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Journal of Bacteriology, May 2005, p. 3249-3254, Vol. 187, No. 9
0021-9193/05/$08.00+0     doi:10.1128/JB.187.9.3249-3254.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.

Isolation and Characterization of FecA- and FeoB-Mediated Iron Acquisition Systems of the Spirochete Leptospira biflexa by Random Insertional Mutagenesis

Hélène Louvel, Isabelle Saint Girons, and Mathieu Picardeau*

Laboratoire des Spirochètes, Institut Pasteur, Paris, France

Received 17 September 2004/ Accepted 28 January 2005

The specific mechanisms by which Leptospira spp. acquire iron from their ecological niches are unknown. A major factor contributing to our ignorance of spirochetal biology is the lack of methods for genetic analysis of these organisms. In this study, we have developed a system for random transposon mutagenesis of Leptospira biflexa using a mariner transposon, Himar1. To demonstrate the validity of Himar1 in vivo transposon mutagenesis in L. biflexa, a screen of mutants for clones impaired in amino acid biosynthesis was first performed, enabling the identification of tryptophan and glutamate auxotrophs. To investigate iron transporters, 2,000 L. biflexa transposon mutants were screened onto media with and without hemin, thus allowing the identification of five hemin-requiring mutants, and the putative genes responsible for this phenotype were identified. Three mutants had distinct insertions in a gene encoding a protein which shares homology with the TonB-dependent receptor FecA, involved in ferric citrate transport. We also identified two mutants with a Himar1 insertion into a feoB-like gene, the product of which is required for ferrous iron uptake in many bacterial organisms. Interestingly, the growth inhibition exhibited by the fecA and feoB mutants was relieved by deferoxamine, suggesting the presence of a ferric hydroxamate transporter. These results confirm the importance of iron for the growth of Leptospira and its ability to use multiple iron sources.

* Corresponding author. Mailing address: Laboratoire des Spirochètes, Institut Pasteur, 28 rue du docteur Roux, 75724 Paris Cedex 15, France. Phone: 33 (1) 45 68 83 68. Fax: 33 (1) 40 61 30 01. E-mail: mpicard{at}pasteur.fr.

Journal of Bacteriology, May 2005, p. 3249-3254, Vol. 187, No. 9
0021-9193/05/$08.00+0     doi:10.1128/JB.187.9.3249-3254.2005
Copyright © 2005, American Society for Microbiology. All Rights Reserved.



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