Use of synthesized double-stranded gene fragments as qPCR standards for the quantification of antibiotic resistance genes

Xu, Like; Chen, Hannah; Canales, Melisa; Ciric, Lena

doi:10.1016/j.mimet.2019.105670

Cited by 21 publications

(22 citation statements)

References 36 publications

(21 reference statements)

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“…Raw water samples were included to provide background information on the ARGs profile. In general, the concentration of ARGs in raw water ranged from 10 3 to 10 8 copies/L, which is comparable to ARGs levels (10 3 to 10 7 copies/L) reported by the authors in a previous study investigating the same types of surface water in London (Xu et al, 2019). The levels of ARGs/integron abundance fluctuated over time, and 10 out of 15 target genes were present in all batch samples.…”

Section: Comparisons Of Args Profiles In Biofilm and Aqueous Samplessupporting

confidence: 79%

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Drinking water biofiltration: Behaviour of antibiotic resistance genes and the association with bacterial community

Campos

Canales

et al. 2020

Water Research

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Antibiotic resistance genes (ARGs) are being detected in drinking water frequently, constituting a major public health issue. As a typical drinking water treatment process, the biofilter may harbour various ARGs due to the filter biofilms established during the filtration process. The objective of this study was to investigate the behaviour of ARGs (blaCTX-M, blaOXA-1, blaTEM, ermB, tetA, tetG, tetQ, tetW, tetX, sul 1, sul 2, dfrA1 and dfrA12) and their possible association with bacteria in a bench-scale biofiltration system. The impact of filter media on horizontal gene transfer (HGT) was also explored using a model conjugative plasmid, RP1.The biofiltration system comprised four types of biofilters, including sand, granular activated carbon (GAC), GAC sandwich, and anthracite-sand biofilters. Results showed that although the absolute abundance of ARGs decreased (0.97-log reduction on average), the ARGs' abundance normalised to bacterial numbers showed an increasing trend in the filtered water.Biofilms collected from the surface layer revealed the lowest relative abundance of ARGs (p < 0.01) compared to the deeper layer biofilms, indicating that the proportion of ARG-carrying bacteria was greater in the lower position. Most chosen ARG numbers correlated to Proteobacteria, Acidobacteria and Nitrospirae phyla, which accounted for 51.9%, 5.2% and 2.0% of the biofilm communities, respectively. GAC media revealed the highest transfer frequency (2.60 × 10 -5 ), followed by anthracite (5.31 × 10 -6 ) and sand (2.47 × 10 -6 ).Backwashing can reduce the transferability of RP1 plasmid significantly in biofilms but introduces more transconjugants into the planktonic phase. Overall, the results of this study could enhance our understanding of the prevalence of ARGs in drinking water biofiltration treatment.

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Section: Comparisons Of Args Profiles In Biofilm and Aqueous Samplessupporting

confidence: 79%

“…In-house qPCR assays were established to quantify the target ARGs and two integron genes. Details of qPCR procedures were as described in a previous study by the authors (Xu et al, 2019).…”

Section: Sample Collection Dna Extraction and Qpcrmentioning

confidence: 99%

Drinking water biofiltration: Behaviour of antibiotic resistance genes and the association with bacterial community

Campos

Canales

et al. 2020

Water Research

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“…For example, White et al (2019) detected three sulfonamide antibiotics (sulfadiazine, sulfamethoxazole, and sulfanilamide) along the Thames River (England) up to the mouth after seven wastewater treatment plants, potentially exerting selection pressure on bacteria and leading to sulfonamide-resistant strains. In addition, the bla TEM , tetA , sul1 , and intI1 genes were quantified in these waters at 10 6 –10 8 copies.L −1 of water ( Xu et al, 2019 ). The sample with the highest sul1 gene abundance in this area was collected near the French coast, around Calais.…”

