目的 观察云南重楼正丁醇提取物(BEPP)对体外白念珠菌生物膜形成的影响。方法 微量稀释法测定BEPP最低抑菌浓度(MIC),XTT法测定生物膜抑制80%浓度(SMIC₈₀),平板法绘制时间-杀菌曲线,扫描电镜(SEM)观察生物膜形态结构,激光共聚焦显微镜(CLSM)观测生物膜厚度,实时荧光定量PCR(qRT-PCR)法检测生物膜形成相关基因HWP1、MP65、SUN41表达量的变化。结果 BEPP对白念珠菌的MIC为32～128 μg/mL,对白念珠菌生物膜的SMIC₈₀为128～512 μg/mL,时间-杀菌曲线显示BEPP具有良好的杀菌作用,SEM观察到BEPP有效抑制白念珠菌生物膜形成,CLSM显示BEPP可以降低生物膜厚度,qRT-PCR检测显示在BEPP作用下,HWP1、MP65、SUN41基因表达量均显著下调。结论 BEPP可以有效抑制体外白念珠菌生物膜的形成。

Objective To investigate the effects of butanol extract from Paris polyphylla var. yunnanensis (BEPP) on the biofilm formation of Candida albicans in vitro. Methods Microdilution method was used to determine the minimum inhibitory concentration (MIC). XTT assay was used to determine the SMIC₈₀. Flat band method was used to protract the time-kill curve. Scanning electron microscopy (SEM) was used to observe the morphological changes of the biofilm. Confocal laser scanning microscopy (CLSM) was used to detect the thickness of the biofilm. The quantification real-time PCR (qRT-PCR) was used to detect the expression changes of genes (EAP1, MP65, and SUN41) of the biofilm cells. Results The MIC of BEPP against C. albicans strains was determined as 32—128 μg/mL. The SMIC₈₀ of BEPP against the biofilm of C. albicans strains was determined as 128—512 μg/mL. Time-sterilization curve results indicated that BEPP had a promise bactericidal effect. SEM results showed that the formation of C. albicans biofilm was inhibited by BEPP, and the morphology of biofilm was also affected by BEPP. The thickness of C. albicans biofilm was reduced by BEPP according to CLSM results. Furthermore, qRT-PCR results indicated that expressions of EAP1, MP65, and SUN41 were significantly down-regulated by BEPP. Conclusion Our results demonstrate that BEPP inhibits the biofilm formation of C. albicans in vitro.

安徽省自然科学基金项目(1408085MH165)