiocarcinoma prevention significantly higher than before participation and in the control group. In conclusion, this successful health education modification program for liver fluke and cholangiocarcinoma, therefore may useful for further work behavior modification in other epidemic areas.

The liver fluke Opisthorchis viverrini is a serious health problem in Thailand. Infection is associated with cholangiocarcinoma (CCA), endemic among human populations in northeast and north Thailand where raw fish containing fluke metacercariae are frequently consumed. Recently, Thailand public health authorities have been organized to reduce morbidity and mortality particularly in the northeast through O. viverrini and CCA screening projects. Health modfication is one of activities included in this campaign, but systemic guidelines of modifying and developing health behavior for liver flukes and CCA prevention in communities towards health belief and social support theory are still various and unclear. Here we review the guidelines for modifying and developing health behavior among populations in rural communities to strengthen understanding regarding perceived susceptibility, severity, benefits, and barriers to liver fluke and CCA prevention. This model may be useful for public health of cancers and related organizations to further health behavior change in endemic areas.

Opisthorchiasis is a major problem in Thailand particularly in northeast region which also has a high incidence of cholangiocarcinomas. Since health modification is needed, this quasi-experimental study aimed to assess the effectiveness of a health education program based on self-efficacy and social support in Tha Tum district, Surin province, Thailand. A total of 70 participants were purposive selected with a history of opisthorchiasis. Participants were divided into experimental and control groups, each with 35 subjects. The experimental group received a health education program composed of: (1) knowledge improvement, lectured with multimedia, demonstration, brochure, and handbook; (2) group discussion about their health beliefs, sharing their ideas and experience; and (3) social support from village health volunteers (VHV), heads of villages (HV), friends, and members of families, and public health officer (PHO). Follow-up was by PHO/VHV/HV, with provision of certificates and flasg for household that did not eat raw fish. Data were collected by predesigned questionnaires after implementation of the program for 3 months. Comparative data was analyzed by paired simple t-test and independent t-test. The results revealed that the experimental group had mean score of knowledge higher more than before the experiment (mean difference=3.1, t=3.915, 95%CI-3.3, -1.8 p-value=0.001), and the control group (mean difference=2.5, t=4.196, 95%CI=1.4, 3.6, p-value=0.001) with statistical significance. The mean scores of practice were higher than before the experiment (mean difference=4.6, t=4.331, 95%CI-5.3, -3.1, p-value=0.001), and control group (mean difference=4.4, t=6.142, 95%CI=4.2, 7.9, p-value=0.001) . The mean scores of perceived susceptibility and perceived severity of opisthorchiasis, al well as perceived benefits and perceived barriers to prevention of opisthorchiasis, were also higher than before the experiment and in the control group (p-value <0.001). In conclusion, this was a successful health education program for liver fluke avoidance. Therefore, it may useful for further behavior modification in the other epidemic areas.

This cross-sectional descriptive and qualitative study was aimed to study the people participation and their approaches toward the human carcinogenic liver fluke, Opisthorchis viverrini, prevention and control in Ban Chaun sub-district administrative organization (BCSAO) and Bamnet Narong sub-district municipality (BNSM), Bamnet Narong district, Chaiyaphum Province, Thailand between June 2013 and February 2014. Participants were purposive selected, included chiefs of sub-district administrative organizations, sub-district municipalities, sub-district health promotion hospitals (SHPHs), heads of village, and a further sample was selected with a multi- stage random sampling for public health volunteers, and villagers. The pre-designed questionnaire contained items for individualized status and the participatory steps of sharing ideas, decision-making, and planning, procession, evaluation, and mutual benefit, for the project O. viverrini prevention and control (POPC). In-depth interviews were used for collection of need approaches to POPC. With 375 participants who completed the questionnaire, it was found that people had a high level regarding to participate in the POPC, particularly in the process stage (X&#95; =3.78, S.D. = 0.56), but the lowest level was found in sharing ideas, decision making, and planning step (X&#95; =3.65, S.D. = 0.63). By comparison, participant status and organization did not significantly differ with people participation. In each step, Ban Chaun sub-district had a high level of participation in the step of sharing ideas, decision making, and planning toward POPC, more than Bamnet Narong sub-district municipality (t=2.20, p=0.028). Approaches for POPC in Ban Chaun sub-district and Bamnet Narong sub-district municipality included requirements for budget support, annual campaigns for liver fluke prevention and control, campaign promotion, risk group observation, home visiting, community rules regarding reducing raw fish consumption in their communities, and a professional public health officer for working in their communities, BCSAO, BNSM, and SHPH, for O. viverrini prevention and control. This study indicates that people realize that eradication of the liver fluke needs a continuous people participation for O. viverrini prevention and control in their communities.

