Molecular Mechanisms in Metal Oxide Nanoparticle-Tryptophan Interactions.

One of the crucial metabolic processes for both plant and animal kingdoms is the oxidation of the amino acid tryptophan (TRP) that regulates plant growth and controls hunger and sleeping patterns in animals. Here, we report revolutionary insights into how this process can be crucially affected by interactions with metal oxide nanoparticles (NPs), creating a toolbox for a plethora of important biomedical and agricultural applications. Molecular mechanisms in TRP-NP interactions were revealed by NMR and optical spectroscopy for ceria and titania and by X-ray single-crystal study and a computational study of model TRP-polyoxometalate complexes, which permitted the visualization of the oxidation mechanism at an atomic level. Nanozyme activity, involving concerted proton and electron transfer to the NP surface for oxides with a high oxidative potential, like CeO2 or WO3, converted TRP in the first step into a tricyclic organic acid belonging to the family of natural plant hormones, auxins. TiO2, a much poorer oxidant, was strongly binding TRP without concurrent oxidation in the dark but oxidized it nonspecifically via the release of reactive oxygen species (ROS) in daylight.

Reusable magnetic mixture of CuFe2O4-Fe2O3 and TiO2 for photocatalytic degradation of pesticides in water.

Photocatalysis is a promising treatment method to remove pollutants from water. TiO2-P25 is a commercially available model photocatalyst, which very efficiently degrades organic pollutants under UVA light exposure. However, the collection and the recovery of TiO2-P25 from cleaned water poses significant difficulties, severely limiting its usability. To address this challenge, we have prepared a sintered mixture of TiO2-P25 nanomaterials and magnetic CuFe2O4-Fe2O3 nanocomposites. The mixture material was shown to contain spinel ferrite, hematite and maghemite structures, copper predominantly in Cu2+ and iron predominantly in Fe3+ state. The CuFe2O4-Fe2O3 and TiO2-P25 mixture demonstrated magnetic collectability from processed water and photocatalytic activity, which was evidenced through the successful photodegradation of the herbicide 2,4-D. Our findings suggest that the sintered mixture of CuFe2O4-Fe2O3 and TiO2-P25 holds a promise for improving photocatalytic water treatment, with the potential to overcome current photocatalyst recovery issues.

Antibacterial activity of solid surfaces is critically dependent on relative humidity, inoculum volume, and organic soiling.

Antimicrobial surface materials potentially prevent pathogen transfer from contaminated surfaces. Efficacy of such surfaces is assessed by standard methods using wet exposure conditions known to overestimate antimicrobial activity compared to dry exposure. Some dry test formats have been proposed but semi-dry exposure scenarios e.g. oral spray or water droplets exposed to ambient environment, are less studied. We aimed to determine the impact of environmental test conditions on antibacterial activity against the model species Escherichia coli and Staphylococcus aureus. Surfaces based on copper, silver, and quaternary ammonium with known or claimed antimicrobial properties were tested in conditions mimicking microdroplet spray or larger water droplets exposed to variable relative air humidity in the presence or absence of organic soiling. All the environmental parameters critically affected antibacterial activity of the tested surfaces from no effect in high-organic dry conditions to higher effect in low-organic humid conditions but not reaching the effect size demonstrated in the ISO 22169 wet format. Copper was the most efficient antibacterial surface followed by silver and quaternary ammonium based coating. Antimicrobial testing of surfaces using small droplet contamination in application-relevant conditions could therefore be considered as one of the worst-case exposure scenarios relevant to dry use surfaces.

Antiviral efficacy of nanomaterial-treated textiles in real-life like exposure conditions.

