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Biomedical subjects

G Prévost

Publications and source records attributed to G Prévost.

At least 19 recordsLinked to original sources
PubMed2006

[Incidence of meticillin-resistant Staphylococcus aureus in neonatal care unit of departmental hospital centre of Zou Collines in Benin].

UNLABELLED: Meticillin resistant Staphylococcus aureus (MRSA) is among the most important causes of nosocomial infections. It possesses a particular ability to spread in hospitals worldwide. OBJECTIVE: To analyze the proportion of MRSA among S. aureus isolated from specimen taken for diagnosis purposes. To make the medical staff aware of the problem of MRSA infections and to take a better care of patients. PATIENTS AND METHODS: During 3 months, a prospective study was carried out in the neonatal unit of centre hospitalier départemental du Zou et Collines in Benin. We identified newborn carriers of SA, particularly MRSA and factors associated with the carriage. Two hundred and ninety patients were admitted in the 3 divisions of the neonatal unit. From 195 specimens examined for diagnosis purposes 48 h after hospitalization, 112 patients were detected by nose swabbing. Concurrently, swabbing of environment was achieved. RESULTS: Among patients'specimens, 141 isolations of S. aureus were observed. The proportion of MRSA was 36% amongst diagnostic specimens. MRSA represented 39% of the environment specimens. None of the isolated MRSA produces Panton Valentine leukocidin. CONCLUSION: Our survey revealed a high level of MRSA among S. aureus isolated from diagnostic specimens. Consecutive to such findings and for decreasing nosocomial infection, an appropriate prevention program was installed, including intensive promotion of hands hygiene, correct sterilization and disinfection of materials and patients.

Benin↗
PubMed2006

Site-directed mutagenesis to assess the binding capacity of class s protein of Staphylococcus aureus leucotoxins to the surface of polymorphonuclear cells.

Staphylococcal leucotoxins result from the association of class S components and class F component inducing the activation and the permeabilization of the target cells. Like alpha-toxin, the leucotoxins are pore-forming toxins with more than 70% beta-sheet. This was confirmed by attenuated total reflectance Fourier transform infrared (ATR-FTIR) spectroscopy. In addition, threonine 28 of a predicted and conserved beta-sheet at the N-terminal extremity of class S proteins composing leucotoxins aligns with histidine 35 of alpha-toxin, which has a key role in oligomerization of the final pore. Flow cytometry was used to study different aminoacid substitutions of the threonine 28 in order to evaluate its role in the biological activity of these class S proteins. Finally, results show that threonine 28 of the leucotoxin probably plays a role similar to that of histidine 35 of alpha-toxin. Mutations on this threonin largely influenced the secondary interaction of the class F component and led to inactive toxin.

Journal Article↗
PubMed2005

[Education for chronic cardiologic diseases in a transversal multidisciplinary unit: the experience of a general hospital center].

Treatments for congestive heart failure, hypertension and cardiovascular risk have significantly changed and have become more complex. The have also become more and more effective thanks to the results of great clinical studies that have enabled European and North-American societies to issues recommendations. The observance of the pharmacological and non-pharmacological treatments requires the education of patients and their family following guidelines that have been clearly defined by the European Society of Cardiology. This education, in which the technic of communication is very important, is common to a lot of chronic diseases and requires adequate material and human resources in order to have an optimal quality of treatment. In a society in which spending is on rise, getting such resources is not easy. However, putting in common resources of several departments can be a good solution. The experience of the Hospital Center of Douai (France) lead to the creation of a Transversal Education Unit at the end of the year 2003. This unit centralizes the efforts of several departments of care like pneumology, pediatrics, diabetology, nutrition and cardiology and allows patients suffering from co-morbidities to have access to various programs of this unit.

Cardiac Rehabilitation↗
PubMed2005

Control of cardiovascular risk factors in patients with type 2 diabetes and hypertension in a French national study (Phenomen).

