Objectively delimiting species boundaries remains an important challenge in systematics and becomes urgent when unresolved taxonomy complicates conservation and recovery efforts. We examined species boundaries in the imperiled freshwater mussel genus Cyclonaias (Bivalvia: Unionidae) using morphometrics, molecular phylogenetics, and multispecies coalescent models to help guide pending conservation assessments and legislative decisions. Congruence across multiple lines of evidence indicated that current taxonomy overestimates diversity in the C. pustulosa species complex. The only genetically and morphologically diagnosable species in the C. pustulosa species complex were C. pustulosa and C. succissa and we consider C. aurea , C. houstonensis, C. mortoni , and C. refulgens to be synonyms of C. pustulosa . In contrast, all three species in the C. nodulata complex ( C. necki , C. nodulata , and C. petrina ) were genetically, geographically, and morphologically diagnosable. Our findings have important conservation and management implications, as three nominal species ( C. aurea , C. houstonensis , and C. petrina ) are being considered for protection under the Endangered Species Act.

Inaccurate systematics confound our ability to determine evolutionary processes that have led to the diversification of many taxa. The North American freshwater mussel tribe Lampsilini is one of the better-studied groups in Unionidae, however, many supraspecific relationships between lampsiline genera remain unresolved. Two genera previously hypothesized to be non-monophyletic that have been largely overlooked are Leptodea and Potamilus. We set out to resolve supraspecific relationships in Lampsilini and test the monophyly of Leptodea and Potamilus by integrating molecular, morphological, and life history data. Our molecular matrix consisted of four loci: cytochrome c oxidase subunit 1 (CO1), NADH dehydrogenase subunit 1 (ND1), internal transcribed spacer 1 (ITS1), and 28S ribosomal RNA. Secondly, we performed both traditional and Fourier shape morphometric analyses to evaluate morphological differences and finally, we compared our results with available life history data. Molecular data supported the paraphyly of both Leptodea and Potamilus, but nodal support was insufficient to make any conclusions regarding generic-level assignments at this time. In contrast, inference from our integrative taxonomic assessment depicts significant support for the recognition of a new species, Potamilus streckersoni sp. nov., the Brazos Heelsplitter. Our data show clear separation of three taxonomic entities in the P. ohiensis species complex: P. amphichaenus, P. ohiensis, and P. streckersoni sp. nov.; all molecularly, geographically, and morphologically diagnosable. Our findings have profound implications for unionid taxonomy and will aid stakeholders in establishing effective conservation and management strategies.http://www.zoobank.org/urn:lsid:zoobank.org:pub:502647C0-418B-4CC4-85A8-BD89FC3F674F

Bivalves exhibit an astonishing diversity of sexual systems and sex-determining mechanisms. They can be gonochoric, hermaphroditic or androgenetic, with both genetic and environmental factors known to determine or influence sex. One unique sex-determining system involving the mitochondrial genome has also been hypothesized to exist in bivalves with doubly uniparental inheritance (DUI) of mtDNA. However, the link between DUI and sex determination remains obscure. In this study, we performed a comparative gonad transcriptomics analysis for two DUI-possessing freshwater mussel species to better understand the mechanisms underlying sex determination and DUI in these bivalves. We used a BLAST reciprocal analysis to identify orthologs between Venustaconcha ellipsiformis and Utterbackia peninsularis and compared our results with previously published sex-specific bivalve transcriptomes to identify conserved sex-determining genes. We also compared our data with other DUI species to identify candidate genes possibly involved in the regulation of DUI. A total of ∼12,000 orthologous relationships were found, with 2,583 genes differentially expressed in both species. Among these genes, key sex-determining factors previously reported in vertebrates and in bivalves (e.g., Sry, Dmrt1, Foxl2) were identified, suggesting that some steps of the sex-determination pathway may be deeply conserved in metazoans. Our results also support the hypothesis that a modified ubiquitination mechanism could be responsible for the retention of the paternal mtDNA in male bivalves, and revealed that DNA methylation could also be involved in the regulation of DUI. Globally, our results suggest that sets of genes associated with sex determination and DUI are similar in distantly-related DUI species.