Antibacterial Activity of ZnSe, ZnSe-TiO2 and TiO2 Particles Tailored by Lysozyme Loading and Visible Light Irradiation.

ZnSe, ZnSe-TiO2 microspheres and nanostructured TiO2 obtained by hydrothermal and sol-gel methods were tested against Staphylococcus aureus ATCC 25923 and Micrococcus lysodeikticus ATCC 4698 before and after lysozyme (Lys) loading. Morphological characterization of inorganic matrices and hybrid organic-inorganic complexes were performed by microscopy techniques (SEM, AFM and Dark Field Hyperspectral Microscopy). Light absorption properties of ZnSe, ZnSe-TiO2 and TiO2 powders were assessed by UV-visible spectroscopy and their ability to generate reactive oxygen species (•OH and O2•-) under visible light irradiation was investigated. Antibacterial activity of ZnSe, ZnSe-TiO2, TiO2, Lys/ZnSe, Lys/ZnSe-TiO2 and Lys/TiO2 samples under exposure to visible light irradiation (λ > 420 nm) was tested against Staphylococcus aureus and Micrococcus lysodeikticus and correlated with ROS photogeneration.

Characterization of Some Salt-Tolerant Bacterial Hydrolases with Potential Utility in Cultural Heritage Bio-Cleaning.

Salt-tolerant enzymes produced by halophilic and halotolerant microorganisms have been proposed to be used in various applications that involve high saline conditions. Considering their biotechnological significance and the current need for more efficient producers of such catalysts, the present study aimed to evaluate the extracellular proteolytic, esterolytic, cellulolytic and xylanolytic activities of some halotolerant strains, and to characterize their functional parameters. A total of 21 bacterial and fungal strains belonging to the genera Bacillus, Virgibacillus, Salinivibrio, Salinicoccus, Psychrobacter, Nocardiopsis, Penicillium, Aspergillus, and Emericellopsis were assayed by quantitative methods. Among them, the members of the Bacillus genus exhibited the highest catalytic activities. The exoenzymes produced by three selected Bacillus strains were active over wide ranges of salinity, temperature and pH. Proteases were active at 20-80 °C, pH 6-10, and 0-1 M NaCl, while esterases showed good catalytic activities at 20-80 °C, pH 7.5-10, and 0-4 M NaCl. Cellulases and xylanases were active at 20-80 °C, pH 5-10, and 0-5 M NaCl. Due to such properties, these hydrolases could be used in a newly proposed application, namely to clean aged consolidants and organic deposits accumulated over time from the surfaces of salt-loaded wall paintings.

Characterization by polyphasic approach of some indigenous halophilic archaea of Djelfa's rock salt "Hadjr el Meelh", Algeria.

Background and Objectives: Hadjr El Melh of Djelfa is an example of hypersaline ecosystems, which can harbor a wide variety of microorganisms under hostile physicochemical conditions. Given the importance of the study of halophilic microorganisms present there in terms of fundamental and applied microbiology, the purpose of this study was to characterize some halophilic archaea isolated from the brines of this environment. Materials and Methods: Eight water samples were chosen randomly and collected for physicochemical and microbiological analyses. Isolation of halophilic archaea was carried out by membrane filter technique. Ten strains were identified by polyphasic approach and tested for enzymes production. Results: Water samples of Djelfa's rock salt were slightly acidic to neutral in pH (6.55-7.36) with salinity ranging from 258.68 g/l to 493.91 g/l. Phenotypic, biochemical, taxonomic and phylogenetic characteristics indicated that all strains were classified within the family of Halobacteiaceae. Based on the comparison of DNA sequences encoded 16S rRNA, it was determined that seven strains were affiliated to the genus Haloarcula, two strains were related to the genus Halobacterium and one strain within the genus Haloferax. Production of different enzymes such as protease, amylase, esterase, lipase, lecithinase, gelatinase and cellulase on solid medium indicated that one strain (S2-2) produced amylase, esterase, lecithinase and protease. However, no strains showed cellulolytic or lipolytic activity. Gelatinase was found in all tested strains. Conclusion: This report constitutes the first preliminary study of culturable halophilic archaea recovered from the brines of Djelfa's rock salt with a promising enzymatic potential in various fields of biotechnology.

Effect of ZnO on Properties of Gels for Heritage Objects Conservation.

