• 关键词：膜-生物反应器  微生物多样性  PCR-DGGE  克隆测序

• 基金项目：天津市自然科学基金重点项目(No.07JCZDJC02100)

• 张斌
• 天津大学环境科学与工程学院, 天津 300072; 2. 军事医学科学院卫生学环境医学研究所, 天津 300050

• 孙宝盛
• 天津大学环境科学与工程学院, 天津 300072

• 季民
• 天津大学环境科学与工程学院, 天津 300072

• 赵祖国
• 军事医学科学院卫生学环境医学研究所, 天津 300050

• 摘要：为了揭示膜-生物反应器中微生物群落结构多样性的演变过程,通过细胞裂解法直接提取不同时期污泥中的基因组DNA,利用基于16SrDNA的PCR-DGGE技术获得了微生物群落的DNA特征指纹图谱,并对条带进行了统计分析和切胶测序,使用序列数据进行了同源性分析并建立了系统发育树.DGGE分析表明,在反应器运行前17d内污泥中微生物群落结构变化很大,与接种污泥的相似性系数下降到了29.2%,从而说明MBR中处理工艺和进水水质的改变导致微生物群落结构多样性降低.在试验过程中,Pseudomonas和Aeromonas hydrophila等种群一直保持着较为稳定的优势地位,也有原始种群如Bacillus sp.的消亡和以Enterococcus faecalis、Comamonassp sp.、Fusobacterium sp.等为代表的次级种群的强化和演变.UPGMA聚类分析将DGGE图谱区分为3大类群并对应于各自的运行时期.测序结果表明,MBR中微生物菌群间进化距离较大,其中Proteobacteria纲和Bacillus属细菌较多.在反应器运行后期演变为优势地位的菌群(如Comamonassp.sp)加剧了膜污染物的产生和积累.

• Abstract：In order to reveal the succession of microbial community diversity in a membrane bioreactor(MBR),the genome DNA of sludge at different period was directly extracted through chemical splitting.PCR-amplified 16SrDNA and denaturing gradient gel electrophoresis(DGGE)were applied to obtain the DNAfingerprint profile of the microbial communities.The bands in the gel were analyzed by statistical methods and excised from the gel for sequencing.The sequences were then used for homology analysis and phylogenetic trees were constructed.DGGEanalysis indicated that a noticeable change took place in microbial structure after 17 days of MBR operation.This suggests that the treatment process and influent wastewater composition has a significant impact on bacterial community structures.Populations such as Pseudomonas and Aeromonas hydrophila were the predominant species throughout the study,while primary communities such as Bacillus sp.died out,and secondary microbial communities such as Enterococcus faecalis、Comamonas sp.and Fusobacterium sp.increased.Cluster analysis of DGGE by UPGMA (unweighted pair group method,arithmetic mean)divided all lanes into three clusters,which corresponded to three manipulation periods during the whole experiment.The sequences indicated that the phylogenetic tree presented the longer genetic distance among the bacteria in the MBR biomass,where Proteobacteria and Bacillus were the dominant species.The preponderant bacteria(such as Comamonas sp.)that increased during the later stage of operation had a greater impact on the production and accumulation of membrane pollutants.

• Key words：MBR  microbial diversity  PCR-DGGE  cloning and sequencing

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