Functional Analysis of Polydnavirus and Insect Immune Cell Genes

The primary immune responses toward parasites are encapsulation and phagocytosis. Larger, multicellular parasites are usually killed by encapsulation which involves attachment of multiple layers of hemocytes to the foreign target. Smaller parasites, in contrast, are ingested by individual hemocytes during phagocytosis. Despite the fundamental importance of these responses to insects, our understanding of their regulation and the counter strategies parasites use to evade the host immune system is limited. Parasitoid wasps and flies are among the most important natural enemies of insect agricultural pests. Parasitoids are usually killed in nonpermissive hosts by encapsulation, whereas parasitoids survive in permissive hosts by either possessing surface features that passively protect the parasitoid from being encapsulated or by immunosuppressing the host's encapsulation response. Immunosuppression is mediated by symbiotic polydnaviruses (PDVs) in many species of parasitoids in the families Braconidae and Ichneumonidae. PDV s do not replicate in the wasp's host but PDV s play an essential role in suppressing the ability of hosts to encapsulate the parasitoid. The viral genes responsible for immunosuppression and the mechanism(s) by which they disrupt capsule formation, however, have until recently been poorly understood. In this proposal, we seek funds to functionally characterize the immune suppressive genes of Microplitis demo/itor bracovirus (MdBV) and Campo/etis sonorensis ichnovirus (CsIV). Our prior results provide considerable information on the regulation of insect cellular immune responses and the immunosuppressive effects caused by MdBV and CsIV. Insight into the molecular mechanisms underlying PDV-mediated immunosuppression also derives from our recent genomic and functional studies. We have sequenced the CsIV genome and are in the finishing phase for MdBV (>90% complete). The CsIV and MdBV genomes share several organizational features that include segmentation, increased copy number of selected DNA segments and genes, and divergence of virulence genes into multiple variants that together form gene families. Yet remarkably, CsIV and MdBV share no sequence homology with one another. Based on these results, we are now positioned to undertake a systematic functional analysis of PDV -insect immune interactions. Specific objectives of this proposal are to: 1) Perform a genome wide analysis of MdBV and CsIV immune regulatory genes by RNA interference (RNAi ) and gain of function experimentation, and 2) Conduct a transcriptional and proteomic analysis of host gene expression in PDV -infected immune cells. This work will be the first truly functional genomics analysis of insect pathogens and their interactions with the insect cellular immune system.