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Background Screening for opisthorchiasis, a parasitic worm infection affecting many millions of people in Southeast Asia, has traditionally relied on faecal egg examination such as the formalin-ethyl acetate concentration technique (FECT) and Kato-Katz method. Although the urinary enzyme-linked immunosorbent assay (ELISA) has been used more recently, we developed a urinary antigen-based rapid diagnostic test (RDT) to simplify diagnosis and as a point-of-care testing (POCT) and field applications for surveillance and control of opisthorchiasis. Methods A urinary Opisthorchis viverrini (OV)-RDT was developed using immunochromatographic methodology with a specific monoclonal antibody against OV. The diagnostic performance of the urinary OV-RDT was compared to that of quantitative faecal FECT and urinary antigen ELISA ( n  = 493). Cross-reactivities of urinary OV-RDT with other helminthiases coexisted with O. viverrini were determined ( n  = 96). A field trial in the application of urinary OV-RDT was compared with urinary antigen ELISA at baseline screening and assessment of drug treatment outcomes in opisthorchiasis ( n  = 1629). The McNemar chi-square, Kruskal–Wallis and Cohen’s kappa coefficient ( κ -value) tests were used for statistical analyses. Results Urinary OV-RDT had sensitivity of 94.2% and specificity of 93.2%, compared to faecal FECT. Urinary OV-RDT had high diagnostic agreement (Kappa = 0.842–0.874, P  < 0.001) and quantitative correlation with urinary antigen ELISA (Kruskal–Wallis tests = 316.2, P  < 0.0001) and faecal FECT (Kruskal–Wallis tests = 362.3, P  < 0.0001). The positive rates by OV-RDT, ELISA and FECT were 48.9%, 52.5% and 49.3%, respectively. Cross-reactions of urinary OV-RDT with other helminthiases were few (2%). Field trials of urinary OV-RDT yielded comparable prevalence of O. viverrini between urinary OV-RDT (53.2%) and urinary antigen ELISA (54.0%). OV screening showed high diagnostic agreement (kappa > 0.8, P  < 0.0001) between urinary OV-RDT and urinary antigen ELISA. The cure rates of opisthorchiasis at 1 month post-praziquantel treatment determined by urinary OV-RDT (86.6%) and urinary antigen ELISA (80.5%) were similar ( P  > 0.05). Conclusions The urinary OV-RDT test has high potential as a new tool for screening and evaluating treatment outcomes in opisthorchiasis. The ease of sample collection and simplicity of urinary OV-RDT may facilitate mass screening, control and elimination of opisthorchiasis, thereby contributing to a reduction in the disease burden in Southeast Asia. Graphical Abstract

Objectives: The current study was performed to determine the antioxidant, anti-inflammatory, and anti-cholangiocarcinoma (CCA) properties of Antidesma thwaitesianum, also known as “MAO,” whole plant extract on Opisthorchiasis in animal models and CCA cell lines. Materials and Methods: Ethanol was used to extract compounds from the whole ripe fruit. The phytochemical investigation of MAO extract was done to evaluate antioxidant activity, and high-performance liquid chromatography was used to identify the active compounds. The efficacy of MAO extract against OV was evaluated in vivo. The anti- CCA activity was evaluated using superoxide dismutase (SRB), cell cycle arrest, apoptosis, and western blot analyses. Results: MAO extract possessed flavonoid and phenolic contents, antioxidant activity, and an expressed cyanidin-3-O-glycosides content of 0.08 μg/mg extract. MAO extract demonstrated hepatoprotective effects through raised alanine transaminase and alkaline phosphatase levels, as well as an influence on oxidative stress via decreased MDA and increased glutathione, superoxide dismutase, and catalase levels. MAO extract significantly inhibited the migration of CCA cells in a concentration- and time-dependent manner, as well as triggered cell cycle arrest on G1 and activated apoptosis pathways via upregulation of C3, downregulation of cyclin-dependent kinase inhibitor p21, cyclin D, and cyclin-dependent kinases 2 expression. MAO extract inhibited inflammation, which in turn decreased fibrosis in hamsters. It also increased hepatoprotective activity. Conclusion: Our findings demonstrate the potential benefits of MAO extract in both in vitro and animal studies of hamster opisthorchiasis. However, more research should be done to ascertain the mechanisms of activity of MAO extracts and elute bioactive components in order to confirm their safety and examine their clinical applications.

