Development of eDNA markers for Coregonids in the Great Lakes

Aaron Maloy (USFWS, Aaron_Maloy@fws.gov), Sam Silverbrand (FWS), Stacey Nerkowski (FWS), Amanda Ackiss (USGS)

The abundance and distribution of coregonids within the Great Lakes has declined, with some species functionally absent from lakes previously occupied. Initial restoration efforts have focused on description and identification of species, their current range, assessment to identify population distributions and status, and captive propagation and rearing efforts for supplementing existing populations or reintroductions into lakes where the species was no longer detected. Environmental DNA (eDNA) can be used to locate unknown populations, monitor spawning locations and provide a more complete estimate of contemporary distributions. In order to fully leverage eDNA methods, additional baseline sequencing data is needed, and additional species-specific markers must be validated. Currently, eDNA species-specific markers exists for lake whitefish and round whitefish species which are easily distinguished form other Coregonid species. However, development of species-specific markers to distinguish the deepwater ciscoes has been a challenge. The USFWS NEFC has previously developed eDNA markers for C. artedi from C. hoyi. These markers are routinely used in Chaumount Bay and other embayments of Lake Ontario for monitoring spawning reefs and have identified low levels of C. artedi and C. hoyi DNA in other locations. While these markers have performed well, they have not been field validated in the upper lakes and do not distinguish between the two species. There are no species-specific markers for other deepwater ciscoes (C. kiyi, C. reighardi) or other rare species (C. nipigon, C. zenithicus). Existing mitochondrial sequence data for Cisco species are limited and publicly available data on GenBank can be misleading due to morphologically similar species being easily misidentified. These limitations in public data necessitate further sequencing data from Cisco specimens with accurate species identification. Fortunately, previous work (USGS, Amanda Ackiss) developed a panel of SNPs (single nucleotide polymorphism) that provide high resolution taxonomic identifications based on nuclear DNA. The SNP based identification has already been completed for multiple specimens of each north America cisco species along with additional whitefish species. The sequencing data used to obtain the SNP data is based on whole genome shotgun sequencing, which provide both nuclear and mitochondrial DNA. eDNA markers are specifically designed to target mitochondrial DNA because a cell can contain up to 2000 copies of each mitochondrial gene as opposed to just two copies of each nuclear gene. Targeting mitochondrial DNA in the environment greatly increase the probability of detection over that of nuclear DNA. Existing sequence data obtained for SNP analysis contains mitochondrial DNA but has not yet been used to assemble the mitochondrial genome of sequenced individuals. In addition, DNA exists for additional specimens that could be sequenced to obtain the mitochondrial genome data needed to design new cisco eDNA markers. Using existing sequencing data and DNA resources, minimal physical collection of new specimens should be needed.