This proposal seeks to (1) characterize Borrelia burgdorferi genes that are selectively expressed in the tick and then (2) investigate the role of the corresponding antigens, and the immune response to them, in spirochete transmission. B. burgdorferi alters its surface proteins in both Ixodes scapularis and the mammalian host. OspA is synthesized by B. burgdorferi within the gut of flat ticks but downregulated as spirochetes migrate to the salivary glands in engorging ticks. In contrast, OspC is not present on B. burgdorferi within flat ticks but is made by spirochetes within feeding ticks. During vertebrate infection, p35, p37, ospE/F homologues, among other genes, are then expressed. In this proposal, an immunologic screening strategy will be used to systematically identify B. burgdorferi genes that are expressed within the arthropod vector. A B. burgdorferi genomic DNA expression library will be probed with hyperimmune sera generated against B. burgdorferi that have been directly isolated from ticks - this hyperimmune sera will therefore contain antibodies to spirochete antigens synthesized within the vector. Specific intervals in the B. burgdorferi life cycle within the arthropod including spirochetes within flat or engorging nymphal ticks, and B. burgdorferi entering larval ticks from the mammalian host will be examined. Direct screening of the B. burgdorferi within ticks will be used to complement the immunologic screening strategy. The temporal expression of these B. burgdorferi genes within the vector will be assessed using RNA-PCR and IFA. We will then determine whether spirochete gene expression within ticks is dependent, at least in part, upon the density of B. burgdorferi - this postulate is supported by the preliminary data. The role of antibodies to the antigens encoded by B. burgdorferi genes expressed within the vector in the prevention of infection will then be investigated. We will determine whether these antibodies can prevent (a) the transmission of B. burgdorferi from tick to the mammalian host, (b) the larval acquisition of spirochetes, or (c) the ability to B. burgdorferi to survive within the vector during the molting process. We will also determine whether OspA, which is preferentially expressed within flat ticks, is involved in binding of B. burgdorferi to the tick gut. This project lead to a greater understanding of differential B. burgdorferi gene expression in ticks, spirochete transmission from the vector, and the role of antibodies in the destruction of B. burgdorferi throughout its complete lifecycle in the vector.