Semaprochilodus insignis is an Amazonian migratory fish species, moving in large shoals between white- and black-water rivers. It has long been classified as a detritivorous fish. However, it is possible that the trophic plasticity of S. insignis could be higher than previously assumed. The objective of this study was to investigate the relative contributions of autotrophic energy sources to the diet of S. insignis in the Negro and Solimões rivers and to determine if the species undergoes an ontogenetic change in the diet. We found variations between the δ13C and δ15N values of S. insignis between the rivers. In the Negro River, periphyton (84%) was the principal energy source for adults, while juveniles in the Solimões River foraged predominantly on terrestrial plants/C3 macrophytes (50%) and phytoplankton (42%). These variations in isotopic signatures are likely associated with migratory movements of S. insignis at different life stages and hydrological periods. Instead of the previously assumed dietary classification, we suggest that S. insignis varies its diet ontogenetically, with adults acting as illiophagous in black-water while the young are detritivorous in white-water rivers. The results show that this species creates complex links between food chains, thus emphasizing the importance of conserving flooded areas.(AU)