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  Bordetel


  Director : GUISO Nicole (nguiso@pasteur.fr)


  abstract

 

Bacteria from the Bordetella genus are responsible for respiratory infections in humans (Bordetella pertussis and Bordetella parapertussis and whooping cough) and in animals (Bordetella bronchiseptica and atrophic rhinitis and kennel cough). The researches developed in our laboratory are aimed to characterize the determinants involved in the pathogenicity of these bacteria analyse immune responses induced in the host after infection and after vaccination, surveillance of polymorphism of clinical isolates and setting up of new prevention and new diagnosis for whooping cough and other bordetellosis



  report

cale

I - Bordetella pertussis VIRULENCE DETERMINANTS

  • Does adenylate cyclase-hemolysin toxin interact with a receptor on eucaryotic cells ?

[N. Khelef, in collaboration with P. Guermonprez and C. Leclerc]

We previously shown that adenylate cyclase-hemolysin (AC-Hly) toxin, secreted by B. pertussis, is responsible in vivo and in vitro of alveolar macrophages death via apoptosis but is not cytotoxic for epithelial cells. For this reason we made the hypothesis that a specific receptor for AC-Hly exists on the macrophage surface. In fact, using different specific antibodies we recently showed that the integrin Mac-1 is the receptor of AC-Hly. Since epithelial cells do not express Mac-1, they are resistant to AC-Hly toxin.

  • What is the localisation of adenylate cyclase-hemolysin inside macrophages or epithelial cells ? [N. Khelef in collaboration with P. Gounon]

It was important using confocal microscopy and immunofluorescence labelling what was the AC-Hly localisation inside the macrophage. Our results show that after fixation on its receptor AC-Hly enters the cell by macropinocytosis. This mechanim is required for AC-Hly cytotoxic activity.

II - PHYSIOPATHOLOGY OF BORDETELLA INFECTIONS

  • Interactions of Bordetella pertussis with human tracheal epithelial cells [L. Bassinet, P. Gueirard in collaboration with P. Gounon]

Epithelial cell plays a role in the recrutment of inflammary cells via cytokines secretion. For this reason our aim is to analyse which cytokines are secreted by human tracheal epithelial cells infected by B. pertussis or in contact with purified toxins or adhesins. Our preliminary results show that B. pertussis strains induce IL-6 secretion while there is no induction or inhibition of secretion of IL-8.

  • Do Bordetella pertussis or Bordetella bronchiseptica survive intracellularly ? In which cell ? [P. Gueirard, S. Thiberge in collaboration with P. Ave, M. Huerre, AM. Balazuc and G. Milon]

The study is performed in vivo using our murine respiratory model and in vitro. Is the target cell a dendritic leukocyte ? Our first results indicate the recruitment of dendritic leucocytes in the pulmonary parenchymus and bronchoalveolar fluid early after infection as well as the presence of intracellular bacteria.

III - POLYMORPHISM OF BORDETELLA PERTUSSIS, PARAPERTUSSIS, AND BRONCHISEPTICA ISOLATES

  • Is there a genetical drift in Bordetella pertussis population over vaccinal pressure ? [C. Boursaux-Eude, F. Rimlinger, C. Weber]

    • Analysis, by typing technique or sequencing of toxin and adhesin structural genes, of isolates circulating in France, a vaccinated country since more than thirty five years with a whole-cell vaccine, confirmed that B. pertussis isolates circulating before and after introduction of generalized vaccination are different. However, a precise analysis suggests that B. pertussis population could vary regularly every three years.
    • Analysis of isolates circulating in Japan, a vaccinated country using acellular vaccines since twenty years showed that circulating isolates are similar to the vaccinal strain. Use of acellular vaccines does not seem to induce a strong selection pressure. However, a greater number of isolates has to be analysed before to draw any conclusion.
    • We are currently analizing B. pertussis isolates from Russia a country with a low vaccine coverage.

  • Polymorphism structural gene encoding pertactin in Bordetella pertussis, parapertussis and bronchiseptica

    • We previously shown that PRN expressed by B. pertussis, B. parapertussis and B. bronchiseptica are very similar but do not induce a cross reactive protective immunity. The differences between the three proteins are located on two regions of the PRN protein (I and II) containing repeated regions. Analysis of several B. pertussis, parapertussis and bronchiseptica PRN indicates that B. pertussis PRN variation is located on region I whereas B. bronchiseptica PRN variation is mainly located on region II. This result shows the importance of PRN region II for the induction of protective immunity. One can suppose that variation of this region could induce vaccine failure.

