Exploratory Studies of lp17-encoded Genetic Factors Important for Tick Colonization by the Lyme Disease Spirochete

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Project Summary Borrelia burgdorferi is an obligate parasite, and is maintained in nature through a complex cycle involving both a tick and mammalian host. The transition between these two very different host types requires the ability to rapidly adapt through changes in gene expression. B. burgdorferi possesses a segmented genome comprised of a single linear chromosome and upwards of 23 linear and circular plasmids. Previous studies have provided information on a number of genes that may be differentially expressed under conditions intended to mimic that of the vertebrate or arthropod host. However, investigation has only just begun to elucidate the importance of lp17 for colonization of the host and tick vector by B. burgdorferi. Published and preliminary work in our lab has recently shown that a small non-coding RNA and the bbd21 gene encoded on lp17 have roles in the regulation of genes important for host adaptation. Despite these advances, there remains a fundamental gap in our understanding of the role and function of many genes for adaptation to the tick vector. Our long-term goal is to identify and characterize factors necessary for B. burgdorferi adaptation to the tick and mammalian host environments. The overall objective of this application is to identify lp17-resident genes required for colonization of the tick vector by B. burgdorferi. Based on published work and preliminary data, the central hypothesis of this proposal is that a number of lp17-resident genes of B. burgdorferi are important for tick colonization, survival, and/or transmission. The rationale for the proposed research is that these identified factors will represent potential targets for the development of a vaccine and/or therapeutics against human infection by the Lyme disease pathogen, as well as targeting factors critical for its enzootic life cycle. Thus, the proposed research is relevant to that part of NIH’s mission that pertains to developing fundamental knowledge that will potentially help to reduce the burdens of human illness and disability. Guided by cited work and preliminary data, our hypothesis will be tested by pursuing the following specific aim and two sub aims: 1) Determine the requirement of lp17-resident genes for tick acquisition, survival, and/or transmission of Bb. Under the first sub aim, Ixodes scapularis tick larvae will first be infected with either wild type or lp17 mutant B. burgdorferi clones using a novel continuous flow tick feeding system to identify genetic regions of lp17 important for tick acquisition and transstadial survival. Under the second sub aim, nymphal ticks infected with either a lp17 mutant or wild type control will be allowed to feed for a given time using our tick feeding system, after which the bacterial burden in isolated midgut and salivary glands will be assessed via immunofluorescence analysis. When applied, the results from the proposed studies are expected to ultimately lead to new control measures to disrupt the pathogen’s enzootic cycle and prevent infection by the Lyme disease spirochete.