The Three Gorges Reservoir (TGR) of China, the largest hydropower project over the world, has attracted much attention to the water impoundment and water-level manipulation. In this study, we evaluated potential effects of water impoundment and seasonal water-level manipulation on the bioaccumulation, trophic transfer and health risk of HMs (Cu, Fe, Zn, Hg, Cd and Pb) in food web components (seston, aquatic invertebrate and fish) in TGR. Our results show that, after the impoundment for eight years (2003–2010), all of the six metal concentrations in aquatic biota fell within the criteria of safety quality guidelines. The concentrations of Cu, Fe, Zn and Hg in fish and aquatic invertebrates were higher than those before impoundment, whereas Cd and Pb were lower than those before impoundment. Nonetheless, Hg, Cd and Pb in aquatic consumers underwent an increasing trend during the entire impoundment, implying potential reservoir effect in the future. Only the concentrations of Hg, Cd and Pb in aquatic consumers exhibited a declining trend towards the dam, showing consistent with the background level at the three reaches. Seasonal variations in HM concentrations of fish and aquatic invertebrates were ascribed to the water-level manipulation associated with reservoir management. Our findings show that Hg or Cd biomagnified through aquatic food web during different hydrological periods, whereas Pb, Cu, Fe and Zn exhibited weak biomagnification power. Overall, Hg, Cd and Pb showed a higher risk than that of Cu, Fe and Zn.