Distinct differences in repertoires of low-molecular-mass secreted antigens of Mycobacterium avium complex and Mycobacterium tuberculosis

Antigens in a 4-week-old culture filtrate (CF) of Mycobacterium avium subsp , avium were separated in sodium dodecyl sulfate-polyacrylamide gel electro phoresis and identified by Western blotting. The culture had minimal lysis of bacilli, giving a CF preparation consisting mainly of secreted proteins. Comparison with a similar CF of Mycobacterium tuberculosis with almost no contamination with intracellular proteins showed the presence of cross-reac tive antigens homologous to the four components of the antigen 85 complex, as well as MPT32. These were major constituents of the M. avium subsp, aviu m CF, In addition, there were several low-molecular-mass bands (<15 kDa) in both species that did not cross-react with polyclonal and polyvalent rabbi t antibodies in Western blotting. Furthermore, these bands were not detecte d in corresponding sonicate preparations, indicating high localization inde xes, which is typical of soluble secreted proteins. A 14-kDa protein was se lected for purification and more detailed characterization. The N-terminal amino acid sequence was determined, and a matching gene was found within th e genomic sequence of M avium subsp. avium which was highly homologous to R v0455c of M. tuberculosis, The gene encoded a signal peptide typical of sec reted mycobacterial proteins. A rabbit antiserum was raised against the pur ified protein, and the antigen was demonstrated by Western blotting in CFs of M. avium subsp. avium, Mycobacterium avium subsp, paratuberculosis, Myco bacterium intracellulare, and Mycobacterium scrofulaceum but was not detect ed in M tuberculosis. This is a new example of a highly homologous gene bei ng differentially expressed by different mycobacterial species.