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As antibiotic resistance continues to spread globally, there is growing interest in the potential to limit the spread of antibiotic resistance genes (ARGs) from wastewater sources. In particular, operational conditions during sludge digestion may serve to discourage selection of resistant bacteria, reduce horizontal transfer of ARGs, and aid in hydrolysis of DNA. This study applied metagenomic analysis to examine the removal efficiency of ARGs through thermophilic and mesophilic anaerobic digestion using bench-scale reactors. Although the relative abundance of various ARGs shifted from influent to effluent sludge, there was no measureable change in the abundance of total ARGs or their diversity in either the thermophilic or mesophilic treatment. Among the 35 major ARG subtypes detected in feed sludge, substantial reductions (removal efficiency >90 %) of 8 and 13 ARGs were achieved by thermophilic and mesophilic digestion, respectively. However, resistance genes of aadA, macB, and sul1 were enriched during the thermophilic anaerobic digestion, while resistance genes of erythromycin esterase type I, sul1, and tetM were enriched during the mesophilic anaerobic digestion. Efflux pump remained to be the major antibiotic resistance mechanism in sludge samples, but the portion of ARGs encoding resistance via target modification increased in the anaerobically digested sludge relative to the feed. Metagenomic analysis provided insight into the potential for anaerobic digestion to mitigate a broad array of ARGs. [ABSTRACT FROM AUTHOR]

Class 1 integrons and gene cassettes containing antibiotic resistance genes (ARGs) in five different sewage treatment plants (STPs) were characterized and quantified using polymerase chain reaction (PCR), sequencing, and quantitative real-time PCR (qRT-PCR) in this study. Class 1 integronase gene ( intI1) was found commonly occurring in all of activated sludge samples from the five STPs, as well as in influent and effluent of two STPs at Hong Kong. One hundred and nine lactose-fermenting Enterobacteriaceae (LFE) strains were isolated from activated sludge of Shatin STP. Among them, 36 strains (33.0%) were found to carry class 1 integrons. PCR assays showed that 11 of the 36 intI1-carrying isolates harbored a common type of gene cassette array of about 1,600 bps, as well as the static genes ( sulI and qacEΔ1) on class 1 integrons. This gene cassette array was found phylogenetically close to antibiotic resistance genes dfr17 and aadA5, encoding dihydrofolate reductase conferring resistance to trimethoprim and adenylyltransferase conferring resistance to spectinomycin/streptomycin, respectively. Antimicrobial susceptibility analysis demonstrated that all the 11 LFEs carrying gene cassette were multi-resistant, especially having common resistance to trimethoprim and streptomycin. qRT-PCR assay showed that genes copies of both class 1 integron and the gene cassette varied significantly among the activated sludge sampled from different STPs, at different time points or different treatment steps. More than 90% of class 1 integrons and the gene cassette were removed by activated sludge processes in two STPs, while the disinfection process removed 94% integron and 77% gene cassette in one STP. [ABSTRACT FROM AUTHOR]

In the present study, a newly developed metagenomic analysis approach was applied to investigate the abundance and diversity of antibiotic resistance genes (ARGs) and mobile genetic elements (MGEs) in aquaculture farm sediments, activated sludge, biofilm, anaerobic digestion sludge, and river water. BLASTX analysis against the Comprehensive Antibiotic Resistance Database was conducted for the metagenomic sequence data of each sample and then the ARG-like sequences were sorted based on structured sub-database using customized scripts. The results showed that freshwater fishpond sediment had the highest abundance (196 ppm), and anaerobic digestion sludge possessed the highest diversity (133 subtypes) of ARGs among the samples in this study. Significantly, rifampin resistance genes were universal in all the diverse samples and consistently accounted for 26.9~38.6 % of the total annotated ARG sequences. Furthermore, a significant linear correlation ( R = 0.924) was found between diversities (number of subtypes) of ARGs and diversities of plasmids in diverse samples. This work provided a wide spectrum scan of ARGs and MGEs in different environments and revealed the prevalence of rifampin resistance genes and the strong correlation between ARG diversity and plasmid diversity for the first time. [ABSTRACT FROM AUTHOR]

