H56 but not H28 vaccination provides post-exposure protection against relapse.

<p>CB6F1 mice (n=12-18) were infected by the aerosol route and at week 6 post-infection (p.i.) treated with antibiotics given in the drinking water <i>ad </i><i>libitum</i>. Mice were vaccinated three times s.c. with CAF01 alone, H28/CAF01 or H56/CAF01 at week 10, 13 and 16 p.i. (A) PBMCs (pooled from 16 mice) were obtained by submandibular bleeding one week after the last vaccination and the frequency and quality of antigen specific CD4 T cell responses were determined by six hour culturing of lung lymphocytes with Ag85B, TB10.4 or ESAT-6 from adjuvant control (black bars), H56/CAF01 (grey bars) and H28/CAF01 (white bars) animals. After culturing, cells were stained for CD4, IFN-γ, IL-2 and TNF-α, analyzed by ICS and subpopulations were divided according to their cytokine profile. Similar patterns of immune responses have been observed in two other experiments. (B) Protection was assessed at week 44 p.i. by culturing lung homogenates (n=12 for test groups and n=18 for control group) for 2-3 weeks at 37°C and enumerating the number of bacteria (expressed as log₁₀). Data displayed are box and whiskers showing median values and minimum and maximum CFU levels. Values marked with an asterisk are significantly different from naïve control animals as assessed by non-parametric Kruskall-Wallis multiple-comparison test (**, <i>p</i><0.01, *, <i>p</i><0.05). Data is representative of two independent experiments, although individual H28 and H56 vaccine evaluations have been done in several other experiments as illustrated in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0080579#pone-0080579-g006" target="_blank">Figure 6</a>.