Exposing the third chromosome of <i>Burkholderia cepacia</i> complex strains as a virulence plasmid

Agnoli, Kirsty; Schwager, Stephan; Uehlinger, Susanne; Vergunst, Annette C.; Viteri, DF; Nguyen, D.T.; Sokol, Pamela A.; Carlier, Aurélien; Eberl, Leo

doi:10.1111/j.1365-2958.2011.07937.x

Cited by 90 publications

(139 citation statements)

References 61 publications

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“…The two largest of these major replicons are chromosomes 1 and 2, which range in size from 3.3 to 3.9 Mb and 2.4 to 3.6 Mb, respectively, in the sequenced Bcc strains. The third replicon ranges in size from 0.5 to 1.4 Mb in sequenced Bcc strains, and we have shown that this element is not a bacterial chromosome, as was previously believed, but is in fact a megaplasmid involved in virulence and antifungal activity (6). This third replicon was originally designated chromosome 3; however, we have proposed the name pC3 to reflect its true status as a megaplasmid.…”

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confidence: 95%

The Third Replicon of Members of the Burkholderia cepacia Complex, Plasmid pC3, Plays a Role in Stress Tolerance

Agnoli

Frauenknecht

Freitag

et al. 2014

Appl Environ Microbiol

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The metabolically versatile Burkholderia cepacia complex (Bcc) occupies a variety of niches, including the plant rhizosphere and the cystic fibrosis lung (where it is often fatal to the patient). Bcc members have multipartite genomes, of which the third replicon, pC3 (previously chromosome 3), has been shown to be a nonessential megaplasmid which confers virulence and both antifungal and proteolytic activity on several strains. In this study, pC3 curing was extended to cover strains of 16 of the 17 members of the Bcc, and the phenotypes conferred by pC3 were determined. B. cenocepacia strains H111, MCO-3, and HI2424 were previously cured of pC3; however, this had not proved possible in the epidemic strain K56-2. Here, we investigated the mechanism of this unexpected stability and found that efficient toxin-antitoxin systems are responsible for maintaining pC3 of strain K56-2. Identification of these systems allowed neutralization of the toxins and the subsequent deletion of K56-2pC3. The cured strain was found to exhibit reduced antifungal activity and was attenuated in both the zebrafish and the Caenorhabditis elegans model of infection. We used a PCR screening method to examine the prevalence of pC3 within 110 Bcc isolates and found that this replicon was absent in only four cases, suggesting evolutionary fixation. It is shown that plasmid pC3 increases the resistance of B. cenocepacia H111 to various stresses (oxidative, osmotic, high-temperature, and chlorhexidine-induced stresses), explaining the prevalence of this replicon within the Bcc.

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confidence: 95%

The Third Replicon of Members of the Burkholderia cepacia Complex, Plasmid pC3, Plays a Role in Stress Tolerance

Agnoli

Frauenknecht

Freitag

et al. 2014

Appl Environ Microbiol

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“…To our surprise, only one of the disrupted genes coded for a previously described virulence factor (the QS regulator CepR), while 19 of the mutations were in metabolic genes, three mutations were in regulatory genes, and three mutants were found to have lost an entire replicon (R12, R33, and R40). The latter three mutants have been described elsewhere (42).…”

Section: Identification Of B Cenocepacia H111 Mutants Attenuated In Cmentioning

confidence: 99%

Identification of Burkholderia cenocepacia Strain H111 Virulence Factors Using Nonmammalian Infection Hosts

et al. 2013

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c Burkholderia cenocepacia H111, a strain isolated from a cystic fibrosis patient, has been shown to effectively kill the nematode Caenorhabditis elegans. We used the C. elegans model of infection to screen a mini-Tn5 mutant library of B. cenocepacia H111 for attenuated virulence. Of the approximately 5,500 B. cenocepacia H111 random mini-Tn5 insertion mutants that were screened, 22 showed attenuated virulence in C. elegans. Except for the quorum-sensing regulator cepR, none of the mutated genes coded for the biosynthesis of classical virulence factors such as extracellular proteases or siderophores. Instead, the mutants contained insertions in metabolic and regulatory genes. Mutants attenuated in virulence in the C. elegans infection model were also tested in the Drosophila melanogaster pricking model, and those also attenuated in this model were further tested in Galleria mellonella. Six of the 22 mutants were attenuated in D. melanogaster, and five of these were less pathogenic in the G. mellonella model. We show that genes encoding enzymes of the purine, pyrimidine, and shikimate biosynthesis pathways are critical for virulence in multiple host models of infection.

