Wastewater-based monitoring has been described as a non-invasive approach to assess virus distribution in a specific geographic area. In this study we assess the genetic diversity and concentration of human adenovirus (HAdV) and human polyomavirus JC (JCPyV) in wastewater samples in Rio de Janeiro, Brazil, during the Olympic Games, Rio 2016. Wastewater samples (50 mL) obtained from domestic and hospital sewages were concentrated by the skimmed milk flocculation method and processed using molecular tools. Quantitative polymerase chain reaction using the ABI PRISM 7500 Real Time TaqMan System and TaqMan Universal Master Mix II detected 18 HAdV and 17 JCPyV strains in 95% of those samples (18/19). A mean viral load of 8.6x105 genomic copies (GC)/L and 1.2x107 GC/L, was achieved for HAdV and JCPyV, respectively. Partial nucleotide sequencing using Sanger methodology revealed three HAdV species/ eight serotypes (HAdV B, D, D10, D17, D19, D22, F40 and F41) and seven genotypes/nine subtypes (JCPyV-1B, 2A, 3A, 3B, 4, 6, 7A, 8A and 8B). HAdV-D17 and -D22, as well as genotypes JCPyV-7 and -8, were detected for the first time in the country. The detection of previously undetected viruses in the region demonstrates the importance of our findings, adding data to the epidemiology of those viruses and corroborating the importance of environmental surveillance carried out from wastewater.