Liu Ming-Che, Tsai Yi-Lin, Huang Yi-Wei, Chen Hsing-Yu, Hsueh Po-Ren, Lai Szu-Yu, Chen Li-Chia, Chou Yi-Hwa, Lin Wen-Yuan, Liaw Shwu-Jen
Department and Graduate Institute of Clinical Laboratory Sciences and Medical Biotechnology, College of Medicine, National Taiwan University, Taipei, Taiwan, Republic of China.
Department of Biotechnology and Laboratory Science in Medicine, National Yang-Ming University, Taipei, Taiwan, Republic of China.
PLoS One. 2016 May 9;11(5):e0153753. doi: 10.1371/journal.pone.0153753. eCollection 2016.
Stenotrophomonas maltophilia, a gram-negative bacterium, has increasingly emerged as an important nosocomial pathogen. It is well-known for resistance to a variety of antimicrobial agents including cationic antimicrobial polypeptides (CAPs). Resistance to polymyxin B, a kind of CAPs, is known to be controlled by the two-component system PhoPQ. To unravel the role of PhoPQ in polymyxin B resistance of S. maltophilia, a phoP mutant was constructed. We found MICs of polymyxin B, chloramphenicol, ampicillin, gentamicin, kanamycin, streptomycin and spectinomycin decreased 2-64 fold in the phoP mutant. Complementation of the phoP mutant by the wild-type phoP gene restored all of the MICs to the wild type levels. Expression of PhoP was shown to be autoregulated and responsive to Mg2+ levels. The polymyxin B and gentamicin killing tests indicated that pretreatment of low Mg2+ can protect the wild-type S. maltophilia from killing but not phoP mutant. Interestingly, we found phoP mutant had a decrease in expression of SmeZ, an efflux transporter protein for aminoglycosides in S. maltophilia. Moreover, phoP mutant showed increased permeability in the cell membrane relative to the wild-type. In summary, we demonstrated the two-component regulator PhoP of S. maltophilia is involved in antimicrobial susceptibilities and low Mg2+ serves as a signal for triggering the pathway. Both the alteration in membrane permeability and downregulation of SmeZ efflux transporter in the phoP mutant contributed to the increased drug susceptibilities of S. maltophilia, in particular for aminoglycosides. This is the first report to describe the role of the Mg2+-sensing PhoP signaling pathway of S. maltophilia in regulation of the SmeZ efflux transporter and in antimicrobial susceptibilities. This study suggests PhoPQ TCS may serve as a target for development of antimicrobial agents against multidrug-resistant S. maltophilia.
嗜麦芽窄食单胞菌是一种革兰氏阴性菌，已日益成为一种重要的医院病原体。它因对包括阳离子抗菌肽（CAPs）在内的多种抗菌剂具有抗性而闻名。已知对一种CAPs——多粘菌素B的抗性由双组分系统PhoPQ控制。为了阐明PhoPQ在嗜麦芽窄食单胞菌对多粘菌素B抗性中的作用，构建了一个phoP突变体。我们发现，phoP突变体中多粘菌素B、氯霉素、氨苄青霉素、庆大霉素、卡那霉素、链霉素和壮观霉素的最低抑菌浓度（MIC）降低了2至64倍。用野生型phoP基因对phoP突变体进行互补，可使所有MIC恢复到野生型水平。结果表明，PhoP的表达是自动调节的，并对Mg2+水平有反应。多粘菌素B和庆大霉素杀菌试验表明，低Mg2+预处理可保护野生型嗜麦芽窄食单胞菌不被杀死，但对phoP突变体无效。有趣的是，我们发现phoP突变体中嗜麦芽窄食单胞菌氨基糖苷类外排转运蛋白SmeZ的表达有所降低。此外，相对于野生型，phoP突变体的细胞膜通透性增加。总之，我们证明了嗜麦芽窄食单胞菌的双组分调节因子PhoP参与抗菌敏感性，低Mg2+作为触发该途径的信号。phoP突变体中膜通透性的改变和SmeZ外排转运蛋白的下调都导致了嗜麦芽窄食单胞菌对药物敏感性的增加，尤其是对氨基糖苷类药物。这是首次报道描述嗜麦芽窄食单胞菌的Mg2+感应PhoP信号通路在调节SmeZ外排转运蛋白和抗菌敏感性中的作用。这项研究表明，PhoPQ双组分系统可能作为开发针对多重耐药嗜麦芽窄食单胞菌的抗菌剂的靶点。
