HOME HELP FEEDBACK SUBSCRIPTIONS ARCHIVE SEARCH TABLE OF CONTENTS		QUICK SEARCH:   [advanced] Author: Keyword(s): Year:  Vol:  Page:

This Article Full Text Full Text (PDF) Supplementary data Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Google Scholar Google Scholar Articles by McAdam, R. A. Articles by Duncan, K. Search for Related Content PubMed PubMed Citation Articles by McAdam, R. A. Articles by Duncan, K. Agricola Articles by McAdam, R. A. Articles by Duncan, K.

Characterization of a Mycobacterium tuberculosis H37Rv transposon library reveals insertions in 351 ORFs and mutants with altered virulence^b

GlaxoSmithKline, Medicines Research Centre, Gunnels Wood Road, Stevenage SG1 2NY, UK¹
London School of Hygiene & Tropical Medicine, Keppel St, London WC1E 7HT, UK²
Albert Einstein College of Medicine, 1300 Morris Park Avenue, Bronx, NY 10461, USA³

Author for correspondence: Ken Duncan. Tel: +44 1438 763841. Fax: +44 1438 764799. e-mail: kd9430{at}gsk.com

A library of Mycobacterium tuberculosis insertional mutants was generated with the transposon Tn5370. The junction sequence between the transposon and the mycobacterial chromosome was determined, revealing the positions of 1329 unique insertions, 1189 of which were located in 351 different ORFs. Transposition was not completely random and examination of the most susceptible genome regions revealed a lower-than-average G+C content ranging from 54 to 62 mol%. Mutants were obtained in all of the recognized M. tuberculosis functional protein-coding gene classes. About 30% of the disrupted ORFs had matches elsewhere in the genome that suggested redundancy of function. The effect of gene disruption on the virulence of a selected set of defined mutants was investigated in a severe combined immune deficiency (SCID) mouse model. A range of phenotypes was observed in these mutants, the most notable being the severe attenuation in virulence of a strain disrupted in the Rv1290c gene, which encodes a protein of unknown function. The library described in this study provides a resource of defined mutant strains for use in functional analyses aimed at investigating the role of particular M. tuberculosis genes in virulence and defining their potential as targets for new anti-mycobacterial drugs or as candidates for deletion in a rationally attenuated live vaccine.

Keywords: gene disruption, severe combined immune deficiency (SCID) mouse, attenuation

Abbreviations: BCG, Bacille Calmette–Guérin; SCID, severe combined immune deficiency

^b The precise locations of all of the insertions examined in this study can be found as supplementary data in Microbiology Online (http://mic.sgmjournals.org).

^a Present address: Department of Molecular Biology and Microbiology, Tufts University, Boston, MA 02111, USA.

This article has been cited by other articles:

				T. Mogi, Y. Murase, M. Mori, K. Shiomi, S. Omura, M. P. Paranagama, and K. Kita Polymyxin B Identified as an Inhibitor of Alternative NADH Dehydrogenase and Malate: Quinone Oxidoreductase from the Gram-positive Bacterium Mycobacterium smegmatis J. Biochem., October 1, 2009; 146(4): 491 - 499. [Abstract] [Full Text] [PDF]

				J. Krawczyk, T. A. Kohl, A. Goesmann, J. Kalinowski, and J. Baumbach From Corynebacterium glutamicum to Mycobacterium tuberculosis--towards transfers of gene regulatory networks and integrated data analyses with MycoRegNet Nucleic Acids Res., August 1, 2009; 37(14): e97 - e97. [Abstract] [Full Text] [PDF]

				G. L. Newton, N. Buchmeier, and R. C. Fahey Biosynthesis and Functions of Mycothiol, the Unique Protective Thiol of Actinobacteria Microbiol. Mol. Biol. Rev., September 1, 2008; 72(3): 471 - 494. [Abstract] [Full Text] [PDF]

				A. Gurvitz, J. K. Hiltunen, and A. J. Kastaniotis Function of Heterologous Mycobacterium tuberculosis InhA, a Type 2 Fatty Acid Synthase Enzyme Involved in Extending C20 Fatty Acids to C60-to-C90 Mycolic Acids, during De Novo Lipoic Acid Synthesis in Saccharomyces cerevisiae Appl. Envir. Microbiol., August 15, 2008; 74(16): 5078 - 5085. [Abstract] [Full Text] [PDF]

				F. Laval, R. Haites, F. Movahedzadeh, A. Lemassu, C. Y. Wong, N. Stoker, H. Billman-Jacobe, and M. Daffe Investigating the Function of the Putative Mycolic Acid Methyltransferase UmaA: DIVERGENCE BETWEEN THE MYCOBACTERIUM SMEGMATIS AND MYCOBACTERIUM TUBERCULOSIS PROTEINS J. Biol. Chem., January 18, 2008; 283(3): 1419 - 1427. [Abstract] [Full Text] [PDF]

				J. Lynett and R. W. Stokes Selection of transposon mutants of Mycobacterium tuberculosis with increased macrophage infectivity identifies fadD23 to be involved in sulfolipid production and association with macrophages Microbiology, September 1, 2007; 153(9): 3133 - 3140. [Abstract] [Full Text] [PDF]

				S. J. Shin, C.-w. Wu, H. Steinberg, and A. M. Talaat Identification of Novel Virulence Determinants in Mycobacterium paratuberculosis by Screening a Library of Insertional Mutants Infect. Immun., July 1, 2006; 74(7): 3825 - 3833. [Abstract] [Full Text] [PDF]

