Different routes, same pathways: molecular mechanisms under silver ion and nanoparticle exposures in the soil sentinel Eisenia fetida

Use of nanotechnology products is increasing; with silver (Ag) nanoparticles particularly widely used. A key uncertainty surrounding the risk assessment of AgNPs is whether their effects are driven through the same mechanism of action that underlies the toxic effects of Ag ions. We present the first full transcriptome study of the effects of Ag ions and NPs in an ecotoxicological model soil invertebrate, the earthworm Eisenia fetida. Gene expression analyses indicated similar mechanisms for both silver forms with toxicity being exerted through pathways related to ribosome function, sugar and protein metabolism, molecular stress, disruption of energy production and histones. The main difference seen between Ag ions and NPs was associated with potential toxicokinetic effects related to cellular internalisation and communication, with pathways related to endocytosis and cilia being significantly enriched. These results point to a common final toxicodynamic response, but initial internalisation driven by different exposure routes and toxicokinetic mechanisms

Mixed messages from benthic microbial communities exposed to nanoparticulate and ionic silver: 3D structure picks up nano-specific effects, while EPS and traditional endpoints indicate a concentration-dependent impact of silver ions

Silver nanoparticles (AgNP) are currently defined as emerging pollutants in surface water ecosystems. Whether the toxic effects of AgNP towards freshwater organisms are fully explainable by the release of ionic silver (Ag+) has not been conclusively elucidated. Long-term effects to benthic microbial communities (periphyton) that provide essential functions in stream ecosystems are unknown. The effects of exposure of periphyton to 2 and 20 μg/L Ag+ (AgNO3) and AgNP (polyvinylpyrrolidone stabilised) were investigated in artificial indoor streams. The extracellular polymeric substances (EPS) and 3D biofilm structure, biomass, algae species, Ag concentrations in the water phase and bioassociated Ag were analysed. A strong decrease in total Ag was observed within 4 days. Bioassociated Ag was proportional to dissolved Ag indicating a rate limitation by diffusion across the diffusive boundary layer. Two micrograms per liter of AgNO3 or AgNP did not induce significant effects despite detectable bioassociation of Ag. The 20-μg/L AgNO3 affected green algae and diatom communities, biomass and the ratio of polysaccharides to proteins in EPS. The 20-μg/L AgNO3 and AgNP decreased biofilm volume to about 50 %, while the decrease of biomass was lower in 20 μg/L AgNP samples than the 20-μg/L AgNO3 indicating a compaction of the NP-exposed biofilms. Roughness coefficients were lower in 20 μg/L AgNP-treated samples. The more traditional endpoints (biomass and diversity) indicated silver ion concentration-dependent effects, while the newly introduced parameters (3D structure and EPS) indicated both silver ion concentration-dependent effects and effects related to the silver species applied

Aging reduces the toxicity of pristine but not sulphidised silver nanoparticles to soil bacteria

In the environment engineered nanoparticles (ENPs) are subject to chemical and physical transformation processes. Thus, to understand their impact, it is important to consider how bioavailability and toxicity are influenced by these “aging” transformations with relation to environmental conditions and ENP properties. Here, two soil bacteria were exposed to Ag ENPs in ISO media (± fulvic acid) and soil pore water extracts with pH 6 and pH 8. The ENPs tested were 49 nm unfunctionalised, citrate stabilised (Ag-citr), 58 nm PVP-coated (Ag-PVP) and 36 nm sulphidised (Ag2S-PVP); AgNO3 was used as a positive control. Exposures were carried out using pristine (unaged) and 24 h aged ENPs, and the 24 h soluble fraction. Overall, toxicity was ranked AgNO3 > Ag-PVP ≥ Ag-citr ≫ Ag2S. Aging of AgNO3, Ag-PVP and Ag-citr in the ISO medium caused little change from unaged exposures and growth inhibition was mainly caused by soluble silver. Added fulvic acid decreased silver toxicity after aging and reduced the contribution of dissolution; as was the case in the soil pore waters where toxicity could not be attributed to ionic silver. Ag2S toxicity to A. globiformis in both ISO variants increased after aging, yet followed the same patterns as the metallic ENPs in the pore waters. For all ENPs pH effects were species dependent. Together this data showed that aging reduced toxicity in media with organic matter and despite soluble silver being the main driver of pristine ENP toxicity in the standard ISO medium, dissolution did not fully explain toxicity in the presence of organic matter

Evaluating environmental risk assessment models for nanomaterials according to requirements along the product innovation Stage-Gate process

Nanomaterial risk governance requires models to estimate the material flow, fate and transport as well as uptake/bioavailability, hazard and risk in the environment. This study assesses the fit of such available models to different stages during the innovation of nano-enabled products. Through stakeholder consultations, criteria were identified for each innovation stage from idea conception to market launch and monitoring. In total, 38 models were scored against 41 criteria concerning model features, applicability, resource demands and outcome parameters. A scoring scheme was developed to determine how the models fit the criteria of each innovation stage. For each model, the individual criteria scores were added, yielding an overall fit score to each innovation stage. Three criteria were critical to stakeholders and incorporated as multipliers in the scoring scheme; the required time/costs and level of expertise needed to use the model, and for risk assessment models only, the option to compare PEC and PNEC. Regulatory compliance was also identified as critical, but could not be incorporated, as a nanomaterial risk assessment framework has yet to be developed and adopted by legislators. In conclusion, the scoring approach underlined similar scoring profiles across stages within model categories. As most models are research tools designed for use by experts, their score generally increased for later stages where most resources and expertise are committed. In contrast, stakeholders need relatively simple models to identify potential hazards and risk management measures at early product development stages to ensure safe use of nanomaterials without costs and resource needs hindering innovation

