Production and Characterisation of an Exopolysaccharide by Bacillus amyloliquefaciens: Biotechnological Applications.

The Bacillus amyloliquefaciens RT7 strain was isolated from an extreme acidic environment and identified. The biodegradation capabilities of the strain using different carbon sources (glucose, oleic acid, Tween 80, PEG 200, and the combination of glucose-Tween 80) were evaluated via an indirect impedance technique. The glucose-Tween 80 combination was further studied using nuclear magnetic resonance (NMR). The exopolysaccharide (EPSRT7) that had been produced with the strain when biodegrading glucose-Tween 80 was isolated and characterised using different techniques (GC-MS, HPLC/MSMS, ATR-FTIR, TGA, and DSC), and its molecular weight was estimated. The results show that the average molecular weight of EPSRT7 was approximately 7.0794 × 104 Da and a heteropolysaccharide composed of mannose, glucose, galactose, and xylose (molar ratio, 1:0.5:0.1:0.1) with good thermostability. EPSRT7 showed good emulsifying activity against different natural oils and hydrocarbons at high concentrations (2 mg/mL) and at the studied pH range (3.1-7.2). It also presented good emulsifying activity compared to that of commercial emulsifiers. Lastly, EPSRT7 showed antioxidant capacity for different free radicals, a lack of cytotoxicity, and antioxidant activity at the cellular level. EPSRT7 has promising applications in bioremediation processes and other industrial applications.

Potential Applications of an Exopolysaccharide Produced by Bacillus xiamenensis RT6 Isolated from an Acidic Environment.

The Bacillus xiamenensis RT6 strain was isolated and identified by morphological, biochemical and molecular tests from an extreme acidic environment, Rio Tinto (Huelva). Optimisation tests for exopolysaccharide (EPS) production in different culture media determined that the best medium was a minimal medium with glucose as the only carbon source. The exopolymer (EPSt) produced by the strain was isolated and characterised using different techniques (GC-MS, HPLC/MSMS, ATR-FTIR, TGA, DSC). The molecular weight of EPSt was estimated. The results showed that the average molecular weight of EPSt was approximately 2.71 × 104 Da and was made up of a heteropolysaccharide composed of glucose (60%), mannose (20%) and galactose (20%). The EPSt showed antioxidant capabilities that significantly improved cell viability. Metal chelation determined that EPSt could reduce the concentration of transition metals such as iron at the highest concentrations tested. Finally, the emulsification study showed that EPSt was able to emulsify different natural polysaccharide oils, reaching up to an 80% efficiency (olive and sesame oil), and was a good candidate for the substitution of the most polluting emulsifiers. The EPSt was found to be suitable for pharmaceutical and industrial applications.

Anion effects on kinetics and thermodynamics of CO2 absorption in ionic liquids.

A thermogravimetric technique based on a magnetic suspension balance operating in dynamic mode was used to study the thermodynamics (in terms of solubility and Henry's law constants) and kinetics (i.e., diffusion coefficients) of CO2 in the ionic liquids [bmim][PF6], [bmim][NTf2], and [bmim][FAP] at temperatures of 298.15, 308.15, and 323.15 K and pressures up to 20 bar. The experimental technique employed was shown to be a fast, accurate, and low-solvent-consuming method to evaluate the suitability of the ionic liquids (ILs) to be used as CO2 absorbents. Thermodynamic results confirmed that the solubility of CO2 in the ILs followed the order [bmim][FAP] > [bmim][NTf2] > [bmim][PF6], increasing with decreasing temperatures and increasing pressures. Kinetic data showed that the diffusion coefficients of CO2 in the ILs followed a different order, [bmim][NTf2] > [bmim][FAP] > [bmim][PF6], increasing with increasing temperatures and pressures. These results evidenced the different influence of the IL structure and operating conditions on the solubility and absorption rate of CO2, illustrating the importance of considering both thermodynamic and kinetic aspects to select adequate ILs for CO2 absorption. On the other hand, the empirical Wilke-Chang correlation was successfully applied to estimate the diffusion coefficients of the systems, with results indicating the suitability of this approach to foresee the kinetic performance of ILs to absorb CO2. The research methodology proposed herein might be helpful in the selection of efficient absorption solvents based on ILs for postcombustion CO2 capture.

