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Project description:Throughout its enzootic cycle, the Lyme disease spirochete Borreliella (Borrelia) burgdorferi, senses and responds to changes in its environment using a small repertoire of transcription factors that coordinate the expression of genes required for infection of Ixodes ticks and various mammalian hosts. Among these transcription factors, the DnaK suppressor protein (DksA) plays a pivotal role in regulating gene expression in B. burgdorferi during periods of nutrient limitation and is required for mammalian infectivity. In many pathogenic bacteria, the gene regulatory activity of DksA, along with the alarmone guanosine penta- and tetra-phosphate ((p)ppGpp), coordinate the stringent response to various environmental stresses, including nutrient limitation. In this study, we sought to characterize the role of DksA in regulating the transcriptional activity of RNA polymerase and its role in the regulation of RpoS-dependent gene expression required for B. burgdorferi infectivity. Using in vitro transcription assays, we observed recombinant DksA inhibits RpoD-dependent transcription by B. burgdorferi RNA polymerase independent of ppGpp. Additionally, we determined the pH-inducible expression of RpoS-dependent genes relies on DksA, but this relationship is independent of (p)ppGpp produced by Relbbu. Subsequent transcriptomic and western blot assays indicate DksA regulates the expression of BBD18, a protein previously implicated in the post-transcriptional regulation of RpoS. Moreover, we observed DksA was required for infection of mice following intraperitoneal inoculation or for transmission of B. burgdorferi by Ixodes scapularis nymphs. Together, these data suggest DksA plays a central role in coordinating transcriptional responses in B. burgdorferi required for infectivity through DksA's interactions with RNA polymerase and post-transcriptional control of RpoS.
| S-EPMC7924775 | biostudies-literature
Project description:Borrelia (B.) burgdorferi sensu lato, including the tick-transmitted agents of human Lyme borreliosis, have particularly complex genomes, consisting of a linear main chromosome and numerous linear and circular plasmids. The number and structure of plasmids is variable even in strains within a single genospecies. Genes on these plasmids are known to play essential roles in virulence and pathogenicity as well as host and vector associations. For this reason, it is essential to explore methods for rapid and reliable characterisation of molecular level changes on plasmids. In this study we used three strains: a low passage isolate of B. burgdorferi sensu stricto strain B31(-NRZ) and two closely related strains (PAli and PAbe) that were isolated from human patients. Sequences of these strains were compared to the previously sequenced reference strain B31 (available in GenBank) to obtain proof-of-principle information on the suitability of next generation sequencing (NGS) library construction and sequencing methods on the assembly of bacterial plasmids. We tested the effectiveness of different short read assemblers on Illumina sequences, and of long read generation methods on sequence data from Pacific Bioscience single-molecule real-time (SMRT) and nanopore (Oxford Nanopore Technologies) sequencing technology.Inclusion of mate pair library reads improved the assembly in some plasmids as did prior enrichment of plasmids. While cp32 plasmids remained refractory to assembly using only short reads they were effectively assembled by long read sequencing methods. The long read SMRT and nanopore sequences came, however, at the cost of indels (insertions or deletions) appearing in an unpredictable manner. Using long and short read technologies together allowed us to show that the three B. burgdorferi s.s. strains investigated here, whilst having similar plasmid structures to each other (apart from fusion of cp32 plasmids), differed significantly from the reference strain B31-GB, especially in the case of cp32 plasmids.Short read methods are sufficient to assemble the main chromosome and many of the plasmids in B. burgdorferi. However, a combination of short and long read sequencing methods is essential for proper assembly of all plasmids including cp32 and thus, for gaining an understanding of host- or vector adaptations. An important conclusion from our work is that the evolution of Borrelia plasmids appears to be dynamic. This has important implications for the development of useful research strategies to monitor the risk of Lyme disease occurrence and how to medically manage it.
| S-EPMC5450258 | biostudies-literature
Project description:Borrelia burgdorferi spirochetes that cause Lyme borreliosis survive for a long time in human serum because they successfully evade the complement system, an important arm of innate immunity. The outer surface protein E (OspE) of B. burgdorferi is needed for this because it recruits complement regulator factor H (FH) onto the bacterial surface to evade complement-mediated cell lysis. To understand this process at the molecular level, we used a structural approach. First, we solved the solution structure of OspE by NMR, revealing a fold that has not been seen before in proteins involved in complement regulation. Next, we solved the x-ray structure of the complex between OspE and the FH C-terminal domains 19 and 20 (FH19-20) at 2.83 ? resolution. The structure shows that OspE binds FH19-20 in a way similar to, but not identical with, that used by endothelial cells to bind FH via glycosaminoglycans. The observed interaction of OspE with FH19-20 allows the full function of FH in down-regulation of complement activation on the bacteria. This reveals the molecular basis for how B. burgdorferi evades innate immunity and suggests how OspE could be used as a potential vaccine antigen.
