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目的模拟体内环境,体外建立细菌生物膜模型,为进一步深入研究细菌生物膜生物学特点提供基础。方法将粘附载体置于气溶胶法和摇床法模拟体内细菌生物膜形成的微环境中,将铜绿假单胞菌株培养3 d后,取出标本分别进行通过FITC-ConA染色及SYTO9/PI染色,然后分别进行荧光显微镜检测及激光共聚焦检测,观察细菌生物膜的形成情况;进行电子显微镜扫描观察形成的细菌生物膜的形态特点。结果在气溶胶的微环境下,FITC-ConA染色后在荧光显微镜观察到明亮成片状的细菌生物膜;SYTO9/PI染色后在激光共聚焦检测,观察到片状,层叠如积云状,棉絮样的细菌生物膜;在电子显微镜扫描观察到大量细菌成团聚集,团状丛生突出表面,具有立体结构的细菌生物膜。在摇床法的微环境下,用3种检测方法都观察到成流线状的细菌生物膜。结论运用气溶胶法﹑摇床法可成功建立分别模拟体内呼吸系统及循环、泌尿系统的微环境下生物膜形成模型。
Objective To simulate the environment in vivo and establish a bacterial biofilm model in vitro, which provides the basis for further study of the biological characteristics of bacterial biofilm. Methods The adherent carrier was placed in the microenvironment where bacterial biofilm formation was simulated by aerosol method and shaker method. After the Pseudomonas aeruginosa strain was cultured for 3 days, the specimens were removed and stained with FITC-ConA and SYTO9 / PI , Respectively, and then were detected by fluorescence microscopy and confocal laser scanning to observe the formation of bacterial biofilm; scanning electron microscopy to observe the formation of bacterial biofilm morphological characteristics. Results In the aerosol microenvironment, FITC-ConA staining showed bright flake bacterial biofilm after being stained by fluorescence microscope. After laser scanning confocal microscopy with SYTO9 / PI staining, lamellae and lamellae were observed, Cotton-like bacterial biofilm; bacterial mass biofilm with three-dimensional structure was observed on the electron microscope scanning when a large number of bacteria were agglomerated, the surface of clusters was prominent, and the three-dimensional structure was observed. In the micro-environment of the shaker method, linear biofilms were observed in all three assays. Conclusion The biofilm formation model in the microenvironment of respiratory system, circulatory system and urinary system can be successfully established by aerosol method and shaker method respectively.