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Tuberculosis vaccine strain Mycobacterium bovis BCG Russia is a natural recA mutant.

Keller PM, Böttger EC, Sander P - BMC Microbiol. (2008)

Bottom Line: Homologous gene replacement was achieved successfully in three out of four strains.At the protein level BCG Russia failed to express RecA.We hypothesize that recA inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to M. bovis AF2122/97 wild-type.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Gloriastrasse 30/32, CH-8006 Zürich, Switzerland. pkeller@immv.uzh.ch

ABSTRACT

Background: The current tuberculosis vaccine is a live vaccine derived from Mycobacterium bovis and attenuated by serial in vitro passaging. All vaccine substrains in use stem from one source, strain Bacille Calmette-Guérin. However, they differ in regions of genomic deletions, antigen expression levels, immunogenicity, and protective efficacy.

Results: As a RecA phenotype increases genetic stability and may contribute restricting the ongoing evolution of the various BCG substrains while maintaining their protective efficacy, we aimed to inactivate recA by allelic replacement in BCG vaccine strains representing different phylogenetic lineages (Pasteur, Frappier, Denmark, Russia). Homologous gene replacement was achieved successfully in three out of four strains. However, only illegitimate recombination was observed in BCG substrain Russia. Sequence analyses of recA revealed that a single nucleotide insertion in the 5' part of recA led to a translational frameshift with an early stop codon making BCG Russia a natural recA mutant. At the protein level BCG Russia failed to express RecA.

Conclusion: According to phylogenetic analyses BCG Russia is an ancient vaccine strain most closely related to the parental M. bovis. We hypothesize that recA inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to M. bovis AF2122/97 wild-type.

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Genealogy of BCG vaccine substrains. Genealogy of BCG vaccine substrains, modified from [6] with permission, copyright (2007) National Academy of Sciences, U.S.A., displaying the original virulent ancestor strain Mycobacterium bovis (isolated by Nocard in 1908) and the subsequent series of genomic alteration including deletions of regions of difference (RD), single nucleotide polymorphisms, and duplications of genomic regions. The recA alteration (recA_D140*), the mmaA3 point mutation [8], and the 22-bp deletion in Rv3405c in BCG Japan [39] have been added to the scheme.
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Figure 4: Genealogy of BCG vaccine substrains. Genealogy of BCG vaccine substrains, modified from [6] with permission, copyright (2007) National Academy of Sciences, U.S.A., displaying the original virulent ancestor strain Mycobacterium bovis (isolated by Nocard in 1908) and the subsequent series of genomic alteration including deletions of regions of difference (RD), single nucleotide polymorphisms, and duplications of genomic regions. The recA alteration (recA_D140*), the mmaA3 point mutation [8], and the 22-bp deletion in Rv3405c in BCG Japan [39] have been added to the scheme.

Mentions: Replication errors, transpositions and recombination events contribute to genetic alterations and drive genome evolution. For BCG various differences in morphology, growth rate, protein expression and genetic make-up have been noted among commercially available substrains [16]. This is presumably a result of the numerous serial passages on natural (potato slices trenched in ox bile) and artificial media, which have led to the acquisition of genomic alterations and further attenuation. Of note, meta-analyses of BCG vaccination trials have indicated protective efficacies ranging from 0–80%. A correlation between the number of serial in vitro passages and the decrease in protective efficacy has been observed [17]. Several reasons have been put forward to explain the varying protective efficacies of BCG, among others genetic differences between vaccine substrains as well as within an individual substrain [18]. More recently it has been suggested that the protective efficacy of ancient vaccine strains charcterized by few regions of difference (Figure 4) may be superior to that of the later ones that are more widely used [6]. We have added the newly identified single nucleotide polymorphism to an existent phylogenetic tree. It will be of interest to see whether recA is functional in other ancient strains (e.g. BCG Moreau or Japan). Sequencing of recA may also help to clarifiy the uncertain origin of the two subcultures of BCG Japan [6].


Tuberculosis vaccine strain Mycobacterium bovis BCG Russia is a natural recA mutant.