This study aimed to modify behavior regarding liver fluke and cholangiocarcinoma prevention in Chumphuang district, Nakhon Ratchasima province, Thailand through integrated learning. A total of 180 participants were included through purposive selection of high-risk scores on verbal screening. Participants attended the health education program which applied the health belief model included family based, knowledge station based, academic merit based and community based learning. Data were collected using a questionnaire composed of 4 parts: 1) personal information, 2) knowledge, 3) perceived susceptibility, severity, benefits, and barriers, 4) practice regarding liver fluke and cholangiocarcinoma prevention. The result revealed that the majority were female (79.9%), age ≥60 years old (33.2%), primary school educational level (76.1%), and agricultural occupation (70.1%). The mean scores of knowledge, perception, and practice to liver fluke and cholangiocarcinoma prevention, before participated the integrative learning were low, moderate, and low, respectively. Meanwhile, the mean score of knowledge, perceived susceptibility, severity, benefits, and barriers, and practice regarding liver fluke and cholangiocarcinoma prevention, were higher with statistical significance after participation in the integrated learning. This finding indicates that health education programs may successfully modify health behavior in the rural communities. Therefore they may useful for further work behavior modification in other epidemic areas.

Opisthorchiasis is still a major health problem in rural communities of Thailand. Infection is associated with cholangiocarcinoma (CCA), which is found frequently in Thailand, particularly in the northeastern. Therefore, this study aimed to evaluate the effectiveness of health intervention in the population at risk for opisthorchiasis and CCA. A quasi-experimental study was conducted in Meuang Yang district, Nakhon Ratchasima province, northeastern Thailand, between June and October 2015. Participants were completed health intervention comprising 4 stations; 1, VDO clip of moving adult worm of liver fluke; 2, poster of life cycle of liver fluke; 3, microscopy with adult and egg liver fluke; and 4, brochure with the knowledge of liver fluke containing infection, signs, symptoms, related disease, diagnosis, treatment, prevention, and control. Pre-and-post-test questionnaires were utilized to collect data from all participants. Students paired t-tests were used to analyze differences between before and after participation in the health intervention. Knowledge (mean difference=-7.48, t=-51.241, 95% CI, -7.77, -7.19, p-value =0.001), attitude (mean difference=-9.07, t=-9.818, 95% CI=-10.9, -7.24, p-value=0.001), and practice (mean difference=-2.04, t=-2.688, 95% CI=-3.55, -0.53, p-value=0.008), changed between before and after time points with statistical significance. Community rules were concluded regarding: (1) cooked cyprinoid fish consumption; (2) stop under cooked cyprinoid fish by household cooker; (3) cooked food consumption; (4) hygienic defecation; (5) corrected knowledge campaign close to each household; (6) organizing a village food safety club; (7) and annual health check including stool examination featuring monitoring by village health volunteers and local public health officers. The results indicates that the present health intervention program was effective and easy to understand, with low cost and taking only a short time. Therefore, this program may useful for further work at community and provincial levels for liver fluke prevention and control.

Background: Opisthorchiasis, caused by Opisthorchis viverrini, is of considerable public health importance in Southeast Asia, particularly in Lao PDR and Thailand., Objective: This study aims to analyze the risk areas for liver flukes in Surin province, Thailand using Geographic Information Systems (GIS)., Material and Method: The 5 main factors (131 variables) were: (1) personal data (2) knowledge, attitude and practice, (3) health service unit, (4) environmental data, and (5) climate data. The relationship between these mainfactors and liver fluke infection was analyzed using multiple regression analysis. Potential surface analysis (PSA) with geographic information systems (GIS) was performed to create maps displaying areas at risk for liver fluke infection in Surin province., Results: The population density (148-169 pop/km2; X73), human attitude (< 50%; X111), and land use (wetland; X64), were statistically significant with liver fluke infection by a percentage of 88.60% (Adjusted R2 = 0.886). The heavy risk areas covered 221.39 km2 and including 8 districts; Si Narong, Sangkha, Phnom Dong Rak, Mueang Surin, Non Narai, Samrong Thap, Chumphon Buri and Rattanaburi., Conclusion: Present study identifies the increased risk areas for liver flukes in Surin Province and may be useful for future planning of prevention and control.

