Aims: This study was designed to investigate the plasmid bearing multiple antibiotic resistant bacteria from different aquatic sources. Place and Duration of Study: This research work was carried out in Akure South Local Government Area of Ondo state, Nigeria between January and June, 2018. Methodology: The pathogenic bacteria associated with water samples collected from different sources in Akure, Nigeria were isolated and characterized. A total of 521 water samples were collected from sources such as wells, taps, streams, rivers, boreholes and rain. All the samples were subjected to presumptive, confirmed and completed tests to evaluate their microbiological quality. The microbial types in the samples were determined using standard microbiological techniques. All isolates obtained in this study were subjected to antibiotic sensitivity analysis and screened for Beta-lactamase production (ESBL). Plasmid profile analysis of the resistance isolates was carried out using standard method. Furthermore, post-curing of the plasmid mediated antibiotic resistance isolates was performed and data obtained were analyzed and presented using analysis of variance. Results: Bacterial isolates such as Acinetobacter baumanni, Citrobacter freundii, Escherichia coli, Enterobacter aerogenes, Klebsiella pneumoniae, Pseudomonas aeruginosa, Salmonella typhi, Salmonella typhimurum, Salmonella paratyphi, Shigella dysenteriae, Serratia marcescens, Proteus vulgaris and Vibrio cholerae were identified from the water samples. The isolate E. coli had the highest percentage distribution of 24.10% in well water and 26.19% in stream water while Salmonella species had the highest occurrence of 53.85% in rain water. The Beta-lactamase producing (ESBL) isolates were resistant to multiple antibiotics except Ciprofloxacin, Gentamycin and Pefloxacin that conferred antibacterial effect. Plasmid-gene profile analysis of the isolates revealed that S. typhimurium, K. pneumoniae, P. aeruginosa and P. vulgaris possess single plasmid each while only E. coli contain two plasmid bands. The post plasmid-curing antibiotic sensitivity test of the isolates revealed that the initial antibiotic resistance of the bacterial isolates were plasmid mediated. Conclusion: Findings from this study suggest the purification of water from these sources before consumption is important as most microbes found in these samples are potential pathogens that are capable of causing infectious diseases with multiple antibiotic resistant features.