Knowledge of the habitat suitability of freshwater mussels (family Unionidae) is necessary for effective decision making in conservation and management. We empirically measured microhabitat use for 10 unionid mussel species, including the U.S. federally endangered Alasmidonta heterodon, at 20 sites in the Tar River basin, North Carolina, USA. We also quantified habitat availability at each site, and calculated habitat suitability for each mussel species. The majority of available habitat across all sites consisted of shallow, slow-moving water with penetrable silt or sand substrate. Among species, mean water depth of occupied habitats ranged 0.23 – 0.54 m, mean bottom velocity ranged 0.001 – 0.055 m/s, average mean-column velocity ranged 0 – 0.055 m/s, and mean substrate penetrability ranged 0.11 – 11.67 on an index scale. The most commonly measured dominant substrate materials were silt, sand, very coarse sand, pea gravel, and coarse gravel. The most common cover types were coarse woody debris and fine woody debris. These findings revealed a relationship between the niche breadth and conservation status of four species. Federally endangered A. heterodon consistently showed a narrower suite of suitable microhabitats than the common mussel Elliptio complanata. The range of suitable habitat characteristics for Fusconaia masoni and Villosa constricta, listed as North Carolina (USA) state endangered and special concern, respectively, was typically narrower than those of E. complanata and wider than those of A. heterodon. These habitat suitability criteria and relationships will be useful to guide identification of suitable sites for habitat protection, mussel relocation, or site restoration.