Investigation On Microbiology Diversity In Activated Sludge Of Biological Nitrogen And Phosphorus Removal Procedure

The bacterial community composition of two activated sludge samples retrieved from A~2/O and reversed A~2/O procedure were investigated by cultivation method and PCR-DGGE (Denaturing gradient gel electrophoresis) technique. And the microbial ecological communities in different reaction regions of biological phosphorus removal systems were investigated by PCR-DGGE technique.The Influence of DNA extraction methods from activated sludge for denaturing gradient gel electrophoresis on microbial diversity analysis was investigated. The results showed that the method of SDS-based DNA extraction from activated sludge (Zhou method) can be used as a rapid and dependable protocol in molecular microbial ecology study.34 bacterial strains were isolated from two activated sludge samples. Based on BIOLOG technique and 16S rRNA gene analysis, these bacterial strains were identified as five lineages of the domain bacteria:α-proteobacteria;β-proteobacteria;γ-proteobacteria; Bacilli and Actinobacteria. The subclass of the Proteobacteria, especiallyβ,γ-Proteobacteria and Actinobacteria predominated in this study. In two activated sludge samples, the composition and distribution of bacterial strains isolated from two activated sludge samples were different. The Shewanella algae; Klebsiella and Rhodobacter sphaeroides ofγ-proteobacteria were only in reversed A~2/O procedure. And the Micrococcus luteus of Actinobacteria was only in A~2/O procedure. The activated sludge samples of areobic region in A~2/O and reversed A~2/O procedure were analyzed by scanning electron microscopy. The results showed that there were lots of round- shaped and rod-shaped bacteria in two procedures. There were more filamentous bacteria in A~2/O procedure than in reversed A~2/O procedure. Moniliform bacteria was observed only in A~2/O procedure. The zoogloea of A~2/O procedure was compact, while that of reversed A~2/O procedure was loose.Using PCR-DGGE technique, the sequences of the V3 region of 16S rRNA gene retrieved from two activated sludges were separated. Examination of 16S rDNA sequences obtained from DGGE bands showed that bacterial phylogenetic composition of activated sludge in the A~2/O and reversed A~2/O procedure was diverse. 16S rDNA sequences corresponding to 20 excised bands from two activated sludge samples were analyzed and classified into four lineages of the domain bacteria: Proteobacteria; Bacteroides; Firmicutes; Candidate division TM7. The number of Nitrosospira and Rhodocyclus was more in reversed A~2/O procedure than in A~2/O procedure.Using PCR-DGGE technique, twelve activated sludge samples from lab-scale and full-scale process were investigated. The results showed that there were four lineages of the domain bacteria in lab-scale and full-scale process, including Bacteroides(54.55%); Proteobacteria (27.27%); Candidate division TM; (9.09%); Firmicutes(9.09%). The difference of microbial ecological communities between lab-scale and full-scale process was that there were more Halomonas bacteria in full-scale process than in lab-scale process, while more Rhodocyclus,γ-Proteobacteria, Bacteroidetes bacteria in lab-scale process than in full-scale process. In lab-scale process, there were less Prevotella, Sphingobacterium, Eubacterium, Nitrosospira in A~2/O procedure than in reversed A~2/O procedure. In full-scale process, both Prevotella and Flavobacterium existed in A~2/O procedure, but only existed in the anoxic region. The most differences of microbial communities in each reaction region of lab-scale and full-scale process were Flavobacteriaceae and Acinetobacter.