Due to the growing interest towards reducing the number of potentially infectious agents on critical high-touch surfaces, the popularity of antimicrobially and antivirally active surfaces, including textiles, has increased. The goal of this study was to create antiviral textiles by spray-depositing three different nanomaterials, two types of CeO2 nanoparticles and quaternary ammonium surfactant CTAB loaded SiO2 nanocontainers, onto the surface of a knitted polyester textile and assess their antiviral activity against two coronaviruses, porcine transmissible gastroenteritis virus (TGEV) and severe acute respiratory syndrome virus (SARS CoV-2). Antiviral testing was carried out in small droplets in semi-dry conditions and in the presence of organic soiling, to mimic aerosol deposition of viruses onto the textiles. In such conditions, SARS CoV-2 stayed infectious at least for 24 h and TGEV infected cells even after 72h of semi-dry deposition suggesting that textiles exhibiting sufficient antiviral activity before or at 24 h, can be considered promising. The antiviral efficacy of nanomaterial-deposited textiles was compared with the activity of the same nanomaterials in colloidal form and with positive control textiles loaded with copper nitrate and CTAB. Our results indicated that after deposition onto the textile, CeO2 nanoparticles lost most of their antiviral activity, but antiviral efficacy of CTAB-loaded SiO2 nanocontainers was retained also after deposition. Copper nitrate deposited textile that was used as a positive control, showed relatively high antiviral activity as expected. However, as copper was effectively washed away from the textile already during 1 h, the use of copper for creating antiviral textiles would be impractical. In summary, our results indicated that antiviral activity of textiles cannot be predicted from antiviral efficacy of the deposited compounds in colloid and attention should be paid on prolonged efficacy of antivirally coated textiles.

Antimicrobial Nano Coatings.

History has demonstrated that the uncontrolled fast thriving of potentially pathogenic microorganisms may lead to serious consequences and, thus, the approaches helping to control the microbial numbers in infectional hot-spots are necessary [...].

Antiviral efficacy of cerium oxide nanoparticles.

Nanomaterials are prospective candidates for the elimination of viruses due to their multimodal mechanisms of action. Here, we tested the antiviral potential of a largely unexplored nanoparticle of cerium dioxide (CeO2). Two nano-CeO2 with opposing surface charge, (+) and (-), were assessed for their capability to decrease the plaque forming units (PFU) of four enveloped and two non-enveloped viruses during 1-h exposure. Statistically significant antiviral activity towards enveloped coronavirus SARS-CoV-2 and influenza virus was registered already at 20 mg Ce/l. For other two enveloped viruses, transmissible gastroenteritis virus and bacteriophage φ6, antiviral activity was evidenced at 200 mg Ce/l. As expected, the sensitivity of non-enveloped viruses towards nano-CeO2 was significantly lower. EMCV picornavirus showed no decrease in PFU until the highest tested concentration, 2000 mg Ce/l and MS2 bacteriophage showed slight non-monotonic response to high concentrations of nano-CeO2(-). Parallel testing of antiviral activity of Ce3+ ions and SiO2 nanoparticles allows to conclude that nano-CeO2 activity was neither due to released Ce-ions nor nonspecific effects of nanoparticulates. Moreover, we evidenced higher antiviral efficacy of nano-CeO2 compared with Ag nanoparticles. This result along with low antibacterial activity and non-existent cytotoxicity of nano-CeO2 allow us to propose CeO2 nanoparticles for specific antiviral applications.

Effects of environmentally relevant concentrations of microplastics on amphipods.

Lack of microplastics (MP) toxicity studies involving environmentally relevant concentrations and exposure times is concerning. Here we analyzed the potential adverse effects of low density polyethylene (LDPE) MP at environmentally relevant concentration in sub-chronic exposure to two amphipods Gmelinoides fasciatus and Gammarus lacustris, species that naturally compete with each other for their habitats. 14-day exposure to 2 µg/L (8 particles/L corresponding to low exposure) and 2 mg/L (∼8400 particles/L, corresponding to high exposure) of 53-100 µm LDPE MP were used to assess ingestion and egestion of MP, evaluate its effects on amphipod mortality, swimming ability and oxidative stress level. Both amphipod species were effectively ingesting and egesting LDPE MP. On the average, 0.8 and 2.5 MP particles were identified in the intestines of each amphipod exposed to 2 µg/L and 2 mg/L LDPE MP, respectively. Therefore, intestinal MP after 14-day exposure did not fully reflect the differences in LDPE MP exposure concentrations. Increased mortality of both amphipods was observed at 2 mg/L LDPE MP and in case of G. lacustris also at 2 µg/L exposure. The effect of LDPE on swimming activity was observed only in case of G. fasciatus. Oxidative stress marker enzymes SOD, GPx and reduced glutathione GSH varied according to amphipod species and LDPE MP concentration. In general G. lacustris was more sensitive towards LDPE MP induced oxidative stress. Overall, the results suggested that in MP polluted environments, G. lacustris may lose its already naturally low competitiveness and become overcompeted by other more resistant species. The fact that in the sub-chronic foodborne exposure to environmentally relevant and higher LDPE MP concentrations all the observed toxicological endpoints were affected refers to the potential of MP to affect and disrupt aquatic communities in the longer perspective.

