Effects of Ageing in Disinfectant Solution on the Corrosion Resistance and Antimicrobial Behavior of Copper Alloys.

This work studies two copper-based alloys as potential antimicrobial weapons for sectors where surface hygiene is essential. Effects of different alloying elements addition at the same Cu content (92.5% by weight) on the corrosion resistance and the antibacterial performance of two copper alloys were studied in an aerated disinfectant solution (0.25% v/v Aniosurf Premium (D)) by electrochemical corrosion, X-ray photoelectron spectroscopy (XPS), time-of-flight secondary ion mass spectroscopy (ToF-SIMS) and antibacterial tests. Results showed that the nature of the alloying elements had a clear influence on the corrosion resistance and antibacterial performance. Electrochemical impedance results and surface analyses demonstrate the presence of organic compounds bound on the substrate and that a film covers part of the total active surface and may act as a protective barrier by preventing the interaction between metal and solution, decreasing the antimicrobial performance of copper-based materials. Low zinc and silicon contents in copper alloys allows for better aging behavior in D solution while maintaining good antibacterial performance. The XPS and ToF-SIMS results indicated that artificial aging in disinfectant enhanced Cu enrichment in the organic film formed, which could effectively stimulate the release of Cu ions from the surface.

Inactivation of Enveloped Bovine Viral Diarrhea Virus and Non-Enveloped Porcine Parvovirus Using Low-Pressure Non-Thermal Plasma.

As the worldwide population has been experiencing since 2020, viruses represent a serious threat to global well-being. To avoid viral transmission through surgery or medical examination, sterilization of medical material is needed. From emerging sterilization processes, the use of non-thermal plasma (NTP) arises as a promising technique to efficiently reduce microbial burden on medical devices, including new complex polymers as thermosensitive ones. Thus, we evaluated the antiviral efficacy of a low-pressure NTP process taking place in a sealed bag. For this purpose, two different plasmas, O2 100% plasma and Ar 80%-O2 20% plasma, were tested against two viruses: the bovine viral diarrhea virus and the porcine parvovirus, surrogates of human hepatitis C virus and human parvovirus B19, respectively. The efficacy of both NTP treatments on viral load can be detected after only five minutes. Moreover, the longer the NTP treatments last, the more the load decreases. The most effective load reduction was obtained with a 120-min O2 plasma treatment inducing a minimum of four-log viral load reduction. So, this process demonstrated strong virucidal capacity inside a sealed bag and represents a very interesting opportunity in the field of fragile medical devices sterilization or disinfection.

Contribution of Fluorescence Techniques in Determining the Efficiency of the Non-thermal Plasma Treatment.

We have recently developed a non-thermal plasma (NTP) equipment intended to sterilize fragile medical devices and maintain the sterile state of items downstream the treatment. With traditional counts on agar plate a six log reduction of Staphylococcus aureus viability was obtained within 120 min of O2, Ar, or N2 NTP treatments. However to determine the best NTP process, we studied the different physiological states of S. aureus by flow cytometry (FC) and confocal laser scanning microscopy (CLSM) focusing on the esterasic activity and membrane integrity of the bacteria. Two fluorochromes, 5-(and-6)-carboxy-2',7'-dichlorofluorescein diacetate and propidium iodide were used in order to distinguish three sub-populations: metabolically active, permeabilized, and damaged bacteria that can be in the viable but nonculturable state. FC and CLSM highlight that O2 and Ar NTP treatments were the most attractive processes. Indeed, a 5 min of Ar NTP generated a high destruction of the structure of bacteria and a 120 min of O2 NTP treatment led to the higher decrease of the total damaged bacteria population. SEM observations showed that in presence of clusters, bacteria of upper layers are easily altered compared to bacteria in the deeper layers. In conclusion, the plate counting method is not sufficient by itself to determine the best NTP treatment. FC and CLSM represent attractive indicator techniques to select the most efficient gas NTP treatment generating the lowest proportion of viable bacteria and the most debris.

Innovative non-thermal plasma disinfection process inside sealed bags: Assessment of bactericidal and sporicidal effectiveness in regard to current sterilization norms.

In this work, we developed a device capable to generate a non-thermal plasma discharge inside a sealed bag. The aim of this study was to assess the effectiveness of the oxygen, nitrogen and argon plasma sterilization on Pseudomonas aeruginosa, Staphylococcus aureus and Bacillus subtilis spores according to the NF EN 556 Norm. Moreover the bag integrity which is a critical key to maintain the sterile state of items after the end of the process was verified by Fourier Transform Infrared (FTIR) and X-ray Photoelectron Spectrometry (XPS) analyses. After plasma treatments, the bacterial counting showed a 6 log reduction of P. aeruginosa and S. aureus in 45 min and 120 min respectively whatever the gas used and a 4 log reduction of B. subtilis spores in 120 min with only oxygen plasma. These results were confirmed by Scanning Electron Microscopy (SEM) observations showing altered bacteria or spores and numerous debris. Taking into account the studied microorganisms, the oxygen plasma treatment showed the highest efficiency. FTIR and XPS analyses showed that this treatment induced no significant modification of the bags. To conclude this non-thermal plasma sterilization technique could be an opportunity to sterilize heat and chemical-sensitive medical devices and to preserve their sterile state after the end of the process.

Endophytic fungi associated with Sudanese medicinal plants show cytotoxic and antibiotic potential.