This study demonstrates how a transdisciplinary learning approach provided new insights for explaining persistent Opisthorchis viverrini infection in northern Thailand, as well as elucidating problems of focusing solely on the parasite as a means of addressing high prevalence of cholangiocarcinoma. Researchers from diverse backgrounds collaborated to design an investigative homestay program for 72 Singaporean and Thai university students in five northeast Thai villages. The students explored how liver fluke infection and potential cholangiocarcinoma development are influenced by local landscape dynamics, aquatic ecology, livelihoods, food culture and health education. Qualitative fieldwork was guided daily by the researchers in a collaborative, co-learning process that led to viewing this health issue as a complex system, influenced by interlinked multidimensional factors. Our transdisciplinary experience has led us to believe that an incomplete understanding of these linkages may reduce the efficacy of interventions. Further, viewing liver fluke infection and cholangiocarcinoma as the same issue is inadvisable. Although O. viverrini infection is an established risk factor for the development of cholangiocarcinoma, multiple factors are known to influence the likelihood of acquiring either. Understanding the importance of the current livelihood transition, landscape modification and the resulting mismatch between local cultures and new socio-ecological settings on cholangiocarcinoma initiation and liver fluke transmission is of critical importance as it may help readjust our view of the respective role of O. viverrini and other socioeconomic risk factors in cholangiocarcinoma etiology and refine intervention strategies. As demonstrated in this study, transdisciplinary approaches have the potential to yield more nuanced perspectives to complex diseases than research that focuses on specific aspects of their epidemiology. They may therefore be valuable when designing effective solutions to context-sensitive diseases such as liver fluke infection and cholangiocarcinoma.

Opisthorchis viverrini infection is an important, human, parasitic disease along the middle and lower Mekong River of Southeast Asia. Despite vast efforts in epidemiological research, the wide geographical variation in O. viverrini prevalence remains unexplained. To investigate the potential influence of village location, prevalence data recorded during the year 2009 for 90 villages in northeast Thailand were analysed by a geographical information systems approach based on a digital elevation model for altitude determination. Land use compositions of village surroundings were derived from images produced by the Thematic Mapper of the Landsat satellite and landscape metrics were applied to quantify spatial patterns. Pearson's correlation coefficients were used to evaluate the associations between the environmental factors and O. viverrini prevalence. In spite of no statistical difference (P = 0.117), higher-ground villages generally had a lower level of opisthorchiasis than those situated lower down, while a significant negative correlation was detected between O. viverrini prevalence and the composition of farmland with low water content (P = 0.028), indicating the potential influence of agricultural lands with drought-tolerant crops. Strengthening this hypothesis, a significant positive association with O. viverrini prevalence was found for the combined land use classes of water and of farmland with high water content (P = 0.032), indicating that the dominance of a large land use patch capable of providing suitable habitats for the intermediate hosts might contribute to the disease. This study highlights the influence of landscapes on the epidemiology of O. viverrini infection and emphasises the need to incorporate land use planning into control strategies.

Opisthorchis viverrini infection is associated with human cholangiocarcinoma and northeast Thailand has the highest incidence of this disease in the world. Bithynia siamensis goniomphalos is the major freshwater snail intermediate host of O. viverrini in this area and an analysis based on geographical information systems was used to determine the effect of variation in soil surface salt on the density and distribution of this snail. A malacological survey was carried out in 56 water bodies in the Khorat basin, northeast Thailand at locations with various soil surface salt levels. Mollusk samples were collected from 10 ecologically representative water body sites with 10-20 sampling stations in each. The shoreline of clear, shallow water bodies was found to be the preferred B. s. goniomphalos habitat. The snails were exclusively found in water with salinity levels ranging between 0.05 and 22.11 parts per thousand (ppt), which supports the notion that B. s. goniomphalos prefers water with some saline content over pure, freshwater. The highest snail population densities were in rice fields, ponds, road-side ditches and canals within a water salinity range of 2.5-5.0 ppt. However, the presence of B. s. goniomphalos was negatively correlated with water salinity (P ≤0.05), both with regard to density and distribution. The areas with the highest density of B. s. goniomphalos were those with less than 1% soil surface salt (potential index = 0.314), while the lowest densities were found in areas exceeding 50% soil surface salt (potential index = 0.015).

[This corrects the article DOI: 10.3389/fimmu.2024.1281544.].

Background. Throughout Asia, there is an unprecedented link between cholangiocarcinoma and infection with the liver fluke Opisthorchis viverrini. Multiple processes, including chronic inflammation and secretion of parasite proteins into the biliary epithelium, drive infection toward cancer. Until now, the mechanism and effects of parasite protein entry into cholangiocytes was unknown. Methods. Various microscopy techniques were used to identify O. viverrini extracellular vesicles (EVs) and their internalization by human cholangiocytes. Using mass spectrometry we characterized the EV proteome and associated changes in cholangiocytes after EV uptake, and we detected EV proteins in bile of infected hamsters and humans. Cholangiocyte proliferation and interleukin 6 (IL-6) secretion was measured to assess the impact of EV internalization. Results. EVs were identified in fluke culture medium and bile specimens from infected hosts. EVs internalized by cholangiocytes drove cell proliferation and IL-6 secretion and induced changes in protein expression associated with endocytosis, wound repair, and cancer. Antibodies to an O. viverrini tetraspanin blocked EV uptake and IL-6 secretion by cholangiocytes. Conclusions. This is the first time that EVs from a multicellular pathogen have been identified in host tissues. Our findings imply a role for O. viverrini EVs in pathogenesis and highlight an approach to vaccine development for this infectious cancer.

