Rise of lasR mutant Pseudomonas aeruginosa keratitis

lasR突变型铜绿假单胞菌角膜炎的兴起

基本信息

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
项目类别：
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

项目摘要

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.

项目概要/摘要美国国立卫生研究院 (NIH) 资助的类固醇治疗角膜溃疡研究发现，自然发生的激光引起的角膜炎铜绿假单胞菌突变体导致的视力结果明显比角膜炎差由野生型铜绿假单胞菌引起。所有分离株均来自印度，于 2006 年至 2010 年期间获得。使用匹兹堡大学坎贝尔实验室的巨大储存库，可追溯到 1992 年观察到令人担忧的趋势，lasR 突变体在铜绿假单胞菌角膜炎中急剧增加在美国被隔离。这是一个潜在的公共卫生问题，这项研究将有助于解决与眼科医生和验光师沟通。这项研究的长期目标是开发预防感染引起的视力丧失的方法。该应用程序的总体目标是确定铜绿假单胞菌 lasR 突变体导致更差临床结果的机制。我们的中心假设 LasR 转录因子抑制影响铜绿假单胞菌存活的基因表达粗糙的眼表，使得突变体中上调的基因赋予增加的感染性表型。这这项研究的基本原理是确定 LasR 控制眼部毒力的机制将提供制定对抗视力丧失的治疗策略的科学依据。制定了三个具体目标旨在质疑我们的中心假设：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显性或隐性毒力表型； 2) 测试特定铜绿假单胞菌基因的重要性 lasR 突变体的表达增加，在眼表存活中发挥作用，从而在兔隐形眼镜感染模型，并诱导角膜炎症，3) 测试 lasR 突变体是否存在对眼表先天防御和眼科相关抗生素治疗的抵抗力升高。这研究将结合先进的分子遗传学来操纵细菌基因组，包括临床分离株、下一代测序以及完善的体外和体内模型。这项研究具有创新性因为目前的诊断方法无法区分引起角膜炎的铜绿假单胞菌菌株，除了抗生素敏感性之外，这项研究将引入一种新方法来识别高度视力威胁性铜绿假单胞菌分离株。这项研究还将评估几种候选毒力因子未经过眼睛测试。这项研究的意义基于两点：A）它的高阐明细菌用来建立致盲性眼部感染的新致病机制的潜力，由此产生的知识最终可用于开发预防因眼部疾病引起的视力丧失的方法感染，以及 B) 引入一种可行的个性化医疗形式，可用于提醒医疗保健工作人员和最有视力丧失风险的患者。