Riverbed sediment is a vital component of river ecosystems and plays an important role in many geomorphological and ecological processes. However, when re-suspension occurs, pathogenic bacteria associated with sediment particles may be released into the water column, thus creating a health risk to those who use such water for drinking, household and recreational purposes. The aim of this study was to investigate the presence of bacterial pathogens Salmonella spp. and Shigella spp. in the Apies River and to ascertain whether there was any level of genetic relatedness between river water and riverbed sediment isolates of these pathogenic bacteria. A total of 124 water and sediment samples were collected from a site located on the Apies Rivers upstream of the Daspoort Wastewater Treatment Works, Pretoria, Gauteng, South Africa, between August and November 2014. In order to detect and identify the target bacteria, samples were analysed by culture-dependent and culture-independent techniques (quantitative real-time PCR). Genetic relatedness was established using Sanger sequencing of the invA gene of Salmonella spp. and ipaH of Shigella spp. Results of this study displayed the presence of the target bacteria both in the water and sediment of the river. The phylogenetic tree of Salmonella spp. revealed a ≥ 99% and 99% genetic relatedness between river water and riverbed sediment isolates for Salmonella spp. and Shigella spp., respectively. The degree of genetic relatedness between sediment and water pathogen isolates suggests that these organisms could possibly have a common origin and that there could be possible movement of microorganisms between the water column and the sediments.