Silver nanoparticles inhibit denitrification by altering the viability and metabolic activity of Pseudomonas stutzeri

Abstract

The environmental toxicity of silver nanoparticles (AgNPs) is currently the focus of intensive research. However, the mechanisms underlying the cytotoxic effects of AgNPs on denitrifying microbes have yet to be explicitly demonstrated. Herein, Pseudomonas stutzeri was used to explore the effects of AgNPs on denitrification and cytotoxicity. The denitrification efficiency decreased from 94.91% in the AgNP-free treatment to 87.66%, 60.51% and 36.10% with treatments of 3.125, 6.25 and 12.5 mg/L AgNPs, respectively. The inhibition and delay in the denitrification process from treatment with AgNPs likely occurred through alteration of the viability and metabolic activity of P. stutzeri. Flow cytometry analysis indicated that the early apoptotic rates of P. stutzeri were 8.72%, 30.60%, and 48.60% with treatments of 3.125, 6.25, and 12.5 mg/L AgNPs, respectively. Results for scanning electron microscope (SEM) imaging, ζ-potential analysis, lactate dehydrogenase (LDH) release and malondialdehyde (MDA) production assays demonstrated adsorption of AgNPs on the cell surface, which altered membrane potential and mediated lipid peroxidation; these events eventually resulted in the aberration of cell morphology. Transmission electron microscopy (TEM) images and measurements of Ag content distribution by ICP-MS indicated that AgNPs were easily internalized by P. stutzeri, which increased the accumulation of reactive oxygen species (ROS). Furthermore, the presence of AgNPs also greatly inhibited expression of genes napA, nirS, cnorB, and nosZ, thereby reducing the activities of nitrate reductase (NAR) and nitrite reductase (NIR). These findings will help further our understanding of the mechanism underlying AgNPs cytotoxicity, and provide the means to evaluate the negative effect of nanoparticles in the environment.