Genome biology of Francisella tularensis populations

描述(申请人提供)：图拉氏方济氏菌是一种高传染性、A组风险的精选细菌剂，有可能被武器化。它是图拉热症的原因。申请者的长期目标是了解驱动图拉氏丝虫四个亚种独特毒力和传播性特征的基因和基因产物的途径。这项R21应用的目的是识别与这种细菌最具毒力的亚种的毒力和传播性有关的基因。核心假设是，毒力和传播力的增加可归因于遗传差异。这一假设将通过追求三个具体目标来验证：1)设计和表征代表图拉氏镰刀菌亚种的参考菌株集合。Tularensis和苏门冬青(Supsp.)具有时间和空间表示的Holarctica；2)构建具有代表性的图拉氏菌和Holarctica亚种的参考芯片。并使用阵列系统地探测菌株集合的基因组多样性；以及3)使用配对末端序列作图来补充来自微阵列研究的数据。将使用的主要途径和方法包括系统地编目特定亚种基因组片段的替代战略和策略。这将使用DNA微阵列分析(从具有代表性的亚种制备的鸟枪库获得的阵列)来完成。Tularens和Holarctica)和一种新的数据分类算法，然后通过Southern blotting和PCR进行DNA序列分析和确认。其次，来自具有代表性的Holarctica菌株的一个窄(10kb)大小的文库将被用来产生成对的末端序列读取，之后该列表将被映射到亚种。图拉尔人基因组序列。这将允许识别特定于亚种的长度差异。这些结果具有集体意义，因为它们有望提供强有力的初步证据，证明毒力和传播力的菌株差异可归因于基因组差异。这些数据有望成为最终的R01水平研究的基础，涉及候选基因的功能分析。这些研究将大大增加我们对这种生物的病理生物学的了解，为疫苗和治疗提供候选基因产品，并为改进的诊断和法医标记的发展提供基因座。

DESCRIPTION (provided by applicant): Francisella tularensis is a highly infectious, risk-group A select bacterial agent that has the potential to be weaponized. It is the cause of tularemia. The applicants' long-term goal is to understand the pathways of genes and gene products that drive unique virulence and transmissibility traits of the four subspecies of F. tularensis. The objective of this R21 application is to identify genes that are responsible for virulence and transmissibility of the most virulent subspecies of this bacterium. The central hypothesis is that increased virulence and transmissibility are attributable to genetic differences. This hypothesis will be tested by pursuing three specific aims: 1) Design and characterize a reference strain collection representing both F. tularensis subsp. tularensis and supsp. holarctica with temporal and spatial representation; 2) Fabricate representative reference microarrays of the tularensis and holarctica subsp. and systematically probe genome diversity of the strain collection using arrays; and 3) Use paired-end sequence mapping to complement data from the microarray-based studies. The principal approaches and methods to be used include alternative strategies and tactics to systematically catalogue subspecies-specific genome segments. This will be accomplished using DNA microarray analyses (arrays derived from shotgun libraries prepared from the representative subsp. tularensis and holarctica) and a novel algorithm for data sorting followed by DNA sequence analyses and confirmation by Southern blotting and PCR. Second, a narrow (10Kb)-size library from a representative holarctica strain will be used to generate paired-end sequence reads, after which the list will be mapped to the subsp. tularensis genome sequence. This will allow length differences that are subspecies-specific to be identified. The outcomes are collectively significant, because they are expected to provide strong preliminary evidence that strain differences in virulence and transmissibility are attributable to genomic differences. Such data are expected to become the foundation for definitive R01-level investigations involving the functional analyses of candidate genes. These studies will significantly increase our understanding of the pathobiology of this organism, provide candidate gene products for vaccine and therapeutics, and provide loci for development of improved diagnostic and forensic markers.