Visceral leishmaniasis, caused by the protozoan Leishmania donovani, is a significant cause of mortality in many areas of the world. Cellular immune mechanisms are critical for recovery from leishmaniasis in both humans and mice, and for the development of long-term immunity against reinfection. Ironically, cellular immune mechanisms can also cause an exacerbation of leishmaniasis in mouse models, and in some forms of human disease. Thus the balance between protective and exacerbating immune T cells may be critical in determining the outcome of disease. The purpose of this project is to identify parasite antigens that are responsible for the development of cellular immune responses during infection with the South American strain of L. donovani chagasi (Ldc). Leishmania survive in mammalian macrophages, but macrophages can be activated to kill intracellular parasites by a subset of parasite antigen- specific T lymphocytes. These lymphocytes are likely to be directed toward antigens present in the amastigote stage of the organism, since this is the form present in a leishmania-infected host. In murine models, a correlation has been found between protection against L. donovani and the secretion of interferon-gamma by lymphocytes. Clinical studies suggest a prominent role for interferon-gamma in recovery from human disease as well. Therefore, we would like to define the amastigote antigens that stimulate interferon-gamma production by immune T lymphocytes. Since amastigotes are difficult to obtain in purified form we have constructed a cDNA library from Ldc amastigote RNA. Expressed products of this library will be studied for their abilities to elicit the secretion of interferon-gamma by T cells from mice immunized against Ldc. The library will initially be immunoscreened with antisera that recognize a large proportion of parasite proteins, to identify fusion protein-producing clones. These recombinant proteins will then be tested for their abilities to stimulate the secretion of interferon-gamma by immune T lymphocytes. Finally, the effects of immunization with these proteins on the subsequent course of Ldc infection will be tested. The project represents a systematic initial approach to identifying recombinant T cell antigens in Ldc amastigotes, that might be important in determining the outcome of visceral leishmaniasis.
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