Detailed information on species delineation and population genetic structure is a prerequisite for designing effective restoration and conservation strategies for imperiled organisms. Phylogenomic and population genomic analyses based on genome-wide ddRAD-Seq data has identified three allopatric lineages in the North American freshwater mussel genus Cyprogenia. Cyprogenia stegaria is restricted to the Eastern Highlands and displays little genetic structuring within this region. However, two allopatric clusters of C. aberti in the Ozark and Ouachita highlands exhibit substantial levels (mean FST = 0.120) of genetic differentiation and each warrants recognition as a distinct evolutionary lineage. Lineages of Cyprogenia in the Ouachita and Ozark highlands are further subdivided reflecting structuring at the level of river systems. Species tree inference and species delimitation in a Bayesian framework using SNP data supported results from phylogenetic analyses, which favors recognizing three species of Cyprogenia over the currently recognized two species.

A comparison of SNPs generated from both destructively and non-destructively collected samples revealed no significant difference in the SNP error rate, quality and amount of ddRAD sequence reads, indicating that nondestructive or trace samples can be effectively utilized to generate SNP data for organisms for which destructive sampling is not permitted.