Tychoplanktonic and epilithic diatom samples were taken monthly during one year at 18 sites in small lowland rivers in the northern central region of Portugal. Canonical correspondence analysis was used to explore the relationships between measured environmental variables and patterns in the diatom assemblages. Conductivity, HCO 3 −, and chemical oxygen demand (COD) were the most significant environmental variables influencing the structure of the tychoplankton and of the epilithon. Other less important variables, such as K +, Fe, N(NO 3 −), and Na + also explained significant (P < 0.05) amounts of variance. The ordination diagrams showed very similar distribution of the tychoplanktonic and epilithic diatom samples. At sites with similar geomorphology as in this study, it is possible to use only one of these samples for water quality evaluation. The optima and tolerances of some diatom taxa were calculated for the most influential variables. Although the diatoms showed high tolerances to some environmental parameters, it was possible to establish groups of taxa with defined and distinctive ecological preferences. Two species groups stood out depending on the alkalinity and mineralization of the water. With a moderate conductivity (> 600 μS·cm −1), alkaline pH, HCO 3 − concentrations equal or higher than 150 mg·l −1, the first group includes Achnanthes hungarica Grunow, Amphora pediculus (Kützing) Grunow, Bacillaria paxillifera (O. F. Müller) Hendey, Cymatopleura solea (Brébisson) W. Smith, Navicula accomoda Hustedt, Navicula lanceolata (Agardh) Ehrenberg, Navicula trivialis Lange-Bertalot, Nitzschia hungarica Grunow. In waters of lower conductivity (varying between 72 and 262 μS·cm −1) slightly acid pH (6 to 6.5) and HCO 3 −1 lower than 46 mg·l −1 the following species were common: Achnanthes oblongella Östrup, Achnanthes subatomoides (Hustedt) Lange-Bertalot, et Archibald, Cymbella gracilis (Ehrenberg) Kützing, Cymbella naviculiformis Auerswald, Diatoma mesodon (Ehrenberg) Kützing, and Eunotia exigua (Brébisson) Rabenhorst. Considering COD, two further groups were distinguished. At high values (> 40 mg·l −1) the following taxa were observed: Achnanthes delicatula (Kützing) Grunow, Navicula capitata Ehrenberg var. hungarica (Grunow) Ross, Nitzschia nana Grunow, Pinnularia interrupta W. Smith, Thalassiosira pseudonana Hasle & Heimdal, Thalassiosira weisflogii (Grunow) Fryxell et Hasle. For lower values than 19 mg·l −1, we found Cymbella naviculiformis Auerswald, Diatoma mesodon (Ehrenberg) Kützing, Eunotia exigua (Brébisson) Rabenhorst, Fragilaria arcus (Ehrenberg) Cleve, and Nitzschia epithemioides Grumow var. disputata (Carter) Lange-Bertalot.