We determined whether immunization decreased lung injury compared to that in unimmunized mice. better than the canine vaccine Bronchicine and provided protection against a strain isolated from a Voglibose dog with kennel cough. Th1/17-polarized immune responses correlate with long-lasting protection against bordetellae and other respiratory pathogens. Notably, BcfA strongly attenuated the Th2 responses elicited by FHA and Prn, resulting in Th1/17-skewed responses in inherently Th2-skewed BALB/c mice. Thus, BcfA functions as both an antigen and an adjuvant, providing protection as a single-component vaccine. BcfA-adjuvanted vaccines may improve the efficacy and durability of vaccines against bordetellae and other pathogens. is an animal pathogen with a wide host range, infecting farm and companion animals (1,C6). It is one of the etiologic agents of kennel cough, or canine infectious respiratory disease (CIRD) (7). is also increasingly isolated from immunocompromised humans, such as those with HIV/AIDS, cancer, or cystic fibrosis. In many of these cases, the infections are linked to exposure to pets with (8,C10). A nasal live attenuated vaccine (11) and a parenteral cellular antigen extract (CAe) vaccine (Bronchicine) (12) against are widely used to minimize kennel cough outbreaks. The CAe formulation replaced more reactogenic whole-cell inactivated vaccines in parallel with the development of acellular pertussis vaccines (aPV). However, a human vaccine against is not available. Although antigens expressed by are present in aPV against the human pathogen (13, 14), these vaccines are only partially effective against (15). While considerable efforts have been devoted to evaluation of the immune response and effectiveness of aPV, there is insufficient research to determine the effectiveness of CAe vaccines. Dogs vaccinated Voglibose with CAe produced serum IgG and IgA and had reduced bacterial burdens compared Voglibose to unvaccinated dogs (16, 17). However, minor vaccine-related side effects were observed, and coughing in 20% of immunized animals was reported, suggesting that the vaccine does not provide complete protection against disease (17). Furthermore, information on the immune response and protective efficacy of these vaccines is limited (12). Thus, there is an urgent need for well-defined, immunogenic acellular vaccines against for veterinary and human use. Together, Th1/17 cellular responses and Th1-skewed antibody responses provide long-lasting protective immunity against bordetellae (18). At present, all aPV are adjuvanted with alum (13, 14), which elicits Th2-skewed cellular and humoral responses with suboptimal and short-lived protection (18, 19). While alum does not cause pyrexia and has the strongest safety record of any adjuvant used Voglibose in human vaccines (20), there have been reports of adverse reactions in animals and humans (21, 22). Thus, development of improved adjuvants is a pressing objective for the more effective control of both veterinary and human diseases. We previously reported identification of colonization factor A (BcfA) (23), an outer membrane protein expressed by but not by the human pathogen (24). BcfA is a paralog of outer membrane protein BipA and has significant homology to intimins and invasins of other bacteria (24). We showed that an experimental vaccine containing BcfA adsorbed to alum elicited protective immune responses against (25). BcfA is also an adjuvant that elicits Th1/Th17 cytokine responses and Th1-type antibodies to protein antigens (26), potentially serving as an alternative adjuvant to alum. In the present study, we tested the efficacy of BcfA as a monovalent vaccine and of BcfA in combination with virulence factors FHA and Prn. We found that Th2-prone BALB/c mice immunized with BcfA as an antigen and without an additional adjuvant elicited Th1/17-polarized responses and efficiently cleared a infection from the lungs and trachea. A combination vaccine containing BcfA and two proteins, FHA and EFNB2 Prn (14), also provided protection against laboratory and canine isolates of for veterinary and human applications. Additionally, BcfA can function as an adjuvant to enhance immune responses against pathogens for which Th1/Th17 immune responses correlate with better protection (27, 28). RESULTS Immunization with BcfA as a single antigen in the absence of another adjuvant reduces colonization of the mouse respiratory tract. We previously reported that immunization with BcfA-alum protected mice against challenge (25). BcfA also enhanced immune responses to heterologous antigens and to vaccine antigens FHA and Prn (26). These results suggested a dual protective function of BcfA as an antigen and an adjuvant. Here, we first tested the hypothesis that BcfA as the sole component would protect against infection in the absence of alum. BALB/c mice (male and female) were immunized intramuscularly (i.m.) with BcfA-alum or BcfA alone (as described in Materials and Methods) and challenged with the prototype laboratory strain RB50 (originally isolated from a rabbit) (29). The numbers of CFU in the lungs and trachea were enumerated at 4?days postinfection (dpi). Both immunizations protected the lungs and trachea of mice but not those of naive unimmunized mice. The bacterial burden was similar in both organs from mice immunized with BcfA-alum or BcfA alone.