ResearchPad - borrelia https://www.researchpad.co Default RSS Feed en-us © 2020 Newgen KnowledgeWorks <![CDATA[Factors affecting the microbiome of <i>Ixodes scapularis</i> and <i>Amblyomma americanum</i>]]> https://www.researchpad.co/article/elastic_article_14701 The microbial community composition of disease vectors can impact pathogen establishment and transmission as well as on vector behavior and fitness. While data on vector microbiota are accumulating quickly, determinants of the variation in disease vector microbial communities are incompletely understood. We explored the microbiome of two human-biting tick species abundant in eastern North America (Amblyomma americanum and Ixodes scapularis) to identify the relative contribution of tick species, tick life stage, tick sex, environmental context and vertical transmission to the richness, diversity, and species composition of the tick microbiome. We sampled 89 adult and nymphal Ixodes scapularis (N = 49) and Amblyomma americanum (N = 40) from two field sites and characterized the microbiome of each individual using the v3-v4 hypervariable region of the 16S rRNA gene. We identified significant variation in microbial community composition due to tick species and life stage with lesser impact of sampling site. Compared to unfed nymphs and males, the microbiome of engorged adult female I. scapularis, as well as the egg masses they produced, were low in bacterial richness and diversity and were dominated by Rickettsia, suggesting strong vertical transmission of this genus. Likewise, microbiota of A. americanum nymphs and males were more diverse than those of adult females. Among bacteria of public health importance, we detected several different Rickettsia sequence types, several of which were distinct from known species. Borrelia was relatively common in I. scapularis but did not show the same level of sequence variation as Rickettsia. Several bacterial genera were significantly over-represented in Borrelia-infected I. scapularis, suggesting a potential interaction of facilitative relationship between these taxa; no OTUs were under-represented in Borrelia-infected ticks. The systematic sampling we conducted for this study allowed us to partition the variation in tick microbial composition as a function of tick- and environmentally-related factors. Upon more complete understanding of the forces that shape the tick microbiome it will be possible to design targeted experimental studies to test the impacts of individual taxa and suites of microbes on vector-borne pathogen transmission and on vector biology.

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<![CDATA[The intergenic small non-coding RNA <i>ittA</i> is required for optimal infectivity and tissue tropism in <i>Borrelia burgdorferi</i>]]> https://www.researchpad.co/article/elastic_article_14635 Lyme disease is a tick-borne infection mediated by the spirochetal bacterium, Borrelia burgdorferi, that is responsible for greater than 300,000 infections in the United States per year. As such, additional knowledge regarding how this pathogen modulates its regulatory armamentarium is needed to understand how B. burgdorferi establishes and maintains infection. The identification and characterization of small, non-coding RNA molecules in living systems, designated as sRNAs, has recalibrated how we view post-transcriptional regulation. Recently, over 1,000 sRNAs were identified in B. burgdorferi. Despite the identification of these sRNAs, we do not understand how they affect infectivity or B. burgdorferi pathogenesis related outcomes. Here, we characterize the ittA B. burgdorferi sRNA and show that it is essential for optimal infection using murine experimental infection as our readout. We also track the effect of this sRNA on the transcriptional and proteomic profile as the first step in providing mechanistic insight into how this important sRNA mediates its regulatory effect.

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<![CDATA[Optimization of tissue sampling for Borrelia burgdorferi in white-footed mice (Peromyscus leucopus)]]> https://www.researchpad.co/article/Nff220985-8630-4822-8507-6b577103a931