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“…The core genome of B. cenocepacia HI2424 has a high %GC content (66.8%) and harbors three chromosomes, each containing rDNA operons (LiPuma et al 2002), although the third chromosome can be eliminated under certain conditions (Agnoli et al 2012). The primary chromosome (Chr1) is 3.48 Mb and contains 3253 genes; the secondary chromosome (Chr2) is 3.00 Mb and contains 2709 genes; and the tertiary chromosome (Chr3) is 1.06 Mb and contains 929 genes.…”

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confidence: 99%

The Rate and Molecular Spectrum of Spontaneous Mutations in the GC-Rich Multichromosome Genome of Burkholderia cenocepacia

et al. 2015

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Spontaneous mutations are ultimately essential for evolutionary change and are also the root cause of many diseases. However, until recently, both biological and technical barriers have prevented detailed analyses of mutation profiles, constraining our understanding of the mutation process to a few model organisms and leaving major gaps in our understanding of the role of genome content and structure on mutation. Here, we present a genome-wide view of the molecular mutation spectrum in Burkholderia cenocepacia, a clinically relevant pathogen with high %GC content and multiple chromosomes. We find that B. cenocepacia has low genome-wide mutation rates with insertion-deletion mutations biased toward deletions, consistent with the idea that deletion pressure reduces prokaryotic genome sizes. Unlike prior studies of other organisms, mutations in B. cenocepacia are not AT biased, which suggests that at least some genomes with high %GC content experience unusual base-substitution mutation pressure. Importantly, we also observe variation in both the rates and spectra of mutations among chromosomes and elevated G:C . T:A transversions in late-replicating regions. Thus, although some patterns of mutation appear to be highly conserved across cellular life, others vary between species and even between chromosomes of the same species, potentially influencing the evolution of nucleotide composition and genome architecture.KEYWORDS mutation rate; genome evolution; replication timing; mutation spectra A S the ultimate source of genetic variation, mutation is implicit in every aspect of genetics and evolution. However, as a result of the genetic burden imposed by deleterious mutations, remarkably low mutation rates have evolved across all of life, making detection of these rare events technologically challenging and accurate measures of mutation rates and spectra exceedingly difficult (Kibota and Lynch 1996;Lynch and Walsh 1998;Sniegowski et al. 2000;Lynch 2011;Fijalkowska et al. 2012;Zhu et al. 2014). Until recently, most estimates of mutational properties have been derived indirectly using comparative genomics at putatively neutral sites (Graur and Li 2000;Wielgoss et al. 2011) or by extrapolation from small reporter-construct studies (Drake 1991). Both of these methods are subject to potentially significant biases, as many putatively neutral sites are subject to selection and mutation rates can vary substantially among different genomic regions (Lynch 2007).To avoid the potential biases of these earlier methods, pairing classic mutation accumulation (MA) with wholegenome sequencing (WGS) has become the preferred method for obtaining direct measures of mutation rates and spectra (Lynch et al. 2008;Denver et al. 2009;Ossowski et al. 2010;Lee et al. 2012; Sung et al. 2012a,b;Heilbron et al. 2014). Using this strategy, a single clonal ancestor is used to initiate several replicate lineages that are subsequently passaged through repeated single-cell bottlenecks for several thousand generations. The complete genomes of ea...

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Exposing the third chromosome of Burkholderia cepacia complex strains as a virulence plasmid

Cited by 90 publications

References 61 publications

The Third Replicon of Members of the Burkholderia cepacia Complex, Plasmid pC3, Plays a Role in Stress Tolerance

The Third Replicon of Members of the Burkholderia cepacia Complex, Plasmid pC3, Plays a Role in Stress Tolerance

Identification of Burkholderia cenocepacia Strain H111 Virulence Factors Using Nonmammalian Infection Hosts

The Rate and Molecular Spectrum of Spontaneous Mutations in the GC-Rich Multichromosome Genome of Burkholderia cenocepacia

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