title: Genetic Determinants of Salmonella enterica Serovar Typhimurium Proliferation in the Cytosol of Epithelial Cells.
abstract: Intestinal epithelial cells provide an important colonization niche for Salmonella enterica serovar Typhimurium during gastrointestinal infections. In infected epithelial cells, a subpopulation of S Typhimurium bacteria damage their internalization vacuole, leading to escape from the Salmonella-containing vacuole (SCV) and extensive proliferation in the cytosol. Little is known about the bacterial determinants of nascent SCV lysis and subsequent survival and replication of Salmonella in the cytosol. To pinpoint S Typhimurium virulence factors responsible for these steps in the intracellular infectious cycle, we screened a S Typhimurium multigene deletion library in Caco-2 C2Bbe1 and HeLa epithelial cells for mutants that had an altered proportion of cytosolic bacteria compared to the wild type. We used a gentamicin protection assay in combination with a chloroquine resistance assay to quantify total and cytosolic bacteria, respectively, for each strain. Mutants of three S Typhimurium genes, STM1461 (ydgT), STM2829 (recA), and STM3952 (corA), had reduced cytosolic proliferation compared to wild-type bacteria, and one gene, STM2120 (asmA), displayed increased cytosolic replication. None of the mutants were affected for lysis of the nascent SCV or vacuolar replication in epithelial cells, indicating that these genes are specifically required for survival and proliferation of S Typhimurium in the epithelial cell cytosol. These are the first genes identified to contribute to this step of the S Typhimurium infectious cycle.
authors.affiliation: Paul G. Allen School for Global Animal Health, College of Veterinary Medicine, Washington State University, Pullman, Washington, USA.
authors.forename: Marie
authors.initials: M
authors.lastname: Wrande
authors.affiliation: Department of Microbial Pathogenesis and Immunology, College of Medicine, Texas A&M University, Bryan, Texas, USA.
authors.forename: Helene
authors.initials: H
authors.lastname: Andrews-Polymenis
authors.affiliation: Paul G. Allen School for Global Animal Health, College of Veterinary Medicine, Washington State University, Pullman, Washington, USA.
authors.forename: Donna J
authors.initials: DJ
authors.lastname: Twedt
authors.affiliation: Laboratory of Bacteriology, Rocky Mountain Laboratories, NIAID, NIH, Hamilton, Montana, USA.
authors.forename: Olivia
authors.initials: O
authors.lastname: Steele-Mortimer
authors.affiliation: Department of Microbiology and Molecular Genetics, University of California-Irvine, Irvine, California, USA.
authors.forename: Steffen
authors.initials: S
authors.lastname: Porwollik
authors.affiliation: Department of Microbiology and Molecular Genetics, University of California-Irvine, Irvine, California, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: McClelland
authors.affiliation: Paul G. Allen School for Global Animal Health, College of Veterinary Medicine, Washington State University, Pullman, Washington, USA lknodler@vetmed.wsu.edu.
authors.forename: Leigh A
authors.initials: LA
authors.lastname: Knodler
mesh_terms: D000818:Animals; D002460:Cell Line; D049109:Cell Proliferation / Q000235:genetics*; D003600:Cytosol / Q000382:microbiology*; D004847:Epithelial Cells / Q000382:microbiology*; D006801:Humans; D051379:Mice; D009154:Mutation; D012486:Salmonella typhimurium / Q000235:genetics* / Q000472:pathogenicity*