Metabolon, Inc, Durham, North Carolina, United States of America.
PLoS One. 2013 Aug 5;8(8):e71025. doi: 10.1371/journal.pone.0071025. Print 2013.
Methicillin-resistant Staphylococcus aureus (MRSA) has emerged as one of the most important pathogens both in health care and community-onset infections. The prerequisite for methicillin resistance is mecA, which encodes a β-lactam-insensitive penicillin binding protein PBP2a. A characteristic of MRSA strains from hospital and community associated infections is their heterogeneous expression of resistance to β-lactam (HeR) in which only a small portion (≤ 0.1%) of the population expresses resistance to oxacillin (OXA) ≥ 10 µg/ml, while in other isolates, most of the population expresses resistance to a high level (homotypic resistance, HoR). The mechanism associated with heterogeneous expression requires both increase expression of mecA and a mutational event that involved the triggering of a β-lactam-mediated SOS response and related lexA and recA genes. In the present study we investigated the cellular physiology of HeR-MRSA strains during the process of β-lactam-mediated HeR/HoR selection at sub-inhibitory concentrations by using a combinatorial approach of microarray analyses and global biochemical profiling employing gas chromatography/mass spectrometry (GC/MS) and liquid chromatography/mass spectrometry (LC/MS) to investigate changes in metabolic pathways and the metabolome associated with β-lactam-mediated HeR/HoR selection in clinically relevant heterogeneous MRSA. We found unique features present in the oxacillin-selected SA13011-HoR derivative when compared to the corresponding SA13011-HeR parental strain that included significant increases in tricarboxyl citric acid (TCA) cycle intermediates and a concomitant decrease in fermentative pathways. Inactivation of the TCA cycle enzyme cis-aconitase gene in the SA13011-HeR strain abolished β-lactam-mediated HeR/HoR selection demonstrating the significance of altered TCA cycle activity during the HeR/HoR selection. These results provide evidence of both the metabolic cost and the adaptation that HeR-MRSA clinical strains undergo when exposed to β-lactam pressure, indicating that the energy production is redirected to supply the cell wall synthesis/metabolism, which in turn contributes to the survival response in the presence of β-lactam antibiotics.
耐甲氧西林金黄色葡萄球菌（MRSA）已成为医院和社区获得性感染中最重要的病原体之一。耐甲氧西林的前提是 mecA，它编码一种β-内酰胺不敏感的青霉素结合蛋白 PBP2a。来自医院和社区相关感染的 MRSA 菌株的一个特征是它们对β-内酰胺（HeR）的耐药性呈异质性表达，其中只有一小部分（≤0.1%）人群对≥10μg/ml 的苯唑西林（OXA）表达耐药性，而在其他分离株中，大多数人群对高水平（同型耐药性，HoR）表达耐药性。与异质性表达相关的机制既需要 mecA 的表达增加，也需要涉及触发β-内酰胺介导的 SOS 反应和相关的 lexA 和 recA 基因的突变事件。在本研究中，我们通过使用微阵列分析和基于气相色谱/质谱（GC/MS）和液相色谱/质谱（LC/MS）的全局生化谱分析的组合方法，在亚抑菌浓度下研究了 HeR/MRSA 菌株在β-内酰胺介导的 HeR/HoR 选择过程中的细胞生理学，以研究与β-内酰胺介导的 HeR/HoR 选择相关的代谢途径和代谢组的变化在临床上相关的异质 MRSA 中。与相应的 SA13011-HeR 亲本菌株相比，我们发现 SA13011-HoR 衍生物在耐苯唑西林选择中存在独特的特征，包括三羧酸柠檬酸（TCA）循环中间产物的显著增加和发酵途径的相应减少。在 SA13011-HeR 菌株中失活 TCA 循环酶顺乌头酸酶基因消除了β-内酰胺介导的 HeR/HoR 选择，证明了在 HeR/HoR 选择过程中改变的 TCA 循环活性的重要性。这些结果提供了证据，证明了 HeR-MRSA 临床菌株在暴露于β-内酰胺压力下所经历的代谢成本和适应，表明能量产生被重新定向以供应细胞壁合成/代谢，这反过来又有助于在β-内酰胺抗生素存在下的存活反应。