UNLABELLED: For some years now, the recommendations of scientific societies have significantly reduced the therapeutic targets for blood pressure, glycaemia and lipid levels in diabetic patients. However, little is known regarding the synchronization between effective risk factor management and the guidelines. To examine this issue, the Phenomen survey was conducted between January and July 2001 on 16358 patients suffering from hypertension followed by a general practitioner in France. AIM: To evaluate the control of cardiovascular risk factors in patients with diabetes and hypertension according to the French guidelines. METHODS AND PATIENTS: 8177 general practitioners, selected from a national database according to quotas, taking into account age, practice and area, had to include the first two hypertensive patients they came across in their practice and to collect their demographic data, cardiovascular risk factors and medications. RESULTS: 2346 out of 16358 hypertensive patients presented with type 2 diabetes (14.3% of the cohort). The number of GP consultations in the last 12 months averaged 8.31. According to the French guidelines, 6.5% had a blood pressure<140/80 mmHg, a total of 38.7% patients met the goal of LDL cholesterol level and 26.6% of patients had an HbA1c<6.5%, 53.4% of patients had an HbA1c between 6.6 and 8%. 37.1% of patients continued to receive antihypertensive monotherapy but only 3% in this monotherapy group reached the target of 130/85 mmHg. 29% of the patients were on antiplatelet therapy. 64.6% of these hypertensive diabetic patients presented with more than three other cardiovascular risk factors. Based on WHO recommendations, 0.3% of the patients met all of the blood pressure, lipid and glycaemic treatment objectives. CONCLUSION: Despite frequent monitoring by a general practitioner, the overall management of modifiable risk factors in this diabetic hypertensive population is clearly inadequate. The impact of the guidelines on effective management remains limited and additional information is required to understand why physicians are not more aggressive in managing modifiable risk factors in diabetic patients.

Aged↗
PubMed2004

Identification of an aspartylglucosaminidase-like protein in the venom of the parasitic wasp Asobara tabida (Hymenoptera: Braconidae).

This study was designed to identify one of the main components of venomous secretions of the endoparasitic wasp Asobara tabida. By using electrophoretic methods, partial amino acid sequencing and immunostaining, we demonstrated the presence of an aspartylglucosaminidase (AGA)-like protein in the venom of this insect. The enzyme had a polymeric conformation and was formed of 30 and 18 kDa subunits. The relative positions of several amino acids involved in substrate binding and catalytic activity of known AGA-proteins, which are usually lysosomal enzymes, were conserved in the NH(2)-terminal ends of these subunits. Antibodies raised against human AGA recognized the two subunits of the protein and a 44 kDa protein, suggesting the presence of a precursor molecule of the enzyme in the venom. However, no reliable measurement of the AGA activity could be performed on the venom extracts, which could be explained by the fact the enzyme would be stored in the reservoir of the venom apparatus under an inactive form. These results constitute the first description of an AGA-like protein in an insect venom and are discussed with respect to the knowledge acquired on lysosomal and venom enzymes.

Amino Acid Sequence↗
PubMed2004

[Virulence factors of clinical Aeromonas caviae isolates].

Aeromonas caviae, an ubiquitous aquatic organism, has long been considered to be of low pathogenicity, and its virulence mechanisms are still not clearly understood. Twenty-eight A. caviae isolates of clinical origin, most often monomicrobic, were identified in our university hospital over a four year period. Patients, mostly immunocompromised, were: eight diarrhoeal infants, 13 diarrhoeal adults, seven bacteraemic adults. Adults were frequently suffering from underlying intestinal malignancy, hepatobiliary disease, gastrectomy. Virulence factors were investigated. Adherence, studied by use of tissue culture HEp-2 cells, and staining of characteristic lateral flagella, were observed in diarrhoeal strains. Extracellular hemolytic activity was tested on rabbit erythrocytes suspensions at 25 and 37 degrees C. One blood culture isolate showed an important hemolytic activity at 25 degrees C, but none at 37 degrees C. Treatment with furin activated the aerolysin precursor and resulted in significant hemolysis at 37 degrees C, and fluid accumulation in rabbit ileal loops similar to that of A. hydrophila as control. The presence of the hemolysin gene was confirmed in this strain by PCR. In conclusion, A. caviae was shown to be a pathogen isolated from diarrhoea and bacteraemia in immunocompromised patients with malignancies and low gastric acidity as favouring factors. Virulence including the ability to adhere to cells and the production of lateral flagella was observed in diarrhoeal strains. The expression and the production of extracellular hemolytic activity and enterotoxicity at 37 degrees C depended on the activation of the pore forming toxin aerolysin precursor by furin. In vivo the protoxin is probably processed to its mature form by host proteases.

Adult↗
PubMed2003

Ion channels and bacterial infection: the case of beta-barrel pore-forming protein toxins of Staphylococcus aureus.