Peromyscus leucopus (the white-footed mouse) is a known reservoir of the Lyme disease spirochete Borrelia burgdorferi. Sampling of white-footed mice allows for year-round B. burgdorferi surveillance as well as opportunities to establish the diversity of the different variants in a geographic region. This study explores the prevalence of B. burgdorferi infections in the tissues of white-footed mice, investigates the correlations between B. burgdorferi infected tissues, and determines the optimum field methods for surveillance of B. burgdorferi in P. leucopus. A total of 90 mice and 573 tissues (spleen, liver, ear, tongue, tail, heart, and kidney) were screened via nested PCR for B. burgdorferi infections. A large number of infections were found in the 90 mice as well as multiple infections within individual mice. Infections in a single mouse tissue (spleen, liver, ear, tongue and tail) were predictive of concurrent infection in other tissues of the same mouse at a statistically significant level. Ear tissue accounted for 68.4% of detected infections, which increased to 78.9% of the infected mice with the inclusion of tail samples. The use of ear punch or tail snip samples (used individually or in tandem) have multiple advantages over current Lyme disease ecological studies and surveillance methodologies, including lower associated costs, minimization of delays, year-round B. burgdorferi testing opportunities, as well as longitudinal monitoring of B. burgdorferi in defined geographic regions. In the absence of an effective vaccine, personal prevention measures are currently the most effective way to reduce Lyme disease transmission to humans. Thus, the identification and monitoring of environmental reservoirs to inform at-risk populations remains a priority. The sampling methods proposed in this study provide a reasonable estimate of B. burgdorferi in white-footed mice in a timely and cost-effective manner.

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<![CDATA[Detection of municipalities at-risk of Lyme disease using passive surveillance of Ixodes scapularis as an early signal: A province-specific indicator in Canada]]> https://www.researchpad.co/article/5c75ac66d5eed0c484d086db

Lyme disease, the most commonly reported vector-borne disease in North America, is caused by the spirochete Borrelia burgdorferi sensu stricto, which is transmitted by Ixodes scapularis in eastern Canada and Ixodes pacificus in western Canada. Recently, the northward range expansion of I. scapularis ticks, in south-eastern Canada, has resulted in a dramatic increase in the incidence of human Lyme disease. Detecting emerging areas of Lyme disease risk allows public health to target disease prevention efforts. We analysed passive tick surveillance data from Ontario and Manitoba to i) assess the relationship between the total numbers of I. scapularis submissions in passive surveillance from humans, and the number of human Lyme disease cases, and ii) develop province-specific acarological indicators of risk that can be used to generate surveillance-based risk maps. We also assessed associations between numbers of nymphal I. scapularis tick submissions only and Lyme disease case incidence. Using General Estimating Equation regression, the relationship between I. scapularis submissions (total numbers and numbers of nymphs only) in each census sub-division (CSD) and the number of reported Lyme disease cases was positively correlated and highly significant in the two provinces (P ≤ 0.001). The numbers of I. scapularis submissions over five years discriminated CSDs with ≥ 3 Lyme disease cases from those with < 3 cases with high accuracy when using total numbers of tick submission (Receiver Operating Characteristics area under the curve [AUC] = 0.89) and moderate accuracy (AUC = 0.78) when using nymphal tick submissions only. In Ontario the optimal cut-off point was a total 12 tick submissions from a CSD over five years (Sensitivity = 0.82, Specificity = 0.84), while in Manitoba the cut-off point was five ticks (Sensitivity = 0.71, Specificity = 0.79) suggesting regional variability of the risk of acquiring Lyme disease from an I. scapularis bite. The performances of the acarological indicators developed in this study for Ontario and Manitoba support the ability of passive tick surveillance to provide an early signal of the existence Lyme disease risk areas in regions where ticks and the pathogens they transmit are expanding their range.

