Abstract
Species delimitation has been divided by two approaches: DNA barcoding that focuses on standardization of the genetic marker and multilocus methods that place a premium on genomic coverage and conceptual rigor in modeling the divergence process. Most multilocus methods fail as barcodes, however, because few assay the same marker set and are therefore not readily comparable across studies and databases. We introduce ultraconserved elements (UCEs) as potential genomic barcodes that allow rigorous species delimitation and a bridge to DNA barcoding database to allow both rigorous species delimitation and standardized identification of delimited taxa. UCEs query thousands of loci across the nuclear genome in way that is replicable across broad taxonomic groups (i.e., vertebrates). We apply UCEs to species delimitation in a species complex of frogs found in the Mexican Highlands. Sarcohyla contains 24 described species, many of which are critically endangered and known only from their type localities. Evidence suggests that one broadly distributed member of the genus, S. bistincta, might contain multiple species. We generated data from 1,891 UCEs, which contained 1,742 informative SNPs for S. bistincta and closely related species. We also captured mitochondrial genomes for most samples as off-target bycatch of the UCE enrichment process. Phylogenies from UCEs and mtDNA agreed in many ways, but differed in that mtDNA suggested a more complex evolutionary history perhaps influenced by reticulate processes. The species delimitation method we used identified eight putative species (which we call lineages pending further study) within S. bistincta. Being able to compare linked mtDNA data to existing sequences on Genbank allowed us to identify one of these lineages nested within S. bistincta as an already-described species, S. pentheter. Another lineage nested within S. bistincta is currently being described as a new species (referred to here as sp. nov.). The remaining six lineages fell into two non-sister clades, one containing the core S. bistincta mostly in Oaxaca and Guerrero, and another in the Transvolcanic Belt. The latter clade, at 10% divergence in mtDNA and paraphyletic with respect to other S. bistincta, is a clear candidate for species status. Our study demonstrates not only that UCEs can be used as effective genomic DNA barcodes, but that combining multilocus genomic data with mtDNA is a powerful approach for both delimiting species and identifying them in poorly described and phenotypically challenging groups.
