Proper water quality monitoring is a valuable tool for water resource management, helping to identify polluting sources and risks related to the use of water resources. One of the main types of contamination found in Brazilian water bodies is fecal contamination, which originates mainly from point source pollution through wastewater disposal. Thus, this study analyzed water quality monitoring data from the responsible environmental body (Minas Gerais Institute of Water Management, IGAM), related to the fecal contamination indicator (FCI), for the years 2000–2018. This was done for the Minas Gerais portion of the São Francisco River basin (SFRB-MG), one of the most important basins in the country. The 10 sub-basins in the area were compared using statistical tools. The work found significant differences between the sub-basins in terms of FCI concentration, highlighting the most impacted ones (SF2, SF3, and SF5) as also being the most densely populated. It is necessary to invest in sanitation measures in order to ensure that water resources are preserved, as well as to reduce the public health risks of downstream municipalities that are supplied with previously contaminated water.