Vaccination with outer membrane vesicles and the fimbrial protein FlfA offers improved protection against lesions following challenge with Gallibacterium anatis

Gallibacterium anatis is an opportunistic poultry pathogen belonging to the Pasteurellaceae family. It has been shown to cause oophoritis, salpingitis and peritonitis in hens, as well as being associated with reduced semen quality in cockerels. Widespread multidrug resistance and substantial antigenic variation among strains of Gallibacterium anatis is a major constraint to treatment with antimicrobials and prevention of infection by vaccination. Novel vaccine strategies targeting G. anatis are therefore necessary. Outer membrane vesicles (OMVs) are nanosized vesicles formed from the outer membrane of Gram-negative bacteria. These vesicles have shown promising potential as both adjuvants and as vaccine candidates against numerous bacterial species. A high vesiculating mutant of G. anatis (G. anatis ΔtolR) has previously been made, enabling production of OMVs in large scale. In this study, we elucidated the potential of G. anatis ΔtolR OMVs as adjuvant for the conserved antigens GtxA-N (the N-terminal part of the RTX like toxin Gallibacterium toxin A) and FlfA (F17-like fimbria), as well as evaluated if combinations of OMVs together with antigens could facilitate cross-protective immunity against three different strains of G. anatis. We showed that ΔtolR OMVs function as an adjuvant for GtxA-N by inducing antigen specific antibody production. However, OMVs in combination with GtxA-N failed to induce protection against lesions after challenge infection. In contrast, vaccination with OMVs in combination with FlfA protected against lesions, especially in the salpinx, caused by two diverse strains of G. anatis, thereby indicating a cross-protective potential. No protection against the third G. anatis strain 7990 could be obtained in any of the experimental settings. In conclusion, ΔtolR OMVs and FlfA could serve as potential future vaccine components againt G. anatis.