Chiral Pesticides Stereoselectively Stimulate N₂O Emissions: An Overlooked Environmental Risk in Agricultural Soils

Agricultural soil is a major source of N₂O emissions due to soil denitrification. However, the influence of pesticides, particularly chiral pesticides, on soil denitrification has often been overlooked. In this study, we investigated the effects and regulatory mechanisms of the widely used chiral pesticide cis-epoxiconazole on denitrification and N₂O emissions. The 2R, 3S-(+)-cis-epoxiconazole ((+)-EPO) enantiomer reduced nitrate removal efficiency by 15.04%-54.49% after 3 days and increased N₂O emissions by 109.27%-163.68% after 35 days of exposure in a microcosm system, whereas the 2S, 3R-(-)-cis-epoxiconazole ((-)-EPO) enantiomer had minimal impact. At a concentration of 0.5 mg/kg, (+)-EPO inhibited β-glucosidase activity by 34.80% and decreased electron transport system activity by 43.70%, thereby impairing electron generation and transfer during denitrification. Expression of the nosZ gene was suppressed by 53.34%, while nitric oxide reductase and nitrous oxide reductase activities were upregulated by 118.89% and downregulated by 46.70%, respectively. These key regulatory changes directly contribute to the increased N₂O emissions observed with (+)-EPO treatment. Additionally, although (+)-EPO caused only minor changes in denitrifying microbial taxa, it increased the relative abundance of Nitrososphaerota and Euryarchaeota by 0.08%-0.10% and 0.40%-0.45%, respectively, after 35 days, enhancing nitrification and indirectly stimulating N₂O emissions. Overall, this study deepens our understanding of how cis-epoxiconazole disrupts denitrification in agricultural soils through microbial, genetic, electronic, and enzymatic pathways.