Short-Term Simulated NH4+/NO3− Inputs Reorganize Soil Microbial Communities and Modify N-Cycling Functional Potential in Temperate Forests

Forest soil nitrogen (N) turnover and nitrous oxide (N2O) release are strongly influenced by atmospheric N deposition. Although the effects of individual inorganic N species on soil biogeochemistry are well documented, the consequences of changing their relative supply for N2O formation and the underlying microbial pathways remain less clear. We therefore examined soil N2O emissions, microbial communities, and N-transformation potential across a gradient of ammonium-to-nitrate (NH4+/NO3−) inputs in a temperate forest soil from Northeast China. Seven treatments covering NH4+/NO3− ratios from 1:5 to 5:1 were established. Soil physicochemical properties and N2O emissions were measured, while microbial taxonomic profiles and N-cycling genes were characterized by metagenomic sequencing. Altering the composition of inorganic N reorganized the microbial community, including the relative representation of archaeal, bacterial, and eukaryotic groups. N2O release was greatest when NH4+ and NO3− were supplied in equal proportions, a pattern that may reflect a stronger contribution of nitrification-associated processes. Across the treatment gradient, the response of N2O emissions to inorganic N composition was nonlinear. By contrast, the 1:2.5 NH4+/NO3− treatment produced comparatively low N2O emissions and contained a greater representation of genes associated with N2O reduction, consistent with enhanced microbial potential for N2O consumption. Under the more nitrate-dominated 1:5 treatment, genes involved in dissimilatory nitrate reduction to ammonium (DNRA) were enriched, indicating a shift in the potential routes of nitrate transformation. Metagenomic data further revealed marked variation in genes linked to nitrification, denitrification, and biological N fixation. In comparison with these compositional and functional responses, microbial diversity changed relatively little, possibly because overlapping functions were distributed among different taxa under contrasting N inputs. Overall, the findings show that the balance between ammonium and nitrate can affect forest-soil N2O emissions by restructuring microbial assemblages and their potential to transform N. These responses emphasize the need to consider the composition, rather than only the amount, of N deposition when evaluating microbial regulation of ecosystem N cycling and greenhouse-gas release. Field studies are still needed to test the generality of these patterns and their relevance to N management.