Decomposing in-situ grown switchgrass roots as hotspots of microbial activity and N2O emission: the combination of dual-isotope labeling and zymography

High temporal and spatial variability of nitrous oxide (N₂O) emission from soils has been a challenge for the systematic prediction of global climate change. It is attributed to multiple hotspots occurring simultaneously and affecting the N dynamics cumulatively on an ecosystem scale. Understanding the mechanisms and contributing factors of N₂O emission in single hotspots is a prerequisite to overcoming this problem.

We investigated the decomposing switchgrass roots as N₂O hotspots, using isotope dual-labeling (¹⁵N and ¹³C) and zymography. Our main objectives were i) to quantify the contribution of decomposing roots to N₂O emission along with the N contents in the soil (total, organic, and inorganic N) and microbial pools, and ii) to differentiate the extracellular enzyme activity in decomposing roots from the bulk soil, and test if the ‘spatially differentiated’ hotspot enzyme activity indeed related to ‘isotopically differentiated’ hotspot N₂O emissions. We treated the soils of the same origin to have different moisture contents (40% and 70% water-filled pore space, WFPS) and pore size distributions (dominant pores of >30 Ø and < 10 mm Ø, referred to as coarse and fine soil), to evaluate how these variables change the contribution of decomposing roots to the N₂O production.

Our results showed that up to 0.4 % of the root driven N can be emitted as N₂O gas, only within 21 days of the decomposition. Approximately 21 ~35% of root N was transformed to dissolved organic N, while less than 1 % of the root N remained as ammonium (NH₄⁺) and nitrate (NO₃^-) during the incubation. Decreasing NH₄⁺ and increasing NO₃^- suggested nitrification. Surprisingly, both inorganic and organic N content was greater in coarse soil, which likely led to intense hotspots of enzyme activity and N₂O emission. However, there was no difference in microbial biomass between the soil materials. Higher chitinase activity and relatively large pores in coarse soils suggest that the fungal activity was higher in coarse soils compared to the fine soils. Root chitinase activity was positively correlated with the root driven N₂O emission rate (p< 0.01, R²=0.22), supporting that the microbial hotspot formed near the root is the hotspots of N₂O emission.

Our study showed that the intensity of root driven N₂O hotspots can highly depend on the soil physical characteristics, being mediated by decomposed substances, and enzyme activity. Tracking the fate of N during the plant root decomposition can provide a new perspective on the strategies to minimize N₂O emissions in bioenergy systems.