One of the current research objectives is the development of new films for the conservation of glass heritage objects. The value of historical glass objects is given by the technology and raw materials used in production as well as their transparency and color. Their colors are correlated with oxide composition rich in transitional metals, which decrease resistance of corrosive agents from the atmosphere. In this paper, SiO2-ZnO gels have been designed to protect historical glass objects. The sol-gel method used to obtain gels is a powerful tool for functionalizing different materials. An important functionalization is the antibacterial activity. By applying a gel, the coated material is able to decrease the growth of bacteria. After deposition, some gels must be strengthened by heat treatment. The effect of ZnO content (10 mol% and 20 mol%) on the properties of the studied gels was investigated by Differential scanning calorimetry (DSC), Fourier transform infrared (FTIR), X-ray diffraction (XRD), Scanning electron microscopy (SEM), and antibacterial tests. Escherichia coli ATCC 25922, Staphylococcus aureus ATCC 25923, and the halotolerant bacterium, Virgibacillus halodenitrificans, isolated from a salt crystal from Unirea mine, Slanic Prahova, Romania, were used. The gel Gel 2 (SiO2-ZnO (20 mol%)) showed the best properties.

Bioprospecting for Novel Halophilic and Halotolerant Sources of Hydrolytic Enzymes in Brackish, Saline and Hypersaline Lakes of Romania.

Halophilic and halotolerant microorganisms represent promising sources of salt-tolerant enzymes that could be used in various biotechnological processes where high salt concentrations would otherwise inhibit enzymatic transformations. Considering the current need for more efficient biocatalysts, the present study aimed to explore the microbial diversity of five under- or uninvestigated salty lakes in Romania for novel sources of hydrolytic enzymes. Bacteria, archaea and fungi were obtained by culture-based approaches and screened for the production of six hydrolases (protease, lipase, amylase, cellulase, xylanase and pectinase) using agar plate-based assays. Moreover, the phylogeny of bacterial and archaeal isolates was studied through molecular methods. From a total of 244 microbial isolates, 182 (74.6%) were represented by bacteria, 22 (9%) by archaea, and 40 (16.4%) by fungi. While most bacteria synthesized protease and lipase, the most frequent hydrolase produced by fungi was pectinase. The archaeal isolates had limited hydrolytic activity, being able to produce only amylase and cellulase. Among the taxonomically identified isolates, the best hydrolytic activities were observed in halotolerant bacteria belonging to the genus Bacillus and in extremely halophilic archaea of the genera Haloterrigena and Halostagnicola. Therefore, the present study highlights that the investigated lakes harbor various promising species of microorganisms able to produce industrially valuable enzymes.

Biocatalytic Strategy for Grafting Natural Lignin with Aniline.

This paper presents an enzyme biocatalytic method for grafting lignin (grafting bioprocess) with aniline, leading to an amino-derivatized polymeric product with modified properties (e.g., conductivity, acidity/basicity, thermostability and amino-functionalization). Peroxidase enzyme was used as a biocatalyst and H2O2 was used as an oxidation reagent, while the oxidative insertion of aniline into the lignin structure followed a radical mechanism specific for the peroxidase enzyme. The grafting bioprocess was tested in different configurations by varying the source of peroxidase, enzyme concentration and type of lignin. Its performance was evaluated in terms of aniline conversion calculated based on UV-vis analysis. The insertion of amine groups was checked by 1H-NMR technique, where NH protons were detected in the range of 5.01-4.99 ppm. The FTIR spectra, collected before and after the grafting bioprocess, gave evidence for the lignin modification. Finally, the abundance of grafted amine groups was correlated with the decrease of the free -OH groups (from 0.030 to 0.009 -OH groups/L for initial and grafted lignin, respectively). Additionally, the grafted lignin was characterized using conductivity measurements, gel permeation chromatography (GPC), thermogravimetric analysis (TGA), temperature-programmed desorption (TPD-NH3/CO2) and scanning electron microscopy (SEM) analyses. The investigated properties of the developed lignopolymer demonstrated its disposability for specific industrial applications of derivatized lignin.

Exploring the hydrolytic potential of cultured halophilic bacteria isolated from the Atacama Desert.

Considering that most industrial processes are carried out under harsh physicochemical conditions, which would inactivate enzymes from commonly isolated mesophilic organisms, current studies are geared toward the identification of extremophilic microorganisms producing enzymes resistant to extreme salt concentrations, temperature and pH. Among the extremophiles, halophilic microorganisms are an important source of salt-tolerant enzymes that can be used in varying biotechnological applications. In this context, the aim of the present work was to isolate and identify halophiles producing hydrolases from the Atacama Desert, one of the harshest environments on Earth. Isolates were recovered from halite samples and screened for the presence of seven different hydrolase activities (amylase, caseinase, gelatinase, lipase, pectinase, cellulase and inulinase) using agar plate-based assays. From a total of 23 halophilic bacterial isolates, most showed lipolytic (19 strains) and pectinolytic (11 strains) activities. The molecular identification of eight selected isolates showed a strong similarity to members of the Halomonas and Idiomarina genera. Therefore, the present study represents a preliminary, but essential, step to identify novel biological sources of extremozymes in an environment once thought to be devoid of life.

