BETA-LACTAMASE AND WOUND INFECTION PATHOGENESIS

尽管在手术中常规使用预防性抗体， 金黄色葡萄球菌伤口感染每年发生在美国。 而感染相关的S.金黄色葡萄球菌通常表现为 在常规药敏试验、专门试验中对抗生素敏感 证明它们通常能够有效地水解 抗生素体外 临床研究表明葡萄球菌存活率 接种到伤口后，部分由β-内酰胺酶介导， 预防性抗生素在体内介导的降解。 主要 目的是阐明β-内酰胺酶在 对S.金黄色伤口脓毒症。 最近的生物化学研究 表明有八种动力学上可区分的S。金黄色 β-内酰胺酶 根据初步数据，假设 尽管表现出明显的结构相似性， 不同β-内酰胺酶侧翼部分的异质性 活性位点裂隙产生了它们的动力学差异。 这一假设将 通过鉴定蛋白质一级结构的差异进行评价， 从核苷酸序列推断，在六个以前未测序的 葡萄球菌β-内酰胺酶的类型。 其次，结构-功能 关系，包括氨基酸取代的鉴定 解释动力学异质性的方法将通过定点 突变和嵌合β-内酰胺酶的构建 与两个β- 内酰胺酶类型。 最后，各种β-内酰胺酶在 介导S.伤口接种后金黄色葡萄球菌存活率 感染将使用皮下脓肿的体内模型进行探索 形成和等基因菌株的S.金黄色葡萄球菌的区别仅在于 和产生的β-内酰胺酶的量。 对《公约》的进一步了解 细菌逃避抗生素的分子水平 预防感染可能有助于制定减少发生的策略 这些病态和潜在致命的感染。

Despite the routine use of prophylactic antibodies in surgery, 35,000 Staphylococcus aureus wound infections occur annually in the United States. Whereas infection-associated strains of S. aureus usually appear to be antibiotic-susceptible in routine susceptibility tests, specialized assays demonstrate that they are often capable of efficiently hydrolyzing the antibiotic in vitro. Clinical studies suggest that staphylococcal survival after inoculation into the wound is mediated, in part, by beta-lactamase- mediated degradation of the prophylactic antibiotic in vivo. The major objective of the proposed work is to clarify the role of beta-lactamase in the pathogenesis of S. aureus wound sepsis. Recent biochemical studies suggest that there are eight kinetically-distinguishable types of S. aureus beta-lactamase. Based upon preliminary data, it is hypothesized that despite exhibiting marked structural similarity, small regions of heterogeneity in the portions of the various beta-lactamase that flank the active site cleft produce their kinetic differences. This hypothesis will be evaluated by identifying the differences in primary protein structure, as deduced from the nucleotide sequence, of the six previously-unsequenced types of staphylococcal beta-lactamase. Subsequently, structure-function relationships including identification of the amino acid substitutions(s) that account for kinetic heterogeneity will be explored by site-directed mutagenesis and the construction of chimeric beta-lactamases after recombination of portions of the structural gene associated with two beta- lactamase types. Finally, the role of the various beta-lactamases in mediating S. aureus survival following wound inoculation with subsequent infection will be explored using an in vivo model of subcutaneous abscess formation and isogenic strains of S. aureus that differ only in the type and amount of beta-lactamase produced. An improved understanding at the molecular level of the means by which bacteria evade antibiotics given to prevent infection may help in developing strategies to reduce the occurance of these morbid and potentially mortal infections.