Intra- and extracellular DNA as carriers of antibiotic resistance genes and class 1 integrons in river waters

Increasing attention is being given to the role of extracellular DNA (eDNA) in the spread of antibiotic resistance genes (ARGs). However, few studies have investigated the content of ARGs in both intracellular DNA (iDNA) and eDNA in freshwater ecosystems. In this study, we analyzed the abundance of class 1 integrons and selected ARGs (ermB, blaCTXM, tetA, and sulII) using qPCR across five rivers (Toce, San Giovanni, San Bernardino, Bardello, and Maggia) located in the subalpine area of Northwestern Italy, characterized by different levels of anthropogenic pollution, as determined by chemical analysis. The River Bardello exhibited a higher concentration of chemical parameters associated with anthropogenic pollution (e.g., ammonium, phosphorus compounds). The abundance of class 1 integrons and ARGs was significantly higher in this river, with iDNA generally being the main carrier of ARGs. However, both DNA fractions contributed to the spread of antibiotic resistance. Overall, our results demonstrated that eDNA is essential for a comprehensive assessment of antibiotic resistance in river ecosystems and that anthropogenic pollution significantly affected ARG abundance.