More robust gut immune responses induced by combining intranasal and sublingual routes for prime-boost immunization.

Norovirus causes acute and debilitating gastroenteritis, characterized by vomiting and diarrhea. We recently reported a recombinant GII. 4 P domain particle (Pd) vaccine adjuvanted with a flagellin, Vibrio vulnificus FlaB, effectively promoting both humoral and cell-mediated immune responses. In the previous study, we found that sublingual (SL) immunization induced higher fecal secretory IgA (SIgA) responses while intranasal (IN) route provided higher amplitude of humoral and cellular immune responses in the systemic compartment. We hypothesized that the combination of IN and SL routes should induce more potent and sustained SIgA responses in the gut. In this study, we have tried combinatorial prime-boost immunization employing both IN and SL routes. The IN priming and SL boosting with the Pd+FlaB vaccine enhanced highest SIgA responses in feces, accompanying increased Pd-specific memory B cells and plasma cells in spleen and bone marrow, respectively. Notably, the strongest long-lasting SIgA response in feces was induced by combined IN prime and SL boost vaccination, which was sustained for more than 3 months. Significantly enhanced gut-homing B cell and follicular helper T cell responses in mesenteric lymph nodes (mLNs) were observed in the IN prime and SL boost combination. IN priming was a requisite for the robust induction of Pd-specific IFNγ, IL-2, IL-4 and IL-5 cytokine responses in the systemic immune compartment. Collectively, the IN prime and SL boost combination was the best option for inducing balanced long-lasting immune responses against the norovirus antigen in both enteric and systemic compartments. These results suggest that immune responses in specific mucosal compartments may be programmed by employing different prime-boost immunization routes.