Mapping of T cell epitopes of the 30-kDa α antigen of Mycobacterium bovis strain bacillus calmette-guérin in purified protein derivative (PPD)-positive individuals

The fibronectin-binding 30-kDa α Ag is a major secretory protein of growing mycobacteria that stimulates in vitro lymphocyte blastogenesis in most healthy purified protein derivative-positive individuals, but only a minority of patients with active tuberculosis. T cell epitopes of the a Ag were assessed using blastogenic responses of PBMC from 12 healthy purified protein derivative-positive subjects to a set of synthetic peptides based on the 325-amino acid sequence of the a Ag of Mycobacterium bovis BCG. Because epitope-specific precursor cells are infrequent and randomly distributed, we used Poisson analysis to determine positive responses to 10 μg/ml of each peptide in 12 replicate culture wells. Seven immunodominant regions of the a Ag were identified. Each subject responded to at least one of the two most dominant epitopes, which correspond to amino acids 131-155 and 233-257 (from N terminus). Peptides of these two epitopes induced production of IFN-γ by sorted CD4⁺ T cells. The immunodominant peptides may have use as components of a vaccine and as tools to study the evolution of the immune response to M. tuberculosis. The two most dominant epitopes both occur in regions of the α Ag that differ from those of the atypical pathogens M. avium and M. kansasii. In addition, the M. bovis epitope of amino acids 133-155 differs from that of M. tuberculosis by a single amino acid. It maY be possible to exploit the sequence differences for development of diagnostic tests with increased specificity.