Chronic high-dose silver nanoparticle exposure stimulates N₂O emissions by constructing anaerobic micro-environment

Silver nanoparticles (Ag-NPs) were found to be responsible for nitrous oxide (N₂O) generation; however, the mechanism of Ag-NP induced N₂O production remains controversial and needs to be elucidated. In this study, chronic Ag-NP exposure experiments were conducted in five independent sequencing batch biofilm reactors to systematically assess the effects of Ag-NPs on N₂O emission. The results indicated that a low dose of Ag-NPs (< 1 mg/L) slightly suppressed N₂O generation by less than 22.99% compared with the no-Ag-NP control method. In contrast, a high dose (5 mg/L) of Ag-NPs stimulated N₂O emission by 67.54%. ICP-MS and SEM-EDS together revealed that high Ag-NP content accumulated on the biofilm surface when exposed to 5 mg/L Ag-NPs. N₂O and DO microelectrodes, as well as N₂O isotopic composition analyses, further demonstrated that the accumulated Ag-NPs construct the anaerobic zone in the biofilm, which is the primary factor for the stimulation of the nitrite reduction pathway to release N₂O. A metagenomic analysis further attributed the higher N₂O emissions under exposure to a high dose of Ag-NPs to the higher relative abundance of narB and nirK genes (i.e. 1.52- and 1.29-fold higher, respectively). These findings collectively suggest that chronic exposure to high doses of Ag-NPs could enhance N₂O emissions by forming anaerobic micro-environments in biofilms.