The rivers from the region of Catalão, Southeast Goiás State, Brazil, are exposed to intense anthropogenic influences including agricultural activities, industry, and urban waste. The aim of this study was to determine the severity of water pollution by conducting an experiment involving in situ biomonitoring of water constituents on genotoxicity in fish inhabiting these sites. The genotoxicity of three sites of the region were analyzed utilizing the micronucleus (MN) test. It was of interest to determine whether there were differences between sampling sites such as urban perimeter, agriculture, and fertilizer industry in control, and monitored species including Astyanax fasciatus, Astyanax altiparanae, and Characidium fasciatum. Data demonstrated that the species at sites 1, 2, and 3 exhibited a marked increase in frequency of MN compared to fish from site 4. Significant elevation in frequency of MN occurred in erythrocytes of A. fasciatus and A. altiparanae at sites 1 and 2. At site 3 higher frequencies of MN were observed in C. fasciatum. MN induction in C. fasciatum was correlated with chromium levels in water and sediment, while A. fasciatum and A. altiparanae showed an association with zinc in water and sediment. Data suggest that benthic and nektonic fish species display different sensitivities in relation to anthropogenic contaminant influences.