Dataset Information

Exploration of Environmental DNA (eDNA) to Detect Kirtland's Snake (Clonophis kirtlandii).

ABSTRACT: Environmental DNA (eDNA) surveys utilize DNA shed by organisms into their environment in order to detect their presence. This technique has proven effective in many systems for detecting rare or cryptic species that require high survey effort. One potential candidate for eDNA surveying is Kirtland's Snake (Clonophis kirtlandii), a small natricine endemic to the midwestern USA and threatened throughout its range. Due to its cryptic and fossorial lifestyle, it is also a notoriously difficult snake to survey, which has limited efforts to understand its ecology. Our goal was to utilize eDNA surveys for this species to increase detection probability and improve survey efficiency to assist future conservation efforts. We conducted coverboard surveys and habitat analyses to determine the spatial and temporal activity of snakes, and used this information to collect environmental samples in areas of high and low snake activity. In addition, we spiked artificial crayfish burrows with Kirtland's Snake feces to assess the persistence of eDNA under semi-natural conditions. A quantitative PCR (qPCR) assay using a hydrolysis probe was developed to screen the environmental samples for Kirtland's Snake eDNA that excluded closely related and co-occurring species. Our field surveys showed that snakes were found in the spring during the first of two seasons, and in areas with abundant grass, herbaceous vegetation, and shrubs. We found that eDNA declines within a week under field conditions in artificial crayfish burrows. In environmental samples of crayfish burrow water and sediment, soil, and open water, a single detection was found out of 380 samples. While there may be physicochemical and biological explanations for the low detection observed, characteristics of assay performance and sampling methodology may have also increased the potential for false negatives. We explored these outcomes in an effort to refine and advance the successful application of eDNA surveying in snakes and groundwater microhabitats.

SUBMITTER: Ratsch R

PROVIDER: S-EPMC7341209 | biostudies-literature | 2020 Jun

REPOSITORIES: biostudies-literature

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Publications

Exploration of Environmental DNA (eDNA) to Detect Kirtland's Snake (<i>Clonophis kirtlandii</i>).

Ratsch Rikki R Kingsbury Bruce A BA Jordan Mark A MA

Animals : an open access journal from MDPI 20200619 6

Environmental DNA (eDNA) surveys utilize DNA shed by organisms into their environment in order to detect their presence. This technique has proven effective in many systems for detecting rare or cryptic species that require high survey effort. One potential candidate for eDNA surveying is Kirtland's Snake (<i>Clonophis kirtlandii</i>), a small natricine endemic to the midwestern USA and threatened throughout its range. Due to its cryptic and fossorial lifestyle, it is also a notoriously difficul ...[more]

PMID: 32575432

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Project description:The environmental DNA (eDNA) method is the practice of collecting environmental samples and analyzing them for the presence of a genetic marker specific to a target species. Little is known about the sensitivity of the eDNA method. Sensitivity is the probability that the target marker will be detected if it is present in the water body. Methods and tools are needed to assess the sensitivity of sampling protocols, design eDNA surveys, and interpret survey results. In this study, the sensitivity of the eDNA method is modeled as a function of ambient target marker concentration. The model accounts for five steps of sample collection and analysis, including: 1) collection of a filtered water sample from the source; 2) extraction of DNA from the filter and isolation in a purified elution; 3) removal of aliquots from the elution for use in the polymerase chain reaction (PCR) assay; 4) PCR; and 5) genetic sequencing. The model is applicable to any target species. For demonstration purposes, the model is parameterized for bighead carp (Hypophthalmichthys nobilis) and silver carp (H. molitrix) assuming sampling protocols used in the Chicago Area Waterway System (CAWS). Simulation results show that eDNA surveys have a high false negative rate at low concentrations of the genetic marker. This is attributed to processing of water samples and division of the extraction elution in preparation for the PCR assay. Increases in field survey sensitivity can be achieved by increasing sample volume, sample number, and PCR replicates. Increasing sample volume yields the greatest increase in sensitivity. It is recommended that investigators estimate and communicate the sensitivity of eDNA surveys to help facilitate interpretation of eDNA survey results. In the absence of such information, it is difficult to evaluate the results of surveys in which no water samples test positive for the target marker. It is also recommended that invasive species managers articulate concentration-based sensitivity objectives for eDNA surveys. In the absence of such information, it is difficult to design appropriate sampling protocols. The model provides insights into how sampling protocols can be designed or modified to achieve these sensitivity objectives.

Project description:Environmental DNA (eDNA) methods are used to detect DNA that is shed into the aquatic environment by cryptic or low density species. Applied in eDNA studies, occupancy models can be used to estimate occurrence and detection probabilities and thereby account for imperfect detection. However, occupancy terminology has been applied inconsistently in eDNA studies, and many have calculated occurrence probabilities while not considering the effects of imperfect detection. Low detection of invasive giant constrictors using visual surveys and traps has hampered the estimation of occupancy and detection estimates needed for population management in southern Florida, USA. Giant constrictor snakes pose a threat to native species and the ecological restoration of the Florida Everglades. To assist with detection, we developed species-specific eDNA assays using quantitative PCR (qPCR) for the Burmese python (Python molurus bivittatus), Northern African python (P. sebae), boa constrictor (Boa constrictor), and the green (Eunectes murinus) and yellow anaconda (E. notaeus). Burmese pythons, Northern African pythons, and boa constrictors are established and reproducing, while the green and yellow anaconda have the potential to become established. We validated the python and boa constrictor assays using laboratory trials and tested all species in 21 field locations distributed in eight southern Florida regions. Burmese python eDNA was detected in 37 of 63 field sampling events; however, the other species were not detected. Although eDNA was heterogeneously distributed in the environment, occupancy models were able to provide the first estimates of detection probabilities, which were greater than 91%. Burmese python eDNA was detected along the leading northern edge of the known population boundary. The development of informative detection tools and eDNA occupancy models can improve conservation efforts in southern Florida and support more extensive studies of invasive constrictors. Generic sampling design and terminology are proposed to standardize and clarify interpretations of eDNA-based occupancy models.

Dataset Information

Exploration of Environmental DNA (eDNA) to Detect Kirtland's Snake (Clonophis kirtlandii).

Publications

Exploration of Environmental DNA (eDNA) to Detect Kirtland's Snake (<i>Clonophis kirtlandii</i>).

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