				T. Yano, L.-S. Li, E. Weinstein, J.-S. Teh, and H. Rubin Steady-state Kinetics and Inhibitory Action of Antitubercular Phenothiazines on Mycobacterium tuberculosis Type-II NADH-Menaquinone Oxidoreductase (NDH-2) J. Biol. Chem., April 28, 2006; 281(17): 11456 - 11463. [Abstract] [Full Text] [PDF]

				Y. Hu, F. Movahedzadeh, N. G. Stoker, and A. R. M. Coates Deletion of the Mycobacterium tuberculosis {alpha}-Crystallin-Like hspX Gene Causes Increased Bacterial Growth In Vivo Infect. Immun., February 1, 2006; 74(2): 861 - 868. [Abstract] [Full Text] [PDF]

				H. Yesilkaya, J. W. Dale, N. J. C. Strachan, and K. J. Forbes Natural Transposon Mutagenesis of Clinical Isolates of Mycobacterium tuberculosis: How Many Genes Does a Pathogen Need? J. Bacteriol., October 1, 2005; 187(19): 6726 - 6732. [Abstract] [Full Text] [PDF]

				S. L. Tran and G. M. Cook The F1Fo-ATP Synthase of Mycobacterium smegmatis Is Essential for Growth J. Bacteriol., July 15, 2005; 187(14): 5023 - 5028. [Abstract] [Full Text] [PDF]

				D. M. Collins, B. Skou, S. White, S. Bassett, L. Collins, R. For, K. Hurr, G. Hotter, and G. W. de Lisle Generation of Attenuated Mycobacterium bovis Strains by Signature-Tagged Mutagenesis for Discovery of Novel Vaccine Candidates Infect. Immun., April 1, 2005; 73(4): 2379 - 2386. [Abstract] [Full Text] [PDF]

				E. A. Weinstein, T. Yano, L.-S. Li, D. Avarbock, A. Avarbock, D. Helm, A. A. McColm, K. Duncan, J. T. Lonsdale, and H. Rubin Inhibitors of type II NADH:menaquinone oxidoreductase represent a class of antitubercular drugs PNAS, March 22, 2005; 102(12): 4548 - 4553. [Abstract] [Full Text] [PDF]

				A. R. Flores, L. M. Parsons, and M. S. Pavelka Jr. Characterization of Novel Mycobacterium tuberculosis and Mycobacterium smegmatis Mutants Hypersusceptible to {beta}-Lactam Antibiotics J. Bacteriol., March 15, 2005; 187(6): 1892 - 1900. [Abstract] [Full Text] [PDF]

				R. H. Copenhaver, E. Sepulveda, L. Y. Armitige, J. K. Actor, A. Wanger, S. J. Norris, R. L. Hunter, and C. Jagannath A Mutant of Mycobacterium tuberculosis H37Rv That Lacks Expression of Antigen 85A Is Attenuated in Mice but Retains Vaccinogenic Potential Infect. Immun., December 1, 2004; 72(12): 7084 - 7095. [Abstract] [Full Text] [PDF]

				E. A. Roberts, A. Clark, S. McBeth, and R. L. Friedman Molecular Characterization of the eis Promoter of Mycobacterium tuberculosis J. Bacteriol., August 15, 2004; 186(16): 5410 - 5417. [Abstract] [Full Text] [PDF]

				S. S. Dawes, D. F. Warner, L. Tsenova, J. Timm, J. D. McKinney, G. Kaplan, H. Rubin, and V. Mizrahi Ribonucleotide Reduction in Mycobacterium tuberculosis: Function and Expression of Genes Encoding Class Ib and Class II Ribonucleotide Reductases Infect. Immun., November 1, 2003; 71(11): 6124 - 6131. [Abstract] [Full Text] [PDF]

				T. Hsu, S. M. Hingley-Wilson, B. Chen, M. Chen, A. Z. Dai, P. M. Morin, C. B. Marks, J. Padiyar, C. Goulding, M. Gingery, et al. The primary mechanism of attenuation of bacillus Calmette-Guerin is a loss of secreted lytic function required for invasion of lung interstitial tissue PNAS, October 14, 2003; 100(21): 12420 - 12425. [Abstract] [Full Text] [PDF]

				D. Schnappinger, S. Ehrt, M. I. Voskuil, Y. Liu, J. A. Mangan, I. M. Monahan, G. Dolganov, B. Efron, P. D. Butcher, C. Nathan, et al. Transcriptional Adaptation of Mycobacterium tuberculosis within Macrophages: Insights into the Phagosomal Environment J. Exp. Med., September 2, 2003; 198(5): 693 - 704. [Abstract] [Full Text] [PDF]

				J. Otero, W. R. Jacobs Jr., and M. S. Glickman Efficient Allelic Exchange and Transposon Mutagenesis in Mycobacterium avium by Specialized Transduction Appl. Envir. Microbiol., September 1, 2003; 69(9): 5039 - 5044. [Abstract] [Full Text] [PDF]

				T. Parish, D. A. Smith, S. Kendall, N. Casali, G. J. Bancroft, and N. G. Stoker Deletion of Two-Component Regulatory Systems Increases the Virulence of Mycobacterium tuberculosis Infect. Immun., March 1, 2003; 71(3): 1134 - 1140. [Abstract] [Full Text] [PDF]

				D. B. Young Mycobacteria research in the post-genomic era Microbiology, October 1, 2002; 148(10): 2915 - 2917. [Full Text] [PDF]