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A high-resolution view of genome-wide pneumococcal transformation.

Croucher NJ, Harris SR, Barquist L, Parkhill J, Bentley SD - PLoS Pathog. (2012)

Bottom Line: Those recombinations importing the selected marker were independent of unselected events elsewhere in the genome, the positions of which were not significantly affected by local sequence similarity between donor and recipient or mismatch repair processes.However, both types of recombinations were sometimes mosaic, with multiple non-contiguous segments originating from the same molecule of donor DNA.The lengths of the unselected events were exponentially distributed with a mean of 2.3 kb, implying that recombinations are stochastically resolved with a fixed per base probability of 4.4×10(-4) bp(-1).

View Article: PubMed Central - PubMed

Affiliation: Pathogen Genomics, Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, United Kingdom. nc3@sanger.ac.uk

ABSTRACT
Transformation is an important mechanism of microbial evolution through which bacteria have been observed to rapidly adapt in response to clinical interventions; examples include facilitating vaccine evasion and the development of penicillin resistance in the major respiratory pathogen Streptococcus pneumoniae. To characterise the process in detail, the genomes of 124 S. pneumoniae isolates produced through in vitro transformation were sequenced and recombination events detected. Those recombinations importing the selected marker were independent of unselected events elsewhere in the genome, the positions of which were not significantly affected by local sequence similarity between donor and recipient or mismatch repair processes. However, both types of recombinations were sometimes mosaic, with multiple non-contiguous segments originating from the same molecule of donor DNA. The lengths of the unselected events were exponentially distributed with a mean of 2.3 kb, implying that recombinations are stochastically resolved with a fixed per base probability of 4.4×10(-4) bp(-1). This distribution of recombination sizes, coupled with an observed under representation of large insertions within transferred sequence, suggests transformation has the potential to reduce the size of bacterial genomes, and is unlikely to act as an efficient mechanism for the uptake of accessory genomic loci.

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Related in: MedlinePlus

Cartoon representing the structure of the selected transformations around the pneumococcal cps locus.The kanamycin resistance cassette (orange) on the donor DNA molecule is flanked by pbp genes (yellow), both of which are β lactam-sensitive alleles. The equivalent locus in the recipient chromosome is occupied by the much longer type 23F capsule biosynthesis cluster (represented by a series green boxes to indicate the relatively large number of genes comprising this cluster), which is flanked by pbp alleles that confer β lactam resistance on the host cell. The selected transformations involve in the exchange of DNA from the donor into the recipient, with boundaries in the regions shaded blue.
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ppat-1002745-g001: Cartoon representing the structure of the selected transformations around the pneumococcal cps locus.The kanamycin resistance cassette (orange) on the donor DNA molecule is flanked by pbp genes (yellow), both of which are β lactam-sensitive alleles. The equivalent locus in the recipient chromosome is occupied by the much longer type 23F capsule biosynthesis cluster (represented by a series green boxes to indicate the relatively large number of genes comprising this cluster), which is flanked by pbp alleles that confer β lactam resistance on the host cell. The selected transformations involve in the exchange of DNA from the donor into the recipient, with boundaries in the regions shaded blue.

Mentions: The first experiment involved the transformation of an isolate of S. pneumoniae ATCC 700669, a serotype 23F strain of the penicillin-resistant PMEN1 lineage [39] (henceforth referred to as 23F-R), with genomic DNA from an acapsular derivative of the penicillin-sensitive S. pneumoniae TIGR4 strain, which has a kanamycin resistance marker in place of a capsule biosynthesis (cps) locus [40] (henceforth referred to as TIGR4Δcps) (Figure 1). Multiple transformations were performed using a concentration of either 5 ng mL−1 or 500 ng mL−1 of donor genomic DNA. Recombinations affecting the cps locus were detected either through selection with kanamycin alone, or kanamycin supplemented with ampicillin. This latter condition was used to test whether the transfer of capsule loci from the β lactam-sensitive donor to the resistant recipient may be inhibited by selection against any co-transfer of β lactam-sensitive PBPs [35].


A high-resolution view of genome-wide pneumococcal transformation.