In this study, we isolated 15 endophytic fungi from five Sudanese medicinal plants. Each fungal endophytic strain was identified by sequencing of internal transcribed spacer (ITS) regions of rDNA. Ethyl acetate extracts were prepared from each endophyte cultivated in vitro and tested for their respective antibacterial activities and antiproliferative activities against human cancer cells. Antibacterial screening was carried out against two bacterial strains: Gram-negative Escherichia coli and Gram-positive methicillin-resistant Staphylococcus aureus, by the broth dilution method. Cell viability was evaluated by the MTT procedure after exposure of MCF7 breast cancer cells and HT29 or HCT116 human colon adenocarcinoma cells to each endophytic extract. Of interest, Byssochlamys spectabilis isolated from Euphorbia prostata showed cytotoxicity (IC50 = 1.51 ± 0.2 µg mL(-1)) against MCF7 cells, but had a low effect against HT29 or HCT116 cells (IC50 > 20 µg mL(-1)). Cladosporium cladosporioides 2, isolated from Vernonia amygdalina leaves, showed antiproliferative activities against MCF7 cells (IC50 = 10.5 ± 1.5 µg mL(-1)) only. On the other hand, B. spectabilis and Alternaria sp. extract had antibacterial activities against the S. aureus strain. The findings of this work revealed that endophytic fungi associated with medicinal plants from Sudan could be considered as an attractive source of new therapeutic compounds.

Effect of ball-milling and Fe-/Al-doping on the structural aspect and visible light photocatalytic activity of TiO2 towards Escherichia coli bacteria abatement.

Escherichia coli abatement was studied in liquid phase under visible light in the presence of two commercial titania photocatalysts, and of Fe- and Al-doped titania samples prepared by high energy ball-milling. The two commercial titania photocatalysts, Aeroxide P25 (Evonik industries) exhibiting both rutile and anatase structures and MPT625 (Ishihara Sangyo Kaisha), a Fe-, Al-, P- and S-doped titania exhibiting only the rutile phase, are active suggesting that neither the structure nor the doping is the driving parameter. Although the MPT625 UV-visible spectrum is shifted towards the visible domain with respect to the P25 one, the effect on bacteria is not increased. On the other hand, the ball milled iron-doped P25 samples exhibit low activities in bacteria abatement under visible light due to charge recombinations unfavorable to catalysis as shown by photoluminescence measurements. While doping elements are in interstitial positions within the rutile structure in MPT625 sample, they are located at the surface in ball milled samples and in isolated octahedral units according to (57)Fe Mössbauer spectrometry. The location of doping elements at the surface is suggested to be responsible for the sample cytotoxicity observed in the dark.

TiO2 photocatalysis damages lipids and proteins in Escherichia coli.

This study investigates the mechanisms of UV-A (315 to 400 nm) photocatalysis with titanium dioxide (TiO2) applied to the degradation of Escherichia coli and their effects on two key cellular components: lipids and proteins. The impact of TiO2 photocatalysis on E. coli survival was monitored by counting on agar plate and by assessing lipid peroxidation and performing proteomic analysis. We observed through malondialdehyde quantification that lipid peroxidation occurred during the photocatalytic process, and the addition of superoxide dismutase, which acts as a scavenger of the superoxide anion radical (O2·(-)), inhibited this effect by half, showing us that O2·(-) radicals participate in the photocatalytic antimicrobial effect. Qualitative analysis using two-dimensional electrophoresis allowed selection of proteins for which spot modifications were observed during the applied treatments. Two-dimensional electrophoresis highlighted that among the selected protein spots, 7 and 19 spots had already disappeared in the dark in the presence of 0.1 g/liter and 0.4 g/liter TiO2, respectively, which is accounted for by the cytotoxic effect of TiO2. Exposure to 30 min of UV-A radiation in the presence of 0.1 g/liter and 0.4 g/liter TiO2 increased the numbers of missing spots to 14 and 22, respectively. The proteins affected by photocatalytic oxidation were strongly heterogeneous in terms of location and functional category. We identified several porins, proteins implicated in stress response, in transport, and in bacterial metabolism. This study reveals the simultaneous effects of O2·(-) on lipid peroxidation and on the proteome during photocatalytic treatment and therefore contributes to a better understanding of molecular mechanisms in antibacterial photocatalytic treatment.

On the use of capillary cytometry for assessing the bactericidal effect of TiO2. Identification and involvement of reactive oxygen species.

The photocatalytic antimicrobial properties of TiO2 were studied on Staphylococcus aureus, Escherichia coli, and Pseudomonas aeruginosa bacterial strains taken as model strains for pathogenic species mainly implied in nosocomial infections. Capillary cytometry, coupled to a double-staining method for visualizing membrane integrity as a cell viability indicator, was highlighted as a rapid, easy-to-use, and automated numeration technique for quantitative and reproducible determination of cellular viability and thus, was able to give an accurate evaluation of the bactericidal effect of UV-A photocatalysis. Cytometry also enabled the study of TiO2-bacteria interactions and aggregation in the dark as well as TiO2 cytotoxicity. Compared with the traditional agar plate cultivation method, a significatively weaker reduction in cell viability was recorded by cytometry whatever the bacteria, TiO2 concentration, and duration of the photocatalytic treatment. The mismatch between both numeration methods was attributed to: (i) the presence of mixed bacteria-TiO2 aggregates that could interfere with bacteria measurement on plates, (ii) prolonged contact of the bacteria with TiO2 during incubation, which could cause additional cytotoxic damage to the bacterial wall, and (iii) the counting of viable but non-culturable bacteria as live bacteria in cytometry, whereas they cannot grow on solid media. A more pronounced difference was observed for P. aeruginosa and S. aureus bacteria, for which 2.9 and 1.9 log10 survival reduction overestimations were measured by plate counting, respectively. Using chemiluminescence, full restoration of cell viability by controlled addition of the O2˙(-) scavenger superoxide dismutase enzyme suggests that O2˙(-) acts, in our conditions, as the main reactive oxygen species responsible for the photocatalytic attack towards the targeted bacteria.