Nitrate content of surface waters results from complex mixing of multiple sources, whose signatures can be modified through N reactions occurring within the different compartments of the whole catchment. Despite this complexity, the determination of nitrate origin is the first and crucial step for water resource preservation. Here, for the first time, we combined at the catchment scale stable isotopic tracers (δ15N and δ18O of nitrate and δ11B) and fecal indicators to trace nitrate sources and pathways to the stream. We tested this approach on two rivers in an agricultural region of SW France. Boron isotopic ratios evidenced inflow from anthropogenic waters, microbiological markers revealed organic contaminations from both human and animal wastes. Nitrate δ15N and δ18O traced inputs from the surface leaching during high flow events and from the subsurface drainage in base flow regime. They also showed that denitrification occurred within the soils before reaching the rivers. Furthermore, this study highlighted the determinant role of the soil compartment in nitrate formation and recycling with important spatial heterogeneity and temporal variability.