We investigated two inter-related questions: How is the fruit consumption rate affected by fish assembly and how does an increase in fruit production alter fish composition and richness? To answer these questions, we carried out two complementary experiments in a karst tropical stream, Brazil, South America. First, we measured the fish assemblage composition before, during and after the addition of fruit under fruit producing trees. Added fruit increased differentiation of fish between up-stream and down-stream sites over time. Fish composition under fruit-producing trees was distinct from adjacent sites. Second, we set Inga laurina (Fabaceae) fruits on the stream bottom and measured the consumption time. We also analyzed if the fishes that consumed the fruits differed among 41 sites. Fish assemblage structure partially explained the observed consumption rate for the fish assemblages. Our study provides new information on how fruit input and the location of fruiting trees can induce spatial segregation within freshwater fish communities. Our results can help improving management plans for conservation and restoration of headwater streams, including the locations of riparian fruit trees that provide resources and promote hotspots of ecological interactions for freshwater communities.