Negligible particle-specific antibacterial activity of silver nanoparticles.

For nearly a decade, researchers have debated the mechanisms by which AgNPs exert toxicity to bacteria and other organisms. The most elusive question has been whether the AgNPs exert direct "particle-specific" effects beyond the known antimicrobial activity of released silver ions (Ag(+)). Here, we infer that Ag(+) is the definitive molecular toxicant. We rule out direct particle-specific biological effects by showing the lack of toxicity of AgNPs when synthesized and tested under strictly anaerobic conditions that preclude Ag(0) oxidation and Ag(+) release. Furthermore, we demonstrate that the toxicity of various AgNPs (PEG- or PVP- coated, of three different sizes each) accurately follows the dose-response pattern of E. coli exposed to Ag(+) (added as AgNO(3)). Surprisingly, E. coli survival was stimulated by relatively low (sublethal) concentration of all tested AgNPs and AgNO(3) (at 3-8 µg/L Ag(+), or 12-31% of the minimum lethal concentration (MLC)), suggesting a hormetic response that would be counterproductive to antimicrobial applications. Overall, this work suggests that AgNP morphological properties known to affect antimicrobial activity are indirect effectors that primarily influence Ag(+) release. Accordingly, antibacterial activity could be controlled (and environmental impacts could be mitigated) by modulating Ag(+) release, possibly through manipulation of oxygen availability, particle size, shape, and/or type of coating.

Differential effect of common ligands and molecular oxygen on antimicrobial activity of silver nanoparticles versus silver ions.

The antibacterial activity of silver nanoparticles (AgNPs) is partially due to the release of Ag(+), although discerning the contribution of AgNPs vs Ag(+) is challenging due to their common co-occurrence. We discerned the toxicity of Ag(+) versus a commercially available AgNP (35.4 ± 5.1 nm, coated with amorphous carbon) by conducting antibacterial assays under anaerobic conditions that preclude Ag0 oxidation, which is a prerequisite for Ag(+) release. These AgNPs were 20× less toxic to E. coli than Ag(+) (EC50: 2.04 ± 0.07 vs 0.10 ± 0.01 mg/L), and their toxicity increased 2.3-fold after exposure to air for 0.5 h (EC50: 0.87 ± 0.03 mg/L) which promoted Ag(+) release. No significant difference in Ag(+) toxicity was observed between anaerobic and aerobic conditions, which rules out oxidative stress by ROS as an important antibacterial mechanism for Ag(+). The toxicity of Ag(+) (2.94 µmol/L) was eliminated by equivalent cysteine or sulfide; the latter exceeded the solubility product equilibrium constant (K(sp)), which is conducive to silver precipitation. Equivalent chloride and phosphate concentrations also reduced Ag(+) toxicity without exceeding K(sp). Thus, some common ligands can hinder the bioavailability and mitigate the toxicity of Ag(+) at relatively low concentrations that do not induce silver precipitation. Furthermore, low concentrations of chloride (0.1 mg/L) mitigated the toxicity of Ag(+) but not that of AgNPs, suggesting that previous reports of higher AgNPs toxicity than their equivalent Ag(+) concentration might be due to the presence of common ligands that preferentially decrease the bioavailability and toxicity of Ag(+). Overall, these results show that the presence of O2 or common ligands can differentially affect the toxicity of AgNPs vs Ag(+), and underscore the importance of water chemistry in the mode of action of AgNPs.

Effect of bare and coated nanoscale zerovalent iron on tceA and vcrA gene expression in Dehalococcoides spp.

