Vegetation is an underutilized medium for environmental DNA (eDNA) sampling.eDNA methods leveraging water as a substrate exclude application to many terrestrial species. The use of eDNA to detect small mammals can complement current survey approaches (live capturing, track plating, and camera trapping) while reducing risks to the animals. The endangered New Mexico meadow jumping mouse (Zapus hudsonius luteus) is specialized to herbaceous riparian zones, making it an ideal candidate for developing a terrestrial eDNA detection method. We developed a species-specific assay for quantitative real-time PCR, then tested the long-term persistence of jumping mouse eDNA on plant material using four herbaceous day nests collected three to six months after occupancy. We conducted a field trial using sterile cotton swabs at six locations along two occupied streams to evaluate our assay's capability to detect present-day eDNA. Each of 60 swabs was used to swab a 0.50 m 2 area along streamside transects that included vegetation such as forbs, grasses, and sedges. We also opportunistically swabbed plants (n = 9) following visual observation of jumping mice.We determined the limit of detection for both assays are fewer than eight copies per reaction. We detected eDNA in three of four nests. From field trial samples, we successfully detected the species from randomly swabbed vegetation (N = 3), and four of nine swabs from vegetation recently used by individuals. Further work is required to develop a robust survey method using this eDNA detection approach. Our study demonstrated that mammalian eDNA can persist on nest vegetation long after the animal was present, highlighting the promise of using eDNA from plants to detect rare or endangered terrestrial species.
Leptonycteris nivalis (the Mexican long-nosed bat) is an endangered nectar-feeding bat species that follows “nectar corridors” as it migrates from Mexico to the southwestern United States. Locating these nectar corridors is key to their conservation and may be possible using environmental DNA (eDNA) from these bats. Hence, we developed and tested DNA metabarcoding and qPCR eDNA assays to determine whether L. nivalis could be detected by sampling the agave flowers on which it feeds. We sampled plants with known bat visitations in the Sierra Madre Oriental in Laguna de Sanchez (LS), Nuevo León, Mexico, and in the Chisos Mountains in Big Bend National Park, Texas, USA (CB). A total of 13 samples included both swabs of agave umbels and cuttings of individual flowers. DNA metabarcoding was performed as a PCR multiplex that targeted bats (SFF-COI), arthropods (ANML-COI), and plants (ITS2 and rbcL). We targeted arthropods and plants in parallel with bats because future metabarcoding studies may wish to examine all the pollinators and plants within the nectar corridor. We developed and tested the sensitivity and specificity of two qPCR assays. We found that both DNA metabarcoding and qPCR were highly successful at detecting L. nivalis (11 of 13 for DNA metabarcoding and 12 of 13 for qPCR). Swabs and flower cuttings and both qPCR assays detected the species over four replicates. We suggest that L. nivalis leaves substantial DNA behind as it forages for nectar. We also suggest that future studies examine the time since sampling to determine its effect on detection success. The DNA metabarcoding multiplex will be useful for parallel questions regarding pollination ecology, while, with further testing, the qPCR assays will be effective for large-scale sampling for the detection of migration corridors and foraging areas. This work may be relevant to other nectar-feeding bat species, which can likely be detected with similar methodologies.
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