Figure 4. ΔznuA B. melitensis and RB51 vaccines stimulate enhanced IFN-γ and IL-22 production by BALB/c CD8⁺ T cells and IL-17 production by IFN-γ^−/− CD4⁺ T cells after challenge with wild-type B. melitensis 16M.

(A, C, E, G, I, K) BALB/c and (B, D, F, H, J, L) IFN-γ^−/− mice (18/group) were nasally vaccinated with 10⁹ CFUs of ΔznuA B. melitensis, RB51, or sPBS and nasally challenged six weeks later with 5 × 10⁴ CFUs of wild-type B. melitensis 16M. Four weeks post-challenge, CD4⁺ and CD8⁺ T cells were isolated (pooled from 2–3 mice/culture and at least three cultures/experiment) and cultured in the presence of syngenic mitomycin C-treated Ag-presenting cells. T cells were restimulated with media or 10⁹ CFUs of HKRB51 for 3 days, and cell culture supernatants were evaluated for (A–D) IFN-γ, (E–H) IL-17, and (I–L) IL-22 production by cytokine-specific ELISAs. Results depict the mean ± SEM of triplicate cultures from two independent experiments; ND, not detected. Significant differences in IFN-γ, IL-17, and IL-22 production were determined: *P≤0.001, **P<0.05 (versus PBS-dosed mice); ^†P≤0.001, P<0.05 (differences between CD4⁺ and CD8⁺ T cells for the same vaccine group); and ^‡P≤0.001, ^‡‡P<0.05 (differences between RB51 and ΔznuA B. melitensis-vaccinated mice); ^# P≤0.001, ^## P<0.05 (differences between BALB/c and IFN-γ^−/− mice to the same vaccine).