Understanding how the harvest of migrating diadromous fishes alters species interactions, energy flow, and community composition within freshwater fish assemblages is vital for obtaining a robust evaluation of fishery sustainability. In New Zealand, the ‘whitebait’ fishery harvests post-larvae of five Galaxias species as they migrate from the sea into freshwater ecosystems. We investigated if fishing-induced reductions in post-larvae biomass of three endemic kōkopu species (Galaxias fasciatus, G. argenteus, and G. postvectis) altered freshwater fish community composition and food web characteristics by limiting a potentially important marine-derived prey subsidy. We compared mass-abundance relationships, and diet (using stable isotope analyses) of fish assemblages in five fished and three unfished streams on the West Coast of New Zealand’s South Island. We found that fish assemblages within unfished streams were composed of more small-bodied fish, resulting from greater influxes of migrating kōkopu, but densities of large-bodied fish were similar between fished and unfished streams. However, initial stable isotope analyses indicated that the diet of larger-bodied fish, including adult kōkopu, was enriched by eating migrating whitebait. Therefore, whitebait are utilised as a marine-derived prey subsidy within freshwater fish assemblages, but we found no evidence that their harvest decreases the density of large-bodied fish, likely because habitat availability is more limiting than food within these streams.