During 2009–2010, we qualitatively surveyed new and existing locations in the James River Basin, Missouri, to update the distribution and status of the freshwater mussel fauna and determine if changes had occurred over a 25-year period. We calculated a suite of metrics to measure richness, diversity, and community composition. We observed significantly fewer live taxa in the James River mainstem and significantly fewer live individuals in Finley Creek than in a 1982–1984 survey effort, including a nearly complete collapse of the mussel fauna in Finley Creek. Basin-wide and across species, values for probability of extirpation were significantly higher than values for probability of colonization, and all tribes had higher probabilities of local extirpation than colonization. Even locations in the James River Basin with relatively high species richness represented low diversity. In the James River mainstem, declines in diversity and abundance were seen throughout the river, but especially below the confluence with Wilsons Creek.