© 2017 by The Society for Freshwater Science. Anthropogenic stressors, such as urban development and intensification of agricultural and industrial activities, can alter physical, chemical, and biological conditions in aquatic systems. Mounting concern over cumulative effects of multiple stressors has stimulated the need to track ecological responses and quantify degradation. Periphytic diatoms have potential to serve as early indicators of ecological degradation in aquatic systems. We examined relations between periphytic diatom community composition, water chemistry, and the magnitude of anthropogenic stressors in lakes of the Muskoka River watershed. These relations were used to develop a bioassessment framework for tracking changes in biological integrity in a region undergoing growth of tourism and related urban and recreational activities. We examined nearshore periphyton and water-chemistry samples from 86 lakes to assess relations between diatom community composition, water chemistry, and measures of 9 anthropogenic stressors (e.g., % urban area, % agricultural area, roadway proximity) based on uni- and multivariate numerical methods. Composition of periphytic diatom communities was associated with the concentration of ions from anthropogenic sources (i.e., road salt, dust suppressants). We demonstrated that periphytic diatoms in the nearshore regions of the lakes may be sensitive early warning indicators of ecological change because the most dramatic shift in diatom community composition was observed with modest increases in anthropogenic stressors. This relationship was weaker at sites where natural variation in pH among lakes was high. pH often acts in conjunction with other chemical factors to influence diatom community composition. Thus, we recommend stratifying monitoring efforts to account for confounding natural variation in pH. We anticipate that periphytic diatoms can be used as a powerful indicator of environmental stress in lakes beyond the MRW.