IV - WHOOPING COUGH PREVENTION

  • What is the prevalence of whooping cough in adults in France ? [E. Njamkepo, A. Le Flèche, P. Gueirard in collaboration with I. Parent (AMP), S. Gilberg (SFTG) and Aventis-Pasteur and Aventis Pasteur-MSD]

In France, since several years a resurgence of whooping cough is observed with infants contamined by adolescents and adults. In order to confirm this epidemiology we undertook with 80 general practitioners, a study to determine the prevalence of whooping cough in adults, describe the clinical symptoms during the disease and evaluate the ability of biological diagnosis such as culture, PCR and serology to diagnose the disease in adult. A prevalence of 32 % was found. All confirmed case were characterized by : a higher frequency of work stops, cough with whoops PCR on experations is a more sensitive diagnosis than culture. Measurement of anti-pertussis toxin IgG and IgA by ELISA seems also a very useful diagnosis.

  • Is Bordetella pertussis polymorphism affecting vaccine efficacy ? [S. Thiberge in collaboration with GlaxoSmithKline and Aventis-Pasteur Laboratories]

Our murine intranasal model, was used to compare the ability of different whole cell and acellular vaccines. Till now, all vaccines are able to induce a protective immunity against infections due to different variants.

  • Whooping cough and immunotherapy [E. Coëffier-Vicart, S. Thiberge in collaboration with F. Nato and P. Lafaye]

The new epidemiology in countries using generalized vaccination since many years is characterized by an increase in new-born mortality (too young to be vaccinated) contaminated by adolescents or adults whom immunity waned with time. The use of specific antibodies able to neutralize toxins and adhesins activities could be a useful tool to decrease symptoms during infection and to stop transmission of the bacteria. For these reason we tested the ability of different antibodies to decrease symptoms after Bordetella infections using our respiratory murine model.

V - BORDETELLA BRONCHISEPTICA HUMAN AND ANIMAL INFECTIONS

  • Bordetella bronchiseptica human and animal infections [P. Gueirard, S. Thiberge, L. Guillemot in collaboration with C. Barny]

B. bronchiseptica is a respiratory pathogen for many mamals species, including human. In human, B. bronchiseptica is considered as an opportunist pathogen for immunodepressed patient or patient with respiratory problems. We previously described that this bacterium can induce chronic infections. We confirmed the zoonotic character of this infection with two patients contaminated with either infected rabbit or dog. Furthermore, samples from dogs showed that the bacterium can be isolated on vaccinated dogs.

Studies will continue because of the increase of immunocompromised patients, absence of cross protection of new pertussis vaccine, low efficacy of veterinary vaccines and antibiotic resistances carried by this bacterium.

VI - CENTRE NATIONAL DE REFERENCE

  • CNR [G. Coralie, P. Gueirard, E. Njamkepo, S. Thiberge]

The missions of this Centre is to confirm the identification of Bordetella isolates collected in France, to maintain the collection, to teach French bacteriologists, to analyse isolates by Pulsed-Field-Gel Electrophoresis, as well as the toxins and adhesins they express, to set up sensitive and specific biological diagnosis techniques and to be part of the National surveillance net, RENACOQ, coordinated by the Ministery of Health.

VII - QUALITY INSURANCE

  • Quality Insurance [F. Fouque]

The set up of procedures and quality insurance in the laboratory was pursued.



  publications

puce Publications of the unit on Pasteur's references database


  personnel

  Office staff Researchers Scientific trainees Other personnel
 

Laurence Langlais –langlais@pasteur.fr – du 11/12/00 à maintenant

Eliane COËFFIER-VICART – coeffier@pasteur.fr – Chargée de Recherche IP

Pascale GUEIRARD – pgueirar@pasteur.fr – Chargée de Recherche IP

Nicole GUISO – nguiso@pasteur .fr - Chef de laboratoire IP

Valérie Caro – post-doc – vcaro@pasteur.fr – stagiaire post-doctorale

Laurence Bassinet – laurence.bassinet@chicreteil.fr – étudiante en thèse

Gilberte CORALIE – gcoralie@pasteur.fr – Technicienne IP

Elodie DENISE – denise@pasteur.fr – Technicienne supérieure à partir de novembre 2001

Françoise FOUQUE – ffouque@pasteur.fr – Ingénieur IP

Laurent GUILLEMOT – guillemo@pasteur.fr - Technicien supérieur IP.

Elisabeth NJAMKEPO – enjamkep@pasteur.fr – Ingénieur IP François RIMLINGER – frimling@pasteur.fr – Technicien supérieur IP

Guillaume SOUBIGOU – soubigou@pasteur.fr – Technicien supérieur IP à partir de octobre 2001

Sabine THIBERGE – thiberge@pasteur.fr – Technicienne supérieure IP jusqu'en septembre 2001

Christian WEBER – cweber@pasteur.fr – Technicien supérieur IP jusqu'en avril 2001

Martine BLERIOT – Aide de laboratoire, (en commun avec le laboratoire Agents antibactériens)

Sandrine FAVRE-ROCHEX – Aide de laboratoire,( en commun avec le laboratoire Agents antibactériens)

Marie-Reine LAURET – Agent de laboratoire, (en commun avec le laboratoire Agents antibactériens)

Pierre SAGOT– Agent de laboratoire, (en commun avec le laboratoire Agents antibactériens)


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