The aim of the study was to use of geochemical, chemical, ecotoxicological and biological indicators for a comprehensive assessment of ecological risks related to the mobility, ecotoxicity and bioavailability of trace elements in the bottom sediment of the Rożnów reservoir. The study found three elements deserving attention in the sediments: cadmium, nickel and chromium. Cadmium proved to be the most mobile and bioavailable, although the total cadmium content and geochemical indicators did not reveal any risk to organisms. Geochemical indicators showed that the sediments are contaminated with nickel and chromium, but both elements had a low bioaccumulation factor. Fractional analysis also revealed relatively low mobility of Cr and Ni and a higher potential risk of bioavailability for nickel. Most of the tested sediment samples had low toxicity in relation to the tested organisms. For H. incongruens, 11% of the samples were non-toxic, 50% of the samples had low toxicity, and 39% of the samples were toxic. For A. fischeri, no toxicity was found in 7% of the samples, low toxicity in 76% of the samples and toxicity in 17% of the sediment samples. The As, Cd, Cu content in the F1 fraction correlated significantly positively with the content of these metals in mussel tissues. Both biotesting and chemical analysis can reveal a potential risk to aquatic organisms. For a real assessment of the ecological risks associated with trace elements, it is necessary to use bioindicators taken from the environment and exposed to trace elements in situ.