Transposon mutagenesis of Mb0100 at the ppe1-nrp locus in Mycobacterium bovis disrupts phthiocerol dimycocerosate (PDIM) and glycosylphenol-PDIM biosynthesis, producing an avirulent strain with vaccine properties at least equal to those of M-bovis BCG

被引:49
作者
Hotter, GS
Wards, BJ
Mouat, P
Besra, GS
Gomes, J
Singh, M
Bassett, S
Kawakami, O
Wheeler, PR
de Lisle, GW
Collins, DM
机构
[1] Wallaceville Anim Res Ctr, AgRes, Upper Hutt, New Zealand
[2] Univ Birmingham, Sch Biosci, Birmingham, W Midlands, England
[3] Vet Labs Agcy, Addlestone, Surrey, England
关键词
D O I
10.1128/JB.187.7.2267-2277.2005
中图分类号
Q93 [微生物学];
学科分类号
071005 ; 100705 ;
摘要
The unusual and complex cell wall of pathogenic mycobacteria plays a major role in pathogenesis, with specific complex lipids acting as defensive, offensive, or adaptive effectors of virulence. The phthiocerol and phthiodiolone dimycocerosate esters (PDIMs) comprise one such category of virulence-enhancing lipids. Recent work in several laboratories has established that the Mycobacterium tuberculosis fadD26-mmpL7 (Rv2930-Rv2942) locus plays a major role in PDIM biosynthesis and secretion and that PDIM is required for virulence. Here we describe two independent transposon mutants (WAg533 and WAg537) of Mycobacterium bovis, both of which carry an insertion in Mb0100 (= M. tuberculosis Rv0097) to reveal a new locus involved in PDIM biosynthesis. The mutations have a polar effect on expression of the downstream genes Mb0101, Mb0102 (fadD10), Mb0103, and Mb0104 (nrp), and Mb0100 is shown to be in an operon comprising these genes and Mb0099. Reverse transcription-PCR analysis shows elevated transcription of genes in the operon upstream from the transposon insertion sites in both mutants. Both mutants have altered colony morphology and do not synthesize PDIMs or glycosylphenol-PDIM. Both mutants are avirulent in a guinea pig model of tuberculosis, and when tested as a vaccine, WAg533 conferred protective immunity against M. bovis infection at least equal to that afforded by M. bovis bacillus Calmette-Guerin.
引用
收藏
页码:2267 / 2277
页数:11
相关论文
共 41 条
[1]   Gene knockout reveals a novel gene cluster for the synthesis of a class of cell wall lipids unique to pathogenic mycobacteria [J].
Azad, AK ;
Sirakova, TD ;
Fernandes, ND ;
Kolattukudy, PE .
JOURNAL OF BIOLOGICAL CHEMISTRY, 1997, 272 (27) :16741-16745
[2]   Targeted replacement of the mycocerosic acid synthase gene in Mycobacterium bovis BCG produces a mutant that lacks mycosides [J].
Azad, AK ;
Sirakova, TD ;
Rogers, LM ;
Kolattukudy, PE .
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA, 1996, 93 (10) :4787-4792
[3]  
Besra GS, 1998, METH MOL B, V101, P91, DOI 10.1385/0-89603-471-2:91
[4]   Identification of a peptide synthetase involved in the biosynthesis of glycopeptidolipids of Mycobacterium smegmatis [J].
Billman-Jacobe, H ;
McConville, MJ ;
Haites, RE ;
Kovacevic, S ;
Coppel, RL .
MOLECULAR MICROBIOLOGY, 1999, 33 (06) :1244-1253
[5]   Crystal structure of PapA5, a phthiocerol dimycocerosyl transferase from Mycobacterium tuberculosis [J].
Buglino, J ;
Onwueme, KC ;
Ferreras, JA ;
Quadri, LEN ;
Lima, CD .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2004, 279 (29) :30634-30642
[6]   Identification of a virulence gene cluster of Mycobacterium tuberculosis by signature-tagged transposon mutagenesis [J].
Camacho, LR ;
Ensergueix, D ;
Perez, E ;
Gicquel, B ;
Guilhot, C .
MOLECULAR MICROBIOLOGY, 1999, 34 (02) :257-267
[7]   Analysis of the phthiocerol dimycocerosate locus of Mycobacterium tuberculosis -: Evidence that this lipid is involved in the cell wall permeability barrier [J].
Camacho, LR ;
Constant, P ;
Raynaud, C ;
Lanéelle, MA ;
Triccas, JA ;
Gicquel, B ;
Daffé, M ;
Guilhot, C .
JOURNAL OF BIOLOGICAL CHEMISTRY, 2001, 276 (23) :19845-19854
[8]   Predictive, structure-based model of amino acid recognition by nonribosomal peptide synthetase adenylation domains [J].
Challis, GL ;
Ravel, J ;
Townsend, CA .
CHEMISTRY & BIOLOGY, 2000, 7 (03) :211-224
[9]   New tuberculosis vaccines based on attenuated strains of the Mycobacterium tuberculosis complex [J].
Collins, DM .
IMMUNOLOGY AND CELL BIOLOGY, 2000, 78 (04) :342-348
[10]   Different susceptibility of two animal species infected with isogenic mutants of Mycobacterium bovis identifies phoT as having roles in tuberculosis virulence and phosphate transport [J].
Collins, DM ;
Kawakami, RP ;
Buddle, BM ;
Wards, BJ ;
de Lisle, GW .
MICROBIOLOGY-SGM, 2003, 149 :3203-3212