Nanoscale zerovalent iron (NZVI) can be used to dechlorinate trichloroethylene (TCE) in contaminated aquifers. Dehalococcoides spp. is the only microbial genus known to dechlorinate TCE to ethene as a respiratory process. However, little is known about how NZVI affects the expression of genes coding for reductive dechlorination. We examined a high-rate TCE-dechlorinating mixed culture which contains organisms similar to known Dehalococcoides to study the effects of NZVI on the expression of two model genes coding for reductive dehalogenases (tceA and vcrA). A novel pretreatment approach, relying on magnetic separation of NZVI prior to reverse transcription qPCR (to avoid RNA adsorption by NZVI), was developed and used with relative quantification (relative to 16S rRNA as endogenous housekeeping gene) to quantify reductive dehalogenase gene expression. Both tceA and vcrA were significantly down-regulated (97- and 137-fold, respectively) relative to baseline (time 0) conditions after 72-h exposure to chlorinated ethenes (0.12 ± 0.03 mg/L cis-DCE, 0.69 ± 0.11 mg/L t-DCE, and 0.54 ± 0.16 mg/L VC) and bare-NZVI (1 g-NZVI/L). However, coating NZVI with an olefin maleic acid copolymer (a common approach to enhance its mobility in aquifers) overcame this significant inhibitory effect, and both tceA and vcrA were up-regulated (3.0- and 3.5-fold, respectively) after 48-h exposure. Thus, NZVI coating might enhance the expression of dechlorinating genes and the concurrent or sequential participation of Dehalococcoides spp. in the remediation process.

Effects of nano-scale zero-valent iron particles on a mixed culture dechlorinating trichloroethylene.

Nano-scale zero-valent iron particles (NZVI) are increasingly being used to treat sites contaminated with chlorinated solvents. This study investigated the effect of NZVI on dechlorinating microorganisms that participate in the anaerobic bioremediation of such sites. NZVI can have a biostimulatory effect associated with water-derived cathodic H(2) production during its anaerobic corrosion (730+/-30 micromol H(2) was produced in 166 h in abiotic controls with 1 g/L NZVI) or an inhibitory effect upon contact with cell surfaces (assessed by transmission electron microscopy). Methanogens, which are known to compete for H(2) with dechlorinators, were significantly biostimulated by NZVI and methane production increased relative to NZVI-free controls from 58+/-5 to 275+/-2 micromol. In contrast, bacteria dechlorinating TCE were inhibited by NZVI, and the first-order degradation rate coefficient decreased from 0.115+/-0.005 h(-1) (R(2)=0.99) for controls to 0.053+/-0.003 h(-1) (R(2)=0.98) for treatments with 1 g/L NZVI. Ethene production from TCE was initially inhibited by NZVI, but after 331 h increased to levels observed for an NZVI-free system (7.6+/-0.3 micromol ethene produced in 502 h compared to 11.6+/-0.5 mmol in the NZVI-free system and 3.8+/-0.3 micromol ethene for NZVI alone). Apparently, cathodic H(2) was utilized as electron donor by dechlorinating bacteria, which recovered following the partial oxidation and presumably passivation of the NZVI. Overall, these results suggest that reductive treatment of chlorinated solvent sites with NZVI might be enhanced by the concurrent or subsequent participation of bacteria that exploit cathodic depolarization and reductive dechlorination as metabolic niches.

[Microbial reductive dechlorination of TCE with nano iron serving as electron donor].

A trichloroethylene (TCE) dechlorinating enrichment (Dehalococcoides spp.), which was isolated from soil of chlorinated ethene contaminated site, was used to investigate whether nano-scale zero valent iron (NZVI) could serve as electron donor for this consortium via cathodic H2 production during anaerobic corrosion. The results show that in the presence of methanol serving as electron donor, dechlorinating culture of 25 fold dilution [(2.0 +/- 0.44) x 10(5) cell/mL] degraded 20 mg/L TCE completely in 96 h, which was accompanied by the production of 2.706 micromol ethene in 190 h. Methanol-free control caused partial degradation of TCE to primarily cis-DCE in 96 h, with only 0.159 micromol ethene produced in 190 h. This indicates bacteria cannot reduce TCE to ethene without electron donor. But when 4 g/L NZVI was added as sole electron donor, this dechlorinating culture degraded 20 mg/L TCE into ethene and vinyl chloride (VC) in 131 h at a speed higher than that by NZVI alone. Compared to 2.706 micromol ethene produced by Dehalococcoides spp. with methanol added as the electron donor, there was only 1.187 micromol ethene produced by bacteria with NZVI serving as the electron donor, which means NZVI has a potential toxicity on Dehalococcoides spp.. At the meantime, 0.109 micromol acetylene was produced in 190 h, which was relatively lower than 0.161 micromol produced by NZVI alone, indicating bacteria competed with NZVI under electron deficient condition. In conclusion, NZVI could serve as electron donor and support dechlorination activity for Dehalococcoides spp. which could enhance the application of NZVI and usage of dechlorinating culture as a polishing strategy in future ground water remediation.