Identification Of Dominant Functional Members In Efficient Wastewater Treatment Bioreactors

An important object of microbial ecology is to linking bacterial phylogenetics with the biological functionality. With the development of the molecular techniques, more and more 16S rRNA genes were sequenced and submitted to the gene sequence databases. These sequences provide enormous information about the uncultured bacteria. But the functions and the environmental roles of most of these bacteria are unknown. At the same time, many studies are finding that the microbial community composition and function often shift together in response to environmental stimuli. Utilizing this paradigm, functional important strains can be identified through functionally monitoring and dissecting the microbial communities.In this study, the biofilm reactors treating quinoline-containing wastewater were set up using the same seed sludge. The microbial communities were analyzed by the molecular techniques. An 18L tank was used as the quinoline reactor and the hydraulic retention time (HRT) was kept at 24h. All conditions were same except the nitrate was added to the denitrifying reactor. After 6 weeks acclimations, the reactor reached the stable stage. The average quinoline removal efficiency was 90.2% and the average COD removal efficiency was 81.1% in the denitrifying reactor. In the anaerobic reactor the average quinoline removal efficiency was 53.6% and the average COD removal efficiency was 60.4% in the denitrifying reactor. The DGGE profiling indicated that dramatic microbial community structure shift was occurred in the two reactors. The dominant bands in the gel were excised and sequenced, the most dominant band in the denitrifying reactor was affiliated with the genera Thauera and Azoarcus. While in the anaerobic reactor Gamma Proteobacteria and Desulfobacter postgatei were the most dominant band sequences. 16S rRNA gene libraries were constructed for the three samples. Both of the two estimators reached an asymptote for the libraries. This indicated that the libraries were large enough to yield stable phylotype richness estimates. Two statistic indices, the Shannon Wiener index (H) and the reciprocal of Simpson's index (1/D), were calculated for the three libraries and indicated that the order of the phylotype richness is SS>DR>AR. Phylogenetic analyses indicated that all clones from the DR belonged to theβsubclass of Proteobacteria, while eight classes were identified in the SS. Clones affiliated with the genera Thauera and Azoarcus were 74% in DR and only 4% in SS. The Gamma Proteobacteria and Desulfobacter postgatei become dominant during the acclimation period in the anaerobic reactor.In order to verify the population level changes of the microorganisms belong to genus Thauera and Azoarcus. Real-time quantitative PCR (RTQ-PCR) was performed to the seeding sludge sample and the biofilm sample in the quinoline-containing denitrifying reactor. The copy number of a Thauera and Azoarcus specific gene fragment were 3.70±0.16×105 and 3.69±0.97×106 per microgram of wet weight of biofilm samples in SS and DR, respectively. This indicated that the cell number of the genera Thauera and Azoarcus increased by about one order of magnitude during the acclimation. The greater abundance of Thauera and Azoarcus in association with higher efficiency after acclimation confirmed that these phylotypes might play an important role for quinoline and COD removal under denitrifying conditions.Using the similar methods, two reactors treating coking wastewater were analyzed. The COD removal efficiencies in the anaerobic and the denitrifying reactors were 18.3% and 51.4%, respectively. GC/MS analysis showed that phenol, toluene and pyridine were more efficiently removed in denitrifying reactor than in anaerobic reactor, while carbazole and aniline had higher removal efficiency in anaerobic reactor. DGGE and 16S rDNA library analysis indicated distinct microbial communities formed under the anaerobic and denitrifying conditions. Phylogenetic analysis of the 16S rRNA gene libraries indicated that the most dominant population is Thauera in denitrifying reactor and an unclassified sulfate-reducing bacterium in anaerobic reactor.In this study,the microbial community structure changes were monitored by the molecular techniques, through linking the bacterial phylogenetics with the overall functionality of the reactors, the dominant functional members were identified.