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Novel genome polymorphisms in BCG vaccine strains and impact on efficacy.

Leung AS, Tran V, Wu Z, Yu X, Alexander DC, Gao GF, Zhu B, Liu J - BMC Genomics (2008)

Bottom Line: Moreover, we have uncovered various polymorphisms in the phoP-phoR locus.Our study demonstrates that major virulence factors are different among BCG strains, which provide molecular mechanisms for important vaccine phenotypes including adverse effect profile, tuberculin reactivity and protective efficacy.These findings have important implications for the development of a new generation of vaccines.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Genetics, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada. andreas.leung@utoronto.ca

ABSTRACT
Bacille Calmette-Guérin (BCG) is an attenuated strain of Mycobacterium bovis currently used as a vaccine against tuberculosis. Global distribution and propagation of BCG has contributed to the in vitro evolution of the vaccine strain and is thought to partially account for the different outcomes of BCG vaccine trials. Previous efforts by several molecular techniques effectively identified large sequence polymorphisms among BCG daughter strains, but lacked the resolution to identify smaller changes. In this study, we have used a NimbleGen tiling array for whole genome comparison of 13 BCG strains. Using this approach, in tandem with DNA resequencing, we have identified six novel large sequence polymorphisms including four deletions and two duplications in specific BCG strains. Moreover, we have uncovered various polymorphisms in the phoP-phoR locus. Importantly, these polymorphisms affect genes encoding established virulence factors including cell wall complex lipids, ESX secretion systems, and the PhoP-PhoR two-component system. Our study demonstrates that major virulence factors are different among BCG strains, which provide molecular mechanisms for important vaccine phenotypes including adverse effect profile, tuberculin reactivity and protective efficacy. These findings have important implications for the development of a new generation of vaccines.

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IS6110 insertion in the phoP promoter in BCG-Russia, -Moreau, and -Japan. (A) Schematic representation of the phoP-phoR locus with IS6110 inserted in an inverse orientation 18 bp upstream from phoP start codon. (B) Nucleotide sequence surrounding IS6110. The IS6110 sequence is boxed. The GAA direct repeats flanking the IS6110 insertion site is underlined and in boldface. The ATG start codons of phoP and phoR are indicated by arrows and in boldface.
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Figure 2: IS6110 insertion in the phoP promoter in BCG-Russia, -Moreau, and -Japan. (A) Schematic representation of the phoP-phoR locus with IS6110 inserted in an inverse orientation 18 bp upstream from phoP start codon. (B) Nucleotide sequence surrounding IS6110. The IS6110 sequence is boxed. The GAA direct repeats flanking the IS6110 insertion site is underlined and in boldface. The ATG start codons of phoP and phoR are indicated by arrows and in boldface.

Mentions: Our sequence analysis revealed a number of polymorphisms in the phoP-phoR locus in various BCG strains compared to the genome sequence of M. bovis. The three early BCG substrains, BCG-Russia, -Japan, and -Moreau, contain an identical IS6110 (1,356 bp) insertion at nucleotide 851593 of the M. tb genome, which is 18 bp upstream of the start codon of phoP (Fig. 2). This IS6110 element is identical to many other copies of IS6110 found in various locations in the M. tb genome. It is flanked by a 3-bp direct repeat (GAA) on both sides and is in an inverse orientation of phoP-phoR (Fig. 2). The presence of an IS6110 element in the promoter region of phoP in BCG-Russia, -Japan, and -Moreau has been described previously, but its insertion site and orientation were not determined until now [49]. Although not present in M. tb H37Rv or M. bovis AF2122/97, an IS6110 insertion in the phoP promoter was found in a clinical strain of M. bovis termed B strain, which was responsible for a severe nosocomial outbreak of multidrug resistant TB in humans in Spain [50,51]. However, unlike the three BCG strains, the IS6110 insertion in the M. bovis B strain is located at 75 bp upstream of the start codon of phoP and is in the same orientation as phoP-phoR [50]. The potential effect of IS6110 on phoP expression is described in the 'Discussion' section.


Novel genome polymorphisms in BCG vaccine strains and impact on efficacy.

