No evidence that crayfish carcasses produce detectable environmental DNA (eDNA) in a stream enclosure experiment

Abstract

Environmental DNA (eDNA) is an emerging tool for monitoring invasive and imperiled species, particularly at low densities. However, the factors that control eDNA production, transport, and persistence in aquatic systems remain poorly understood. For example, the extent to which carcasses produce detectable eDNA is unknown. If positive detections are associated with dead organisms, this could confound monitoring for imperiled or invasive species. Here, we present results from one of the first studies to examine carcass eDNA in situ by deploying carcasses of the invasive red swamp crayfish (Procambarus clarkii) in a stream enclosure experiment for 28 days. We predicted that carcasses would initially produce eDNA that would decline over time as carcasses decayed. Unsurprisingly, crayfish carcasses lost biomass over time, but at the conclusion of our experiment much of the carapace and chelae remained. However, no eDNA of P. clarkii was detected in any of our samples at the crayfish density (15 P. clarkii carcasses at ∼615 g of biomass initially), stream flow (520–20,319 L/s), or temperature (∼14–25 °C) at our site. Subsequent analyses demonstrated that these results were not the consequence of PCR inhibition in our field samples, poor performance of the eDNA assay for intraspecific genetic diversity within P. clarkii, or due to the preservation and extraction procedure used. Therefore, our results suggest that when crayfish are relatively rare, such as in cases of new invasive populations or endangered species, carcasses may not produce detectable eDNA. In such scenarios, positive detections from field studies may be more confidently attributed to the presence of live organisms. We recommend that future studies should explore how biomass, flow, and differences in system (lentic vs. lotic) influence the ability to detect eDNA from carcasses.

Limitations of eDNA analysis for Carcinus maenas abundance estimations

“…Possible inhibition of PCR due to enzymatic activity released during decay was investigated, and no inhibition of primers was occurring. Such findings are similar to examinations by Curtis and Larson [ 53 ] of invasive red swamp crayfish ( Procambarus clarkia ). In a swamp enclosure, the carcasses of this crayfish released no detectable eDNA into the water column.…”

High stream flows dilute environmental DNA (eDNA) concentrations and reduce detectability

“…(2015), we extracted DNA from filters using a chloroform–isoamyl alcohol extraction procedure in a clean room, free from high‐copy DNA and isolated from the PCR laboratory. This extraction method (Renshaw et al., 2015) has been shown to be robust to inhibition from tannins or humic acids (Curtis & Larson, 2020; Schrader et al., 2012) and often produces higher DNA yields than other extraction procedures (Deiner et al., 2015). One extraction blank was used for every ~25 samples.…”

“…to be robust to inhibition from tannins or humic acids (Curtis & Larson, 2020;Schrader et al, 2012) and often produces higher DNA yields than other extraction procedures (Deiner et al, 2015). One extraction blank was used for every ~25 samples.…”

Environmental DNA detection of the giant freshwater crayfish (Astacopsis gouldi)

“…Environmental DNA‐based detection has already been applied to several crayfish species including the signal crayfish Pacifastacus leniusculus (Dana, 1852), Japanese crayfish Cambaroides japonicus (De Haan, 1841), red swamp crayfish Procambarus clarkii (Girard, 1852), spiny‐cheek crayfish Faxonius limosus (Rafinesque, 1817), Shasta crayfish Pacifastacus fortis (Faxon, 1914), and the rusty crayfish Faxonius rusticus (Girard, 1852) (Cowart et al, 2018; Harper et al, 2018; Ikeda et al, 2016; Mauvisseau et al, 2018; Tréguier et al, 2014). Despite concerns that eDNA detection of invertebrates may be challenging due to presence of exoskeletons which may hamper eDNA shedding (Curtis & Larson, 2020; Dunn et al, 2017; Tréguier et al, 2014), as well as poor assay performance when detecting other rare freshwater crayfish species (i.e., Procambarus clarkia (Girard, 1852); Tréguier et al, 2014, and Faxonius eupunctus Williams, 1952; Rice et al, 2018), studies demonstrate that eDNA‐based tools can be effective for crayfish surveillance, even at low densities (Dougherty et al, 2016; Harper et al, 2018). In this study, we developed and validated a sensitive and specific qPCR probe‐based assay to enable detection of A. gouldi from eDNA water samples.…”