Spatial analysis of cholangiocarcinoma in relation to diabetes mellitus and Opisthorchis viverrini infection in Northeast Thailand.

Cholangiocarcinoma (CCA) exhibits a heightened incidence in regions with a high prevalence of Opisthorchis viverrini infection, with previous studies suggesting an association with diabetes mellitus (DM). Our study aimed to investigate the spatial distribution of CCA in relation to O. viverrini infection and DM within high-risk populations in Northeast Thailand. Participants from 20 provinces underwent CCA screening through the Cholangiocarcinoma Screening and Care Program between 2013 and 2019. Health questionnaires collected data on O. viverrini infection and DM, while ultrasonography confirmed CCA diagnoses through histopathology. Multiple zero-inflated Poisson regression, accounting for covariates like age and gender, assessed associations of O. viverrini infection and DM with CCA. Bayesian spatial analysis methods explored spatial relationships. Among 263,588 participants, O. viverrini infection, DM, and CCA prevalence were 32.37%, 8.22%, and 0.36%, respectively. The raw standardized morbidity ratios for CCA was notably elevated in the Northeast's lower and upper regions. Coexistence of O. viverrini infection and DM correlated with CCA, particularly in males and those aged over 60 years, with a distribution along the Chi, Mun, and Songkhram Rivers. Our findings emphasize the association of the spatial distribution of O. viverrini infection and DM with high-risk CCA areas in Northeast Thailand. Thus, prioritizing CCA screening in regions with elevated O. viverrini infection and DM prevalence is recommended.

Influence of misconceptions and inappropriate eating behaviors on Opisthorchis viverrini infection among at-risk populations undergoing cholangiocarcinoma screening in Northeastern Thailand.

Raw or undercooked freshwater fish consumption contributes to persistent Opisthorchis viverrini infection in Northeast Thailand. This study aims to assess the relationship between misconceptions, unhealthy eating habits, and O. viverrini infection. Data were obtained from the Cholangiocarcinoma Screening and Care Program in Northeast Thailand from 2019 to 2021. Participants were screened for O. viverrini annually over the following 2 years using the Kato-Katz technique. Misconceptions and unhealthy eating habits were assessed through questionnaires. The relationship between these factors and O. viverrini infection was evaluated using adjusted risk ratios (ARRs) and 95% confidence intervals (CIs) from generalized estimating equations under binomial regression framework. Of 5375 participants screened for O. viverrini over 3 years, infection rates were 21.53%, 10.7%, and 4.6% each year, respectively. Out of those, 636 participants responded to questions regarding misconceptions. Results showed that participants who believed in the efficacy of putting lime or red ants in Koi pla (raw fish salad) or eating Koi pla with white whiskey to kill parasites, and early-stage cholangiocarcinoma can be cured, were 41% (ARR, 1.41; 95% CI, 1.03-1.94) and 57% (ARR, 1.57; 95% CI, 1.06-2.33), respectively, more likely to be infected with O. viverrini. Our study confirms that belief in using lime or red ants in Koi pla or eating Koi pla with white whiskey to make it cooked, or early-stage cholangiocarcinoma can be cured, increases O. viverrini infection risk in high-risk populations. Changing health beliefs and eating habits is necessary to reduce O. viverrini infection and its risk to cholangiocarcinoma.

Opisthorchis viverrini and Strongyloides stercoralis mono- and co-infections: Bayesian geostatistical analysis in an endemic area, Thailand.

Parasitic infections caused by Opisthorchis viverrini and Strongyloides stercoralis remain a major public health threat in the Greater Mekong Sub-region. An understanding of climate and other environmental influences on the geographical distribution and emergence of parasitic diseases is a crucial step to guide targeted control and prevention programs. A parasitological survey was conducted from 2008 to 2013 and included 12,554 individuals (age between 20 and 60 years) from 142 villages in five districts in Khon Kaen Province, Thailand. Geographical information systems, remote sensing technologies and a Bayesian geostatistical framework were used to develop models for O. viverrini and S. stercoralis mono- and co-infections in areas where both parasites are known to co-occur. The results indicate that male sex, increased age, altitude, precipitation, and land surface temperature have influenced the infection rate and geographical distribution of mono- and co-infections of O. viverrini and S. stercoralis in this area. Males were 6.69 times (95% CrI: 5.26-8.58) more likely to have O. viverrini - S. stercoralis co-infection. We observed that O. viverrini and S. stercoralis mono-infections display distinct spatial pattern, while co-infection is predicted in the center and southeast of the study area. The observed spatial clustering of O. viverrini and S. stercoralis provides valuable information for the spatial targeting of prevention interventions in this area.

Reduction of Reinfection Rates with Opisthorchis viverrini through a Three-Year Management Program in Endemic Areas of Northeastern Thailand.