(1) Background: Opisthorchis viverrini is a significant health problem in the Mekong subregion of Southeast Asia, causing aggressive cholangiocarcinoma. Current diagnostic procedures do not cover early diagnosis and low infection. Hence, an effective diagnostic tool is still required. Immunodiagnosis seems promising, but attempts to generate monoclonal antibodies have not yet been successful. This study aims to develop a single-chain variable antibody fragment (scFv) against Rhophilin-associated tail protein 1-like (ROPN1L), the sperm-specific antigen of adult O. viverrini, which has not been reported elsewhere. (2) Methods: The target epitope for phage screening was L3-Q13 of OvROPN1L, which showed the highest antigenicity to human opisthorchiasis analyzed in a previous study. This peptide was commercially synthesized and used for phage library screening. The isolated phage was produced in a bacterial expression system and tested for specificity in vitro and in silico. (3) Results: One of fourteen phages, named scFv anti-OvROPN1L-CL19, significantly bound to rOvROPN1L compared with non-infected hamster fecal extracts. This phage clone was successfully produced and purified using Ni-NTA chromatography. Indirect ELISA demonstrated that scFv anti-OvROPN1L-CL19 has a high reactivity with O. viverrini-infected hamster fecal extracts (12 wpi, n = 6) in comparison with non-infected hamster fecal extracts (0 wpi, n = 6), while the polyclonal rOvROPN1L antibodies did not show such a difference. Molecular modeling and docking confirmed our in vitro findings. (4) Conclusion: scFv anti-OvROPN1L-CL19 could be used as an effective material for developing O. viverrini-immunodiagnostic procedures in the future.

The foodborne liver trematode Opisthorchis felineus (Rivolta, 1884) is a member of the triad of phylogenetically related epidemiologically important Opisthorchiidae trematodes, which also includes O. viverrini (Poirier, 1886) and Clonorchis sinensis (Loos, 1907). Despite similarity in the life cycle, Opisthorchiidae liver flukes also have marked differences. Two species (O. viverrini and C. sinensis) are recognized as Group 1A biological carcinogens, whereas O. felineus belongs to Group 3A. In this review, we focus on these questions: Are there actual differences in carcinogenicity among these 3 liver fluke species? Is there an explanation for these differences? We provide a recent update of our knowledge on the liver fluke O. felineus and highlight its differences from O. viverrini and C. sinensis. In particular, we concentrate on differences in the climate of endemic areas, characteristics of the life cycle, the range of intermediate hosts, genomic and transcriptomic features of the pathogens, and clinical symptoms and morbidity of the infections in humans. The discussion of these questions can stimulate new developments in comparative studies on the pathogenicity of liver flukes and should help to identify species-specific features of opisthorchiasis and clonorchiasis pathogenesis.

Foodborne trematodes (FBT) of public health significance include liver flukes (Clonorchis sinensis, Opisthorchis viverrini, O. felineus, Fasciola hepatica and F. gigantica), lung flukes (Paragonimus westermani and several other Paragonimus spp.) and intestinal flukes, which include heterophyids (Metagonimus yokogawai, Heterophyes nocens and Haplorchis taichui), echinostomes (Echinostoma revolutum, Isthmiophora hortensis, Echinochasmus japonicus and Artyfechinostomum malayanum) and miscellaneous species, including Fasciolopsis buski and Gymnophalloides seoi. These trematode infections are distributed worldwide but occur most commonly in Asia. The global burden of FBT diseases has been estimated at about 80 million, however, this seems to be a considerable underestimate. Their life cycle involves a molluscan first intermediate host, and a second intermediate host, including freshwater fish, crustaceans, aquatic vegetables and freshwater or brackish water gastropods and bivalves. The mode of human infection is the consumption of the second intermediate host under raw or improperly cooked conditions. The major pathogenesis of C. sinensis and Opisthorchis spp. infection includes inflammation of the bile duct which leads to cholangitis and cholecystitis, and in a substantial number of patients, serious complications, such as liver cirrhosis and cholangiocarcinoma, may develop. In lung fluke infections, cough, bloody sputum and bronchiectasis are the most common clinical manifestations. However, lung flukes often migrate to extrapulmonary sites, including the brain, spinal cord, skin, subcutaneous tissues and abdominal organs. Intestinal flukes can induce inflammation in the intestinal mucosa, and they may at times undergo extraintestinal migration, in particular, in immunocompromised patients. In order to control FBT infections, eating foods after proper cooking is strongly recommended.