Increased eDNA detection sensitivity using a novel high-volume water sampling method

Environmental DNA analysis has revolutionized the way we study rare, invasive, and endangered taxa. However, if eDNA testing is to become an increasingly reliable tool, high detection sensitivity is crucial. Current eDNA sampling methods, like filtration and precipitation, can only process small volumes of water per sample. If only a few samples are collected, eDNA from the target organism might be missed, leading to false-negative results. We developed an eDNA collection method for lentic systems that improved detection sensitivity while keeping the total number of samples low. Unlike filtration and precipitation, which mainly target extracellular DNA, this method specifically targets eDNA in larger particle sizes and is not limited to processing small volumes of water. A 64-micrometer mesh tow net was used to process >3,000 L of water per eDNA sample. We compared the tow net method to a common collection method, a 0.45 μm cellulose nitrate filter that processes about 1 L of water per eDNA sample. Paired tow and filter samples were collected at 37 locations and tested for two taxa: an aquatic plant, Northern watermilfoil (Myriophyllum sibiricum), and aquatic mollusks, including Helisoma anceps, using Kompetitive Allele Specific PCR (KASP) assays. We detected M. sibiricum significantly more frequently in tow samples than filter samples. Mollusks were detected in all eDNA samples (tow nets and filters), but when eDNA samples were diluted 25-fold to mimic a low target concentration scenario, mollusk DNA was detected significantly more frequently in tow samples than filter samples. This high-volume eDNA sampling method, using a tow net to process thousands of liters of water, can improve detection sensitivity for multiple taxa, making it a useful tool for researchers and managers.