Relationship Between Pesticide Exposure Factors and Health Symptoms Among Chili Farmers in Northeast Thailand.

OBJECTIVES: The unsafe use of pesticides in agriculture represents a major hazard to human health. This study was conducted to investigate the association between pesticide exposure and health symptoms among chili farmers in northeast Thailand. METHODS: This cross-sectional analytic study included 141 chili farmers in Sakon Nakhon Province, in northeast Thailand. Data regarding health symptoms were gathered using a self-report questionnaire. A medical technician tested blood cholinesterase activity using an erythrocyte method, and an occupational medicine specialist at Ramathibodi Hospital in Bangkok, Thailand reviewed the results. Associations between personal characteristics, pesticide exposure factors, and health symptoms were analyzed using multiple logistic regression. RESULTS: Of the 141 chili farmers studied, 66.7% experienced pesticide poisoning, as indicated by below-normal cholinesterase levels. Fatigue was the most frequently reported symptom associated with pesticide exposure, affecting 37.6% of participants. This was followed by nausea and vomiting (31.9%), dizziness (14.9%), and dry throat (14.9%). Multivariate logistic regression analysis revealed that several factors were significantly associated with adverse symptoms: amount of work experience, volume of pesticides used, use of chemical pesticides, use of leaking containers during spraying, direct pesticide exposure while working, contact with pesticide-soaked clothing, consumption of food and drinks in the fields, and blood cholinesterase level indicating risk. CONCLUSIONS: This study suggests potential health risks for chili farmers stemming from exposure to and contamination by pesticides used in agricultural practices. To mitigate these risks, it is essential to supply personal protective equipment and to implement educational programs aimed at improving protective behaviors among farmers.

Examination of Diagnostic Performance of New IgG4 Rapid Test Compared with IgG- and IgG4-ELISAs to Investigate Epidemiology of Strongyloidiasis in Northeast Thailand.

Strongyloidiasis, caused by Strongyloides stercoralis, is a neglected tropical disease with a global distribution. The infection can be fatal in immunocompromised individuals, and accurate diagnosis leading to timely treatment can save lives. Serodiagnosis is a sensitive method for diagnosis and is recommended for screening high-risk individuals. A point-of-care rapid test will facilitate the screening activities, especially in low-resource settings. This study aims to apply a new IgG4 immunochromatographic test using S. stercoralis recombinant antigen (SsRapid® cassette test) and to compare it with in-house IgG and IgG4 enzyme-linked immunosorbent assays (IgG- and IgG4-ELISAs) using native Strongyloides ratti antigen to investigate the epidemiology of strongyloidiasis in northeast Thailand. A total of 300 people participated, with 136 males and 164 females of a similar mean age. The reference tests were fecal examinations using the formalin-ethyl acetate concentration technique and an agar plate culture technique. The prevalence of S. stercoralis determined by SsRapid (81.7%) was significantly higher than that by fecal examinations (43.3%) or by antibody detection by IgG-ELISA (53.0%) or IgG4-ELISA (44.0%). The diagnostic sensitivities of SsRapid, IgG-ELISA, and IgG4-ELISA were found to be 93.9%, 77.7%, and 63.1%, respectively. The rate of positive tests by the SsRapid was significantly correlated to the levels of Strongyloides-specific IgG4 and IgG antibodies. By all diagnostic methods, male participants had a significantly higher prevalence of strongyloidiasis than females. Age was significantly associated with the concentration of specific serum IgG but not with the SsRapid grading score. In conclusion, SsRapid was shown to be a sensitive and valuable diagnostic test for the epidemiology study of strongyloidiasis.

Accuracy of a new rapid diagnostic test for urinary antigen detection and assessment of drug treatment in opisthorchiasis.

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.

Diagnostic performance of Strongyloides-specific IgG4 detection in urine for diagnosis of human strongyloidiasis.

