Mussel die-offs have been noted in recent years in Pickwick Reservoir, Tennessee River, Alabama. The primary affected species was Fusconaia ebena, but also affected to lesser degrees were Ellipsaria lineolata, Quadrula pustulosa, and Quadrula quadrula. These events were characterized by large numbers of empty shells—fresh-dead and live individuals that were presumed to be diseased because of weak and slow valve closure responses to external stimuli. Anecdotal evidence suggested the possible involvement of an etiological agent, such as a bacterial pathogen. The die-offs have occurred in Pickwick Reservoir (river miles 236–256) in sequential years during the past approximately 10 y. These timeframes have coincided with reduced basin inflows and warmer water temperatures. The majority of the moribund and freshly dead F. ebena were females possibly predisposed to infection and disease from ongoing reproductive activity. Affected and healthy-cohort mussels were collected to characterize the bacterial flora prior to, during, and after a July 2006 die-off, and during a subsequent die-off in September 2008. The numbers of total bacteria from both the 2006 and 2008 die-offs were significantly greater from the diseased specimens. For example, from the September 2008 die-off, the mean count from diseased F. ebena soft tissues was 9.75 × 10⁶ cfu/g, which was more than 100 times greater (P = 0.025) than the mean from healthy cohorts (6.74 × 10⁴ cfu/g). The predominant bacteria from affected F. ebena from July 2006 were Hafnia alvei and Aeromonas sobria, whereas from September 2008 the predominant bacteria were Enterobacter spp., Aeromonas schubertii, Aeromonas veronii bv. veronii, and Aeromonas veronii bv. sobria.