Environmental DNA detection of the male mitochondrial genome of freshwater mussels (Unionidae)

Environmental DNA (eDNA) has shown promise for the detection of threatened and endangered species and has been implemented for monitoring aquatic spawning events. Freshwater unionid mussels exhibit a rare form of mitochondrial inheritance, in which males possess a unique mitochondrial mitotype that is divergent from the female mitotype. As freshwater mussels are spermcasters, the detection of male mitotype eDNA may provide critical conservation information related to timing of sperm release. This study re-purposed an existing eDNA metabarcoding dataset to detail the unique detection of eDNA pertaining to the male mitotype. Water samples collected alongside an extensive mussel salvage within the Walhonding River, Ohio, detected 16 distinct male mitotypes. However, several constraints limit the proper interpretation of these detections. There is currently a lack of reporting on assay compatibility with the male mitotype within freshwater mussel eDNA literature. Reference genetic databases are critically lacking, with only four of the 16 male eDNA sequences in this study able to be discerned to a species. This study highlights the importance of detailing these detections as the unique inheritance system provides opportunities to document difficult to record spawning behaviors, and eDNA may be employed as a survey tool to evaluate patterns of metapopulation geneflow.