Establishing detection thresholds for environmental DNA using receiver operator characteristic (ROC) curves
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METHODS AND RESOURCES ARTICLE
Establishing detection thresholds for environmental DNA using receiver operator characteristic (ROC) curves Natasha R. Serrao1,2 · Scott M. Reid2 · Chris C. Wilson2
Received: 24 May 2016 / Accepted: 28 July 2017 © Springer Science+Business Media B.V. 2017
Abstract Environmental DNA (eDNA) detection is increasingly being used to assess the potential presence of aquatic species. Setting defensible thresholds for qPCR assays are necessary to differentiate “detection” from “nondetection”, but threshold rationales are rarely reported in eDNA studies. Detection thresholds for data inclusion and exclusion have implications for downstream data analysis; uncritical acceptance could result in false positive results (Type I error), whereas overly conservative thresholds could result in high false negative interpretation (Type II error). We assessed the use of the receiver operator characteristics (ROC) framework to identify a detection threshold for balancing sensitivity and specificity of eDNA data. The ROC framework was tested using empirical eDNA data from redside dace (Clinostomus elongatus), an endangered freshwater fish in southwestern Ontario. Based on negative controls, positive controls, and quality assurance
Data accessibility: Details of the eDNA primer and probe design and sequences are provided in Supplemental File 1. Data for the quantitative PCR trials summarized in Table 1 and Fig. 2 will be deposited in the Dryad data repository (provisional Dryad doi:10.5061/dryad.2h023). Electronic supplementary material The online version of this article (doi:10.1007/s12686-017-0817-y) contains supplementary material, which is available to authorized users. * Chris C. Wilson [email protected] 1
Environmental and Life Sciences Graduate Program, Trent University, 2140 East Bank Drive, Peterborough, ON K9L 0G2, Canada
2
Aquatic Research and Monitoring Section, Ontario Ministry of Natural Resources and Forestry, Trent University, 2140 East Bank Drive, Peterborough, ON K9L 0G2, Canada
guidelines, a minimum concentration of 1 copy reaction−1 was identified as the most appropriate threshold for this study. Although optimal thresholds will likely vary between studies and labs based on empirical data, the ROC framework provides a statistical method for establishing defensible thresholds for data acceptance or rejection as well as tradeoffs between the sensitivity and specificity of eDNA data. Keywords Environmental DNA (eDNA) · Thresholds · False positives · False negatives · Receiver operator characteristic (ROC) · Sensitivity · Specificity
Introduction Detection of environmental DNA (eDNA) to infer the presence of species in aquatic systems is increasingly being used to document potential occurrences of invasive and endangered species (Ficetola et al. 2008; Wilcox et al. 2013; Moyer et al. 2014). Although the method’s sensitivity and indirect detection of species occurrences make eDNA surveillance a potent tool for monitoring species, potential errors from false positive a
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