Ecology and physiology of halophilic microorganisms - Thematic issue based on papers presented at Halophiles 2019 - 12th International Conference on Halophilic Microorganisms, Cluj-Napoca, Romania, 24-28 June, 2019.

In June 2019, the 12th International Conference on Halophilic Microorganisms - Halophiles 2019, was held in Cluj-Napoca, Romania. This thematic issue of FEMS Microbiology Letters contains papers based on lectures and posters presented at the conference. We here provide a short overview of past research on hypersaline environments in Romania and the microorganisms inhabiting them, and briefly present the papers published in this thematic issue.

One-Pot Enzymatic Production of Lignin-Composites.

A novel and efficient one-pot system for green production of artificial lignin bio-composites has been developed. Monolignols such as sinapyl (SA) and coniferyl (CA) alcohols were linked together with caffeic acid (CafAc) affording a polymeric network similar with natural lignin. The interaction of the dissolved SA/CA with CafAc already bound on a solid support (SC2/SC6-CafAc) allowed the attachment of the polymeric product direct on the support surface (SC2/SC6-CafAc-L1 and SC2/SC6-CafAc-L2, from CA and SA, respectively). Accordingly, this procedure offers the advantage of a simultaneous polymer production and deposition. Chemically, oxi-copolymerization of phenolic derivatives (SA/CA and CAfAc) was performed with H2O2 as oxidation reagent using peroxidase enzyme (2-1B mutant of versatile peroxidase from Pleurotus eryngii) as catalyst. The system performance reached a maximum of conversion for SA and CA of 71.1 and 49.8%, respectively. The conversion is affected by the system polarity as resulted from the addition of a co-solvent (e.g., MeOH, EtOH, or THF). The chemical structure, morphology, and properties of the bio-composites surface were investigated using different techniques, e.g., FTIR, TPD-NH3, TGA, contact angle, and SEM. Thus, it was demonstrated that the SA monolignol favored bio-composites with a dense polymeric surface, high acidity, and low hydrophobicity, while CA allowed the production of thinner polymeric layers with high hydrophobicity.

Spectroscopic Investigation of the Interaction of the Anticancer Drug Mitoxantrone with Sodium Taurodeoxycholate (NaTDC) and Sodium Taurocholate (NaTC) Bile Salts.

The focus of the present work was to investigate the interaction of the anticancer drug mitoxantrone with two bile salts, sodium taurodeoxycholate (NaTDC) and sodium taurocholate (NaTC). Ultraviolet-visible (UV-Vis) absorption and electron paramagnetic resonance (EPR) spectroscopy were used to quantify the interaction and to obtain information on the location of mitoxantrone in bile salt micelles. The presence of submicellar concentrations of both bile salts induces mitoxantrone aggregation and the extent of drug aggregation in NaTDC is higher than in NaTC. For micellar bile salts concentrations, mitoxantrone monomers are entrapped in the micellar core. Binding constants, micelle/water partition coefficients and the corresponding thermodynamic parameters for binding and partitioning processes were estimated using the changes in monomer absorbance in the presence of bile salts. Binding interaction of mitoxantrone is stronger for NaTDC than NaTC micelles, whereas partitioning efficiency is higher for NaTC micelles for all investigated temperatures. Thermodynamic parameters indicate that both binding and partitioning processes are spontaneous and entropy controlled. The spectral behavior and thermodynamic parameters indicate distinct types of mitoxantrone interaction with NaTDC and NaTC micelles supported by the differences in nature and structure of bile salts micelles.

Mitoxantrone-Surfactant Interactions: A Physicochemical Overview.

Mitoxantrone is a synthetic anticancer drug used clinically in the treatment of different types of cancer. It was developed as a doxorubicin analogue in a program to find drugs with improved antitumor activity and decreased cardiotoxicity compared with the anthracyclines. As the cell membrane is the first barrier encountered by anticancer drugs before reaching the DNA sites inside the cells and as surfactant micelles are known as simple model systems for biological membranes, the drugs-surfactant interaction has been the subject of great research interest. Further, quantitative understanding of the interactions of drugs with biomimicking structures like surfactant micelles may provide helpful information for the control of physicochemical properties and bioactivities of encapsulated drugs in order to design better delivery systems with possible biomedical applications. The present review describes the physicochemical aspects of the interactions between the anticancer drug mitoxantrone and different surfactants. Mitoxantrone-micelle binding constants, partitions coefficient of the drug between aqueous and micellar phases and the corresponding Gibbs free energy for the above processes, and the probable location of drug molecules in the micelles are discussed.

