The Ciliate Protist Tetrahymena pyriformis as a Cellular Adhesion Model for the Pathogenic Bacterium Staphylococcus aureus

Staphylococcus aureus is one of the main pathogenic agents responsible for nosocomial and community-acquired bacterial infections. The pathogenicity of this Gram-positive bacterium is ensured by its different adhesion factors. Collagen and the extracellular glycoprotein adhesin are among the Staphylococcus most important virulence factors. It has been shown that most of the S. aureus strains carry the ica operon, responsible for biofilm production. However, the coexpression of the icaA and the icaD genes is necessary for complete biofilm synthesis. The aim of our study was to study a collection of 15 clinical strains of S. aureus from different sources for the presence of cna and icaD genes coding intercellular adhesion proteins. We also intended to estimate the strains¿ ability to form biofilms by the red Cong method and to test the adhesion ability of S. aureus to the ciliated protist Tetrahymena pyriformis, which we used as a novel cellular adhesion model. Finally, we checked the adhesion¿s inhibition capacity of some plants extracts. The molecular detection of adhesion genes revealed that 80% of strains are cna positive, and 73% are icaD positive. Qualitative biofilm production of S. aureus revealed that 66.6% of strains were slime producers. The adhesion test revealed that 20% of strains are strongly adhering to T. pyriformis and that the Clematis cirrhosa extract has an anti-adhering effect of S. aureus to the ciliate T. pyriformis.Peer Reviewe

Effect of Origanum vulgare Essential Oil Supplementation on the Advanced Parameters of Mobility and on the Integrity of Human Sperm DNA

The reduced sperm mobility is one of the most important causes of male infertility. Several reports have indicated that the treatment of subnormal sperm samples with certain agents prior to artificial insemination significantly improves the fertilizing potential of sperm. We have among others some stimulants such as pentoxifylline, relaxin, prostaglandin E, and diltiazem. In our precedent work, we had tested the effect of supplementation with three essential oils, namely, sage (Salvia officinalis), oregano (Origanum vulgare), and eucalyptus (Eucalyptus globulus), on sperm cell mobility and vitality. Oregano oil had shown interesting biological properties by giving the best values of progressive mobility and vitality. In this study, we aim to verify the effect of oregano oil supplementation on the advanced parameters of mobility and on the integrity of the sperm DNA of 25 male infertile volunteers. Our results showed that oregano oil over an incubation period of 5 to 10 min of exposure significantly improves the advanced parameters of mobility, namely, curvilinear velocity (VCL), linear velocity (VSL), the mean velocity of the path (VAP), and the amplitude of the displacement (ALH). The effect of the increase in the VCL decreased the linearity (LIN), the mean line (STR), and the mean wobble (WOB). Oregano oil at 5 min had no significant effect on the DNA fragmentation index (DFI) and sperm decondensation index (SDI). However, at 10 min, it had a significant effect on both DFI and SDI. The analysis of our results showed that this plant oil rich in terpenoids and phenolic antioxidants could be a quite good in vitro additive with high potential for the world of medically assisted reproduction

Cloning and heterologous overexpression of three gap genes encoding different glyceraldehyde-3-phosphate dehydrogenases from the plant pathogenic bacterium Pseudomonas syringae pv. tomato strain DC3000

The gammaproteobacterium Pseudomonas syringae pv. tomato DC3000 is the causal agent of bacterial speck, a common disease of tomato. The mode of infection of this pathogen is not well understood, but according to molecular biological, genomic and proteomic data it produces a number of proteins that may promote infection and draw nutrients from the plant. Glyceraldehyde-3- phosphate dehydrogenase (GAPDH) is a major enzyme of carbon metabolism that was reported to be a surface antigen and virulence factor in other pathogenic microorganisms, but its possible role in the infection process of P. syringae has so far not been studied. Whole-genome sequence analyses revealed the occurrence in this phytopathogenic bacterium of three paralogous gap genes encoding distinct GAPDHs, namely two class I enzymes having different molecular mass subunits and one class III bifunctional D-erythrose-4-phosphate dehydrogenase/GAPDH enzyme. By using genome bioinformatics data, as well as alignments of both DNA and deduced protein sequences, the three gap genes of P. syringae were one-step cloned with a His-Tag in pET21a vector using a PCR-based strategy, and its expression optimized in E. coli BL21 to achieve high yield of the heterologous proteins. In accordance with their distinct molecular phylogenies, these bacterial gap genes encode functional GAPDHs of diverse molecular masses and nicotinamide-coenzyme specificities, suggesting specific metabolic and/or cellular roles.Peer Reviewe