Application of molecular techniques to the elucidation of the microbial community structure of antique paintings.

This paper uses molecular techniques to describe the microstructure and microbiological communities of sixteenth century artwork and their relationships. The microbiological populations, analysed by denaturing gradient gel electrophoresis (DGGE), were highly influenced by the chemical composition of the pictorial layers detected by energy-dispersive X-ray analysis. DGGE revealed that the diversity of microbial communities was lower in pictorial layers composed of pigments with metals, such as Pb, Cu and Hg, than in those found in pictorial layers without such compounds. The number of cultivable microorganisms, mainly fungi and bacteria, was very low in comparison to those found by DGGE, revealing the presence of both cultivable and as-yet-uncultivated (or not viable) species in the samples analysed. Both fungi and bacteria were present in a non-random spatial distribution. Environmental scanning electron microscopy and fluorescent in situ hybridisation analyses revealed that bacterial populations were usually found in close contact with the surface of the pictorial layers, and fungal populations were located on the bacterial biofilm. This work shows, for the first time, the correlation between the diversity of the microbial populations and the chemical composition of the pictorial layers of an artwork.

Cwp2p, the plasma membrane receptor for Pichia membranifaciens killer toxin.

PMKT is a channel-forming killer toxin secreted by Pichia membranifaciens. To identify novel genes that mediate cellular resistance to PMKT we screened a collection of 288 deletion mutants. We found 29 open reading frames (ORFs) that, when deleted, confer resistance to PMKT. In addition, the deletion of 15 ORFs was observed to increase protoplast resistance, in agreement with the initial assumption that a plasma membrane receptor for PMKT exists. Whole cells and protoplasts of a cwp2Delta mutant were found to be completely resistant to PMKT and were unable to bind PMKT, indicating that Cwp2p interacts with it. A protein with a molecular mass of 11.7 kDa was purified from PMKT-affinity columns. This protein was sequenced and identified as Cwp2p. Glycosylphosphatidylinositol (GPI) anchoring-defective mutants were much less sensitive to PMKT, as were wild-type protoplasts pretreated with phosphatidylinositol-specific phospholipase C to remove GPI-anchored proteins, indicating that the GPI-anchored precursor of Cwp2p is also necessary for PMKT activity. Carboxyfluorescein-entrapped liposomes containing a purified GFP-Cwp2p fusion protein in their membranes were much more sensitive to PMKT than protein-free liposomes. Cwp2p and its GPI-anchored precursor are proposed for the first time to be involved as PMKT secondary receptors.

The transcriptional response of Saccharomyces cerevisiae to Pichia membranifaciens killer toxin.

The transcriptional response of Saccharomyces cerevisiae to Pichia membranifaciens killer toxin (PMKT) was investigated. We explored the global gene expression responses of the yeast S. cerevisiae to PMKT using DNA microarrays, real time quantitative PCR, and Northern blot. We identified 146 genes whose expression was significantly altered in response to PMKT in a non-random functional distribution. The majority of induced genes, most of them related to the high osmolarity glycerol (HOG) pathway, were core environmental stress response genes, showing that the coordinated transcriptional response to PMKT is related to changes in ionic homeostasis. Hog1p was observed to be phosphorylated in response to PMKT implicating the HOG signaling pathway. Individually deleted mutants of both up- (99) and down-regulated genes (47) were studied for altered sensitivity; it was observed that the deletion of up-regulated genes generated hypersensitivity (82%) to PMKT. Deletion of down-regulated genes generated wild-type (36%), resistant (47%), and hypersensitive (17%) phenotypes. This is the first study that shows the existence of a transcriptional response to the poisoning effects of a killer toxin.