烧伤临床鲍曼不动杆菌生物膜形成对外排泵调节基因adeS和adeR及其功能基因簇adeAB表达的影响

Effect of biofilm formation of Acinetobacter baumannii strains isolated from burn patients on expressions of efflux pump regulatory genes adeS and adeR and corresponding functional gene cluster adeAB

目的了解烧伤临床鲍曼不动杆菌分离株在游离、生物膜成膜状态下,其外排泵上游调节基因ade S和adeR的表达变化以及相应功能基因簇ade AB的改变,探讨生物膜形成对细菌主动外排功能和耐药的影响。方法选取2013年3—9月上海交通大学医学院附属瑞金医院烧伤住院患者创面、血液和静脉导管来源的鲍曼不动杆菌耐药菌9株、敏感菌9株,试管摇菌法培养24 h后采用实时荧光定量PCR技术分别检测外排泵基因ade A、ade B以及调节基因ade S、adeR的mRNA相对表达量。结果耐药株和敏感株在游离态(游离菌)下ade A、ade B、ade S、adeR的表达量均高于各自成膜后(膜内菌)的表达量(均P<0.05)。耐药株在游离态(游离菌)和成膜后(膜内菌)ade A、ade B的表达量均高于敏感株(均P<0.05)。结论主动外排作用是游离态鲍曼不动杆菌耐药的主要机制之一。生物膜形成后,鲍曼不动杆菌膜内菌通过ade S感应膜内环境变化后下调其自身表达,继而钝化adeR,减少功能基因ade AB的表达,使得生物膜成为细菌耐药的主要原因。

Objective To explore changes of expressions of efflux pump upstream regulatory genes adeS and adeR and corresponding functional gene cluster adeAB of Acinetobacter baumannii（AB） isolates under free and biofilm formation conditions and investigate the effect of biofilm formation on the active efflux function and drug resistance.Methods Nine drug-resistant and nine drug-sensitive AB strains isolated from wound excretion,blood,and venous catheter of burn patients hospitalized in Ruijin Hospital of Shanghai Jiao Tong University School of Medicine from March to September 2013 were collected.The strains were cultured for 24 h by tube method under shaking condition and the relative mRNA expressions of efflux pump gene adeA and adeB and regulatory genes adeS and adeR were detected by quantitative real-time PCR.Results Expressions of adeA,adeB,adeS,and adeR of drug-resistant and drug-sensitive strains under free condition（free bacteria） were higher than those under biofilm formation condition（biofiim bacteria）（P〈0.05）.Expressions of adeA and adeB of drug-resistant strains under free and biofilm formation conditions were higher than those of sensitive strains（P〈0.05）.Conclusion The active efflux function is a major mechanism of drug resistance of AB under free condition.After biofilm was formed,AB can down-regulate the expression of adeS by sensing the changes of biofilm condition,inactivating the adeR,and decreasing the expression of functional gene adeAB,which makes the biofilm being the leading cause for drug resistance.