Synergistic antibacterial effect of copper and silver nanoparticles and their mechanism of action.

Bacterial infections are one of the leading causes of death worldwide. In the case of topical bacterial infections such as wound infections, silver (Ag) has historically been one of the most widely used antibacterials. However, scientific publications have demonstrated the adverse effects of silver on human cells, ecotoxicity and insufficient antibacterial effect for the complete elimination of bacterial infections. The use of Ag in the form of nanoparticles (NPs, 1-100 nm) allows to control the release of antibacterial Ag ions but is still not sufficient to eliminate infection and avoid cytotoxicity. In this study, we tested the potency of differently functionalized copper oxide (CuO) NPs to enhance the antibacterial properties of Ag NPs. The antibacterial effect of the mixture of CuO NPs (CuO, CuO-NH2 and CuO-COOH NPs) with Ag NPs (uncoated and coated) was studied. CuO and Ag NP combinations were more efficient than Cu or Ag (NPs) alone against a wide range of bacteria, including antibiotic-resistant strains such as gram-negative Escherichia coli and Pseudomonas aeruginosa as well as gram-positive Staphylococcus aureus, Enterococcus faecalis and Streptococcus dysgalactiae. We showed that positively charged CuO NPs enhanced the antibacterial effect of Ag NPs up to 6 times. Notably, compared to the synergy of CuO and Ag NPs, the synergy of respective metal ions was low, suggesting that NP surface is required for the enhanced antibacterial effect. We also studied the mechanisms of synergy and showed that the production of Cu+ ions, faster dissolution of Ag+ from Ag NPs and lower binding of Ag+ by proteins of the incubation media in the presence of Cu2+ were the main mechanisms of the synergy. In summary, CuO and Ag NP combinations allowed increasing the antibacterial effect up to 6 times. Thus, using CuO and Ag NP combinations enables to retain excellent antibacterial effects due to Ag and synergy and enhances beneficial effects, since Cu is a vital microelement for human cells. Thus, we suggest using combinations of Ag and CuO NPs in antibacterial materials, such as wound care products, to increase the antibacterial effect of Ag, improve safety and prevent and cure topical bacterial infections.

Comparison of Toxicity and Cellular Uptake of CdSe/ZnS and Carbon Quantum Dots for Molecular Tracking Using Saccharomyces cerevisiae as a Fungal Model.

Plant resource sharing mediated by mycorrhizal fungi has been a subject of recent debate, largely owing to the limitations of previously used isotopic tracking methods. Although CdSe/ZnS quantum dots (QDs) have been successfully used for in situ tracking of essential nutrients in plant-fungal systems, the Cd-containing QDs, due to the intrinsic toxic nature of Cd, are not a viable system for larger-scale in situ studies. We synthesized amino acid-based carbon quantum dots (CQDs; average hydrodynamic size 6 ± 3 nm, zeta potential -19 ± 12 mV) and compared their toxicity and uptake with commercial CdSe/ZnS QDs that we conjugated with the amino acid cysteine (Cys) (average hydrodynamic size 308 ± 150 nm, zeta potential -65 ± 4 mV) using yeast Saccharomyces cerevisiae as a proxy for mycorrhizal fungi. We showed that the CQDs readily entered yeast cells and were non-toxic up to 100 mg/L. While the Cys-conjugated CdSe/ZnS QDs were also not toxic to yeast cells up to 100 mg/L, they were not taken up into the cells but remained on the cell surfaces. These findings suggest that CQDs may be a suitable tool for molecular tracking in fungi (incl. mychorrhizal fungi) due to their ability to enter fungal cells.

Nanomaterials and Microorganisms: From Green Synthesis to Antibacterial Applications in Medicine and Agriculture.

Nanomaterial-based solutions for microorganism-related issues are gaining interest in medical fields, consumer applications, and agriculture [...].

Aminocatalysts are More Environmentally Friendly than Hydrogen-Bonding Catalysts.

