Fig. 2.

Molecular characterization of MTBVAC, MTBVAC-L2 and MTBVAC-L3. (a) Expression of representative genes from the PhoP-regulon (mcr7, pks2 and pks3) measured by qRT-PCR in MTBVAC, MTBVAC-L2 and MTBVAC-L3 compared to their respective parental strains. Each gene was normalized against sigA expression in each sample. Bars represent the mean and standard deviation from three independent experiments. (b) Western-blot of ESAT-6 and CFP-10 proteins in whole-cell lysates (left panel) and secreted fractions (right panel) in the MTBVAC vaccines and their wild type strains. Note the absence of ESAT-6 secretion as a consequence of the phoP mutation in all vaccine strains. Also note the presence of CFP-10 in the secreted fraction in all MTBVAC strains. Detection of SigA and SigB serves as loading control in the whole-cell lysate and also as cell integrity control in the secreted fraction. (c) Western-blot of PE_PGRS in total (left panels) and secreted fractions (right panel) from the MTBVAC vaccine set. Note the differential absence of PE_PGRS secretion in MTBVAC-L2. (d) Expression of the ppsAB genes belonging to the PDIM biosynthetic operon measured by qRT-PCR. Bars represent the mean and standard deviation from three independent experiments. Each sample was normalized relative to the endogenous control sigA.