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Int. J. Mol. Sci. 2014, 15(6), 10083-10100; doi:10.3390/ijms150610083

A Comprehensive Insight into Tetracycline Resistant Bacteria and Antibiotic Resistance Genes in Activated Sludge Using Next-Generation Sequencing

State Key Laboratory of Pollution Control and Resource Reuse, Environmental Health Research Center, School of the Environment, Nanjing University, Nanjing 210023, China
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Received: 30 March 2014 / Revised: 9 May 2014 / Accepted: 22 May 2014 / Published: 5 June 2014
(This article belongs to the Special Issue Metagenomics: a Powerful Lens Viewing the Microbial World)
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Abstract

In order to comprehensively investigate tetracycline resistance in activated sludge of sewage treatment plants, 454 pyrosequencing and Illumina high-throughput sequencing were used to detect potential tetracycline resistant bacteria (TRB) and antibiotic resistance genes (ARGs) in sludge cultured with different concentrations of tetracycline. Pyrosequencing of 16S rRNA gene revealed that tetracycline treatment greatly affected the bacterial community structure of the sludge. Nine genera consisting of Sulfuritalea, Armatimonas, Prosthecobacter, Hyphomicrobium, Azonexus, Longilinea, Paracoccus, Novosphingobium and Rhodobacter were identified as potential TRB in the sludge. Results of qPCR, molecular cloning and metagenomic analysis consistently indicated that tetracycline treatment could increase both the abundance and diversity of the tet genes, but decreased the occurrence and diversity of non-tetracycline ARG, especially sulfonamide resistance gene sul2. Cluster analysis showed that tetracycline treatment at subinhibitory concentrations (5 mg/L) was found to pose greater effects on the bacterial community composition, which may be responsible for the variations of the ARGs abundance. This study indicated that joint use of 454 pyrosequencing and Illumina high-throughput sequencing can be effectively used to explore ARB and ARGs in the environment, and future studies should include an in-depth investigation of the relationship between microbial community, ARGs and antibiotics in sewage treatment plant (STP) sludge.
Keywords: antibiotic resistant bacteria; antibiotic resistance genes; sewage treatment plant; tetracycline; 454 pyrosequencing; metagenomic analysis antibiotic resistant bacteria; antibiotic resistance genes; sewage treatment plant; tetracycline; 454 pyrosequencing; metagenomic analysis
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).

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Huang, K.; Tang, J.; Zhang, X.-X.; Xu, K.; Ren, H. A Comprehensive Insight into Tetracycline Resistant Bacteria and Antibiotic Resistance Genes in Activated Sludge Using Next-Generation Sequencing. Int. J. Mol. Sci. 2014, 15, 10083-10100.

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