Habitat fragmentation impedes dispersal of aquatic fauna, and barrier removal is increasingly used to increase stream network connectivity and facilitate fish dispersal. Improved understanding of fish community response to barrier removal is needed, especially in fragmented agricultural streams where numerous antiquated dams are likely destined for removal. We examined post-removal responses in two distinct fish communities formerly separated by a small aging mill dam. The dam was removed midway through the 6 year study, enabling passage for downstream fishes affiliated with a connected reservoir into previously inaccessible habitat, thus creating the potential for taxonomic homogenization between upstream and downstream communities. Both communities changed substantially post-removal. Two previously excluded species (white sucker, yellow perch) established substantial populations upstream of the former dam, contributing to a doubling of total fish biomass. Meanwhile, numerical density of pre-existing upstream fishes declined. Downstream, largemouth bass density was inversely correlated with prey fish density throughout the study, while post-removal declines in bluegill density coincided with cooler water temperature and increased suspended and benthic fine sediment. Upstream and downstream fish communities became more similar post-removal, represented by a shift in Bray-Curtis index from 14 to 41 % similarity. Our findings emphasize that barrier removal in highly fragmented stream networks can facilitate the unintended and possibly undesirable spread of species into headwater streams, including dispersal of species from remaining reservoirs. We suggest that knowledge of dispersal patterns for key piscivore and competitor species in both the target system and neighboring systems may help predict community outcomes following barrier removal.