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<![CDATA[The growing importance of lone star ticks in a Lyme disease endemic county: Passive tick surveillance in Monmouth County, NJ, 2006 – 2016]]> https://www.researchpad.co/article/5c6c75cbd5eed0c4843d01e0

As human cases of tick-borne disease continue to increase, there is a heightened imperative to collect data on human-tick encounters to inform disease prevention. Passive tick surveillance programs that encourage members of the public to submit ticks they have encountered can provide a relatively low-cost means of collecting such data. We report the results of 11 years of tick submissions (2006–2016) collected in Monmouth County, New Jersey, an Atlantic coastal county long endemic for Lyme disease. A total of 8,608 ticks acquired in 22 U.S. states were submitted, 89.7% of which were acquired in Monmouth County, from 52 of the County’s 53 municipalities. Seasonal submission rates reflected known phenology of common human-biting ticks, but annual submissions of both Amblyomma americanum and Dermacentor variabilis increased significantly over time while numbers of Ixodes scapularis remained static. By 2016, A. americanum had expanded northward in the county and now accounted for nearly half (48.1%) of submissions, far outpacing encounters with I. scapularis (28.2% of submissions). Across all tick species and stages the greatest number of ticks were removed from children (ages 0–9, 40.8%) and older adults (ages 50+, 23.8%) and these age groups were also more likely to submit partially or fully engorged ticks, suggesting increased risk of tick-borne disease transmission to these vulnerable age groups. Significantly more people (43.2%) reported acquiring ticks at their place of residence than in a park or natural area (17.9%). This pattern was more pronounced for residents over 60 years of age (72.7% acquired at home). Education that stresses frequent tick checks should target older age groups engaged in activity around the home. Our results strongly suggest that encounter rates with ticks other than I. scapularis are substantial and increasing and that their role in causing human illness should be carefully investigated.

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<![CDATA[Association of small fiber neuropathy and post treatment Lyme disease syndrome]]> https://www.researchpad.co/article/5c6c75a6d5eed0c4843cff72

Objectives

To examine whether post-treatment Lyme disease syndrome (PTLDS) defined by fatigue, cognitive complaints and widespread pain following the treatment of Lyme disease is associated with small fiber neuropathy (SFN) manifesting as autonomic and sensory dysfunction.

Methods

This single center, retrospective study evaluated subjects with PTLDS. Skin biopsies for assessment of epidermal nerve fiber density (ENFD), sweat gland nerve fiber density (SGNFD) and functional autonomic testing (deep breathing, Valsalva maneuver and tilt test) were performed to assess SFN, severity of dysautonomia and cerebral blood flow abnormalities. Heart rate, end tidal CO2, blood pressure, and cerebral blood flow velocity (CBFv) from middle cerebral artery using transcranial Doppler were monitored.

Results

10 participants, 5/5 women/men, age 51.3 ± 14.7 years, BMI 27.6 ± 7.3 were analyzed. All participants were positive for Lyme infection by CDC criteria. At least one skin biopsy was abnormal in all ten participants. Abnormal ENFD was found in 9 participants, abnormal SGNFD in 5 participants, and both abnormal ENFD and SGNFD were detected in 4 participants. Parasympathetic failure was found in 7 participants and mild or moderate sympathetic adrenergic failure in all participants. Abnormal total CBFv score was found in all ten participants. Low orthostatic CBFv was found in 7 participants, three additional participants had abnormally reduced supine CBFv.

Conclusions

SFN appears to be associated with PTLDS and may be responsible for certain sensory symptoms. In addition, dysautonomia related to SFN and abnormal CBFv also seem to be linked to PTLDS. Reduced orthostatic CBFv can be associated with cerebral hypoperfusion and may lead to cognitive dysfunction. Autonomic failure detected in PTLDS is mild to moderate. SFN evaluation may be useful in PTLDS.

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<![CDATA[The economic burden of Lyme disease and the cost-effectiveness of Lyme disease interventions: A scoping review]]> https://www.researchpad.co/article/5c390bc8d5eed0c48491e514

Introduction

While Lyme disease (LD) is mostly treatable, misdiagnosed or untreated LD can result in debilitating sequelae and excessive healthcare usage. The objective of this review was to characterize the body of literature on the economic burden of Lyme disease (LD) and the cost-effectiveness of LD interventions, such as antibiotic treatment and vaccination.

