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Biomedical subjects

C Rouquette

Publications and source records attributed to C Rouquette.

At least 19 recordsLinked to original sources
PubMed2001

Genetic organization and regulation of antimicrobial efflux systems possessed by Neisseria gonorrhoeae and Neisseria meningitidis.

Efflux pumps can make a significant contribution to the capacity of bacteria to resist the action of antibiotics. Certain efflux pumps also recognize antimicrobial agents that are present in their respective hosts and their ability to export toxic agents could enhance bacterial survival during infection prior to appearance of cellular or humoral host defensive systems. This review is concerned with the principal efflux pumps possessed by two closely related strict human pathogens, Neisseria gonorrhoeae and Neisseria meningitidis. Specific emphasis is placed on the organization of the structural genes encoding the mtr and far efflux pumps, the substrates (often host-derived) recognized by these pumps, and the cis- and trans-acting transcriptional factors that regulate efflux pump gene expression in gonococci and meningococci. The overriding theme of this review is that the efflux pumps possessed by these pathogens likely contribute to their pathogenic mechanisms by providing a means to escape a number of antimicrobial compounds that bathe mucosal surfaces.

Anti-Bacterial Agents↗
PubMed1999

Induction of the mtrCDE-encoded efflux pump system of Neisseria gonorrhoeae requires MtrA, an AraC-like protein.

The mtr (multiple transferable resistance) gene complex in Neisseria gonorrhoeae encodes an energy-dependent efflux pump composed of the MtrC-MtrD-MtrE cell envelope proteins that serves to export structurally diverse antimicrobial, hydrophobic agents (HAs). Many of these agents have membrane-acting detergent activity. Using Triton X-100 (TX-100) as a representative HA, we found that the mtrCDE efflux pump operon could be induced to higher levels of expression when an HA-sensitive strain was exposed to sublethal concentrations of this non-ionic detergent and the structurally related spermicide, nonoxynol-9. This induction was at the level of mtrCDE gene transcription and was independent of the MtrR repressor, which normally decreases mtrCDE gene expression. However, the enhanced resistance of gonococci to TX-100 was dependent on the expression of a previously undescribed gonococcal protein that belonged to the AraC/XylS family of transcriptional activators. We have termed this protein MtrA to signify its likely role in the activation of mtrCDE gene expression. Taken together with previous studies dealing with the genetic control of mtrCDE gene expression, we propose that gonococci can modulate their resistance to HAs through both positive and negative transcriptional control processes. The action of these regulatory processes is probably of importance in determining the survival capacity of gonococci at mucosal surfaces that contain detergent-like HAs.

ATP-Binding Cassette Transporters↗
PubMed1998

The ClpC ATPase of Listeria monocytogenes is a general stress protein required for virulence and promoting early bacterial escape from the phagosome of macrophages.

Under stress conditions, the facultative intracellular pathogen Listeria monocytogenes produces a ClpC ATPase, which is a general stress protein encoded by clpC and belonging to the HSP-100/Clp family. A ClpC-deficient mutant was obtained by gene disruption in strain LO28, which became highly susceptible to stress conditions in vitro. Intracellular growth of this mutant was restricted within macrophages, one of the major target cells of L. monocytogenes, during the infectious process. A quantitative electron microscope study showed that, contrary to wild-type bacteria that rapidly gain access to the cytoplasm of macrophages, mutant bacteria remained confined to membrane-bound phagosomes. Only a few mutant bacteria disrupted the phagosome membrane after 4h of incubation, then polymerized actin filaments and multiplied within the cytoplasm. The ClpC ATPase, therefore, promotes early bacterial escape from the phagosome of macrophages, thus enhancing intracellular survival. The ClpC ATPase was produced in vivo during experimental infection by wild-type bacteria. The virulence of the ClpC-deficient mutant was severely attenuated in mice, with a three-log decrease in its 50% lethal dose compared with wild-type bacteria. Bacterial growth of mutant bacteria was strongly restricted in organs, presumably because of an impairment of intracellular survival in host tissues. Our results provide evidence that a general stress protein is required for the virulence of L. monocytogenes, which behaves as a virulence factor promoting intracellular survival of this pathogen.

Adenosine Triphosphatases↗
PubMed1997

Genetic basis of the MbrC "ploidy" phenotype in Escherichia coli.

The mbrC17 mutation in Escherichia coli had been shown to cause conditional growth defects and an increase in the quantity of DNA per cell. The present work was aimed at identifying the mutation. Sequencing showed that the MbrC17 phenotype does not involve glr (murI), as previously suggested. P1 transduction data indicated that the mbrC17 mutation is closely linked to rpoB, and allele exchange showed it to lie within the secE-nusG operon. A single change relative to wild type was found in the secE-nusG region from the mbrC17 strain, a G-->A mutation 23 bp upstream of the secE coding sequence. This mutation causes a two-fold increase in the concentration of secE-nusG mRNA.

