Combining effects of nitrogen fertilizer and biochar on soil N2O emissions and microbial community in a subtropical rapeseed-soybean rotation

Reducing nitrogen fertilizer application is a key strategy for mitigating soil N₂O emissions in agriculture. Biochar has great potential in reducing excessive fertilizer use. However, the interactive effects of biochar and nitrogen (N) fertilization on N₂O emissions are poorly understood. This study investigated how varying N fertilization (H: 100 %, M: 75 %, and L: 50 % of the conventional urea application rate) and biochar application rates (B0: 0 t ha⁻², B1: 15 t ha⁻², and B2: 60 t ha⁻²) affect N₂O emissions, microbial community, and the abundance of N₂O-related functional genes (amoA, nirS, nirK, and nosZ) in a subtropical oilseed rape (Brassica napus L.)-soybean (Glycine max (L.) Merrill) rotation system. Compared to control treatments (B0), biochar increased soil cumulative N₂O emissions by 55–61.5 % in the oilseed season and 200–245 % in the soybean season. Biochar also significantly increased microbial diversity and altered bacterial community composition, with notable shifts in the relative abundance of key phyla such as Bacillota, Bacteroidota, Armatimonadota, and Nitrospirota. These effects were more pronounced under higher biochar application, likely driven by increases in soil total carbon (TC), nitrogen (TN), and ammonium (NH₄⁺-N). Biochar increased the abundance of N₂O-related genes but had no significant effect on the (nirS + nirK)/nosZ ratio. Co-occurrence network analysis further revealed that biochar altered microbial interactions in a rate- and season-dependent manner, with high rate simplifying the network and potentially disrupting community stability, especially during the soybean season. In contrast, N fertilizer had limited effects on N₂O emissions, microbial diversity or community structure. Partial least squares path modeling (PLS-PM) suggested that biochar increased N₂O emissions primarily by enhancing soil TC, TN, pH, and denitrification process, while N fertilizer may affect N₂O emissions through nitrification. These findings highlight the need for optimized biochar and fertilizer management strategies and emphasize the importance of identifying N₂O production pathways and conducting long-term field studies to ensure the sustainable use of biochar in agriculture.