Lin Ming-Feng, Tsai Pei-Wen, Chen Jeng-Yi, Lin Yun-You, Lan Chung-Yu
Department of Medicine, National Taiwan University Hospital Chu-Tung Branch, Hsin-Chu County, Taiwan.
Institute of Molecular and Cellular Biology, National Tsing Hua University, Hsin-Chu City, Taiwan.
PLoS One. 2015 Oct 20;10(10):e0141107. doi: 10.1371/journal.pone.0141107. eCollection 2015.
Multidrug-resistant Acinetobacter baumannii has recently emerged as an important pathogen in nosocomial infection; thus, effective antimicrobial regimens are urgently needed. Human antimicrobial peptides (AMPs) exhibit multiple functions and antimicrobial activities against bacteria and fungi and are proposed to be potential adjuvant therapeutic agents. This study examined the effect of the human cathelicidin-derived AMP LL-37 on A. baumannii and revealed the underlying mode of action. We found that LL-37 killed A. baumannii efficiently and reduced cell motility and adhesion. The bacteria-killing effect of LL-37 on A. baumannii was more efficient compared to other AMPs, including human ß-defensin 3 (hBD3) and histatin 5 (Hst5). Both flow cytometric analysis and immunofluorescence staining showed that LL-37 bound to A. baumannii cells. Moreover, far-western analysis demonstrated that LL-37 could bind to the A. baumannii OmpA (AbOmpA) protein. An ELISA assay indicated that biotin-labelled LL-37 (BA-LL37) bound to the AbOmpA74-84 peptide in a dose-dependent manner. Using BA-LL37 as a probe, the ~38 kDa OmpA signal was detected in the wild type but the ompA deletion strain did not show the protein, thereby validating the interaction. Finally, we found that the ompA deletion mutant was more sensitive to LL-37 and decreased cell adhesion by 32% compared to the wild type. However, ompA deletion mutant showed a greatly reduced adhesion defect after LL-37 treatment compared to the wild strain. Taken together, this study provides evidence that LL-37 affects A. baumannii through OmpA binding.
多重耐药鲍曼不动杆菌最近已成为医院感染中的一种重要病原体；因此，迫切需要有效的抗菌方案。人抗菌肽（AMPs）具有多种功能以及针对细菌和真菌的抗菌活性，被认为是潜在的辅助治疗剂。本研究检测了人源cathelicidin衍生的抗菌肽LL-37对鲍曼不动杆菌的作用，并揭示了其潜在的作用方式。我们发现LL-37能有效杀死鲍曼不动杆菌，并降低其细胞运动性和黏附性。与其他抗菌肽相比，包括人β-防御素3（hBD3）和组蛋白5（Hst5），LL-37对鲍曼不动杆菌的杀菌效果更显著。流式细胞术分析和免疫荧光染色均显示LL-37能与鲍曼不动杆菌细胞结合。此外，Far-Western分析表明LL-37能与鲍曼不动杆菌外膜蛋白A（AbOmpA）结合。酶联免疫吸附测定（ELISA）表明生物素标记的LL-37（BA-LL37）以剂量依赖的方式与AbOmpA74-84肽结合。以BA-LL37为探针，在野生型中检测到约38 kDa的OmpA信号，但ompA缺失菌株未显示该蛋白，从而验证了两者的相互作用。最后，我们发现ompA缺失突变体对LL-37更敏感，与野生型相比细胞黏附性降低了32%。然而，与野生菌株相比，ompA缺失突变体在LL-37处理后的黏附缺陷明显减少。综上所述，本研究提供了证据表明LL-37通过与OmpA结合来影响鲍曼不动杆菌。