Staphylococcus aureus strains causing human pathologies produce several toxins, including a pore-forming protein family formed by the single-component alpha-hemolysin and the bicomponent leukocidins and gamma-hemolysins. The last comprise two protein elements, S and F, that co-operatively form the active toxin. alpha-Hemolysin is always expressed by S. aureus strains, whereas bicomponent leukotoxins are more specifically involved in a few diseases. X-ray crystallography of the alpha-hemolysin pore has shown it is a mushroom-shaped, hollow heptamer, almost entirely consisting of beta-structure. Monomeric F subunits have a very similar core structure, except for the transmembrane stem domain which has to refold during pore formation. Large deletions in this domain abolished activity, whereas shorter deletions sometimes improved it, possibly by removing some of the interactions stabilizing the folded structure. Even before stem extension is completed, the formation of an oligomeric pre-pore can trigger Ca(2+)-mediated activation of some white cells, initiating an inflammatory response. Within the bicomponent toxins, gamma-hemolysins define three proteins (HlgA, HlgB, HlgC) that can generate two toxins: HlgA+HlgB and HlgC+HlgB. Like alpha-hemolysin they form pores in planar bilayers with similar conductance, but opposite selectivity (cation instead of anion) for the presence of negative charges in the ion pathway. gamma-Hemolysin pores seem to be organized as alpha-hemolysin, but should contain an even number of each component, alternating in a 1:1 stoichiometry.

Animals↗
PubMed2002

Retrieving biological activity from LukF-PV mutants combined with different S components implies compatibility between the stem domains of these staphylococcal bicomponent leucotoxins.

Bicomponent leucotoxins, such as Panton-Valentine leucocidin, are composed of two classes of proteins, a class S protein such as LukS-PV, which bears the cell membrane binding function, and a class F protein such as LukF-PV, which interacts to form a bipartite hexameric pore. These leucotoxins induce cell activation, linked to a Ca(2+) influx, and pore formation as two consecutive and independently inhibitable events. Knowledge of the LukF-PV monomer structure has indicated that the stem domain is folded into three antiparallel beta-strands in the water-soluble form and has to refold into a transmembrane beta-hairpin during pore formation. To investigate the requirements for the cooperative assembly of the stems of the S and F components to produce biological activity, we introduced multiple deletions or single point mutations into the stem domains of LukF-PV and HlgB. While the binding of the mutated proteins was weakly dependent on these changes, Ca(2+) influx and pore formation were affected differently, confirming that they are independent events. Ca(2+) entry into human polymorphonuclear cells requires oligomerization and may follow the formation of a prepore. The activity of some of the LukF-PV mutants, carrying the shorter deletions, was actually improved. This demonstrated that a crucial event in the action of these toxins is the transition of the prefolded stem into the extended beta-hairpins and that this step may be facilitated by small deletions that remove some of the interactions stabilizing the folded structure.

Amino Acid Sequence↗
PubMed2001

Mode of action of beta-barrel pore-forming toxins of the staphylococcal alpha-hemolysin family.

Staphylococcal alpha-hemolysin is the prototype of a family of bacterial exotoxins with membrane-damaging function, which share sequence and structure homology. These toxins are secreted in a soluble form which finally converts into a transmembrane pore by assembling an oligomeric beta-barrel, with hydrophobic residues facing the lipids and hydrophilic residues facing the lumen of the channel. Besides alpha-hemolysin the family includes other single chain toxins forming homo-oligomers, e.g. beta-toxin of Clostridium perfringens, hemolysin II and cytotoxin K of Bacillus cereus, but also the staphylococcal bi-component toxins, like gamma-hemolysins and leucocidins, which are only active as the combination of two similar proteins which form hetero-oligomers. The molecular basis of membrane insertion has become clearer after the determination of the crystal structure of both the oligomeric pore and the soluble monomer. Studies on this family of beta-barrel pore-forming toxins are important for many aspects: (i) they are involved in serious pathologies of humans and farmed animals, (ii) they are a good model system to investigate protein-membrane interaction and (iii) they are the basic elements for the construction of nanopores with biotechnological applications in various fields.

Amino Acid Sequence↗
PubMed2001

Flow cytometric determination of Panton-Valentine leucocidin S component binding.

