Environmental DNA enables detection of terrestrial mammals from forest pond water

Terrestrial animals must have frequent contact with water to maintain their lives, implying that environmental DNA (eDNA) originating from terrestrial animals should be detectable from places containing water in terrestrial ecosystems. Aiming to detect the presence of terrestrial mammals using forest water samples, we applied a set of universal PCR primers (MiMammal, a modified version of fish universal primers) for metabarcoding mammalian eDNA. After verifying the primers usefulness in silico and using water samples from zoo cages of animals with known species compositions, we collected five 500-ml water samples from ponds in two cool-temperate forests in Hokkaido, northern Japan. Using eDN