The effects of the herbicide Diuron (DIU) and the bactericide Triclosan (TCS) were assessed on laboratory-grown stream biofilms. Four week-old biofilms were exposed in mesocosms to 48-hours of short pulses of either DIU or TCS. The direct and indirect effects of each toxicant on the biofilms, and the subsequent recovery of the biofilms, were evaluated according to structural and functional biomarkers. These parameters were analyzed immediately before exposure, immediately after exposure, and 9 and 16 days post-exposure. DIU caused an increase in diatom mortality (+ 79%), which persisted until the end of the experiment. TCS also affected diatom mortality (+ 41%), although the effect did not appear until 1 week post-exposure. TCS caused an increase in bacterial mortality (+ 45%); however, this parameter returned to normal values 1 week post-exposure. TCS compromised the cellular integrity of the green alga Spirogyra sp., whereas DIU did not. TCS also strongly inhibited phosphate uptake (− 71%), which did not return to normal values until 2 weeks post-exposure. DIU directly affected algae, but barely affected the heterotrophs, whereas TCS seriously impaired bacteria (direct effect) as well as autotrophs (indirect effect). However, the biofilms recovered their normal structure and function within only a few days to a few weeks. These findings demonstrate the capacity of biofilms to cope with periodic inputs of toxicants, but also the risks associated to repeated exposure or multi-contamination in aquatic ecosystems.