To investigate the diversities and abundances of nitrifiers and to apply the highthroughput
sequencing technologies to analyze the overall microbial community
structures and functions in the wastewater treatment bioreactors were the major
objectives of this study. Specifically, this study was conducted: (1) to investigate the
diversities and abundances of AOA, AOB and NOB in bioreactors, (2) to explore the
bacterial communities in bioreactors using 454 pyrosequencing, and (3) to analyze the
metagenomes of activated sludge using Illumina sequencing.
A lab-scale nitrification bioreactor was operated for 342 days under low DO (0.15~0.5
mg/L) and high nitrogen loading (0.26~0.52 kg-N/(m3d)). T-RFLP and cloning analysis
showed there were only one dominant AOA, AOB and NOB species in the bioreactor,
respectively. The amoA gene of the dominant AOA had a similarity of 89.3% with the
isolated AOA species Nitrosopumilus maritimus SCM1. The AOB species detected in the
bioreactor belonged to Nitrosomonas genus. The abundance of AOB was more than 40
times larger than that of AOA. The percentage of NOB in total bacteria increased from
not detectable to 30% when DO changed from 0.15 to 0.5 mg/L. Compared with
traditional methods, pyrosequencing analysis of the bacteria in this bioreactor provided
unprecedented information. 494 bacterial OTUs was obtained at 3% distance cutoff.
Furthermore, 454 pyrosequencing was applied to investigate the bacterial communities of
activated sludge samples from 14 WWTPs of Asia (mainland China, Hong Kong, and
Singapore) and North America (Canada and the United States). The results revealed huge
amounts of OTUs in activated sludge, i.e. 1183~3567 OTUs in one sludge sample at 3%
distance cutoff. Clear geographical differences among these samples were observed. The
AOB amoA genes in different WWTPs were found quite diverse while the 16S rRNA
genes were relatively conserved.
To explore microbial community structures and functions in the abovementioned labscale
bioreactor and a full-scale bioreactor, over six gigabases of metagenomic sequence
data and 150,000 paired-end reads of PCR amplicons were generated from the activated
sludge in the two bioreactors on Illumina HiSeq2000 platform. Three kinds of sequences
(16S rRNA amplicons, 16S rRNA gene tags and predicted genes) were used to conduct
taxonomic assignment and their applicabilities and reliabilities were compared. Specially,
based on 16S rRNA and amoA gene sequences, AOB were found more abundant than
AOA in the two bioreactors. Furthermore, the analysis of the metabolic profiles and
pathways indicated that the overall pathways in the two bioreactors were quite similar.
However, the abundances of some specific genes in the two bioreactors were different.
In addition, 454 pyrosequencing was also used to detect potentially pathogenic bacteria in
environmental samples. It was found most abundant potentially pathogenic bacteria in the
WWTPs were affiliated with Aeromonas and Clostridium. Aeromonas veronii,
Aeromonas hydrophila and Clostridium perfringens were species most similar to the
potentially pathogenic bacteria found in this study. Overall, the percentage of the
sequences closely related to known pathogenic bacteria sequences was about 0.16% of
the total sequences. Additionally, a Java application (BAND) was developed for
graphical visualization of microbial abundance data. / published_or_final_version / Civil Engineering / Doctoral / Doctor of Philosophy
| Identifer | oai:union.ndltd.org:HKU/oai:hub.hku.hk:10722/184237 |
| Date | January 2012 |
| Creators | Ye, Lin, 叶林 |
| Publisher | The University of Hong Kong (Pokfulam, Hong Kong) |
| Source Sets | Hong Kong University Theses |
| Language | English |
| Detected Language | English |
| Type | PG_Thesis |
| Source | http://hub.hku.hk/bib/B48079649 |
| Rights | The author retains all proprietary rights, (such as patent rights) and the right to use in future works., Creative Commons: Attribution 3.0 Hong Kong License |
| Relation | HKU Theses Online (HKUTO) |
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