连花清瘟胶囊水提物对耐甲氧西林金黄色葡萄球菌细菌生物膜的影响

Inhibitory effect of Lianhua Qingwen water extract on methicillin-resistant Staphylococcus aureus biofilm in vitro

目的观察连花清瘟胶囊水提物对耐甲氧西林金黄色葡萄球菌（MRSA）细菌生物膜的影响,探讨连花清瘟的抗菌作用机制,为临床抗菌药物的使用提供参考。方法建立体外MRSA生物膜模型,利用酶标仪检测连花清瘟胶囊水提物作用下生物膜生物总量及膜内活菌数的改变,并进一步将体外生物膜中活菌、死菌细菌外基质进行荧光染色,以激光共聚焦观测在药物作用下生物膜中活菌、死菌、细菌外基质3种组分的变化和生物膜厚度的改变。结果连花清瘟胶囊水提物173-10.81mg/ml对MRSA形成期的生物膜有抑制作用,并且可以破坏MRSA成熟生物膜43.25-0.34mg/ml,此浓度下,酶标仪检测到的连花清瘟组OD值与对照组相比减小,且未见甲氧西林组对成熟生物膜有明显的抑制结果,数据差异有统计学意义（P〈0.05）,激光共聚焦结果显示,连花清瘟胶囊水提物173mg/ml能够降低体外生物膜厚度,并可以显著减少生物膜中活菌和死菌数量。结论连花清瘟对MRSA细菌生物膜有破坏作用,提示其可用于耐药菌的临床治疗。

OBJECTIVE To investigate the effect of Lianhua Qingwen（LHQW）water extract on methicillinresistant Staphylococcus aureus（MRSA）biofilm model and explore antibacterial mechanism in order to provide the evidence for its clinical use.METHODS MRSA-biofilm in vitro model was built,and microplate reader was used to evaluate changes in biomass and the number of living bacteria under the effect of LHQW water extract.Then the living,dead bacteria and bacterial matrix were dyed with fluorescer,and the changes of the living,dead bacteria and the change of thickness of biofilm under the effect of drugs were observed with confocal laser scanning microsoopy.RESULTS In comparison with control group,MRSA biofilm was inhibited in the formation stage by LHQW water extract from 173 to 10.81mg/ml,and the inhibition result was also observed in the mature stage from 43.25 to 0.34mg/ml.In this concentration,the enzyme labeling instrument detected LHQW OD value was decreased compared with the control group.While negative result was obtained from the methicillin group at the mature stage of MRSA biofilm.Therefore,data difference was significant（P〈0.05）.Further imaging results showed that LHQW water extract could decrease the thickness of MRSA biofilm and significantly reduced the number of live and dead bacteria in the biofilm.CONCLUSIONLHQW has an anti-biofilm effect on MRSA which suggests it can be used in clinical treatment for drug resistant bacteria.