1887

Abstract

The gene is the last cistron in the operon and functions as a negative regulator of the Pho regulon. The authors previously identified a mutant of extraintestinal pathogenic strain CFT073 and demonstrated that this mutant was attenuated for survival in the murine model of ascending urinary tract infection. It is hypothesized that the PhoU protein might serve as a urovirulence factor by indirectly affecting the expression of virulence-related genes. In this study, the mutant was further characterized and PhoU was confirmed as a virulence factor. Western blot analysis demonstrated that insertion of the transposon in the gene disrupted the expression of PhoU. The mutant had derepressed alkaline phosphatase activity under phosphate-excess and -limiting conditions. In single-challenge murine ascending urinary tract infection experiments, quantitative cultures of urine, bladder and kidney revealed no significant differences between the mutant strain and the wild-type strain CFT073. However, in competitive colonization experiments, the mutant strain was significantly out-competed by the wild-type strain in the kidneys and urine and recovered in lower amount in the bladder. Complementation of the mutant with a plasmid containing the wild-type gene restored the expression of PhoU and alkaline phosphate activity to wild-type levels and no significant difference in colonization was observed between the mutant containing the complementing plasmid and wild-type in competitive colonization experiments. In human urine, the mutant and wild-type grew comparably when inoculated independently, indicating that the attenuation observed was not due to a general growth defect. However, as observed , the wild-type out-competed the mutant in competition growth experiments in human urine. These data indicate that PhoU contributes to efficient colonization of the murine urinary tract and add PhoU to a short list of confirmed urovirulence factors.

Loading

Article metrics loading...

/content/journal/micro/10.1099/mic.0.28281-0
2006-01-01
2024-03-29
Loading full text...

Full text loading...

/deliver/fulltext/micro/152/1/153.html?itemId=/content/journal/micro/10.1099/mic.0.28281-0&mimeType=html&fmt=ahah