Methods

We followed Joanna Briggs Institute scoping review methodologies. We systematically searched terms related to LD, economic evaluations, costs, and cost-effectiveness in Medline, Embase, PsycInfo, Cochrane Library, and the grey literature up to November 2017. We included primary economic evaluations conducted in North America and Europe, reporting LD-related costs or cost-effectiveness of human interventions. Two reviewers screened articles and charted data independently. Costs were standardized to 2017 United States dollars (USD).

Results

We screened 923 articles, and included 10 cost-effectiveness analyses (CEA) and 11 cost analyses (CA). Three CEAs concluded LD vaccination was likely cost-effective only in endemic areas (probability of infection ≥1%). However, LD vaccination is not currently available as an intervention in the US or Europe. Six studies assessed economic burden from a societal perspective and estimated significant annual national economic impact of: 735,550 USD for Scotland (0.14 USD per capita, population = 5.40M), 142,562 USD in Sweden (0.014 USD per capita, 9.96M), 40.88M USD in Germany (0.51 USD per capita, 80.59M), 23.12M USD in the Netherlands (1.36 USD per capita, 17.08M), and up to 786M USD in the US (2.41 USD per capita, 326.63M).

Conclusions

Lyme disease imposes an economic burden that could be considered significant in the US and other developed countries to justify further research efforts in disease control and management. Societal costs for Lyme disease can be equally impactful as healthcare costs, but are not fully understood. Economic literature from countries with historically high incidence rates or increasing rates of Lyme disease are limited, and can be useful for future justification of resource allocation.

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<![CDATA[Borrelia miyamotoi Infections in Small Mammals, California, USA]]> https://www.researchpad.co/article/5c1686a6d5eed0c48444437d

Surveillance to investigate the wildlife–vector transmission cycle of the human pathogen Borrelia miyamotoi in California, USA, revealed infections in dusky-footed woodrats, brush mice, and California mice. Phylogenetic analyses suggest a single, well-supported clade of B. miyamotoi is circulating in California.

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<![CDATA[Genetic manipulation of Leishmania donovani threonyl tRNA synthetase facilitates its exploration as a potential therapeutic target]]> https://www.researchpad.co/article/5b3099a1463d7e6a0ede32bd

Background

Aminoacyl tRNA synthetases are central enzymes required for protein synthesis. These enzymes are the known drug targets in bacteria and fungi. Here, we for the first time report the functional characterization of threonyl tRNA synthetase (LdThrRS) of Leishmania donovani, a protozoan parasite, the primary causative agent of visceral leishmaniasis.

Methodology

Recombinant LdThrRS (rLdThrRS) was expressed in E. coli and purified. The kinetic parameters for rLdThrRS were determined. The subcellular localization of LdThrRS was done by immunofluorescence analysis. Heterozygous mutants of LdThrRS were generated in Leishmania promastigotes. These genetically manipulated parasites were checked for their proliferation, virulence, aminoacylation activity and sensitivity to the known ThrRS inhibitor, borrelidin. An in silico generated structural model of L. donovani ThrRS was compared to that of human.

Conclusions

Recombinant LdThrRS displayed aminoacylation activity, and the protein is possibly localized to both the cytosol and mitochondria. The comparison of the 3D-model of LdThrRS to human ThrRS displayed considerable similarity. Heterozygous parasites showed restrictive growth phenotype and had attenuated infectivity. These heterozygous parasites were more susceptible to inhibition by borrelidin. Several attempts to obtain ThrRS homozygous null mutants were not successful, indicating its essentiality for the Leishmania parasite. Borrelidin showed a strong affinity for LdThrRS (KD: 0.04 μM) and was effective in inhibiting the aminoacylation activity of the rLdThrRS (IC50: 0.06 μM). Borrelidin inhibited the promastigotes (IC50: 21 μM) stage of parasites. Our data shows that LdThrRS is essential for L. donovani survival and is likely to bind with small drug-like molecules with strong affinity, thus making it a potential target for drug discovery efforts.

