单宁酸联合氟康唑抑制金黄色葡萄球菌与白念珠菌混合生物膜的作用及机制研究

摘要：目的 分析单宁酸联合氟康唑降低金黄色葡萄球菌与白念珠菌混合生物膜的作用及其机制。方法 收集2019年临床分离的3株耐甲氧西林金黄色葡萄球菌(MRSA)，分别命名为SA1，SA2和SA3；利用分光光度法检测单宁酸单独或联合氟康唑对金黄色葡萄球菌与白念珠菌DAY185混合菌群生长能力的影响，进一步采用结晶紫染色法测定对混合生物膜形成能力的影响；扫描电子显微镜观察单宁酸联合氟康唑对混合生物膜结构影响；实时荧光定量PCR(qRT-PCR)检测白念珠菌DAY185生物膜形成相关基因(ALS1、ALS3和RBT1)和SA1生物膜形成相关基因(icaA、sarA和cidA)的表达量变化。结果 单宁酸可以降低金黄色葡萄球菌与白念珠菌混合菌群的生长能力和混合生物膜形成能力，当联合氟康唑时抑制作用更加明显；扫描电子显微镜显示单宁酸联合氟康唑可以明显减少金黄色葡萄球菌在白念珠菌菌丝上的黏附；qRT-PCR结果表明：单宁酸联合氟康唑主要可以降低混合生物膜中白念珠菌ALS3基因(P＜0.05)和金黄色葡萄球菌icaA和sarA基因(P＜0.05)的表达量。结论 单宁酸联合氟康唑可以降低金黄色葡萄球菌与白念珠菌混合生物膜的形成，其机制主要通过降低白念珠菌菌丝形成相关基因ALS3和金黄色葡萄球菌黏附相关基因icaA和sarA的表达水平。

Investigation of the inhibitory function and mechanism of tannic acid combined with fluconazole on Staphylococcus aureus and Candida albicans polymicrobial biofilms

Abstract Objective To investigate the inhibitory function and mechanism of tannic acid (TA) combinedwith fluconazole on Staphylococcus aureus and Candida albicans polymicrobial biofilms. Methods Three clinicalmethicillin-resistant S. aureus (MRSA) strains were collected in 2019 and named as SA1, SA2, and SA3, respectively.The influence of growth ability of S. aureus and C. albicans DAY185 mixed strains after the action of tannic acidor combined with fluconazole were measured using the spectrophotometer method, and their effects against thepolymicrobial biofilm formation ability were further analyzed using the crystal violet staining method. ScanningElectron Microscope (SEM) was used to assess the impact of tannic acid combined with fluconazole on polymicrobialbiofilms structure. Real-time quantitative PCR (qRT-PCR) was conducted to detect the expression of biofilm-relatedgenes ALS1, ALS3, and RBT1 of C. albicans DAY185 and icaA, sarA, and cidA of SA1. Results Tannic acid couldreduce the growth ability and biofilm formation ability of S. aureus and C. albicans mixed strains. Its inhibitory effectwas more significant when combined with fluconazole. SEM showed the attachment of S. aureus on hyphae of C.albicans was reduced significantly after tannic acid and fluconazole treatment. qRT-PCR revealed that tannic acidcombined with fluconazole could mainly reduce the expression levels of adhesion related genes ALS3 of C. albicansand icaA and sarA of S. aureus. Conclusion Tannic acid combined with fluconazole could reduce the polymicrobialbiofilms formation by S. aureus and C. albicans, and its mechanism was mainly by diminishing the genes expression(ALS3) involved in the formation of C. albicans hyphae and adhesion related genes icaA and sarA of S. aureus.