Flow alteration of streams from anthropogenic impacts is ubiquitous worldwide. In this paper, we examine flow alteration in a single watershed, the Wabash River and suggest potential impacts on aquatic assemblages. The Indicators of Hydrologic Alteration (IHA) software was used to evaluate hydrological variables generated from daily discharge data of 80 USGS gauging stations, during the past several decades. We used the approach of regressions of hydrological variables against time to identify hydrologic change. An average of 6.9 hydrologic variables (of 33 variables) were significantly altered at each station, and stations with larger watersheds had an increased number of altered variables. A principal components (PC) analysis of altered hydrologic variables resulted in three of four PC axes that were correlated with watershed area. Significant results were: (1) larger streams were more likely to have hydrologic alteration than smaller streams; (2) streams with upstream dams had increased minimum flows for 1-, 3-, 7- and 30-day intervals, decreased maximum flows for 1-, 3-, 7- and 30-day intervals, increased fall rates, decreased summer monthly flows and decreased high pulse counts; (3) the presence of agriculture in upstream watersheds resulted in an increased number of zero flow days, increased low pulse counts and decreased high flows during October and April. No significant differences were detected in hydrologic alteration based on urban land-use of upstream watersheds. Hydrologic alterations in the Wabash River watershed have undoubtedly resulted in ecological degradation. We suggest that changes in assemblages during the past century are linked to these hydrologic alterations. Copyright © 2008 John Wiley & Sons, Ltd.