, Volume 17, Issue 6, pp 725–733 | Cite as

Identification of Fur regulated genes in the bacterial fish pathogen Photobacterium damselae ssp. piscicida using the Fur titration assay

  • Carlos R. Osorio
  • Manuel L. Lemos
  • Volkmar Braun


Bacteria have developed a series of iron-scavenging and transport systems. The expression of many of the iron utilization genes is tightly regulated by the Fe2+ loaded Fur repressor protein. In this study, the Fur titration assay (FURTA) was used to screen for DNA fragments from a genomic DNA library of Photobacterium damselae ssp. piscicida containing potential Fe2 +Fur binding sites or iron binding-proteins which withdraw iron from Fur. One of the clones encoded a tonB gene and adjacent a functionally related exbB gene. An additional and complete tonB exbB exbD gene cluster was identified and sequenced. A gene homologous to the ferritin gene was found whose FURTA-positive phenotype may be explained by its iron-binding ability. Genes encoding a putative complete iron-regulated outer membrane transport protein and a pseudogene of a transport protein were found. The FURTA assay also revealed iron regulation of the AraC type transcriptional regulation.

fur iron FURTA Photobacterium damselae 


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ankenbauer RG, Quan HN. 1994 FptA, the Fe(III)-pyochelin receptor of Pseudomonas aeruginosa: a phenolate siderophore receptor homologous to hydroxamate siderophore receptors. J Bacteriol 176, 307–319.Google Scholar
  2. Bobrov AG, Geoffroy VA, and Perry RD. 2002 Yersiniabactin production requires the thioesterase domain of HMWP2 and YbtD, a putative phosphopantetheinylate transferase. Infect Immun 70, 4204–4214.Google Scholar
  3. Braun V, Hantke K. 1991 Genetics of bacterial iron transport. In: Winkelmann G, ed. CRC Handbook of microbial iron chelates. Boca Raton: CRC Press: 107–138.Google Scholar
  4. Chen CY, Wu KM, Chang YC, et al. 2003 Comparative genome analysis of Vibrio vulnificus, a marine pathogen. Genome Res 13, 2577–2587.Google Scholar
  5. Crosa JH. 1997 Signal transduction and transcriptional and posttran-scriptional control of iron-regulated genes in bacteria. Microbiol Mol Biol Rev 61, 319–336.Google Scholar
  6. Do Vale A, Ellis AE, Silva MT. 2001 Electron microscopic evidence that expression of capsular polysaccharide by Photobacterium damselaesubsp. piscicidais dependent on iron availability and growth phase. Dis Aquat Org 44, 237–240.Google Scholar
  7. Fassbinder F, van Vliet AHM, Gimmel V, Kusters JG, Kist M, Bereswill S. 2000 Identification of iron-regulated genes of Helicobacter pyloriby a modified Fur titration assay (FURTA-Hp). FEMS Microbiol Lett 184, 225–229.Google Scholar
  8. Fetherston JD, Bearden SW, and Perry RD. 1996 YbtA, an AraC-type regulator of the Yersinia pestispesticin/yersiniabactin receptor. Mol Microbiol 22, 315–325.Google Scholar
  9. Fetherston JD, Bertolino VJ, and Perry RD. 1999 YbtP and YbtQ: two ABC transporters required for iron uptake in Yersinia pestis. Mol Microbiol 32, 289–299.Google Scholar
  10. Gallegos MT, Michán C, and Ramos JL. 1993 The XylS/AraC family of regulators. Nucl Acids Res 21, 807–810.Google Scholar
  11. Hantke K. 1981 Regulation of ferric iron transport in Escherichia colik-12: isolation of a constitutive mutant. Mol Gen Genet 182, 288–292.Google Scholar
  12. Hantke K. 1987 Selection procedure for deregulated iron transport mutants (fur)inEscherichia colik12: furnot only affects iron metabolism. Mol Gen Genet 210, 135–139.Google Scholar
  13. Heidelberg JF, Eisen JA, Nelson WC, et al. DNA sequence of both chromosomes of the cholera pathogen Vibrio cholerae. Nature 406, 477–483, 2000.Google Scholar
  14. Heinrichs DE, Poole K. 1993 Cloning and sequence analysis of a gene (pchr) encoding an AraC family activator of pyochelin and ferripyochelin receptor synthesis in Pseudomonas aeruginosa. J Bacteriol 175, 5882–5889.Google Scholar
