Abstract
The overuse or misuse of antibiotics has accelerated antibiotic resistance, creating a major challenge for the public health in the world. Sewage treatment plants (STPs) are considered as important reservoirs for antibiotic resistance genes (ARGs) and activated sludge characterized with high microbial density and diversity facilitates ARG horizontal gene transfer (HGT) via mobile genetic elements (MGEs). However, little is known regarding the pool of ARGs and MGEs in sludge microbiome. In this study, the transposon aided capture (TRACA) system was employed to isolate novel plasmids from activated sludge of one STP in Hong Kong, China. We also used Illumina Hiseq 2000 high-throughput sequencing and metagenomics analysis to investigate the plasmid metagenome. Two novel plasmids were acquired from the sludge microbiome by using TRACA system and one novel plasmid was identified through metagenomics analysis. Our results revealed high levels of various ARGs as well as MGEs for HGT, including integrons, transposons and plasmids. The application of the TRACA system to isolate novel plasmids from the environmental metagenome, coupled with subsequent high-throughput sequencing and metagenomic analysis, highlighted the prevalence of ARGs and MGEs in microbial community of STPs.
Highlights
The wide use of antibiotics results in environmental releases, accelerating antibiotic resistance development in hospital wastewater, domestic sewage and livestock manure
Quantitative real time PCR indicated that the level of tetracycline resistance gene type tetG increased from 1.16105 copies/ng of total DNA to 4.16106 copies/ng of plasmid DNA after being purified by QIAGEN Plasmid Purification Kit and treated by Plasmid Safe DNase (Figure S1), indicating that the plasmids DNA were concentrated by about 37 fold after the treatments since the gene tetG is found to be only located on Mobile genetic elements (MGEs), such as plasmids
Annotation of all the reads showed that the majority was of bacterial origin while very small fractions came from fungi (548 reads,0.005%) and protozoa (499 reads,0.005%)
Summary
The wide use of antibiotics results in environmental releases, accelerating antibiotic resistance development in hospital wastewater, domestic sewage and livestock manure. We employed the transposon aided capture (TRACA) system [19] to isolate novel plasmids from microbial communities in activated sludge of a STP of Hong Kong, China.
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