The biofilm formation mechanism and the chemical tolerance effect of the Legionella on the solid wall accompanied by a flow

本研究旨在通过阐明军团菌在人工水文环境下的生物膜形成机制和化学耐受性作用，提出军团菌病的感染预防措施。为了达到上述目的，解决了模拟实际温水环境的流动中管道内部生物膜的形成机制，并进一步解决了生物膜对药物的防御机制，重点研究了以下两点。(1)对比静态培养系统固壁生物膜形成机理与流动介质系统固壁生物膜形成机理，明确温度条件和流动条件下生物膜形成机理的差异。(2)阐明了化学效应（生物膜的化学渗透性、Ella菌的存活率、生物膜的回收率）和耐化学细菌的存在，以及流动介质系统中固体壁上形成的生物膜。工作成果如下。制作了流体式温度梯度试验装置（流动式温度梯度培养箱：FTGI）和模拟实际温水环境的军团菌培养装置，并进行了性能试验。将培养至10cm培养皿的1份琼脂培养基放入培养皿中，对军团菌进行好氧培养48小时后，用FTGI进行培养。将壁表面温度和流体温度分别改变为37℃和42℃，每隔24 h抽取一次采样水，通过ATP测量测量军团菌数量的变化。暂停设备约48小时后，观察军团菌的生物膜粘附在玻璃壁玻片表面。提取的生物膜采用BacLive/Dead染色和荧光显微镜观察，共聚焦激光扫描显微镜观察深度方向军团菌的生死状态。澄清了军团菌在温度为37℃和生物膜温度为42℃时的生与死差异。少

This study was aimed at proposing the infection preventive measures of legionellosis by the elucidation of the biofilm formation mechanism and the chemical tolerance effect of the legion Ella in artificial hydrological environment. In order to achieve the purpose of above which solves the biofilm formation mechanism in the piping intine accompanied by the flow which imitated actual warm water environment, and solves the defense mechanism over the drugs of a biofilm further, emphasis was put on the following two points. (1) Compare the biofilm formation mechanism in the solid wall of a static culture system with the biofilm formation mechanism on the solid wall of a flow medium system, and clarify a difference of the formation mechanism by temperature conditions and a flow condition. (2) Clarify existence of an appearance of the chemical effect (the chemical permeability into a biofilm, the survival ratio of Legion Ella, rate of recovery of a biofilm), and a chemical resistance bacteriu … More m to the biofilm formed on the solid wall in the flow medium system. Fruits of work are described below. The fluid type thermal gradient test equipment (Flow type Temperature Gradient Incubator: FTGI) and the Legionella culture apparatus which imitated actual warm water environment were manufactured, and the performance test was performed. After putting in and carrying out the aerobic culture of the Legionella of one agar medium cultivated to 10cm Petri-dish to a culture vessel for 48 hours, it cultivated by FTGI. Surface-of-a-wall temperature and fluid temperature were changed to 37 degrees and 42 degrees, respectively, sampled water was extracted every 24 hours, and the change in the number of legionella was measured by ATP measurement. Equipment was suspended about 48 hours afterward and biofilm adhesion of Legionella was observed to the surface-of-a-wall slide glass. The extracted biofilm was observed with BacLive/Dead dyeing and a fluorescence microscope, and the Legionella life-and-death status of the depth direction was observed by a confocal laser scanning microscope. The difference of the Legionella life-and-death in the temperature of 37 degrees and the biofilm by 42 degrees was clarified. Less