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.

Recent advances on the immunobiology of Bithynia spp. hosts of Opisthorchis viverrini.

This article reviews the past and present scientific reports regarding Bithynia spp. focusing on the biology, ecology and life cycle of Bithynia snails and their responses to Opisthorchis viverrini infection. Moreover, new data regarding comparative molecular genomics and proteomic approaches have recently revealed novel molecular components involved in the immune defence responses from Bithynia spp., providing additional perspectives for future studies. Studies on the specific interaction between Bithynia snails and their trematodes will contribute to further understanding the snail-parasite relationship with regards to epidemiology and control of Opisthorchiasis and broaden the scope on comparative immunology of gastropod snails.

Bayesian spatial analysis of cholangiocarcinoma in Northeast Thailand.

Cholangiocarcinoma (CCA) is a malignant neoplasm of the biliary tract. Thailand reports the highest incidence of CCA in the world. The aim of this study was to map the distribution of CCA and identify spatial disease clusters in Northeast Thailand. Individual-level data of patients with histopathologically confirmed CCA, aggregated at the sub-district level, were obtained from the Cholangiocarcinoma Screening and Care Program (CASCAP) between February 2013 and December 2017. For analysis a multivariate Zero-inflated, Poisson (ZIP) regression model was developed. This model incorporated a conditional autoregressive (CAR) prior structure, with posterior parameters estimated using Bayesian Markov chain Monte Carlo (MCMC) simulation with Gibbs sampling. Covariates included in the models were age, sex, normalized vegetation index (NDVI), and distance to water body. There was a total of 1,299 cases out of 358,981 participants. CCA incidence increased 2.94 fold (95% credible interval [CrI] 2.62-3.31) in patients >60 years as compared to ≤60 years. Males were 2.53 fold (95% CrI: 2.24-2.85) more likely to have CCA when compared to females. CCA decreased with a 1 unit increase of NDVI (Relative Risk =0.06; 95% CrI: 0.01-0.63). When posterior means were mapped spatial clustering was evident after accounting for the model covariates. Age, sex and environmental variables were associated with an increase in the incidence of CCA. When these covariates were included in models the maps of the posterior means of the spatially structured random effects demonstrated evidence of spatial clustering.

Ecological Niche Model based on Maximum Entropy for mapping distribution of Bithynia siamensis goniomphalos, first intermediate host snail of Opisthorchis viverrini in Thailand.

The snail Bithynia siamensis goniomphalos acts as first intermediate host of the liver fluke, Opisthorchis viverrini, which causes opisthorchiasis in humans. In this study, we used a geographic information system (GIS), remote sensing (RS) and software using the maximum entropy (MaxEnt) algorithm to predict the distribution of B. s. goniomphalos in Thailand on the basis of environmental and climatic factors. The MaxEnt model for B. s. goniomphalos was excellent, with average test AUC values of 0.89. The predicted distribution of B. s. goniomphalos was affected by altitude, land cover, normalized difference vegetation index (NDVI), precipitation in the driest month (BIO 14), land surface temperature (LST) and soil pH. The areas suitable for B. s. goniomphalos were mostly in Northeast Thailand and some northern parts of the country. The presence of B. s. goniomphalos decreases with increasing altitude and increasing NDVI value. Bithynia s. goniomphalos is most likely to occur in paddy fields and cropland. Opisthorchiasis prevalence was directly proportional to the likelihood of snail occurrence as predicted by the model. This is the first time this ecological niche model has been used to predict Bithynia snail distribution and hence to provide a basis for future work of opisthorchiasis prevention in opisthorchiasis-endemic countries.

Differential Protein Expression in the Hemolymph of Bithynia siamensis goniomphalos Infected with Opisthorchis viverrini.

Bithynia siamensis goniomphalos is a freshwater snail that serves as the first intermediate host of the human liver fluke Opisthorchis viverrini. This parasite is a major public health problem in different countries throughout the Greater Mekong sub-region (Thailand, southern Vietnam, Lao PDR and Cambodia). Chronic O. viverrini infection also results in a gradual increase of fibrotic tissues in the biliary tract that are associated with hepatobiliary diseases and contribute to cholangiocarcinoma (a fatal type of bile duct cancer). Infectivity of the parasite in the snail host is strongly correlated with destruction of helminths by the snail's innate immune system, composed of cellular (hemocyte) and humoral (plasma) defense factors. To better understand this important host-parasite interface we applied sequential window acquisition of all theoretical spectra mass spectrometry (SWATH-MS) to identify and quantify the proteins from the hemolymph of B. siamensis goniomphalos experimentally infected with O. viverrini and compare them to non-infected snails (control group). A total of 362 and 242 proteins were identified in the hemocytes and plasma, respectively. Of these, 145 and 117 proteins exhibited significant differences in expression upon fluke infection in hemocytes and plasma, respectively. Among the proteins with significantly different expression patterns, we found proteins related to immune response (up-regulated in both hemocyte and plasma of infected snails) and proteins belonging to the structural and motor group (mostly down-regulated in hemocytes but up-regulated in plasma of infected snails). The proteins identified and quantified in this work will provide important information for the understanding of the factors involved in snail defense against O. viverrini and might facilitate the development of new strategies to control O. viverrini infection in endemic areas.

