Detection of terrestrial mammals using environmental DNA during heavy rainfall events and associated influencing factors

Recent developments in environmental DNA (eDNA) analyses have facilitated non-invasive and cost-effective ecological monitoring. Based on eDNA of terrestrial species released into water upon contact, simultaneous detection of aquatic and terrestrial species is feasible. However, an efficient sampling design for terrestrial vertebrate eDNA in aquatic environments has not yet been established because DNA is rarely released into these environments. In this study, we targeted eDNA transported from land to rivers through surface runoff during rainfall in three rivers and one irrigation channel within the Kiyotake River system, Japan. We quantified the eDNA concentration of a specific terrestrial vertebrate (<i>Bos taurus</i>) using digital polymerase chain reaction (PCR) and examined the efficiency of using filter papers with different pore sizes (0.7 µm and 2.7 µm). We also assessed the influence of various environmental factors (<i>e.g.</i>, rainfall characteristics described by the parameters of Gaussian distribution, water turbidity) on eDNA detection across different rainfall events. During the surveys, target DNA was detected in 42 out of 47 samples, suggesting the feasibility of consistently detecting terrestrial mammals from stormwater runoff. Overall, compared with the glass fiber filter with larger pore size, the smaller pore size filter captured more eDNA. The generalized linear mixed model revealed that prolonged rainfall duration, turbidity, and pH had a significant positive effect on eDNA concentration, whereas the distance from the assumed point of entry into the river to the sampling point had a significant negative effect. These results suggest that the runoff and transport of eDNA from terrestrial areas to rivers are enhanced under prolonged rainfall conditions, although eDNA degrades while transported along a longer watercourse by biochemical decomposition and sedimentation.