Comparative and functional genomic analyses of iron transport and regulation in Leptospira spp.

Laboratoire des Spirochètes, Institut Pasteur, 28 rue du docteur Roux, 75724 Paris Cedex 15, France.
Journal of Bacteriology (Impact Factor: 2.69). 12/2006; 188(22):7893-904. DOI: 10.1128/JB.00711-06
Source: PubMed

ABSTRACT The spirochetes of the Leptospira genus contain saprophytic and pathogenic members, the latter being responsible for leptospirosis. Despite the recent sequencing of the genome of the pathogen L. interrogans, the slow growth of these bacteria, their virulence in humans, and a lack of genetic tools make it difficult to work with these pathogens. In contrast, the development of numerous genetic tools for the saprophyte L. biflexa enables its use as a model bacterium. Leptospira spp. require iron for growth. In this work, we show that Leptospira spp. can acquire iron from different sources, including siderophores. A comparative genome analysis of iron uptake systems and their regulation in the saprophyte L. biflexa and the pathogen L. interrogans is presented in this study. Our data indicated that, for instance, L. biflexa and L. interrogans contain 8 and 12 genes, respectively, whose products share homology with proteins that have been shown to be TonB-dependent receptors. We show that some genes involved in iron uptake were differentially expressed in response to iron. In addition, we were able to disrupt several putative genes involved in iron acquisition systems or iron regulation in L. biflexa. Comparative genomics, in combination with gene inactivation, gives us significant functional information on iron homeostasis in Leptospira spp.

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    ABSTRACT: Leptospira are spirochete bacteria comprising both pathogenic and free living species. The saprophyte L. biflexa is a model bacterium for studying leptospiral biology due to relative ease of culturing and genetic manipulation. In this study, we constructed a library of 4996 random transposon mutants in L. biflexa. We screened the library for increased susceptibility to the DNA intercalating agent, ethidium bromide (EtBr), in order to identify genetic determinants that reduce L. biflexa susceptibility to antimicrobial agents. By phenotypic screening, using subinhibitory EtBr concentrations, we identified 29 genes, that when disrupted via transposon insertion, led to increased sensitivity of the bacteria to EtBr. At the functional level, these genes could be categorized in regulation and signaling (11), transport (6), membrane structure (5), stress response (2), DNA damage repair (1), other processes (3), and one gene had no predicted function. Genes involved in transport (including efflux pumps) and regulation (two-component systems, anti-sigma factor antagonists, etc.) were overrepresented, demonstrating that these genes are major contributors to EtBr tolerance. This finding suggests that transport genes which would prevent EtBr to enter the cell cytoplasm are critical for EtBr resistance. We identified genes required for the growth of L. biflexa in the presence of sublethal EtBr concentration, and characterized their potential as antibiotic resistance determinants. This study will help to delineate mechanisms of adaptation to toxic compounds, as well as potential mechanisms of antibiotic resistance development in pathogenic L. interrogans.
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