Concerns about rising greenhouse gas (GHG) concentrations have spurred the promotion of no-tillage practices as a means to stimulate carbon storage and reduce CO2 emissions in agro-ecosystems. Recent research has ignited debate about the effect of earthworms on the GHG balance of soil. It is unclear how earthworms interact with soil management practices, making long-term predictions on their effect in agro-ecosystems problematic. Here we show, in a unique two-year experiment, that earthworm presence increases the combined cumulative emissions of CO2 and N2O from a simulated no-tillage (NT) system to the same level as a simulated conventional tillage (CT) system. We found no evidence for increased soil C storage in the presence of earthworms. Because NT agriculture stimulates earthworm presence, our results identify a possible biological pathway for the limited potential of no-tillage soils with respect to GHG mitigation.