Microplastics pollution is widespread in marine ecosystems and a major threat to biodiversity. Nevertheless, our knowledge of the impacts of microplastics in freshwater environments and biota is still very limited. The interaction of microplastics with freshwater organisms and the risks associated with the human consumption of organisms that ingested microplastics remain major knowledge gaps. In this study, we assessed the ingestion of microplastics by Hoplosternum littorale, a common freshwater fish heavily consumed by humans in semi-arid regions of South America. We assessed the abundance and diversity of both plastic debris and other food items found in the gut of fishes caught by local fishermen. We observed that 83% of the fish had plastic debris inside the gut, the highest frequency reported for a fish species so far. Most of the plastic debris (88.6%) recovered from the guts of fish were microplastics (<5 mm), fibres being the most frequent type (46.6%). We observed that fish consumed more microplastics at the urbanized sections of the river, and that the ingestion of microplastics was negatively correlated with the diversity of other food items in the gut of individual fish. Nevertheless, microplastics ingestion appears to have a limited impact on H. littorale, and the consequences of human consumption of this fish were not assessed. Our results suggest freshwater biota are vulnerable to microplastics pollution and that urbanization is a major factor contributing to the pollution of freshwater environments with microplastics. We suggest the gut content of fish could be used as a tool for the qualitative assessment of microplastics pollution in freshwater ecosystems. Further research is needed to determine the processes responsible for the high incidence of microplastics ingestion by H. littorale, and to evaluate the risk posed to humans by the consumption of freshwater fish that ingested microplastics.