The importance of asymmetric organocatalysis in contemporary organic synthesis is widely acknowledged. However, there are practically no data on the environmental safety of organocatalysts, although this aspect is crucial for the sustainability of all new materials, chemicals, and technologies. To start to fill this data-gap, a library of 26 organocatalysts containing hydrogen-bonding catalysts [(thio)ureas and squaramides] and aminocatalysts (primary or secondary amines) was evaluated for their toxicity using the naturally luminescent Vibrio fischeri bacteria (ISO assay; one of the most widely used ecotoxicity tests). Thioureas and squaramides were shown to be relatively toxic: none of them was ranked as "not harmful" (i. e., half maximal effective concentration EC50 >100 mg L-1 ), whereas the presence of the trifluoromethyl moiety increased their toxic effect. Importantly, the aminocatalysts, whose EC50 values ranged from 25 to >300 mg L-1 , could be considered remarkably more environmentally safe or green alternatives.

Virgin and UV-weathered polyamide microplastics posed no effect on the survival and reproduction of Daphnia magna.

Although evidence suggests that microplastic (MP) particles pose a risk to organisms, the effects of virgin and weathered MP should be evaluated separately as their effects may be different. In this work, we provide new information on the toxic potential of virgin and UV-weathered polyamide, one of the commonly used plastics worldwide. Polyamide MP particles were subjected to UV-weathering in wet conditions over 26 days in a customized irradiation chamber equipped with UV-C light tubes (15 W each, max. wavelength 254 nm). The toxicity of virgin and UV-weathered polyamide MP (< 180 µm in one dimension, 100 and 300 mg L-1) was evaluated by studying Daphnia magna reproduction in natural lake water spiked with MP, following the 21-day OECD 211 test guideline. In parallel, a nonionic surfactant Tween 20 (7 mg L-1) was added to the test medium to improve the suspendability of the MP. The results of the tests showed no adverse effects of either virgin or UV-weathered polyamide MP on the reproduction of D. magna. In addition, presence of Tween 20 in the test medium had no effects on the test results. These results bring a new perspective on the potential long-term impact of polyamide particles on aquatic organisms, especially considering that the polyamide has received marginal attention in the ecotoxicological research. However, standard test endpoints (survival and reproduction) may still miss long-term adverse effects of insoluble e.g., plastic particles and additional studies may be necessary.

Visible-Light Active Flexible and Durable Photocatalytic Antibacterial Ethylene-co-vinyl Acetate-Ag/AgCl/α-Fe2O3 Composite Coating.

When particles are mixed in polymer, particle surfaces become passivated by polymer matrix, leading to significantly reduced photocatalytic and, thus, also reduced antibacterial activity, as the catalytic particles become isolated from the outer environment and microorganisms reaching the surface. Herein, we demonstrate a facile and rapid approach for coating preparation at room temperature, yielding good adhesion of particles in combination with the particles' interface location. Flexible ethylene-co-vinyl acetate Ag/AgCl/α-Fe2O3 composite coatings were prepared by the spin-coating method. The synthesized photocatalytically active coating surface exhibited a distinct and rapid inhibition of bacterial growth, with at least a 7-log reduction of gram-positive bacteria Staphylococcus aureus viability after 30 min of visible-light illumination. We also analyzed the shedding of the Ag-ions and reactive oxygen species production from the composite coating and showed that reactive oxygen species played the main role in the photocatalytic bacterial inactivation, destroying the bacteria cell as proven by the Confocal Laser Scanning Microscopy.

Polyamide microplastic exposure elicits rapid, strong and genome-wide evolutionary response in the freshwater non-biting midge Chironomus riparius.