Amino Acid Isomerases↗
PubMed1996

Identification of a ClpC ATPase required for stress tolerance and in vivo survival of Listeria monocytogenes.

We identified a new chromosomal locus involved in the virulence of the facultative intracellular pathogen Listeria monocytogenes. This locus displays the same genetic organization as that of the clpC/mecB locus of Bacillus subtilis. It contains a thermoregulated operon of four genes, whose transcription is upregulated at 42 degrees C. The last gene of this operon is clpC, which encodes a protein of 826 amino acid residues, identified as a ClpC ATPase, sharing a strong peptide sequence identity (78%) with ClpC/MecB of B. subtilis. Tn917 insertions inactivating the entire operon, or only clpC, gave mutants highly susceptible to stress, including iron limitation, elevated temperatures and high osmolarity. The virulence of these mutants was severely impaired in the mouse. A clpC insertional mutant was also restricted in its capacity to grow in bone-marrow-derived macrophages. These results demonstrate that the ClpC ATPase of L. monocytogenes is a general stress protein involved in intracellular growth and in vivo survival of this pathogen in host tissues.

Adenosine Triphosphatases↗
PubMed1996

The pathogenesis of infection by Listeria monocytogenes.

Listeria monocytogenes is a Gram-positive bacterium responsible for severe infections in human and a large variety of animal species. It is a facultative intracellular pathogen which invades macrophages and most tissue cells of infected hosts where it can proliferate. The molecular basis of this intracellular parasitism has been to a large extent elucidated. The virulence factors, including internalin, listeriolysin O, phospholipases and a bacterial surface protein, ActA, are encoded by chromosomal genes organised in operons. Following internalisation into host cells, the bacteria escape from the phagosomal compartment and enter the cytoplasm. They then spread from cell to cell by a process involving actin polymerisation. In infected hosts, the bacteria cross the intestinal wall at Peyer's patches to invade the mesenteric lymph nodes and the blood. The main target organ is the liver, where the bacteria multiply inside hepatocytes. Early recruitment of polymorphonuclear cells lead to hepatocyte lysis, and thereby bacterial release. This causes prolonged septicaemia, particularly in immunocompromised hosts, thus exposing the placenta and brain to infection. The prognosis of listeriosis depends on the severity of meningoencephalitis, due to the elective location of foci of infection in the brain stem (rhombencephalitis). Despite bactericidal antibiotic therapy, the overall mortality is still high (25 to 30%).

Actins↗
PubMed1995

An iron-dependent mutant of Listeria monocytogenes of attenuated virulence.

A bank of Tn917-insertional mutants from the facultative intracellular pathogen Listeria monocytogenes was screened by an original method based on bacterial growth on synthetic medium under iron-limiting conditions. One mutant, whose in vitro growth in synthetic medium was specifically dependent upon the availability of iron in its environment, was isolated and characterized. The insertional event occurred in a non-coding region, upstream of a rrn operon and located within a 1100-kb NotI fragment of the physical map, where the virulence genes already identified in L. monocytogenes were also present. Protein analysis by SDS-PAGE revealed a pleiotropic effect of the insertional event on cell-associated proteins, suggesting a polar effect of the transposon on adjacent unknown gene(s). The virulence in the mouse of this mutant was strongly impaired, although it was capable in vitro of growing intracellularly and of spreading from cell to cell, as shown by the production of lytic plaques on cell culture.

Animals↗
PubMed1986

[A rapid method for the determination of cyclosporin A in blood by high performance liquid chromatography].

The purpose of this paper is to propose a method for the determination of cyclosporin in whole blood by means of HPLC with UV detection. After a one step extraction into ether, the organic layer is washed NaOH and evaporated to dryness. The residue is reconstituted in a mixture of acetonitrile and water (40/60) (V/V) and injected into the chromatograph. The C8 reversed phase column is warming at 75 degrees C temperature. The mobile phase consists of acetonitrile and water (60/40) (V/V) and the wavelength is set at 210 nm. The technique is rapid, sensitive, reproducible and well suited for therapeutic monitoring and for pharmacokinetic studies.

Chromatography, High Pressure Liquid↗
PubMed1985

[Plasma assay of haloperidol by high performance liquid chromatography].

Haloperidol is extracted with hexane-isoamyl alcohol in the presence of two internal standards, chlorohaloperidol and spiroperidol. For chromatographic separation, a C18 reversed-phase column and a mobile phase of acetonitrile/methanol/phosphate buffer are used. Detection is by UV absorption spectrophotometry at 245 nm, and the limit of detection was set to 1 ng/ml. To study hydroxyhaloperidol, a metabolite of haloperidol which has much less UV-absorption than haloperidol, detection must be seat 220 nm.

Chromatography, High Pressure Liquid↗