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ABSTRACT: Clinical isolates of Neisseria meningitidis from cases of meningococcal disease, collected between January 2000 and December 2004, were identified and typed at the French National Reference Centre. A representative subset of 546 isolates from among 2882 isolates was further genotyped by multilocus sequence typing to determine their genetic lineages (clonal complexes) and the degree of diversification among different clonal complexes. Representative isolates of the main clonal complexes were tested for their virulence in mice and for proapoptotic effects on human epithelial cells. High genetic diversity in some genetic lineages (ST-32 and ST-41/44) was correlated with heterogeneity in virulence in mice and proapoptotic effects on human epithelial cells. In contrast, the homogeneous genetic structure of isolates of the ST-11 clonal complex, regardless of their serogroup, correlated positively with a fatal outcome of the infection, increased virulence in mice and increased proapoptotic effects on human epithelial cells.
Clinical Microbiology and Infection 06/2008; 14(5):467-72. · 4.54 Impact Factor
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ABSTRACT: Invasive serogroup W135 Neisseria meningitidis strains were isolated in Portugal between 1995 and 2002. Nine of 12 isolates showed "Hajj-2000"-associated phenotypes and belonged to the ST-11/ET-37 clonal complex. Pulsed-field gel electrophoresis clustered the ST-11/ET-37 isolates together with the "Hajj-2000" strain although the profiles were distinguishable.
Pathologie Biologie 04/2008; 56(2):94-6. · 1.53 Impact Factor
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ABSTRACT: PilC1, a pilus-associated protein in Neisseria menin- gitidis, is a key element in initial meningococcal adhesion to target cells. A promoter element (CREN, contact regulatory element of Neisseria) is responsible for the transient induction of this gene upon cell contact. crgA (contact-regulated gene A) encodes a transcriptional regulator whose expression is also induced upon cell contact from a promoter region similar to the CREN of pilC1. CrgA shows significant sequence homologies to LysR-type transcriptional regulators. Its inactivation in meningococci provokes a dramatic reduction in bacterial adhesion to epithelial cells. Moreover, this mutant is unable to undergo intimate adhesion to epithelial cells or to provoke effacing of microvilli on infected cells. Purified CrgA is able to bind to pilC1 and crgA promoters, and CrgA seems to repress the expression of pilC1 and crgA. Our results support a dynamic model of bacteria-cell interaction involving a network of regulators acting in cascade. CrgA could be an intermediate regulator in such a network.
The EMBO Journal 04/2000; 19(5):1068-78. · 9.20 Impact Factor
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ABSTRACT: Pilus-mediated adherence makes an essential contribution to the pathogenesis of Neisseria meningitidis by allowing the initial localized adherence. Pili are assembled from a protein subunit called pilin. Two proteins, PilC1 and PilC2, are also key elements in the formation of pili as the production of at least one PilC protein is required for pilus assembly. In addition, PilC1 but not PilC2 modulates adhesiveness, most probably by being the adhesin. Recently, both genes have been demonstrated to be controlled by different promoters, pilC2 is expressed from a single transcription starting point (TSP), whereas pilC1 has three TSPs. One of these, PC1.1, corresponds to the unique TSP of pilC2, and two others, PC1.2 and PC1.3, are located in a region upstream of pilC1 but not pilC2. This suggests that both genes may be under the control of separate regulatory pathways. In this work, by engineering pilC1-lacZ and pilC2-lacZ transcriptional fusions, we provide evidence that expression of pilC1, but not that of pilC2, is transiently induced by bacterial cell contact. This induction required viable cells, did not need the presence of pili and relied on the expression of pilC1 from PC1.3. Destruction of this TSP by site-directed mutagenesis did not significantly diminish the piliation level or the basal expression of PilC1, but led to the loss of cell contact-dependent upregulation of pilC1 and to a dramatic decrease in bacterial adhesiveness. Taken together, these data demonstrate that cell contact-dependent upregulation of the transcription of pilC1 at PC1.3 is essential for meningococcal pilus-mediated adhesion.
Molecular Microbiology 07/1998; 28(6):1153-63. · 5.01 Impact Factor
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ABSTRACT: Strains of Neisseria meningitidis of serogroup B isolated in the Czech Republic frequently belong to serotype 22. We analyzed the genetic relationships among strains of this serotype by using the multilocus enzyme electrophoresis technique and the polymorphism of the pilA gene. Our results indicate that these strains correspond to a highly heterogeneous population rather than to the expansion of a single clone.
Journal of Clinical Microbiology 03/1998; 36(2):563-5. · 4.15 Impact Factor
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ABSTRACT: Due to the spread of the meningococcal infections, a good epidemiological surveillance is needed. Prophylactic measures should be undertaken because of the high transmissibility of these bacteria. One problem which hinders the epidemiological characterization is that the responsible strain should be isolated. The aim of this work is to develop a rapid and non culture typing method of Neisseria meningitidis.
Six cerebrospinal fluids were obtained from 5 different patients with meningococcal meningitis. A specific locus, pil A, for N. meningitidis was amplified by polymerization chain reaction (PCR). The polymorphism of this locus was then analyzed by digesting the PCR products with one of three different restriction enzymes.
The polymorphism of this locus allowed us to establish the clonal relationships between the meningococcal strains involved in the infection. Three CSF corresponded to epidemiological strains.