Infectivity and development of Opisthorchis viverrini metacercariae in immunosuppressed Barbonymus gonionotus fingerlings (Cyprinidae).

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.

Reprint of "Experimental infection of Opisthorchis viverrini cercariae to the cyprinid fish, Barbonymus gonionotus".

Opisthorchis viverrini is an important public health problem, a major cause of cholangiocarcinoma in the Greater Mekong subregion including Lao PDR, Cambodia, Vietnam and Thailand. Humans acquire the infection by consumption of raw, fermented or partially cooked freshwater cyprinid fish containing infective metacercariae. This study examined the effect of cercarial infection dosage (25-200 cercariae), age (1-60 day) and size (1-24 mm) of Barbonymus gonionotus fish on infection success of O. viverrini cercariae. Additionally, the site of cyst formation of O. viverrini cercariae and subsequent development to the metacercariae was examined. The result showed that cercarial infection dose (F4,95=8.52, P<0.001) age (P<0.001, ƛLR(2)=954.72) and size (P<0.001, ƛLR(2)=1204.84) were significantly associated with number of O. viverrini metacercariae recovery with in every additional fish age, the O. viverrini metacercarial recovery rate increased by 5.4% (P<0.001). Most metacercariae were found in the body of the fish (81.4%). We concluded that O. viverrini cercarial infection dosage, and age and size of fish were important determinants for a successful infection to fish.

Localization of the cystogenous glands of Opisthorchis viverrini cercariae.

Opisthorchiasis in northeastern Thailand is an important etiology of cholangiocarcinoma. To form the infectious stage, free swimming cercariae penetrate cyprinid fish, shed their tails, and then secret a cystic substance to cover their larval stage to form metacercariae in the fish body. We determined the location of the cystogenous glands in Opisthorchis viverrini cercariae. The cercariae and metacercariae were obtained from the naturally infected snail host, Bithynia siamensis goniomphalos and from cyprinid fish, respectively. The cyst walls of the metacercariae were separated and used to immunize inbred male BALB/c mice to obtain cyst wall antibodies. The general characteristics of the O. viverrini cercariae and metacercariae were studied by hematoxylin and eosin (H&E) staining of sections. The location and ultrastructure of the cystogenous glands of cercariae were studied by immunoperoxidase, immunofluorescence and transmission electron microscopy. The structures and organelles of cercariae and metacercariae could be identified, but the cystogenous glands could not be detected in H&E sections. The immunoperoxidase and immunofluorescence sections revealed positive reactions for cystogenous glands predominanted in the lateral part of the cercariae and were clearly seen in the cyst wall of the metacercariae. The ultrastructure of the cystogenous glands contained semitranslucent electron dense oval shaped granules. If interference occurs during the formation of the cysts by fish immune response, the metacercariae may not develop to maturity. It may be easily digested or degraded by human stomach acid and pepsin. This may be an efficient method for control of O. viverrini infection which requires further detailed study.

Experimental infection of Opisthorchis viverrini cercariae to the cyprinid fish, Barbonymus gonionotus.

Opisthorchis viverrini is an important public health problem, a major cause of cholangiocarcinoma in the Greater Mekong subregion including Lao PDR, Cambodia, Vietnam and Thailand. Humans acquire the infection by consumption of raw, fermented or partially cooked freshwater cyprinid fish containing infective metacercariae. This study examined the effect of cercarial infection dosage (25-200 cercariae), age (1-60 day) and size (1-24mm) of Barbonymus gonionotus fish on infection success of O. viverrini cercariae. Additionally, the site of cyst formation of O. viverrini cercariae and subsequent development to the metacercariae was examined. The result showed that cercarial infection dose (F4,95=8.52, P<0.001) age (P<0.001, ƛLR(2)=954.72) and size (P<0.001, ƛLR(2)=1204.84) were significantly associated with number of O. viverrini metacercariae recovery with in every additional fish age, the O. viverrini metacercarial recovery rate increased by 5.4% (P<0.001). Most metacercariae were found in the body of the fish (81.4%). We concluded that O. viverrini cercarial infection dosage, and age and size of fish were important determinants for a successful infection to fish.

Effect of soil surface salt on the density and distribution of the snail Bithynia siamensis goniomphalos in northeast Thailand.

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).