Detection, movement and persistence of invertebrate eDNA in groundwater.

Sampling groundwater biodiversity is difficult because of limited access and issues with species identification. Environmental DNA (eDNA) provides a viable alternative to traditional sampling approaches, however limited knowledge of the abundance and fate of DNA in groundwater hinders the interpretation of data from these environments. Groundwater environments are dark and have lower oxygen concentrations and microbial activity than surface waters. Consequently, assumptions about DNA fate in surface ecosystems may not apply to groundwaters. Here, we test the longevity and transport of eDNA in groundwater within a static microcosm and a flow-through mesocosm. A variety of invertebrates were placed within a mesocosm and microcosm to enable DNA shedding, and then removed. DNA persisted for up to 5 weeks after their removal in the static experiment and was detected between 9 and 33 days in the flow-through experiment. Sediments and water both proved important for eDNA detection. Crustacean DNA was detected sporadically and unpredictably, whereas non-crustacean DNA was detected more frequently despite their lower densities. We suggest that detecting crustaceans poses a challenge to utilising eDNA approaches for stygofauna monitoring. This is confounded by the scarcity of sequences for stygofauna in reference databases. Further research is needed before eDNA alone can be routinely employed for stygofauna detection.