Defining the bacterial factors that modulate sensitivity to B-lactam in CA-MRSA

定义调节 CA-MRSA 对 B-内酰胺敏感性的细菌因素

• 批准号: 7754871
• 负责人: Ambrose Lin Yau Cheung
• 金额: $ 23.75万
• 依托单位: DARTMOUTH COLLEGE
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-01-05 至 2011-09-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7754871
• 关键词: Accounting; Affect; Animal Model; Antibiotics; Autolysis; Binding; Binding Proteins; Cefoxitin; Cell Wall; Chromosomes; Communities; Complex; Data; Development; Disease; Elements; Enzymes; Epidemic; Exhibits; Figs - dietary; Genes; Genetic Determinism; Genetic Transcription; Genome; Genomics; Goals; Health; Heel; High Pressure Liquid Chromatography; Hospitals; Human; IS Elements; Individual; Infection; Lactams; Manuscripts; Maps; Mediating; Metabolism; Methicillin; Methicillin Resistance; Mind; Mobile Genetic Elements; Morphology; Nafcillin; Oxacillin; Penicillin Binding Protein 4; Penicillin Resistance; Penicillins; Peptidoglycan; Phenotype; Pilot Projects; Plasmids; Play; Proteins; Regulation; Resistance; Resistance development; Sepsis; Staphylococcus aureus; Virulence; Virulent; antimicrobial; base; beta-Lactams; crosslink; killings; methicillin resistant Staphylococcus aureus; mutant; novel; pathogen; public health relevance; recombinase; response

DESCRIPTION (provided by applicant): Recent cases of infections caused by community-acquired methicillin-resistant Staphylococcus aureus (CA-MRSA) strains in healthy individuals have raised concerns worldwide. CA-MRSA strains differ from hospital-acquired MRSAs (HA-MRSA) by virtue of their genomic background and increased virulence in animal models. Resistance to methicillin in S. aureus is mediated by mecA which is embedded within a mobile genetic element (21-67 kb) called SCCmec. There are five types of SCCmec in MRSA, which differ with respect to the mec complex (mecI-mecR1-mecA), integrated plasmids, transposons, IS elements and other accessory genes. USA300 and MW2 (USA400), the two most common CA-MRSA isolates, carry the SCCmec type IV while USA100 and USA200, which are common HA-MRSAs, contain type II. Recent studies from our lab have shown that a loss of penicillin-binding protein 4 (PBP4) is sufficient to cause a 16-fold reduction in oxacillin and nafcillin resistance in CA-MRSA, but not in HA- MRSA. This finding was confirmed with cefoxitin, a semi-synthetic beta-lactam that binds PBP4 irreversibly, which exhibits synergistic killing with oxacillin against CA-MRSA strains but not against HA-MRSA strains. As all MRSA strains contain mecA, these data indicate that that mecA encoding PBP2A is not the sole determinant of methicillin resistance in CA-MRSA strains. Based on the differences in SCCmec types, we hypothesize that the accessory genes within SCCmec type IV in CA-MRSA strains USA300 and USA400 confer sensitivity to oxacillin (also nafcillin) in the absence of pbp4 or in the presence of cefoxitin. To define the bacterial determinants, we have developed two aims: I) mapping the bacterial determinants that confer sensitivity to oxacillin/nafcillin in the absence of pbp4 or in the presence of PBP4-binding beta-lactam such as cefoxitin; II) characterization of the genetic determinants that confer oxacillin sensitivity in the presence of cefoxitin in CA-MRSA strains USA300 and USA400. With these studies, we will identity the genes in type IV SCCmec responsible for this phenotype (Aim 1). Mutants of these genes in USA300 and USA400 will be constructed and characterized with respect to antibiotic sensitivity, cell wall metabolism and cell wall morphology (Aim II). Upon completion of these studies, we seek to identify genetic determinants that impact oxacillin resistance in CA-MRSA. These genetic determinants may be targets for the development of novel antimicrobial therapy. PUBLIC HEALTH RELEVANCE: Community-acquired methicillin resistant Staphylococcus aureus (CA-MRSA) has become a major health problem. Contrary to hospital-acquired methicillin resistant S. aureus, CA-MRSAs are more virulent and can affect healthy individuals. We have found an "Achilles Heel" in CA-MRSA in that penicillin binding protein 4 (PBP4) is required for methicillin resistance in CA-MRSA. There are genes in CA-MRSA involved in this oxacillin sensitivity upon deletion of the pbp4 gene. Our goal is to identify and characterize these genes that confer oxacillin sensitivity.

