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Volume 150, Issue 4

Low-proline environments impair growth, proline transport and survival of strain-specific mutants Free

Abstract

is a common cause of disease in humans, particularly in hospitalized patients. This species needs to import several amino acids to survive, including proline. Previously, it was shown that an insertion mutation in the high-affinity proline uptake gene in strain RN6390 affected proline uptake by the bacteria as well as reducing their ability to survive . To further delineate the effect of the mutation on growth of strain RN6390, a proline uptake assay that spanned less than 1 min was done to measure transport. An eightfold difference in proline levels was observed between the wild-type strain and the high-affinity proline transport mutant strain after 15 s, indicating that the defect was only in proline transport and not a combination of proline transport, metabolism and accumulation that would have been assessed with longer assays. A mutant of strain RN4220 was then grown in minimal medium with different concentrations of proline. When compared to the wild-type strain, the mutant strain was significantly growth impaired when the level of proline was decreased to 1·74 μM. An assessment of proline concentrations in mouse livers and spleens showed proline concentrations of 7·5 μmol per spleen and 88·4 μmol per liver. To verify that the effects on proline transport and bacterial survival were indeed caused solely by a mutation in , the mutation was complemented by cloning a full-length gene on a plasmid that replicates in . Complementation of the mutant strains restored proline transport, growth in low-proline medium, and survival within mice. These results show that the mutation in led to attenuated growth in low-proline media and by corollary low-proline murine organ tissues due to less efficient transport of proline into the bacteria.

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2004-04-01
2024-04-19
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References

