Investigation Of Microbial Community Structure In Biofilm Reactors For Raw Water Pretreatment

Biological pretreatment process in the water treatment train could improve the conventional treatment processes for better dissolved organics and ammonia removal. Communities of prokaryotic microorganisms present in the biological pretreatment reactors are responsible for most of the carbon and nutrient removal from raw water and thus represent the core component of the reactors. Nitrification is the process of converting ammonia to nitrate via nitrite and is catalyzed by aerobic chemoautotrophic ammonia-oxidizing bacteria (AOB) and nitrite-oxidizing bacteria. Ammonia oxidation is thought to be the rate-limiting step for nitrification in most systems. Microbial community structure in the biological water pretreatment reactors may be unique because of the much lower substrate concentration in the influent water and the different operational parameters compared to other systems. However, the microbial communities, which directly govern substrate utilization performance of the process, are poorly understood.In this study, microbial community structures in full-scale aerated submerged biofilm reactors for micropolluted raw water pretreatment were investigated using molecular techniques, including nucleic acids extraction, clone library construction, 16S rDNA sequence homology analysis, reverse transcription-PCR, denaturing gradient gel electrophoresis (DGGE), and quantitative real-time PCR techniques. Investigations of community composition and population dynamics of AOB were emphasized due to the particular ammonia removal require in the processes. The relationship between AOB populations, specific reactor operational characteristics and the reactor performance was examined.16S rRNA gene clone libraries revealed 13 bacterial divisions in the biofilm reactor. The majority of clone sequences were related to the Alpha-, Beta- and Gamma-proteobacteria. A variety of oligotrophic bacterial sequences were identified, some sequences related to bacteria owning high potential metabolic capacities were detected in biofilm samples, such as Rhodobacter-like rRNA gene sequences. Nitrospira-like bacteria was found to be the nitrite-oxidizing bacteria in the reactor. There was a significant difference between results of the bacterial community diversity gained from culture-dependent and culture-independent methods. AOB communities in the biological water pretreatment reactors of Huinan and Hangtou Waterworks were characterized by analysis of 16S rRNA gene and the functional gene amoA, respectively. Results of different investigation routes demonstrated that it is important to apply suitable molecular markers, useful primers or probes and reasonable strategies in investigating the ecology of AOB in environments. Phylogenetic analysis revealed at least three AOB groups in the biofilm reactors, which affiliated with the Nitrosomonas oligotropha lineage, Nitrosomonas communis lineage (Nitrosomonas nitrosa-like AOB) and an unknown Nitrosomonas group. DGGE profiles of both molecular markers showed identical temporal shifts of AOB communities in Huinan and Hangtou bioreactors, indicating their identical ecological adaption directions in both reactors. DGGE analysis based on the RNA approaches exhibited more variable patterns of temporal changes of AOB communities than the DNA-derived approaches during the study, and the RNA approaches were more functional to reflect the dynamics and physiological conditions of AOB communities. The results also suggested that the activity of N. nitrosa-like AOB was more sensitive to low temperature.The population sizes of total bacteria and betaproteobacterial AOB in the biofilm reactor of Hangtou Waterworks were quantified with 16S rRNA gene real-time PCR assay. The results showed that bacterial number detected throughout a year varied substantially, by up to four orders of magnitude. Cell numbers of AOB corresponded to 0.23-1.8% of the total bacterial fraction. Water temperature was shown to have major influence on AOB population size in the reactor by the statistic analysis, and a positive correlation between AOB cell numbers and ammonia removal efficiency was suggested.Based on the quantitative results of the three specific AOB groups by real-time PCR assays, a change in competitive dominance between AOB of N. oligotropha lineage and N. communis lineage was observed. A positive correlation between cell numbers of N. communis lineage and ammonia removal efficiency was suggested. And a more significant positive correlation between cell numbers of the unknown Nitrosomoans group and ammonia removal efficiency was also revealed. Statistic analysis showed that variation of water temperature did not correlate to population size of this unknown AOB group, which is significative for optimizing the ammonia removal performance in the reactor in winter. Quantitative real-time PCR technique was proved to be a quick and effective molecular monitor method for quantifying microbial communities in the reactors, which will give directions for forecast and improvement of reactor performance in the future.