Environmental DNA storage and extraction method affects detectability for multiple aquatic invasive species

Environmental DNA (eDNA) refers to genetic material released by organisms into their surrounding environment. Collecting and identifying eDNA has gained popularity for monitoring and surveillance of aquatic invasive species. Invasive species management is most successful when an invasion is identified early while population size is likely to be low, highlighting the importance of eDNA detection sensitivity. Various factors influence DNA yield recovered from environmental samples. Environmental DNA storage and extraction methods, for example, can be adjusted to maximize DNA yield, thereby improving detectability. In this study, we compared the performance of two eDNA storage and extraction methods in detecting three common aquatic invasive species (Bythotrephes longimanus, Dreissena polymorpha, and Faxonius rusticus) across five natural ecosystems of Minnesota, United States. One method involved storing filters in 95% ethanol (EtOH) and extracting DNA using a DNeasy PowerSoil Pro Kit (Qiagen, Hilden, Germany), whereas the other method used cetyl trimethylammonium bromide (CTAB) for storage and a phenol–chloroform–isoamyl (PCI) procedure for DNA extraction. We also investigated the effect of DNA extract volume (1 μL relative to 3 μL) in qPCR reactions on eDNA detections for the commercial kit method. The CTAB‐PCI method yielded significantly more positive detections, across all three species, compared to the EtOH‐Qiagen method. Moreover, we found that using 1 μL of DNA extract in qPCR reactions was equally effective as using 3 μL. To improve detections of aquatic invasive species, we recommend that researchers store eDNA sample filters in CTAB or a similar lysis buffer such as Longmire's solution and extract with PCI when feasible, but note that lower extract volumes might be used without negative effect when either increasing technical replicates or repurposing samples for the detection of multiple species. In this study, we compared the performance of two eDNA storage and extraction methods in detecting three common aquatic invasive species across five natural ecosystems. We found that CTAB storage with a phenol–chloroform–isoamyl (PCI) extraction protocol out‐performed storage in ethanol and extraction with a common commercial kit. We recommend that when possible researchers use this CTAB‐PCI approach for surveillance of especially difficult‐to‐detect and high‐priority species.