The objective of this research was to evaluate land use and occupation in the landscape surrounding two neotropical streams of the Upper Paraná River in Brazil and to investigate if the water from these streams induces erythrocyte nuclear abnormalities in Astyanax lacustris. We observed that the areas surrounding the streams are predominantly used for agricultural purposes (73.46% of the total area), followed by buildings (15.21%). Only 8.78% of the area consisted of forest fragments. We observed higher frequencies of micronuclei in erythrocytes of A. lacustris exposed to water from impacted sites (IMP) compared with less impacted sites and the negative control (NC) (p < 0.05). Analysis of nuclear alterations in A. lacrustis erythrocytes showed significant differences in the frequency of notched nuclei and nuclear pyknosis only between the NC and the IMP sites (p < 0.05). Our results indicate that water from the most impacted sites of the microbasin can induce mutagenic and genotoxic effects in A. lacustris.