Optimizing Threatened Wood Turtle (Glyptemys insculpta) Environmental DNA Surveillance with Experimental Control Standardization

Abstract

Turtles are experiencing declines global while their distributions are poorly understood by modern science. Environmental DNA (eDNA) has proven to be a valuable tool in rapid species detection. However, in such a rapidly growing field, standardized practices have not yet taken hold, and rigorous comparisons among methodologies have not been evaluated. This thesis consists of two chapters that seek to answer questions facing eDNA methodologies used for monitoring of cryptic and endangered turtles. Chapter 1 is a literature review developed to provide comparisons on the current methods used for turtle eDNA research. Among current literature, turtles represent the most well-represented group of reptiles to date and most studies have been conducted in the global north with slow growth in the number of publications conducted south of the equator which includes several areas of high turtle diversity. Taxonomic representation of eDNA work is also disproportionate with the family Emydidae being the focus of more than 53% of the studies found in the literature to date. There is also a wide diversity of approaches in experimental design and the implementation of laboratory and field protocols. Factors such as sample type, genetic marker selection, and implementation of experimental controls are also highly variable between publications. Chapter 2 is a research chapter focusing on a threatened North American species. Wood Turtle (Glyptemys insculpta) is one of many turtle species currently experiencing declines across their range. However, their species status is still not fully understood. eDNA has been demonstrated to be an effective and sensitive noninvasive surveillance approach for detecting turtles that can provide rapid information on detection and occupancy for conservation monitoring and management. The field of eDNA requires dealing with several challenges as it relies on rigorous validation of the results by carefully assessing the sources of error that produce false results (false positives and false negatives) when conducting work to determine species presence. This chapter focuses on the ongoing issue involving field blank false positive amplifications which led to the development of a new standardized protocol. This new protocol has led to a reduction in false positives in field blanks showcasing the importance of standard eDNA practices concerning control usage at every stage of sampling.