Physical factors, such as hydrologic variability, are major structuring forces of prairie stream communities. Macroinvertebrate and algae densities can both decrease sharply in response to floods and drying. Less is known about the influences of biological factors, such as fishes. The influence of fishes on macroinvertebrate communities varies with environmental factors and other biologic interactions, ranging from neutral to strong negative effects on some populations, and the strength of these interactions sometimes appear linked to hydrology. Drying intermittent streams leave permanent water refugia that may be hotspots for interactions between fishes and invertebrates. Effects of fishes on macroinvertebrate communities may vary with invertebrate life cycle stages (e.g., larvae, emerging adults, colonizing adults). I examined macroinvertebrate communities (benthic and emerging) and algal biomass across a range of permanent stream pool sites at Konza Prairie Biological Station with naturally varying densities of fishes. I also manipulated fish densities in a mesocosm experiment to address how fishes may also be effecting colonization during recovery from hydrologic disturbance. Fish biomass had a negative impact on invertebrate abundance, but not biomass or taxa richness, in natural pools. Total fish biomass was not correlated with total insect emergence in natural pools, but orangethroat darter (Etheostoma spectabile) biomass was inversely correlated with emerging Chironomidae biomass (r2 = 0.43, p = 0.047) and individual midge body size (r2 = 0.61, p = 0.014). Predatory fish biomass and a date interaction appeared in top linear models, indicating fish may also delay insect emergence from natural pools. Fish presence reduced abundance of colonizing insects (p < 0.001) and total invertebrate biomass (p = 0.001) in mesocosms. Mesocosm insect communities in pools without fishes were characterized by more Chironomidae, Culicidae, and Corduliidae (p < 0.001 for all). Chlorophyll-a increased between sampling dates in mesocosms, but did not differ between treatments. Not all life stages showed the same response to fishes, illustrating the need for understanding life histories in order to interpret the influence of fishes. Understanding how fishes in prairie streams affect ecosystem structure and function is critical for conservation and management of remaining grassland streams. Results suggest fishes can influence colonization and community structure in prairie stream pools, which serve as important refugia during hydrologic disturbance and source areas for colonists during recovery.