Section: Discussionmentioning

confidence: 99%

Occurrence of Indicator Genes of Antimicrobial Resistance Contamination in the English Channel and North Sea Sectors and Interactions With Environmental Variables

et al. 2022

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The marine environment is a potential natural reservoir of antimicrobial resistance genes (ARGs), subject to anthropogenic effluents (wastewater, industrial, and domestic), and known as a final receiving system. The aim of this study was to investigate the abundance and geographical distribution of the three blaTEM, sul1, and intI1 genes, proposed as indicators of contamination to assess the state of antimicrobial resistance in environmental settings, added to the tetA gene and the microbial population (tuf gene) in the English Channel and North Sea areas. Bacterial DNA was extracted from 36 seawater samples. The abundance of these genes was determined by quantitative PCR (qPCR) and was analyzed in association with environmental variables and geographical locations to determine potential correlations. The blaTEM and tetA genes were quantified in 0% and 2.8% of samples, respectively. The sul1 and intI1 genes were detected in 42% and 31% of samples, respectively, with an apparent co-occurrence in 19% of the samples confirmed by a correlation analysis. The absolute abundance of these genes was correlated with the microbial population, with results similar to the relative abundance. We showed that the sul1 and intI1 genes were positively correlated with dissolved oxygen and turbidity, while the microbial population was correlated with pH, temperature and salinity in addition to dissolved oxygen and turbidity. The three tetA, sul1, and intI1 genes were quantified in the same sample with high abundances, and this sample was collected in the West Netherlands coast (WN) area. For the first time, we have shown the impact of anthropogenic inputs (rivers, man-made offshore structures, and maritime activities) and environmental variables on the occurrence of three indicators of environmental contamination by antimicrobial resistance in the North Sea and English Channel seawaters.

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“…Xu et al demonstrated the versatility of chemically synthesized double-stranded (ds) DNA, which can be employed as a qPCR standard for ARGs offering comparable performance, in terms of sensitivity and reliability, to natural DNA. This qPCR method has been successfully used with various sample types, such as animal feces, soil, and surface water [41]. A multiplex real-time PCR was used for AMR characterization in Neisseria gonorrhoeae including resistance to ciprofloxacin, ceftriaxone, cefixime, azithromycin, and spectinomycin.…”

Section: Pcr-based Methodsmentioning

confidence: 99%

Rapid Methods for Antimicrobial Resistance Diagnostics

et al. 2021

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Antimicrobial resistance (AMR) is one of the most challenging threats in public health; thus, there is a growing demand for methods and technologies that enable rapid antimicrobial susceptibility testing (AST). The conventional methods and technologies addressing AMR diagnostics and AST employed in clinical microbiology are tedious, with high turnaround times (TAT), and are usually expensive. As a result, empirical antimicrobial therapies are prescribed leading to AMR spread, which in turn causes higher mortality rates and increased healthcare costs. This review describes the developments in current cutting-edge methods and technologies, organized by key enabling research domains, towards fighting the looming AMR menace by employing recent advances in AMR diagnostic tools. First, we summarize the conventional methods addressing AMR detection, surveillance, and AST. Thereafter, we examine more recent non-conventional methods and the advancements in each field, including whole genome sequencing (WGS), matrix-assisted laser desorption/ionization time-of-flight (MALDI-TOF) spectrometry, Fourier transform infrared (FTIR) spectroscopy, and microfluidics technology. Following, we provide examples of commercially available diagnostic platforms for AST. Finally, perspectives on the implementation of emerging concepts towards developing paradigm-changing technologies and methodologies for AMR diagnostics are discussed.

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Use of synthesized double-stranded gene fragments as qPCR standards for the quantification of antibiotic resistance genes

Cited by 21 publications

References 36 publications

Drinking water biofiltration: Behaviour of antibiotic resistance genes and the association with bacterial community

Drinking water biofiltration: Behaviour of antibiotic resistance genes and the association with bacterial community

Occurrence of Indicator Genes of Antimicrobial Resistance Contamination in the English Channel and North Sea Sectors and Interactions With Environmental Variables

Rapid Methods for Antimicrobial Resistance Diagnostics

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