Hazard of polystyrene micro-and nanospheres to selected aquatic and terrestrial organisms.

Plastics contamination in the environment is a major concern. Risk assessment of micro- and nanoplastics (MPL and NPL) poses significant challenges due to MPL and NPL heterogeneity regarding compositional polymers, particle sizes and morphologies in the environment. Yet, there exists considerable toxicological literature on commercial polystyrene (PS) micro- and nanospheres. Although such particles do not directly represent the environmental MPL and NPL, their toxicity data should be used to advance the hazard assessment of plastics. Here, toxicity data of PS micro- and nanospheres for microorganisms, aquatic and terrestrial invertebrates, fish, and higher plants was collected and analyzed. The evaluation of 294 papers revealed that aquatic invertebrates were the most studied organisms, nanosized PS was studied more often than microsized PS, acute exposures prevailed over chronic exposures, the toxicity of PS suspension additives was rarely addressed, and ∼40 % of data indicated no organismal effects of PS. Toxicity mechanisms were mainly studied in fish and nematode Caenorhabditis elegans, providing guidance for relevant studies in higher organisms. Future studies should focus on environmentally relevant plastics concentrations, wide range of organisms, co-exposures with other pollutants, and method development for plastics identification and quantification to fill the gap of bioaccumulation assessment of plastics.

Preparation and Characterization of Photocatalytically Active Antibacterial Surfaces Covered with Acrylic Matrix Embedded Nano-ZnO and Nano-ZnO/Ag.

In the context of healthcare-acquired infections, microbial cross-contamination and the spread of antibiotic resistance, additional passive measures to prevent pathogen carryover are urgently needed. Antimicrobial high-touch surfaces that kill microbes on contact or prevent their adhesion could be considered to mitigate the spread. Here, we demonstrate that photocatalytic nano-ZnO- and nano-ZnO/Ag-based antibacterial surfaces with efficacy of at least a 2.7-log reduction in Escherichia coli and Staphylococcus aureus viability in 2 h can be produced by simple measures using a commercial acrylic topcoat for wood surfaces. We characterize the surfaces taking into account cyclic wear and variable environmental conditions. The light-induced antibacterial and photocatalytic activities of the surfaces are enhanced by short-term cyclic wear, indicating their potential for prolonged effectivity in long-term use. As the produced surfaces are generally more effective at higher relative air humidity and silver-containing surfaces lost their contact-killing properties in dry conditions, it is important to critically evaluate the end-use conditions of materials and surfaces to be tested and select application-appropriate methods for their efficacy assessment.

Cellular binding, uptake and biotransformation of silver nanoparticles in human T lymphocytes.

Our knowledge of uptake, toxicity and detoxification mechanisms as related to nanoparticles' (NPs') characteristics remains incomplete. Here we combine the analytical power of three advanced techniques to study the cellular binding and uptake and the intracellular transformation of silver nanoparticles (AgNPs): single-particle inductively coupled mass spectrometry, mass cytometry and synchrotron X-ray absorption spectrometry. Our results show that although intracellular and extracellularly bound AgNPs undergo major transformation depending on their primary size and surface coating, intracellular Ag in 24 h AgNP-exposed human lymphocytes exists in nanoparticulate form. Biotransformation of AgNPs is dominated by sulfidation, which can be viewed as one of the cellular detoxification pathways for Ag. These results also show that the toxicity of AgNPs is primarily driven by internalized Ag. In fact, when toxicity thresholds are expressed as the intracellular mass of Ag per cell, differences in toxicity between NPs of different coatings and sizes are minimized. The analytical approach developed here has broad applicability in different systems where the aim is to understand and quantify cell-NP interactions and biotransformation.