The binding of the S component (LukS-PV) from the bicomponent staphylococcal Panton-Valentine leucocidin to human polymorphonuclear neutrophils (PMNs) and monocytes was determined using flow cytometry and a single-cysteine substitution mutant of LukS-PV. The mutant was engineered by replacing a glycine at position 10 with a cysteine and was labeled with a fluorescein moiety. The biological activity of the mutant was identical to that of the native protein. It has been shown that LukS-PV has a high affinity for PMNs (Kd = 0.07 +/- 0.02 nM, n = 5) and monocytes (Kd = 0.020 +/- 0.003 nM, n = 3) with maximal binding capacities of 197,000 and 80,000 LukS-PV molecules per cell, respectively. The nonspecifically bound molecules of LukS-PV do not form pores in the presence of the F component (LukF-PV) of leucocidin. LukS-PV and HlgC share the same receptor on PMNs, but the S components of other staphylococcal leukotoxins, HlgA, LukE, and LukM, do not compete with LukS-PV for its receptor. Extracellular Ca2+ at physiological concentrations (1 to 2 nM) has only a slight influence on the LukS-PV binding, in contrast to its complete inhibition by Zn2+. The down-regulation by phorbol 12-myristate 13-acetate (PMA) of the binding of LukS-PV was blocked by staurosporine, suggesting that the regulatory effect of PMA depends on protein kinase C activation. The labeled mutant form of LukS-PV has proved very useful for detailed binding studies of circulating white cells by flow cytometry. LukS-PV possesses a high specific affinity for a unique receptor on PMNs and monocytes.

Bacterial Toxins↗
PubMed2001

Staphylococcus aureus isolated in cases of impetigo produces both epidermolysin A or B and LukE-LukD in 78% of 131 retrospective and prospective cases.

Clinical symptoms of impetigo and staphylococcal scalded skin syndrome may not only be expressed as the splitting of cell layers within the epidermis but are often accompanied by some localized inflammation. Toxin patterns of Staphylococcus aureus isolates originating from patients with impetigo and also from those with other primary and secondary skin infections in a retrospective isolate collection in France and a prospective isolate collection in French Guiana revealed a significant association (75% of the cases studied) of impetigo with production of at least one of the epidermolysins A and B and the bicomponent leucotoxin LukE-LukD (P < 0.001). However, most of the isolates were able to produce one of the nonubiquitous enterotoxins. Pulsed-field gel electrophoresis (PFGE) of genomic DNA hydrolyzed with SmaI showed a polymorphism of the two groups of isolates despite the fact that endemic clones were suspected in French Guiana and France. The combination of toxin patterns with PFGE fingerprinting may provide further discrimination among isolates defined in a given cluster or a given pulsotype and account for a specific virulence. The new association of toxins with a clinical syndrome may reveal principles of the pathological process.

Adolescent↗
PubMed2000

Racing against host's immunity defenses: a likely strategy for passive evasion of encapsulation in Asobara tabida parasitoids.

The hymenopteran Asobara tabida Nees (Braconidae, Alysiinae) develops as a solitary endophagous parasite in larvae of several Drosophila species. Most A. tabida eggs possess a sticky chorion which attaches to the tissue of the host organs within a few hours following oviposition. A. tabida sticky eggs usually avoid encapsulation, though the probability of survival decreases in hosts carrying a larger number of circulating hemocytes. Here, we hypothesized that the elicitation of the encapsulation reaction may result from a race between two phenomena: the host's hemocytic reaction and the embedment of the parasitic egg within the host tissues. In order to test this hypothesis, we measured the speed of capsule formation in D. melanogaster larvae of different ages, knowing that the number of circulating hemocytes increases with the age of the larvae. Using a non-virulent A. tabida strain, the eggs of which do not attach to the host tissue, we found that the speed of capsule formation increased correlatively with the age of the D. melanogaster larva. Therefore, the hypothesis of a physiological race between host's immunity defenses and parasite's avoidance of host's defenses is strongly supported by our results. Also, A. tabida eggs which attach to the host's tissue before the attack by the hemocytes has taken place may be considered as a strategy of passive evasion from encapsulation.

Journal Article↗
PubMed2000

Crystal structure of the F component of the Panton-Valentine leucocidin.

Leucocidins and gamma-hemolysins are bi-component staphylococcal toxins that form lytic transmembrane pores. Their cytotoxic activities involve the synergistic association of a class S and a class F component, produced as water-soluble monomers which assemble on the surface of specific cells. The structure of the F protein from Panton-Valentine leucocidin, solved at 2.0 A resolution, and sequence alignment suggest that it represents the fold of any secreted protein in this family of toxins. The comparison of this structure to that of the homoheptameric alpha-hemolysin provides some insights into the molecular events that may occur during pore formation.

