The study was designed to determine if spatial distributional patterns of lotic macroinvertebrates differed more strongly between eastern deciduous forest (EDF) and grassland (GRASS) biomes than among sites along rivers within a biome. Macroinvertebrates were collected in spring at five sites along each of three rivers within the EDF of southwestern Ontario and three rivers within the grasslands of south-central Alberta. Differences in total density, taxonomic composition, and functional feeding groups of macroinvertebrates were not consistent among sites and between biomes, as shown by the significance of the interaction term (biomes x sites). Variation in total density among rivers within biomes was attributable to differences in land use practices. Only three taxa (Leptophlebiidae, EDF; Ephemerellidae, GRASS; Pulmonata, GRASS) showed significant (negative curvilinear) relationships between density and distance from a river's source. Densities of shredders (EDF), collector-gatherers (GRASS), and predators (GRASS) also decreased in a curvilinear fashion with increasing river distance from source. The longitudinal river profiles of both taxonomic composition and functional feeding groups were weak. Despite differences in land use types within biomes, an overriding influence of biomes on the macroinvertebrate community was evident.