Resumo. AIM: This research aimed to study the composition of epilithic diatom flora in headwater areas of the sub-basin of the Andreas stream, RS, Brazil, and their relation with eutrophication processes. METHODS: Quarterly excursions (March, June, September, December 2012 and Mach 2013) were performed in ten sampling points selected in the sub-basin, to collect samples for the identification and counting the organisms in the group of diatoms (Class Bacillariophyceae). RESULTS: The results indicated the occurrence of 243 taxa, distributed in 53 genera. Of these, 59 were considered abundant, being distributed in 29 genera. Seven species showed elevated tolerance levels to organic pollution and eutrophication: Adlafia drouetiana (R. M. Patrick) Metzeltin & Lange-Bertalot, Amphipleura lindheimeri Grunow; Fallacia monoculata (Hustedt) D. G. Mann, Navicula cryptotenella Lange-Bertalot, Navicula symmetrica R. M. Patrick, Nitzschia palea (Kützing) W. Smith and Sellaphora auldreekie D. G. Mann & S. M. McDonald in Mann et al. CONCLUSION: Although this research has been conducted in headwater areas, the occurrence of these seven species could be explained by considering the use of these areas for agricultural and livestock purposes, compromising the stability of these aquatic ecosystems, due to the significant contribution of fertilizer and organic matter, a condition that characterizes a process of eutrophication.