Exposure to silver nanoparticles (AgNPs) may alter the structure and function of freshwater ecosystems. However, there remains a paucity of studies investigating the effects of AgNP exposure on freshwater communities in the natural environment where interactions with the ambient environment may modify AgNP toxicity. We used nutrient diffusing substrates to determine the interactive effects of AgNP exposure and phosphorus (P) enrichment on natural assemblages of periphyton in three Canadian Shield lakes. The lakes were all phosphorus poor and spanned a gradient of dissolved organic carbon availability. Ag slowly accumulated in the exposed periphyton, which decreased periphyton carbon and chlorophyll a content and increased periphyton C:P and N:P in the carbon rich lakes. We found significant interactions between AgNP and P treatments on periphyton carbon, autotroph standing crop and periphyton stoichiometry in the carbon poor lake such that P enhanced the negative effects of AgNPs on chlorophyll a and lessened the impact of AgNP exposure on periphyton stoichiometry. Our results contrast with those of other studies demonstrating that P addition decreases metal toxicity for phytoplankton, suggesting that benthic and pelagic primary producers may react differently to AgNP exposure and highlighting the importance of in situ assays when assessing potential effects of AgNPs in fresh waters.