To investigate the effect of the food sources and trophic position of fish in different anthropogenic interference of Three Gorges Reservoir (TGR) and Changshou Reservoir, we used stable isotopes analysis to investigate the δ¹³C and δ¹⁵N signatures of fish in these two reservoirs. Meanwhile, differences in isotopic values and trophic position of fish were also compared. The result showed that little variation in δ¹³C values of fish was observed between TGR and Changshou Reservoir. The average δ¹³C values of fish observed in Changshou Reservoir (-22.66‰±1.94‰) was similar to that in TGR (-22.59‰±1.87‰). While, the δ¹⁵N values of fish in Changshou Reservoir (13.95‰ ±3.02‰) were significantly higher than that in TGR (11.98‰ ±2.25‰). We calculated the trophic level among different fishes based on the value of δ¹⁵N. The trophic level decreased in order of carnivorous fish > omnivorous fish > planktivore fish > herbivorous fish in TGR. While, a relative difference was occurred at Lake Changshou in terms of the trophic level which was conducted as planktivore fish > carnivorous fish > omnivorous fish > herbivorous fish when compared with those in TGR. The result indicated the uncertainty of trophic level derived from value of δ¹⁵N for aqueous systems with intensive influences of human activities.