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<![CDATA[Head Lice of Pygmies Reveal the Presence of Relapsing Fever Borreliae in the Republic of Congo]]> https://www.researchpad.co/article/5989db2aab0ee8fa60bd12a7

Background

Head lice, Pediculus humanus capitis, occur in four divergent mitochondrial clades (A, B, C and D), each having particular geographical distributions. Recent studies suggest that head lice, as is the case of body lice, can act as a vector for louse-borne diseases. Therefore, understanding the genetic diversity of lice worldwide is of critical importance to our understanding of the risk of louse-borne diseases.

Methodology/Principal Findings

Here, we report the results of the first molecular screening of pygmies’ head lice in the Republic of Congo for seven pathogens and an analysis of lice mitochondrial clades. We developed two duplex clade-specific real-time PCRs and identified three major mitochondrial clades: A, C, and D indicating high diversity among the head lice studied. We identified the presence of a dangerous human pathogen, Borrelia recurrentis, the causative agent of relapsing fever, in ten clade A head lice, which was not reported in the Republic of Congo, and B. theileri in one head louse. The results also show widespread infection among head lice with several species of Acinetobacter. A. junii was the most prevalent, followed by A. ursingii, A. baumannii, A. johnsonii, A. schindleri, A. lwoffii, A. nosocomialis and A. towneri.

Conclusions/Significance

Our study is the first to show the presence of B. recurrentis in African pygmies’ head lice in the Republic of Congo. This study is also the first to report the presence of DNAs of B. theileri and several species of Acinetobacter in human head lice. Further studies are needed to determine whether the head lice can transmit these pathogenic bacteria from person to another.

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<![CDATA[Influences of Host Community Characteristics on Borrelia burgdorferi Infection Prevalence in Blacklegged Ticks]]> https://www.researchpad.co/article/5989da0aab0ee8fa60b7762b

Lyme disease is a major vector-borne bacterial disease in the USA. The disease is caused by Borrelia burgdorferi, and transmitted among hosts and humans, primarily by blacklegged ticks (Ixodes scapularis). The ~25 B. burgdorferi genotypes, based on genotypic variation of their outer surface protein C (ospC), can be phenotypically separated as strains that primarily cause human diseases—human invasive strains (HIS)—or those that rarely do. Additionally, the genotypes are non-randomly associated with host species. The goal of this study was to examine the extent to which phenotypic outcomes of B. burgdorferi could be explained by the host communities fed upon by blacklegged ticks. In 2006 and 2009, we determined the host community composition based on abundance estimates of the vertebrate hosts, and collected host-seeking nymphal ticks in 2007 and 2010 to determine the ospC genotypes within infected ticks. We regressed instances of B. burgdorferi phenotypes on site-specific characteristics of host communities by constructing Bayesian hierarchical models that properly handled missing data. The models provided quantitative support for the relevance of host composition on Lyme disease risk pertaining to B. burgdorferi prevalence (i.e. overall nymphal infection prevalence, or NIPAll) and HIS prevalence among the infected ticks (NIPHIS). In each year, NIPAll and NIPHIS was found to be associated with host relative abundances and diversity. For mice and chipmunks, the association with NIPAll was positive, but tended to be negative with NIPHIS in both years. However, the direction of association between shrew relative abundance with NIPAll or NIPHIS differed across the two years. And, diversity (H') had a negative association with NIPAll, but positive association with NIPHIS in both years. Our analyses highlight that the relationships between the relative abundances of three primary hosts and the community diversity with NIPAll, and NIPHIS, are variable in time and space, and that disease risk inference, based on the role of host community, changes when we examine risk overall or at the phenotypic level. Our discussion focuses on the observed relationships between prevalence and host community characteristics and how they substantiate the ecological understanding of phenotypic Lyme disease risk.

