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目的研究群体感应(quorum sensing,QS)系统在铜绿假单胞菌生物膜(biofilm,BF)形成中的作用。方法体外建立QS系统完整的铜绿假单胞菌野生型PAO1菌株与QS系统缺陷(lasI rhlI基因缺陷△lasI△rhlI)型菌株生物膜模型,通过SYT09/PI荧光探针标记,结合激光共聚焦显微镜摄取BF不同层面的图片,经图像结构分析(ISA)软件分析获得QS系统lasI rhlI缺陷株、PAO1菌株BF相关空间结构参数。结果培养第3天PAO1菌株可形成较厚、有孔状通道的成熟BF结构,而△lasI△rhlI菌株仅形成明显稀薄的早期BF结构,△lasI△rhlI菌株的3 dBF厚度为(6.62±0.31)μm,PAO1菌株为(21.64±0.57)μm(t=0.205,P<0.05),区域孔率(areal porosity,AP)分别为0.902±0.006、0.970±0.003;平均扩散距离(average diffusion distance,ADD)和结构熵(textural entropy,TE)在△lasI△rhlI菌株分别为1.571±0.019和6.586±0.110;在PAO1菌株分别为1.467±0.015和5.213±0.111,△la-sI△rhlI菌株AP较PAO1菌株低(t=0.335,P<0.05);而ADD、TE较PAO1菌株高(t=0.289,t=0.300,P<0.05)。结论lasI rhlI基因缺陷明显影响铜绿假单胞菌的BF形成能力,QS系统在铜绿假单胞菌BF形成中发挥重要作用。
Objective To investigate the role of quorum sensing (QS) in the formation of biofilm (BF). Methods A biofilm model of Pseudomonas aeruginosa wild-type PAO1 strain and QS system-deficient strain (lasI rhlI) was established in vitro using SYT09 / PI fluorescent probe and laser scanning confocal microscopy The pictures of different layers of BF were taken and the spatial structure parameters of lasI rhlI-deficient strain and PAO1-strain BF of QS system were obtained by image structure analysis (ISA) software analysis. Results On the third day, the PAO1 strain could form a mature BF structure with thick and porous channels, while the △ lasI △ rhlI strain only formed a distinctly thin early BF structure. The 3 dBF thickness of the △ lasI △ rhlI strain was (6.62 ± 0.31 (21.64 ± 0.57) μm for PAO1 strain (t = 0.205, P <0.05) and 0.902 ± 0.006 for PAO1 strain, and 0.970 ± 0.003 for APO strain. The average diffusion distance (ADD) ) And the entropy of textural entropy (TE) were 1.571 ± 0.019 and 6.586 ± 0.110 respectively in △ lasI △ rhlI strains, 1.467 ± 0.015 and 5.213 ± 0.111 in PAO1 strains, respectively, compared with those in PAO1 strain (T = 0.335, P <0.05), while ADD and TE were higher than PAO1 strains (t = 0.289, t = 0.300, P <0.05). Conclusion The deficiency of lasI rhlI gene significantly affects the BF formation ability of Pseudomonas aeruginosa, and the QS system plays an important role in the formation of Pseudomonas aeruginosa BF.