Susceptibility to hazardous materials and contamination is largely determined by genetic make-up and evolutionary history of affected organisms. Yet evolutionary adaptation and microevolutionary processes triggered by contaminants are rarely considered in ecotoxicology. Using an evolve and resequencing approach, we investigated genome-wide responses of the midge C. riparius exposed to virgin polyamide microplastics (0-180 µm size range, at concentration 1 g kg-1) during seven consecutive generations. The results were integrated to a parallel life-cycle experiment ran under the same exposure conditions. Emergence, life-cycle trait, showed first a substantial reduction in larval survival, followed by a rapid recovery within three generations. On the genomic level, we observed substantial selectively driven allele frequency changes (mean 0.566 ± 0.0879) within seven generations, associated with a mean selection coefficient of 0.322, indicating very strong selection pressure. Putative selection targets were mainly connected to oxidative stress in the microplastics exposed C. riparius population. This is the first multigenerational study on chironomids to provide evidence that upon exposure to polyamide microplastic there are changes on the genomic level, providing basis to rapid adaptation of aquatic organisms to microplastics.

Cubic Iron Core-Shell Nanoparticles Functionalized to Obtain High-Performance MRI Contrast Agents.

Nanoparticles with SiO2 coating were synthesized to have a cubic iron core. These were found to have saturation magnetization very close to the highest possible value of any iron-containing nanoparticles and the bulk iron saturation magnetization. The in vitro toxicology studies show that they are highly biocompatible and possess better MRI contrast agent potential than iron oxide NPs.

Long-Term Toxicity of Gadolinium to the Freshwater Crustacean Daphnia magna.

The lanthanides are considered emerging contaminants but information on their long-term toxicity to aquatic species under environmentally relevant conditions is scarce. We aimed to fill this gap by evaluating the long-term adverse effects of gadolinium on the freshwater model-crustacean Daphnia magna. The exposure of D. magna for up to 39 days to 0.1 mg Gd/L (a 21-days chronic toxicity NOEC value derived by us formerly) in the lake water had no negative effect (p > 0.05) on vitality, size and reproduction of parent animals as well as their offspring. Thus, assumingly the current Gd contamination levels of surface waters pose no hazard to aquatic crustaceans that in general are very sensitive to various pollutants. Moreover, presence of 0.1 mg Gd/L in the lake water even mitigated the long-term toxic effect of 0.2 mg Ni/L (studied as a model co-contaminant) to D. magna's vitality and productivity.

Evaluation of the potential toxicity of UV-weathered virgin polyamide microplastics to non-biting midge Chironomus riparius.

The relevance of the environmental hazard evaluation of virgin plastics particles is problematic, as plastics almost never occur in a virgin state after being discarded into the environment. However, the producers or importers must evaluate the environmental effect of their products as they are produced. Many plastic types e.g., polyamide, polyethylene are already under pre-registration, according to the database of the European Chemicals Agency (ECHA), in order to restrict the placing on the market of polymers (as defined by Article 3(5) of EU's REACH regulation (Registration, Evaluation, Authorization & Restriction of Chemicals), as a substance or in a mixture (ECHA, 2019). However, the hazard of microplastics could not be evaluated without relevant data on its (eco)toxic effects. In this work, the long-term toxicity of virgin polyamide microplastic (PA-MP) (size from 0 to 180 µm) and UV-weathered virgin PA-MP was investigated in the controlled life cycle experiments conducted in accordance with the OECD guidelines for testing of chemicals using Chironomus riparius (OECD test 218). In addition, a three-generation test was conducted to understand the trans-generational toxicity potential of virgin PA-MP. After UV irradiation (26 d) the buoyancy and color of the particles was changed and the share of smaller particles (of a few micrometer size range) increased. The exposure of C. riparius larvae to UV-weathered PA-MP (1000 mg kg-1) during their life cycle (28 d), negatively affected their development and subsequent emergence as adults. However, the exposure to virgin PA-MP throughout the life cycle and also over three consecutive generations did not significantly reduced the number of emerged adults. From the point of view of environmental hazard, the virgin polyamide plastics have probably no long-term hazard to chironomids. While it may not be relevant as environmental pollutant in the strict sense, UV-weathering may turn it hazardous.

