Virulence, Immunogenicity, and Protective Efficacy of Two RecombinantMycobacterium bovisBacillus Calmette-Guérin Strains Expressing the Antigen ESAT-6 fromMycobacterium tuberculosis

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Abstract
We constructed two recombinantMycobacterium bovisBCG (rBCG) strains expressing ESAT-6 ofMycobacterium tuberculosis, named rBCG-1 and rBCG-2. rBCG-1 contained the ESAT-6 gene linked to BCGhsp60and expressed a fusion protein, while rBCG-2, with a secretory sequence, could secret ESAT-6 into the culture medium. There was no evidence for increased virulence of the two rBCG strains when we made a comparison between them and BCG with regard to organ bacterial loads, lung histology, and survival time. rBCG-1 induced significantly higher specific antibody titers and stronger cellular immune response than BCG, whereas rBCG-2 had immunogenicity similar to that of the parental BCG strain. Both rBCG-1 and rBCG-2 conferred marked protection againstM. tuberculosisinfection, yet in terms of protective efficacy, they showed no significant improvements upon conventional BCG vaccine.