Sol-Gel Synthesis of Silica-Poly (Vinylpyrrolidone) Hybrids with Prooxidant Activity and Antibacterial Properties.

The present work deals with the sol-gel synthesis of silica-poly (vinylpyrrolidone) hybrid materials. The nanohybrids (Si-PVP) have been prepared using an acidic catalyst at ambient temperature. Tetramethyl ortosilane (TMOS) was used as a silica precursor. Poly (vinylpyrrolidone) (PVP) was introduced into the reaction mixture as a solution in ethanol with a concentration of 20%. The XRD established that the as-prepared material is amorphous. The IR and 29Si MAS NMR spectra proved the formation of a polymerized silica network as well as the hydrogen bonding interactions between the silica matrix and OH hydrogens of the silanol groups. The TEM showed spherical particle formation along with increased agglomeration tendency. The efficacy of SiO2/PVP nanoparticles as a potential antimicrobial agent against a wide range of bacteria was evaluated as bacteriostatic, using agar diffusion and spot tests. Combined effects of hybrid nanomaterial and antibiotics could significantly reduce the bactericidal concentrations of both the antibiotic and the particles, and they could also eliminate the antibiotic resistance of the pathogen. The registered prooxidant activity of the newly synthesized material was confirmative and explicatory for the antibacterial properties of the tested substance and its synergetic combination with antibiotics. The effect of new hybrid material on Crustacea Daphnia magna was also estimated as harmless under concentration of 0.1 mg/mL.

Fucoidan-Containing, Low-Adhesive Siloxane Coatings for Medical Applications: Inhibition of Bacterial Growth and Biofilm Development.

The deposition of low-adhesive siloxane coatings is a current trend for the non-toxic control of bacterial growth and biofilm formation. Total elimination of biofilm formation has not been reported so far. The aim of this investigation was to study the ability of a non-toxic, natural, biologically active substance, such as fucoidan, to inhibit bacterial growth on similar medical coatings. The fucoidan amount was varied, and its impact on the bioadhesion-influencing surface characteristics, as well as on bacterial cell growth, was investigated. The inclusion of up to 3-4 wt.% brown algae-derived fucoidan in the coatings increases their inhibitory effect, more significantly on the Gram-positive bacterium S. aureus than on the Gram-negative bacterium Escherichia coli. The biological activity of the studied siloxane coatings was ascribed to the formation of a low-adhesive, biologically active surface top layer consisting of siloxane oil and dispersed water-soluble fucoidan particles. This is the first report on the antibacterial activity of fucoidan-containing medical siloxane coatings. The experimental results give reason to expect that relevantly selected, natural biologically active substances can be efficient in the non-toxic control of bacterial growth on medical devices and, as a result, medical device-associated infections.

Investigation of Biological and Prooxidant Activity of Zinc Oxide Nanoclusters and Nanoparticles.

Zinc oxide (ZnO) nanomaterials offer some promising antibacterial effects. In this study, a new form of ZnO is synthesized, named ZnO nanocluster bars (NCs). Herein, ZnO NCs, ZnO nanoparticles (NPs), ZnO coated with silica (ZnO-SiOA, ZnO-SiOB), and SiO2 NPs were prepared, characterized, and their antimicrobial and prooxidant activity were tested. The prooxidant activity of all nanomaterials was studied according to free-radical oxidation reactions (pH 7.4 and pH 8.5) in chemiluminescent model systems. Each form of new synthesized ZnO nanomaterials exhibited a unique behavior that varied from mild to strong prooxidant properties in the Fenton`s system. ZnO NPs and ZnO NCs showed strong antibacterial effects, ZnO-SiOA NPs did not show any antibacterial activity representing biocompatibility. All tested NMs also underwent oxidation by H2O2. ZnO NCs and ZnO NPs exhibited strong oxidation at pH 8.5 in the O2-. generating system. While, SiO2, ZnO-SiOA andZnO-SiOB possessed pronounced 60-80% antioxidant effects, SiO2 NPs acted as a definitive prooxidant which was not observed in other tests. ZnO NCs are strong oxidized, assuming that ZnO NCs provide a slower release of ZnO, which leads to having a stronger effect on bacterial strains.  Thus, ZnO NCs are an important antibacterial agent that could be an emergent replacement of traditional antibiotics.

Comparative Study of the Marinobacter hydrocarbonoclasticus Biofilm Formation on Antioxidants Containing Siloxane Composite Coatings.

