2008 Annual Report 1a.Objectives (from AD-416) To utilize the recently obtained and annotated Babesia bovis genome sequence and functional genomic approaches for the identification of virulence factors and defined vaccine development. 1b.Approach (from AD-416) The genome sequence of the B. bovis virulent strain T2Bo has been sequenced and annotated and we are currently devising stable transfection methods for B. bovis. Our experimental approach towards vaccine development will be based initially in utilizing genome sequences from several B. bovis virulent and attenuated strains to identify genes associated with virulence, cerebral babesiosis and tick transmission. Then, the functional role of identified virulence-associated and transmission- associated genes will be further defined using Babesia transfection technology either by knocking out or complementing the genes of interest. Comparative studies of the phenotypes of the wild type and knock-out transfected strains will be performed. These will include their ability to produce acute and chronic disease, cerebral babesiosis, and tick transmission. The goal will be to obtain genetically defined attenuated and non tick-transmissible parasite strains that can be used in the field to prevent B. bovis caused bovine babesiosis. In addition, functional gene characterization through transfection will aid in the identification of protective antigens that can aid the design of subunit vaccines and the expression of foreign genes, such as tick antigens, in Babesia vaccine strains that may lead to developing dual anti tick-parasite vaccines. Documents SCA with WSU. 3.Progress Report Through use of comparative genomics and the use of syntenic analyses the positional homolog of T. parva p67 was recently discovered in the genome of b. bovis by studying syntenic blocks between the genomes of T. parva and B. bovis. The B. bovis homolog encodes a 517 amino acid protein which shares only 10.6% amino acid identify with T. parva p67. We’ve hypothesized that the immune and functional selection pressure has driven divergence of the B. bovis homolog from that of p67 and supports the testing of B. bovis p67 for its ability to induce protective immune responses. The progress of this subordinate project is monitored through weekly meetings with WSU collaborators.