Tracing Life in Desert Springs: Optimizing eDNA Sampling to Reveal Hidden Aquatic Biodiversity.

Authors: Shohreh Mahdavinia, Duane Moser, Michael Schewmm, Alireza Saidi-Mehrabad, Yusuf Umar

Year: 2025

Abstract

Animals continuously shed DNA into their environment, creating trace genetic material known as environmental DNA (eDNA). This material can be used to non-destructively infer the presence and relative abundance of organisms through molecular detection methods such as quantitative PCR (qPCR) and metabarcoding. Here, we describe experiments designed to optimize eDNA sampling efficiency using a combined qPCR–metabarcoding approach. Fieldwork was conducted at the headwaters of the Amargosa River in Nevada, within the former 7J Ranch (Atwood Preserve). We tested a range of sampling variables, including filter pore sizes (0.45, 0.7, 1.2, and 5 µm), filter materials (polyethersulfone and glass fiber), and water column depth (surface neuston vs. midwater). Most experiments were conducted at Dace Spring, a high-quality spring believed to host only the endemic speckled dace (Rhinichthys osculus nevadensis). Results indicated that while finer pore-sized filters produced higher total DNA yields, larger pore sizes preferentially enriched for target fish DNA despite lower overall concentrations. Neuston samples consistently yielded less DNA, and qPCR detection rates of fish were correspondingly lower, using an adaptation of MiFish 12S rRNA gene primers. Our assays confirmed the absence of invasive mosquitofish (Gambusia affinis) and the presence of pure speckled dace in Dace Spring, while also verifying a cryptic population of dace in a small, previously unconfirmed spring. Additionally, metabarcoding revealed Micropterus salmoides (largemouth bass) in a nearby farm pond, though its DNA signal was absent in the pond’s anoxic hypolimnion. These findings demonstrate how eDNA sampling design, particularly filter selection and water-layer targeting can critically influence detection outcomes in desert spring ecosystems.