Refinement of the selection of physicochemical properties for grouping and read-across of nanoforms

Before placing a new nanoform (NF) on the market, its potential adverse effects must be evaluated. This may e.g. be done via hazard and risk assessment. Grouping and read-across of NFs is a possible strategy to reduce resource consumption, maximising the use of existing data for assessment of NFs. The GRACIOUS project provides a framework in which possible grouping and read-across for NFs is mainly based on an evaluation of their similarity. The impact of NFs on human health and the environment depends strongly on the concentration of the NF and its physicochemical properties, such as chemical composition, size distribution, shape, etc. Hence, knowledge of the most relevant physicochemical properties is essential information for comparing similarity. The presented work aims to refine existing proposals for sets of descriptors (descriptor array) that are needed to describe distinct NFs of a material to identify the most relevant ones for grouping and read-across. The selection criteria for refining this descriptor array are explained and demonstrated. Relevant protocols and methods are proposed for each physicochemical property. The required and achievable measurement accuracies of the refined descriptor array are reviewed, as this information is necessary for similarity assessment of NFs based on individual physicochemical properties

Soil pH effects on the interactions between dissolved zinc, non-nano- and nano-ZnO with soil bacterial communities

Zinc oxide nanoparticles (ZnO NPs) are used in an array of products and processes, ranging from personal care products to antifouling paints, textiles, food additives, antibacterial agents and environmental remediation processes. Soils are an environment likely to be exposed to manmade nanoparticles due to the practice of applying sewage sludge as a fertiliser or as an organic soil improver. However, understanding on the interactions between soil properties, nanoparticles and the organisms that live within soil is lacking, especially with regards to soil bacterial communities. We studied the effects of nanoparticulate, non-nanoparticulate and ionic zinc (in the form of zinc chloride) on the composition of bacterial communities in soil with a modified pH range (from pH 4.5 to pH 7.2). We observed strong pH dependent effects on the interaction between bacterial communities and all forms of zinc, with the largest changes in bacterial community composition occurring in soils with low and medium pH levels (pH 4.8 and 5.9). The high pH soil (pH 7.2) was less susceptible to the effects of zinc exposure. At the highest doses of zinc (2500 mg/kg dw soil) both nano and non-nano particulate zinc applications elicited a similar response in the soil bacterial community, and this differed significantly to the ionic zinc salt treatment. The results highlight the importance of considering soil pH in nanotoxicology studies, although further work is needed to determine the exact mechanisms controlling the toxicity and fate and interactions of nanoparticles with soil microbial communities

Effects of silver nanoparticles on freshwater microbial communities

Nanoparticles (NPs) display special chemical properties because of their size, shape, composition and electronic structure. These properties lend NPs their functionality, but may also lead to toxic effects. Due to their widespread use in consumer products an exposure of the aquatic environment to NPs is anticipated and already proven in first analytical surveys. Especially metal/metaloxide NPs are widely used, of which silver nanoparticles (AgNPs) have gained considerable attention due to their broad microbiocidal properties. This implies a specific hazard for exposed aquatic primary producers (algae) and bacteria. In a case study with AgNPs, the toxicity to natural freshwater microbial communities was determined. The microbial communities were exposed to AgNPs of different sizes (10, 20, 40 and 50 nm) as well as different coatings (non-coated, citrate coated) in concentrations from 0.1 - 5000 nmol/L expcept for one silver nanoparticle type (50 nm, uncoated) which was tested in a range from 100 - 10 000 nmol/L. The selected silver nanoparticles were all purchased from commercial producers and distributers (Amepox, British Biocell, NanoTrade, Tedpella). To distinguish between particle related effects and effects caused by free silver ions silver nitrate was tested as a reference in corresponding total silver concentrations. The effects on the algal parts of the community was studied through pigment profile analysis with HPLC. The bacterial community composition was analysed by metabolic profiling on so called ecologplates[TRADEMARK] containing different carbon sources with a redox dye responding to respiration with colour change. The metabolic activity i.e. the colour change was detected as absorbance and was measured over time in several intervals for a total time period of 96 hours. Inhibition of growth detected as decrease in total biomass was seen in both bacterial and algal communities. All experiments were backed up by analytical measurements, measuring the total silver content, dissolved Ag+ (ultrafiltration) and particle size distribution (Nanosight)

Effects of selected silver nanoparticles on freshwater microbial communities

Due to their widespread use in consumer products an exposure of the aquatic environment to nanoparticles (NPs) is most likely. Especially metal/metaloxide NPs are widely used, of which silver nanoparticles (AgNPs) have gained considerable attention due to their broad microbiocidal properties implying a specific hazard for exposed aquatic algae and bacteria.In a case study with selected AgNPs (different sizes (10, 20 and 40 nm), and different coatings (citrate coated, non-coated)), the toxicity to natural freshwater microbial communities was determined. To distinguish between particle related effects and effects caused by free silver ions silver nitrate (AgNO3) was tested as a reference. The effects on the algal part of the community was studied through pigment profiling with HPLC. The bacterial community composition was analysed by metabolic profiling (ecologplates™). Inhibition of growth detected as a decrease in total biomass was seen in both bacterial and algal communities, for some cases in the range of environmentally realistic concentrations. Differences in toxicity could be determined for the different particles with AgNO3 being for almost all cases the most toxic compound with one excpetion. All experiments were backed up by analytical measurements (total silver content, dissolved silver (ultrafiltration), particle size distribution (Nanosight))