title: Iron Acquisition in Mycobacterium avium subsp. paratuberculosis.
abstract: Mycobacterium avium subsp. paratuberculosis is a host-adapted pathogen that evolved from the environmental bacterium M. avium subsp. hominissuis through gene loss and gene acquisition. Growth of M. avium subsp. paratuberculosis in the laboratory is enhanced by supplementation of the media with the iron-binding siderophore mycobactin J. Here we examined the production of mycobactins by related organisms and searched for an alternative iron uptake system in M. avium subsp. paratuberculosis. Through thin-layer chromatography and radiolabeled iron-uptake studies, we showed that M. avium subsp. paratuberculosis is impaired for both mycobactin synthesis and iron acquisition. Consistent with these observations, we identified several mutations, including deletions, in M. avium subsp. paratuberculosis genes coding for mycobactin synthesis. Using a transposon-mediated mutagenesis screen conditional on growth without myobactin, we identified a potential mycobactin-independent iron uptake system on a M. avium subsp. paratuberculosis-specific genomic island, LSP(P)15. We obtained a transposon (Tn) mutant with a disruption in the LSP(P)15 gene MAP3776c for targeted study. The mutant manifests increased iron uptake as well as intracellular iron content, with genes downstream of the transposon insertion (MAP3775c to MAP3772c [MAP3775-2c]) upregulated as the result of a polar effect. As an independent confirmation, we observed the same iron uptake phenotypes by overexpressing MAP3775-2c in wild-type M. avium subsp. paratuberculosis. These data indicate that the horizontally acquired LSP(P)15 genes contribute to iron acquisition by M. avium subsp. paratuberculosis, potentially allowing the subsequent loss of siderophore production by this pathogen.
  
  
  OBJECTIVE
  Many microbes are able to scavenge iron from their surroundings by producing iron-chelating siderophores. One exception is Mycobacterium avium subsp. paratuberculosis, a fastidious, slow-growing animal pathogen whose growth needs to be supported by exogenous mycobacterial siderophore (mycobactin) in the laboratory. Data presented here demonstrate that, compared to other closely related M. avium subspecies, mycobactin production and iron uptake are different in M. avium subsp. paratuberculosis, and these phenotypes may be caused by numerous deletions in its mycobactin biosynthesis pathway. Using a genomic approach, supplemented by targeted genetic and biochemical studies, we identified that LSP(P)15, a horizontally acquired genomic island, may encode an alternative iron uptake system. These findings shed light on the potential physiological consequence of horizontal gene transfer in M. avium subsp. paratuberculosis evolution.
authors.affiliation: Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada McGill International TB Centre, Montreal, Quebec, Canada.
authors.forename: Joyce
authors.initials: J
authors.lastname: Wang
authors.affiliation: Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada.
authors.forename: Jalal
authors.initials: J
authors.lastname: Moolji
authors.affiliation: Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada.
authors.forename: Alex
authors.initials: A
authors.lastname: Dufort
authors.affiliation: McGill University and Genome Quebec Innovation Centre, Montreal, Quebec, Canada.
authors.forename: Alfredo
authors.initials: A
authors.lastname: Staffa
authors.affiliation: Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada McGill International TB Centre, Montreal, Quebec, Canada.
authors.forename: Pilar
authors.initials: P
authors.lastname: Domenech
authors.affiliation: Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada McGill International TB Centre, Montreal, Quebec, Canada Department of Medicine, McGill University, Montreal, Quebec, Canada.
authors.forename: Michael B
authors.initials: MB
authors.lastname: Reed
authors.affiliation: Department of Microbiology and Immunology, McGill University, Montreal, Quebec, Canada Research Institute of the McGill University Health Centre, Montreal, Quebec, Canada McGill International TB Centre, Montreal, Quebec, Canada Department of Medicine, McGill University, Montreal, Quebec, Canada marcel.behr@mcgill.ca.
authors.forename: Marcel A
authors.initials: MA
authors.lastname: Behr
mesh_terms: D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D001692:Biological Transport; D015964:Gene Expression Regulation, Bacterial / Q000502:physiology; D007501:Iron / Q000378:metabolism*; D009154:Mutation; D016927:Mycobacterium avium subsp. paratuberculosis / Q000378:metabolism*; D010080:Oxazoles / Q000378:metabolism*
chemical_list: D001426:Bacterial Proteins; D010080:Oxazoles; C018608:mycobactins; D007501:Iron