Persistent organic and inorganic contaminants generated by industrial effluent wastes poses a threat to the maintenance of aquatic ecosystems and public health. The Khniss and Hamdoun rivers, located in the central-east of Tunisia, receive regularly domestic and textile wastewater load. The present study aimed to survey the water quality of these rivers using physicochemical, analytical and toxicological approaches. In the physicochemical analysis, the recorded levels of COD and TSS in both samples exceed the Tunisian standards. Using the analytical approach, several metals and some textile dyes were detected. Indeed, 17 metals were detected in both samples in varying concentrations, which do not exceed the Tunisian standards. The sources of metals pollution can be of natural and anthropogenic origin. Three textile disperse dyes were detected with high levels compared to other studies: the disperse orange 37 was detected in the Khniss river with a concentration of 6.438 μg/L and the disperse red 1 and the disperse yellow 3 were detected in the Hamdoun river with concentrations of 3.873 μg/L and 1895 μg/L, respectively. Textile activities were the major sources of disperse dyes. For both samples, acute and chronic ecotoxicity was observed in all the studied organisms, however, no genotoxic activity was detected. The presence of metals and textile disperse dyes could be associated with the ecotoxicological effects observed in the river waters, in particular due to the industrial activity, a fact that could deteriorate the ecosystem and therefore threaten the human health of the population living in the study areas. Combining chemical and biological approaches, allowed the detection of water ecotoxicity in testing organisms and the identification of possible contributors to the toxicity observed in these multi-stressed water reservoirs.