Correlations Of Distribution Patterns Of ARGs And MGEs With Microbial Communities In MSTPs Revealed By High-throughputSequencing

Overuse and misuse of antibiotics in health protection of human and animals have resulted in the emergence and spread of various antibiotic resistance genes (ARGs) and antibiotic resistant bacteria (ARB) in the environment, which is conferring enormous challenge and threat on environmental safety and human health. Municipal sewage treatment plants (MSTPs) are considered important environmental pools for ARB and ARGs, which have received growing environmental health concerns. This thesis aimed to to comprehensively investigate the abundance and diversity of ARGs, mobile genetic elements (MGEs) and bacterial species and their correlations in MSTPs. Experimental methods and results are summarized as follows:Occurrence and abundance of tet genes in JXZ-MSTP:PCRs showed that the 15 tet genes tested in this study occurred in JXZ-MSTP. Among the eight genes quantified, each gene had significantly higher abundance in IW than in AS and EW (P<0.05), and tetE, tetG, tetM, tetO and tetQ seemed to have higher abundance in EW than in AS (P <0.05). Phylogenic analysis of tetG gene showed that the eight OTUs generated from the 132 tetG clones of three samples were mainly grouped into two types (IW Group; AS and EW Group).Abundance and diversity of ARGs in four MSTPs and the relationship between ARGs and MGEs in the MSTPs:BLAST against the ARDB showed that the relative abundance of all ARGs in IW was higher than those of AS and EW in each MSTP. After activated sludge process, over 72% of the ARGs detected in IW were eliminated. In order to explore the correlation between ARGs and MGEs in the MSTPs, we also searched the metagenomes for plasmids, integrons and insertion sequences. Statistical analysis showed that the total abundance of ARGs had significant correlation with the total abundance of each MGE (integrase genes, plasmids and ISs) in the MSTPs (P< 0.001 each).Correlation between ARGs and bacterial communities in MSTPs:Pyrosequencing was used to characterize the microbial structure shift along the treatment processes in the MSTPs. The patterns of Chao 1 and Shannon agreed with the estimation of OTUs, revealing that AS had higher species diversity than IW. The correlation was examined through CCA based on genera and ARGs in the samples collected from four MSTPs. Among the 25 identified ARGs, especially tetE and tetM, were significantly correlated with the distribution of genus in MSTPs (P<0.05), indicating that these ARGs possibly played important roles in shaping the genus distribution in MSTPs. Pearson correlation coefficient revealed that the abundance of each ARG was significantly positively correlated with at least one genus (R>0.5;P<0.05). After activated sludge process, the relative abundance of potential ARB was generally decreased to fewer than 33%.In conclusion, results of qPCR, molecule cloning, shotgun sequencing, pyrosequencing and metagenomic analysis reveal prevalence of a variety of ARGs in MSTPs, and the abundance of ARGs and MGEs are correlated with bacterial community in sludge. Activated sludge process can greatly affect the microbial structure and cause dilution of ARGs, MGEs and ARB in sewage.