Abstract:The drainages of the Iguaçu River basin, as well as the main channel of the river, have peculiar characteristics resulting from geomorphological processes in this area, including the formation of the Iguaçu River Falls. This slope in the lower portion of the basin is a biogeographical barrier to many organisms. In this study was inventoried the fish fauna of streams of the lower Iguaçu River basin, evaluating possible differences in the species composition upstream and downstream of this biogeographical barrier. Sampling were conducted between 2004 and 2013, in five streams, three upstream and two downstream of the Iguaçu River Falls, using electrofishing. The nMDS analysis was run to investigate possible groupings of similar fauna between the streams sampled. The scores of this test were tested as to the significance of groupings with the Hotelling T 2 test. The indicator value method (IndVal) was used to detect the distribution of species among the groups of the streams upstream and downstream of the Iguaçu River Falls. We collected 18,908 individuals of six orders, 11 families, and 40 species. Siluriformes and Characiformes had the highest species richness; Cyprinodontiformes presented the highest abundance. Considering the species recorded, 21 are considered natives to the Iguaçu River basin, including 15 endemic, wich were found only in streams upstream of the falls. Additional 18 species were verified only in the streams downstream of such barrier. Four species were common to both stretches. The axis 1 of the nMDS separated two groups: streams upstream (S1, S2 and S3) and streams downstream of the Iguaçu River Falls (S4 and S5). The indicator species analysis also indicated a distinction between the groups of streams, that were significantly different (Hotelling's T 2 = 234.36, p ˂ 0.0001). The number of endemic species in the streams upstream of the Falls (15 spp.) evidences a significant effect of isolation promoted by the Iguaçu River Falls, and confirms the role of this barrier in the vicariant processes and endemism typical of this basin. These results emphasize the importance of conserving these ecosystems, once the extinction of species in this region means the irreversible loss of them. Resumo: As drenagens da bacia do rio Iguaçu, bem como o canal principal do rio, possuem características peculiares resultantes de processos geomorfológicos dessa área, incluindo a formação das Cataratas do rio Iguaçu. Esse desnível na porção inferior da bacia constitui uma barreira biogeográfica para muitos organismos. Neste trabalho inventariamos a fauna de peixes de riachos na porção do baixo rio Iguaçu, avaliando possíveis diferenças na composição de espécies acima e abaixo dessa barreira biogeográfica. Entre 2004 e 2013, foram amostrados cinco riachos, sendo três localizados acima das Cataratas do Iguaçu e dois abaixo, por meio de pesca elétrica. Para investigar possíveis padrões de agrupamentos de fauna similares entre os riachos amostrados foi