  1. Bae J.-H., Miller K. J. 1992; Identification of two proline transport systems in Staphylococcus aureus and their possible roles in osmoregulation. Appl Environ Microbiol 58:471–475
     [Google Scholar]
  2. Bayer A. S., Coulter S. N., Stover C. K., Schwan W. R. 1999; Impact of the high-affinity proline permease gene (putP) on the virulence of Staphylococcus aureus in experimental endocarditis. Infect Immun 67:740–744
     [Google Scholar]
  3. Boyce J. M. 1997; Epidemiology and prevention of nosocomial infections. In The Staphylococci in Human Disease pp. 309–328Edited by Crossley K. B., Archer G. L. New York: Churchill Livingstone;
     [Google Scholar]
  4. Bradford M. M. 1976; A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein–dye binding. Anal Biochem 72:248–254 [CrossRef]
     [Google Scholar]
  5. Centers for Disease Control & Prevention; 1997; Staphylococcus aureus with reduced susceptibility to vancomycin – United States, 1997. Morbid Mortal Weekly Rep 46:765–766
     [Google Scholar]
  6. Centers for Disease Control & Prevention 1999; Four pediatric deaths from community acquired methicillin-resistant Staphylococcus aureus – Minnesota and North Dakota, 1997–1999. Morbid Mortal Weekly Rep 48:707–710
     [Google Scholar]
  7. Chambers S. T., Kunin C. M. 1987; Isolation of glycine betaine and proline betaine from human urine. J Clin Invest 79:731–737 [CrossRef]
     [Google Scholar]
  8. Coulter S. N., Schwan W. R., Ng E. Y. & 7 other authors; 1998; Staphylococcus aureus genetic loci impacting growth and survival in multiple infection environments. Mol Microbiol 30:393–404 [CrossRef]
     [Google Scholar]
  9. 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]
  10. Durack D. T., Beeson P. B. 1972; Experimental bacterial endocarditis. II. Survival of bacteria in endocardial vegetations. Br J Exp Pathol 53:50–53
     [Google Scholar]
  11. Easmon C. S. F., Adlam C. 1983 Staphylococci and Staphylococcal Infections New York: Academic Press;
  12. Gladstone G. P. 1937; The nutrition of Staphylococcus aureus: nitrogen requirements. Br J Exp Pathol 19:208–226
     [Google Scholar]
  13. Iandolo J. J., Kraemer G. R. 1990; High frequency transformation of Staphylococcus aureus by electroporation. Curr Microbiol 21:373–376 [CrossRef]
     [Google Scholar]
  14. Inoue H., Date Y., Kohashi K., Yoshitomi H., Tsuruta Y. 1996; Determination of total hydroxyproline and proline in human serum and urine by HPLC with fluorescence detection. Biol Pharm Bull 19:163–166 [CrossRef]
     [Google Scholar]
  15. Inoue H., Iguchi H., Kono A., Tsuruta Y. 1999; Highly sensitive determination of N-terminal prolyl dipeptides, proline and hydroxyproline in urine by high-performance liquid chromatography using a new fluorescent labeling reagent, 4-(5,6-dimethoxy-2-phthalimidinyl)-2-methoxyphenylsulfonyl chloride. J Chromatography 724:221–230 [CrossRef]
     [Google Scholar]
  16. Kauffman C. A., Bradley S. F. 1997; Epidemiology of community-acquired infection. In The Staphylococci in Human Disease pp. 287–308Edited by Crossley K. B., Archer G. L. New York: Churchill Livingstone;
     [Google Scholar]
  17. Kloos W. E., Pattee P. A. 1965; Transduction analysis of the histidine region in Staphylococcus aureus. J Gen Microbiol 39:195–207 [CrossRef]
     [Google Scholar]
  18. Luchansky J. B., Muriana P. M., Klaenhammer T. R. 1988; Application of electroporation for transfer of plasmid DNA to Lactobacillus, Lactococcus, Leuconostoc, Listeria, Pediococcus, Bacillus, Staphylococcus, Enterococcus and Propionibacterium. Mol Microbiol 2:637–646 [CrossRef]
     [Google Scholar]
  19. NNIS 1996; National Nosocomial Infections Surveillance (NNIS) report, data summary from October 1986–April 1996, issued May 1996. A report from the National Nosocomial Infections Surveillance (NNIS) System. Am J Infect Control 24:380–388 [CrossRef]
     [Google Scholar]
  20. Novick R. P. 1990; The Staphylococcus as a molecular genetic system. In Molecular Biology of the Staphylococci pp. 1–40Edited by Novick R. P. New York: VCH Publishers;
     [Google Scholar]
  21. Panlilio A. L., Culver D. H., Gaynes R. P., Banerjee S., Henderson T. S., Tolson J. S., Martone W. J. 1992; Methicillin-resistant Staphylococcus aureus in U. S. Hospitals 1975–1991 Infect Control Hosp Epidemiol 13:582–586 [CrossRef]
     [Google Scholar]
  22. Pourkomailian B., Booth I. R. 1994; Glycine betaine transport by Staphylococcus aureus: evidence for feedback regulation of the activity of the two transport systems. Microbiology 140:3131–3138 [CrossRef]
     [Google Scholar]
  23. Reed L. J., Meunch H. 1938; A simple method of estimating fifty per cent endpoints. Am J Hyg 27:493–497
     [Google Scholar]
  24. Rudin L., Sjostrom J. E., Lindberg M., Philipson L. 1974; Factors affecting competence for transformation in Staphylococcus aureus. J Bacteriol 118:155–164
     [Google Scholar]
  25. Sambrook J., Fritsch E. F., Maniatis T. 1989 Molecular Cloning: a Laboratory Manual, 2nd edn. Cold Spring Harbor, NY: Cold Spring Harbor Laboratory;
  26. Schwan W. R., Seifert H. S., Duncan J. L. 1992; Growth conditions mediate differential transcription of fim genes involved in phase variation of type 1 pili. J Bacteriol 174:2367–2375
     [Google Scholar]
  27. Schwan W. R., Demuth A., Kuehn M., Goebel W. 1994; Phosphatidylinositol-specific phospholipase C from Listeria monocytogenes contributes to intracellular survival and growth of Listeria innocua. Infect Immun 62:4795–4803
     [Google Scholar]
  28. Schwan W. R., Coulter S. N., Ng E. Y. W. & 7 other authors; 1998; Identification and characterization of the PutP proline permease that contributes to in vivo survival of Staphylococcus aureus in animal models. Infect Immun 66:567–572
     [Google Scholar]
  29. Schweizer H. P. 1993; Small broad-host-range gentamycin resistance gene cassettes for site-specific insertion and deletion mutagenesis. BioTechniques 15:831–834
     [Google Scholar]
  30. Sieradzki K., Roberts R. B., Haber S. W., Tomasz A. 1999; The development of vancomycin resistance in a patient with methicillin-resistant Staphylococcus aureus infection. N Engl J Med 340:517–523 [CrossRef]
     [Google Scholar]
  31. Smith T. L., Pearson M. L., Wilcox K. R. & 8 other authors; 1999; Emergence of vancomycin resistance in Staphylococcus aureus. N Engl J Med 340:493–501 [CrossRef]
     [Google Scholar]
  32. Townsend D. E., Wilkinson B. J. 1992; Proline transport in Staphylococcus aureus: a high affinity system and a low affinity system involved in osmoregulation. J Bacteriol 174:2702–2710
     [Google Scholar]
  33. Wengender P. A., Miller K. J. 1995; Identification of a PutP proline permease gene homolog in Staphylococcus aureus by expression cloning of the high-affinity proline transport system in Escherichia coli. Appl Environ Microbiol 61:252–259
     [Google Scholar]
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Low-proline environments impair growth, proline transport and in vivo survival of Staphylococcus aureus strain-specific putP mutants
Microbiology 150, 1055 (2004); https://doi.org/10.1099/mic.0.26710-0
/content/journal/micro/10.1099/mic.0.26710-0
/content/journal/micro/10.1099/mic.0.26710-0
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