Resistance to the murine malarial parasite Plasmodium chabaudi adami has been shown to be mediated by antibody-independent mechanisms of immunity. In addition, T lymphocytes have been shown to adoptively transfer this immunity. This provides a model system in which to investigate the plasmodial antigens which activate these cell-mediated immune mechanisms, an approach which may provide new insights into the prophylaxis and treatment of malaria. We propose to fractionate P. chabaudi adami antigens by biochemical techniques and to test the activity of these fractions in a number of assays which reflect cell-mediated immunity. These assays include: the capacity of the antigen fractions to stimulate cell division in IL2-dependent cells which have been shown to adoptively protect mice against challenge infection; the elicitation of delayed-type hypersensitivity skin reactions in mice which have previously been sensitized with this parasite; and the ability to actively immunize naive mice against a P. chabaudi adami challenge infection. Those antigen fractions having activity in these assays will be characterized for their plasmodial polypeptide content. In addition, we will prepare both polyclonal and monoclonal antibodies against these active fractions. The antisera obtained as a result will be used to screen expression libraries of P. chabaudi adami genes. Three different libraries will be constructed using Lambdagtll as the bacteriophage vector: a genomic library constructed with randomly cleaved DNA; a genomic library of DNA digested with mung bean nuclease; and a cDNA library synthesized from poly A+ RNA derived from the parasite. These three expression libraries will be screened with the antisera, and positive clones encoding these epitopes will be expanded. We will then study the nature of these genes and determine whether polypeptides synthesized by the recombinant bacteria can elicit cell-mediated immunity and immunize mice against a P. chabaudi adami challenge.