No systematic study of antioxidant containing coatings and their anti-biofilm action has been reported so far. The utilization of antioxidants in protective coatings to inhibit marine biofilm formation is a current challenge. The aim of this preliminary study was to prepare, characterize and compare the efficiency of low adhesive siloxane composite coatings equally loaded with different antioxidants against mono-species biofilms formation. Most often participating in the marine biofilms formation, Marinobacter hydrocarbonoclasticus was the test bacterium. Both the biofilm covered surface area (BCSA) and corrected total cell fluorescence (CTCF) (by fluorescent microscopy) were selected as the parameters for quantification of the biofilm after 1 h and 4 h incubation. Differing extents of altered surface characteristics (physical-chemical; physical-mechanical) and the specific affection of M. hydrocarbonoclasticus biofilm formation in both reduction and stimulation, were found in the studied antioxidant containing coatings, depending on the chemical nature of the used antioxidant. It was concluded that not all antioxidants reduce mono-species biofilm formation; antioxidant chemical reactivity stipulates the formation of an altered vulcanization network of the siloxane composites and thus microbial adhesion which influences the surface characteristics of the vulcanized coatings; and low surface energy combined with a low indentation elastic modulus are probably pre-requisites of low microbial adhesion.

Prooxidant and antimicrobic effects of iron and titanium oxide nanoparticles and thalicarpine.

The aim is to evaluate the prooxidant and antimicrobial effects of Fe3O4 and TiO2 nanoparticles and thalicarpine by luminescent and standard microbiological assays. Their effect on the kinetics of free-radical oxidation reactions (at pH 7.4 and pH 8.5) is studied in the following model systems, using activated chemiluminescence: chemical, with Fenton's reagent (H2O2-FeSO4)-for the generation of hydroxyl radicals (.OH); chemical, with oxidant hydrogen peroxide (H2O2); chemical (NAD.H-PhMS), for the generation of superoxide radicals (O2.-). Fe3O4 nanoparticles exhibit highly pronounced antioxidant properties; TiO2 nanoparticles exhibit mild to moderate prooxidant properties at neutral and alkaline conditions. Those properties are tested by the chemiluminescent method for the first time. Thalicarpine and its combination with TiO2 nanoparticles exhibit pronounced antioxidant activities at pH 8.5 which are lost and transformed into well-presented prooxidant effects at pH 7.4. That is a result-supported proof on the observed typical properties of thalicarpine and TiO2, namely antibacterial, organic-preserving and anti-pathogenic activities. The antimicrobial effect is tested on Gram-positive and Gram-negative bacteria: two strains of Escherichia coli, Bacillus cereus 1095 and Staphylococcus aureus. All bacteria are destroyed after the application of TiO2, but not Fe3O4 nanoparticles, showing their antibacterial effect. Thalicarpine, in combination with TiO2, showed even synergetic antibacterial effect.

Antibacterial effect of TiO2 :Cu:Ag thin coatings on Pseudomonas strain measured by microbiological and ATP assays.

The aim of this study is to investigate the antimicrobial properties of nanocomposite thin TiO2 :Cu:Ag films on Pseudomonas putida as a natural isolate and an opportunistic pathogen. Several different methods were used to compare the antibacterial effect of thin TiO2 :Cu:Ag layers obtained by radiofrequency magnetron sputter deposition against P. putida: optical density of the bacterial suspension, most probable number of survived cells, dehydrogenase activity inhibition, scanning electron microscopy images, atomic flame absorption spectroscopy, and adenosine triphosphate (ATP) luminescent assay. Optical density measurements and most probable plate count were in agreement and showed a strong bactericidal effect of the as-deposited and only bacteriostatic effect of the annealed coatings with the same metal content on tested bacteria. As the metal quantity in the medium rises during the first hour of the experiment, it could be suggested that this is the main reason for cell death. ATP-luminescent assay showed up to 18-fold reduction of the signal. It was compared with other microbiological and biochemical assays to prove the strong antibacterial effect of nanocomposite thin TiO2 :Cu:Ag coatings with the possibilities of medical applications. Protection of medical devices against infections is a significant current challenge raised by an increasing number of medical devices-associated infections and microbial resistance to conventional antibiotic and multidrug treatments. Deposition of antimicrobial coatings is one of the current approaches to mitigate the problem.

Preparation and Biological Activity of New Collagen Composites, Part I: Collagen/Zinc Titanate Nanocomposites.

The aim of this investigation was to develop new antimicrobial collagen/zinc titanate (ZnTiO3) biomaterials using a sol-gel cryogenic draying technology in keeping the native collagen activity. Broad-spectrum antimicrobial activity was demonstrated against Firmicutes (Staphylococcus epidermidis, Bacillus cereus, and Candida lusitaniae) and Gracilicutes (Escherichia coli, Salmonella enterica, and Pseudomonas putida) microorganisms. The antimicrobial activity as well as the cytotoxicity were specific for the different test microorganisms (Gram-positive and Gram-negative bacteria and fungi) and model eukaryotic cells (osteosarcoma, fibroblast, and keratinocyte cells), respectively, and both were depending on the ZnTiO3 concentration. Three mechanisms of the antimicrobial action were supposed, including (i) mechanical demolition of the cell wall and membrane by the crystal nanoparticles of the ZnTiO3 entrapped in the collagen matrix, (ii) chelation of its metal ions, and (iii) formation of free oxygen radicals due to the interaction between the microbial cells and antimicrobial agent. It was concluded that the optimal balance between antimicrobial activity and cytotoxicity could be achieved by a variation of the ZnTiO3 concentration. The antifungal and broad-spectrum antibacterial activity of the studied collagen/ZnTiO3 nanocomposites, combined with a low cytotoxicity, makes them a promising anti-infection biomaterial.