Human liver fluke, Opisthorchis viverrini, a food-borne trematode is a significant public health problem in Southeast Asia, particularly in Thailand. Despite a long history of control programmes in Thailand and a nationwide reduction, O. viverrini infection prevalence remains high in the northeastern provinces. Therefore, a new strategy for controlling the liver fluke infection using the EcoHealth/One Health approach was introduced into the Lawa Lake area in Khon Kaen province where the liver fluke is endemic. A programme has been carried using anthelminthic treatment, novel intensive health education methods both in the communities and in schools, ecosystem monitoring and active community participation. As a result, the infection rate in the more than 10 villages surrounding the lake has declined to approximate one third of the average of 50% as estimated by a baseline survey. Strikingly, the Cyprinoid fish species in the lake, which are the intermediate host, now showed less than 1% prevalence compared to a maximum of 70% at baseline. This liver fluke control programme, named "Lawa model," is now recognised nationally and internationally, and being expanding to other parts of Thailand and neighbouring Mekong countries. Challenges to O. viverrini disease control, and lessons learned in developing an integrative control programme using a community-based, ecosystem approach, and scaling-up regionally based on Lawa as a model are described., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Background: This study aimed to evaluate the impact of a community-based health education and communication programme on reducing liver fluke infections caused by the consumption of uncooked fish among people in a high-risk area of Thailand., Materials and Methods: The study was quasi-experimental in nature, with three-stages. Stage 1 involved a situational and capacity analysis of designated communities in Khon Kaen province. This was followed by the development of a model for community-based health education and communication to prevent liver fluke infections among high-risk people, and, lastly, implementation and evaluation of the model were performed. Data were collected using both qualitative and quantitative methods. In total, 390 people were surveyed, and quasi-experimental and comparison groups, each with 90 people, were assessed between May 2011 and April 2012. Analysis was using statistical OR, 95 % CI, the Willcoxon matched pairs signed ranks test, the chi-square test, and the Mann-Whitney U test., Results: The findings showed that most respondents had a high level of knowledge and understanding of liver fluke disease (89.5%, 95% CI:86.0-92.4), and positive attitudes toward the prevention of the disease (94.4%, 95% CI:91.6-96.4). However, with regard to changes in consumption of uncooked fish, most respondents were still in the pre-contemplation phase (55.1%, 95% CI:50.0-60.1), followed by the contemplation phase, 22.6%. Furthermore, four factors were found to be associated with the consumption of uncooked fish--the consumption of alcohol (OR 4.16, 95% CI:1.79-9.65), gender (OR 3.17 , 95% CI:1.53-6.54), smoking (OR 3.03, 95% CI:1.31-7.05), and age 40 years and above (OR 2.68, 95% CI:1.02-7.05). After nine months of the health education and communication programme using local media based on local wisdom, culture and persons, the results showed that, compared to the control group, members of the experimental group had a higher level of knowledge, a better attitude and lower levels of ill- advised consumption behaviour. Also, it was found that consumption of uncooked fish, by an assessment of the level of stage of change, was reduced. (p-value 0.002)., Conclusions: The health education and communication programme developed as part of the study was effective in changing the consumption of uncooked fish. Therefore, this approach should be promoted in other high-risk areas in Thailand in the future.

It is urgently necessary to be aware of the distribution and risk areas of liver fluke, Opisthorchis viverrini, for proper allocation of prevention and control measures. This study aimed to investigate the human behavior, and environmental factors influencing the distribution in Surin Province of Thailand, and to build a model using stepwise multiple regression analysis with a geographic information system (GIS) on environment and climate data. The relationship between the human behavior, attitudes (<50%; X111), environmental factors like population density (148-169 pop/km2; X73), and land use as wetland (X64), were correlated with the liver fluke disease distribution at 0.000, 0.034, and 0.006 levels, respectively. Multiple regression analysis, by equations OV=-0.599+0.005(population density (148-169 pop/km2); X73)+0.040 (human attitude (<50%); X111)+0.022 (land used (wetland; X64), was used to predict the distribution of liver fluke. OV is the patients of liver fluke infection, R Square=0.878, and, Adjust R Square=0.849. By GIS analysis, we found Si Narong, Sangkha, Phanom Dong Rak, Mueang Surin, Non Narai, Samrong Thap, Chumphon Buri, and Rattanaburi to have the highest distributions in Surin province. In conclusion, the combination of GIS and statistical analysis can help simulate the spatial distribution and risk areas of liver fluke, and thus may be an important tool for future planning of prevention and control measures.

