Podocnemis unifilis is found throughout the Amazon basin, where it has been harvested as a highly valued source of animal protein since precolonial times. Harvesting rates are also lower during the flooding season due to the availability of habitat for turtle dispersal. This study investigated the effects of the hydrological cycle and human impacts on the abundance and population structure of the species on the middle Xingu River between September 2007 and March 2008. The highest density of basking turtles was recorded during the flooding period, while the number of specimens captured in hand nets was greater during low water. Density and capture rates varied positively with increasing distance from Altamira, the study area's main urban center. Consequently, the abundance of P. unifilis was affected negatively by proximity to urban centers. In the Great Bend area of the Xingu River, densities and abundance were lower, and the mean size of the animals was smaller. These results likely reflect the effects on local stocks of the presence of local gold-prospecting operations and, thus, more harvesting pressure on the P. unifilis population. The sex ratio was biased in favor of males (1.90∶1), possibly as a result of preferential harvesting of females, which are larger in size than males and are often laden with eggs. Further long-term studies are needed to better understand the impact of anthropogenic pressures on long-lived organisms such as turtles. In addition, such information would aid the ecological sustainability of turtles in the Amazon Basin that provide rural communities with subsistence resource.