Dr Megan Shaffer1,2, Dr Elizabeth Andruszkiewicz Allan1, Dr Pedro Brandao1, Dr Elizabeth Brasseale1, Dr Kim Parsons2, Dr Andrew Olaf Shelton2, Dr Tania Valdivia1, Dr Jilian Xiong1, Dr Ryan Kelly1
1University of Washington, Seattle, USA, 2NOAA – Northwest Fisheries Science Center, Seattle, USA
Biography:
Dr Megan Shaffer is a Research Scientist at the University of Washington and NOAA – Northwest Fisheries Science Center in Seattle, WA. She received her PhD in Marine Biology at Te Herenga Waka – Victoria University of Wellington in 2019.
Abstract:
Collection of environmental DNA (eDNA) offers a promising new approach to detect rare organisms in the environment that would likely remain undetected with traditional survey methods. Here, we evaluate critical steps of the eDNA workflow to optimize the detection of rare targets, including: (1) collection (filter pore size and volume filtered), (2) DNA preservation and extraction, and (3) metabarcoding primer choice. Leveraging a managed bottlenose dolphin population (Tursiops truncatus) in an open system, we quantitatively examined the effects of varying parameters for collection (1 versus 5 micron; 1 versus 3 L); preservation (desiccation, storage at -80ºC, Longmire's buffer, RNAlater, DNA/RNA Shield); and extraction (phenol-chloroform, Qiagen DNeasy Blood and Tissue Kit, Zymo Mini Prep Plus Kit), using a species-specific digital droplet PCR assay. Based on DNA copy yields, we found that smaller pore sizes, increased sample volume and the phenol chloroform extraction method led to the capture of more total DNA; however, they did not necessarily yield more target DNA. We then examined the relative performance of three common eDNA metabarcoding primers by sequencing approximately 500 samples collected at various depths (0-500 m) in 2019 along the US West Coast. We amplified samples using two general mitochondrial DNA (mtDNA) 12S markers that capture fish and marine mammals (MiFish, MarVer1) and a marine mammal-specific marker for the mtDNA control region (D-loop) and found substantial differences in performance in terms of both sensitivity and species resolution. Together, these findings contribute to our understanding of how to maximize detections of rare targets using eDNA.