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<![CDATA[Genomic Characteristics of Chinese Borrelia burgdorferi Isolates]]> https://www.researchpad.co/article/5989daa7ab0ee8fa60ba7fca

In China, B. burgdorferi, B.garinii, B. afzelii and B. yangtze sp. nov have been reported; B.garinii and B. afzelii are the main pathogenic genotypes. But until now only one Chinese strain was reported with whole genome sequence. In order to further understand the genomic characteristics and diversity of Chinese Borrelia strains, 5 isolates from China were sequenced and compared with the whole genome sequences of strains in other areas. The results showed a high degree of conservation within the linear chromosome of Chinese strains, whereas plasmid showed a much larger diversity according to the majority genomic information of plasmids. The genome sequences of the five Chinese strains were compared with the corresponding reference strains, respectively, according to the genospecies. Pairwise analysis demonstrates that there are only 70 SNPs between the genomes of CS4 and B31. However, there are many more SNPs between the genomes of QX-S13 and VS116, PD91 and PBi, FP1 and PKo, R9 and Pko, respectively. Gene comparison showed some important different genes. OspA was one of the important different genes. Comparative genomic studies have found that OspA gene sequences of PD91 and R9 had great differences compared with the sequence of B31. OspA gene sequence of R9 had a 96bp deletion; OspA gene of PD91 had two deletions: 9bp and 10 bp. To conclude, we showed the genomic characteristics of four genotype Chinese B. burgdorferi strains. The genomic sequence of B. yangtze sp. nov and differences from B. valaisiana were first reported. Comparative analysis of Chinese strains with the different Borrelia species from other areas will help us to understand evolution and pathogenesis of Chinese Borrelia burgdorferi strains.

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<![CDATA[Environmental Factors Affecting Survival of Immature Ixodes scapularis and Implications for Geographical Distribution of Lyme Disease: The Climate/Behavior Hypothesis]]> https://www.researchpad.co/article/5989daadab0ee8fa60ba9e4d

Recent reports suggest that host-seeking nymphs in southern populations of Ixodes scapularis remain below the leaf litter surface, while northern nymphs seek hosts on leaves and twigs above the litter surface. This behavioral difference potentially results in decreased tick contact with humans in the south, and fewer cases of Lyme disease. We studied whether north-south differences in tick survival patterns might contribute to this phenomenon. Four month old larvae resulting from a cross between Wisconsin males and South Carolina females died faster under southern than under northern conditions in the lab, as has previously been reported for ticks from both northern and southern populations. However, newly-emerged larvae from Rhode Island parents did not differ consistently in mortality under northern and southern conditions, possibly because of their younger age. Survival is lower, and so the north-south survival difference might be greater in older ticks. Larval survival was positively related to larval size (as measured by scutal area), while survival was positively related to larval fat content in some, but not all, trials. The difference in larval survival under northern vs. southern conditions might simply result from faster metabolism under warmer southern conditions leading to shorter life spans. However, ticks consistently died faster under southern than under northern conditions in the laboratory when relative humidity was low (75%), but not under moderate (85%) or high (95%) RH. Therefore, mortality due to desiccation stress is greater under southern than under northern conditions. We hypothesize that mortality resulting from the greater desiccation stress under southern conditions acts as a selective pressure resulting in the evolution of host-seeking behavior in which immatures remain below the leaf litter surface in southern I. scapularis populations, so as to avoid the desiccating conditions at the surface. If this hypothesis is correct, it has implications for the effect of climate change on the future distribution of Lyme disease.

