Environmental DNA Biosurveillance of Chelonid Herpesvirus 5 ( ChHV5 ) in Endangered Sea Turtles Around Singapore

Sea turtle fibropapillomatosis (FP) is a potentially debilitating and neoplastic disease closely associated with the aetiological agent chelonid herpesvirus 5 (ChHV5). FP occurs in all endangered sea turtle species across most oceanic basins, with incidences rising rapidly in many geographic locations. Although FP is an emerging disease threat, the prevalence of FP and ChHV5 in turtle populations around Southeast Asia remains relatively understudied, underscoring the need for effective disease monitoring and surveillance strategies. To address this gap, our study reports the first near‐complete ChHV5 genome from Southeast Asia, which was obtained from an FP‐afflicted turtle stranded in Singapore waters in 2022. Along with the host mitochondrial genome, the viral genome provides a key reference point for ChHV5 distribution and host population origins from the region. To further enhance monitoring efforts, we also explored the application of environmental DNA (eDNA) methods as a non‐invasive biosurveillance tool for detecting ChHV5 DNA in seawater and sediment. Using real‐time PCR, ChHV5 DNA was successfully detected in rehabilitation tank water housing the afflicted turtle, with viral abundance increasing after turtle reintroduction into the tank. This observation is consistent with direct viral shedding as a potential ChHV5 transmission route. Lastly, the feasibility of detecting ChHV5 DNA from sediment samples using real‐time PCR was assessed through a spike‐and‐recovery experiment. This method was subsequently applied to sediment samples collected from hawksbill turtle nesting sites around Singapore, with no positive detections observed. Overall, the eDNA methods and next‐generation sequencing approaches described herein establish a baseline for ChHV5 biosurveillance, providing important reference data to inform future sea turtle monitoring efforts in Southeast Asia.