Correction to: Quantitative Measurement of Cell-Nanoparticle Interactions Using Mass Cytometry.

This chapter was inadvertently published with the acknowledgement section leaving out the following sentence: "This work received funding from South Australian Government PRIF program Project "International Cluster on Nanosafety" of Nicolas H. Voelcker and Enzo Lombi." This correction has been updated in the chapter.

Quantitative Measurement of Cell-Nanoparticle Interactions Using Mass Cytometry.

Mass cytometry is a technique that uses inductively coupled plasma mass spectrometry (ICP-MS) to quantify the isotopic composition of cells in suspension. Traditionally it has been used in conjunction with antibodies labeled with stable lanthanide isotopes to investigate cellular heterogeneity. Here we describe its use to quantify uptake of metal nanoparticles by cells in suspension.

Propidium iodide staining underestimates viability of adherent bacterial cells.

Combining membrane impermeable DNA-binding stain propidium iodide (PI) with membrane-permeable DNA-binding counterstains is a widely used approach for bacterial viability staining. In this paper we show that PI staining of adherent cells in biofilms may significantly underestimate bacterial viability due to the presence of extracellular nucleic acids (eNA). We demonstrate that gram-positive Staphylococcus epidermidis and gram-negative Escherichia coli 24-hour initial biofilms on glass consist of 76 and 96% PI-positive red cells in situ, respectively, even though 68% the cells of either species in these aggregates are metabolically active. Furthermore, 82% of E. coli and 89% S. epidermidis are cultivable after harvesting. Confocal laser scanning microscopy (CLSM) revealed that this false dead layer of red cells is due to a subpopulation of double-stained cells that have green interiors under red coating layer which hints at eNA being stained outside intact membranes. Therefore, viability staining results of adherent cells should always be validated by an alternative method for estimating viability, preferably by cultivation.

Potential ecotoxicological effects of antimicrobial surface coatings: a literature survey backed up by analysis of market reports.

This review was initiated by the COST action CA15114 AMICI "Anti-Microbial Coating Innovations to prevent infectious diseases," where one important aspect is to analyze ecotoxicological impacts of antimicrobial coatings (AMCs) to ensure their sustainable use. Scopus database was used to collect scientific literature on the types and uses of AMCs, while market reports were used to collect data on production volumes. Special attention was paid on data obtained for the release of the most prevalent ingredients of AMCs into the aqueous phase that was used as the proxy for their possible ecotoxicological effects. Based on the critical analysis of 2,720 papers, it can be concluded that silver-based AMCs are by far the most studied and used coatings followed by those based on titanium, copper, zinc, chitosan and quaternary ammonium compounds. The literature analysis pointed to biomedicine, followed by marine industry, construction industry (paints), food industry and textiles as the main fields of application of AMCs. The published data on ecotoxicological effects of AMCs was scarce, and also only a small number of the papers provided information on release of antimicrobial ingredients from AMCs. The available release data allowed to conclude that silver, copper and zinc are often released in substantial amounts (up to 100%) from the coatings to the aqueous environment. Chitosan and titanium were mostly not used as active released ingredients in AMCs, but rather as carriers for other release-based antimicrobial ingredients (e.g., conventional antibiotics). While minimizing the prevalence of healthcare-associated infections appeared to be the most prosperous field of AMCs application, the release of environmentally hazardous ingredients of AMCs into hospital wastewaters and thus, also the environmental risks associated with AMCs, comprise currently only a fraction of the release and risks of traditional disinfectants. However, being proactive, while the use of antimicrobial/antifouling coatings could currently pose ecotoxicological effects mainly in marine applications, the broad use of AMCs in other applications like medicine, food packaging and textiles should be postponed until reaching evidences on the (i) profound efficiency of these materials in controlling the spread of pathogenic microbes and (ii) safety of AMCs for the human and ecosystems.

Antimicrobial potency of differently coated 10 and 50 nm silver nanoparticles against clinically relevant bacteria Escherichia coli and Staphylococcus aureus.