Leung AS, Tran V, Wu Z, Yu X, Alexander DC, Gao GF, Zhu B, Liu J - BMC Genomics (2008)

IS6110 insertion in the phoP promoter in BCG-Russia, -Moreau, and -Japan. (A) Schematic representation of the phoP-phoR locus with IS6110 inserted in an inverse orientation 18 bp upstream from phoP start codon. (B) Nucleotide sequence surrounding IS6110. The IS6110 sequence is boxed. The GAA direct repeats flanking the IS6110 insertion site is underlined and in boldface. The ATG start codons of phoP and phoR are indicated by arrows and in boldface.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2553098&req=5

Figure 2: IS6110 insertion in the phoP promoter in BCG-Russia, -Moreau, and -Japan. (A) Schematic representation of the phoP-phoR locus with IS6110 inserted in an inverse orientation 18 bp upstream from phoP start codon. (B) Nucleotide sequence surrounding IS6110. The IS6110 sequence is boxed. The GAA direct repeats flanking the IS6110 insertion site is underlined and in boldface. The ATG start codons of phoP and phoR are indicated by arrows and in boldface.
Mentions: Our sequence analysis revealed a number of polymorphisms in the phoP-phoR locus in various BCG strains compared to the genome sequence of M. bovis. The three early BCG substrains, BCG-Russia, -Japan, and -Moreau, contain an identical IS6110 (1,356 bp) insertion at nucleotide 851593 of the M. tb genome, which is 18 bp upstream of the start codon of phoP (Fig. 2). This IS6110 element is identical to many other copies of IS6110 found in various locations in the M. tb genome. It is flanked by a 3-bp direct repeat (GAA) on both sides and is in an inverse orientation of phoP-phoR (Fig. 2). The presence of an IS6110 element in the promoter region of phoP in BCG-Russia, -Japan, and -Moreau has been described previously, but its insertion site and orientation were not determined until now [49]. Although not present in M. tb H37Rv or M. bovis AF2122/97, an IS6110 insertion in the phoP promoter was found in a clinical strain of M. bovis termed B strain, which was responsible for a severe nosocomial outbreak of multidrug resistant TB in humans in Spain [50,51]. However, unlike the three BCG strains, the IS6110 insertion in the M. bovis B strain is located at 75 bp upstream of the start codon of phoP and is in the same orientation as phoP-phoR [50]. The potential effect of IS6110 on phoP expression is described in the 'Discussion' section.

Bottom Line: Moreover, we have uncovered various polymorphisms in the phoP-phoR locus.Our study demonstrates that major virulence factors are different among BCG strains, which provide molecular mechanisms for important vaccine phenotypes including adverse effect profile, tuberculin reactivity and protective efficacy.These findings have important implications for the development of a new generation of vaccines.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Molecular Genetics, University of Toronto, 1 King's College Circle, Toronto, Ontario, Canada. andreas.leung@utoronto.ca

ABSTRACT
Bacille Calmette-Guérin (BCG) is an attenuated strain of Mycobacterium bovis currently used as a vaccine against tuberculosis. Global distribution and propagation of BCG has contributed to the in vitro evolution of the vaccine strain and is thought to partially account for the different outcomes of BCG vaccine trials. Previous efforts by several molecular techniques effectively identified large sequence polymorphisms among BCG daughter strains, but lacked the resolution to identify smaller changes. In this study, we have used a NimbleGen tiling array for whole genome comparison of 13 BCG strains. Using this approach, in tandem with DNA resequencing, we have identified six novel large sequence polymorphisms including four deletions and two duplications in specific BCG strains. Moreover, we have uncovered various polymorphisms in the phoP-phoR locus. Importantly, these polymorphisms affect genes encoding established virulence factors including cell wall complex lipids, ESX secretion systems, and the PhoP-PhoR two-component system. Our study demonstrates that major virulence factors are different among BCG strains, which provide molecular mechanisms for important vaccine phenotypes including adverse effect profile, tuberculin reactivity and protective efficacy. These findings have important implications for the development of a new generation of vaccines.

Show MeSH
Related in: MedlinePlus