Leishmania parasite causes human disease (Leishmaniasis) with clinical symptoms ranging from self healing cutaneous lesions to fatal visceral infection. The lack of accurate tools for clinical diagnosis and of a vaccine for leishmaniasis poses a serious health risk for U. S. military personnel, their families and tourists, either living or travelling in endemic areas. In addition, people infected with HIV are particularly prone to be infected by the leishmania parasite. Macrophages are the targets for both type of infectious agents, therefore, it raises the possibility that one infection increases vulnerability to other or accelerating the progression towards full blown disease. Study of modulation of cytokines by the infected macrophages could provide insight in the pathogenesis of parasitic infection. To answer such questions we need first to identifying parasite genes which could be used as potential vaccine in people at risk of being infected or for patients suffering from the disease. We have developed an in vitro culture system for Leishmania donovani. It allows the transformation of promastigotes to amastigotes and vice versa without going through the sand fly and human macrophage interaction. Parasites grown under such conditions are similar to those isolated from in vivo sources. Using this system, we have (a) identified several parasite genes whose expression is regulated during parasite differentiation, (b) some of the proteins synthesized by these genes elicit antibody responses in patients suffering from leishmaniasis, and (c) expressed cloned parasite genes to develop antibodies which can be used for diagnostic purposes, (d) have knocked out genes in parasites by homologous recombination to study the effect of such mutations on the pathogenesis of parasites. We have also developed arbitrary primed-PCR based detection assay for Leishmania parasite. This test allows to identify the geographically distinct isolates of same species as well as different species . Further, the randomly amplified DNA fragments which are generated by this technique, can also be used to identify genes of the parasite with ease. Currently, detailed analysis of some of the genes is being carried out, with an understanding that it can provide some insight in parasite differentiation and development. Further, studies will be initiated to study the effect of cytokines such as IL-2 and IL-12 which are thought to be necessary for the progression of Leishmaniasis.
