Comprehensive DNA barcodes for species identification and discovery of cryptic diversity in mayfly larvae from South Korea: Implications for freshwater ecosystem biomonitoring

DNA barcoding of aquatic macroinvertebrates holds much promise as a tool for taxonomic research and for providing baseline reference for phylogenetic analysis and aquatic ecosystem biomonitoring. We obtained 112 novel sequences of the barcode region of the mitochondrial DNA cytochrome c oxidase subunit I gene representing 11 families, 25 genera, and 43 species of mayfly (Insecta: Ephemeroptera) from South Korea. No species shared barcode sequences and all can be identified with barcodes with a possible exception of some species. Minimum levels of interspecific genetic distances ranged from 6.7 to 32.9% (mean: 23.7%), whereas average levels of intraspecific divergence was 3.7%. The latter value was inflated by the presence of very high divergences within some taxa. In fact, approximately 33.3% (15/45) of the species included two or more haplotype clusters showing greater than 5.0% sequence divergence and some values were as high as 32.9%. Many of the species with high intraspecific divergences are para- or polyphyletic and represent the possibility of species complexes. Our study suggests that type or topotype specimens should be sequenced to identify accurate barcoding clusters with morphological species concepts and also to determine the status of currently synonymized species.