Trophic relationships, distribution and interactions among invasive and native Laurentian Great Lakes biota assessed using metabarcoding of stomach content DNA (scDNA) and environmental DNA (eDNA)

As aquatic invasive species (AIS) proliferate, they pose serious threats to native taxa, foodwebs and ecosystems. Massively-parallel, high-throughput, next-generation sequencing (NGS) and metabarcoding of environmental DNA (eDNA) and predator stomach content DNA (scDNA) are new approaches to detect AIS and can facilitate detailed analyses of AIS impacts. Using such technology, degraded and digested samples, and cryptic taxa can be identified with high sensitivity. The Laurentian Great Lakes (Great Lakes) are highly invaded and to determine occurrence patterns of AIS and native species inhabiting them I used a shortened portion of Cytochrome Oxidase One (CO1) and NGS metabarcoding. I developed, optimized and validated novel target-species CO1 PCR primer sets for AIS invertebrates: Bythotrephes longimanus, Cercopagis pengoi, Dreissena rostriformis bugensis, Dreissena polymorpha, and Hemimysis anomala. I tested primer set sensitivities, specificities, and multiplexing and used these with universal primer sets to analyze field-sampled scDNA and eDNA. I analyzed influences of abiotic (spatiotemporal) and biotic (predator species and size) variables in relation to prey occurrences in scDNA. The lowest AIS DNA in detected with novel primer sets in PCRs was: B. longimanus = 2.07; C. pengoi = 0.0002; D. r. bugensis = 0.0009; D. polymorpha = 0.103; and H. anomala = 0.127 (Chapter 2). Detection limits within mixes of interfering DNA (as percentage of total DNA) were: B. longimanus = 3.90 %, C. pengoi = 0.003 %, D. r. bugensis = 0.020 %, D. polymorpha = 0.170 % and H. anomala = 0.019 %. To determine invertebrate AIS roles in Lake Michigan, I metabarcoded scDNA from alewife (Alosa pseudoharengus), bloater (Coregonus hoyi), ninespine stickleback (Pungitius pungitius), rainbow smelt (Osmerus mordax) and slimy sculpin (Cottus cognatus) sampled Spring 2009 and 2010 at 73-128 m depths from three offshore sites. I detected multiple occurrences of target AIS and three native prey (Leptodiaptomus sicilis, Limnocalanus macrurus, and Mysis diluviana; Chapter 3). Driving variation in occurrences of the AIS and native taxa in scDNA was sample site, highlighting localized ecological and foodweb differences in ongoing AIS roles and impacts. Predator species effects likely reflected realized ecological feeding niches. To metabarcode scDNA from Lake Erie commercially fished predator species walleye (Sander vitreus), white bass (Morone chrysops), white perch (Morone americana) and yellow perch (Perca flavescens), I used novel AIS primer sets (Chapter Two) and a universal primer set to target fishes. Three invertebrate AIS including C. pengoi, B. longimanus, and D. r. bugensis, and AIS prey fishes gizzard shad (Dorosoma cepedianum), rainbow smelt and round goby (Neogobius melanostomus), and native prey fishes channel catfish (Ictalurus punctatus) and emerald shiner (Notropis athernoides) were detected, and all were relatively common components of scDNA. Occurrence patterns varied significantly with season, predator species, year, and predator size; however, the significances of factors also varied with prey species. Prey occurrences revealed dynamics among native and AIS prey and predators, potential competition, or prey selectivity which varied by way of these factors (Chapter 4). In 2013, 43 sites each in the Sydenham and Grand Rivers were sampled and metabarcoded for invertebrate eDNA using a universal primer set. Data revealed spatial patterns of AIS at sites within and among each river. Key AIS identified were: D. r. bugensis, Branchiura sowerbyi, Potamothrix moldaviensis, Craspedacusta sowerbyi and Skistodiaptomus pallidus. Further, I identified rare native mollusks including Quadrula quadrula and Villosa fabalis - threatened and endangered species in lower and middle reaches of the Sydenham. Novel results and information will provide important guidance to those tasked with ongoing challenges of managing, slowing spread of and eradicating AIS, and conserving native species