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Unprecedented nitrogen (N) inputs into terrestrial ecosystems have profoundly altered soil N cycling. Ammonia oxidizers and denitrifiers are the main producers of nitrous oxide (N2O), but it remains unclear how ammonia oxidizer and denitrifier abundances will respond to N loading and whether their responses can predict N-induced changes in soil N2O emission. By synthesizing 101 field studies worldwide, we showed that N loading significantly increased ammonia oxidizer abundance by 107% and denitrifier abundance by 45%. The increases in both ammonia oxidizer and denitrifier abundances were primarily explained by N loading form, and more specifically, organic N loading had stronger effects on their abundances than mineral N loading. Nitrogen loading increased soil N2O emission by 261%, whereas there was no clear relationship between changes in soil N2O emission and shifts in ammonia oxidizer and denitrifier abundances. Our field-based results challenge the laboratory-based hypothesis that increased ammonia oxidizer and denitrifier abundances by N loading would directly cause higher soil N2O emission. Instead, key abiotic factors (mean annual precipitation, soil pH, soil C:N ratio, and ecosystem type) explained N-induced changes in soil N2O emission. Altogether, these findings highlight the need for considering the roles of key abiotic factors in regulating soil N transformations under N loading to better understand the microbially mediated soil N2O emission.