Application of gene transformation technologies to the functional analysis of immunomodulation in Plasmodium

Malaria is the most lethal human parasitic disease, causing more than 2 million deaths annually. Efforts to develop effective anti-malarial therapies have been hindered by an incomplete understanding of how parasites have adapted to proliferate so effectively whilst also evading immune responses. The rodent parasite, Plasmodium chabaudi, provides the best available experimental model for addressing these questions but, until now the lack of gene modification technologies for this parasite has precluded direct functional studies on the gene products that are implicated. In a recent pilot study we have established the conditions for Plasmoduim chabaudi genetic modification and have successfully generated the first fluorescent parasites. These and other parasite lines will immediately be an invaluable tool in experiments that will visualise live interactions between parasite and host immune cells and tissues and that will examine host responses to specific parasite proteins in an in vivo setting.

Work on rodent models of malaria has contributed substantially to our understanding of malaria immunology and to the nature of host-parasite interactions that influence parasite virulence and infectiousness. However, Plasmodium chabaudi, the most appropriate model for in vivo studies of malaria disease has, until now, resisted attempts at transfection. We have recently achieved this and are now in a position to apply the power of post-genomic genetic manipulations to this important model. We propose to use this new technology to directly visualise interactions between parasites and immune cells and tissues, and also study individual parasite protein function, so as to test hypotheses such as (i) do parasite genes with structural similarities to mammalian immune genes function in immunomodulation? (ii) how are memory immune responses against malaria initiated? Specifically, we aim to
? Optimize transfection technologies for the generation of genetically-modified P. chabaudi;
? Use the technology to stereoimage parasite-infected tissues.
? Disrupt genes encoding putative parasite immunomodulatory proteins to determine whether these proteins are virulence determinants and whether they promote parasite survival during acute and/or chronic phases.
? Transfect P. chabaudi parasites with mini-genes coding for defined immunogenic peptides to be able to characterize the nature and regulation of T and B cell responses in infection.