As a One Health issue impacting life across humans, animals, and the environment, antibiotic resistance is found in one of the most basic resources for life – water. Antibiotic-resistant pathogens and their genes have been found in streams, rivers, lakes, and oceans. Here, we used a comprehensive integrated analysis encompassing isolation of carbapenem-resistant bacteria, whole genome, and shotgun metagenomics to track the fate of the resistome, mobilome, and virulome in surface waters from several waterbodies in Australia and the Pacific Islands. To provide globally comparative data sets and to link this study directly to local epidemiological data, qPCR assays were designed to mirror molecular assays currently used for international-wide human surveillance for nosocomial and community-acquired antimicrobial-resistant infections. The class 1 integron-integrase gene (intl1), a proxy for multi-drug resistance, sulfonamide (sul1) and extended-spectrum β-lactam (blactxm32, blaSHV, blaTEM, blaGES, blaKPC) genes increased sharply in peri-urban and urban waterbodies. Whereas tetracyclines (tetW, tetX, tetA), vancomycin (vanA), and quaternary ammonium compounds (qacE) were significantly abundant in agricultural catchments. Interestingly, polymyxin resistance genes (mcr1-5) were abundant in undeveloped, agricultural, peri-urban, and urban areas. Agricultural, peri-urban, and urban waterbodies were carriers of multidrug-resistant and virulent ESKAPE pathogens, including E. coli ST131 and ST69 carrying the plasmid pUTI89. This comprehensive study of waterbodies in Australia and the Pacific Islands provides new insight into drivers of antimicrobial resistance and pinpoints key monitoring targets indicative of where human sources and exposures are likely to be most acute and in need of multi-sector surveillance.