From Water Samples to Entire Fish Community Assessments: A New Next-Generation Sequencing Assay for Environmental DNA

Environmental DNA (eDNA) research is rapidly expanding beyond species detection and into the population genetics and genomics realm, having broad management application. We developed a new eDNA next-generation sequencing assay to simultaneously provide a “community snapshot”, identifying and quantifying relative abundances of all individual fish species in a water sample. Detectable are all Great Lakes fish species, as well as high-risk potential invasive fish species. We developed primers and optimized PCR conditions to amplify diagnostic regions of the cytochrome b and COI mitochondrial DNA genes of all targeted fishes (>200 species), and are adding a nuclear gene. First vetting our assay in the laboratory, Illumina sequencing libraries were prepared for a series of simulated fish communities containing known, varying concentrations of tissue DNA extractions from 10 representative fish species, including native and high-risk potential invasive species. Observed relative abundances of aligned sequence outputs were highly correlated to expected relative abundances, confirming the assay's performance. The assay is being tested using water samples from aquaria and field experiments with known species presence and abundances (the latter from netting and electroshocking), including the Maumee River.