Environmental DNA is effective in detecting the federally threatened Louisiana Pinesnake (Pituophis ruthveni)

Aron D Katz,Jinelle H Sperry,Elizabeth C Sternhagen,Christopher A Melder,Lynsey R Harper,Mark A Davis,Sarah E Pearce

doi:10.1002/edn3.126

Abstract

AbstractSuccessful conservation of rare, threatened, or endangered (RTE) species is dependent upon rapid and accurate assessment of their distribution and abundance. However, assessments are challenging as RTE species typically exist as numerically small populations in often fragmented habitats and can possess complex natural histories. Environmental DNA (eDNA) analysis may provide a rapid, cost‐effective means of assessing RTE species presence/absence in viable habitat patches. We evaluated the efficacy of eDNA surveillance for the Louisiana Pinesnake (Pituophis ruthveni), an elusive, semi‐fossorial, nonvenomous colubroid snake endemic to Louisiana and Texas, USA, that has dramatically declined in both distribution and abundance. We developed two quantitative polymerase chain reaction (qPCR) assays that target the mitochondrial cytochrome c oxidase subunit I (COI) and mitochondrially encoded ATP synthase membrane subunit 6 (ATP6) genes. We validated each assay in silico, in vitro, and in situ, and investigated the influence of eDNA extraction method and genetic marker on assay performance. Both assays were highly sensitive and successfully detected the Louisiana Pinesnake under artificial and field conditions, including bedding samples collected from captive snake enclosures (100%), soil samples from Louisiana Pinesnake release sites (100%), and soil samples from sites where Louisiana Pinesnakes were documented via radio telemetry (45%). Although differences between genetic markers were negligible, assay performance was strongly influenced by eDNA extraction method. Informed by our results, we discuss methodological and environmental factors influencing Louisiana Pinesnake eDNA detection and quantification, broader implications for management and conservation of the Louisiana Pinesnake and other terrestrial reptiles and provide recommendations for future research. We suggest that eDNA surveys can more effectively assess Louisiana Pinesnake occupancy than conventional sampling, highlighting the need for comprehensive eDNA monitoring initiatives to better identify suitable habitat that will promote persistence of this imperiled species going forward.

Highlights

Effective biodiversity conservation in the Anthropocene is reliant upon accurate delimitation of species’ distributions and abundances (Niemiller et al, 2018)
Environmental DNA analysis stands firmly at the forefront of species monitoring as a viable alternative to conventional tools. eDNA is trace DNA released by organisms into their environment via secretions (Ficetola, Miaud, Popanon, & Tab erlet, 2008; Jerde, Mahon, Chadderton, & Lodge, 2011), excretions (Anderson et al, 2011; Martellini, Payment, & Villemur, 2005; Thomsen et al, 2012), or even decomposing carcasses (Merkes, McCalla, Jensen, Gaikowski, & Amberg, 2014, but see Curtis & Larson, 2020) that can be harnessed from environmental samples without observation or direct capture of the target organism itself (Thomsen & Willerslev, 2015)
Our results indicate that QPS performed better than CIA extraction for Louisiana Pinesnake eDNA detection and concentration (Figures 2–4), but the QPS approach may be cost prohibitive for some conservation efforts (USD 5.86 per extraction as of April 2020)

Summary

Introduction

Effective biodiversity conservation in the Anthropocene is reliant upon accurate delimitation of species’ distributions and abundances (Niemiller et al, 2018). These factors can negate our ability to detect species This is an acute problem for amphibians and reptiles (Durso & Seigel, 2015; Steen, 2010), and snakes present a prickly case. Given the problematic nature of inventorying and monitoring elusive, semi-fossorial, and rare species, novel approaches to improve detection are imperative for accurate assessment of occupancy and quantification of distributions and abundances. An eDNA assay was developed for Red Cornsnake, Pantherophis guttatus Linnaeus, 1766, and successfully used for laboratory-based eDNA accumulation/degradation experiments, but the assay was not tested under field conditions (Kucherenko et al, 2018) These studies illustrate the potential and pitfalls of eDNA analysis as an additional tool for surveying snakes

Findings

Discussion

Conclusion