The waters of rivers Swat and Kabul are the main water source for domestic and irrigation purposes in the northwestern part of Pakistan. However, this water has been contaminated due to human activities. This study aimed to analyze the water of these rivers for occurrence of antibiotic resistance genes among Gram-negative bacteria. Samples were collected from 10 different locations of these rivers. The samples were processed for the isolation of Gram-negative bacteria. Isolated bacteria were checked against 12 different antibiotics for susceptibility. The isolates were also analyzed for the presence of seven antibiotic resistance genes. A total of 50 bacterial isolates were recovered that belonged to five different bacterial genera, that is, Escherichia coli, Klebsiella oxytoca, Pseudomonas aeruginosa, Raoultella terrigena (Klebsiella terrigena), and Pseudomonas fluorescens. Antibiotic resistance pattern was cefixime 72%, cephalothin 72%, ampicillin 68%, nalidixic acid 68%, kanamycin 54%, streptomycin 42%, sulfamethoxazole-trimethoprim 28%, chloramphenicol 28%, meropenem 8%, gentamicin 8%, amikacin 2%, and tobramycin 2%. The prevalence of bla-TEM gene was 72% (n = 36), aadA gene 34% (n = 17), sul gene 32% (n = 16), bla-CTXM gene 12% (n = 6), int gene 66% (n = 33), and int1 gene 6% (n = 3). This information highlights the need for controlling and monitoring the release of domestic wastes to rivers.