Stability and Microbial Toxicity of Silver Nanoparticles under Denitrifying Conditions

While the antibacterial effect of silver nanoparticles (AgNPs) on environmentally beneficial microbes has drawn considerable attention, the stability and microbial toxicity of AgNPs in a system where nitrate reduction is the dominant terminal electron-accepting process remain understudied. Here, we explore the impact of citrate-coated AgNPs (cit-AgNPs) on the growth and metabolism of two metal-sensitive and one nonsensitive bacterial strains under denitrifying conditions. Dose–response analysis revealed that in contrast to the bacteriostatic effect exhibited at 1 ppm, 5 ppm cit-AgNPs were bactericidal to the metal-sensitive strains. It was observed that the growth of the cells initiated Ag­(I) formation, and the supplement of chloride (2.7 mM) to the cultures substantially mitigated the bactericidal capacity of cit-AgNPs, indicating that AgNP dissolution to ionic Ag­(I) played a key role in AgNP toxicity. Abiotic experiments confirmed that nitrite, not nitrate, had the capacity to oxidize cit-AgNPs. Transcriptomic analysis revealed that (i) the gene encoding for membrane stress was upregulated proportionally to cit-AgNP concentrations; (ii) cit-AgNPs and Ag­(I) at higher levels upregulated genes involved in oxidative stress and iron–sulfur clusters, whereas expressions of the genes responsible for electron transport, ATP synthesis, and denitrification were substantially repressed; (iii) the addition of chloride significantly altered the level of transcriptional profiles of all of the genes. These results not only provide evidence of abiotic AgNP oxidation by metabolic intermediate nitrogen species but also suggest that AgNPs and Ag­(I) may induce differential toxicity modes to prokaryotes. Our findings reinforce the importance of evaluating the potential ecological toxicity and risks associated with the transformation of nanomaterials.