Antibiotic resistance and metabolic pathways in Chlamydia species

项目3-衣原体的抗生素耐药性和代谢途径 该项目将重点研究衣原体的关键生物合成和代谢功能，因为它们对药物有影响 在寄主中的抗性和生长和存活。它包括三个目标： 具体目标1-建立治疗丙型肝炎首选药物出现耐药性的模型。 沙眼衣原体感染。我们将在体外和体外检测阿奇霉素耐药(Azm*^)突变体的适合性 在活体内。然后我们将筛选体内出现的代偿突变体，并对这些突变体进行表征 使用全基因组测序技术。我们还将选择对四环素自发耐药(TC*^) 豚鼠自然感染模型中的鼠疫杆菌突变株(GPIC)。在自然界中出现的突变 将分析感染环境中的生长特征，如组织培养和竞争中的生长 在没有抗生素的情况下进行体内和体外实验。作为动物模型研究的补充， 作为对耐药菌株的调查，临床设置为“治疗失败”(临床核心)。 特定目标2-生物合成和代谢途径空洞填充-表征生物化学和 衣原体必需的生物合成和代谢途径的遗传成分。这一目标将 将该项目的抗生素耐药性目标与生物合成途径的漏洞填充联系起来，重点放在 肽多聚糖悖论和转肽酶。我们将确定C. 沙眼衣原体对β-内酰胺类抗生素(阿莫西林/青霉素)的耐药性。新陈代谢途径空洞填充成分 这一目标将解决衣原体尚不完整的三个基本生长因子 合成：叶酸、利波酸和NAD。在每种情况下，我们都将检查衣原体表达的可能性 这些因子从头合成的酶，或者它们是否表达了它们的新的运输系统 从寄主细胞液中摄取。遗传和生化方法将用于替代细菌 这是由于无法利用基因技术在衣原体中产生突变体而造成的。 不同病毒感染过程中GPIC基因表达谱的特异靶3转录组定位 豚鼠的解剖部位(眼部与生殖器)。豚鼠将感染两种不同的克隆体 眼睛和生殖器部位的GPIC菌株和微阵列将用于确定特定的利基基因 表达配置文件。要比较的表达谱是：1)眼睛部位与生殖器部位；2)生殖器部位 受感染的男性与生殖器感染的女性。将选择高度差异表达的基因/途径 以探索途径/酶为目标的详细的体外分析(后腹学) 抑制剂和最终的新疗法。

Project 3 - Antibiotic resistance and metabolic pathways in Chlamydia spp This project will focus on key biosynthetic and metabolic functions of Chlamydia as they impact on drug resistance and growth and survival in the host. It consists of three aims: Specific Aim 1 - To model the emergence of resistance to the drugs of choice for the treatment of C. trachomatis infections. We will measure the fitness of azithromycin resistant (AZM*^) mutants in vitro and in vivo. We will then screen for compensatory mutants that arise in vivo and characterize these mutants using whole genome sequencing technology. We will also select for spontaneous tetracycline resistant (Tc*^) mutants of C. caviae (GPIC) in a natural infection model in guinea pigs. The mutants that arise in the natural infection setting will be analyzed for growth characteristics such as growth in tissue culture and competition experiments in vivo and in vitro in the absence of antibiotic. A complement to the animal model studies will be a survey for drug resistant Isolates the clinical setting among "treatment failures" (Clinical Core). Specific Aim 2 - Biosynthetic and metabolic pathway hole filling - characterize the biochemical and genetic components of essential biosynthetic and metabolic pathways of Chlamydia spp. This aim will link the antibiotic resistance aim of the project with biosynthetic pathway hole filling by focusing on the peptidoglycan paradox and transpeptidatlon. We will determine the molecular basis of sensitivity of C. trachomatis to p-lactam antibiotics (amoxicillin/penicillin). The metabolic pathway hole filling component of this aim will address three essential growth factors for which Chlamydia has Incomplete pathways for synthesis: folate, llpolc acid and NAD. In each case we will examine the possibility that Chlamydia express enzymes for de novo synthesis of these factors or whether they express novel transport systems for their uptake from the host cytosol. Genetic and biochemical approaches will be used in surrogate bacterial systems due to the unavailability of genetic techniques for creating mutants in Chlamydia. Specific Aim 3 - Transcriptome mapping of GPIC gene expression profile during infection in different anatomical sites (ocular vs. genital) in guinea pig. Guinea pigs will be infected with two different clonal strains of GPIC at ocular and genital sites and microarrays will be used to determine niche specific gene expression profiles. The expression profiles to be compared are: 1) ocular site vs. genital site; 2) genitally infected males vs. genitally infected females. Highly differentially expressed genes/pathways will be chosen for detailed in vitro analysis (metabdomics) with the goal of exploring pathway/enzyme as targets for inhibitors and eventually new therapeutics.