产生物膜菌株的分离鉴定及其产膜特性分析  

Isolation,Identification and Characterization of a Biofilm Formation Strain

作　　者：李龙杰[1,2] 周刚[2] 施庆珊[2] 欧阳友生[2] 陈仪本[2] 胡文锋[1] 

机构地区：[1]华南农业大学食品学院,广州510642 [2]广东省微生物研究所省部共建华南应用微生物国家重点实验室,广州510070

出　　处：《中国生物工程杂志》2013年第11期38-43,共6页China Biotechnology

基　　金：广东省科技计划项目(2011B010500024;2012B040301043);广东省科学院青年科学研究基金(qnjj201203)资助项目

摘　　要：目的:在含异噻唑啉酮类杀菌剂的腐败变质样品中分离筛选能够产细菌生物膜的菌株,并对其产膜特性进行初步研究。方法:采用平板划线法分离纯化菌株;采用微孔板结晶紫染色法筛选产生物膜菌株;通过生理生化特征和16S rDNA分子进化分析鉴定菌株种属;运用激光共聚焦电子显微镜观察分离得到的菌株在玻璃片表面形成生物膜情况,包括胞外多糖及生物膜内部活、死细菌分布;最后再次采用微孔板结晶紫染色法研究培养时间、pH值和不同碳源对该菌产生物膜的影响。结果:分离筛选出一株产生物膜菌株BF-17,并且该菌株被鉴定为阴沟肠杆菌(Enterobacter cloacae);BF-17分别在96 h和pH为5时产生物膜量最高;静置培养4 d后,BF-17在玻璃片表面能产生典型的生物膜形态和结构;同时该菌株利用α-乳糖和D-果糖产生物膜能力最高,而利用柠檬酸产生物膜能力最低。结论:菌株BF-17产生物膜能力稳定,可以作为生物膜深入研究的材料;同时,BF-17的产膜能力易受外界环境条件的影响。Objective： Isolate and screen a strain that has the ability to form biofilms from the samples of industrial spoilage with isothiazolinone, meanwhile research its preliminary characterization of biofilm formation. Methods：The streak plate and crystal violet staining methods were used to isolate and screen the biofilm-forming strain respectively. Morphological, physiological characteristics and 16S rDNA sequence analysis were adopted to determine the phylogenetic position of the isolated strain. The biofilms formed on the glass surface were observed under confocal scanning laser microscopy, including exopolysaccharides and live or dead cells. The influence of culture time, pH and different carbon sources on biofilm formation was also researched by crystal violet staining method in the microtiter plates. Results ： A biofilm-forming strain was successfully isolated and named as BF-17, which was identified and clarified into Enterobacter cloacae. The highest biofilm biomass was found at the time of 96 h or pH 5.0, respectively. After static cultured for 4 days, biofilms of BF-17 formed on the glass surface exhibited a representative morphological characteristics and structures of the typical biofilms. Meanwhile, the highest biofilm formation was formed in the M9 medium with the supplementation of c^-lactose or D-fructose respectively. Whereas, addition of citric acid in M9 medium made the lowest biofilm formation. Conclusion ： E. cloacae BF-17 shows a stable capacity of biofilm formation and can be used as a sample for further biofilm study. Moreover, the biofilm formation of BF-17 can be influenced by various external environment factors.

关 键 词：阴沟肠杆菌 生物膜 产膜特性 

分 类 号：Q815[生物学—生物工程]