外膜蛋白CsgG影响禽致病性大肠杆菌生物被膜形成

The outer membrane protein CsgG influences biofilm formation of avian pathogenic Escherichia coli

摘要:

【背景】禽致病性大肠杆菌(avian pathogenic Escherichia coli, APEC)能够引起禽类气囊炎、心包炎等症状，严重制约养禽业的健康发展。同时，APEC与人类肠道外致病性大肠杆菌(extraintestinal pathogenic Escherichia coli, ExPEC)具有相似的毒力因子，严重威胁公共卫生。APEC形成生物被膜可抵抗抗生素杀伤及逃避宿主免疫。因此，探究APEC生物被膜形成的关键基因和调控机制，对于防控APEC具有重要的理论和实践意义。【目的】探讨外膜蛋白CsgG在APEC生物被膜形成中的作用，揭示APEC生物被膜形成的分子机制，为禽大肠杆菌病的防治提供理论基础。【方法】利用Red同源重组技术构建的csgG基因缺失株ΔcsgG及其互补株CΔcsgG，分析了csgG对APEC生长、运动能力、膜通透性、胞外多糖(extracellular polysaccharides, EPS)产量及生物被膜形成能力的影响。【结果】csgG的缺失并不影响APEC的生长和细胞膜的通透性，但增强了APEC的运动性。csgG的缺失显著降低了APEC生物被膜形成能力，扫描电子显微镜观察发现缺失株生物被膜细菌组成由多层变为单层，细菌间黏附减少。激光扫描共聚焦显微镜观察发现csgG的缺失导致生物被膜结构松散，黏附的细菌数量减少，同时发现csgG缺失显著降低APEC的EPS产量。实时荧光定量PCR分析显示，csgG的缺失导致与生物被膜形成相关基因的转录水平显著降低。【结论】CsgG在APEC生物被膜形成过程中起着关键作用，严重影响生物被膜结构的完整性。

Abstract:

[Background] Avian pathogenic Escherichia coli (APEC) could cause inflammatory conditions such as avian airsacculitis and pericarditis, which seriously restricts the healthy development of the poultry industry. Meanwhile, APEC and human extraintestinal pathogenic Escherichia coli (ExPEC) have similar virulence factors, posing a serious threat to public health. Biofilm formation can enhance the antibiotic resistance of APEC and help it escape from the host immune system. Therefore, exploring the key genes and regulatory mechanisms of the biofilm formation of APEC are of great theoretical and practical significance for the prevention and control of APEC. [Objective] To elucidate the role of the outer membrane protein CsgG in the biofilm formation of APEC, reveal the molecular mechanism of its biofilm formation, and provide a theoretical basis for the prevention and control of avian diseases associated with APEC. [Methods] The csgG-deleted strain ΔcsgG and its complementary strain CΔcsgG were constructed by Red homologous recombination. The strains were then used to study the effects of csgG on the growth, motility, membrane permeability, extracellular polysaccharide (EPS) production, and biofilm formation of APEC. [Results] The deletion of csgG did not impact the growth or membrane permeability but enhanced the motility of APEC. The deletion of csgG significantly reduced the biofilm formation of APEC. Scanning electron microscopy revealed that the biofilm changed from multilayers to monolayers after the deletion of csgG, with reduced intercellular adhesion. Confocal laser microscope revealed that the deletion of csgG led to a loose structure of the biofilm and a decrease in the number of adherent bacteria. Furthermore, the deletion of csgG significantly reduced the EPS production of APEC. The RT-qPCR results showed that the deletion of csgG led to significant reductions in the transcript levels of genes related to biofilm formation. [Conclusion] CsgG is a pivotal factor in the biofilm formation of APEC, influencing the structural integrity of the biofilm.