Previous freshwater mussel surveys (1915–1997) at sites in the upper North Fork Holston River watershed upstream of Saltville, VA, documented 21 species. To assess current status of the fauna, approximately 363 survey hours were spent qualitatively sampling 44 sites in a 77-km reach of the upper river between Saltville and Ceres during 2000–2004. Thirteen species of live freshwater mussels were collected. Species richness appears to have declined only slightly over the last 100 years in this reach of river. However, declines in abundance are now evident in a 6.4-km reach immediately upstream of Saltville. A die-off of federally endangered Fusconaia cor (shiny pigtoe) and candidate species Lexingtonia dolabelloides (slabside pearlymussel) was documented in the upper river during 1999–2002, but the cause was not identified.