Antibacterial Activity of Positively and Negatively Charged Hematite (α-Fe2O3) Nanoparticles to Escherichia coli, Staphylococcus aureus and Vibrio fischeri.

In the current study, the antibacterial activity of positively and negatively charged spherical hematite (α-Fe2O3) nanoparticles (NPs) with primary size of 45 and 70 nm was evaluated against clinically relevant bacteria Escherichia coli (gram-negative) and Staphylococcus aureus (gram-positive) as well as against naturally bioluminescent bacteria Vibrio fischeri (an ecotoxicological model organism). α-Fe2O3 NPs were synthesized using a simple green hydrothermal method and the surface charge was altered via citrate coating. To minimize the interference of testing environment with NP's physic-chemical properties, E. coli and S. aureus were exposed to NPs in deionized water for 30 min and 24 h, covering concentrations from 1 to 1000 mg/L. The growth inhibition was evaluated following the postexposure colony-forming ability of bacteria on toxicant-free agar plates. The positively charged α-Fe2O3 at concentrations from 100 mg/L upwards showed inhibitory activity towards E. coli already after 30 min of contact. Extending the exposure to 24 h caused total inhibition of growth at 100 mg/L. Bactericidal activity of positively charged hematite NPs against S. aureus was not observed up to 1000 mg/L. Differently from positively charged hematite NPs, negatively charged citrate-coated α-Fe2O3 NPs did not exhibit any antibacterial activity against E. coli and S. aureus even at 1000 mg/L. Confocal laser scanning microscopy and flow cytometer analysis showed that bacteria were more tightly associated with positively charged α-Fe2O3 NPs than with negatively charged citrate-coated α-Fe2O3 NPs. Moreover, the observed associations were more evident in the case of E. coli than S. aureus, being coherent with the toxicity results. Vibrio fischeri bioluminescence inhibition assays (exposure medium 2% NaCl) and colony forming ability on agar plates showed no (eco)toxicity of α-Fe2O3 (EC50 and MBC > 1000 mg/L).

Techniques Used for Analyzing Microplastics, Antimicrobial Resistance and Microbial Community Composition: A Mini-Review.

Antimicrobial resistance (AMR) is a global health threat. Antibiotics, heavy metals, and microplastics are environmental pollutants that together potentially have a positive synergetic effect on the development, persistence, transport, and ecology of antibiotic resistant bacteria in the environment. To evaluate this, a wide array of experimental methods would be needed to quantify the occurrence of antibiotics, heavy metals, and microplastics as well as associated microbial communities in the natural environment. In this mini-review, we outline the current technologies used to characterize microplastics based ecosystems termed "plastisphere" and their AMR promoting elements (antibiotics, heavy metals, and microbial inhabitants) and highlight emerging technologies that could be useful for systems-level investigations of AMR in the plastisphere.

Advances in Nanotoxicology: Towards Enhanced Environmental and Physiological Relevance and Molecular Mechanisms.

Nanotoxicology, a discipline transpired by the need to assess the human and environmental safety of nanoscale materials, has evolved over the past 15 years into a mature area of toxicology [...].

Sample preparation considerations for surface and crystalline properties and ecotoxicity of bare and silica-coated magnetite nanoparticles.

Magnetite (Fe3O4) nanoparticles (NPs) have widely used in various fields, including in medicine, due to their (super)paramagnetic properties. This requires a thorough evaluation of their possible hazardous effects. However, there is no standard procedure for the preparation of oxidation-prone NPs (such as magnetite) before subjecting them to biological assays. In this study we used Fe3O4 NPs (bare and silica-coated) as test samples to compare different preparation methods (ultrasound, centrifugation and filteration of NPs suspensions) based on X-ray and dynamic light scattering analysis and evaluation of microstructure and surface charge. After oxidation and functionalization, all samples retained their superparamagnetic behaviour. The toxicity of NP suspensions obtained by the methods described for Paramecium caudatum ciliates and Sinapis alba plants was evaluated.

