Defining a Molecular Signature of Uropathogenic Escherichia coli

项目总结 尿路感染(UTIs)是最常见的细菌感染之一，50%的女性受到影响 在他们一生中至少有一次。这些妇女中约有30%将经历反复感染。 大肠埃希氏菌是最常见的原因，占这些感染的75%。尿液中的细菌， 菌尿，是一系列的临床表现，从无症状(ASB)到严重症状(如 如菌血症)。与致泻性大肠杆菌不同，UPEC没有可用于诊断的离散遗传因子 目的。这在诊断中造成了一个瓶颈，当区分无症状的尿路定殖者和 致病分离物。这对老年人或残疾人等患者群体来说尤其具有挑战性 无法传达症状的存在。识别诊断分子的必要性很大 UPEC签名。虽然没有定义UPEC的离散基因，但我们之前已经展示了一种新陈代谢 区分ASB和膀胱炎分离株的特征。这些代谢物中有许多与嘌呤有关。 新陈代谢。虽然已知嘌呤的生物合成与几种细菌的发病有关 在病原体中，嘌呤挽救的作用在很大程度上仍不清楚。这项提议的目标是识别一种分子 UPEC的签名。为了确定潜在的焦点区域，我们首先评估了核苷酸中间体， 次黄嘌呤，随着时间的推移而被吸收。我们的初步数据显示ASB分离株摄取次黄嘌呤的速度快于 膀胱炎分离株。这个项目的主要假设是，微妙的基因组特征决定了 嘌呤代谢的调节，构成了UPEC的定义和诊断遗传特征。至 检验这一假设，我将使用转录组学、蛋白质组学和计算基因组学来评估潜在的 嘌呤新陈代谢的分子特征。目标1中的实验将研究次黄嘌呤的作用 在明确定义的小鼠模型中进行尿路感染时的抢救。我将探索膜蛋白和 在感染过程中参与嘌呤代谢的已知基因的转录丰度，并使用靶向 比较基因组学来寻找这些途径的基因操纵子的变异。在《目标2》中，我将选择一个宽阔的， 无偏基因组学方法识别可能构成分子的其他潜在基因组区域 签名。我将利用储存在我们微生物库中的分离物，与未确认的医疗记录相联系，以及 对他们进行测序和全基因组关联研究(GWAS)。总而言之，这些数据将提供 UPEC的严格定义，同时研究潜在的分子特征。此外，这些研究将 明确嘌呤挽救在UPEC发病机制中的作用。拟议的工作将在以下方面产生广泛影响 嘌呤代谢在细菌发病机制中的作用以及微生物GWA在患者中的新应用 表型。

PROJECT SUMMARY Urinary tract infections (UTIs) are among one of the most common bacterial infections, afflicting 50% of women at least once in their lifetime. Approximately 30% of these women will experience a recurrent infection. Escherichia coli is the most common cause and is responsible for 75% of these infections. Bacteria in the urine, bacteriuria, is a spectrum of clinical presentations, ranging from asymptomatic (ASB), to severe symptoms(such as bacteremia). Unlike diarrheagenic E. coli, UPEC do not have a discrete genetic factor to use for diagnostic purposes. This creates a bottleneck in diagnostics when discerning an asymptomatic urocolonizer from a pathogenic isolate. This is particularly challenging for patient groups such as the elderly or disabled who are unable to communicate the presence of symptoms. There is a significant need to identify a diagnostic molecular signature of UPEC. While there is no discrete gene that defines UPEC, we have previously shown a metabolic signature that does differentiate ASB and cystitis isolates. Many of these metabolites are associated with purine metabolism. While it is known that purine biosynthesis is implicated in the pathogenesis of several bacterial pathogens, the role of purine salvage remains largely unknown. The goal of this proposal is to identify a molecular signature for UPEC. To identify potential areas of focus, we first assessed how the nucleotide intermediate, hypoxanthine, is taken up over time. Our preliminary data show ASB isolates uptake hypoxanthine faster than cystitis isolates. The overarching hypothesis of this project is that subtle genomic features dictate the regulation of purine metabolism, constituting a defining and diagnostic genetic signature for UPEC. To test this hypothesis, I will use transcriptomics, proteomics, and computational genomics to assess the potential of purine metabolism as a molecular signature. Experiments in Aim 1 will investigate the roles of hypoxanthine salvage during a UTI in a well-defined murine model. I will explore differences in membrane proteins and transcript abundance of known genes involved in purine metabolism throughout infection, and use targeted comparative genomics to look for variations in the gene operons of these pathways. In Aim 2, I will take a broad, unbiased genomics approach to identify other potential genomic regions that could constitute a molecular signature. I will utilize isolates banked at our microbial biobank, linked to the de-identified medical record, and subject them to sequencing and a genome-wide association study (GWAS). Collectively, these data will provide a rigorous definition of UPEC, while investigating a potential molecular signature. Additionally, these studies will define the role of purine salvage in UPEC pathogenesis. The proposed work will have broad implications in purine metabolism in bacterial pathogenesis as well as the novel use of microbial GWAS with patient- phenotypes.