15N tracers and microbial analyses reveal in situ N2O sources in contrasting water regimes of a drained peatland forest

Managed peatlands are a significant source of nitrous oxide (N₂O), a powerful greenhouse gas and stratospheric ozone depleter. Due to the complexity and diversity of microbial N₂O processes, different methods such as tracer, isotopomer, and microbiological technologies are required to understand these processes. The combined application of different methods helps to precisely estimate these processes, which is crucial for the future management of drained peatlands, and to mitigate soil degradation and negative atmospheric impact. In this study, we investigated N₂O sources by combining tracer, isotopomer, and microbial analysis in a drained peatland forest under flooded and drained treatments. On average, the nitrification genes showed higher abundances in the drained treatment, and the denitrification genes showed higher abundances in the flooded treatment. This is consistent with the underlying chemistry, as nitrification requires oxygen while denitrification is anaerobic. We observed significant differences in labelled N₂O fluxes between the drained and flooded treatments. The emissions of N₂O from the flooded treatment were nearly negligible, whereas the N₂O evolved from the nitrogen-15 (¹⁵N)-labelled ammonium (¹⁵NH₄⁺) in the drained treatment peaked at 147 μg ¹⁵N m^-2 h^-1. This initially suggested nitrification as the driving mechanism behind N₂O fluxes in drained peatlands, but based on the genetic data, isotopic analysis, and N₂O mass enrichment, we conclude that hybrid N₂O formation involving ammonia oxidation was the main source of N₂O emissions in the drained treatment. Based on the ¹⁵N-labelled nitrate (¹⁵NO₃^-) tracer addition and gene copy numbers, the low N₂O emissions in the flooded treatment came possibly from complete denitrification producing inert dinitrogen. At atomic level, we observed selective enrichment of mass 45 of N₂O molecule under ¹⁵NH₄⁺ amendment in the drained treatment and enrichment of both masses 45 and 46 under ¹⁵NO₃^- amendment in the flooded treatment. The selective enrichment of mass 45 in the drained treatment indicated the presence of hybrid N₂O formation, which was also supported by the high abundances of archaeal genes.