Silver nanoparticles (nanoAg) are effective antimicrobials and promising alternatives to traditional antibiotics. This study aimed at evaluating potency of different nanoAg against healthcare infections associated bacteria: Gram-negative Escherichia coli and Gram-positive Staphylococcus aureus. A library of differently coated nanoAg of two different sizes (10 and 50 nm) were prepared using coating agents poly-L-Lysine (PLL), cetyltrimethyl-ammonium bromide (CTAB), citrate (CIT), polyvinyl-pyrrolidone (PVP), polysorbate 80 (Tween 80), and dioctyl-sodium sulfosuccinate (AOT). Stability evaluation by means of agglomeration and dissolution behaviour was performed for all nanoAg under conditions relevant for this study. Antibacterial properties of nanoAg were addressed by determining their minimal bactericidal concentrations (MBC) in deionised (DI) water to minimise the influence of silver speciation on its bioavailability. In parallel, AgNO3 was analysed as an ionic control. Studied nanoAg were efficient antimicrobials being remarkably more potent towards E. coli than to S. aureus (4 h MBC values for different nanoAg ranged from 0.08 to 5.0 mg Ag/L and 1.0-10 mg Ag/L, respectively). The toxicity of all nanoAg to S. aureus (but not to E. coli) increased with exposure time (4 h vs 24 h). 10 nm sized nanoAg released more Ag-ions and were more toxic than 50 nm nanoAg. Coating-dependent toxicity was more prominent for 50 nm nanoAg coated with Tween 80 or CTAB rendering the least toxic nanoAg. Obtained results showed that the antimicrobial effects of nanoAg were driven by shed Ag-ions, depended on target bacteria, exposure time and were the interplay of NP size, solubility and surface coating.

Rapid in situ assessment of Cu-ion mediated effects and antibacterial efficacy of copper surfaces.

Release of metal ions from metal-based surfaces has been considered one of the main drivers of their antimicrobial activity. Here we describe a method that enables parallel assessment of metal ion release from solid metallic surfaces and antimicrobial efficacy of these surfaces in a short time period. The protocol involves placement of a small volume of bioluminescent bacteria onto the tested surface and direct measurement of bioluminescence at various time points. In this study, two recombinant Escherichia coli strains, one expressing bioluminescence constitutively and applicable for general antimicrobial testing, and the other induced by Cu ions, were selected. Decrease in bioluminescence of constitutive E. coli on the surfaces showed a good correlation with the decrease in bacterial viability. Response of Cu-inducible E. coli showed a correlation with Cu content in the tested surfaces but not with Cu dissolution suggesting the role of direct bacteria-surface contact in Cu ion-driven antibacterial effects. In summary, the presented protocol enables the analysis of microbial toxicity and bioavailability of surface-released metal ions directly on solid surfaces within 30-60 min. Although optimized for copper and copper alloy surfaces and E. coli, the method can be extended to other types of metallic surfaces and bacterial strains.

UVA-induced antimicrobial activity of ZnO/Ag nanocomposite covered surfaces.

Application of efficient antimicrobial surfaces has been estimated to decrease both, the healthcare-associated infections and the spread of antibiotic-resistant bacteria. In this paper, we prepared ZnO and ZnO/Ag nanoparticle covered surfaces and evaluated their antimicrobial efficacy towards a Gram-negative bacterial model (Escherichia coli), a Gram-positive bacterial model (Staphylococcus aureus) and a fungal model (Candida albicans) in the dark and under UVA illumination. The surfaces were prepared by spin coating aliquots of ZnO and ZnO/Ag nanoparticle suspensions onto glass substrates. Surfaces contained 2 or 20 µg Zn/cm2 and 0-0.02 µg Ag/cm2. No significant antimicrobial activity of the surfaces, except of those with the highest Ag or Zn content was observed in the dark. On the other hand, UVA illuminated surfaces containing 20 µg Zn/cm2 and 2 µg Zn plus 0.02 µg Ag/cm2 caused >3 log decrease in the viable counts of E. coli and S. aureus in 30 min. As proven by brilliant blue FCF dye degradation and elemental analysis of the surfaces, this remarkable antimicrobial activity was a combined result of photocatalytic effect and release of Zn and Ag ions from surfaces. Surfaces retained significant antibacterial and photocatalytic properties after several usage cycles. Compared to bacteria, yeast C. albicans was significantly less sensitive to the prepared surfaces and only about 1 log reduction of viable count was observed after 60 min UVA illumination. In conclusion, the developed ZnO/Ag surfaces exhibit not only high antibacterial activity but also some antifungal activity.

