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Box turtles are long-lived, inhabit both aquatic and terrestrial habitats, and have relatively small home ranges making them a suitable candidate as a sentinel. To characterize their changes in response to environmental health, assessment of observed variation of this species is required. Thus, a comparative health assessment was employed in 825 Eastern box turtles in east central Illinois and Oak Ridge, Tennessee, to establish a baseline health assessment with regard to sex, age class, season, and location, identify temporal trends, and map parameters. Hematological and plasma biochemical variables measured included packed cell volume, total solids, white blood cell and differential counts, calcium, phosphorus, aspartate aminotransferase, bile acids, creatine kinase, and uric acid. These variables were reduced to four principle components that explained 68.8 % of the cumulative variance. The top model included the main effects of year, location, and sex, but no interactions. Spatial analysis of turtles in Tennessee demonstrated increased WBC and decreased PCV in 2011 associated with a clear-cut silviculture site. The results of this health assessment can serve as a baseline of population health in future studies and aid in the utility of this species as a sentinel.