Abstract
The superior antimicrobial properties of silver nanoparticles (Ag NPs) are well-documented, but the exact mechanisms underlying Ag-NP microbial toxicity remain the subject of intense debate. Here, we show that Ag-NP concentrations as low as 10 ppm exert significant toxicity against Bacillus subtilis, a beneficial bacterium ubiquitous in the soil. Growth arrest and chromosomal DNA degradation were observed, and flow cytometric quantification of propidium iodide (PI) staining also revealed that Ag-NP concentrations of 25 ppm and above increased membrane permeability. RedoxSensor content analysis and Phag-GFP expression analysis further indicated that reductase activity and cytosolic protein expression decreased in B. subtilis cells treated with 10–50 ppm of Ag NPs. We conducted X-ray absorption near-edge structure (XANES) and extended X-ray absorption fine structure (EXAFS) analyses to directly clarify the valence and fine structure of Ag atoms in B. subtilis cells placed in contact with Ag NPs. The results confirmed the Ag species in Ag NP-treated B. subtilis cells as Ag2O, indicating that Ag-NP toxicity is likely mediated by released Ag+ ions from Ag NPs, which penetrate bacterial cells and are subsequently oxidized intracellularly to Ag2O. These findings provide conclusive evidence for the role of Ag+ ions in Ag-NP microbial toxicity, and suggest that the impact of inappropriately disposed Ag NPs to soil and water ecosystems may warrant further investigation.
Highlights
Silver nanoparticles (Ag NPs) are the most widely used nanomaterial in healthcare today, with total annual worldwide production estimated to be in the range of 500 tons [1]
Ag NPs can vary broadly in terms of size and morphology [32], and as Ag-NP size has been cited as a determining factor in toxicity [23], it is important to assess the diameter and structure of Ag-NPs used in toxicity experiments, in order to allow for meaningful comparison across studies
Ag NPs used in this study were derived via pulse microwave-assisted synthesis and characterized by field emission scanning electron microscopy (FE-SEM), X-ray diffraction (XRD), and transmission electron microscopy (TEM)
Summary
Silver nanoparticles (Ag NPs) are the most widely used nanomaterial in healthcare today, with total annual worldwide production estimated to be in the range of 500 tons [1]. Nanosilver Microbial Toxicity Mediated by Ag+ Ions products [5,6,7]. Dissolved Ag+ ion concentrations have been found to dictate the toxicity of Ag NPs [12,13], with nanosilver serving as a source for Ag+ ions [14,15]. A recent study further showed that Ag NPs leach Ag+ ions under aerobic but not anaerobic conditions, while Ag-NP microbial toxicity was limited to aerobic conditions [16]. Positive surface charge has been found to protect Bacillus species bacteria against Ag-NP toxicity [17,18]. These studies offer indirect evidence for the role of Ag+ ions in Ag-NP antimicrobial activity
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