The Clinch River, Virginia is known for high aquatic biodiversity, particularly Unionidae which are declining at remarkable rates. Studies conducted on the mainstem have only addressed effects of point-source stressors (various toxic spills and effluents from the Clinch River Plant (CRP), Carbo, Virginia) that have been introduced into the Clinch River. It is hypothesized that the tributaries of the Clinch River deliver a variety of stressors to the mainstem, which may affect the diverse fauna. The aquatic health of 19 upper Clinch River tributaries, Virginia, was assessed via ecotoxicological ratings that indicated the least healthy catchments were associated with mining activity (Dumps, Russell and Coal Creeks). Tributaries were categorized by land use and mining streams were significantly different from agricultural and forested streams (F = 9.63, p<0.0001). Tributaries with ecotoxicological ratings (ETR) <80 from 100 were deemed suboptimal and thus studied further. Using identical response variables and upstream and downstream sites, resulting ETRs for nine streams indicated no model significance regarding land use, year, or site. Variability within treatments and low sampling sizes contributed to lack of significance, and results indicate that future studies need to be designed incorporating sites with analogous land use stressors. This first assessment of upper Clinch River tributaries indicates the catchments requiring remediation are Dumps, Russell and Coal Creeks, while tributaries requiring extensive evaluations are Big, Lick, Swords, Big Spring, Guest River, Cavitts and Middle Creeks. Tributaries that were deemed healthy (ETRs >80) were Big Stony, Copper, Indian, Stock, Little River and Cove Creeks.