Ligand-Doped Copper Oxo-hydroxide Nanoparticles are Effective Antimicrobials.

Bacterial resistance to antimicrobial therapies is an increasing clinical problem. This is as true for topical applications as it is for systemic therapy. Topically, copper ions may be effective and cheap antimicrobials that act through multiple pathways thereby limiting opportunities to bacteria for resistance. However, the chemistry of copper does not lend itself to facile formulations that will readily release copper ions at biologically compatible pHs. Here, we have developed nanoparticulate copper hydroxide adipate tartrate (CHAT) as a cheap, safe, and readily synthesised material that should enable antimicrobial copper ion release in an infected wound environment.First, we synthesised CHAT and showed that this had disperse aquated particle sizes of 2-5 nm and a mean zeta potential of - 40 mV. Next, when diluted into bacterial medium, CHAT demonstrated similar efficacy to copper chloride against Escherichia coli and Staphylococcus aureus, with dose-dependent activity occurring mostly around 12.5-50 mg/L of copper. Indeed, at these levels, CHAT very rapidly dissolved and, as confirmed by a bacterial copper biosensor, showed identical intracellular loading to copper ions derived from copper chloride. However, when formulated at 250 mg/L in a topically applied matrix, namely hydroxyethyl cellulose, the benefit of CHAT over copper chloride was apparent. The former yielded rapid sustained release of copper within the bactericidal range, but the copper chloride, which formed insoluble precipitates at such concentration and pH, achieved a maximum release of 10 ± 7 mg/L copper by 24 h.We provide a practical formulation for topical copper-based antimicrobial therapy. Further studies, especially in vivo, are merited.

Microfluidic Cell Microarray Platform for High Throughput Analysis of Particle-Cell Interactions.

With advances in nanotechnology, particles with various size, shape, surface chemistry, and composition can be easily produced. Nano- and microparticles have been extensively explored in many industrial and clinical applications. Ensuring that the particles themselves are not possessing toxic effects to the biological system is of paramount importance. This paper describes a proof of concept method, in which a microfluidic system is used in conjunction with a cell microarray technique aiming to streamline the analysis of particle-cell interaction in a high throughput manner. Polymeric microparticles, with different particle surface functionalities, were first used to investigate the efficiency of particle-cell adhesion under dynamic flow. Silver nanoparticles (AgNPs, 10 nm in diameter) perfused at different concentrations (0 to 20 µg/mL) in parallel streams over the cell microarray exhibited a higher toxicity compared to the static culture in the 96-well-plate format. This developed microfluidic system can be easily scaled up to accommodate a larger number of microchannels for high throughput analysis of the potential toxicity of a wide range of particles in a single experiment.

Uptake and transcytosis of functionalized superparamagnetic iron oxide nanoparticles in an in vitro blood brain barrier model.

Two major hurdles in nanomedicine are the limited strategies for synthesizing stealth nanoparticles and the poor efficacy of the nanoparticles in translocating across the blood brain barrier (BBB). Here we examined the uptake and transcytosis of iron oxide nanoparticles (IONPs) grafted with biomimetic phosphorylcholine (PC) brushes in an in vitro BBB model system, and compared them with bare, PEG or PC-PEG mixture grafted IONPs. Hyperspectral imaging indicated IONP co-localization with cells. Quantitative analysis with total reflection X-ray fluorescence spectrometry showed that after 24 h, 78% of PC grafted, 68-69% of PEG or PC-PEG grafted, and 30% of bare IONPs were taken up by the BBB. Transcytosis of IONPs was time-dependent and after 24 h, 16-17% of PC or PC-PEG mixture grafted IONPs had passed the BBB model, significantly more than PEG grafted or bare IONPs. These findings point out that grafting of IONPs with PC is a viable strategy for improving the uptake and transcytosis of nanoparticles.