Influence of <i>Bradyrhizobia</i> and <i>Calonectria ilicicola</i> on N ₂ O content in soybean nodules

Abstract Nitrous oxide (N 2 O) production presents a significant challenge to achieve agricultural net-zero due to the reliance on nitrogen fertilizers. While nitrogen fertilizers fulfill the high nitrogen demands for producing protein-rich seeds, soybean ( Glycine max ) also forms nodules with Bradyrhizobia to facilitate biological nitrogen fixation. However, some Bradyrhizobia can perform denitrification, a process that reduces NO 3 − to N 2 ; and if the final step in denitrification – mediated by nitrous oxide reductase (encoded by the nosZ gene) – is disrupted, N 2 O will be produced instead of N 2 . This study showed that soybean nodules formed by Bradyrhizobium diazoefficiens USDA110 (which contains nosZ gene) produce less aqueous N 2 O than those formed by Bradyrhizobium japonicum USDA6 (which lacks nosZ gene), underscoring the critical role of nosZ gene in reducing N 2 O content in nodules. Additionally, inoculation with the fungal pathogen Calonectria ilicicola reduced the nodule biomass and the nosZ gene expression of B. diazoefficiens , leading to higher N 2 O content in nodules. Nonetheless, the increase of N 2 O content caused by C. ilicicola became irrelevant in the presence of nosZ gene in soybean nodules. Therefore, field surveys were conducted to reveal regional of nosZ gene prevalence, with lower frequencies observed in the Kaohsiung and Pingtung regions, which are the major vegetable soybean (edamame)-producing areas in Taiwan. These findings highlight the importance of using Bradyrhizobia strains containing nosZ gene to promote sustainable soybean production prior to controlling soil-borne pathogens like C. ilicicola for mitigating N 2 O production, and the integrated consideration of both bacterial and fungal effects would support the net-zero of sustainable farming practices.