  15. Juíz-Río S, Osorio CR, Lemos ML. 2004 Identification and char-acterisation of the furgenes in Photobacterium damselaessp. piscicidaand ssp. damselae. Dis Aquat Org 58, 157–164.Google Scholar
  16. Magariños B, Santos Y, Romalde JL, Rivas C, Barja JL, Toranzo AE. 1992 Pathogenic activities of the live cells and extracellu-lar products of the fish pathogen Pasteurella piscicida. J Gen Microbiol 138, 2491–2498.Google Scholar
  17. Magariños B, Romalde JL, Lemos ML, Barja JL, Toranzo AE. 1994 Iron uptake by Pasteurella piscicidaand its role in pathogenicity for fish. Appl Environ Microbiol 60, 2990–2998.Google Scholar
  18. Magariños B, Bonet R, Romalde JL, Martínez MJ, Congregado F, Toranzo AE. 1996a Influence of the capsular layer on the virulence of Pasteurella piscicidafor fish. Microb Pathog 21, 289–297.Google Scholar
  19. Magariños B, Toranzo AE, Romalde JL. 1996b Phenotypic and pathobiological characteristics of Pasteurella piscicida. Annu Rev Fish Dis 6, 41–64.Google Scholar
  20. Makino K, Oshima K, Kurokawa K, et al. 2003 Genome sequence of Vibrio parahaemolyticus: a pathogenic mechanism distinct from that of iV. cholerae. Lancet 361, 743–749.Google Scholar
  21. Matthijs S, Baysse C, Koedam N, Tehrani KA, Verheyden L, Budzikiewicz H, Schafer M, Hoorelbeke B, Meyer JM, De Greve H, and Cornelis P. 2004 The Pseudomonassiderophore quino-lobactin is synthesized from xanthurenic acid, an intermediate of the kynurenine pathway. Mol Microbiol 52, 371–384.Google Scholar
  22. Neilands JB. 1990 Molecular aspects of regulation of high affinity iron absortion in microorganisms. Adv Inorg Biochem 8, 63–90.Google Scholar
  23. Neilands JB. 1995 Siderophores: structure and function of microbial iron transport compounds. J BiolChem 270, 26723–26726.Google Scholar
  24. Occhino DA, Wyckoff EE, Henderson DP, Wrona TJ, Payne SM. 1998 Vibrio choleraeiron transport: haem transport genes are linked to one of two sets of tonB, exbB, exbDgenes. Mol Microbiol 29, 1493–1507.Google Scholar
  25. O'Malley SM, Mouton SL, Occhino DA, et al. 1999 Comparison of the heme iron utilization systems of pathogenic vibrios. J Bacteriol 181, 3594–3598.Google Scholar
  26. Panina EM, Mironov AA, and Gelfand MS. 2001 Comparative ana-lysis of Fur regulons in gamma-proteobacteria. Nuc Acids Res 29, 5195–5206.Google Scholar
  27. Sambrook J, Rusell DW. 2001 Molecular cloning: a laborat-ory manual.cold Spring Harbor, N. Y.: Cold Spring Harbor Laboratory Press.Google Scholar
  28. Sauer U, Hantke K, and Braun V. 1990 Sequence of the FhuE outer-membrane receptor gene of Escherichia colik-12 and properties of mutants. Mol Microbiol 4, 427–437.Google Scholar
  29. Stojiljkovic I, Bäumler AJ, Hantke K. 1994 Fur regulon in Gram-negative Bacteria. Identification and characterization of new iron-regulated Escherichia coligenes by a Fur titration assay. J Mol Biol 236, 531–545.Google Scholar
  30. Tabor S, Richardson CC. 1985 A bacteriophage T7 RNA polymerase-promoter system for controlled exclusive expression of specific genes. Proc Natl Acad Sci USa. 82, 1074–1078.Google Scholar
  31. van Vliet AHM, Wooldridge KG, Ketley JM. 1998 Iron-responsive gene regulation in a Campylobacter jejuni furmutant. J Bacteriol 180, 5291–5298.Google Scholar
  32. Wertheimer AM, Verweij W, Chen Q, et al. 1999 Characterization of the angRgene of Vibrio anguillarum: essential role in virulence. Infect Immun 67, 6496–6509.Google Scholar

Copyright information

© Kluwer Academic Publishers 2004

Authors and Affiliations

  • Carlos R. Osorio
    • 1
    • 2
  • Manuel L. Lemos
    • 1
  • Volkmar Braun
    • 2
  1. 1.Departamento de Microbioloxía e Parasitoloxía, Instituto de AcuiculturaUniversidade de Santiago de Compostela, 15782, Santiago de CompostelaGaliciaSpain
  2. 2.Mikrobiologie/MembranphysiologieUniversität TübingenTübingenGermany

Personalised recommendations