References

  1. Bahrani-Mougeot F. K, Buckles E. L, Lockatell C. V, Hebel J. R, Johnson D. E, Tang C. M, Donnenberg M. S. 2002; Type-1 fimbriae and extracellular polysaccharides are preeminent uropathogenic Escherichia coli virulence determinants in the murine urinary tract. Mol Microbiol 45:1079–1093 [CrossRef]
    [Google Scholar]
  2. Bochner B. R, Gadzinski P, Panomitros E. 2001; Phenotype microarrays for high-throughput phenotypic testing and assay of gene function. Genome Res 11:1246–1255 [CrossRef]
    [Google Scholar]
  3. Brinkman E, Beckwith J. 1975; Analysis of the regulation of Escherichia coli alkaline phosphatase synthesis using deletions and ϕ 80 transducing phages. J Mol Biol 96:307–316 [CrossRef]
    [Google Scholar]
  4. Burall L. S, Harro J. M, Li X, Lockatell C. V, Himpsl S. D, Hebel J. R, Johnson D. E, Mobley H. L. 2004; Proteus mirabilis genes that contribute to pathogenesis of urinary tract infection: identification of 25 signature-tagged mutants attenuated at least 100-fold. Infect Immun 72:2922–2938 [CrossRef]
    [Google Scholar]
  5. Connell H, Agace W, Klemm P, Schembri M, Mårild S., Svanborg C. 1996; Type 1 fimbrial expression enhances Escherichia coli virulence for the urinary tract. Proc Natl Acad Sci U S A 93:9827–9832 [CrossRef]
    [Google Scholar]
  6. Culham D. E, Dalgado C, Gyles C. L, Mamelak D, MacLellan S, Wood J. M. 1998; Osmoregulatory transporter ProP influences colonization of the urinary tract by Escherichia coli . Microbiology 144:91–102 [CrossRef]
    [Google Scholar]
  7. Daigle F, Fairbrother J. M, Harel J. 1995; Identification of a mutation in the pst-phoU operon that reduces pathogenicity of an Escherichia coli strain causing septicemia in pigs. Infect Immun 63:4924–4927
    [Google Scholar]
  8. Foxman B, Barlow R, D'Arcy H, Gillespie B, Sobel J. D. 2000; Urinary tract infection: self-reported incidence and associated costs. Ann Epidemiol 10:509–515 [CrossRef]
    [Google Scholar]
  9. Guyer D. M, Radulovic S, Jones F. E, Mobley H. L. T. 2002; Sat, the secreted autotransporter toxin of uropathogenic Escherichia coli , is a vacuolating cytotoxin for bladder and kidney epithelial cells. Infect Immun 70:4539–4546 [CrossRef]
    [Google Scholar]
  10. Johnson J. R, Russo T. A. 2002; Uropathogenic Escherichia coli as agents of diverse non-urinary tract extraintestinal infections. J Infect Dis 186:859–864 [CrossRef]
    [Google Scholar]
  11. Litwin M. S, Saigal C. S, Yano E. M. 8 other authors 2005; Urologic diseases in America Project: analytical methods and principal findings. J Urol 173:933–937 [CrossRef]
    [Google Scholar]
  12. Lucas R. L, Lostroh C. P, DiRusso C. C, Spector M. P, Wanner B. L, Lee C. A. 2000; Multiple factors independently regulate hilA and invasion gene expression in Salmonella enterica serovar typhimurium. J Bacteriol 182:1872–1882 [CrossRef]
    [Google Scholar]
  13. Mobley H. L. T, Green D. M, Trifillis A. L, Johnson D. E, Chippendale G. R, Lockatell C. V, Jones B. D, Warren J. W. 1990; Pyelonephritogenic Escherichia coli and killing of cultured human renal proximal tubular epithelial cells: role of hemolysin in some strains. Infect Immun 58:1281–1289
    [Google Scholar]
  14. Morohoshi T, Maruo T, Shirai Y, Kato J, Ikeda T, Takiguchi N, Ohtake H, Kuroda A. 2002; Accumulation of inorganic polyphosphate in phoU mutants of Escherichia coli and Synechocystis sp. strain PCC6803. Appl Environ Microbiol 68:4107–4110 [CrossRef]
    [Google Scholar]
  15. Muda M, Rao N. N, Torriani A. 1992; Role of PhoU in phosphate transport and alkaline phosphatase regulation. J Bacteriol 174:8057–8064
    [Google Scholar]
  16. Nagy G, Dobrindt U, Schneider G, Khan A. S, Hacker J, Emody L. 2002; Loss of regulatory protein RfaH attenuates virulence of uropathogenic Escherichia coli . Infect Immun 70:4406–4413 [CrossRef]
    [Google Scholar]
  17. Nakata A, Amemura M, Shinagawa H. 1984; Regulation of the phosphate regulon in Escherichia coli K-12: regulation of the negative regulatory gene phoU and identification of the gene product. J Bacteriol 159:979–985
    [Google Scholar]
  18. Polissi A, Pontiggia A, Feger G, Altieri M, Mottl H, Ferrari L, Simon D. 1998; Large-scale identification of virulence genes from Streptococcus pneumoniae . Infect Immun 66:5620–5629
    [Google Scholar]
  19. Redford P, Roesch P. L, Welch R. A. 2003; degS is necessary for virulence and is among extraintestinal Escherichia coli genes induced in murine peritonitis. Infect Immun 71:3088–3096 [CrossRef]
    [Google Scholar]
  20. Rippere-Lampe K. E, O'Brien A. D, Conran R, Lockman H. A. 2001; Mutation of the gene encoding cytotoxic necrotizing factor type 1 (cnf [sub]1[/sub]) attenuates the virulence of uropathogenic Escherichia coli . Infect Immun 69:3954–3964 [CrossRef]
    [Google Scholar]
  21. Sambrook J, Fritsch E. F, Maniatis T. 1989 Molecular Cloning: a Laboratory Manual Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
    [Google Scholar]
  22. Snyder J. A, Haugen B. J, Buckles E. L, Lockatell C. V, Johnson D. E, Donnenberg M. S, Welch R. A, Mobley H. L. 2004; Transcriptome of uropathogenic Escherichia coli during urinary tract infection. Infect Immun 72:6373–6381 [CrossRef]
    [Google Scholar]
  23. Steed P. M, Wanner B. L. 1993; Use of the rep technique for allele replacement to construct mutants with deletions of the pstSCAB-phoU operon: evidence of a new role for the PhoU protein in the phosphate regulon. J Bacteriol 175:6797–6809
    [Google Scholar]
  24. Surin B. P, Rosenberg H, Cox G. B. 1985; Phosphate-specific transport system of Escherichia coli : nucleotide sequence and gene-polypeptide relationships. J Bacteriol 161:189–198
    [Google Scholar]
  25. Torres A. G, Redford P, Welch R. A, Payne S. M. 2001; TonB-dependent systems of uropathogenic Escherichia coli : aerobactin and heme transport and TonB are required for virulence in the mouse. Infect Immun 69:6179–6185 [CrossRef]
    [Google Scholar]
  26. Wang R. F, Kushner S. R. 1991; Construction of versatile low-copy-number vectors for cloning, sequencing and gene expression in Escherichia coli . Gene 100:195–199 [CrossRef]
    [Google Scholar]
  27. Wanner B. L. others 1996 Phosphorus assimilation and control of the phosphate regulon In Escherichia coli and Salmonella: Cellular and Molecular Biology pp. 1357–1381 Edited by Neidhardt F. C. Washington, DC: American Society for Microbiology;
    [Google Scholar]
  28. Welch R. A, Burland V, Plunkett G., III. & 16 other authors 2002; Extensive mosaic structure revealed by the complete genome sequence of uropathogenic Escherichia coli . Proc Natl Acad Sci U S A 99:17020–17024 [CrossRef]
    [Google Scholar]
http://instance.metastore.ingenta.com/content/journal/micro/10.1099/mic.0.28281-0
Loading
/content/journal/micro/10.1099/mic.0.28281-0
Loading

Data & Media loading...

This is a required field
Please enter a valid email address
Approval was a Success
Invalid data
An Error Occurred
Approval was partially successful, following selected items could not be processed due to error