BACKGROUND: Detection of parasite-specific IgG in urine is a sensitive method for diagnosis of strongyloidiasis and gives similar accuracy to serum IgG. However, there are no data concerning detection of IgG subclass in urine. To further explore the utility of diagnosis from urine samples, we evaluated the diagnostic performance of IgG4 in urine compared with parasitological and other immunological methods. METHODS: The urine and sera included proven strongyloidiasis (group 1, n = 93), other parasitic infections (group 2, n = 40) and parasite negatives (group 3, n = 93). The performance of Strongyloides-specific IgG4 in urine for diagnosis of strongyloidiasis using fecal examinations as the reference standard was assessed. RESULTS: With fecal examination as a gold standard, Strongyloides-specific IgG4 in urine had 91.4% sensitivity and 93.2% specificity while serum IgG4 had 93.6% sensitivity and 91.0% specificity. IgG4 in both urine and serum had almost perfect diagnostic agreements with fecal examination (Cohen's kappa coefficient was > 0.8). Cross-reactivity to Opisthorchis viverrini and Taenia spp. of IgG4 in urine were 7.5% and 12.5% in serum. Concurrent analyses of total IgG in urine and serum showed that the sensitivities (97.9-100%) and specificities (88.7-91.0%) were similar (P > 0.05). The sensitivity for parasitological examination by the formalin-ethyl acetate concentration technique (FECT) was 49.5% and that for agar plate culture technique (APC) it was 92.6%. CONCLUSION: Our findings showed that specific IgG4 detection in urine yielded similar diagnostic performance to the same biomarkers in serum. This suggests that accurate diagnosis of strongyloidiasis can be performed using urine samples and IgG4 is a valid choice of diagnostic marker. Further assessment is required to assess the utility of urine IgG4 for measuring the response treatment in strongyloidiasis.

Human Fecal Carriage of Streptococcus agalactiae Sequence Type 283, Thailand.

Streptococcus agalactiae (group B Streptococcus) sequence type 283 bacteremia, found almost exclusively in Southeast Asia, is associated with consuming raw freshwater fish, but some patients deny consumption. We detected fecal carriage in 5/184 (2.7%) persons in northeast Thailand. Human carriers might contribute to transmission or be the original source of this sequence type.

Comparison of a Urine Antigen Assay and Multiple Examinations with the Formalin-Ethyl Acetate Concentration Technique for Diagnosis of Opisthorchiasis.

Detection of worm antigen in urine is a sensitive diagnostic method for opisthorchiasis, particularly for light-intensity infections; however, the presence of eggs in feces is essential for validating results from the antigen assay. To address the issue of low sensitivity of fecal examination, we modified the protocol for the formalin-ethyl acetate concentration technique (FECT) and compared it against urine antigen measurements for detection of the parasite Opisthorchis viverrini. First, we optimized the FECT protocol by increasing the number of drops for examinations from the standard two drops to a maximum of eight. We were able to detect additional cases after examination of ≥ 3 drops, and the prevalence of O. viverrini saturated after examination of ≥ 5 drops. We then compared the optimized FECT protocol (examining five drops of suspension) against urine antigen detection for the diagnosis of opisthorchiasis in field-collected samples. The optimized FECT protocol detected O. viverrini eggs in 25 of 82 individuals (30.5%) who had positive urine antigen tests but were fecal egg negative by the standard FECT protocol. The optimized protocol also retrieved O. viverrini eggs in 2 of 80 antigen-negative cases (2.5%). In comparison with the composite reference standard (combined FECT and urine antigen detection), the diagnostic sensitivity of examining two and five drops of FECT and the urine assay was 58.2, 67, and 98.8%, respectively. Our results show that multiple examinations of fecal sediment increase the diagnostic sensitivity of FECT and thus provide further support for the reliability and utility of the antigen assay for diagnosis and screening of opisthorchiasis.

Long-term Persistence of Opisthorchis viverrini Antigen in Urine: A Prospective Study in Northeast Thailand.

Antigen detected in urine for the diagnosis of opisthorchiasis has a low daily variation; however, the longer term variability in antigen concentrations is unknown. In this study, we prospectively monitored Opisthorchis viverrini antigen concentrations for 30 consecutive days and at subsequent monthly intervals in a cohort of opisthorchiasis-positive individuals. On the basis of the monoclonal antibody-based ELISA, the profiles of antigen-positive rate and antigen concentration exhibited no significant change over 30 days with a mean proportion positive of 87.1% (range 73.7%-100%), and the average antigen concentration was 29.7 ± 2.2 ng/mL (mean ± SE). The urine antigen concentration at baseline was similar to the subsequent measurements at 2, 4, 6, and 10 months in the follow-up study (P > 0.05). The consistency and low daily and long-term fluctuation of O. viverrini antigen in urine demonstrates the reliability of urine assay for diagnosis of opisthorchiasis.

Effects of day-to-day variation of Opisthorchis viverrini antigen in urine on the accuracy of diagnosing opisthorchiasis in Northeast Thailand.

