Most of the adenylate cyclase (AC) present in Bordetella species is localized in the outer membrane, partly exposed to the cell surface. An isolation procedure to obtain the cell-bound enzyme was applied to Bordetella bronchiseptica, Bordetella pertussis and Bordetella parapertussis. Passive transfer of B. bronchiseptica anti-AC antibody, either in the form of monoclonal antibody or antibody transferred to offspring from female mice previously immunized with immunopurified B. bronchiseptica AC, prevented both death and the development of "atrophic rhinitis" in mice infected by an aerosol of virulent B. bronchiseptica. Immunopurified AC from both B. bronchiseptica and B. pertussis protected mice against intracerebral challenge with strain 18323, although five to six times more protein of B. bronchiseptica was necessary to produce the same level of protection as with B. pertussis-derived protein. The AC of Bordetella species differ in molecular weight and, despite sharing at least two common non-overlapping epitopes (BB05, BB07), antigenic differences are likely: sera from piglets artificially infected with virulent B. bronchiseptica developed high ELISA titers against AC from B. bronchiseptica but low titers against B. pertussis-derived antigen. Strains of B. bronchiseptica which did not induce "atrophic rhinitis" in pigs were found to be AC-less variants. When tested in our laboratory model, such a strain was also unable to induce "atrophic rhinitis" in mice. We conclude that AC is a mandatory component of any future subcellular vaccine against Bordetella species.