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Estuaries provide vital habitat to a wide variety of fish species, so understanding how human activities impact estuarine habitats has important implications for management and conservation of fish stocks. We used nationwide datasets on anthropogenic disturbance to perform a quantitative assessment of habitat stressors in US estuaries. Habitat stressors were characterized by four categories of indicator datasets: (1) land cover/land use, (2) alteration of river flows, (3) pollution sources, and (4) eutrophication. These datasets were combined using a multiscale hierarchical spatial framework to provide a composite stressor index for 196 estuaries throughout the contiguous USA. Investigation of indicator patterns among 13 defined USA coastal subregions revealed clear differences across the USA attributable to both natural variation as well as differences in anthropogenic activities. We compared the mean composite scores for each subregion and found the lowest stressor index scores in the Downeast Maine and the Oregon Coast subregions. Subregions with the highest stressor index scores were the Southern California Bight (due to land cover changes, river flow alteration, and pollution) and Mid-Atlantic Bight (due to land cover changes, pollution, and eutrophication). Inland-based measures of pollutants, river flow, and land use all showed strong correlations with eutrophication measured within estuaries. Our approach provides an indicator-based assessment for a larger number of estuaries than has been possible in previous assessments, and in the case of river flow, for variables which previously have not been evaluated at a broad spatial scale. The results of this assessment can be applied to help prioritize watershed and estuarine restoration and protection across the contiguous USA.