Rise of lasR mutant Pseudomonas aeruginosa keratitis

lasR突变型铜绿假单胞菌角膜炎的兴起

• 批准号: 10652442
• 负责人: ROBERT M SHANKS
• 金额: $ 39.82万
• 依托单位: UNIVERSITY OF PITTSBURGH AT PITTSBURGH
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-07-01 至 2025-06-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10652442
• 关键词: Address; Alleles; Animal Model; Antibiotic Resistance; Antibiotic Therapy; Antibiotic susceptibility; Antibiotics; Awareness; Back; Bacteria; Bacterial Antibiotic Resistance; Bacterial Genome; Bacterial Infections; Blindness; Cicatrix; Clinical; Collection; Communication; Complement; Contact Lenses; Cornea; Corneal Ulcer; Disease; Dose; Eye; Eye Infections; Eye diseases; Frequencies; Funding; Gene Expression; Gene Expression Regulation; Genes; Genetic; Genotype; Goals; Growth; Health Personnel; Immune response; Immune system; In Vitro; India; Infection; Inflammation; Inflammatory; Keratitis; Laboratories; Link; Lipase; Measures; Mission; Modeling; Molecular Genetics; Muramidase; Mus; Mutation; Ophthalmologist; Optometrist; Organism; Oryctolagus cuniculus; Outcome; Pathogenesis; Pathogenicity; Patient-Focused Outcomes; Patients; Phenotype; Phylogenetic Analysis; Pilum; Predisposition; Pseudomonas aeruginosa; Public Health; Publishing; Reagent; Research; Resistance; Risk; Role; Signal Induction; Source; Steroids; Testing; Therapeutic; United States; United States National Institutes of Health; Universities; Up-Regulation; Virulence; Virulence Factors; Virulent; Vision; Visual; antibiotic tolerance; antimicrobial peptide; bacterial resistance; clinical diagnostics; combat; cystic fibrosis patients; derepression; design; diagnostic strategy; improved; in vivo; in vivo Model; innovation; knowledge of results; loss of function; mutant; next generation sequencing; novel strategies; novel therapeutic intervention; ocular surface; pathogen; personalized medicine; prevent; quorum sensing; repository; success; targeted sequencing; transcription factor; treatment strategy; trend; whole genome

Project Summary / Abstract The NIH funded Steroids for Corneal Ulcers Study found that keratitis caused by naturally occurring lasR mutants of the bacteria Pseudomonas aeruginosa resulted in significantly worse vision outcomes than keratitis caused by wild-type P. aeruginosa. All the isolates, from India, were obtainted between 2006-2010. Using the vast repository at the University of Pittsburgh Campbell Laboratory, which has isolates back to 1992, a concerning trend was observed that lasR mutants have dramatically increased among P. aeruginosa keratitis isolates in the United States. This is a potential public health problem that this study would help to communicate to ophthalmologists and optometrists. The long-term goal of this research is to develop approaches to prevent vision loss caused by infections. The overall objective of this application is to determine the mechanism by which lasR mutants of P. aeruginosa cause worse clinical outcomes. Our central hypothesis is that the LasR transcription factor inhibits expression of genes that influence P. aeruginosa survival on the harsh ocular surface, such that genes upregulated in the mutant confer an increased infectivity phenotype. The rationale for this study is that identifying the mechanisms by which LasR controls ocular virulence will provide a scientific basis to develop therapeutic strategies to combat vision loss. Three specific aims have been designed to interrogate our central hypothesis: 1) to characterize the genetic basis of lasR mutations among the clinical isolate collection at the Campbell Laboratory, and evaluate the extent to which the mutations confer dominant or recessive virulence phenotypes; 2) to test the importance of specific P. aeruginosa genes that have increased expression in lasR mutants for a role in ocular surface survival, establishing corneal infection in a rabbit contact lens infection model, and in inducing corneal inflammation, and 3) to test whether lasR mutants have elevated resistance to ocular surface innate defenses and ophthalmically relevant antibiotic therapy. This study will use a combination of advanced molecular genetics to manipulate bacterial genomes including clinical isolates, next generation sequencing, and well-established in vitro and in vivo models. This study is innovative because current diagnostic approaches do not differentiate between P. aeruginosa strains that cause keratitis, other than antibiotic susceptibility, and this study will introduce a new approach to identify highly vision threatening P. aeruginosa isolates. This study will also evaluate several candidate virulence factors that have not been tested with respect to the eye. The significance of this study is based on two items: A) its high potential to elucidate new pathogenic mechanisms that bacteria wield to establish blinding ocular infections, and the resulting knowledge can ultimately be used to develop approaches to prevent vision loss due to ocular infections, and B) introducing a feasible form of personalized medicine that can be used to alert health care workers to patients most at risk for vision loss.

项目摘要 /摘要 NIH资助的角膜溃疡类固醇研究发现，天然发生的LASR引起的角膜炎 铜绿假单胞菌的突变体与角膜炎相比，视力结果明显差 由野生型P.铜绿假单胞菌引起。来自印度的所有分离株在2006年至2010年之间均获得了捕获。使用 匹兹堡大学坎贝尔大学实验室的大量存储库，该实验室隔离到1992年， 关于趋势的观察，铜绿假单胞菌角膜炎中的LASR突变体显着增加 在美国的分离株。这是一个潜在的公共卫生问题，这项研究将有助于 与眼科医生和验光师交流。这项研究的长期目标是发展 防止感染引起的视力丧失的方法。该应用程序的总体目的是确定 铜绿假单胞菌的LASR突变体引起较差的临床结果的机制。我们的中心假设 是LASR转录因子抑制了影响铜绿假单胞菌存活的基因的表达 苛刻的眼表，使突变体中上调的基因赋予了增加的感染表型。这 这项研究的基本原理是确定LASR控制眼毒力的机制将提供 科学的基础，以制定治疗策略来打击视力丧失。三个具体目标是 旨在审问我们的中心假设：1）表征LASR突变的遗传基础 坎贝尔实验室的临床分离株收集，并评估突变赋予的程度 主导或隐性毒力表型； 2）测试特定铜绿假单胞菌基因的重要性 在LASR突变体中表达增加，以在眼部表面存活中发挥作用，从而建立角膜感染 兔隐形眼镜感染模型和诱导角膜炎症，以及3）测试拉斯尔突变体是否 对眼表面先天防御和眼科相关的抗生素疗法的耐药性升高。这 研究将使用晚期分子遗传学的组合来操纵细菌基因组，包括临床 分离株，下一代测序以及在体外和体内模型建立良好的。这项研究是创新的 因为当前的诊断方法没有区分引起角膜炎的铜绿假单胞菌菌株，所以 除了抗生素易感性以外，这项研究将引入一种新的方法来识别高度视力 威胁铜绿假单胞菌分离株。这项研究还将评估几种具有的候选毒力因素 没有对眼睛进行测试。这项研究的意义基于两个项目：a） 潜力阐明细菌挥舞着眼睛感染的新致病机制， 最终可以使用所得的知识来开发方法，以防止视力丧失。 感染，b）引入可行的个性化药物形式，可用于警告医疗保健 最有视力丧失风险的患者的工人。