聚氯乙烯材料表面细菌生物膜结构观察

Study on the structure of bacterial biofilm on PVC surface

目的建立聚氯乙烯(PVC)材料表面细菌生物膜(BF)体外模型,并探讨激光共聚焦显微镜(CLSM)和环境扫描电子显微镜(SEM)对研究BF的应用价值。方法用生物膜形成阳性的表皮葡萄球菌(表葡菌)RP62A,在TBS培养基中,进行体外PVC材料表面BF形成实验动态观察,分别于培育6、12、18、24、30、48h后,用CLSM观察BF的厚度、单位面积BF群落数量、断层扫描图像、BF中活和死菌荧光比例及其三维重建图像,用环境SEM观察BF表面精细结构。结果表葡菌BF是具有高度组织化的多细胞群体结构,其中细菌密集,由活菌和死菌组成;PVC材料表面BF的形成是动态的过程,12～18h细菌黏附达到高峰,24h形成成熟BF;成熟BF中,内层、中间层及外层的活、死菌百分率比较差异有统计学意义(P<0.05)。结论表葡菌生物膜结构复杂,CLSM与环境SEM的有机结合是观察PVC材料表面细菌BF的理想方法。

Objective To establish the model of staphylococcus epidermidis （SE） biofilms （BF） on the surface of polyvinyl chloride （PVC） material, and evaluate the value of confocal laser scanning microscope （CLSM） and scanning electron microscope （SEM） in studying the configuration of SE-BF. Methods The in vivo dynamic study was performed after SE-RP62A BF formation on the surface of PVC material culture in TBS at 37 ℃ at 6,12,18,24,30,48 h respectively. The thickness of BF, colony number,percentage of vital bacterial and scanning picture of SE-RP62A biofilm on PVC of each group were measured by CLSM,and the configuration of SE biofilm was observed by SEM. Results It was shown that SE BF had highly organized structure with multi-cell colony population. A lot of vital and dead bacterium gathered densely in BF. The BF formation on the surface of PVC material was a dynamic process,the peak of bacterial adhesion needed 12 - 18 hours,and formed mature BF in 24 hours. There was significant difference of the percentage of vital bacterial among interior, middle and outer layers （P 〈 0.05 ） of material. Conclusion The SE BF had a complicated structure, combined performing CLSM and SEM is the ideal method for study BF on PVC material.