Microbial community development of biofilm in Amaranth decolourization technology analysed by FISH.

The aim of this study was to elucidate the role, the space distribution and the relationships of the bacteria from the genus Pseudomonas in a biofilm community during semi-continuous Amaranth decolourization process in model sand biofilters. The examined parameters of the process were as follows: technological parameters; key enzyme activities (azoreductase, succinate dehydrogenase, catechol-1,2-dioxygenase, catechol-2,3-dioxygenase); the number of azo-degrading bacteria and the bacteria from genus Pseudomonas (plate count technique); the amount and the location of Pseudomonas sp. using fluorescent in situ hybridization (FISH). The results showed that the increase of the Amaranth removal rate with 120% was accompanied with increase of the enzyme activities of the biofilm (azoreductase activity - with 25.90% and succinate dehydrogenase - with 10.61%). The enzyme assays showed absence of activity for сatechol-1,2-dioxygenase and catechol-2,3-dioxygenase at the early phase and high activities of the same oxygenases at the late phase (2.76 and 1.74 µmol/min mg protein, respectively). In the beginning of the process (0-191 h), the number of the culturable microorganisms from genus Pseudomonas was increased with 48.76% but at the late phase (191-455 h) they were decreased with 15.25% while the quantity of the non-culturable bacteria from this genus with synergetic relationships was increased with 23.26%. The dominant microbial factors were identified in the structure of the biofilm during the azo-degradation process by using FISH analysis. Furthermore, the inner mechanisms for increase of the rate and the range of the detoxification were revealed during the complex wastewater treatment processes.

Water quality assessment of aquatic ecosystems using ecological criteria - case study in Bulgaria.

Four aquatic ecosystems (two rivers and two dams) situated in the western part of Bulgaria were investigated over a three years' period. The River Egulya and Petrohan dam are situated in mountainous regions at about 1000 m altitude, and are not influenced by any anthropogenic sources. Petrohan dam is a site for long-term ecosystem research as a part of Bulgarian long-term ecological research network. The other two systems belong to populated industrial areas. The River Martinovska flows through a region with former long-term mining activity, while Ogosta dam is near a battery production factory. Both the geochemical and geographical ecosystems' conditions are different, and their social usage as well. Ogosta dam water is used for irrigation and Petrohan dam for electric supply. The ecosystem sensitivity to heavy metals was evaluated by a critical load approach. Two criteria were used for risk assessment: critical load exceedance and microbial toxicity test. All studied ecosystems were more sensitive to cadmium than to lead deposition. The potential risk of Cd damage is higher for Petrohan dam and the River Egulya, where critical load exceedance was calculated for two years. Pseudomonas putida growth inhibition test detected a lack of toxicity for all studied ecosystems at the time of investigation with the exception of the low water September sample of the River Martinovska. The fast bacterial test is very suitable for a regular measurement of water toxicity because of its simplicity, lack of sophisticated equipment and clear results.

Biomarkers for the prediction of the resistance and susceptibility of grapevine leaves to downy mildew.

We examined metabolic profiles of acetone and butanol extracts obtained from the leaves of 18 seedlings of the Bulgarian wine-making cultivar Storgozia. The acetone extracts contained the components from the leaf surface, while the butanol extracts were enriched with polar components from inside the leaf tissue. The leaves displayed different degrees of resistance and susceptibility to the etiological agent downy mildew, Plasmopara viticola. Based on the statistically significant correlations (P<0.05) between the GC-MS data of the identified metabolites and the estimated leaf resistances, 10 individual components were proposed as possible biomarkers for the downy mildew resistance and susceptibility in grapevine. All were found in the butanol extracts, and can be considered to form two groups: compounds with high correlations (r=+/-0.50 to +/-1.00) - 3-hydroxybutanoic acid, 2,3,4-trihydroxybutanoic acid, 2,3,4-trihydroxybutanoic acid (isomer), hexadecanoic acid, 3-hydroxyhexanoic acid and myo-inositol, and compounds with moderate correlations (r=+/-0.30 to +/-0.49) hydroxybutanedioic acid, alanine, glutamine, arabinoic acid and aldohexoses. Among them, the more polar compounds were related to sensitivity, and only hexadecanoic and the monohydroxycarboxylic acids were related to resistance in grapevine.