Freshwater fish assemblages are sensitive to changes in land use, but it is unclear how rapidly assemblages respond to such change or how closely they track physical changes in the stream environment. We monitored fish assemblages at the outflow of three watersheds on the outer coastal plain of Mississippi over a period of 8 y. The watersheds differed in patterns of land use, being predominantly forested, rapidly urbanizing, and long-urbanized, respectively. Watersheds were distinct in fish species composition, differing primarily in abundance of Etheostoma lynceum, E. stigmaeum, Luxilus chrysocephalus and Lythrurus roseipinnis. Species richness was consistently greatest at the predominantly forested site and least at the urbanized site. Fish assemblages were distinguishable in terms of substrate preference, water-speed, silt tolerance, and trophic habit. The stream draining the urbanizing watershed showed an increase in richness of species of high silt tolerance and a decrease in richness of those preferring a gravel substrate over the sampling period. Contrary to expectation, annual variation in composition was modest relative to differences among sites. Annual variation in richness and numbers was unrelated to measures of streamflow or seasonal precipitation. Changes in stream character related to land use change appear to have impacted the fish assemblages of these streams, but the transition to an urban fish assemblage in the urbanizing stream occurred before the study period, well before the obvious signs of physical degradation appeared at the site.