Fitness of gram-negative pathogens during bacteremia

我们正在输掉与我们医院和医院的抗药性革兰氏阴性细菌的战斗 临床护理设施。美国疾病控制与预防中心估计，每年有200多万人感染17种猪瘟。 抗药性细菌病原体，每年导致23,000人死亡。超过一半的物种是革兰氏- 耐碳青霉烯肠杆菌科(CRE)及非发酵菌等阴性病原菌 条件致病菌鲍曼不动杆菌。尽管疾控中心描述了碳青霉烯类耐药(CR) 细菌是“噩梦细菌”，这些病原体或其抗生素的毒力所需的因素-- 在血液感染期间易感的对应者在很大程度上是未知的。因此，迫切需要 确定独特的(物种特有的)和共同的(六种革兰氏阴性物种中的大多数需要的)适合性和 菌血症所需的毒力因子，并绘制关键代谢途径和必需基因图谱 这些病原体在体内使用的集合。我们的长期目标是阐明糖尿病的发病机制。 引起医院获得性感染的革兰氏阴性细菌。此应用程序的总体目标是 对肺炎克雷伯氏菌代表菌株进行RNA-SEQ和体内生长速率测定， 小鼠粘质沙雷氏菌、弗劳地柠檬酸杆菌、阴沟肠杆菌和鲍曼不动杆菌 菌血症模型，在该模型中，这些分离物的TN-SEQ已基本完成。我们的中心假设是 根据CR种在科(肠杆菌科)和纲(鲍曼曲霉)水平上的亲缘关系， 这些病原体需要同源核心功能和物种特有的适应因子的组合才能 在菌血症期间获取营养并避开宿主的反应。建议进行这些研究的理由是 导致我们健康中血液感染的革兰氏阴性病原体的抗生素耐药性正在迅速上升 护理系统。我们计划通过以下方式客观地识别对血液感染至关重要的独特和共同的基因 并检测它们在体内的基因表达。独特而常见的毒力 将对决定因素进行调查，以确定发病机制。我们将检验我们的中心假设 并通过完成这些特定目的来实现本申请的目的：1)定义活跃的代谢 菌血症期间六种革兰氏阴性病原菌的生长途径和生长动力学。2)识别 六种革兰氏阴性病原体菌血症所需的共同和独特的途径。具体预期 结果将包括：(A)精确测量每种病原体在菌血症期间的生长动力学，(B) 确定体内相同生长阶段的优先途径和所需途径，以及(C)详细的 这些通路带有病原体特有的基因或在多个病原体之间共享的基因。这个 这些研究的积极影响将是巨大的。我们将揭示我们六个人的致病机制 其中大多数涉及医院耐药病原体，并确定开发新疗法的目标。

We are losing the battle against antibiotic-resistant Gram-negative bacterial pathogens in our hospitals and clinical care facilities. The CDC estimates that over 2 million people are infected annually with 17 species of antibiotic-resistant bacterial pathogens, killing 23,000 people per year. Over half of the species are Gram- negative pathogens including carbapenem-resistant Enterobacteriaceae (CRE) and the non-fermenting opportunistic pathogen Acinetobacter baumannii. Although the CDC described carbapenem-resistant (CR) bacteria as “nightmare bacteria”, the factors required for virulence of these pathogens or their antibiotic- susceptible counterparts during bloodstream infections are largely unknown. Thus, there is an urgent need to identify unique (species-specific) and common (required by most of the six Gram-negative species) fitness and virulence factors required by these species for bacteremia, and map key metabolic pathways and essential gene sets used by these pathogens in vivo. Our long-term goal is to delineate the mechanisms of pathogenesis in Gram-negative bacteria that cause hospital-acquired infections. The overall objective of this application is to conduct RNA-seq and measure in vivo growth rates in representative isolates of E. coli, Klebsiella pneumoniae, Serratia marcescens, Citrobacter freundii, and Enterobacter cloacae as well as A. baumannii, in the murine model of bacteremia in which Tn-seq has been largely completed for these isolates. Our central hypothesis is that based on the relatedness of CR species at the family (Enterobacteriaceae) and class (A. baumannii) levels, these pathogens require a combination of orthologous core functions and species-specific fitness factors to acquire nutrients and evade host responses during bacteremia. The rationale for these proposed studies is that antibiotic resistance is rising rapidly in Gram-negative pathogens that cause bloodstream infections in our health care systems. We plan to objectively identify unique and common genes critical for bloodstream infection by antibiotic-susceptible and CR bacteria and measure their in vivo gene expression. Unique and common virulence determinants will be investigated to determine mechanisms of pathogenesis. We will test our central hypothesis and attain the objective of this application by completing these specific aims: 1) Define the active metabolic pathways and resultant growth kinetics across six Gram-negative pathogens during bacteremia. 2) Identify shared and unique pathways required for bacteremia by six Gram-negative pathogens. Specific expected outcomes will include: (a) precise measurement of growth kinetics of each pathogen during bacteremia, (b) identification of preferred and required pathways at equivalent growth phases in vivo, and (c) detailed maps of these pathways annotated with genes that are pathogen-specific or shared across multiple pathogens. The positive impact of these studies will be substantial. We will uncover mechanisms of pathogenesis for six of our most concerning antibiotic-resistant hospital pathogens and identify targets for developing new therapeutics.