Antigen detection in urine using an enzyme-linked immunosorbent assay (ELISA) is more sensitive than fecal examination for diagnosis of opisthorchiasis and for assessment of the effects of drug treatment. It is not known whether day-to-day variation of urine composition, including levels of Opisthorchis viverrini antigen, influences the urine assay. We investigated this topic with the cooperation of participants from two localities in Northeast Thailand. Project participants were screened for parasite infections for three consecutive days using the quantitative formalin-ethyl acetate concentration technique (FECT) to detect O. viverrini eggs and the urine ELISA for detection of O. viverrini antigen. A subset of participants (n = 801) with matched fecal and urine samples were analyzed for comparison of inter-day prevalence estimates and the performance of the urine assay compared against FECT for diagnosis of opisthorchiasis. The daily prevalence measured by the urine assay ranged between 29.0%-30.2% while those by FECT ranged between 11.9%-20.2%. The cumulative three-day prevalence estimate determined by the urine antigen assay was 30.3%, which was significantly higher than that by FECT (20.2%, p < 0.05). A significant positive correlation was found between the concentration of antigen in urine and fecal egg counts (p < 0.001). Overall, the urine assay had better diagnostic performance for opisthorchiasis than fecal examination by FECT. The high sensitivity plus negligible daily variation of O. viverrini antigen in urine indicates the utility of the urine assay for diagnosis, as well as population screening, of opisthorchiasis.

Performance of Mini Parasep® SF stool concentrator kit, Kato-Katz, and formalin-ethyl acetate concentration methods for diagnosis of opisthorchiasis in Northeast Thailand.

BACKGROUND: Control and elimination of the liver fluke (Opisthorchis viverrini) is a primary preventive strategy against cholangiocarcinoma in Southeast Asia. A sensitive parasitological diagnostic method is required to facilitate a surveillance and control program. In this study, we evaluated the performance of Mini Parasep® SF stool concentrator kit (stool kit) compared with Kato-Katz (KK) and the quantitative formalin-ethyl acetate concentration technique (FECT) for detection of O. viverrini and co-endemic parasitic infections. METHODS: A cross-sectional survey for parasitic infection in residents aged > 15 years in a community in Kalasin province, Northeast Thailand, was conducted in 2018. Fecal samples were collected and screened by KK method, and a subset of samples was further examined by the stool kit and FECT methods. The results were analyzed for prevalence of parasitic infections in addition to the diagnostic performance of the methods for qualitative and quantitative detection of helminthiases. RESULTS: The initial survey of parasitic infection determined by the KK method (n = 567) showed the prevalence of O. viverrini was 32.63%, followed by Taenia 2.65%, echinostomes 1.76%, hookworms 1.41%, Trichuris trichiura 0.53% and Strongyloides stercoralis 0.53%. Within a subset of samples tested with multiple diagnostics (n = 150), the detection rates of O. viverrini by the stool kit, FECT and KK methods were 27.3%, 30.7% and 28.7%, respectively. The diagnostic sensitivity for opisthorchiasis was similar for FECT (75.5%), KK(66.0%) and the stool kit (67.3%). For other parasitic infections, FECT and stool kit methods performed better than KK, particularly in detecting minute intestinal flukes (MIF), S. stercoralis and coinfections. When measuring the intensity of O. viverrini infection (fecal egg counts), the stool kit results showed a significant positive correlation with KK and FECT (P < 0.05). CONCLUSIONS: As the stool kit is simple to use and shows a comparable performance to FECT, it may serve as an alternative method of fecal examination for screening of helminthiasis including opisthorchiasis.

Application of Urine and Copro Antigen Assays after Primary Infection and Drug Treatment in an Experimental Opisthorchiasis Animal Model.

Infection by Opisthorchis viverrini causes significant health problems, including cholangiocarcinoma (CCA); thus control and elimination of this trematode is an important strategy for the reduction of CCA. Currently, urine and copro antigen detection is more sensitive than parasitological examination of the feces for the diagnosis of opisthorchiasis. Given limitations in human studies, we used an animal model to quantify the parasite antigen profiles in urine and feces in O. viverrine-infected hamsters, and postdrug treatment. The positive detections of O. viverrini antigen began from week 1 in urine and week 2 in feces after infection until week 28 of the study. The recoveries of O. viverrini worms were detected starting from week 1 and eggs of O. viverrini were detected in feces from week 3 after infection and remained detectable throughout the study period. There was a significant positive correlation of urine and copro antigen levels with the number of fecal egg counts (P < 0.01) and worm recovery (P < 0.01). In the drug-treatment experiment, treatment of infected hamsters with praziquantel significantly reduced worm burden, fecal egg output, and antigen in urine and feces compared with the untreated controls (P < 0.001). At 4 weeks posttreatment, the egg and worm reduction rates were 100% and 95.5%, respectively. The positive antigen detections in urine and feces corresponded with partial worm clearance from praziquantel treatment. This study demonstrated a direct link of urine and copro antigen tests with worms infecting the liver thereby reaffirming the reliability of urine and copro antigen assay in opisthorchiasis diagnosis.