描述（由申请人提供）：最近由社区获得性耐甲氧西林金黄色葡萄球菌（CA-MRSA）菌株在健康个体中引起的感染病例引起了全世界的关注。 CA-MRSA 菌株与医院获得性 MRSA (HA-MRSA) 的不同之处在于其基因组背景和动物模型中毒力的增加。金黄色葡萄球菌对甲氧西林的抗性是由 mecA 介导的，mecA 嵌入称为 SCCmec 的可移动遗传元件 (21-67 kb) 中。 MRSA中有五种类型的SCCmec，其不同之处在于mec复合物（mecI-mecR1-mecA）、整合质粒、转座子、IS元件和其他辅助基因。 USA300 和 MW2 (USA400) 是两种最常见的 CA-MRSA 分离株，携带 IV 型 SCCmec，而常见的 HA-MRSA 菌株 USA100 和 USA200 则携带 II 型。我们实验室最近的研究表明，青霉素结合蛋白 4 (PBP4) 的缺失足以导致 CA-MRSA 中苯唑西林和萘夫西林耐药性降低 16 倍，但在 HA-MRSA 中则不然。这一发现得到了头孢西丁的证实，头孢西丁是一种半合成的 β-内酰胺，可不可逆地结合 PBP4，它与苯唑西林对 CA-MRSA 菌株具有协同杀伤作用，但对 HA-MRSA 菌株没有作用。由于所有 MRSA 菌株都含有 mecA，这些数据表明编码 PBP2A 的 mecA 并不是 CA-MRSA 菌株中甲氧西林耐药性的唯一决定因素。基于 SCCmec 类型的差异，我们假设 CA-MRSA 菌株 USA300 和 USA400 中 SCCmec IV 型内的辅助基因在不存在 pbp4 或存在头孢西丁的情况下赋予对苯唑西林（也是萘夫西林）的敏感性。为了定义细菌决定簇，我们制定了两个目标：I) 绘制在不存在 pbp4 或存在 PBP4 结合 β-内酰胺（如头孢西丁）的情况下赋予苯唑西林/萘夫西林敏感性的细菌决定簇； II)在CA-MRSA菌株USA300和USA400中头孢西丁存在下赋予苯唑西林敏感性的遗传决定因素的表征。通过这些研究，我们将鉴定 IV 型 SCCmec 中导致该表型的基因（目标 1）。将构建 USA300 和 USA400 中这些基因的突变体，并对其抗生素敏感性、细胞壁代谢和细胞壁形态进行表征（目标 II）。完成这些研究后，我们寻求确定影响 CA-MRSA 苯唑西林耐药性的遗传决定因素。这些遗传决定因素可能是开发新型抗菌疗法的目标。 公共卫生相关性：社区获得性耐甲氧西林金黄色葡萄球菌 (CA-MRSA) 已成为一个主要的健康问题。与医院获得的耐甲氧西林金黄色葡萄球菌相反，CA-MRSA 的毒性更强，可以影响健康个体。我们发现 CA-MRSA 的“致命弱点”在于 CA-MRSA 的甲氧西林耐药性需要青霉素结合蛋白 4 (PBP4)。 CA-MRSA 中有一些基因在删除 pbp4 基因后与苯唑西林敏感性有关。我们的目标是识别和表征这些赋予苯唑西林敏感性的基因。