Université Paris Descartes

Faculté de Médecine
Paris Descartes (site Necker)

Unité de Pathogénie
des Infections Systémiques
UMR 570

Head : Prof. Xavier Nassif

156, rue de Vaugirard
75730 Paris Cedex 15
FRANCE
Extra-cellular bacterial pathogens are the main cause of infections in developed countries and are often responsible for nosocomial infections. Paradoxically these bacteria are mostly commensal, and become invasive only in certain circumstances. Among the great variety of commensal bacteria that colonize a human, only few strains are able to disseminate. Furthermore some of these bacteria once septicemic can cross the blood-brain barrier and be responsible for meningitis.


Our goal is, using Neisseria meningitidis as a model, to identify the mechanisms by which (i) a bacteria can stop being a commensal and disseminate from its normal niche, and (ii) once it is septicemic how it can cross the blood brain barrier. N.meningitis is responsible for septicemia and meningitis after crossing of the blood brain barrier. Meningococcal meningitis remain a health threat in both developped and developping countries, and there is an urgent need for the development of a new vaccine strategy. Paradoxically, it is a commensal of human throat and colonizes a significant percentage of a population as healthy carrier. The reason why in some circumstances N.meningitidis can invade the bloodstream and become pathogenic remain unknown. Host factors are obviously important, however recently some clonal complexes have been shown to be more likely than others to be invasive.


We concentrate our work in several directions :


1. Identification of the bacterial attributes linked to invasiveness.


By comparative genomics, we have recently identified using a large collection of strains isolated from carriers and patients during the same period and in the same area that strains isolated from diseases were most likely to contain a prophage, designated Meningococcal Disease Associated (MDA) phage. We subsequently demonstrated that this phage belongs to the family of filamentous phage, like M13 and the CTX phage of Vibrio cholereae. Further work is ongoing in order to characterize the molecular mechanisms by which this phage increases invasiveness.


2. Identification of the interaction of N.meningitidis with the host cells in relation to the crossing of the blood-brain-barrier

Our contribution has been to demonstrate that N. meningitidis cross this barrier by interacting directly with the endothelial cells of the BBB. Recently, we combined several approaches: (i) in vitro modeling of the interaction of the bacterium with endothelial cells in the context of flowing liquid with a laminar flow chamber; and (ii) live, in vivo imaging of cerebral microcirculation. Current results on Neisseria meningitidis indicate that the mechanical properties of neisserial adhesion and blood microcirculation target meningococci to transiently underperfused cerebral capillaries and thus determine disease development. This integrated approach will be further exploited for N. meningitidis and will be extended to other pathogens that interact with the BBB.


We have shown, in collaboration with the laboratory of Sandrine Bourdoulous and Pierre-Olivier Couraud (Institut Cochin, Paris, France) that during adhesion to host cells, N. meningitidis triggers a local remodeling of the eukaryotic cell surface resulting in a meshwork of cellular projections that can be found, in vitro and in vivo, inside and around microcolonies adhering to the cell surface (Figure). Numerous host cell proteins are recruited within minutes under the bacterial microcolony, including actin-associated proteins. The signaling modulates the ability of the bacteria to invade host cells and possibly to cross cellular barriers placing this process at the center of pathogenesis. We are pursuing this work with the aim to identify the consequence of this signaling onto the tight junctions and the host cell proteins responsible for the initiation of the signaling.

3. Identification of the bacterial attributes involved in the host cell interaction

Although the potent and rapid bacteria-induced cellular response is well described morphologically and some cellular participants identified, the bacterial components involved in triggering this signaling remain to be identified. In contrast to several Gram-negative bacterial pathogens, N. meningitidis does not express type III or type IV secretion systems. Numerous outermembrane components have been identified as allowing the interaction of N. meningitidis with cells, however the only one whose role has been demonstrated to be essential for the crossing of the BBB are the type IV pili. In order to extensively identify the bacterial components involved in piliation, internalization and transcytose we used a comparative and functional genomic approaches. This approach identified the type IV pili as the only bacterial attribute required for bacterial adhesion and signaling. We are pursuing this work by deciphering the function of the pilus-associated components involved in the interaction with the host cells and identifying the subsequent eukaryotic counterparts.


The ability to adhere to and grow on the surface of human host cells is an absolute requirement for N. meningitidis to colonize new hosts and to disseminate inside its host. However, how N. meningitidis adapts to growth on host cells remains poorly understood. Identification of the function of the meningococcal genes differentially expressed upon initial interaction with host cells and of the regulation pathways controlling the expression of these genes are prerequisites to the understanding of how efficient colonization can be achieved and thereby explain in part the host-specificity of the meningococcus.





Xavier Nassif (Professeur, Université Paris 5)
Emmanuelle Bille (AHU, AP-HP)
Marek Drab (Professor)
Eric Frapy (Ingénieur, U-Paris Descartes)
Mathieu Coureuil (Post-Doc)
Anne Jamet (PhD student, U-Paris Descartes)
Herve Lecuyer (PhD student, U-Paris Descartes)
Hélene Omer (PhD student, U-Paris Descartes)



Tinsley C. R., Bille E., Nassif X. 2006.Bacteriophages and pathogenicity: more than just providing a toxin? Microbes Infect., 8: 1365-71

Bourdoulous S., Couraud P.-O., Nassif X. 2002.Methods for studying the mechanisms of Microbial entry into the central nervous system. In Methods in Microbiology, 131: 420-437

Nassif X., Bourdoulous S., Eugene E., and Couraud P.-O. 2002. How do extracellular pathogens cross the blood-brain barrier ? Trends in Microbiology, 10: 227-232

Tinsley C. , Nassif X. 2001. Meningococcal pathogenesis: at the boundary between the pre and post genomic eras. Current Opinion in Microbiology, 4: 47-52.

