The drastic decline of the eastern oyster (Crassostrea virginica) in Chesapeake Bay and other estuarine areas along the Atlantic coast of the United States has prompted officials to consider the possibility of introducing the Suminoe oyster Crassostrea ariakensis into these waters. However, the introduction of an exotic oyster species may have unforeseen and potentially harmful effects on the Bay's ecosystem. An important question to be addressed is the potential for the nonnative oyster to become a reservoir for human, fish, and shellfish bacterial pathogens. The purpose of this study is to establish and optimize methodology, and carry out a preliminary screen of C. ariakensis and C. virginica from Chesapeake Bay for the presence of Vibrio parahaemolyticus, V. vulnificus, mycobacteria, and enterobacteria, using cultivation on semiselective media and PCR. In spite of the high prevalence of Vibrio spp. in soft tissues and the shell surface of both oyster species, V. parahaemolyticus and V. vulnificus were absent. Coliform enterobacteria were detected in tissues from both oyster species by cultivation in a semiselective medium. In contrast, mycobacteria were not detected in oyster soft tissues by both cultivation and PCR, but they were abundant on the shell surface. Although oysters have been reported to harbor V. cholerae and other Vibrio species as shell biofilm-associated bacteria, this report provides the first direct evidence for the presence of mycobacteria in oyster shell-associated biofilms. Further, this preliminary study revealed no substantial differences between C. virginica from coastal Maine and C. ariakensis from Chesapeake Bay waters concerning selected components of their associated microbial flora.