The public water supply of the Rio de Janeiro metropolitan region is highly dependent on transposition from the Paraíba do Sul River: 70% of the water is diverted to the Piraí River and then passes through a series of other rivers and reservoirs, finally discharging in the Guandu River. During this path, the water is exposed to many sources of pollution. This makes the quality of the raw water that reaches the Guandu Treatment Station (WTS) highly vulnerable. This article reports the analysis of the cytogenotoxic potential of water samples collected at four different points along the Piraí River downstream from the transposition point, utilizing the Allium cepa test system. The samples were collected in two periods, the dry and wet seasons. The water at all four collection points presented some level of cytogenotoxicity, with the presence in the test cells of large nucleoli, multiple nucleoli, nuclear buds, lagging chromosomes, sticky chromosomes, karyorrhexis, cytoplasmic shrinkage and changes of the mitotic index. The samples collected during the dry season had a larger number of cells with alterations, indicating that the cytogenotoxic potential varies in function of the time of year, depending on the volume of contaminated effluents. The results obtained along with data from the Rio de Janeiro State Environmental Institute (INEA) for the same period reveal the importance of monitoring along with proper sanitation and sewage treatment, and that the presence of pollutants not only hampers water treatment, but also poses risks to organisms at different trophic levels, including humans.