Use of signature-tagged mutagenesis to identify genes associated with colonization of sheep by E. coli O157:H7

Vet Microbiol. 2017 Mar:201:177-182. doi: 10.1016/j.vetmic.2017.01.031. Epub 2017 Jan 30.

Abstract

Outbreaks of Escherichia coli O157:H7 in the United States due to contaminated foods are a public health issue and a continuing problem. The major reservoir for these organisms is the gastrointestinal tract of ruminants where they are a member of the resident microbiota. Several factors that contribute to the colonization of cattle have been identified, but a systematic screen of genes that might contribute to the colonization and persistence phenotype in mature ruminants has not been reported. Using a sheep model of persistence, signature tagged mutagenesis (STM) was used to screen 1326 mutants for a persistence-negative phenotype of E. coli O157:H7. We identified 9 genes by STM that appeared to be required for colonization and/or survival in sheep. Three of the genes had functions associated with central metabolism (thiK, ftrA and nrdB), one was involved with LPS formation (wbdP), one encodes a non-LEE encoded effector protein (nleB) and one was a methyltransferase encoded on a prophage (Z2389). The remaining three genes did not have homology with any known genes. Six sheep given ΔwbdP and 2 sheep each were given mutants (ΔthiK (Z1745), ΔftrA (Z2164) and Z2389). The ΔwbdP mutant was recovered from the feces of 4/6 sheep at 6 days pi with a mean number of 1.42log10CFU/g feces compared to 4.6log10CFU/g feces for the wild type strain. This difference was significant (P<0.001) over the time course of the experiment (days 6-23). Both ΔthiK and ΔftrA mutants were recovered from 1 of 2 sheep at 9 days PI by enrichment procedures (<50CFU/g feces) whereas mutant Z2389 was not recovered from either animal past 2 days pi. The roles of all of these gene products require further study to determine how the persistence phenotype of a given strain of E. coli O157:H7 interacts with host factors.

Keywords: Colonization; E. coli O157:H7; Sheep.

MeSH terms

  • Animals
  • Bacterial Adhesion / genetics
  • Colony Count, Microbial / veterinary
  • Escherichia coli Infections / microbiology
  • Escherichia coli Infections / veterinary*
  • Escherichia coli O157 / genetics*
  • Escherichia coli O157 / pathogenicity
  • Escherichia coli Proteins / genetics*
  • Feces / microbiology
  • Gastrointestinal Tract / microbiology
  • Mutagenesis, Insertional
  • Sheep
  • Sheep Diseases / microbiology*
  • Virulence Factors / genetics

Substances

  • Escherichia coli Proteins
  • Virulence Factors