Bifunctional carbohydrate biopolymers entrapped lipase as catalyst for the two consecutive conversions of α-pinene to oxy-derivatives.

Bifunctional catalysts designed as carbohydrate biopolymers entrapping lipase have been investigated for the biotransformation of a natural compound (α-pinene) to oxy-derivatives. Lipases assisted the epoxidation of α-pinene using H2O2 as oxidation reagent and ethyl acetate as both acetate-supplier and solvent affording α-pinene oxide as the main product. Further, the biopolymer promoted the isomerization of α-pinene oxide to campholenic aldehyde and trans-carenol. In this case, the biopolymers played double roles of the support and also active part of the bifunctional catalyst. Screening of enzymes and their entrapping in a biopolymeric matrix (e.g. Ca-alginate and κ-carrageenan) indicated the lipase extracted from Aspergillus niger as the most efficient. In addition, the presence of biopolymers enhanced the catalytic activity of the immobilized lipase (i.e. 13.39×10(3), 19.76×10(3)and 26.46×10(3) for the free lipase, lipase-carrageenan and lipase-alginate, respectively). The catalysts stability and reusability were confirmed in eight consecutively reaction runs.

Highly thermostable exopolysaccharide produced by the moderately halophilic bacterium isolated from a man-made young salt lake in Romania.

Halophilic bacterial strains isolated from a man-made salt lake in Romania produce a specific exopolysaccharide. This product is synthesized under both static and stirring conditions, and the yield of this exoploysaccharide depends on the composition of the culture medium. The highest amount of polymer was obtained in the presence of casamino acids and yeast extract, under stirring conditions. This polymer has high thermostability, with a melting point at 207 degrees C. The melting process is associated with the thermal degradation of the compound. This polymer is characterized by maximum absorbtion at 260 nm and fluorescence emission at 530 nm. The FT-IR analysis of the polymer revealed the presence of a saccharide structure and of amine and sulfate groups throughout the sugar backbone.

Phylogenetic relationships within the family Halobacteriaceae inferred from rpoB' gene and protein sequences.

In order to clarify the current phylogeny of the haloarchaea, particularly the closely related genera that have been difficult to sort out using 16S rRNA gene sequences, the DNA-dependent RNA polymerase subunit B' gene (rpoB') was used as a complementary molecular marker. Partial sequences of the gene were determined from 16 strains of the family Halobacteriaceae. Comparisons of phylogenetic trees inferred from the gene and protein sequences as well as from corresponding 16S rRNA gene sequences suggested that species of the genera Natrialba, Natronococcus, Halobiforma, Natronobacterium, Natronorubrum, Natrinema/Haloterrigena and Natronolimnobius formed a monophyletic group in all trees. In the RpoB' protein tree, the alkaliphilic species Natrialba chahannaoensis, Natrialba hulunbeirensis and Natrialba magadii formed a tight group, while the neutrophilic species Natrialba asiatica formed a separate group with species of the genera Natronorubrum and Natronolimnobius. Species of the genus Natronorubrum were split into two groups in both the rpoB' gene and protein trees. The most important advantage of the use of the rpoB' gene over the 16S rRNA gene is that sequences of the former are highly conserved amongst species of the family Halobacteriaceae. All sequences determined so far can be aligned unambiguously without any gaps. On the other hand, gaps are necessary at 49 positions in the inner part of the alignment of 16S rRNA gene sequences. The rpoB' gene and protein sequences can be used as an excellent alternative molecular marker in phylogenetic analysis of the Halobacteriaceae.

Haloferax prahovense sp. nov., an extremely halophilic archaeon isolated from a Romanian salt lake.

A novel halophilic archaeon, strain TL6T, was isolated from Telega Lake, a hypersaline environment in Prahova county, Romania. Strain TL6T was able to grow in media with a salt concentration of between 2.5 and 5.2 M, with optimum growth at a concentration of 3.5 M. The novel strain was able to grow at concentrations of 1 M MgCl2 or less, with an optimum of 0.4 M Mg2+. Growth of the novel strain occurred between pH 6.0 and 8.5, with an optimum of pH 7.0-7.5. The G+C content of the total DNA was 63.7 mol%. The 16S rRNA gene sequence of the novel strain was most closely related to species of the genus Haloferax (97.3-99.3 % sequence similarity). The lipid profile of the novel strain corresponded to that of other species belonging to the genus Haloferax. A comparative analysis of the phenotypic properties and DNA-DNA hybridization between the novel strain and other species of the genus Haloferax strongly supported the conclusion that strain TL6T represents a novel species within this genus, for which the name Haloferax prahovense sp. nov., is proposed. The type strain is TL6T (=JCM 13924T=DSM 18310T).