大肠杆菌密度感应调节子C（QseC）在生物材料植入感染中的作用研究

Escherichia coli is one of the most common microorganisms associated with implant-associated infections. Bacterial biofilm has been shown to play an important role in implant-associated infections. In the biofilm state, microorganisms are relatively immune to antibodies and resistant to conventional antimicrobial agents. Regulation of the biofilm formation involves the qseC quorum-sensing system. The quorum sensing E.coli regulator C (QseC) is the histidine sensor kinase that detect autoinducer-3/epinephrine/norepinephrine. Herein we directly examined the effects of epinephrine/norepinephrine on biofilm formation of the qseC-deleted mutant of Escherichia coli on indwelling devices..We investigated biofilm formation on the surface of medical-grade polyvinyl chloride (PVC) by using the E.coli K-12 strain as well as corresponding qseC mutant. E.coli qseC mutant was constructed using lambda Red mutagenesis. LB with epinephrine and without epinephrine was used, and then the experiment of bacterial biofilm formation on PVC squares was taken. The thickness of bacterial community and bacterial community quantity in the unit area on PVC materials were measured by confocal laser scanning microscope (CLSM), and the surface structure of biofilm formation was observed by scanning electron microscope (SEM). Additionally, a rat model of nonlethal hemorrhagic shock was used. Whether the ability of E.coli to sense epinephrine/norepinephrine by QseC would have the effect on the bacterial translocation to indwelling devices in a rat model of hemorrhagic shock was studied..These results serve to give a deeper understanding of how Escherichia coli biofilm on indwelling devices works. Our results are helpful for understanding and preventing biofilm formation, as well as helpful for combating implant-associated infections.

大肠杆菌是临床生物材料植入感染的常见致病菌之一。生物材料表面细菌生物膜的形成是导致大肠杆菌感染难治性的重要因素。大肠杆菌密度感应调节子C（QseC）能识别外部信号分子，在浮游状态下细菌团块形成的密度感应调节中起重要作用。然而，在生物材料植入引发的细菌生物膜感染中，大肠杆菌QseC的作用如何？目前相关报道甚少。.本研究采用Red同源重组技术敲除大肠杆菌qseC基因，从分子、细胞和器官水平进行生物材料植入感染中大肠杆菌qseC基因功能研究；探讨大肠杆菌QseC及其信号分子对生物材料表面细菌附着能力及细菌生物膜形成的作用；通过失血性休克建立大鼠肠道大肠杆菌移位及腹膜腔内生物材料植入感染动物模型，探讨大肠杆菌QseC在肠道细菌移位引发生物材料植入感染中的作用；揭示防治大肠杆菌引发的生物材料植入感染可能作用的分子药物靶标，为通过基因沉默技术防治生物材料植入感染等后续研究提供实验依据。

大肠杆菌（Escherichia coli）是人体肠道中的条件致病菌，心脏大血管手术体外循环期间、神经外科或骨科等手术控制性降压期间、以及临床上常见的失血性休克，均 可导致肠通透性升高及肠道细菌移位，造成菌血症，血液中的细菌为细菌在生物材料表面黏附提供菌源。细菌在生物材料表面一旦形成细菌生物膜，就能有效地抵御机体的防御反应和抗生素的治疗，是导致生物材料植入感染难以控制的根源。本研究采用Red同源重组技术构建大肠杆菌K-12菌株qseC阴性突变菌株，在前期建立的聚氯乙烯材料表面细菌生物膜模型基础上，采用激光扫描等技术，探讨大肠杆菌QseC对生物材料表面细菌生物膜形成能力的影响；通过失血性休克建立肠道细菌移位动物模型，探讨大肠杆菌QseC在肠道细菌移位引发生物材料植入感染中的作用；揭示防治大肠杆菌引发的生物材料植入感染的有效分子药物靶标，为防治生物材料植入感染提供实验依据。. 研究结果提示：①大肠杆菌QseC缺失后运动能力显著下降。儿茶酚胺对大肠杆菌运动能力有直接促进作用，而且该作用是通过QseC介导的。②生物材料表面大肠杆菌生物膜的形成是一个动态过程，生物材料表面细菌群落数量及细菌生物膜厚度的增加与时间的变化相关，这可能与生物材料植入感染的临床感染特征有关。③大肠杆菌QseC在生物材料表面细菌生物膜的形成中具有重要作用。儿茶酚胺对大肠杆菌在生物材料表面细菌生物被膜的形成有直接促进作用，而且该作用是通过QseC介导的。④大肠杆菌QseC缺失后，运动能力降低，同时QseC信号链中断，细菌对肠粘膜的侵袭力和穿透力减弱，导致细菌移位发生率下降。QseC-肾上腺素信号通路在肠道大肠杆菌移位并在生物材料表面定植中起重要作用。