Animal Model Core E

核心E.动物模型核心 项目负责人：克莱尔史密斯 摘要 了解结核分枝杆菌（Mtb）代谢产物影响人类的机制 结核病需要实施既易处理又 忠实地复制在人类中观察到的结核病状态。在这里，我们建议利用协作 交叉（CC），一组遗传多样性重组近交系小鼠，可重复和无限期地 重生了我们以前已经表明，CC面板包括广泛的结核病 特征和感染微环境。CC小鼠提供了一种研究特定结核分枝杆菌的易处理模型 遗传代谢物对，与表现出有限表型变异的标准近交系小鼠相比。为 例如，在全基因组TnSeq实验中，我们发现在19个高价值Mtb代谢基因中，只有 一种在常规C57 BL/6 J（BL 6）小鼠品系中控制生长。然而，超过一半的变种人 研究显示，当在CC小鼠品系中筛选时，体内生长表型。此外，通过研究 宿主遗传背景，其中单个细菌基因控制或不控制体内Mtb存活， Smith实验室可以开始研究宿主因素在控制Mtb应答中的作用。动物核心E将进行 实验感染方法，以支持项目1和3，重点是毒力相关的脂质和 诊断，分别。为了支持Mtb代谢物作为项目3中诊断测试的鉴定，我们将 表征来自感染高和低结核分枝杆菌负荷的小鼠的肺和血清。支持努力 了解项目1中宿主压力对分枝杆菌包膜内容物的作用，我们将生产Mtb 在小鼠体内传代的菌株。扩展我们现有的Tnseq方法来鉴定分枝杆菌基因， 为了在体内控制Mtb生长，我们将选择并测试50个合并的Mtb CRISPR敲低菌株，用于肺结核。 感染CC菌株。从这些中，获得了具有最强体内生长的五种Mtb CRISPR敲低菌株。 表型将作为单基因敲除进一步详细研究，以确定它们的特定功能。

Core E. Animal Model Core Project Leader: Clare Smith ABSTRACT Understanding the mechanisms through which Mycobacterium tuberculosis (Mtb) metabolites impact human tuberculosis (TB) disease requires implementation of model animal systems that are both tractable and faithfully replicate the TB disease states observed in humans. Here we propose to leverage the Collaborative Cross (CC), a genetically diverse panel of recombinant inbred mice that can be reproducibly and indefinitely regenerated. We have previously shown that the CC panel encompasses a broad spectrum of TB disease traits and infection microenvironments. CC mice provide a tractable model in which to study specific Mtb genetic-metabolite pairs, compared to standard inbred mice that show limited phenotypic variability. For example, in a genome-wide TnSeq experiment, we found that among 19 high value Mtb metabolic genes, only one controlled growth in the conventional C57BL/6J (BL6) mouse strain. However, more than half of mutants studied showed in vivo growth phenotypes when screened across CC mouse strains. Further, through study of the host genetic backgrounds in which individual bacterial genes do or do not control in vivo Mtb survival, the Smith laboratory can begin to study host factors in control of Mtb response. The Animal Core E will conduct experimental infection approaches to support Projects 1 and 3 that focus on virulence associated lipids and diagnostics, respectively. To support identification of Mtb metabolites as diagnostic tests in Project 3, we will characterize lungs and serum from mice infected with a high and low burden of Mtb. Supporting efforts to understand the role of host pressure on the mycobacterial envelope content in Project 1, we will produce Mtb strains passaged in vivo in mice. Extending our existing Tnseq approach to identify mycobacterial genes that control Mtb growth in vivo, we will select and test up 50 pooled Mtb CRISPR knockdown strains for pulmonary infection in CC strains. From these, five Mtb CRISPR knockdown strains with the strongest in vivo growth phenotypes will be further studied in detail as single gene knockdowns to determine their specific functions.