Virulent and avirulent Aeromonas salmonicida strains grown inside intraperitoneal implants in Rainbow trout (Oncorhynchus mykiss) were examined for unique antigen expression. Western blots (immunoblots), performed with immune rabbit serum raised against in vivo-grown cells, revealed several unique antigens. With the exception of lipopolysaccharide (LPS), these novel antigens were destroyed after proteinase K treatment. The majority of these antigens were not induced in vitro in response to either iron limitation or anaerobiosis. In addition, electron microscopy demonstrated the presence of a putative capsule on in vivo-grown cells. Purification and fractionation of this carbohydrate material from cells grown in carbon-rich synthetic media resulted in the isolation and separation of an antigenically distinct LPS not seen with cells grown in standard media. Antiserum raised against in vivo-grown cells recognized both this LPS and the typical LPS of A. salmonicida apparent in in vitro-grown cells. Antiserum raised against in vitro-grown cells recognized only the LPS expressed in vitro. Antiserum directed against in vivo-grown cells was approximately 10 times more sensitive than serum directed against in vitro-grown cells in detecting A. salmonicida in infected fish kidney tissue.