Genetic potential for N₂O metabolism in tree tissues: Insights into nitrogen cycling gene abundance and nosZ diversity across trees

Nitrous oxide (N ₂ O) is a potent greenhouse gas, and microorganisms play a crucial role in its metabolism. While soil microbial roles in N ₂ O cycling are well studied, there is a major knowledge gap regarding the distribution and diversity of these microbes within tree ecosystems. In this study, we aimed to comprehensively assess the nitrogen (N) cycling gene abundance and the diversity of N ₂ O-reducing microorganisms in shoots (leaves and terminal branches) and wood cores of four tree categories — European beech ( Fagus sylvatica ), European hornbeam ( Carpinus betulus ), birch ( Betula pendula and Betula pubescens ) and Norway spruce ( Picea abies ) across long transect. We assessed N ₂ O exchange through shoot incubation experiments and measured internal N ₂ O concentrations in stem wood. Inorganic N compounds were studied as indicators of microbial transformation, and a targeted metagenomic approach was used to analyze the relative abundance of N-cycling genes and nosZ clade I and II diversity. Our study revealed that hornbeam shoots showed potential N₂O emissions, while beech shoots indicated N₂O consumption in the incubation study. Birch had the highest internal stem wood N₂O concentration, and beech the lowest when compared to the ambient concentration. Metagenomic analysis confirmed the presence of key nitrification and denitrification genes in both tissues, with nosZ genes abundant in spruce shoots, birch wood cores, and beech wood cores—clade I dominating over clade II and Rhizobiales prevalent within clade I. These findings provide new insights into tree microbiome and its contribution to N ₂ O exchange in tree-associated environments.