The specific aim of this proposal is to characterize the surface lipids of WHO-reference stocks of species of the protozoan parasite Leishmania, responsible for cutaneous, mucocutaneous and visceral disease of man in many temperature and tropical regions of the world, and to determine the roles of those lipids in the host-parasite relationships of the life-cycle stages.
This aim i s to be realized by (1) in vivo propagation of the amastigote stage in laboratory animals and in vitro in cultures of macrophage-like cells, (2) in vitro propagation of the promastigote stage (characteristic of the vector sand fly) in culture under a variety of environmental stresses chosen to mimic those to which the parasite is likely to be exposed (Delta T, Delta pH, Delta osmolarity, Delta 02 and CO2), (3) isolation, collection and purification of amastigotes from macrophages by density gradient centrifugation and lectin agglutination techniques, (4) isolation, collection and purification of plasma membranes of leishmanias and macrophages by fractionation (N2 cavitation, vesiculation, polyacrylamide bead complexing) and centrifugation methods, (5) analysis of the lipid constituents of parasites and their surface membranes, in detail, by chromatographic, chemical degradative and spectrometric means (GC-MS, PNMR, IR, UV), and (6) assessment of the antigenicity of surface glycolipids by attempts to raise antibodies to them in rabbits.
