Documenting species occurrences and habitat occupancy of unionid mussels can often be challenging. Environmental DNA (eDNA) has been shown to be a reliable tool for detecting unionids with comparable or greater sensitivity than conventional sampling and has the added advantages of not disturbing individuals or occupied habitats. However, single‐species eDNA assays are limited to targeting individual species of interest and are functionally blind to the presence of other species. Community eDNA assays have the potential to characterize local species assemblages simultaneously but are currently less extensively developed and implemented than single‐species eDNA testing. We tested the effectiveness of community eDNA markers to identify unionid species assemblages, using two overlapping conserved primers that target the maternal mitochondrial 16S rDNA region. Both primer sets were optimized using three mock communities and successfully amplified 62.5%–81.6% of species with largely consistent results between the primer sets. Following optimization, eDNA from water samples from 24 reference sites with known mussel communities was amplified and sequenced to quantify species richness and diversity within and among sites. Metabarcoding results from the monitoring sites largely mirrored those from the mock communities, with >80% of species detections identified by both assays. The results were broadly consistent with species data from quadrat‐based manual field surveys, although both community eDNA and conventional sampling detected some species that the other method did not. These results demonstrate that community eDNA assays using conserved primers and next‐generation sequencing have the potential to simultaneously target eDNA from multiple unionid species and provide a powerful tool for complementing or augmenting conventional field surveys to characterize and monitor unionid species assemblages.

Watershed inventories and population monitoring are essential components of efforts to conserve and recover freshwater mussel diversity in Canada. We used two datasets to assess the efficacy of a quadrat-based sampling protocol for: (1) detecting mussel species at risk; (2) characterizing species composition; (3) providing accurate estimates of abundance; and (4) detecting changes in density. The protocol is based on a systematic design (with random starts) that samples 20% of monitoring sites with visual-tactile surface searches and excavation of 1 m² quadrats. The first dataset included 40 sampling sites in five Ontario rivers, and the second dataset consisted of complete census sampling at two 375 m² sites that represented contrasting mussel assemblages. Our results show that the protocol can be expected to detect the majority of species present at a site and provide accurate and precise estimates of total mussel density. Excavation was essential for detection of small individuals and to accurately estimate abundance. However, the protocol was of limited usefulness for reliable detection of most species at risk. Furthermore, imprecise density estimates precluded detection of all but the most extreme changes in density of most individual species. Meeting monitoring objectives will require either substantially greater sampling effort under the current protocol, or a fundamental revision of the sampling approach.

Alterations to watercourses affect connectivity in aquatic systems and can influence dispersal of aquatic biota. Dams fragment populations and act as isolating barriers, but canals create connections between waterbodies that can be used as corridors for dispersal by opportunistic invaders. The Niagara Peninsula of Ontario, Canada, has a 200-y history of canal operation, resulting in major modification of the watercourses in the region. This modification allowed numerous invasive species to enter the upper Great Lakes (e.g., sea lamprey) and probably has facilitated dispersal in native species. The purpose of our study was to explore the effects of canal and dam construction on the genetic structure of Mapleleaf (Quadrula quadrula), a widespread and relatively common species in the central Great Lakes that has been found only recently in several western Lake Ontario harbors. Establishment of Q. quadrula in Lake Ontario may have been a recent event, facilitated by the Niagara Peninsula’s history of canal operation. We used analyses of microsatellite DNA genotypes to examine the effect of canals on the genetic structure of mussel populations. Structure analysis revealed a pattern of gene flow between lakes that cannot be explained by watercourse connections prior to the creation of the Welland Canal. Evidence suggestive of historical bottlenecks at some Lake Ontario sites may indicate that these populations became established after canal creation. After considering genetic structure, hydrogeography and isolation-by-distance (IBD) analysis, the first iteration of the canal (1829–1833) is most supported as the configuration that facilitated colonization. However, weak IBD signals across canal models may signify continued gene flow across configurations. Our study demonstrates the connective effect of canals on freshwater mussel populations and has the potential to improve conservation strategies for this and other unionid species at risk.

Wabash Pigtoe, Fusconaia flava, and the related Round Pigtoe, Pleurobema sintoxia, are freshwater mussels (Bivalvia: Unionidae: Pleurobemini) native to the Great Lakes region of North America. Fusconaia flava is considered widespread and relatively common while P. sintoxia is considered an imperiled species. These species are similar in shell shape and coloration and have confounded many freshwater malacologists, resulting in frequent misidentifications. We sought to determine if morphometric analyses could be used to reliably distinguish between these species. Two hundred and forty-six specimens were collected from rivers in Michigan and Ontario. For each specimen, a preliminary identification was made, shell measurements and foot color (orange or white) were documented, and photos of the left shell valve were taken. A genetic sample was taken from 133 specimens for cytochrome c oxidase subunit 1 (COI) barcoding. COI sequences were used for species identification by comparing to sequences on GenBank. Twenty-one digitized landmarks along the outline of the left valve were analyzed and compared to the results of the DNA barcoding. Landmark data correctly assigned 99.2% of specimens to their DNA-confirmed species identity, compared to 82.0% accuracy of field identifications and 77.0% accuracy for foot color. The creation of a DNA-confirmed morphometric database will aid freshwater malacologists across the Great Lakes region in differentiating between these species.

Simpsonaias ambigua (Salamander Mussel) is a freshwater mussel of the Family Unionidae endemic to North America, and it is considered endangered across most of its range. This species is unique among the Unionidae, because it uses the salamander, Necturus maculosus (Mudpuppy), as its larval host rather than fish like all other unionids. The overall goal of this study was to obtain baseline data on the current distribution and life history traits of S. ambigua in Ontario, Canada. These data are critical for future recovery efforts to protect, restore, or augment populations of S. ambigua across its range. Conventional survey methods were adapted to target S. ambigua, and an additional method was employed to detect this species: trapping N. maculosus and inspecting for signs of encysted glochidia. Both methods were successful at detecting S. ambigua when they were present. Furthermore, Simpsonaias ambigua’s life history traits were investigated, and more information is now available on the reproductive timing windows related to gonad and glochidia development and the host infestation period, as well as on longevity.