A recombinant DNA strategy has been used systematically to survey the Mycobacterium tuberculosis genome for sequences that encode specific antigens detected by monoclonal antibodies. M. tuberculosis genomic DNA fragments with randomly generated endpoints were used to construct a large Xgtll recombinant DNA expression library. Sufficient numbers of recombinants were produced to contain inserts whose endpoints occur at nearly every base pair in the pathogen genome. Protein antigens specified by linear segments of pathogen DNA and produced by the recombinant phage of Escherichia coli were screened with monoclonal antibody probes. This approach was coupled with an improved detection method for gene isolation using antibodies to clonally isolate DNA sequences that specify polypeptide components of M. tuberculosis. The methodology described here, which is applicable to other pathogens, offers possibilities for the development of more sensitive and specific immunodiagnostic and seroepidemiological tests for tuberculosis and, ultimately, for the development of more effective vaccines.

African trypanosomiasis is accompanied by a profound general immunosuppression in which both suppressive T cells and macrophages (M phi) have been implicated. The present studies define changes in the M phi surface, endocytic and secretory properties, during the infection of mice by Trypanosoma brucei. Peritoneal M phi obtained after the control of the first wave of parasitemia displayed characteristics similar to those activated by intracellular pathogens, such as Mycobacterium bovis bacillus Calmette-Guérin, e.g., the enhanced expression of Ia antigen, decreased M phi-specific antigens, receptors mediating the pinocytosis of mannose-terminal glycoproteins, and an increased ability to secrete plasminogen activator, superoxide anion, and H2O2. Some markers of macrophage activation persisted during the subpatent period before the recurrence of parasitemia, whereas others reverted to normal. Mature T cell function appears not to be essential for M phi activation by T. brucei since the infection of athymic nude mice also induced Ia antigens and plasminogen activator. These studies show that M phi activated by different pathways express common features which may contribute to immune dysfunction observed in trypanosomiasis, as well as in other infections.

African trypanosomiasis is accompanied by profound general immunosuppression. The experiments described here were designed to characterize the contribution of macrophages to the immune pathology of this disease. We used peptonestimulated, uninfected mice and injected them intraperitoneally with lethally irradiated and 3S-labeled Trypanosoma brucei and parasite-specific antisera.

DNA hybridization studies of genomic DNA indicated that, while different isolates of armadillo-derived Mycobacterium leprae have a high degree of homology, binding of M. leprae genomic DNA to DNA of other species of mycobacteria or to corynebacteria was low, establishing that M. leprae is only remotely genetically related to any of the species examined. Several candidate leprosy vaccine mycobacterial strains were similarly found to have little genetic similarity to M. Ieprae. In contrast, the DNAs of the slow-growing mycobacteria M. tuberculosis, M. africanum, M. bovis, and M. microti were found to be very closely related. In the course of these studies, an M. keprae-specific repetitive sequence, greater than 15-fold per genome equivalent, was identified that might be useful for diagnostic and epidemiological studies. Percent binding of probe DNA Sample DNA M. leprae M. tuberculosis, Low stringency High stringency high stringency M. leprae grown in armadillos Single inoculum Mixed inoculum Leprosy-associated Corynebacterium strains 2628 L.B. Shep IV Other Corynebacterium strains C. diphtheriae type gravis C. diphtheriae type mitis Corynebacterium sp. strain 813 Candidate leprosy vaccine strains ICRC bacillus Mycobacterium strain w R1-7 FMR bacillus M. tuberculosis complex strains M. tuberculosis (Erdman) M. tuberculosis H37Rv M. bovis (TMC 409) M. bovis BCG (Pasteur) M. africanum (TMC 5122) M. microti (TMC 1601) Other mycobacteria M. avium (TMC 724) M. chelonei (TMC 1544) M. fortuitum (TMC 1529) M. gordonae (TMC 1324) M. intracellulare (TMC 1406) M. kansasii (TMC 1204) "M. Iufu" M. marinum (TMC 1218) M. nonchromojenicum (TMC 1481) M. phlei (TMC 1548) M. scrofulaceum (TMC 1320) M. smegmatis (TMC 1515) M. vaccae (TMC 1526

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