School of Pharmacy, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong.
Department of Microbiology, Faculty of Medicine, The Chinese University of Hong Kong, Hong Kong.
Int J Biol Macromol. 2024 Oct;278(Pt 2):134670. doi: 10.1016/j.ijbiomac.2024.134670. Epub 2024 Aug 14.
Endolysins (lysins), a novel class of antibacterial agents derived from bacteriophages, efficiently lyse bacteria by degrading the peptidoglycan layer within the bacterial wall. Colistin, a classic peptide antibiotic with the ability to permeabilize the outer membrane, has recently shown great promise in synergizing with lysins against gram-negative bacteria. However, the exact mechanisms responsible for their synergy remain unclear. Here, we first demonstrated the synergistic bacterial killing of various lysin and colistin combinations. With a model lysin, LysAB2, we then confirmed that there is a threshold concentration of colistin causing sufficient permeabilization of the outer membrane for lysin to access the peptidoglycan layer and subsequently exert its lytic ability. The threshold colistin concentrations were found to range 0.2-0.8 μM for the tested bacteria, with the exact value largely depending on the density of lipopolysaccharides on the outer membrane. Beyond the threshold colistin level, LysAB2 could synergize with colistin at a concentration as low as 0.31 μM. Next, we proved for the first time that lysin-induced degradation of the peptidoglycan layer facilitated the disruption of cytoplasmic membrane by colistin, elevated the level of reactive oxygen species in bacterial cells, and boosted the killing effect of colistin. Additionally, the colistin-lysin combination could effectively eliminate established biofilms due to the biofilm dispersal ability of lysin. The in-vivo efficacy was preliminary confirmed in a Galleria mellonella infection model for combination with colistin doses (≥ 1.8 μg/larvae), which could reach beyond the threshold concentration, and a fixed LysAB2 dose (10 μg/larvae). In summary, our study provided the first experimental evidence unravelling the mechanisms behind the synergy of colistin and lysins. All these findings provided important insights in guiding the dosing strategy for applying this combination in future development.
溶菌素（lysin）是一类来源于噬菌体的新型抗菌剂，通过降解细菌细胞壁中的肽聚糖层，有效地裂解细菌。黏菌素是一种经典的肽类抗生素，具有破坏外膜的能力，最近在与溶菌素协同作用对抗革兰氏阴性菌方面显示出巨大的潜力。然而，其协同作用的确切机制尚不清楚。在这里，我们首先证明了各种溶菌素和黏菌素组合的协同杀菌作用。然后，我们使用模型溶菌素 LysAB2 证实，存在一个引起外膜足够通透性的黏菌素阈值浓度，使溶菌素能够进入肽聚糖层并随后发挥其裂解能力。发现测试细菌的阈值黏菌素浓度范围为 0.2-0.8 μM，确切值在很大程度上取决于外膜上脂多糖的密度。超过阈值黏菌素水平后，LysAB2 可以在低至 0.31 μM 的浓度下与黏菌素协同作用。接下来，我们首次证明溶菌素诱导的肽聚糖层降解有助于黏菌素破坏细胞质膜，增加细菌细胞内活性氧的水平，并增强黏菌素的杀菌效果。此外，由于溶菌素具有生物膜分散能力，该黏菌素-溶菌素组合可以有效消除已建立的生物膜。在与黏菌素剂量（≥1.8μg/幼虫）结合的 Galleria mellonella 感染模型中，初步证实了体内疗效可以达到阈值浓度以上，并且固定的 LysAB2 剂量（10μg/幼虫）。综上所述，我们的研究提供了第一个实验证据，揭示了黏菌素和溶菌素协同作用的机制。所有这些发现为指导未来开发中应用该组合的剂量策略提供了重要的见解。
