We investigated photoinhibition on natural communities of ammonia oxidizing (AO) archaea (AOA) and bacteria (AOB) embedded in complex stream biofilms, and its implications on nitrogen uptake at biofilm scale. Based on the strong photoinhibition previously exhibited by free living and cultured AOA and AOB, we expected AO activity to decrease in biofilms experimentally exposed to light, reducing the contribution of microbial nitrification to ammonium uptake. To test it, we conducted light manipulation experiments in mesocosms using biofilms naturally developed on stream cobbles sides both exposed to light (light-side) and facing the sediment (dark-side). We observed a strong AO photoinhibition in dark-side biofilms, accompanied by either biofilm-scale decreases in nitrification or increases in nitrogen uptake likely by heterotrophic activity. Conversely, in light-side biofilms photoinhibition was not observed suggesting that photoautotrophic layers may protect AO in situ by a sunshade effect. Experimental light and dark alternation cycles stimulated AO, enhancing both nitrification and ammonium uptake. These results support photoinhibition effects on natural AO communities, especially in biofilms developed under ambient dark conditions, whereas this effect seems to be buffered in biofilms developed under light conditions. Therefore, the contribution of nitrification to ammonium uptake at biofilm scale should consider not only the physiological study of AOA and AOB, but also the environmental conditions and community structure at the habitat microscale, since they may counterbalance, among others, the damaging light effects under natural conditions.