Denitrifier activity drives diurnal N2O flux dynamics in a thawing arctic permafrost

Diurnal variation of nitrous oxide (N2O) fluxes was recently shown to occur across terrestrial ecosystems, typically with higher daytime emissions. However, the underlying microbial mechanisms remain poorly understood. Here, we investigated the microbial drivers of daily N2O dynamics in a permafrost palsa mire, where a higher net N2O sink has been observed during dark than under light conditions. By integrating day night flux measurements with molecular analyses, we could directly link short-term temporal patterns of N2O exchange with diurnal shifts in microbial activity. Quantitative PCR revealed the presence of denitrification genes nirK and nirS, associated with N2O emissions in soils, and nosZ clades I and II, involved in N2O consumption. However, only nirK and nosZ clade I were transcriptionally active. The diurnal pattern of nirK/nosZ gene expression largely aligned with the diurnal pattern of N2O fluxes, with elevated ratios during daytime, particularly in wet and productive fen habitats. Metatranscriptomic analyses further revealed higher daytime expression ratios of N2O producing relative to N2O reducing genes ((p450nor + norB + hao)/nosZ), in addition to high nirK/nosZ expression ratios. Members of Alphaproteobacteria, particularly Rhizobiales and Bradyrhizobium, consistently expressed both nirK and nosZ across the diel cycle. Analysis of 1,753 metagenome-assembled genomes obtained from the present and previous work further indicate that partial denitrifiers are key players in regulating N2O fluxes in arctic permafrost palsa mire. Together, our results provide new insights into the microbial basis for diurnal N2O dynamics, which should be considered when estimating ecosystem-climate feedback in terrestrial ecosystems in the Arctic and beyond.