Croucher NJ, Harris SR, Barquist L, Parkhill J, Bentley SD - PLoS Pathog. (2012)

Cartoon representing the structure of the selected transformations around the pneumococcal cps locus.The kanamycin resistance cassette (orange) on the donor DNA molecule is flanked by pbp genes (yellow), both of which are β lactam-sensitive alleles. The equivalent locus in the recipient chromosome is occupied by the much longer type 23F capsule biosynthesis cluster (represented by a series green boxes to indicate the relatively large number of genes comprising this cluster), which is flanked by pbp alleles that confer β lactam resistance on the host cell. The selected transformations involve in the exchange of DNA from the donor into the recipient, with boundaries in the regions shaded blue.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3375284&req=5

ppat-1002745-g001: Cartoon representing the structure of the selected transformations around the pneumococcal cps locus.The kanamycin resistance cassette (orange) on the donor DNA molecule is flanked by pbp genes (yellow), both of which are β lactam-sensitive alleles. The equivalent locus in the recipient chromosome is occupied by the much longer type 23F capsule biosynthesis cluster (represented by a series green boxes to indicate the relatively large number of genes comprising this cluster), which is flanked by pbp alleles that confer β lactam resistance on the host cell. The selected transformations involve in the exchange of DNA from the donor into the recipient, with boundaries in the regions shaded blue.
Mentions: The first experiment involved the transformation of an isolate of S. pneumoniae ATCC 700669, a serotype 23F strain of the penicillin-resistant PMEN1 lineage [39] (henceforth referred to as 23F-R), with genomic DNA from an acapsular derivative of the penicillin-sensitive S. pneumoniae TIGR4 strain, which has a kanamycin resistance marker in place of a capsule biosynthesis (cps) locus [40] (henceforth referred to as TIGR4Δcps) (Figure 1). Multiple transformations were performed using a concentration of either 5 ng mL−1 or 500 ng mL−1 of donor genomic DNA. Recombinations affecting the cps locus were detected either through selection with kanamycin alone, or kanamycin supplemented with ampicillin. This latter condition was used to test whether the transfer of capsule loci from the β lactam-sensitive donor to the resistant recipient may be inhibited by selection against any co-transfer of β lactam-sensitive PBPs [35].

Bottom Line: Those recombinations importing the selected marker were independent of unselected events elsewhere in the genome, the positions of which were not significantly affected by local sequence similarity between donor and recipient or mismatch repair processes.However, both types of recombinations were sometimes mosaic, with multiple non-contiguous segments originating from the same molecule of donor DNA.The lengths of the unselected events were exponentially distributed with a mean of 2.3 kb, implying that recombinations are stochastically resolved with a fixed per base probability of 4.4×10(-4) bp(-1).

View Article: PubMed Central - PubMed

Affiliation: Pathogen Genomics, Wellcome Trust Sanger Institute, Wellcome Trust Genome Campus, Hinxton, Cambridge, United Kingdom. nc3@sanger.ac.uk

ABSTRACT
Transformation is an important mechanism of microbial evolution through which bacteria have been observed to rapidly adapt in response to clinical interventions; examples include facilitating vaccine evasion and the development of penicillin resistance in the major respiratory pathogen Streptococcus pneumoniae. To characterise the process in detail, the genomes of 124 S. pneumoniae isolates produced through in vitro transformation were sequenced and recombination events detected. Those recombinations importing the selected marker were independent of unselected events elsewhere in the genome, the positions of which were not significantly affected by local sequence similarity between donor and recipient or mismatch repair processes. However, both types of recombinations were sometimes mosaic, with multiple non-contiguous segments originating from the same molecule of donor DNA. The lengths of the unselected events were exponentially distributed with a mean of 2.3 kb, implying that recombinations are stochastically resolved with a fixed per base probability of 4.4×10(-4) bp(-1). This distribution of recombination sizes, coupled with an observed under representation of large insertions within transferred sequence, suggests transformation has the potential to reduce the size of bacterial genomes, and is unlikely to act as an efficient mechanism for the uptake of accessory genomic loci.

Show MeSH
Related in: MedlinePlus