||Artificial light at night (ALAN) is recognized as a contributor to environmental change and a biodiversity threat on a global scale. Despite its widespread use and numerous potential ecological effects, few studies have investigated the impacts on aquatic ecosystems and primary producers. Light is a source of energy and information for benthic autotrophs that form the basis of food webs in clear, shallow waters. Artificial night-time illumination may thus affect biomass and community composition of primary producers. We experimentally mimicked the light conditions of a light-polluted area (approximately 20 lux, white LED) in streamside flumes on a sub-alpine stream. We compared the biomass and community composition of periphyton grown under ALAN with periphyton grown under a natural light regime in two seasons using communities in early (up to 3 weeks) and later (4â??6 weeks) developmental stages. In early periphyton, ALAN decreased the biomass of autotrophs in both spring (57% at 3 weeks) and autumn (43% at 2 weeks), decreased the proportion of cyanobacteria in spring (54%), and altered the proportion of diatoms in autumn (11% decrease at 2 weeks and 5% increase at 3 weeks). No effects of ALAN were observed for later periphyton. Further work is needed to test whether streams with frequent physical disturbances that reset the successional development of periphyton are more affected by ALAN than streams with more stable conditions. As periphyton is a fundamental component of stream ecosystems, the impact of ALAN might propagate to higher trophic levels and/or affect critical ecosystem functions.