Sulphurous mineral water oral therapy: effects on erythrocyte metabolism

Sulphurous mineral water oral therapy: Effects on erythrocyte metabolism Modern medical research appears to confirm the validity of the use of sulphurous spring waters as therapeutic and preventive remedies for a large variety of diseases affecting the respiratory tract, skin, liver, intestine, gynaecological apparatus and osteoarticular system. The ingestion of water containing hydrogen sulphide (H2S) is common in spring sulphurous mineral water (SMW) therapy. We hypothesized that observed detrimental effects are related to the alteration of erythrocytes metabolism caused by H2S. To verify our hypothesis, we treated 20 healthy volunteers with SMW and evidenced an increase of methemoglobin concentration, an inhibition of both erythrocyte glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and glucose-6-phosphate dehydrogenase (G6PDH) activities. To investigate the mechanism of H2S effect on GAPDH activity, an in vitro study was performer by incubating both erythrocytes from 12 healthy volunteers and purified GAPDH with buffered [35S]- H2S labelled sulphurous water. The interaction between H2S and NAD+(H), was also investigated. The results indicate that a direct reaction between GAPDH and H2S does not occur and the observed decrease of GAPDH activity is to ascribe to the reaction between H2S and NAD+(H). This may lead to GAPDH inhibition by two ways, namely (i) cellular NAD+(H) reduced availability and (ii) catalytic site blockage. In conclusion, our results show that among the detrimental effects of SMW administration are erythrocyte GAPDH and G6PDH activity inhibition and increased methemoglobin concentration. A mechanism to explain the occurrence of these biochemical events is also proposed. These data can also give supplemental information on the adverse events and, at least in part, explain the clinical symptoms observed. Furthermore, it appears possible to use the assay of erythrocyte GAPDH to monitor the risk to undergo adverse events

The Effect of Supplementation with Some Essential Oils on the Mobility and the Vitality of Human Sperm

The objective of this work is to study the improvement effect of some essential oils of sage (Salvia officinalis), oregano (Origanum vulgare), and eucalyptus (eucalyptus globulus) on the physiological parameters characterizing the quality of human sperm (mobility and vitality). We find natural biomolecules to improve sperm quality to increase the chances of success of very low in vitro fertilization (IVF) that stagnate around 20%. Sperm samples were mixed with different concentrations of essential oils. The effect of these essential oils on the motility and vitality of spermatozoa has been analyzed. The mobility was determined using a Computer Assisted Sperm Analysis (CASA). In the other side, the evaluation of sperm vitality was performed by staining eosin 2% and the microscopic examination is carried out via optical microscope. A drop of sperm will be mixed with a drop of eosin solution 2%, spread between the slip and coverslip, then allowed to air dry, and examined under a microscope. A significant improvement in the mobility and vitality of human spermatozoa has been noted with oregano. Eucalyptus after 10 min of exposure also significantly improves the mobility and vitality of the spermatozoa. Sage does not improve mobility for these incubation times but significantly improves vitality

Structural, Kinetic, and Pharmacodynamic Mechanisms of d-Amino Acid Oxidase Inhibition by Small Molecules.

We characterized the mechanism and pharmacodynamics of five structurally distinct inhibitors of d-amino acid oxidase. All inhibitors bound the oxidized form of human enzyme with affinity slightly higher than that of benzoate (Kd 48 2-4 \u3bcM). Stopped-flow experiments showed that pyrrole-based inhibitors possessed high affinity (Kd 48 100-200 nM) and slow release kinetics (k < 0.01 s(-1)) in the presence of substrate, while inhibitors with pendent aromatic groups altered conformations of the active site lid, as evidenced by X-ray crystallography, and showed slower kinetics of association. Rigid bioisosteres of benzoic acid induced a closed-lid conformation, had slower release in the presence of substrate, and were more potent than benzoate. Steady-state d-serine concentrations were described in a PK/PD model, and competition for d-serine sites on NMDA receptors was demonstrated in vivo. DAAO inhibition increased the spatiotemporal influence of glial-derived d-serine, suggesting localized effects on neuronal circuits where DAAO can exert a neuromodulatory role