Application of Signature-Tagged Mutagenesis for Identification of Escherichia coli K1 Genes That Contribute to Invasion of Human Brain Microvascular Endothelial Cells
- 1 September 2000
- journal article
- research article
- Published by American Society for Microbiology in Infection and Immunity
- Vol. 68 (9), 5056-5061
- https://doi.org/10.1128/iai.68.9.5056-5061.2000
Abstract
Escherichia coli K1 is the leading cause of gram-negative bacterial meningitis in neonates. It is principally due to our limited understanding of the pathogenesis of this disease that the morbidity and mortality rates remain unacceptably high. To identify genes required for E. coli K1 penetration of the blood-brain barrier (BBB), we used the negative selection strategy of signature-tagged transposon mutagenesis (STM) to screen mutants for loss or decreased invasion of human brain microvascular endothelial cells (HBMEC) which comprise the BBB. A total of 3,360 insertion mutants of E. coli K1 were screened, and potential HBMEC invasion mutants were subjected to a secondary invasion screen. Those mutants that failed to pass the serial invasion screens were then tested individually. Seven prototrophic mutants were found to exhibit significantly decreased invasive ability in HBMEC. We identified traJ and five previously uncharacterized loci whose gene products are necessary for HBMEC invasion by E. coli K1. In addition, cnf1 , a gene previously shown to play a role in bacterial invasion, was identified. More importantly, a traJ mutant was attenuated in penetration of the BBB in the neonatal rat model of experimental hematogenous meningitis. This is the first in vivo demonstration that traJ is involved in the pathogenesis of E. coli K1 meningitis.Keywords
This publication has 19 references indexed in Scilit:
- Identification of Escherichia coli K1 genes contributing to human brain microvascular endothelial cell invasion by differential fluorescence inductionMolecular Microbiology, 2000
- Complex lipid determines tissue-specific replication of Mycobacterium tuberculosis in miceNature, 1999
- The Gene Locus yijP Contributes to Escherichia coli K1 Invasion of Brain Microvascular Endothelial CellsInfection and Immunity, 1999
- The Complete Genome Sequence of Escherichia coli K-12Science, 1997
- Simultaneous Identification of Bacterial Virulence Genes by Negative SelectionScience, 1995
- Cloning of the YenI restriction endonuclease and methyltransferase from Yersinia enterocolitica serotype O8 and construction of a transformable R−M+ mutantGene, 1993
- Structural and functional analyses of the FinP antisense RNA regulatory system of the F conjugative piasmidMolecular Microbiology, 1993
- Induction of phagocytic behaviour in human epithelial cells by Escherichia coli cytotoxic necrotizing factor type1Molecular Microbiology, 1993
- The K1 capsule is the critical determinant in the development of Escherichia coli meningitis in the rat.JCI Insight, 1992
- A Broad Host Range Mobilization System for In Vivo Genetic Engineering: Transposon Mutagenesis in Gram Negative BacteriaBio/Technology, 1983