High-throughput sequence analysis of vlsE recombination events in Borrelia burgdorferi

这个项目的目标是使用高通量测序和计算分析来获得更好的 了解VLS抗原变异系统，致病莱姆病细菌通过该系统逃避人类 免疫系统。VLS系统存在于所有莱姆病细菌中，是逃避适应性的必需系统。 机体在感染后的头两周的免疫力和存活率。VLS基因座由单一的 编码表面脂蛋白VlsE的表达位点和连续的沉默DNA盒阵列。穿过 一种鲜为人知的基因转换机制，来自无声磁带的随机DNA片段取代了 表达部位的相应区域，导致VlsE蛋白的变化和宿主的逃避 抗体反应。免疫系统逃避会导致长期感染和可能持续的症状 几个月到几年。将DNA测序与我们的分析软件结合使用，该项目将提供 对VLS抗原中DNA重组事件的最全面、统计准确的分析 引起莱姆病的伯氏疏螺旋体变异系统。由此产生的信息将 帮助理解哺乳动物宿主中莱姆病重组事件所涉及的过程 逃避宿主免疫系统。此外，利用VLS系统开发和演示的方法 后来可以应用于其他使用类似策略躲避人类和动物的致病系统 宿主免疫系统。对这一变异系统的透彻理解对其发展具有重要意义 阻断莱姆病传播周期和预防莱姆病长期表现的策略 疾病。这项研究的单一目的是研究大量的DNA序列(&gt；120万) 从不同时间感染莱姆病的动物中提取，并与最初接种的莱姆病疫苗进行比较 他们被感染了。这些样本之前是从感染莱姆病的小鼠身上采集的 导致克隆7、14、28和90天；来自体外培养和感染组织外植体的样本将被 包括在内以供比较。PacBio DNA测序数据将使用一种新的序列进行比对和分析 分析算法(Varaligner)，深入了解每个VlsE变异群体 样本。在不同的时间点和组织中获得的结果将被比较以评估重组 模式和动力学，这可能揭示这一基因转化过程中涉及的关键序列。这个 开发的软件将提供给科学界进行类似的深入抗原性分析 其他细菌和原生动物病原体的变异系统。

The goal of this project is to use high throughput sequencing and computational analysis to gain a better understanding of the vls antigenic variation system, by which Lyme disease-causing bacteria evade the human immune system. The vls system is found in all Lyme disease bacteria and is required for evasion of adaptive immunity and survival of the organism beyond the first two weeks of infection. The vls locus consists of a single expression site encoding the surface lipoprotein VlsE and a contiguous array of silent DNA cassettes. Through a poorly understood gene conversion mechanism, random DNA segments from the silent cassettes replace the corresponding regions in the expression site, causing changes in the VlsE protein and evasion of the host antibody response. Immune system evasion leads to long-term infection and manifestations that may persist for months to years. Using DNA sequencing in combination with our analysis software, this project will provide the most comprehensive, statistically accurate analysis of the DNA recombination events in the vls antigenic variation system of Borrelia burgdorferi, a bacterium that causes Lyme disease. The resulting information will aid in understanding the processes involved in Lyme disease recombination events in mammalian hosts and evasion of host immune systems. Moreover, the approaches developed and demonstrated with the vls system can later be applied to other pathogenic systems which use similar tactics for evasion of the human and animal host immune systems. A thorough understanding of this variation system is important for the development strategies for interrupting the transmission cycle and preventing debilitating long-term manifestations of Lyme disease. The single aim of this research is to study a large number of DNA sequences (> 1.2 million) derived from animals with differing times of Lyme disease infection, and comparison to the original inoculates with which they were infected. The samples were previously collected from mice infected with Lyme disease causing clones for 7, 14, 28, and 90 days; samples from in vitro cultures and infected tissue explants will be included for comparison. PacBio DNA sequencing data will be aligned and analyzed using a novel sequence analysis algorithm (Varaligner) to provide an in-depth view of vlsE variant population present within each sample. Results obtained in different time points and tissues will be compared to assess the recombination patterns and kinetics, which may reveal the critical sequences involved in this gene conversion process. The software developed will be provided to the scientific community for similar in-depth analyses of antigenic variation systems in other bacterial and protozoal pathogens.