Population dynamics and diversity of trematode infections in Bithynia siamensis goniomphalos in an irrigated area in northeast Thailand.

Several trematodes including Opisthorchis viverrini utilize Bithynia siamensis goniomphalos as a snail intermediate host in their life cycles. In order to capture a comprehensive range of host­parasite interactions and their transmission dynamic patterns, B. s. goniomphalos were sampled monthly over 4 consecutive years in an irrigated paddy-field habitat in northeast Thailand. Using a standard cercarial shedding method, a high diversity of trematodes (17 types) was recovered. Virgulate xiphidiocercariae were the most prevalent (7.84%) followed by O. viverrini (0.71%). In addition to seasonal and environmental factors, the quantity of irrigation water for rice cultivation correlated with transmission dynamics of trematodes in B. s. goniomphalos. The peak prevalence of all trematode infections combined in the snails shifted from the cool-dry season in 2010­2012 to the hot-dry season in 2013 associated with an increasing quantity of water irrigation. A low frequency of mixed trematode infections was found, indicating that the emergence of virgulate cercariae, but not of O. viverrini, was negatively impacted by the presence of other trematodes in the same snail. Taken together, the observed results suggest that interactions between host and parasite, and hence transmission dynamics, depend on specific characteristics of the parasite and environmental factors including irrigated water for rice cultivation.

Concentration of Urine Samples Improves Sensitivity in Detection of Strongyloides-Specific IgG Antibody in Urine for Diagnosis of Strongyloidiasis.

Detection of IgG in urine is an efficient method comparable to that in serum for diagnosis of strongyloidiasis, but the effects of daily variation in urine dilution on diagnostic accuracy are not clearly known. This study evaluated the effects of urine concentration on the detection of parasite-specific IgG by urine enzyme-linked immunosorbent assay (ELISA), particularly in individuals with borderline results or false-negative diagnosis. Optimal concentration conditions were established by comparing Strongyloides-specific IgG antibody levels between unconcentrated and concentrated urine in participants with different infection intensities, namely, healthy control (HC), low-negative (LN), high-negative (HN), and low-positive (LP) groups. The optimal condition was selected and validated in a field trial study. The final urine concentration protocol required centrifugation at 4,000 × g at 4°C for 10 mins using the Amicon concentrator tube. This protocol was validated in groups of participants with various diagnoses according to urine ELISA and fecal examination (n = 148). The concentrated-urine ELISA increased the proportion of positive results in the LN group by 68.2% and by 100% in the HN group. Significantly elevated IgG antibody levels were seen in the LP group. In the group that was false negative by urine ELISA but positive by fecal examination (n = 28), concentrated-urine ELISA yielded 100% positive results. Overall, the frequency estimates of Strongyloides stercoralis were 23.6% by fecal culture, 27% by standard urine ELISA, and 90.5% by concentrated-urine ELISA. The concentration of urine samples prior to analysis by ELISA improved the sensitivity for diagnosis and is potentially useful in the diagnosis of strongyloidiasis in immunocompromised individuals or in low-prevalence areas.

Development and Efficacy of Droplet Digital PCR for Detection of Strongyloides stercoralis in Stool.

Human strongyloidiasis is one of the neglected tropical diseases caused by infection with soil-transmitted helminth Strongyloides stercoralis. Conventional stool examination, a method commonly used for diagnosis of S. stercoralis, has low sensitivity, especially in the case of light infections. Herein, we developed the droplet digital polymerase chain reaction (ddPCR) assay to detect S. stercoralis larvae in stool and compared its performance with real-time PCR and stool examination techniques (formalin ethyl-acetate concentration technique [FECT] and agar plate culture [APC]). The ddPCR results showed 98% sensitivity and 90% specificity, and real-time PCR showed 82% sensitivity and 76.7% specificity when compared with the microscopic methods. Moreover, ddPCR could detect a single S. stercoralis larva in feces, and cross-reactions with other parasites were not observed. In conclusion, a novel ddPCR method exhibited high sensitivity and specificity for detection of S. stercoralis in stool samples. This technique may help to improve diagnosis, particularly in cases with light infection. In addition, ddPCR technique might be useful for screening patients before starting immunosuppressive drug therapy, and follow-up after treatment of strongyloidiasis.

