Polymerase chain reaction detection of Opisthorchis viverrini metacercaria in freshwater cyprinid fish from local markets in Nakhonnayok province, Thailand

Abstract Background/Objectives: Opisthorchis viverrini , a foodborne zoonotic trematode endemic to Southeast Asia, is a major cause of cholangiocarcinoma and an environmental public health concern. Its transmission relates to aquatic ecosystems, waterborne diseases, and health–environment interactions. This study assessed prevalence and distribution in freshwater cyprinid fish from Nakhon Nayok, Thailand, using polymerase chain reaction (PCR)-based diagnostics. Materials and Methods: A total of 359 fish representing five cyprinid species were collected from four districts. Tissue samples (gills, scales, and flesh) underwent enzymatic digestion for metacercarial recovery, followed by deoxyribonucleic acid extraction. Species-specific PCR targeting a 330 bp fragment was conducted, and products were visualized by agarose gel electrophoresis. Results: Among the 359 fish, Henicorhynchus siamensis was most common (27.6%), followed by Barbodes schwanenfeldi (25.9%), Barbodes gonionotus (24.5%), Trichopodus trichopterus (10.6%), and Anabas testudineus (11.4%). PCR revealed species- and tissue-specific infections: H. siamensis showed the highest prevalence (15.2% scales, 9.1% flesh, and 6.1% gills), T. trichopterus moderate (12.9% gills, 6.5% scales), and B. gonionotus low (2.3%), while none were detected in A. testudineus or B. schwanenfeldi . Ban Na showed the highest positivity (28%), followed by Ongkharak (12%) and Pak Phli (5%), with none in Mueang. Conclusion: This study highlights species- and region-specific variations of O. viverrini infection in freshwater fish from Nakhon Nayok. PCR provided reliable early detection, supporting molecular tools for surveillance and targeted public health interventions in opisthorchiasis control.