Borrelia afzelii differential gene expression in the course of tick blood-feeding, and during chronic infection in mice was studied. Temperature effect on B. afzelii gene expression and infectivity was investigated. Infection rates of mice immunized with B. afzelii tick gut antigen at various stages of tick blood-intake were analyzed. This work was funded by the Grant Agency of the Czech Republic, Project No. 17-27393S to Radek Šíma.

The prevalence of chronic Lyme disease (CLD) remains relatively unknown in Connecticut because there is not an agreement on what CLD is and how it should be diagnosed in addition to which pathological agent causes CLD. The aim of this quantitative study was to assess whether there were significant differences between two groups of primary care physicians (PCP) working in Connecticut from two different points in time regarding their knowledge in the diagnosis, treatment, and management of CLD. A knowledge, attitude, and practice model was used as the underlying theoretical framework for this study. A random cross-sectional survey was mailed out to the 1,726 PCPs found in the list of certified medical doctors in Connecticut of 2015. One hundred and forty-five PCPs responses (11.9% response rate) were received and compared to responses from previous data (a 2010 study) of 285 PCPs (39.1% response rate) from the list of certified medical doctors in 2006. The PCP estimated mean number of patients diagnosed and treated for CLD was not significantly different between 2006 and 2015. However, a significantly higher number of PCPs in 2015 reported knowing Lyme disease (LD) symptoms but not feeling comfortable diagnosing LD (χ� = 536.83, p < 0.001), and significantly more PCPs in 2015 reported knowing LD symptoms and feeling comfortable diagnosing CLD (χ� = 265.41, p < 0.001). This study can promote social change by encouraging Connecticut PCPs to recognize CLD as a diagnosis to enable the development of registries and case-control assessments. The findings of this study may also inspire future studies.

Indiana University-Purdue University Indianapolis (IUPUI) / Lyme disease, the most prevalent vector-borne illness in the United States, is a multisystem inflammatory disorder caused by infection with the spirochete Borrelia burgdorferi (Bb). This spirochete is maintained in nature through an enzootic cycle involving ticks and small mammals. The Bb genome encodes a large number of surface lipoproteins, many of which are expressed during mammalian infection. One of these lipoproteins is the major outer surface protein C (OspC) whose production is induced during transmission as spirochetes transition from ticks to mammals. OspC is required for Bb to establish infection in mice and has been proposed to facilitate evasion of innate immunity. However, the exact biological function of OspC remains elusive. Our studies show the ospC-deficient spirochete could not establish infection in NOD-scid IL2rγnull mice that lack B cells, T cells, NK cells, and lytic complement, whereas the wild-type spirochete was fully infectious in these mice. The ospC mutant also could not establish infection in SCID and C3H mice that were transiently neutropenic during the first 48 h post-challenge. However, depletion of F4/80+ phagocytes at the skin-site of inoculation in SCID mice allowed the ospC mutant to establish infection in vivo. In phagocyte-depleted SCID mice, the ospC mutant was capable to colonize the joints and triggered neutrophilia during dissemination in a similar pattern as wild-type bacteria. We then constructed GFP-expressing Bb strains to evaluate the interaction of the ospC mutant with phagocytes. Using flow cytometry and fluorometric assay for phagocytosis, we found that phagocytosis of GFP-expressing ospC mutant spirochetes by murine peritoneal macrophages and human THP-1 cells was significantly higher than parental wild-type Bb strains, suggesting that OspC has an anti-phagocytic property. This enhancement in phagocytosis was not mediated by MARCO and CD36 scavenger receptors and was not associated with changes in mRNA levels of TNFα, IL-1β, and IL-10. Phagocytosis assays with HL60 neutrophil-like cells showed that uptake of Bb strains was independent to OspC. Together, our findings reveal that F4/80+ phagocytes are important for clearance of the ospC mutant, and suggest that OspC promotes spirochetes' evasion of macrophages in the skin of mice during early Lyme borreliosis.

In North America, Borrelia burgdorferi is the primary causative agent of Lymedisease which is a growing health concern. The ability of B. burgdorferi to maintain chronic infection indicates that they are capable of immune evasion. A distinguishing characteristic of B. burgdorferi is the large number of sequences encoding predicted or known lipoproteins, including outer surface protein F (OspF). This study analyzes the specificity of the humoral immune response to B. burgdorferi B3 IMI OspF proteins during murine and human infection. Immunoblot analyses revealed a temporal expression of OspF proteins during infection and mapped the immunodominant epitopes which lie within the variable domains. To determine if OspF-related proteins are produced by other isolates, immunoblot analyses were performed using sera collected from mice and humans infected with diverse B. burgdorferi strains. Differences in the immunoreactivity profile to OspF proteins were seen among the infection sera tested. To identify the molecular basis of these differences, the ospF gene was isolated from several strains, sequenced and evolutionary analyses were conducted. These analyses revealed that OspF proteins show little diversity despite the separate geographic locations from which isolates originated. The high degree of OspF protein conservation seen in isolates from two distinct regions emphasizes the potential for OpsF proteins as vaccinogens or in serodiagnostic assays. Altogether, this study demonstrates the potential contribution of OspF proteins to immune evasion through its temporal expression during infection which may play specific roles at different stages of infection. Studies are underway to determine if inactivation of ospF genes through allelic exchange mutagenesis impacts on the pathogenicity of the Lyme disease spirochetes.

