Targeting resident memory T cell immunity culminates in pulmonary and systemic protection against Brucella infection.

Brucellosis remains the most common zoonotic disease globally. Currently no vaccines for humans exist, and conventional brucellosis vaccines for livestock fail to confer complete protection; hence, an improved vaccine is needed. Although Brucella infections primarily occur following a mucosal exposure, vaccines are administered parenterally. Few studies have considered mucosal vaccinations, or even targeting of tissue-resident memory T (T-RM) cells. T-RM cells protect against viral infections, but less is known of their role in bacterial infections, and even less for brucellosis. Oral prime, nasal boost with a newly developed Brucella abortus double mutant (znBAZ) confers nearly complete protection against pulmonary challenge with wild-type (wt) B. abortus 2308, and its protective efficacy is >2800-fold better than the RB51 vaccine. Vaccination with znBAZ potently stimulated CD8(+) T cells, whereas mucosal vaccination with RB51 induced mostly CD4(+) T cells. Subsequent analysis revealed these pulmonary CD44(+) CD69(+) CD8(+) T cells to be either CD103(+) or CD103(-) T-RM cells, and these sequestered to the lung parenchyma as CXCR3(lo) and to the airways as CXCR3(hi). Both CD8(+) T-RM subsets contained single-positive IFN-gamma and TNF-alpha, as well as, polyfunctional cells. IL-17-producing CD8(+) T-RM cells were also induced by znBAZ vaccination, but in vivo IL-17 neutralization had no impact upon protection. In vivo depletion of CD4(+) T cells had no impact upon protection in znBAZ-vaccinated mice. In contrast, CD4(+) T cell depletion reduced RB51's protective efficacy in spleens and lungs by two- and three-logs, respectively. Although anti-CD8 mAb-treated znBAZ-vaccinated mice showed a significantly reduced pulmonary efficacy, this treatment failed to completely deplete the lung CD8(+) T cells, leaving the CD103(+) and CD103(-) CD8(+) T-RM cell ratios intact. Only znBAZ-vaccinated CD8(-/-) mice were fully sensitive to pulmonary challenge with virulent wt B. abortus 2308 since CD8(+) T-RM cells could not be induced. Collectively, these data demonstrate the key role of mucosal vaccination for the generation of CD8(+) T-RM cells in protecting against pulmonary challenge with virulent B. abortus. Author summary Brucellosis is the most common zoonotic disease worldwide and is transmitted via the consumption of unpasteurized dairy products or exposure to Brucella-laden aerosols. In fact, mucosal exposure is the most common route of infection for humans and animals, yet parenteral vaccination of livestock remains the preferred route of immunization. To determine whether development of a mucosal vaccination regimen could effectively generate immunity against pulmonary challenge with virulent B. abortus, a double-mutant B. abortus vaccine was administered mucosally, and found to induce CD8(+) T-RM cells. These conferred complete protection against pulmonary infection and prevented systemic brucellae spread even in the absence of immune recirculating CD8(+) T cells. These data show that mucosal vaccination can stimulate the induction of T-RM cells, which should be considered as a more effective means to protect against brucellosis. Furthermore, brucellosis needs to be considered a mucosal pathogen to warrant development of approaches different from conventional methods to protect humans and livestock against this disease.