Genetic basis for persistence of Borrelia burgdorferi

摘要 在脊椎动物宿主中建立持续感染所需的遗传因素是最少的因素之一。 了解伯氏疏螺旋体感染的各个方面。用于建立持久性的多个触发器 已提出然而，由于缺乏敏感的血清学和分子检测， 研究持续性螺旋体的生物学及其在治疗后莱姆病中的直接作用的工作 综合征（PTLDS）。我们已经开发了两种检测方法，克服了这些主要的限制，可以促进 持续感染的研究。TBDCapSeq是下一代捕获测序测定法， 灵敏度上级于定量PCR，并能进行B。宿主组织样品中的伯氏螺旋体基因组组装。 TBD-血清芯片是一种肽阵列，每次检测可以识别对170，000个线性肽的抗体反应， 为B细胞对B主要抗原的反应提供了前所未有的一瞥。burgdorferi。本提案中 这些检测将用于三个特定目标，将检查一个人的分子和遗传足迹。 持久性B。伯氏感染在具体目标1中，我们将对以下疾病进行全面的血清学分析： 来自充分表征的PTLDS患者队列的血清和脑脊液。我们将确定所有主要的 免疫反应性表位。这些数据将与以下血清学数据进行区分： 其他非PTLDS莱姆病队列涉及PTLDS的潜在特异性生物标志物。在具体目标2中， 我们将使用TBDCapSeq来识别B的分子足迹。在PTLDS患者队列中的burgdorferi。 TBDCapSeq的上级灵敏度与高测序深度的结合将提供 对B. burgdorferi和PTLDS。在具体目标3中，我们 建立B三种不同基因型动物模型。C_3H/HeN、BALB/c和穿孔鼠中的burgdorferi 白足鼠每个B的容量。用于建立持久性的burgdorferi基因型， 将评估抗生素治疗，并将使用TBDCapSeq来鉴定 持久性螺旋体沿着基因组分析，我们将进行第一次转录组研究， 持久性螺旋体结合起来，这些目标将提供深入了解患者的免疫反应， PTLDS和识别螺旋体持续存在的机制触发因素。

ABSTRACT The genetic factors required to establish a persistent infection in vertebrate hosts is one of the least understood aspects of an infection with Borrelia burgdorferi. Multiple triggers for establishment of persistence have been proposed. However, the lack of sensitive serologic and molecular assays has substantially impeded work examining the biology of persistent spirochetes and their direct role in Post-Treatment Lyme Disease Syndrome (PTLDS). We have developed two assays that overcome these major limitations and can facilitate studies of persistent infections. The TBDCapSeq, a next generation capture sequencing assay, has a sensitivity superior to quantitative PCR and enables B. burgdorferi genome assembly in host tissue samples. The TBD-Serochip, a peptide array that can identify antibody responses to 170,000 linear peptides per assay, provides unprecedented glimpses into B cell responses to the major antigens of B. burgdorferi. In this proposal these assays will be employed in three Specific Aims that will examine molecular and genetic footprints of a persistent B. burgdorferi infection. In Specific Aim 1, we will perform a comprehensive serologic analysis of sera and cerebrospinal fluid from a well characterized PTLDS patient cohort. We will identify all major immunoreactive epitopes in both specimen types. These data will be differentiated with serology data from other non-PTLDS Lyme disease cohorts to implicate potential specific biomarkers of PTLDS. In Specific Aim 2, we will employ the TBDCapSeq to identify molecular footprints of B. burgdorferi in the PTLDS patient cohort. The combination of superior sensitivity of TBDCapSeq coupled with high sequencing depth will provide the most robust molecular analysis of the link between B. burgdorferi and PTLDS. In Specific Aim 3, we will establish an animal model for three distinct genotypes of B. burgdorferi in C3H/HeN, BALB/c and Peromyscus leucopus mice. The capacity of each B. burgdorferi genotype for establishing persistence with and without antibiotic treatment will be assessed and the TBDCapSeq will be used to identify genetic features unique to persistent spirochetes. Along with genomic analysis, we will perform the first transcriptome investigation of persistent spirochetes. Combined, these Aims will provide insight into the immune response of patients with PTLDS and identify mechanistic triggers of spirochete persistence.