title: Comprehensive Identification of Meningococcal Genes and Small Noncoding RNAs Required for Host Cell Colonization.
abstract: Neisseria meningitidis is a leading cause of bacterial meningitis and septicemia, affecting infants and adults worldwide. N. meningitidis is also a common inhabitant of the human nasopharynx and, as such, is highly adapted to its niche. During bacteremia, N. meningitidis gains access to the blood compartment, where it adheres to endothelial cells of blood vessels and causes dramatic vascular damage. Colonization of the nasopharyngeal niche and communication with the different human cell types is a major issue of the N. meningitidis life cycle that is poorly understood. Here, highly saturated random transposon insertion libraries of N. meningitidis were engineered, and the fitness of mutations during routine growth and that of colonization of endothelial and epithelial cells in a flow device were assessed in a transposon insertion site sequencing (Tn-seq) analysis. This allowed the identification of genes essential for bacterial growth and genes specifically required for host cell colonization. In addition, after having identified the small noncoding RNAs (sRNAs) located in intergenic regions, the phenotypes associated with mutations in those sRNAs were defined. A total of 383 genes and 8 intergenic regions containing sRNA candidates were identified to be essential for growth, while 288 genes and 33 intergenic regions containing sRNA candidates were found to be specifically required for host cell colonization.
  
  
  
  Meningococcal meningitis is a common cause of meningitis in infants and adults. Neisseria meningitidis (meningococcus) is also a commensal bacterium of the nasopharynx and is carried by 3 to 30% of healthy humans. Under some unknown circumstances, N. meningitidis is able to invade the bloodstream and cause either meningitis or a fatal septicemia known as purpura fulminans. The onset of symptoms is sudden, and death can follow within hours. Although many meningococcal virulence factors have been identified, the mechanisms that allow the bacterium to switch from the commensal to pathogen state remain unknown. Therefore, we used a Tn-seq strategy coupled to high-throughput DNA sequencing technologies to find genes for proteins used by N. meningitidis to specifically colonize epithelial cells and primary brain endothelial cells. We identified 383 genes and 8 intergenic regions containing sRNAs essential for growth and 288 genes and 33 intergenic regions containing sRNAs required specifically for host cell colonization.
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France.
authors.forename: Elena
authors.initials: E
authors.lastname: Capel
authors.affiliation: Department of Infectious Diseases and Immunology, Faculty of Veterinary Medicine, Utrecht University, Utrecht, The Netherlands.
authors.forename: Aldert L
authors.initials: AL
authors.lastname: Zomer
authors.affiliation: CUBE Division of Computational Systems Biology, Department of Microbiology and Ecosystem Science, University of Vienna, Vienna, Austria.
authors.forename: Thomas
authors.initials: T
authors.lastname: Nussbaumer
authors.affiliation: Plateforme Génomique de l'Institut Imagine, Hôpital Necker, Paris, France.
authors.forename: Christine
authors.initials: C
authors.lastname: Bole
authors.affiliation: Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France INSERM U1016, Institut Cochin, CNRS UMR8104, Paris, France.
authors.forename: Brigitte
authors.initials: B
authors.lastname: Izac
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France.
authors.forename: Eric
authors.initials: E
authors.lastname: Frapy
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France.
authors.forename: Julie
authors.initials: J
authors.lastname: Meyer
authors.affiliation: Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France INSERM U1016, Institut Cochin, CNRS UMR8104, Paris, France.
authors.forename: Haniaa
authors.initials: H
authors.lastname: Bouzinba-Ségard
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France Assistance Publique-Hôpitaux de Paris, Hôpital Necker Enfants Malades, Paris, France.
authors.forename: Emmanuelle
authors.initials: E
authors.lastname: Bille
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France Unidade de Microbiologia Molecular e Infecção, Instituto de Medicina Molecular, Lisbon, Portugal Assistance Publique-Hôpitaux de Paris, Hôpital Necker Enfants Malades, Paris, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Jamet
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France.
authors.forename: Anne
authors.initials: A
authors.lastname: Cavau
authors.affiliation: Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France INSERM U1016, Institut Cochin, CNRS UMR8104, Paris, France.
authors.forename: Franck
authors.initials: F
authors.lastname: Letourneur
authors.affiliation: Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France INSERM U1016, Institut Cochin, CNRS UMR8104, Paris, France.
authors.forename: Sandrine
authors.initials: S
authors.lastname: Bourdoulous
authors.affiliation: CUBE Division of Computational Systems Biology, Department of Microbiology and Ecosystem Science, University of Vienna, Vienna, Austria.
authors.forename: Thomas
authors.initials: T
authors.lastname: Rattei
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France Assistance Publique-Hôpitaux de Paris, Hôpital Necker Enfants Malades, Paris, France xavier.nassif@inserm.fr.
authors.forename: Xavier
authors.initials: X
authors.lastname: Nassif
authors.affiliation: Institut Necker Enfants-Malades, INSERM U1151, Equipe 11, Paris, France Université Paris Descartes, Sorbonne Paris Cité, Faculté de Médecine, Paris, France.
authors.forename: Mathieu
authors.initials: M
authors.lastname: Coureuil
mesh_terms: D002460:Cell Line; D004251:DNA Transposable Elements; D004705:Endocytosis; D042783:Endothelial Cells / Q000382:microbiology*; D004847:Epithelial Cells / Q000382:microbiology*; D055786:Gene Knockout Techniques; D015723:Gene Library; D006801:Humans; D016254:Mutagenesis, Insertional; D009345:Neisseria meningitidis / Q000235:genetics* / Q000254:growth & development / Q000472:pathogenicity*; D058727:RNA, Small Untranslated / Q000235:genetics*; D037521:Virulence Factors / Q000235:genetics*
chemical_list: D004251:DNA Transposable Elements; D058727:RNA, Small Untranslated; D037521:Virulence Factors