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目的研究群体感应(QS)系统在铜绿假单胞菌生物膜(BF)形成中的作用。方法体外建立3 d QS系统完整的铜绿假单胞菌野生型PA01菌株与QS系统缺陷(lasRrhlR基因缺陷△lasR△rhlR)型菌株生物模型,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lasRrhlR缺陷株,PA01菌株BF相关空间结构参数。结果培养第3天PA01菌株可形成较厚、有孔状通道的成熟BF结构,而△lasR△rhlR菌株仅形成明显稀薄的早期BF结构,△lasR△rhlR菌株的3 d BF厚度为(7.36±0.2)μm,PA01菌株为(21.64±0.57)μm(P<0.05),区域孔率(areal porosity,AP)分别为:(0.902±0.006)、(0.928±0.002);平均扩散距离(average diffusion distance,ADD)和结构熵(textural entropy,TE)在△lasR△rhlR菌株分别为:(1.503±0.029)和(5.706±0.190);在PA01菌株分别为:(1.467±0.015)和(5.213±0.111),△lasR△rhlR菌株AP较PA01菌株低(P<0.05);而ADD、TE较PA01菌株高(P<0.05)。结论△lasR△rhlR基因缺陷明显影响铜绿假单胞菌的BF形成能力,QS系统lasRrhlR基因在铜绿假单胞菌BF形成中发挥重要作用。
Objective To investigate the role of population-responsive (QS) system in the formation of Pseudomonas aeruginosa biofilm (BF). Methods The whole model of Pseudomonas aeruginosa wild-type PA01 strain and QS system-deficient strain (lasRrhlR gene defective △ lasR △ rhlR) strain was established in vitro for 3 days by SYS09 / PI fluorescent probe and combined with laser scanning confocal microscopy The images of different layers of BF were taken and the spatial structure parameters related to BF of lasRrhlR strain and PA01 strain of QS system were obtained by image structure analysis (ISA) software. Results On the third day after cultivation, the PA01 strain formed a thick, porous BF-like structure. However, the △ lasR △ rhlR strain formed only a distinctly thin early BF structure. The Δ BFR of rhsR △ rhlR strain was (7.36 ± (P <0.05). The average PAI of PA01 was (0.902 ± 0.006) and (0.928 ± 0.002), respectively. The average diffusion distance , ADD and TE were respectively (1.503 ± 0.029) and (5.706 ± 0.190) in the lasRΔ rhlR strains, and (1.467 ± 0.015) and (5.213 ± 0.111) in the PA01 strains respectively , AP lasR rhlR strain AP lower than the PA01 strain (P <0.05), while ADD, TE higher than the PA01 strain (P <0.05). Conclusion The △ lasR △ rhlR gene deficiency significantly affects the BF formation ability of Pseudomonas aeruginosa, and the lasRrhlR gene of QS system plays an important role in the formation of Pseudomonas aeruginosa BF.