Keller PM, Böttger EC, Sander P - BMC Microbiol. (2008)

Genealogy of BCG vaccine substrains. Genealogy of BCG vaccine substrains, modified from [6] with permission, copyright (2007) National Academy of Sciences, U.S.A., displaying the original virulent ancestor strain Mycobacterium bovis (isolated by Nocard in 1908) and the subsequent series of genomic alteration including deletions of regions of difference (RD), single nucleotide polymorphisms, and duplications of genomic regions. The recA alteration (recA_D140*), the mmaA3 point mutation [8], and the 22-bp deletion in Rv3405c in BCG Japan [39] have been added to the scheme.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2483709&req=5

Figure 4: Genealogy of BCG vaccine substrains. Genealogy of BCG vaccine substrains, modified from [6] with permission, copyright (2007) National Academy of Sciences, U.S.A., displaying the original virulent ancestor strain Mycobacterium bovis (isolated by Nocard in 1908) and the subsequent series of genomic alteration including deletions of regions of difference (RD), single nucleotide polymorphisms, and duplications of genomic regions. The recA alteration (recA_D140*), the mmaA3 point mutation [8], and the 22-bp deletion in Rv3405c in BCG Japan [39] have been added to the scheme.
Mentions: Replication errors, transpositions and recombination events contribute to genetic alterations and drive genome evolution. For BCG various differences in morphology, growth rate, protein expression and genetic make-up have been noted among commercially available substrains [16]. This is presumably a result of the numerous serial passages on natural (potato slices trenched in ox bile) and artificial media, which have led to the acquisition of genomic alterations and further attenuation. Of note, meta-analyses of BCG vaccination trials have indicated protective efficacies ranging from 0–80%. A correlation between the number of serial in vitro passages and the decrease in protective efficacy has been observed [17]. Several reasons have been put forward to explain the varying protective efficacies of BCG, among others genetic differences between vaccine substrains as well as within an individual substrain [18]. More recently it has been suggested that the protective efficacy of ancient vaccine strains charcterized by few regions of difference (Figure 4) may be superior to that of the later ones that are more widely used [6]. We have added the newly identified single nucleotide polymorphism to an existent phylogenetic tree. It will be of interest to see whether recA is functional in other ancient strains (e.g. BCG Moreau or Japan). Sequencing of recA may also help to clarifiy the uncertain origin of the two subcultures of BCG Japan [6].

Bottom Line: Homologous gene replacement was achieved successfully in three out of four strains.At the protein level BCG Russia failed to express RecA.We hypothesize that recA inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to M. bovis AF2122/97 wild-type.

View Article: PubMed Central - HTML - PubMed

Affiliation: Institut für Medizinische Mikrobiologie, Universität Zürich, Gloriastrasse 30/32, CH-8006 Zürich, Switzerland. pkeller@immv.uzh.ch

ABSTRACT

Background: The current tuberculosis vaccine is a live vaccine derived from Mycobacterium bovis and attenuated by serial in vitro passaging. All vaccine substrains in use stem from one source, strain Bacille Calmette-Guérin. However, they differ in regions of genomic deletions, antigen expression levels, immunogenicity, and protective efficacy.

Results: As a RecA phenotype increases genetic stability and may contribute restricting the ongoing evolution of the various BCG substrains while maintaining their protective efficacy, we aimed to inactivate recA by allelic replacement in BCG vaccine strains representing different phylogenetic lineages (Pasteur, Frappier, Denmark, Russia). Homologous gene replacement was achieved successfully in three out of four strains. However, only illegitimate recombination was observed in BCG substrain Russia. Sequence analyses of recA revealed that a single nucleotide insertion in the 5' part of recA led to a translational frameshift with an early stop codon making BCG Russia a natural recA mutant. At the protein level BCG Russia failed to express RecA.

Conclusion: According to phylogenetic analyses BCG Russia is an ancient vaccine strain most closely related to the parental M. bovis. We hypothesize that recA inactivation in BCG Russia occurred early and is in part responsible for its high degree of genomic stability, resulting in a substrain that has less genetic alterations than other vaccine substrains with respect to M. bovis AF2122/97 wild-type.

Show MeSH
Related in: MedlinePlus