We developed fish assemblage-based multimetric indices as indicators of ecological conditions for the lower Missouri, impounded upper Mississippi, unimpounded upper Mississippi, and Ohio rivers. Using data from 475 daytime electrofishing samples, we calculated 81 candidate fish metrics. We screened these metrics for their range and their responsiveness to a multimetric stressor gradient that included water chemistry, human disturbance, and landscape-scale indicators of human disturbance and stress. Each reach-specific great-river fish index (GRFIn) included 8-10 nonredundant metrics scored on a continuous scale from 0 to 10 (10 = good). No metrics were common to all four GRFIns; one metric (the number of minnow species) was common to three GRFIns. We determined the least disturbed conditions from the y-intercepts of the quantile regressions between the GRFIn scores and the corresponding stressor gradients. Ecological condition as indicated by GRFIn score varied among reaches; 44 ± 8% (95% confidence interval around the mean) of the lower Missouri River (by length), 51 ± 20% of the unimpounded upper Mississippi River, 43 ± 8% of the impounded upper Mississippi River, and 39 ± 7% of the Ohio River was most disturbed.