This typing method allows a rapid and less expensive epidemiological characterization of meningococcal infections. Moreover, it is a non culture typing method.
La Presse Médicale 11/1997; 26(32):1516-9. · 0.67 Impact Factor
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ABSTRACT: The genetic relationships between 88 meningococcal strains were analyzed by using the polymorphism of the pilA gene and the multilocus enzyme electrophoresis. While a good agreement was observed, correlation with antigenic formula (serogroup, serotype, and serosubtype) was incomplete. The inadequacy of serological classification alone in outbreak surveillance may be overcome by DNA-based approaches.
Journal of Clinical Microbiology 04/1997; 35(3):745-50. · 4.15 Impact Factor
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ABSTRACT: Adherence to eukaryotic cells is essential in the pathogenesis of Neisseria meningitidis. Pilus-mediated adhesion has been shown to play an essential role in this process. Pilin, the pilus major subunit, and two pilus associated proteins, PilC1 and PilC2, are key components in meningococcal adhesiveness. Phase and/or antigenic variation of these molecules are the only identified means by which N. meningitidis modulates pilus-mediated adhesion. PilA/PilB is a pleiotropic regulatory system first characterized in Neisseria gonorrhoeae where it controls pilin gene transcription. Similar alleles are found in N. meningitidis. To address the role of this regulatory pathway in N. meningitidis, we engineered a meningococcal pilA mutant strain and analysed the consequences of this mutation on pilus-mediated adhesion using epithelial Hec-1-B cells. This mutation resulted in a threefold reduction in adhesiveness. As no change in the amount of pilin nor in pilin gene mRNA was detected, we compared the expression of the pilC genes in both pilA and parental strains. Two transcriptional fusions pilC1-lacZ and pilC2-lacZ were constructed. A threefold reduction in beta-galactosidase activity was observed in the pilA mutant strain harbouring the pilC1-lacZ fusion. No effect of the pilA mutation on beta-galactosidase activity was observed in the strain carrying the pilC2-lacZ fusion. Gel retardation experiments confirmed that the PilA protein binds to the promoter region of pilC1 but not of pilC2. Taken together, these data demonstrate that PilA modulates meningococcal adhesiveness via the transcription of pilC1. Thus, in addition to phase variation, a more co-ordinate and responsive system may allow a fine adaptation of adhesiveness of meningococci to various environmental signals.
Molecular Microbiology 04/1996; 19(5):1073-84. · 5.01 Impact Factor
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ABSTRACT: A new molecular typing method for identification and characterization of Neisseria meningitidis is reported. Chromosomal DNA from 20 well-documented meningococcal strains of serogroup A originating from France, Central African Republic, Sudan and Burkina Faso were amplified using the polymerase chain reaction. Primers designed in this study were located in the pilA/pilB locus which has been shown to be conserved in the genus Neisseria. The amplified fragments were subjected to restriction endonuclease analysis using three different enzymes, and the restriction endonuclease patterns obtained were compared. Clonal isolates clustered together in distinct restriction endonuclease patterns which are described in this study and coincided with electrotypes as determined by multi-locus enzyme electrophoresis. This DNA-based typing system for meningococci may be useful for epidemiological studies.
Molecular and Cellular Probes 11/1995; 9(5):297-306. · 2.08 Impact Factor
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ABSTRACT: The transcriptional regulation of the pilE gene, coding for the pilin in Neisseria gonorrhoeae, by PilA/PilB proteins is quite complex. Sequence analysis of PilA suggested that it has multiple domains. PilA appears to have in its N-terminal half a DNA-binding site followed by a region showing sequence similarity with other bacterial transcriptional regulators. In its C-terminal half, PilA has extensive homology with the 54 kDa protein of the eukaryotic signal-recognition particle which is involved in protein secretion. A transcriptional fusion between the promoter of pilE and the lacZ gene was constructed and integrated into the gonococcal chromosome. We show that transcription of the pilE-lacZ fusion is affected in pilA mutants in the absence of any possible interference with pilin secretion. Moreover, pilE transcription depends on a -24/-12-type promoter which could be a member of a family of promoters recognized by the alternative sigma subunit, RpoN, of the RNA polymerase. We also show that PilA binds specifically to the promoter region of pilE and that it is phosphorylated in a manner dependent on acidic residues Glu-59, Asp-149 and Asp-186. The functional organization of PilA suggests that it may be an unusual transcriptional regulator different from other RpoN-dependent activators.
Molecular Microbiology 03/1995; 15(4):667-77. · 5.01 Impact Factor
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ABSTRACT: Sequence analysis has shown that PilA, a transcriptional regulator of pilin gene expression in Neisseria gonorrhoeae, has extensive homology with the 54-kDa protein of the signal recognition particle of eukaryotes and its receptor, as well as with two proteins of Escherichia coli, FtsY and Ffh, which have been proposed to be a part of a signal recognition particle-like apparatus. We tested the putative role of PilA in protein export in N. gonorrhoeae and did not find any effect. However, we did observe induction of a heat shock response and a previously described slow-growth phenotype when PilA function was impaired. We also examined the interference of pilA expression in E. coli with the function of the products of ftsY and ffh and observed an accumulation of pre-beta-lactamase. We argue against a direct role for PilA in protein export in gonococci and propose instead that PilA is involved in the modulation of cell growth rate in response to different environmental conditions.
Journal of Bacteriology 10/1992; 174(18):5978-81. · 3.83 Impact Factor
