Periphyton is commonly used as a biomonitoring tool for stream research. Often only one or few sites are sampled and assumed to be representative of an entire stream reach. Current literature focuses on periphyton heterogeneity at a smaller scale, on individual rocks within a riffle, and larger scales, within a watershed or ecoregion. For the purpose of this research, the understudied intermediate scales, within a single riffle and among riffles, were sampled. Periphyton was collected at four nested spatial scales: eight reaches (each nested within a different stream), ten riffles (nested within a reach), five samples (nested within a riffle), and ten rock scrapings (nested within a sample).  The eight streams included three highly impacted, three moderately impacted, and two unimpacted by acid mine drainage.  Scores from a diatom index of biotic integrity and species composition data was analyzed using UPGMA cluster analysis and DGA to determine heterogeneity of periphyton communities within and among riffles. As expected, these data show little variation within riffles and among riffles of highly impacted streams. The length of a stream represented by the biological monitoring of a single point and whether the represented length is dependent upon the health of the stream is discussed.