This study aimed to investigate the presence and diversity of AmpC β-lactamase and integrase genes among DNA (genomic and plasmid) from bacterial populations in selected aquatic systems. Following an enrichment step, DNA was isolated and subjected to polymerase chain reaction (PCR) and digital droplet PCR. The intI1 gene and AmpC β-lactamase genes were present in genomic and plasmid DNA from all sites in the Mooi, Crocodile and Marico Rivers, with the exception of intI1 in the Marico River. Digital droplet PCR demonstrated that copy numbers varied considerably (0.0 to 29.38 copies per picogram of DNA). Some samples in which ampC was not detected, intI1 was present. Amplicons of ampC genes were subjected to restriction digest using HindIII. Samples where the restriction markers were absent were purified by cloning followed by plasmid extraction, PCR amplification, and sequencing of individual AmpC gene fragments. Phylogenetic analysis identified all positive AmpC genes as Class C β-lactamases, comprising of ampC, CMY- and ACT-families. Detecting AmpC and intl1 genes on plasmids suggests a high risk of horizontal gene transfer and potential dissemination of these and other antibiotic resistance genes surrounding immediate aquatic environments. Consequences of β-lactamase diversity in aquatic ecosystems are relatively unexplored in South African aquatic ecosystems.