Energy flows from land to sea and between pelagic and benthic environments have the potential to increase the connectivity between estuaries and adjacent ecosystems as well as among estuarine habitats. To identify such energy flows and the main trophic pathways of energy transfer in the Minho River estuary, we investigated the spatial and temporal fluctuations of carbon and nitrogen stable isotope ratios in benthic (and their potential food sources) and epibenthic consumers. Sampling was conducted along the estuarine salinity gradient from winter to summer of 2011. We found that the carbon (δC = C/C) and nitrogen (δN = N/N) stable isotope ratios of the most abundant benthic and epibenthic consumers varied along the salinity gradient. The δC values increased seaward, whereas the opposite pattern was found for the δN, especially during the summer. The stable isotope ratios revealed two trophic pathways in the Minho estuary food web. The first pathway is supported by phytoplankton and represented by filter feeders such as zooplankton and some deposit feeders, particularly amphipods and polychaetes. The second pathway is supported by detritus and composed essentially of deposit feeders, which by being consumed, allow detritus to be incorporated into higher trophic levels. Spatial and temporal feeding variations in the estuarine benthic food web are driven by hydrology and proximity to adjacent ecosystems (terrestrial, marine). During high river discharge periods, the δC (ca. -28‰) and C: N (>10) values suggested an increase of terrestrial-derived OM to the particulate OM pool, which was then used by suspension feeders. During low river discharge periods, marine intrusion increased upriver, which was reflected in benthic consumers' C-enriched stable isotope values. No relationship was found between food quality (phytoplankton vs. detritus) and food chain length because the lowest and highest values were associated with freshwater and saltmarsh areas, respectively, both dominated by the detrital pathway. This study demonstrates that benthic consumers enhance the connectivity between estuaries and its adjacent ecosystems by utilizing subsidies of terrestrial and marine origin and that benthic-pelagic coupling is an important energy transfer mechanism to the benthic food web.