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- Inter-laboratory calibration of quantitative analyses of antibiotic resistance genesPublication . Rocha, Jaqueline; Cacace, Damiano; Kampouris, Ioannis; Guilloteau, Hélène; Jäger, Thomas; Marano, Roberto B.M.; Karaolia, Popi; Manaia, Célia M.; Merlin, Christophe; Fatta-Kassinos, Despo; Cytryn, Eddie; Berendonk, Thomas U.; Schwartz, ThomasAntibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARGs) are widely distributed in the environment where they represent potential public health threats. Quantitative PCR (qPCR) is a suitable approach to detect and quantify ARGs in environmental samples. However, the comparison of gene quantification data between different laboratories is challenging since the data are predominantly obtained under non-harmonized conditions, using different qPCR protocols. This study aimed at carrying out an inter-laboratory calibration in order to assess the variability inherent to the qPCR procedures for quantification of ARGs. With this aim, samples of treated wastewater collected in three different countries were analysed based on common DNA extract pools and identical protocols as well as distinct equipment, reagents batches, and operators. The genes analysed were the 16S rRNA, vanA, blaTEM, qnrS, sul1, blaCTXM-32 and intI1 and the artificial pNORM1 plasmid containing fragments from the seven targeted genes was used as a reference. The 16S rRNA gene was the most abundant, in all the analysed samples, followed by intI1, sul1, qnrS, and blaTEM, while blaCTXM-32 and vanA were below the limit of quantification in most or all the samples. For the genes 16S rRNA, sul1, intI1, blaTEM and qnrS the inter-laboratory variation was below 28% (3–8%, 6–18%, 8–21%, 10–24%, 15–28%, respectively). While it may be difficult to fully harmonize qPCR protocols due to equipment, reagents and operator variations, the inter-laboratory calibration is an adequate and necessary step to increase the reliability of comparative data on ARGs abundance in different environmental compartments and/or geographic regions.
- Urban wastewater treatment plants as hotspots for the release of antibiotics in the environment: a reviewPublication . Michael, I.; Rizzo, L.; McArdell, C. S.; Manaia, C. M.; Merlin, C.; Schwartz, T.; Dagot, C.; Fatta-Kassinos, D.Urban wastewater treatment plants (UWTPs) are among the main sources of antibiotics' release into various compartments of the environment worldwide. The aim of the present paper is to critically review the fate and removal of various antibiotics in wastewater treatment, focusing on different processes (i.e. biological processes, advanced treatment technologies and disinfection) in view of the current concerns related to the induction of toxic effects in aquatic and terrestrial organisms, and the occurrence of antibiotics that may promote the selection of antibiotic resistance genes and bacteria, as reported in the literature. Where available, estimations of the removal of antibiotics are provided along with the main treatment steps. The removal efficiency during wastewater treatment processes varies and is mainly dependent on a combination of antibiotics' physicochemical properties and the operating conditions of the treatment systems. As a result, the application of alternative techniques including membrane processes, activated carbon adsorption, advanced oxidation processes (AOPs), and combinations of them, which may lead to higher removals, may be necessary before the final disposal of the effluents or their reuse for irrigation or groundwater recharge.
- Antibiotic resistance genes in treated wastewater and in the receiving water bodies: a pan-European survey of urban settingsPublication . Cacace, Damiano; Fatta-Kassinos, Despo; Manaia, Célia M.; Cytryn, Eddie; Kreuzinger, Norbert; Rizzo, Luigi; Karaolia, Popi; Schwartz, Thomas; Alexander, Johannes; Merlin, Christophe; Garelick, Hemda; Schmitt, Heike; Vries, Daisy de; Schwermer, Carsten U.; Meric, Sureyya; Ozkal, Can Burak; Pons, Marie-Noelle; Kneis, David; Berendonk, Thomas U.There is increasing public concern regarding the fate of antibiotic resistance genes (ARGs) during wastewater treatment, their persistence during the treatment process and their potential impacts on the receiving water bodies. In this study, we used quantitative PCR (qPCR) to determine the abundance of nine ARGs and a class 1 integron associated integrase gene in 16 wastewater treatment plant (WWTP) effluents from ten different European countries. In order to assess the impact on the receiving water bodies, gene abundances in the latter were also analysed. Six out of the nine ARGs analysed were detected in all effluent and river water samples. Among the quantified genes, intI1 and sul1 were the most abundant. Our results demonstrate that European WWTP contribute to the enrichment of the resistome in the receiving water bodies with the particular impact being dependent on the effluent load and local hydrological conditions. The ARGs concentrations in WWTP effluents were found to be inversely correlated to the number of implemented biological treatment steps, indicating a possible option for WWTP management. Furthermore, this study has identified blaOXA-58 as a possible resistance gene for future studies investigating the impact of WWTPs on their receiving water.