SRP412808 PRJNA910298 The riverine resistome, mobilome and virulome in eutrophic South Australian surface waters Eutrophication is a leading cause of impairment of many freshwater and coastal marine ecosystems in the world. The impact of eutrophication on water quality and microbial communities is well documented, however, little is known about its effects on the resistome, mobilome and virulome. Here, we tested the hypothesis that riverine microbial diversity and function varies during eutrophication events, with a focus on antibiotic resistance genes, mobile genetic elements and human opportunistic pathogens. We applied a comprehensive integrated analysis encompassing isolation of carbapenem resistant bacteria, resistance genes quantification (qPCR), whole genome and shotgun metagenomics to track the fate and temporal dynamic of the resistome, mobilome and virulome before and after eutrophication in riverine surface waters. The class 1 integron-integrase gene (intl1) was significantly increased post eutrophication, suggesting that genetic recombination capacity increased in eutrophic waters, along with the likely presence of multi-drug resistant bacteria. Metagenomic and whole genome sequencing analysis confirmed that the resistome, mobilome and virulome of eutrophic-origin were significantly altered. Sulfonamide (sul1), beta-lactam (blaSHV, blaTEM, blaGES, blaKPC, blaOXA-48-like), tetracycline (tetX and tetR) and polymyxin (mcr1, mcr3) resistance genes increased sharply post eutrophication. Similarly, multidrug resistant Acinetobacter baumannii, Klebsiella pneumoniae and Pseudomonas aeruginosa abundance were significantly increased too. Overall, in this study the riverine aquatic resistome, mobilome and virulome were dynamic, as evidenced by the increase of certain gene markers and enteric opportunistic pathogens in eutrophic surface waters. This provides new insights into drivers of riverine microbiome and pinpoints key monitoring targets indicative of eutrophic exposure.