Bacterial community in activated sludge (AS) is diverse and highly dynamic. Little is known about the mechanism shaping bacterial community composition and dynamics of AS and no study had quantitatively compared the contribution of abiotic environmental factors and biotic associations to the temporal dynamics of AS microbial communities with significantly different diversity. In this study, two full-scale sewage treatment plants (STPs) with distinct operational parameters and influent composition were sampled biweekly over 1 year to reveal the correlating factors to whole and sub-groups of AS bacterial community diversity and dynamics. The results show that the bacterial communities of the two STPs were entirely different and correlated with the influent composition and operating configurations. Bacterial associations represented by cohesion metrics and the environmental factor temperature were the primary correlated factors to the temporal bacterial community dynamics within each STP. The STP with high diversity and evenness could treat influent with higher suspended solid and a shorter sludge retention time, and was less correlated with environmental factors, implying the importance of diversity for AS system. [ABSTRACT FROM AUTHOR]

To comprehensively understand the profile of free-living bacteria and potential bacterial pathogens in sewage treatment plants (STPs), this study applied high-throughput sequencing-based metagenomics approaches to investigate the effects of activated sludge (AS) treatment process and ultraviolet (UV) disinfection on the community of bacterial pathogens in two full-scale STPs. A total of 23 bacterial genera were identified as free-living bacteria, and 243 species/OTU97% were identified as potential bacterial pathogens, 6 of which were confidently detected in the STPs (with the total abundances ranging from 0.02 to 14.19%). Both diversity and relative abundance of the detected bacterial pathogens decreased obviously after AS treatment process (p < 0.05), and increased slightly after sedimentation (p < 0.05). UV disinfection shows no obvious effects on the total relative abundance of the free-living pathogenic bacteria in sewage. Although large amounts of the particle-bound pathogens were eliminated through the sewage treatment process, the STPs could not effectively remove the free-living bacterial pathogens, and some pathogenic bacteria (e.g., Pseudomonas aeruginosa) present in the effluent had higher relative abundance after UV disinfection. Overall, the results extend our knowledge regarding the community of potential pathogens (especially free-living pathogens) in STPs. [ABSTRACT FROM AUTHOR]

In this study, we utilized the Illumina high-throughput metagenomic approach to investigate diversity and abundance of both microbial community and copper resistance genes (CuRGs) in activated sludge (AS) which was enriched under copper selective stress up to 800 mg/L. The raw datasets (~3.5 Gb for each sample, i.e., the copper-enriched AS and the control AS) were merged and normalized for the BLAST analyses against the SILVA SSU rRNA gene database and self-constructed copper resistance protein database (CuRD). Also, the raw metagenomic sequences were assembled into contigs and analyzed based on Open Reading Frames (ORFs) to identify potentially novel copper resistance genes. Among the different resistance systems for copper detoxification under the high copper stress condition, the Cus system was the most enriched system. The results also indicated that genes encoding multi-copper oxidase played a more important role than those encoding efflux proteins. More significantly, several potentially novel copper resistance ORFs were identified by Pfam search and phylogenic analysis. This study demonstrated a new understanding of microbial-mediated copper resistance under high copper stress using high-throughput shotgun sequencing technique. [ABSTRACT FROM AUTHOR]

Arsenic contamination in a water system is an urgent environmental problem that has caused severe endemic arsenicosis in south and southeast Asian countries. It is well known that microbially mediated arsenic metabolism can enhance arsenic mobility, bioavailability, and toxicity. Here, for the first time, we applied the Illumina high-throughput metagenomic and metatranscriptomic approaches to study the distribution, diversity, abundance, and expression of microbial arsenic metabolism genes in activated sludge and five coastal sediments. An average depth of ~2.6 Gb clean data for each sample was finally obtained for BLAST analysis after quality filtration and normalization. The results revealed that: (1) highly diverse arsenic metabolism-like genes were found and the overall abundance varied from 0.18 % to 0.26 % in the six metagenomic data sets; (2) arsenic metabolism-like genes were expressed with extremely low levels or no expression at all in activated sludge; and (3) the distribution, diversity, and abundance of aioA-like, arrA-like, arsB-like, ACR3-like, and arsM-like genes varied significantly in the six surveyed environments. This study provided a novel perspective on understanding the ecology of arsenic metabolism in different water environments using high-throughput sequencing technique. [ABSTRACT FROM AUTHOR]