Genetic diversity and population structure of Haemonchus contortus in goats from Thailand.

Haemonchus contortus is one of the most economically important parasitic nematodes affecting small ruminant livestock worldwide. This study was conducted to elucidate the genetic diversity and population structure of this nematode in Thailand based on mitochondrial DNA markers, the nicotinamide dehydrogenase subunit 4 (nad4) and the cytochrome c oxidase subunit 1 (cox1) genes. One hundred and thirty-six adult worms were obtained from 86 abomasa of slaughtered goats from 13 different localities in 5 regions of Thailand. Identification to the genus Haemonchus was done using morphology. DNA sequences of the nuclear ribosomal second internal transcribed spacer (ITS2) identified each specimen to species: three fixed nucleotide (SNP) differences distinguished H. contortus from H. placei. Genetic analysis defined 118 and 122 unique haplotypes in partial sequences of nad4 (alignment length 723 bp) and cox1 (645 bp) genes, respectively. Nucleotide diversities were 0.031 and 0.043 for nad4 and cox1 genes, respectively. Low genetic differentiation was observed among H. contortus samples from various provinces in Thailand. This is the first study on the genetic diversity and population structure of H. contortus of goats in Thailand. This study has provided insights into the transmission dynamics of this parasitic nematode, information which is essential for farm management and parasite control.

Evaluation of a Rapid IgG4 Lateral Flow Dipstick Test to Detect Strongyloides stercoralis Infection in Northeast Thailand.

Strongyloides stercoralis affects more than half a billion people worldwide, and hyperinfection in immunocompromised patients can be fatal. Elimination of this neglected tropical disease requires field-applicable diagnostic tools. We conducted a laboratory evaluation of a lateral flow rapid dipstick test (SsRapid™) using sera samples from a Strongyloides-endemic area in northeast Thailand. Group 1 was S. stercoralis-positive and larvae- and/or antibody-positive (according to the IgG ELISA) (N = 100). Group 2 had negative fecal examination and IgG ELISA results (N = 25). Group 3 had other parasitic infections and negative IgG ELISA results (N = 25). The results showed good diagnostic sensitivity (82%) and excellent specificity (96%). Suggested improvements in the SsRapid™ test include increased diagnostic sensitivity and conversion to the more robust cassette format. Field studies should be performed as well.

Analysis of Daily Variation for 3 and for 30 Days of Parasite-Specific IgG in Urine for Diagnosis of Strongyloidiasis by Enzyme-Linked Immunosorbent Assay.

Recent work has found urine analysis to be as sensitive as serology for diagnosis of strongyloidiasis. Here, we examined the daily variation of Strongyloides-specific IgG in urine by qualitative and quantitative ELISA and its effects on diagnostic accuracy and reliability. In the first part of the study, matched urine and fecal samples were collected from project participants in northeast Thailand for three consecutive days. Urine samples were analyzed for Strongyloides-specific IgG by ELISA using Strongyloides ratti as the antigen source. Performance of urine ELISA was compared with parasitological diagnosis by agar plate culture technique (APCT) and formalin-ethyl acetate concentration technique (FECT). In the second part of the study, urine IgG levels were compared daily for thirty consecutive days. The prevalence of Strongyloides infection, as measured by urine ELISA for three consecutive days, was significantly higher than that found using parasitological methods (63.1% vs. 22%). There was slight daily variation in prevalence estimates according to urine ELISA while there were significant variations according to parasitological examination methods over three consecutive days. For the 3-day experiment, urine ELISA had 83-86% diagnostic sensitivity when compared with the fecal examination method or with a composite standard (combined results from fecal examination methods (APCT or FECT) and/or urine ELISA). The levels of parasite-specific IgG in urine were stable throughout both the 3-day and the 30-day studies. In conclusion, diagnosis of strongyloidiasis by urine ELISA is more sensitive than by fecal methods, with minimal daily variation for qualitative and quantitative diagnosis. Urine ELISA has potential for clinical diagnosis and population screening of strongyloidiasis.

Impact of geography and time on genetic clusters of Opisthorchis viverrini identified by microsatellite and mitochondrial DNA analysis.

