Evolutionary Insights from Analyses of Spatial Genetic Variation in North American Frogs

Comparing patterns of spatial genetic structure across co-distributed species is critical to making inferences about the factors influencing population divergence, persistence, and change over time. The fields of phylogenetics and phylogeography are changing rapidly with the application of high-throughput sequencing to non-model organisms. Cost-effective use of this technology for these studies, which often include a relatively small portion of the genome for several taxa, requires strategies for genome partitioning and sequencing multiple individuals in parallel. In this dissertation, I implement several strategies to develop markers and collect large multi-locus datasets to investigate evolutionary history and spatial genetic structure of North American frogs. In the first study, I estimated a multilocus phylogeny for the North American chorus frog genus Pseudacris using anonymous nuclear loci developed using a reduced representation library approach. I sequenced 27 nuclear loci and three mitochondrial loci for 44 individuals on 1/3 of an Illumina MiSeq run, obtaining 96.5% of the targeted amplicons at less than 20% of the cost of traditional Sanger sequencing. I found heterogeneity among gene trees, although four major clades (Trilling Frog, Fat Frog, crucifer, and West Coast) were consistently supported, and resolved the relationships among these clades for the first time with strong support. Bayesian concordance analysis in BUCKy and species tree analysis in *BEAST produced largely similar topologies, although I identified taxa that require additional investigation in order to clarify taxonomic and geographic range boundaries. In the next study, I further examined one species complex within the chorus frog genus across its range, the Illinois Chorus Frog (Pseudacris illinoensis) and Strecker’s Chorus Frog (P. streckeri). Populations of P. illinoensis, which was originally described as a subspecies of P. streckeri, occur in small, disjunct regions associated with scarce sand prairie habitats that have been impacted by human development and agricultural practices. In addition to the anonymous nuclear and mitochondrial loci described above, I developed 24 microsatellite loci to study the genetic structure of P. illinoensis and P. streckeri, investigate the evolutionary history of this species complex, and help inform conservation efforts. I implemented a Bayesian model comparison approach to investigate previous biogeographic hypotheses while simultaneously inferring the presence of genetic structure in this species complex. Phylogenetic analyses uncovered a deep divergence between populations in the Edwards Plateau of central Texas and all other P. streckeri/P.illinoensis populations, but suggested the disjunct distribution of P. illinoensis occurred more recently. The best-supported migration model was consistent with the hypothesis that central Texas represented a refugium from which populations expanded via multiple routes. This model also indicated that disjunct northern and southern regions of P. illinoensis should be considered genetically distinct management units. This study provides an evolutionary context for future studies and conservation efforts in P. illinoensis, and demonstrates the utility of model-based approaches for phylogeographic inference. In the last two studies, I used targeted sequence capture and high-throughput sequencing of four anurans collected from 36 congruent localities to investigate aspects of phylogeographic concordance in the Southeastern U.S. Coastal Plain. In total, I recovered >375 nuclear loci across species and assembled nearly complete mitochondrial genomes, resulting in one of the most comprehensive phylogeographic datasets to date. Individual nuclear gene trees were poorly resolved even with long (~1400 bp) nuclear sequences, suggesting little concordance among nucleotide sites. The mitochondrial and combined nuclear datasets were largely concordant, and indicated high spatial and temporal discordance among species, which may correspond to differences in natural history. The Southern Toad (Anaxyrus terrestris) exhibited the weakest genetic structure and the Southern Leopard Frog (Lithobates sphenocephalus) exhibited the most resolved patterns of genetic structure. Two treefrogs (Hyla cinerea and H. squirella) had very similar levels of structure, except for two highly divergent H. cinerea individuals in the westernmost part of the sampled range. When compared to paleoclimate niche models, I found the two species with divergent western and eastern clades (H. cinerea and L. sphenocephalus) had more limited areas of climatic niche stability from the Last Glacial Maximum to present. I concluded that differential responses to past climate change combined with differences in natural history characteristics have shaped discordant patterns of genetic structure in anuran amphibians of the Southeastern Coastal Plain. Overall, large-scale genomic datasets combined with ecological and climatic data provide insight into the processes underlying subtle patterns of intraspecific genetic variation.