ε-聚赖氨酸对耐甲氧西林金黄色葡萄球菌USA300的作用机制

Antibacterial effect and mechanism of ε-polylysine against methicillin-resistant Staphylococcus aureus USA300

目的研究ε-聚赖氨酸(ε-polylysine,ε-PL)对耐甲氧西林金黄色葡萄球菌(methicillin-resistant Staphylococcus aureus,MRSA)标准菌株USA300的抑菌作用及其机制。方法依据CLSI微量肉汤稀释法测定最低抑菌浓度(minimum inhibitory concentration,MIC)和最低杀菌浓度(minimum bactericidal concentration,MBC);绘制24 h内不同浓度ε-PL作用后USA300菌株时间-抑菌曲线;SYBR Green I/PI检测ε-PL处理后USA300的生存情况;测定ε-PL处理后菌液电导率、胞外ATP含量、可溶性蛋白含量的变化;利用扫描电镜(scanning electron microscopy,SEM)观察ε-PL对USA300形态的影响。结果ε-PL对USA300的MIC、MBC分别为5.12和10.24 mg/mL。ε-PL处理后细菌死/活比例,菌液电导率,胞外ATP含量,胞外可溶性蛋白含量均增加,表明菌膜破损;经SEM进一步确证。结论ε-PL对USA300生长有良好的抑制作用,且与ε-PL浓度呈正相关。ε-PL处理后细胞膜结构破坏、通透性改变,导致细胞内容物大量渗出,其抑菌机制可能与破坏细菌菌体结构有关。

Objective To investigate the antibacterial activity and mechanism ofε-polylysine(ε-PL)against methicillin-resistant Staphylococcus aureus(MRSA)standard strain USA300.Methods Microbroth dilution was performed to determine the minimal inhibitory concentration(MIC)and the minimum bactericidal concentration(MBC)ofε-PL according to the CLSI document.The time-inhibition curve of USA300 strain was evaluated after the treatment withε-PL at different concentrations within 24 h.SYBR Green I/PI was used to detect bacterial survival afterε-PL treatment.The changes of electrical conductivity,extracellular ATP content and soluble protein content were determined afterε-PL treatment.The effect ofε-PL on the morphology of USA300 was observed by scanning electron microscopy(SEM).Results The MIC and MBC ofε-PL to USA300 were 5.12 and 10.24 mg/mL,respectively.The ratio of dead to alive,electric conductivity,extracellular ATP content and extracellular soluble protein content increased afterε-PL treatment,which indicated the rupture of USA300 and further confirmed by SEM.Conclusionε-PL has a significant inhibitory effect on the growth of USA300,which was positively correlated with the drug concentration.After treatment ofε-PL,the structure of cell membrane was destroyed and the permeability of cell membrane was changed,resulting in a large amount of exudation of cell contents.Its antibacterial mechanism may be related to the destruction of bacterial cell structure.