Infection by the small liver fluke, Opisthorchis viverrini, causes serious public health problems, including cholangiocarcinoma, in Thailand and southeastern Asian countries. Previous studies have reported that O. viverrini represents a species complex with varying levels of genetic differentiation in Thailand and Lao PDR. In this study, we re-examined population genetic structure and genetic diversity of O. viverrini using extensive samples of the parasite collected over 15 years from 12 geographical localities in Thailand and eight localities in Lao PDR. Parasite life-cycle stages of 721 individuals of O. viverrini (91 cercariae, 230 metacercariae and 400 adult worms) were genotyped using 12 microsatellite loci. Metacercariae exhibited genetic diversity comparable with that of experimentally raised adults: metacercariae can therefore be used to represent O. viverrini populations without the need for laboratory definitive hosts. Data obtained from larval as well as adult worms identified two distinct genetic clusters of O. viverrini. Sequences of a portion of the mitochondrial cox1 gene strongly supported the existence of these two clusters. One, the widespread cluster, was found at all sampled sites. The second cluster occurred only in Phang Khon District, Sakon Nakhon Province (SPk), within the Songkram River wetland in Thailand. A striking feature of our data relates to the temporal dynamics of the SPk cluster, which was largely replaced by representatives of the widespread cluster over time. If the SPk cluster is excluded, no marked genetic differences were seen among O. viverrini populations from Thailand and Lao PDR. The underlying causes of the observed population structure and population dynamics of O. viverrini are not known.

Application of urine antigen assay to evaluate outcomes of praziquantel treatment and reinfection in opisthorchiasis in northeast Thailand.

BACKGROUND: A urine antigen assay was applied to evaluate chemotherapeutic outcomes and reinfection patterns of opisthorchiasis in Thailand. METHODS: We used a prospective study design by following opisthorchiasis subjects at baseline and post-treatment using a urine antigen assay and faecal examination by the formalin-ethyl acetate concentration technique (FECT). RESULTS: The antigen of Opisthorchis viverrini in urine diminished within 4 weeks after praziquantel treatment. Concurrent faecal examinations by FECT showed that faecal eggs were negative at 4 weeks after treatment. In a subsequent study, reinfection rates and intensity patterns of O. viverrini were evaluated at 48 weeks after praziquantel treatment. Within a group of subjects with curative treatment (n=137), 16.8% became reinfected according to FECT and 27.7% according to the urine antigen assay (p<0.05). There were significant correlations in intensity of infection between pretreatment and at 48 weeks post-treatment in both faecal egg counts and antigen levels in urine. CONCLUSIONS: The results suggested that in addition to screening, the urine antigen assay is an efficient tool for monitoring outcomes of drug treatment and reinfection in opisthorchiasis. Due to the ease of urine sample collection and handling, the urine assay becomes an alternative method to faecal examination for diagnosis and monitoring of opisthorchiasis.

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

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

Comparing the performance of urine and copro-antigen detection in evaluating Opisthorchis viverrini infection in communities with different transmission levels in Northeast Thailand.

To combat and eventually eliminate the transmission of the liver fluke Opisthorchis viverrini, an accurate and practical diagnostic test is required. A recently established urine antigen detection test using monoclonal antibody-based enzyme-linked-immunosorbent assay (mAb-ELISA) has shown promise due to its high diagnostic accuracy and the use of urine in place of fecal samples. To further test the utility of this urine assay, we performed a cross sectional study of 1,043 people in 3 opisthorchiasis endemic communities in northeast Thailand by applying urine antigen detection together with copro-antigen detection methods. The quantitative formalin-ethyl acetate concentration technique (FECT) was concurrently performed as a reference method. The prevalence of O. viverrini determined by urine antigen detection correlated well with that by copro-antigen detection and both methods showed 10-15% higher prevalence than FECT. Within the fecal negative cases by FECT, 29% and 43% were positive by urine and copro-antigen detection, respectively. The prevalence and intensity profiles determined by antigen detection and FECT showed similar patterns of increasing trends of infection with age. The concentration of antigen measured in urine showed a positive relationship with the concentration of copro-antigen, both of which were positively correlated with fecal egg counts. The data observed in this study indicate that urine antigen detection had high diagnostic accuracy and was in concordance with copro-antigen detection. Due to the ease and noninvasiveness of sample collection, the urine assay has high potential for clinical diagnosis as well as population screening in the program for the control and elimination of opisthorchiasis.