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<![CDATA[Louse-Borne Relapsing Fever (Borrelia recurrentis) in a Somali Refugee Arriving in Italy: A Re-emerging Infection in Europe?]]> https://www.researchpad.co/article/5989da62ab0ee8fa60b912d7 ]]> <![CDATA[Microarray Analyses of Inflammation Response of Human Dermal Fibroblasts to Different Strains of Borrelia burgdorferi Sensu Stricto]]> https://www.researchpad.co/article/5989da6dab0ee8fa60b93889

In Lyme borreliosis, the skin is the key site of bacterial inoculation by the infected tick, and of cutaneous manifestations, erythema migrans and acrodermatitis chronica atrophicans. We explored the role of fibroblasts, the resident cells of the dermis, in the development of the disease. Using microarray experiments, we compared the inflammation of fibroblasts induced by three strains of Borrelia burgdorferi sensu stricto isolated from different environments and stages of Lyme disease: N40 (tick), Pbre (erythema migrans) and 1408 (acrodermatitis chronica atrophicans). The three strains exhibited a similar profile of inflammation with strong induction of chemokines (CXCL1 and IL-8) and IL-6 cytokine mainly involved in the chemoattraction of immune cells. Molecules such as TNF-alpha and NF-κB factors, metalloproteinases (MMP-1, -3 and -12) and superoxide dismutase (SOD2), also described in inflammatory and cellular events, were up-regulated. In addition, we showed that tick salivary gland extracts induce a cytotoxic effect on fibroblasts and that OspC, essential in the transmission of Borrelia to the vertebrate host, was not responsible for the secretion of inflammatory molecules by fibroblasts. Tick saliva components could facilitate the early transmission of the disease to the site of injury creating a feeding pit. Later in the development of the disease, Borrelia would intensively multiply in the skin and further disseminate to distant organs.

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<![CDATA[Evidence for Host-Genotype Associations of Borrelia burgdorferi Sensu Stricto]]> https://www.researchpad.co/article/5989d9feab0ee8fa60b730b0

Different genotypes of the agent of Lyme disease in North America, Borrelia burgdorferi sensu stricto, show varying degrees of pathogenicity in humans. This variation in pathogenicity correlates with phylogeny and we have hypothesized that the different phylogenetic lineages in North America reflect adaptation to different host species. In this study, evidence for host species associations of B. burgdorferi genotypes was investigated using 41 B. burgdorferi-positive samples from five mammal species and 50 samples from host-seeking ticks collected during the course of field studies in four regions of Canada: Manitoba, northwestern Ontario, Quebec, and the Maritimes. The B. burgdorferi genotypes in the samples were characterized using three established molecular markers (multi-locus sequence typing [MLST], 16S-23S rrs-rrlA intergenic spacer, and outer surface protein C sequence [ospC] major groups). Correspondence analysis and generalized linear mixed effect models revealed significant associations between B. burgdorferi genotypes and host species (in particular chipmunks, and white-footed mice and deer mice), supporting the hypotheses that host adaptation contributes to the phylogenetic structure and possibly the observed variation in pathogenicity in humans.

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<![CDATA[A high-throughput genetic screen identifies previously uncharacterized Borrelia burgdorferi genes important for resistance against reactive oxygen and nitrogen species]]> https://www.researchpad.co/article/5989db53ab0ee8fa60bdce51

Borrelia burgdorferi, the causative agent of Lyme disease in humans, is exposed to reactive oxygen and nitrogen species (ROS and RNS) in both the tick vector and vertebrate reservoir hosts. B. burgdorferi contains a limited repertoire of canonical oxidative stress response genes, suggesting that novel gene functions may be important for protection of B. burgdorferi against ROS or RNS exposure. Here, we use transposon insertion sequencing (Tn-seq) to conduct an unbiased search for genes involved in resistance to nitric oxide, hydrogen peroxide, and tertiary-butyl hydroperoxide in vitro. The screens identified 66 genes whose disruption resulted in increased susceptibility to at least one of the stressors. These genes include previously characterized mediators of ROS and RNS resistance (including components of the nucleotide excision repair pathway and a subunit of a riboflavin transporter), as well as novel putative resistance candidates. DNA repair mutants were among the most sensitive to RNS in the Tn-seq screen, and survival assays with individual Tn mutants confirmed that the putative ribonuclease BB0839 is involved in resistance to nitric oxide. In contrast, mutants lacking predicted inner membrane proteins or transporters were among the most sensitive to ROS, and the contribution of three such membrane proteins (BB0017, BB0164, and BB0202) to ROS sensitivity was confirmed using individual Tn mutants and complemented strains. Further analysis showed that levels of intracellular manganese are significantly reduced in the Tn::bb0164 mutant, identifying a novel role for BB0164 in B. burgdorferi manganese homeostasis. Infection of C57BL/6 and gp91phox-/- mice with a mini-library of 39 Tn mutants showed that many of the genes identified in the in vitro screens are required for infectivity in mice. Collectively, our data provide insight into how B. burgdorferi responds to ROS and RNS and suggests that this response is relevant to the in vivo success of the organism.