Nassif X. A furtive pathogen revealed (perspective).2000. Science, 287: 1767-1768.





Mikaty G, Soyer M, Mairey E, Henry N, Dyer D, Forest KT, Morand P, Guadagnini S, Prevost MC, Nassif X and Dumenil G. 2009. Extracellular bacterial pathogen induces host cell surface reorganization to resist shear stress. PLoS Pathog., 5(2):e1000314.

Helaine S, Dyer DH, Nassif X, Pelicic V, Forest KT. 2007. 3D structure/function analysis of PilX reveals how minor pilins can modulate the virulence properties of type IV pili. Proc. Natl. Acad. Sci., 104: 15888-93.

Chamot-Rooke J, Rousseau B, Lanternier F, Mikaty G, Mairey E, Malosse C, Bouchoux G, Pelicic V, Camoin L, Nassif X, Dumenil G. 2007. Alternative Neisseria spp. type IV pilin glycosylation with a glyceramido acetamido trideoxyhexose residue. Proc. Natl. Acad. Sci., 104: 14783-8.

Carbonnelle E., Helaine S., Nassif X., and Pelicic V. 2006. A systematic genetic analysis in Neisseria meningitidis defines the Pil proteins required for assembly, functionality, stabilization and export of type IV pili. Mol Microbiol. 61: 1510-22.

Linhartova I., Basler M., Ichikawa J., Pelicic V., Osicka R., Lory S., Nassif X. and Sebo P. 2006. Meningococcal adhesion suppresses proapoptotic gene expression and promotes expression of genes supporting early embryonic and cytoprotective signaling of human endothelial cells. FEMS Microbiol Lett. 263: 109-18.

Mairey E., Genovesio A., Donnadieu E., Bernard C., Jaubert F., Pinard E., Seylaz J., Olivo-Marin J. C., Nassif X., and Dumenil G. 2006. Cerebral microcirculation shear stress levels determine Neisseria meningitidis attachment sites along the blood-brain barrier. J Exp Med. 203: 1939-50.

Doulet N., Donnadieu E., Laran-Chich M. P., Niedergang F., Nassif X., Couraud P. O., and Bourdoulous S. 2006. Neisseria meningitidis infection of human endothelial cells interferes with leukocyte transmigration by preventing the formation of endothelial docking structures. J Cell Biol. 173: 627-37.

Tinsley C. R., Bille E., and Nassif X.. 2006. Bacteriophages and pathogenicity: more than just providing a toxin? Microbes Infect. 8: 1365-71.

Yasukawa K., Martin P., Tinsley C. R., and Nassif X. 2006. Pilus-mediated adhesion of Neisseria meningitidis is negatively controlled by the pilus-retraction machinery. Mol Microbiol. 59: 579-89.

Hakkarainen J., Toivanen M., Leinonen A., Frangsmyr L., Stromberg N., Lapinjoki S., Nassif X., and Tikkanen-Kaukanen C. 2005. Human and bovine milk oligosaccharides inhibit Neisseria meningitidis pili attachment in vitro. J Nutr.135: 2445-8.

Lambotin M., Hoffmann I., Laran-Chich M. P., Nassif X., Couraud P. O., and Bourdoulous S. 2005. Invasion of endothelial cells by Neisseria meningitidis requires cortactin recruitment by a phosphoinositide-3-kinase/Rac1 signalling pathway triggered by the lipo-oligosaccharide. J Cell Sci. 118: 3805-16.

Dumenil G. and Nassif X. 2005. Extracellular bacterial pathogens and small GTPases of the Rho family: an unexpected combination. Curr Top Microbiol Immunol. 291: 11-28.

Bille E., Zahar J. R., Perrin A., Morelle S., Kriz P., Jolley K. A., Maiden M. C., Dervin C., Nassif X., and Tinsley C. R. 2005. A chromosomally integrated bacteriophage in invasive meningococci. J Exp Med. 201: 1905-13.

Morelle S., Carbonnelle E., Matic I., and Nassif X. 2005. Contact with host cells induces a DNA repair system in pathogenic Neisseriae. Mol Microbiol. 55: 853-61.

Helaine S., Carbonnelle E., Prouvensier L., Beretti J. L., Nassif X., and Pelicic V. 2005. PilX, a pilus-associated protein essential for bacterial aggregation, is a key to pilus-facilitated attachment of Neisseria meningitidis to human cells. Mol Microbiol. 55: 65-77.

Carbonnelle E., Helaine S., Prouvensier L., Nassif X., and Pelicic V. 2005. Type IV pilus biogenesis in Neisseria meningitidis: PilW is involved in a step occurring after pilus assembly, essential for fibre stability and function. Mol Microbiol. 55: 54-64.

Morand P. C, Bille E., Morelle S., Eugene E., Beretti J.L., Wolfgang M., Meyer T. F., Koomey M. and Nassif X. 2004. Type IV pilus retraction in pathogenic Neisseria is regulated by the PilC proteins. EMBO J., 3 : 2009-17.

Geoffroy M. C., Floquet S., Metais A., Nassif X., and Pelicic V. 2003. Large-scale analysis of the meningococcus genome by gene disruption: resistance to complement-mediated lysis. Genome Research, 13: 391-398.