| S-EPMC3696643 | biostudies-literature
Project description:The spirochetal agent of Lyme disease, Borrelia burgdorferi, is transmitted by bites of Ixodes ticks to mammalian reservoir hosts and humans. The mechanism(s) by which the organism is trafficked from vector to host is poorly understood. In this study, we demonstrate that a B. burgdorferi mutant strain deficient in the synthesis of the bba64 gene product was incapable of infecting mice via tick bite even though the mutant was (i) infectious in mice when introduced by needle inoculation, (ii) acquired by larval ticks feeding on infected mice, and (iii) able to persist through tick molting stages. This finding of a B. burgdorferi gene required for pathogen transfer and/or survival from the tick to the susceptible host represents an important breakthrough toward understanding transmission mechanisms involved for the Lyme disease agent.
| S-EPMC2867675 | biostudies-literature
Project description:Oral vaccination strategies are of interest to prevent transmission of Lyme disease as they can be used to deliver vaccines to humans, pets, and to natural wildlife reservoir hosts of Borrelia burgdorferi. We developed a number of oral vaccines based in E. coli expressing recombinant OspC type K, OspB, BBK32 from B. burgdorferi, and Salp25, Salp15 from Ixodes scapularis. Of the five immunogenic candidates only OspC induced significant levels of antigen-specific IgG and IgA when administered to mice via the oral route. Antibodies to OspC did not prevent dissemination of B. burgdorferi as determined by the presence of spirochetes in ear, heart and bladder tissues four weeks after challenge. Next generation sequencing of genomic DNA from ticks identified multiple phyletic types of B. burgdorferi OspC (A, D, E, F, I, J, K, M, Q, T, X) in nymphs that engorged on vaccinated mice. PCR amplification of OspC types A and K from flat and engorged nymphal ticks, and from heart and bladder tissues collected after challenge confirmed sequencing analysis. Quantification of spirochete growth in a borreliacidal assay shows that both types of spirochetes (A and K) survived in the presence of OspC-K specific serum whereas the spirochetes were killed by OspA specific serum. We show that oral vaccination of C3H-HeN mice with OspC-K induced significant levels of antigen-specific IgG. However, these serologic antibodies did not protect mice from infection with B. burgdorferi expressing homologous or heterologous types of OspC after tick challenge.
| S-EPMC4798528 | biostudies-literature
Project description:The blacklegged tick, Ixodes scapularis, is established in several regions of Ontario, Canada, and continues to spread into new geographic areas across the province at a rapid rate. This poses a significant public health risk since I. scapularis transmits the Lyme disease-causing bacterium, Borrelia burgdorferi, and other pathogens of potential public health concern. The objective of this study was to develop species distribution models for I. scapularis and B. burgdorferi to predict and compare the potential distributions of the tick vector and the Lyme disease pathogen as well as the ecological factors most important for species establishment. Ticks were collected via tick dragging at 120 sites across southern, central, and eastern Ontario between 2015 and 2018 and tested for tick-borne pathogens. A maximum entropy (Maxent) approach was used to model the potential distributions of I. scapularis and B. burgdorferi. Two independent datasets derived from tick dragging at 25 new sites in 2019 and ticks submitted by the public to local health units between 2015 and 2017 were used to validate the predictive accuracy of the models. The model for I. scapularis showed high suitability for blacklegged ticks in eastern Ontario and some regions along the shorelines of the Great Lakes, and moderate suitability near Algonquin Provincial Park and the Georgian Bay with good predictive accuracy (tick dragging 2019: AUC = 0.898; ticks from public: AUC = 0.727). The model for B. burgdorferi showed a similar predicted distribution but was more constrained to eastern Ontario, particularly between Ottawa and Kingston, and along Lake Ontario, with similarly good predictive accuracy (tick dragging 2019: AUC = 0.958; ticks from public: AUC = 0.863. The ecological variables most important for predicting the distributions of I. scapularis and B. burgdorferi included elevation, distance to deciduous and coniferous forest, proportions of agricultural land, water, and infrastructure, mean summer/spring temperature, and cumulative annual degree days above 0°C. Our study presents a novel application of species distribution modelling for I. scapularis and B. burgdorferi in Ontario, Canada, and provides an up to date projection of their potential distributions for public health knowledge users.
| S-EPMC7485816 | biostudies-literature
Project description:The restriction-modification (R-M) systems of many bacteria present a barrier to the stable introduction of foreign DNA. The Lyme disease spirochete Borrelia burgdorferi has two plasmid-borne putative R-M genes, bbe02 and bbq67, whose presence limits transformation by shuttle vector DNA from Escherichia coli. We show that both the bbe02 and bbq67 loci in recipient B. burgdorferi limit transformation with shuttle vector DNA from E. coli, irrespective of its dam, dcm, or hsd methylation status. However, plasmid DNA purified from B. burgdorferi transformed naïve B. burgdorferi much more efficiently than plasmid DNA from E. coli, particularly when the bbe02 and bbq67 genotypes of the B. burgdorferi DNA source matched those of the recipient. We detected adenine methylation of plasmid DNA prepared from B. burgdorferi that carried bbe02 and bbq67. These results indicate that the bbe02 and bbq67 loci of B. burgdorferi encode distinct R-M enzymes that methylate endogenous DNA and cleave foreign DNA lacking the same sequence-specific modification. Our findings have basic implications for horizontal gene transfer among B. burgdorferi strains with distinct plasmid contents. Further characterization and identification of the nucleotide sequences recognized by BBE02 and BBQ67 will facilitate efficient genetic manipulation of this pathogenic spirochete.