Lyme disease is one of the most common vector-borne diseases around the world, and the numbers of reported cases are quickly rising. Ixodes ticks are the principal vectors, while Borrelia burgdorferi sensu lato genospecies are the etiological agents of the disease. Climate change, namely global warming, and habitat modification, namely forest fragmentation, are hypothesized to play an active role in this rise in reported cases. An analysis of the primary literature, specifically of studies focused on North America and Europe, was conducted in order to investigate these hypotheses. These studies show that global warming has precipitated a growth in tick populations as well as a northward tick migration, thereby increasing the risk of Lyme disease in emergent and endemic areas alike, for Borrelia spirochetes quickly infect naïve tick populations. Furthermore, published studies support the idea that forest fragmentation near human population centers has also increased the risk of Lyme disease in North America, for edge habitats provide suitable conditions for ticks and provide edible vegetation for the animals on which ticks feed, animals which also serve as hosts for B. burgdorferi sensu lato. In contrast, a decrease in fragmentation was found to facilitate tick invasion and establishment in Europe. These studies demonstrate that anthropogenic habitat modifications of varying types can affect ticks and their host populations and increase the risk of Lyme disease near human population centers. However, more research needs to be done to truly understand the different factors that are precipitating the rising number of cases of Lyme disease since there are significant interactions between climate change, habitat modification, and other drivers not examined here. Furthermore, understanding how these drivers function in specific geographic locations can help scientists and public officials tailor local public health measures appropriately. Finally, researchers and pharmaceutical companies must develop a safe, long-lasting, and effective vaccine against the Lyme disease spirochete, for there is not one currently available. Although easily treatable if diagnosed early, Lyme disease can progress to debilitating disease. Unfortunately, the risk of contracting this illness is currently rising and will continue to rise unless effective preventative measures are employed.

Uninfected larvae of Ixodes dammini, lxodes cookei, and Amblyomma americanum were fed on hamsters that had been injected intraperitoneally with a 0.5.ml sample of Borrelia burgdorferl (2.5 X 107 spirochetes per ml) 21 days earlier. A total of 108 of these larvae comprised of 36 1. dammini, 36 i. cookei, and 36 A. americanum were aseptically dissected and examined by darkfield and immunofluorescent microscopy for the presence of B. burgdorferl within 48 hours of feeding on the B. burgdorferi infected hamsters. The removal and examination of the midgut diverticula revealed that 32/36 (88.9%) of the l. dammini larvae contained B. burgdorferl. Only 5/36 (13.9%) of the l. cookei larvae and 7/36 of the A. americanum larvae harbored spirochetes in their midgut diverticula.A portion of the nymphs that molted from the above larvae were also dissected and examined by darkfleld and indirect immunofluorescent techniques. Borrelia burgdorferi were observed in the midgut diverticula of 94/107 (87.8%) of the l. dammini nymphs. None of the 30 (0%) l. cookei nymphs examined were found positive for spirochetes and only 1/60 (1.7%) of the A. americanum nymphs was found positive for B. burgdorrerl.A total of 83 lL dammini, 53 A. americanum, and 161. cookei nymphs reared from larvae that fed to repletion on hamsters infected with B. burgdorrerl were allowed to feed on uninfected hamsters to assess transmission of B. burgdorrerl. Transmission was demonstrated only by the l. dammlnl nymphs. The findings of this study suggest that it is extremely unlikely that l. cookei can serve as a vector for B. burgdorrerl, but do not rule out completely the possibility that A. americanum may be able to maintain B, burgdorrerl infections transstadially and, under certain conditions, transmit the organisms to vertebrate hosts. / Department of Physiology and Health Science

Uninfected nymphs of Ixodes scapularis and Amblyomma americanum were fed on hamsters that had been injected intraperitoneally with a 0.5 ml sample of Borrelia burgdorferi (2.5 X 10' spirochetes per ml) 30 days earlier. All nymphs fed to repletion and were then housed during the molting process. In order to assess their ability to transmit the spirochetes, the resulting l. scapularis and A. americanum adults were allowed to feed on uninfected rabbits.Dissection of the adult l. scapularis ticks revealed 9/12 (75%) to harbor motile spirochetes, identified as B. burgdorferi by darkfield microscopy, isolation in BSK II medium, and indirect immunofluorescent antibody staining with the monoclonal antibody H5332. Transmission was successful to one of two New Zealand White rabbits by these infected ticks.Dissection of the adult A. americanum ticks revealed 0/150 (0%) to harbor spirochetes. Transmission to each of three rabbits was unsuccessful. However, 5/90 (5.6%) cultures of midgut material from these same ticks, harbored non-motile spirochete-like bodies. The identity of these "spirochetes" is unknown. / Department of Biology

Thesis (M.S.)--University of Tennessee, Knoxville, 2009. / Title from title page screen (viewed on Mar. 18, 2010). Thesis advisor: Graham Hickling. Vita. Includes bibliographical references.

Thesis (Ph. D.)--University of Missouri-Columbia, 1999. / Typescript. Vita. Includes bibliographical references (leaves 70-85). Also available on the Internet.

Thesis (Ph. D.)--University of Missouri-Columbia, 1999. / Typescript. Vita. Includes bibliographical references (leaves 70-85). Also available on the Internet.