Correction: Sample preparation considerations for surface and crystalline properties and ecotoxicity of bare and silica-coated magnetite nanoparticles.

[This corrects the article DOI: 10.1039/D1RA05703K.].

Nanotoxicology and Nanomedicine: The Yin and Yang of Nano-Bio Interactions for the New Decade.

Nanotoxicology and nanomedicine are two sub-disciplines of nanotechnology focusing on the phenomena, mechanisms, and engineering at the nano-bio interface. For the better part of the past three decades, these two disciplines have been largely developing independently of each other. Yet recent breakthroughs in microbiome research and the current COVID-19 pandemic demonstrate that holistic approaches are crucial for solving grand challenges in global health. Here we show the Yin and Yang relationship between the two fields by highlighting their shared goals of making safer nanomaterials, improved cellular and organism models, as well as advanced methodologies. We focus on the transferable knowledge between the two fields as nanotoxicological research is moving from pristine to functional nanomaterials, while inorganic nanomaterials - the main subjects of nanotoxicology - have become an emerging source for the development of nanomedicines. We call for a close partnership between the two fields in the new decade, to harness the full potential of nanotechnology for benefiting human health and environmental safety.

Effects of Humic Acids on the Ecotoxicity of Fe3O4 Nanoparticles and Fe-Ions: Impact of Oxidation and Aging.

The magnetite nanoparticles (MNPs) are increasingly produced and studied for various environmental applications, yet the information on their ecotoxicity is scarce. We evaluated the ecotoxicity of MNPs (~7 nm) before and after the addition of humic acids (HAs). White mustard Sinapis alba and unicellular ciliates Paramecium caudatum were used as test species. The MNPs were modified by HAs and oxidized/aged under mild and harsh conditions. Bare MNPs proved not toxic to plants (96 h EC50 > 3300 mg/L) but the addition of HAs and mild oxidation increased their inhibitory effect, especially after harsh oxidation (96 h EC50 = 330 mg/L). Nevertheless, all these formulations could be ranked as 'not harmful' to S. alba (i.e., 96 h EC50 > 100 mg/L). The same tendency was observed for ciliates, but the respective EC50 values ranged from 'harmful' (24 h EC50 = 10-100 mg/L) to 'very toxic' (24h EC50 < 1 mg/L). The ecotoxicity of Fe-ions with and without the addition of HAs was evaluated in parallel: Fe (II) and Fe (III) ions were toxic to S. alba (96 h EC50 = 35 and 60 mg/L, respectively) and even more toxic to ciliates (24 h EC50 = 1 and 3 mg/L, respectively). Addition of the HAs to Fe-ions yielded the respective complexes not harmful to plants (96h EC50 > 100 mg/L) but toxic to ciliates (24h EC50 = 10-100 mg/L). These findings will be helpful for the understanding of the environmental fate and toxicity of iron-based NPs.

Impact of surface functionalization on the toxicity and antimicrobial effects of selenium nanoparticles considering different routes of entry.

Selenium nanoparticles (SeNPs) were first designed as nutritional supplements, but they are attractive also for use in diagnostic and therapeutic systems owing to their biocompatibility and protective effects. This study aimed to examine if different SeNPs stabilization strategies affect their (i) antimicrobial activity against bacteria Escherichia coli and Staphylococcus aureus and yeast Saccharomyces cerevisiae and (ii) toxicity to human cells of different biological barriers i.e., skin, oral and intestinal mucosa. For surface stabilization, polyvinylpyrrolidone (PVP), poly-L-lysine (PLL) and polyacrylic acid (PAA) were used rendering neutral, positively and negatively charged SeNPs, respectively. The SeNPs (primary size ~80 nm) showed toxic effects in human cells in vitro and in bacteria S. aureus, but not in E. coli and yeast S. cerevisiae. Toxicity of SeNPs (24 h IC50) ranged from 1.4 to >100 mg Se/L, depending on surface functionalization (PLL > PAA > PVP) and was not caused by ionic Se. At subtoxic concentrations, all SeNPs were taken up by all human cell types, induced oxidative stress response and demonstrated genotoxicity. As the safety profile of SeNPs was dependent not only on target cells (mammalian cells, bacteria, yeast), but also on surface functionalization, these aspects should be considered during development of novel SeNPs-based biomedical products.

