Identification and analysis of compensatory mutations that support the evolution of antibiotic resistance in Neisseria gonorrhoeae

支持淋病奈瑟菌抗生素耐药性进化的补偿突变的鉴定和分析

• 批准号: 10219082
• 负责人: Yonatan H Grad
• 金额: $ 74.84万
• 依托单位: HARVARD SCHOOL OF PUBLIC HEALTH
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-07-17 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10219082
• 关键词: Active Sites; Address; Alleles; Anabolism; Animal Model; Animals; Antibiotic Resistance; Antibiotic Therapy; Antibiotic susceptibility; Antibiotics; Back; Binding; Biochemical; Biology; Ceftriaxone; Cell physiology; Ciprofloxacin; Clinical; Coculture Techniques; Collection; Data; Dependence; Diagnosis; Diagnostic; Disease; Drug resistance; Essential Genes; Evolution; Female; Financial compensation; Genes; Genetic; Genetic Epistasis; Genome; Genomics; Goals; Gonorrhea; Growth; Guidelines; Human; In Vitro; Infection; Knowledge; Lethal Genes; Link; Maintenance; Mediating; Methods; Modeling; Molecular; Morphology; Mus; Mutate; Mutation; Neisseria gonorrhoeae; Organism; Pathogenesis; Pathway interactions; Phenotype; Phylogeny; Physiological; Physiology; Population; Predisposition; Prevention; Public Health; Research Personnel; Resistance; Testing; Thiamine; Vaccines; Variant; bacterial fitness; base; clinically relevant; co-infection; cost; drug-resistant gonorrhea; experimental analysis; fitness; genome sequencing; improved; infection rate; insight; large datasets; metabolomics; mouse model; mutant; novel strategies; quinolone resistance; reproductive tract; resistance allele; resistance gene; resistance mutation; resistant strain; success; surveillance strategy; transcriptomics; whole genome

PROJECT SUMMARY The increasing rate of infection and spread of antibiotic resistance in Neisseria gonorrhoeae poses an urgent threat to public health. Knowledge of the pathways and genes that support the emergence and spread of antibi- otic resistance is necessary to develop new strategies for surveillance, diagnosis, and treatment. Despite our understanding of the genes and alleles that confer resistance, significant knowledge gaps remain regarding the factors that contribute to the uneven distribution of resistance across the gonococcal species phylogeny. A core issue that remains poorly explored is the impact of resistance determinants on gonococcal fitness: to what extent do resistance determinants impact fitness, and, if they incur a fitness cost, how does the gonococcus adapt and mitigate these costs? In this proposal, we address these gaps through a comprehensive strategy linking experi- mental and computational identification of compensatory mutations with studies of their mechanisms of action. The overall goal of this project is to determine the impact of mutations that increase resistance to the two most clinically relevant antibiotics for treatment of gonorrhea, ciprofloxacin and ceftriaxone, on bacterial fitness. We will achieve this goal through three specific aims. In Aim 1, we will determine the fitness costs of resistance alleles when transformed into susceptible isolates from different niches and with distinct phylogeny and identify compensatory mutations that mitigate these fitness costs through experimental evolution in the female mouse model. We will examine the two most common ciprofloxacin resistance-conferring alleles (gyrAS91F,D95G and parCS87R) in clinical isolates, and four ceftriaxone resistance-conferring alleles (two variants of penA, the lethal target of ceftriaxone, and newly described variants in rpoB and rpoD), and evaluate the dependence of compen- satory pathways on genomic background. In Aim 2, we will leverage our collection of over 7500 gonococcal genomes from clinical isolates for which we have antibiotic-resistance phenotypes and employ population ge- nomics methods to identify potential compensatory mutations and test these in the mouse model. Moreover, we will define the allelic diversity and distribution of candidates identified in Aim 1. In Aim 3, we will determine the mechanism of action of confirmed compensatory mutations arising from the studies in Aims 1 & 2 using an integrative strategy that examines growth and morphology, transcriptomics, metabolomics, and directed studies of biochemical function. We will also build on preliminary data on compensatory mutations in acnB and mleN for ceftriaxone resistance and on the thiamine biosynthesis pathway in gyrA-mediated quinolone resistance. This interdisciplinary project brings together the complementary and non-overlapping expertise of three lead- ing investigators in the biology and genetics of antibiotic resistance in N. gonorrhoeae, linking the mouse model of gonococcal infection (Dr. Jerse), population genomics (Dr. Grad), and biochemical and physiological charac- terization of resistance-related variants (Dr. Nicholas).

项目摘要 淋病奈瑟氏菌的感染率和抗生素耐药性的增长构成紧急 对公共卫生的威胁。了解支持抗体的出现和传播的途径和基因 对于制定监视，诊断和治疗的新策略是必不可少的。尽管我们 了解赋予抗性的基因和等位基因，关于 导致在淋球菌物种系统发育中耐药性分布不平的因素。核心 探索较差的问题是抵抗决定因素对淋球菌健康的影响：多大程度上 阻力决定因素会影响健身，如果它们产生健身成本，淋球菌如何适应和 减轻这些费用？在此提案中，我们通过将实验相关联的全面战略来解决这些差距 - 通过研究其作用机制的研究，对补偿性突变的心理和计算识别。 该项目的总体目标是确定突变的影响，以增加对两者的抵抗力 大多数临床相关的抗生素用于治疗细菌适应性的淋病，环丙沙星和头孢曲松。 我们将通过三个特定目标来实现这一目标。在AIM 1中，我们将确定抵抗的健身成本 等位基因转变为来自不同壁ni的易感分离株并具有独特的系统发育，并识别 通过雌鼠的实验进化来降低这些健身成本的补偿性突变 模型。我们将检查两个最常见的环丙沙星耐药性等位基因（gyras91f，d95g和 临床分离株中的PARCS87R）和四个头孢曲松抗性的等位基因（Pena的两个变体，致死 头孢曲松的靶标，以及在RPOB和RPOD中新描述的变体，并评估组合的依赖性 基因组背景上的卫星途径。在AIM 2中，我们将利用7500多个淋球菌的收藏 我们具有抗生素耐药表型的临床分离株的基因组并采用了种群 识别潜在补偿性突变并在小鼠模型中测试这些方法的方法。而且，我们 将定义AIM 1中确定的候选人的等位基量多样性和分布。在AIM 3中，我们将确定 AIMS 1和2的研究的确认补偿突变的作用机理使用A 研究生长和形态，转录组学，代谢组学和定向研究的综合策略 生化功能。我们还将基于有关ACNB和MLEN中补偿性突变的初步数据 头孢曲松耐药性和硫胺素生物合成途径在Gyra介导的奎诺酮耐药性中。 这个跨学科项目汇集了三个领导者的补充和非重叠专业知识 在淋病链球菌中生物学和遗传学的研究者，将小鼠模型联系起来 淋球菌感染（Jerse博士），人群基因组学（Grad博士）以及生化和生理特征 抗性相关变体的特性（尼古拉斯博士）。