Seasonal hydrology is assumed to be an important reason why the Lower Mekong Basin supports highly productive and biodiverse inland fisheries. We used C and N stable isotope ratios of tissue samples to estimate primary production sources supporting fish biomass in the Mekong and three large tributaries in Cambodia. We used a Bayesian mixing model to estimate relative contributions of four alternative production sources seston, benthic algae, riparian grasses, and riparian macrophytes. There was little seasonal variation in isotopic signatures of riparian plants, but benthic algae and seston showed large seasonal shifts in carbon ratios. Seston and benthic algae were the most important production sources supporting fish biomass overall during the dry season, and riparian vegetation was the most important source during the wet season. Sources contributed differentially to biomass of trophic and habitat guilds, especially during the dry season. A dam on the upper Sesan River has changed hydrology, channel geomorphology, and other factors and, compared with the other three rivers, its fish biomass appears to derive from algae to a greater extent.