Structure and composition of benthic macroinvertebrate assemblages were investigated in seven sampling sites with a gradient of environmental integrity and water quality conditions. Composite samples of the four most representative substrates were collected in order to characterize the riffle-pool dynamic in each sampling site. Spatial and temporal variability of macroinvertebrate assemblages were analyzed at two scales: using substrates and grouping samples for comparing sampling sites. Distribution of macroinvertebrates was influenced primarily by substrate type, but also by environmental integrity, water quality and sampling period. Species occurrence was highly dependent on substrate type. At local spatial scale, environmental degradation measured by the Riparian Channel Environmental Inventory and water chemistry were the determinants of assemblage patterns. We evaluated to which extent the substrates were influenced by environmental integrity and water chemistry, and we found that degradation influenced significantly the macroinvertebrate fauna on the four substrate types, although they were not responding to the same variables. Our results show that qualitatively communities were not influenced by seasonal changes, but abundance was stochastically dependent on rainfall.