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<![CDATA[RpoS Regulates Essential Virulence Factors Remaining to Be Identified in Borrelia burgdorferi]]> https://www.researchpad.co/article/5989d9f1ab0ee8fa60b6ea57

Background

Since the RpoN-RpoS regulatory network was revealed in the Lyme disease spirochete Borrelia burgdorferi a decade ago, both upstream and downstream of the pathway have been intensively investigated. While significant progress has been made into understanding of how the network is regulated, most notably, discovering a relationship of the network with Rrp2 and BosR, only three crucial virulence factors, including outer surface protein C (OspC) and decorin-binding proteins (Dbps) A and B, are associated with the pathway. Moreover, for more than 10 years no single RpoS-controlled gene has been found to be critical for infection, raising a question about whether additional RpoS-dependent virulence factors remain to be identified.

Methodology/Principal Findings

The rpoS gene was deleted in B. burgdorferi; resulting mutants were modified to constitutively express all the known virulence factors, OspC, DbpA and DbpB. This genetic modification was unable to restore the rpoS mutant with infectivity.

Conclusions/Significance

The inability to restore the rpoS mutant with infectivity by simultaneously over-expressing all the three virulence factors allows us to conclude RpoS also regulates essential genes that remain to be identified in B. burgdorferi.

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<![CDATA[Transcriptional Profiling the 150 kb Linear Megaplasmid of Borrelia turicatae Suggests a Role in Vector Colonization and Initiating Mammalian Infection]]> https://www.researchpad.co/article/5989d9fbab0ee8fa60b721f1

Adaptation is key for survival as vector-borne pathogens transmit between the arthropod and vertebrate, and temperature change is an environmental signal inducing alterations in gene expression of tick-borne spirochetes. While plasmids are often associated with adaptation, complex genomes of relapsing fever spirochetes have hindered progress in understanding the mechanisms of vector colonization and transmission. We utilized recent advances in genome sequencing to generate the most complete version of the Borrelia turicatae 150 kb linear megaplasmid (lp150). Additionally, a transcriptional analysis of open reading frames (ORFs) in lp150 was conducted and identified regions that were up-regulated during in vitro cultivation at tick-like growth temperatures (22°C), relative to bacteria grown at 35°C and infected murine blood. Evaluation of the 3’ end of lp150 identified a cluster of ORFs that code for putative surface lipoproteins. With a microbe’s surface proteome serving important roles in pathogenesis, we confirmed the ORFs expression in vitro and in the tick compared to spirochetes infecting murine blood. Transcriptional evaluation of lp150 indicates the plasmid likely has essential roles in vector colonization and/or initiating mammalian infection. These results also provide a much needed transcriptional framework to delineate the molecular mechanisms utilized by relapsing fever spirochetes during their enzootic cycle.

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<![CDATA[Oral Immunization with OspC Does Not Prevent Tick-Borne Borrelia burgdorferi Infection]]> https://www.researchpad.co/article/5989d9e8ab0ee8fa60b6c0cf

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.

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