Nasal vaccination stimulates CD8+ T cells for potent protection against mucosal Brucella melitensis challenge

Brucellosis remains a significant zoonotic threat worldwide. Humans and animals acquire infection via their oropharynx and upper respiratory tract following oral or aerosol exposure. After mucosal infection, brucellosis develops into a systemic disease. Mucosal vaccination could offer a viable alternative to conventional injection practices to deter disease. Using a nasal vaccination approach, the Delta znuA B. melitensis was found to confer potent protection against pulmonary Brucella challenge, and reduce colonization of spleens and lungs by more than 2500-fold, with >50% of vaccinated mice showing no detectable brucellae. Furthermore, 10-fold more brucellae-specific, interferon-gamma (IFN-gamma)-producing CD8(+) T cells than CD4(+) T cells were induced in the spleen and respiratory lymph nodes. Evaluation of pulmonary and splenic CD8(+) T cells from mice vaccinated with Delta znuA B. melitensis revealed that these expressed an activated effector memory (CD44(hi)CD62L(Io)CCR7(Io)) T cells producing elevated levels of IFN-gamma, tumor necrosis factor-alpha, perforin and granzyme B. To assess the relative importance of these increased numbers of CD8(+) T cells, CD8(-/-) mice were challenged with virulent B. melitensis, and they showed markedly increased bacterial loads in organs in contrast to similarly challenged CD4(-/-) mice. Only Delta znuA B. melitensis- and Rev-1-vaccinated CD4(-/-) and wild-type mice, not CD8(-/-) mice, were completely protected against Brucella challenge. Determination of cytokines responsible for conferring protection showed the relative importance of IFN-gamma, but not interleukin-17 (IL-17). Unlike wild-type (wt) mice, IL-17 was greatly induced in IFN-gamma(-/-) mice, but IL-17 could not substitute for IFN-gamma's protection, although an increase in brucellae dissemination was observed upon in vivo IL-17 neutralization. These results show that nasal Delta znuA B. melitensis vaccination represents an attractive means to stimulate systemic and mucosal immune protection via CD8(+) T-cell engagement.