The Great Smoky Mountains National Park, an international biosphere reserve, harbors a very diverse flora of the acidophilic diatom genus Eunotia Ehrenberg. We examined the ecology and distribution patterns of Eunotia collected from bryophytes in streams through out the park amongst a complex geology at a variety of altitudes. Relative abundance and community composition were examined in relation to water chemistry (such as nitrates and metals), pH, and geology. Overall, greater abundances of Eunotia occurred at our higher elevations sites relative to those at lower altitudes. We found species associations with some of the watersheds and disturbance levels in the park, i.e. around the West Prong of the Little Pigeon River area, where E. incisa, E. subarcuatoides, and species from the E. exigua complex consistently grouped together. Other taxa, such as E. rhomboidea that were common in streams throughout the park, were consistently present in greater abundance in three watersheds. We describe new Eunotia species from aquatic habitats in the park and include light and scanning electron micrographs. This work contributes to our understanding of Eunotia taxonomy, while simultaneously providing the ecological information that will be useful in monitoring water systems, such as those in our national parks.