黄芩素联合两性霉素B对烟曲霉菌生物被膜的体外作用

Effects of baicalein combined with amphotericin B on aspergillus fumigatus biofilm in vitro

目的探讨黄芩素联合两性霉素B(AMB)对烟曲霉菌生物被膜的破坏作用和杀菌效果。方法采用微量液基稀释法测定黄芩素及AMB对烟曲霉菌的最低抑菌浓度(MIC)及最低杀菌浓度(MFC);构建烟曲霉菌菌株生物被膜模型,将模型分为空白组、黄芩素组、AMB组、黄芩素+AMB组。成模24 h加入黄芩素组、AMB组、黄芩素+AMB组分别加入相应药物干预(黄芩素浓度128μg/m L,AMB浓度8 g/m L),空白组不干预。药物干预48 h后,结晶紫染色法行生物被膜半定量观察,胞外基质染色法荧光显微镜下观察生物被膜形态。结果生物被膜半定量结果:AMB组与空白组比较,P>0.05;黄芩素组低于AMB组和空白组,P均<0.05;黄芩素+AMB组低于黄芩素组,P<0.05。荧光显微镜下空白组和AMB组可见菌丝密集交叉缠绕,胞外基质丰富;黄芩素组菌丝仍密集交叉在一起,但菌丝细长、光滑,无明显胞外基质;黄芩素+AMB组菌丝明显减少,无明显胞外基质。结论黄芩素可以破坏烟曲霉生物被膜胞外基质。黄芩素与AMB联合应用可增加AMB对烟曲霉菌丝的渗透作用,提高其杀菌效果。

Objective To investigate the destructive effect and sterilizing effect of baicalein combined with amphotericin B （AMB） on aspergillus fumigatus biofilm. Methods The broth microdilution method was employed to test the minimum inhibitory concentration （MIC） and minimum fungicidal concentration （MFC） of baicalein and AMB, and the biofilm models of aspergillus fumigatus were established, and the models were divided into the blank group, baicalein group, AMB group and baicalein ＋ AMB group. After modeling for 24 hours, the corresponding drugs were added to the baicalein group, AMB group and baicalein ＋ AMB group. After 48-hour intervention, crystal violet assay was operated for the semi-quantification and extracellular matrix was used to observe the biofilm morphology under fluorescence microscopy. Results Crystal violet semi-quantification indicated that there was no significantly difference between AMB group and the control group （P 〉 0.05 ）, the biofilm of the baicalein group was significantly lower than that of either AMB group or the control group （ P 〈 0.05 ）, and the baicalein ＋ AMB group was lower than the baicalein group （ P 〈 O. 05）. Fluorescence microscopy showed that the hypha of control group and AMB group was crossed and encoded with rich extracellular matrix, the hypha of the baicalein group was still intensively crossed and became smoother and slender with less extracellular matrix, in the baicalein ＋ AMB group, mycelium was significantly reduced with hardly extracellular matrix. Conclusions Baicalein can destroy the extracellular matrix of aspergillus fumigatus biofilm. Baicalein combined with AMB can increase the penetration of AMB on aspergillus fumigatus and improve its sterilizing effect.