A Practical Framework for Integrating Basic Separation Techniques with Molecular Quantification to Extract and Monitor Antibiotic Resistance Genes in Water Samples

Antibiotic resistance of harmful microorganisms is recognized as a serious threat to human and animal health, which is worsened by the dissemination of antibiotic resistance genes (ARGs). However, it remains challenging to identify ARGs in water samples and to distinguish intracellular (iARGs) and extracellular ARGs (eARGs). In this work, we present a feasibility study to use simple, practical methodologies to extract and distinguish ARGs in water samples using both filtration and centrifugation, followed by quantification using qPCR. Using both synthetic water samples and lake water samples, we demonstrated that the workflow reliably detected and quantified ARGs in synthetic samples. In the lake water sample, ARGs were detected at low concentrations and required a pre-concentration step to achieve detection. These methods may still need further enhancement and consideration of potential inhibitors of qPCR to be considered as a promising approach for monitoring ARGs in aqueous samples.