Human liver fluke, Opisthorchis viverrini (Ov), is the major risk factor of cholangiocarcinoma (CCA) in northeastern Thailand. Our approach focuses on genetic progression and molecular changes in the carcinogenic pathway of liver fluke-associated CCA aimed at assessing patients at risk of CCA and using chemoprevention as the secondary cancer prevention to reduce the incidence of CCA. This review summarizes altered gene expressions, biomolecules and their modification, i.e. DNA adducts, oxidized proteins, oxysterols and fibrotic markers in hamster- and human-CCA. Potential risk biomarker(s) and chemopreventive agent(s) criteria and selection were based on results from experimental and epidemiological studies identifying hepatobiliary disease, including CCA. Laboratory results reveal that oxidative stress induced by Ov infection leads to bimolecular damage, tissue remodeling especially periductal fibrosis and alteration of gene expressions, which could be involved in all steps of CCA carcinogenesis. Some of these molecules are reported to change their levels in opisthorchiasis, periductal fibrosis diagnosed by ultrasonography and CCA. Chemoprevention in experimental CCA tumorigenesis is discussed. These multiple risk biomarkers could now be explored for screening including chemopreventive intervention of subjects living in endemic areas where the prevalence of opisthorchiasis remains high., (© 2014 Japanese Society of Hepato-Biliary-Pancreatic Surgery.)

Opisthorchis viverrini is an ongoing public health problem in Northeast Thailand. Despite continuous efforts for decades by healthcare organizations to overcome this problem, infection rates remain high. To enable related personnel to identify and address the various issues effectively, a cross-sectional study was performed to investigate prevalence and risk factors for opisthorchiasis. The target group was 3,916 Thai residents of Northeast Thailand who were 15 or over. Participants were recruited using the 30 clusters sampling technique. The data were gathered through questionnaires, focus group discussions, in-depth interviews, and stool examinations for parasite eggs (using the Modified Kato Katz method). The data were analyzed using descriptive and inference statistics; in order to ascertain the risk factors and test them using the odds ratio and multiple logistic regressions. The prevalence of opisthorchiasis was 22.7% (95%CI: 0.26 to 0.24). The province with the highest prevalence was Nakhorn Phanom (40.9%; female to male ratio =1:1.2). The age group with the highest prevalence was 40-49 year olds. All age groups had a prevalence >20%. Four of seven provinces had a prevalence >20%. The factors related to opisthorchiasis were (a) sex, (b) age (especially > 50), (c) proximity and duration living near a water body, and (d) eating raw and/or fermented fish. In order to reduce the prevalence of opisthorchiasis, the focus in populations living in upper Northeast Thailand should be changing their eating behaviors as appropriate to their tradition and context.

Opisthorchis viverrini infection causes inflammation and liver injury leading to periductal fibrosis. Little is known about the pathological alterations in bile canaliculi in opisthorchiasis. This study aimed to investigate bile canalicular alterations in O. viverrini-infected hamsters and to examine the chemopreventive effects of curcumin on such changes. Hamsters were infected with O. viverrini and one group of animals was fed with 1% dietary curcumin supplement. Animals were examined during the acute infection phase, days 21 and 30 post-infection (PI) and chronic infection phase (day 90 PI). Scanning electron microscopy revealed that in the infected group fed with a normal diet, bile canaliculi became slightly tortuous by 30 day PI and more tortuous at day 90 PI. Transmission electron microscopy showed a reduction in microvilli density of canaliculi starting at day 30 PI, with a marked loss of microvilli at day 90 PI. These ultrastructral changes were slightly seen at day 21 PI, which was similar to that found in infected animals fed with 1% curcumin-supplemented diet. Notably, curcumin treatment prevented the reduction of microvilli density, reduced the dilation of bile canaliculi, and decreased the tortuosity of the bile canaliculi relative to non-infected animals on a normal diet at days 30 and 90 PI. These results suggest that curcumin reduces alteration of bile canaliculi and may be a promising agent to prevent the onset of bile duct abnormalities induced by O. viverrini infection.