Denitrification and anaerobic ammonium oxidation (anammox) are the major microbial processes responsible for global nitrogen (N) loss. Yet, the relative contributions of denitrification and anammox to N loss across contrasting terrestrial and aquatic ecosystems worldwide remain unclear, hampering capacities to predict the human alterations in the global N cycle. Here, a global synthesis including 3240 observations from 199 published isotope pairing studies is conducted and finds that denitrification governs microbial N loss globally (79.8±0.4%). Significantly, anammox is more important in aquatic than terrestrial ecosystems worldwide and can contribute up to 43.2% of N loss in global seawater. Global maps for N loss associated with denitrification and anammox are further generated and show that the contribution of anammox to N loss decreases with latitude for soils and sediments but generally increases with substrate depth. This work highlights the importance of anammox as well as denitrification in driving ecosystem N losses, which is critical for improving the current global N cycle model and achieving sustainable N management.