Mycobacterium tuberculosis H37Rv (Mtb) excludes phagocyte oxidase (phox) and inducible nitric oxide synthase (iNOS) while preventing lysosomal fusion in macrophages (MΦs). The antigen 85A deficient (Δ fbpA) mutant of Mtb was vaccinogenic in mice and the mechanisms of attenuation were compared with MΦs infected with H37Rv and BCG. Δ fbpA contained reduced amounts of trehalose 6, 6, dimycolate and induced minimal levels of SOCS-1 in MΦs. Blockade of oxidants enhanced the growth of Δ fbpA in MΦs that correlated with increased colocalization with phox and iNOS. Green fluorescent protein-expressing strains within MΦs or purified phagosomes were analysed for endosomal traffick with immunofluorescence and Western blot. Δ fbpA phagosomes were enriched for rab 5, rab 11, LAMP-1 and Hck suggesting enhanced fusion with early, recycling and late endosomes in MΦs compared with BCG or H37Rv. Δ fbpA phagosomes were thus more mature than H37Rv or BCG although, they failed to acquire rab7 and CD63 preventing lysosomal fusion. Finally, Δ fbpA infected MΦs and dendritic cells (DCs) showed an enhanced MHC-II and CD1d expression and primed immune T cells to release more IFN-γ compared with those infected with BCG and H37Rv. Δ fbpA was thus more immunogenic in MΦs and DCs because of an enhanced susceptibility to oxidants and increased maturation.
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