权益分类	功能权益	普通用户	{{item.name}}会员
{{category.name}}	{{benefitItem.name}}

Identification of daptomycin resistance mechanisms in Clostridioides difficile

艰难梭菌达托霉素耐药机制的鉴定

基本信息

批准号：
10688123
负责人：
Craig D Ellermeier
金额：
$ 46.3万
依托单位：
UNIVERSITY OF IOWA
依托单位国家：
美国
项目类别：
财政年份：
2011
资助国家：
美国
起止时间：
2011-02-15 至 2026-07-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10688123
关键词：
Anabolism Antibiotic Resistance Antibiotics Bacillus subtilis Binding Biogenesis Bioinformatics CRISPR interference Categories Cell Wall Cell membrane Cells Cessation of life Charge Clostridium Clostridium difficile Cysteine Daptomycin Data Development Essential Genes Extracellular Domain Foundations Genes Glucosamine Glycolipids Glycopeptide Antibiotics Goals Health Hospitals Human Immune system Immunologic Factors Individual Infection Lipids Mediating Membrane Methods Metronidazole Modeling Nursing Homes Operon Penicillin-Binding Proteins Peptidoglycan Peptidyltransferase Persons Pharmaceutical Preparations Phase III Clinical Trials Phenotype Phosphatidylglycerols Play Polysaccharides Production Proteins Public Health Publishing Regulon Reporter Resistance Role Signal Transduction Stress Structure System Testing United States Vancomycin biological adaptation to stress cell envelope crosslink experimental study follow-up gain of function genetic approach insight knock-down loss of function member membrane biogenesis mutant new therapeutic target novel novel therapeutics overexpression programs resistance gene resistance mechanism response tool transcriptome sequencing transposon sequencing

项目摘要

Project Summary Clostridioides (Clostridium) difficile causes nearly 500,000 infections a year in the United States, leading to nearly 30,000 deaths. The CDC has declared C. difficile an “urgent” threat to public health, the highest threat category. New treatments are sorely needed. Many of our most useful antibiotics target the cell envelope. However, little is known about cell envelope biogenesis or cell envelope stress response in C. difficile. These cell envelope stress response systems may play an important role in antibiotic resistance. By understanding how C. difficile responds to cell envelope stress we may uncover better treatment options for C. difficile infections. Surotomycin, a daptomycin derivative, which targets peptidoglycan synthesis showed promise as a potential treatment for C. difficile. However, conflicting phase 3 clinical trials have halted development of surotomycin as a treatment for C. difficile infections. We used multiple genetic approaches to identify key factors involved in daptomycin resistance. By isolating spontaneous daptomycin resistant mutants and performing Tn-seq we identified two different two component regulatory systems, DraRS and DapRS, that are involved in daptomycin resistance. We will pursue three specific aims to dissect the role of these regulators in controlling daptomycin resistance and cell envelope stress response. In Aim 1 we will define how the DraRS regulatory system contributes to antibiotic resistance. We will test both gain-of-function and loss-of-function draRS mutants for their effect on resistance to a number of cell envelope stresses and on cell envelope biogenesis. We will also define the DraR regulon and determine the contribution of individual genes to antibiotic resistance and cell envelope biogenesis and stress response. In Aim 2 we will dissect how activity of DraR is controlled in response to cell envelope stress. Our preliminary data suggest DraRS is activated by antibiotics disrupt the lipid-II cycle. We will use CRISPRi to test this model by genetically recapitulating the effects of antibiotics and determining the effect on DraR activation. We will also isolate mutants of DraS that are unable to respond to cell envelope stress. In Aim 3 we will define the role of DapRS and the DapRS-regulated genes hexSDF in cell envelope biogenesis. Our data suggest they are required for production of a unique glycolipid which makes up 16% of the polar lipids in the C. difficile membrane. We will define the DapR regulon and how individual regulon members contribute to daptomycin resistance, cell charge, and lipid content and cell envelope stress response. Together these aims will advance our understanding of C. difficile by defining how it resists cell-wall acting antibiotics like daptomycin and vancomycin.

项目摘要艰难梭菌每年在美国造成近50万人感染，导致近 3万人死亡。美国疾病控制与预防中心宣布艰难梭菌是对公众健康的“紧急”威胁，属于最高威胁类别。迫切需要新的治疗方法。我们许多最有用的抗生素都是针对细胞膜的。然而，几乎没有已知艰难梭菌的细胞膜生物发生或细胞膜应激反应。这些细胞包膜应激反应系统可能在抗生素耐药性中发挥重要作用。通过了解艰难梭菌是如何对细胞膜压力的反应我们可能会发现治疗艰难梭菌感染的更好的选择。索洛托霉素，一种针对肽聚糖合成的达托霉素衍生物显示出有望成为治疗C. 艰难抉择。然而，相互冲突的3期临床试验已经阻止了苏托霉素作为治疗糖尿病的药物的发展。艰难梭菌感染。我们使用多种遗传方法来确定与达托霉素有关的关键因素。抵抗。通过分离自发的达托霉素抗性突变株并进行TN-SEQ，我们鉴定了两个参与达托霉素耐药的两个不同的调控系统，即DRARS和DAPRS。我们将追求三个具体目标来剖析这些调节器在控制达托霉素耐药性和细胞包膜应激反应。在目标1中，我们将定义DRRS监管系统如何对抗生素做出贡献抵抗。我们将测试功能获得和功能丧失的draRS突变体对一些细胞被膜压力和对细胞被膜生物发生的影响。我们还将定义Drar规则和确定单个基因在抗生素耐药性、细胞膜生物发生和应激中的作用回应。在目标2中，我们将剖析DRR活性是如何在响应细胞被膜压力时被控制的。我们的初步数据显示，抗生素激活了DRRS，扰乱了脂质-II循环。我们将使用CRISPRi测试这个模型通过基因重现抗生素的作用并确定其对DRAR激活的影响。我们还将分离不能对细胞包膜压力做出反应的DRAS突变体。在目标3中，我们将定义 DapRS及其调控基因在细胞被膜生物发生中的作用我们的数据表明他们是生产一种独特的糖脂所必需的，这种糖脂占艰难梭菌中极性脂的16% 薄膜。我们将定义DAPR调节子以及单个调节子成员如何对达托霉素做出贡献抗性、细胞电荷、脂质含量和细胞包膜应激反应。这些目标将通过定义艰难梭菌如何抵抗细胞壁作用来促进我们对艰难梭菌的理解达托霉素和万古霉素等抗生素。