Bacterial Toxins↗
PubMed2000

Variable expressions of Staphylococcus aureus bicomponent leucotoxins semiquantified by competitive reverse transcription-PCR.

A competitive reverse transcription-PCR method was developed for the semiquantitation of the expression of genes encoding bicomponent leucotoxins of Staphylococcus aureus, e.g., Panton-Valentine leucocidin (lukPV), gamma-hemolysin (hlgA and hlgCB), and LukE-LukD (lukED). The optimization procedure included RNA preparation; reverse transcription; the use of various amounts of enzymes, antisense primer, and RNA; and the final amplification chain reaction. Reproducible results were obtained, with sensitivity for detection of cDNA within the range of 1 mRNA/10(4) CFU to 10(2) mRNA/CFU, depending on the gene. Both specific mRNAs were more significantly expressed at the late-exponential phase of growth. Expression was about 100-fold higher in yeast extract-Casamino Acids-pyruvate medium than in heart infusion medium. Expression of the widely distributed gamma-hemolysin locus in the NTCC 8178 strain was around 10-fold diminished compared with that in the ATCC 49775 strain. Because of the lower level of hlgA expression, the corresponding protein, which is generally not abundant in culture supernatant, should be investigated for its contribution to the leucotoxin-associated virulence. The agr, sar, and agr sar mutant strains revealed a great dependence with regard to leucotoxin expression on the global regulatory system in S. aureus, except that expression of hlgA was not affected in the agr mutant.

Bacterial Proteins↗
PubMed1999

Discoupling the Ca(2+)-activation from the pore-forming function of the bi-component Panton-Valentine leucocidin in human PMNs.

The consecutive cell activation, including Ca(2+)-channel opening, and pore formation leading to human neutrophil lysis were the two functions of the staphylococcal Panton-Valentine leucocidin attempted to be discoupled by site-directed mutagenesis. In a first approach consisting in deletions of the cytoplasmic extremity of the transmembranous domain, we produced a LukF-PV DeltaSer125-Leu128 with a slightly reduced Ca(2+) induction but with a significantly lowered lytic activity when combined with its synergistic protein LukS-PV. The second approach consisted in the modification of charges and/or introduction of a steric hindrance inside the pore, which also led to interesting mutated proteins: LukF-PV G131D, G131W and G130D. The latter had an intact Ca(2+) induction ability while the lytic one was 20-fold diminished. Binding properties and intrinsic pore diameters of these discoupled toxins remained comparable to the wild-type protein. The mutated proteins promoted interleukin-8 secretion, but they were rather inactive in an experimental model. New insights are brought concerning the role of the two functions in the virulence of this bi-component leucotoxin.

Amino Acid Substitution↗
PubMed1999

The structure of a Staphylococcus aureus leucocidin component (LukF-PV) reveals the fold of the water-soluble species of a family of transmembrane pore-forming toxins.

BACKGROUND: Leucocidins and gamma-hemolysins are bi-component toxins secreted by Staphylococcus aureus. These toxins activate responses of specific cells and form lethal transmembrane pores. Their leucotoxic and hemolytic activities involve the sequential binding and the synergistic association of a class S and a class F component, which form hetero-oligomeric complexes. The components of each protein class are produced as non-associated, water-soluble proteins that undergo conformational changes and oligomerization after recognition of their cell targets. RESULTS: The crystal structure of the monomeric water-soluble form of the F component of Panton-Valentine leucocidin (LukF-PV) has been solved by the multiwavelength anomalous dispersion (MAD) method and refined at 2.0 A resolution. The core of this three-domain protein is similar to that of alpha-hemolysin, but significant differences occur in regions that may be involved in the mechanism of pore formation. The glycine-rich stem, which undergoes a major rearrangement in this process, forms an additional domain in LukF-PV. The fold of this domain is similar to that of the neurotoxins and cardiotoxins from snake venom. CONCLUSIONS: The structure analysis and a multiple sequence alignment of all toxic components, suggest that LukF-PV represents the fold of any water-soluble secreted protein in this family of transmembrane pore-forming toxins. The comparison of the structures of LukF-PV and alpha-hemolysin provides some insights into the mechanism of transmembrane pore formation for the bi-component toxins, which may diverge from that of the alpha-hemolysin heptamer.

Amino Acid Sequence↗