IMMUNODOMINANT ANTIGENS OF MYCOBACTERIUM TUBERCULOSIS

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Tuberculosis causes more deaths world-wide than any other infectious pathogen, and its incidence is increasing in the United States because HIV-infected patients are extraordinarily susceptible to tuberculosis. Eradication of tuberculosis requires development of an effective vaccine, which in turn hinges on understanding the mechanism of protective immunity against tuberculosis. The spectrum of manifestations of tuberculosis infection reflects the cell-mediated immune response, and provides an excellent model to characterize the immunologic correlates of protective immunity, delayed-type hypersensitivity (DTH) and ineffective immunity to intracellular pathogens. The goals of this proposal are to characterize T cell subpopulations that mediate protective immunity against tuberculosis and to identify mycobacterial antigens that elicit protective immunity. These antigens are potential candidates for an antituberculosis vaccine and for diagnostic skin test reagents. Our specific aims are: 1) Identification of T cell subsets that mediate resistant and protective immune responses to Mycobacterium tuberculosis. Dominant alpha beta T cell antigen receptor subtypes in tissue and blood of patients throughout the spectrum of tuberculosis infection will be identified by immunolabeling with monoclonal reagents, combined with PCR amplification of alpha beta T cell receptor genes. 2) Identification of purified and recombinant mycobacterial antigens that elicit proliferation by alpha beta T cell subpopulations that mediate resistant and protective immune responses. Antigens will be screened for immunoreactivity by T cell lines, and epitopes defined by testing T cell clones against synthetic peptides. Immunodominant antigens will be identified by limiting dilution analysis and by evaluating immunoreactivity of antigen across multiple HLA types. 3) Investigation of the capacity of immunodominant antigens to induce a functional response by characterization of the lymphokine profile of stimulated T-lymphocytes from patients across the spectrum of tuberculosis infection. Lymphokine concentrations will be measured directly in cell supernatants, and lymphokine mRNA detected by PCR amplification. 4) Evaluation of immunodominant M. tuberculosis-specific antigens as diagnostic skin test reagents to detect patients with tuberculosis infection. Skin testing with these antigens in tuberculin- positive and tuberculin-negative individuals, combined with measurement of lymphocyte proliferation, will allow assessment of the relationship between immunoreactivity in vitro and induction of DTH in vivo.