Acidification of surface water is a persistent threat to aquatic ecosystems throughout the world. Several international policies have led to successful reductions of emissions of acidifying gases, but recovery of surface-water quality has been modest. Associated biological communities continue to suffer, and research on ecosystem functional response is sparse. We investigated shifts in macroinvertebrate communities, shifts in whole-stream metabolism, whether whole-stream metabolism is as sensitive an indicator of acidification as is benthic community structure, and shifts in stream food webs in 9 small forested streams exhibiting a pH gradient from 4.0 to 7.7 and belonging to the GEOchemical MONitoring (GEOMON) Network in the Czech Republic. Acidification was related to changes in macroinvertebrate communities including reduced taxon richness and reduced Ephemeroptera family richness and abundance. Whole-stream metabolism was not significantly related to stream pH and may be less sensitive to acidification than macroinvertebrate community metrics. ¹³C and ¹⁵N isotopic signatures of the Chironomidae and Leuctridae shifted away from periphyton isotopic signatures and toward isotopic signatures of allochthonous food sources at acidified sites, a result suggesting either species-level dietary changes or replacement of species by those that rely more heavily on detritus. Our results document long-lasting changes in the ecosystem structure of these small streams in response to acidification. Differences in the responses of structural and functional metrics to acidification have implications for stream-monitoring programs.