| S-EPMC3067601 | biostudies-literature
Project description:Coexistence of multiple tick-borne pathogens or strains is common in natural hosts and can be facilitated by resource partitioning of the host species, within-host localization, or by different transmission pathways. Most vector-borne pathogens are transmitted horizontally via systemic host infection, but transmission may occur in the absence of systemic infection between two vectors feeding in close proximity, enabling pathogens to minimize competition and escape the host immune response. In a laboratory study, we demonstrated that co-feeding transmission can occur for a rapidly-cleared strain of Borrelia burgdorferi, the Lyme disease agent, between two stages of the tick vector Ixodes scapularis while feeding on their dominant host, Peromyscus leucopus. In contrast, infections rapidly became systemic for the persistently infecting strain. In a field study, we assessed opportunities for co-feeding transmission by measuring co-occurrence of two tick stages on ears of small mammals over two years at multiple sites. Finally, in a modeling study, we assessed the importance of co-feeding on R0, the basic reproductive number. The model indicated that co-feeding increases the fitness of rapidly-cleared strains in regions with synchronous immature tick feeding. Our results are consistent with increased diversity of B. burgdorferi in areas of higher synchrony in immature feeding - such as the midwestern United States. A higher relative proportion of rapidly-cleared strains, which are less human pathogenic, would also explain lower Lyme disease incidence in this region. Finally, if co-feeding transmission also occurs on refractory hosts, it may facilitate the emergence and persistence of new pathogens with a more limited host range.
| S-EPMC5474356 | biostudies-literature
Project description:Lyme disease is caused by members of the Borrelia burgdorferi sensu lato species complex. Arthritis is a well-known late-stage pathology of Lyme disease, but the effects of B. burgdorferi infection on bone at sites other than articular surfaces are largely unknown. In this study, we investigated whether B. burgdorferi infection affects bone health in mice. In mice inoculated with B. burgdorferi or vehicle (mock infection), we measured the presence of B. burgdorferi DNA in bones, bone mineral density (BMD), bone formation rates, biomechanical properties, cellular composition, and two- and three-dimensional features of bone microarchitecture. B. burgdorferi DNA was detected in bone. In the long bones, increasing B. burgdorferi DNA copy number correlated with reductions in areal and trabecular volumetric BMDs. Trabecular regions of femora exhibited significant, copy number-correlated microarchitectural disruption, but BMD, microarchitectural, and biomechanical properties of cortical bone were not affected. Bone loss in tibiae was not due to increased osteoclast numbers or bone-resorbing surface area, but it was associated with reduced osteoblast numbers, implying that bone loss in long bones was due to impaired bone building. Osteoid-producing and mineralization activities of existing osteoblasts were unaffected by infection. Therefore, deterioration of trabecular bone was not dependent on inhibition of osteoblast function but was more likely caused by blockade of osteoblastogenesis, reduced osteoblast survival, and/or induction of osteoblast death. Together, these data represent the first evidence that B. burgdorferi infection induces bone loss in mice and suggest that this phenotype results from inhibition of bone building rather than increased bone resorption.
| S-EPMC5278181 | biostudies-literature
Project description:The laboratory diagnosis of Lyme disease is currently dependent on the detection of IgM and IgG antibodies against Borrelia burgdorferi, the causative agent of the disease. The significance of serum IgA against B. burgdorferi remains unclear. The production of intrathecal IgA has been noted in patients with the late Lyme disease manifestation, neuroborreliosis, but production of antigen-specific IgA during early disease has not been evaluated. In the current study, we assessed serum IgA binding to the B. burgdorferi peptide antigens, C6, the target of the FDA-cleared C6 EIA, and FlaB(211-223)-modVlsE(275-291), a peptide containing a Borrelia flagellin epitope linked to a modified VlsE sequence, in patients with early and late Lyme disease. Specific IgA was detected in 59 of 152 serum samples (38.8%) from early Lyme disease patients. Approximately 50% of early Lyme disease patients who were seropositive for peptide-specific IgM and/or IgG were also seropositive for peptide-specific IgA. In a subpopulation of patients, high peptide-specific IgA could be correlated with disseminated disease, defined as multiple erythema migrans lesions, and neurological disease complications. These results suggest that there may be an association between elevated levels of antigen-specific IgA and particular disease manifestations in some patients with early Lyme disease.
| S-EPMC5440658 | biostudies-literature