The free-flowing Clinch and Powell watershed in Virginia, USA, harbors a high number of endemic mussel and fish species but they are declining or going extinct at an alarming rate. To prioritize resource management strategies with respect to these fauna, a geographical information system was developed and various statistical approaches were used to relate human land uses with available fish, macroinvertebrate, and native mussel assemblage data. Both the Ephemeroptera, Plecoptera, Trichoptera (EPT) family-level index, and the fish index of biotic integrity (IBI) were lowest in a subwatershed with the greatest coal mining activity (analysis of variance [ANOVA], p < 0.05). Limited analyses in two other subwatersheds suggested that urban and agricultural land uses within a specified riparian corridor were more related to mussel species richness and fish IBI than land uses in entire catchments. Based on land uses within a riparian corridor of 200 m x 2 km for each biological site in the watershed, fish IBI was inversely related to percent cropland and urban area and positively related to pasture area (stepwise multiple regression, R2 = 0.55, p < 0.05). Sites less than 2 km downstream of urban areas, major highways, or coal mine activities had a significantly lower mean IBI value than those more than 2 km away (ANOVA, p < .05). Land use effects included poorer instream cover and higher substrate embeddedness (t test, p < 0.05). Weaker land use relationships were observed for EPT and mussel species richness. Episodic spills of toxic materials, originating from transportation corridors, mines, and industrial facilities, also have resulted in local extirpations of native species. particularly mussels. The number of co-occurring human activities was directly related to stream elevation in the Clinch River, with more human land uses in headwater areas. Approximately 60% of known U.S. Fish and Wildlife mussel concentration sites in the watershed are located within 2 km of at least two land use sources identified as potentially stressful in our analyses. Our results indicate that a number of land uses and stressors are probably responsible for the decline in native species. However, protection of naturally vegetated riparian corridors may help mitigate some of these effects.