Application of environmental DNA analysis for the detection of Opisthorchis viverrini DNA in water samples

Opisthorchiasis, which can lead to cholangiocarcinoma in cases of chronic infection, is a major public health problem in Southeast Asian countries. The trematode, Opisthorchis viverrini, is the causative agent of the disease. Accurate and rapid monitoring of O. viverrini is crucial for disease prevention and containment. Therefore, in this study we sought to develop a novel species-specific real-time PCR assay for detecting O. viverrini using environmental DNA (eDNA). The diagnostic sensitivity of the newly developed real-time PCR assay was similar to that of the traditional PCR assay for 50 fecal samples collected in Lao PDR (21 and 19 samples were positive by real-time PCR and traditional PCR, respectively). The efficacy of eDNA analysis and its applicability in the field were tested using a total of 94 environmental water samples collected from 44 sites in Savannakhet, Lao PDR during May and October 2015 and February 2016. O. viverrini eDNA was detected in five samples by real-time PCR, indicating the presence of the fluke in the area and the risk of infection for individuals consuming fish from these water sources. The application of eDNA analysis would facilitate the identification of O. viverrini endemic hotspots and contribute to the ecological control of opisthorchiasis, and this strategy can be applied to other eukaryotic water pathogens.