The U.S. Environmental Protection Agency and Environment Canada are supporting the development of indicators of ecosystem health that can be used to report on progress in restoring and maintaining the Great Lakes ecosystem, as called for in the Great Lakes Water Quality Agreement between the United States and Canada. One indicator under development for Great Lakes mesotrophic environments is based on burrowing mayflies (Hexagenia: Ephemeroptera: Ephemeridae). In this paper, we report the results of a benthic survey in spring 2002 to determine the status of nymphal populations of Hexagenia in two western Lake Superior embayments, the St. Louis River estuary, an area with significant water-use impairments, and Chequamegon Bay, an area with no known water-use impairments. Ponar grab samples collected throughout these embayments showed nymphs were generally abundant in finely particulate, cohesive substrate (clay or mixtures of clay and sand) in both embayments. However, in the St. Louis River estuary nymphs were absent in those preferred substrates at 11 stations in the eastern portion of St. Louis Bay and the adjoining northwestern portion of the Duluth-Superior Harbor, where the sediments were variously contaminated with visible amounts of taconite pellets, paint chips, oil, or combusted coal waste (clinkers). Our results suggest that human activities have rendered those portions of the St. Louis River estuary unsuitable for habitation by Hexagenia nymphs and we recommend that trend monitoring of the nymphal population there be conducted to permit reporting on progress in restoring and maintaining the health and integrity of this Great Lakes ecosystem embayment, consistent with the intent of the Great Lakes Water Quality Agreement.