亚抑菌浓度头孢噻肟增强沙门菌生物被膜形成机制研究

Bacterial biofilm formation is an important adaptive strategy in response to adverse stress, and can increase the development of bacterial multidrug resistance and evasion of host immune surveillance. In turn, resistant bacteria may be more easily exposed to sub-inhibitory concentration of antimicrobials, which will induce the enhancement of biofilm formation. Salmonella is one of the important zoonotic and foodborne bacterial pathogens, and its resistance to the third-generation cephalosporins like cefotaxime has attracted world-wide attention. Sub-inhibitory concentration of cefotaxime has been confirmed to enhance Salmonella biofilm formation, while the mechanism of action remains unclear. In this project, we will firstly intend to discover the potential factors related to cefotaxime-induced enhancement of Salmonella biofilm formation using RNA-seq transcriptomics and bioinformatics. Then, using such techniques as enzyme inhibition assay and laser confocal microscopy, we will identify the role of extracellular matrix components in cefotaxime-induced enhancement of Salmonella biofilm formation, followed by analyze and verify the regulatory pathway. Next we will try to discover the membrane proteins responsible for the binding and transportation of cefotaxime, and clarify the mechanism for signal recognition and transmission. Based on these experiments, the mechanism of enhancement on Salmonella biofilm formation induced by sub-inhibitory concentration of cefotaxime will be interpreted. This project will contribute to the investigation on interactive regulation of drug-pathogen, and will also guide the rational use of antimicrobials and the prevention of biofilm formation, which possesses important theoretical and practical significances.

生物被膜形成是细菌一种重要抗逆性适应策略，可提高细菌多重耐药性和逃避宿主免疫监视；耐药性存在使细菌更易暴露于亚抑菌浓度抗菌药胁迫，进而诱导生物被膜形成增强。沙门菌是一种重要人兽共患和食源性病原菌，其对头孢噻肟等第三代头孢菌素耐药性广受关注，业已证实亚抑菌浓度头孢噻肟可增强沙门菌生物被膜形成，但其作用机制尚不清楚。本项目拟结合RNA-seq转录组和生物信息学分析，挖掘参与头孢噻肟增强沙门菌生物被膜形成的潜在因子；利用酶抑制试验、激光共聚焦显微技术等，鉴定胞外基质组分在介导生物被膜诱导性增强中的作用，进而分析和验证其调控通路；利用荧光标记青霉素竞争结合试验等，鉴定头孢噻肟结合和转运相关细菌膜蛋白，探究信号识别和传导机制；综合试验结果，诠释亚抑菌浓度头孢噻肟增强沙门菌生物被膜形成的机制。本项目有助于研究药物-病原互作调控，也有助于指导抗菌药合理使用和生物被膜防控，具有重要的理论和应用研究。

生物被膜形成是细菌一种重要抗逆性适应策略，可提高多重耐药性和逃避宿主免疫监视；耐药性存在常使细菌更易面临亚抑菌浓度抗菌药胁迫，进而诱导生物被膜形成增强。沙门菌是一种重要人兽共患和食源性病原菌，其对头孢噻肟等第三代头孢菌素耐药性广受关注，业已证实亚抑菌浓度头孢噻肟可增强沙门菌生物被膜形成，但作用机制尚不清楚。本项目结合生物信息学、RNA-seq转录组和转座子突变库构建，挖掘出参与头孢噻肟增强沙门菌生物被膜形成的潜在因子；利用酶抑制试验、激光共聚焦显微技术等，证实了胞外基质组分尤其是纤维素在介导沙门菌生物被膜形成及头孢噻肟诱导生物被膜增强中的作用，进而分析了参与纤维素调控的关键基因；利用基因缺失株和回补株的构建，鉴定了青霉素结合蛋白PBP1b为头孢噻肟结合和转运的膜蛋白，证实了细菌分裂异常和长丝状形态形成参与头孢噻肟对沙门菌生物被膜的增强。综合试验结果，提出亚抑菌浓度头孢噻肟增强沙门菌生物被膜形成的分子机制。本项目有助于研究药物-病原互作调控，也有助于指导抗菌药合理使用和防控生物被膜形成，具有重要的理论和应用研究价值。