Cytokine production by stimulated splenocytes obtained from mice inoculated with PBS or the vaccine strains. Mice were inoculated with PBS or with B. melitensis Rev 1, B. ovis PNV25dA, and B. ovis PNV22A vaccine strains. Spleen cells were obtained 28 days after vaccine inoculation from five mice per group. Splenocytes were cultured as described in Materials and methods and stimulated with heat-killed B. ovis PA (107 CFU/well) for 72 h. Supernatants were analyzed by ELISA tests specific for IFN-γ (A), TNF-α (B), IL-1α (C), IL-10 (D), and IL-12(p40) (E). Results are expressed as means ± SD (n = 5) of each cytokine amount per well. See strain abbreviations and inoculation doses in legend to Figure 1. Significant differences (PLSD test) between mice inoculated with each vaccine and control mice inoculated with PBS are marked as follows: ***, P ≤ 0.0005; **, P ≤ 0.005; *, P ≤ 0.05. Significant differences between mice inoculated with the B. ovis attenuated strains and mice vaccinated with B. melitensis Rev 1 are represented in brackets following the same rule.