T-cell recognition of the 18-kilodalton antigen of Mycobacterium leprae
Infection and Immunity
The 18-kilodalton (kDa) antigen of Mycobacterium leprae was expressed as a fusion protein with a 2-kDa leader peptide and used in proliferation assays with peripheral blood cells. Fifty percent of untreated tuberculoid leprosy patients and 93% of long-term leprosy contacts responded to the recombinant protein in lymphocyte transformation tests. Comparison of the stimulation indices in the two groups showed that the contacts responded more strongly than the tuberculoid leprosy patients. Seventy
... patients. Seventy percent of Mycobacterium bovis BCG-vaccinated European donors responded, although with low stimulation indices. The isolation of 18-kDa antigen-responsive T-cell lines from a BCG-vaccinated British donor confirmed that the 18-kDa antigen contains at least one cross-reactive epitope. These results indicate that the 18-kDa protein is an important antigen in the immune response to leprosy. Evaluation of the contribution of individual leprosy antigens to immunity is now possible through the cloning and expression of Mycobacterium leprae genes in bacteriophage lambda gtll (25). T cells recognizing antigens of 65 (11, 22), 36 (9, 17, 22) and 18 (13) kilodaltons (kDa) have been described. The 18-kDa antigen is of particular interest, as a high proportion of M. leprae-specific T-cell clones from donors immunized with a killed M. leprae vaccine recognized this protein (13). Crude lysates of Escherichia coli expressing recombinant proteins have been used to screen T-cell clones for reactivity (10, 13) but are unsuitable for use with uncloned peripheral blood cells. In addition, P-galactosidase fusion proteins cannot be used with uncloned cells, as there is a variable human T-cell response to the P-galactosidase portion of the molecule (21a). We have therefore recloned the gene for the 18-kDa protein into a plasmid vector so that it is fused to a 2-kDa peptide containing a run of six asparagine residues to reduce proteolytic degradation (Asn-18K). We report here that a high proportion of leprosy contacts, presumed to be immune to M. leprae, showed lymphocyte proliferation to the Asn-18K antigen; a lower proportion of untreated tuberculoid leprosy patients responded. We also show that the 18-kDa protein contains at least one epitope which is not specific to M. leprae. MATERIALS AND METHODS Patients and controls. Long-term leprosy contacts were men and women who had worked at the Marie Adelaide Leprosy Centre, Karachi, Pakistan, for between 2 and 20 years and had developed no signs of clinical leprosy. Tuberculoid leprosy patients were new, untreated patients attending the Marie Adelaide Leprosy Centre who were diagnosed clinically, by bacterial index counted on slit-skin smears and, in the majority of cases, histologically by punch-skin biopsy taken from the edge of an active lesion. European Mycobacterium bovis BCG-vaccinated donors were laboratory personnel in London.