Ingestion and effects of virgin polyamide microplastics on Chironomus riparius adult larvae and adult zebrafish Danio rerio.

Secondary microplastics (MP) produced by fragmentation of plastic in the environment or as a result of human activities can easily be taken up by organisms. The harmful effects of MP depend on e.g., the type, dimensions, sorption capacity and concentration of MP. In this study the ingestion of virgin irregularly-shaped polyamide microplastics (PA-MP; up to180 µm) by two different species was evaluated: 3rd - 4th instar larvae of midge Chironomus riparius and adult fish Danio rerio. More specifically, in the case of C. riparius larvae their feeding strategy, i.e. the ability to differentiate between food and non-food material (PA-MP) and the impact of pseudo-satiation by PA-MP on larval growth, development and emergence was evaluated. Two feeding regimes (with and without food supply) and two PA-MP concentrations (100 mg kg-1 and 1000 mg kg-1) were applied. Fish were exposed to two PA-MP concentrations (30 and 330 mg L-1) for 48 h followed by 48 h of depuration. The fish were fed during both periods. Both, chironomid larvae and adult zebrafish actively ingested PA-MP. Remarkably more PA-MP was ingested when larvae were not fed during the exposure to PA-MP. In the case of fish, the ingested particles were effectively evacuated from the gastrointestinal tract. Even the highest PA-MP concentrations did not cause obvious harmful effects to either species. The obtained data are informative for risk evaluation of PA-MP as polyamide is registered in the database of the European Chemicals Agency (ECHA) in the framework of the EU's REACH (Registration, Evaluation, Authorization & Restriction of Chemicals) regulation.

Surface carboxylation or PEGylation decreases CuO nanoparticles' cytotoxicity to human cells in vitro without compromising their antibacterial properties.

Clinical use of CuO nanoparticles (NPs) as antibacterials can be hampered by their toxicity to human cells. We hypothesized that certain surface functionalizations of CuO NPs may render NPs toxic to bacteria, but still be relatively harmless to human cells. To control this hypothesis, the toxicity of differently functionalized CuO NPs to bacteria Escherichia coli vs human cells (THP-1 macrophages and HACAT keratinocytes) was compared using similar conditions and end points. CuO NPs functionalized with polyethylene glycol (CuO-PEG), carboxyl (CuO-COOH, anionic), ammonium (CuO-NH4+, cationic) and unfunctionalized CuO NPs and CuSO4 (controls) were tested. In general, the toxicity of Cu compounds decreased in the following order: CuO-NH4+ > unfunctionalized CuO > CuSO4 > CuO-COOH > CuO-PEG. Positively charged unfunctionalized CuO and especially CuO-NH4+ proved most toxic (24-h EC50 = 21.7-47 mg/l) and had comparable toxicity to bacterial and mammalian cells. The multivariate analysis revealed that toxicity of these NPs was mostly attributed to their positive zeta potential, small hydrodynamic size, high Cu dissolution, and induction of reactive oxygen species (ROS) and TNF-α. In contrast, CuO-COOH and CuO-PEG NPs had lower toxicity to human cells compared to bacteria despite efficient uptake of these NPs by human cells. In addition, these NPs did not induce TNF-α and ROS. Thus, by varying the NP functionalization and Cu form (soluble salt vs NPs), it was possible to "target" the toxicity of Cu compounds, whereas carboxylation and PEGylation rendered CuO NPs that were more toxic to bacteria than to human cells envisaging their use in medical antibacterial products.