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Investigations into genome diversity of Haemophilus influenzae using whole genome sequencing of clinical isolates and laboratory transformants.

Power PM, Bentley SD, Parkhill J, Moxon ER, Hood DW - BMC Microbiol. (2012)

Bottom Line: Haemophilus influenzae is an important human commensal pathogen associated with significant levels of disease.When compared by Mauve alignment these indicated groupings of H. influenzae that were consistent with previously published analyses; capsule expressing strains fell into two distinct groups and those of serotype b (Hib) were found in two closely positioned lineages.For 18 Hib strains representing both lineages we found many discrete regions (up to 40% of the total genome) displaying sequence variation when compared to a common reference strain.

View Article: PubMed Central - HTML - PubMed

Affiliation: University of Oxford Department of Paediatrics, Medical Sciences Division, John Radcliffe Hospital, Headington, Oxford, UK. .

ABSTRACT

Background: Haemophilus influenzae is an important human commensal pathogen associated with significant levels of disease. High-throughput DNA sequencing was used to investigate differences in genome content within this species.

Results: Genomic DNA sequence was obtained from 85 strains of H. influenzae and from other related species, selected based on geographical site of isolation, disease association and documented genotypic and phenotypic differences. When compared by Mauve alignment these indicated groupings of H. influenzae that were consistent with previously published analyses; capsule expressing strains fell into two distinct groups and those of serotype b (Hib) were found in two closely positioned lineages. For 18 Hib strains representing both lineages we found many discrete regions (up to 40% of the total genome) displaying sequence variation when compared to a common reference strain. Evidence that this naturally occurring pattern of inter-strain variation in H. influenzae can be mediated by transformation was obtained through sequencing DNA obtained from a pool of 200 independent transformants of a recipient (strain Rd) using donor DNA from a heterologous Hib strain (Eagan).

Conclusion: Much of the inter-strain variation in genome sequence in H. influenzae is likely the result of inter-strain exchanges of DNA, most plausibly through transformation.

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Related in: MedlinePlus

SNPs of H. influenzae type b strain sequences when compared with Hib strain 10810. The complete genome sequence of the Hib strain 10810 was used as a reference against which the sequence reads of each strain were mapped using MAQ. Each vertical black line represents the location of a SNP. The equivalent groupings to those identified in Figure 1 are labelled on the right hand side. Regions marked at the bottom of the figure represent genome segments which are present in the reference strain 10810 but that may not be found in all other strains. The brackets on the left hand side of the figure indicate the number of MLST alleles shared between the pairs of genomes indicated; the sequence type (ST) of each strain is indicated to the right of its name.
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Figure 2: SNPs of H. influenzae type b strain sequences when compared with Hib strain 10810. The complete genome sequence of the Hib strain 10810 was used as a reference against which the sequence reads of each strain were mapped using MAQ. Each vertical black line represents the location of a SNP. The equivalent groupings to those identified in Figure 1 are labelled on the right hand side. Regions marked at the bottom of the figure represent genome segments which are present in the reference strain 10810 but that may not be found in all other strains. The brackets on the left hand side of the figure indicate the number of MLST alleles shared between the pairs of genomes indicated; the sequence type (ST) of each strain is indicated to the right of its name.

Mentions: Based on the output from Mauve analysis, we selected Hib strains to analyse, in more depth, the differences in genome content that shape this level of diversity within the species. We used read-mapping by MAQ to investigate single nucleotide polymorphisms (SNPs) between 18 Hib strains included in our genome sequence database and a common reference (Table 1, Figure 2). Strain RM7018, originally designated non-typeable was excluded as it was not a member of this Hib group based on Mauve analysis (Figure 1). Conversely, we included strain PLMIOG2822H-L, a type b strain that had been wrongly classified as H. haemolyticus. Sequence reads were mapped onto a complete reference Hib genome sequence (strain 10810; Genbank FQ312006.1) and used to identify SNPs for all Hib strains. The Hib groupings observed (Figure 2) were essentially the same as those observed by Mauve analysis (Figure 1). Based on the location and number of SNPs, the β1 strains can be sub-grouped into β1a-β1e, and strain RM7598 contains sufficient differences to constitute a separate group (ψ) from the β2 strains (Figure 2). Genome sequence data provides greater resolution in characterising divergence of strains that share identical or similar MLST profiles. For example, when we compared the patterns of SNPs of the sub-grouped β1a-β1e strains to their respective MLSTs, we found that strains RM7578 and DC800 shared similar blocks of SNPs when compared to strain 10810, in a pattern indicative of a common vertical inheritance. Strains RM7578 and DC800 had differed by two MLST alleles (Figure 2). Strains RM7122 and Eagan also differed by two MLST alleles but differed by 4,853 SNPs in comparison to strain 10810.


Investigations into genome diversity of Haemophilus influenzae using whole genome sequencing of clinical isolates and laboratory transformants.

Power PM, Bentley SD, Parkhill J, Moxon ER, Hood DW - BMC Microbiol. (2012)

SNPs of H. influenzae type b strain sequences when compared with Hib strain 10810. The complete genome sequence of the Hib strain 10810 was used as a reference against which the sequence reads of each strain were mapped using MAQ. Each vertical black line represents the location of a SNP. The equivalent groupings to those identified in Figure 1 are labelled on the right hand side. Regions marked at the bottom of the figure represent genome segments which are present in the reference strain 10810 but that may not be found in all other strains. The brackets on the left hand side of the figure indicate the number of MLST alleles shared between the pairs of genomes indicated; the sequence type (ST) of each strain is indicated to the right of its name.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3539920&req=5

Figure 2: SNPs of H. influenzae type b strain sequences when compared with Hib strain 10810. The complete genome sequence of the Hib strain 10810 was used as a reference against which the sequence reads of each strain were mapped using MAQ. Each vertical black line represents the location of a SNP. The equivalent groupings to those identified in Figure 1 are labelled on the right hand side. Regions marked at the bottom of the figure represent genome segments which are present in the reference strain 10810 but that may not be found in all other strains. The brackets on the left hand side of the figure indicate the number of MLST alleles shared between the pairs of genomes indicated; the sequence type (ST) of each strain is indicated to the right of its name.
Mentions: Based on the output from Mauve analysis, we selected Hib strains to analyse, in more depth, the differences in genome content that shape this level of diversity within the species. We used read-mapping by MAQ to investigate single nucleotide polymorphisms (SNPs) between 18 Hib strains included in our genome sequence database and a common reference (Table 1, Figure 2). Strain RM7018, originally designated non-typeable was excluded as it was not a member of this Hib group based on Mauve analysis (Figure 1). Conversely, we included strain PLMIOG2822H-L, a type b strain that had been wrongly classified as H. haemolyticus. Sequence reads were mapped onto a complete reference Hib genome sequence (strain 10810; Genbank FQ312006.1) and used to identify SNPs for all Hib strains. The Hib groupings observed (Figure 2) were essentially the same as those observed by Mauve analysis (Figure 1). Based on the location and number of SNPs, the β1 strains can be sub-grouped into β1a-β1e, and strain RM7598 contains sufficient differences to constitute a separate group (ψ) from the β2 strains (Figure 2). Genome sequence data provides greater resolution in characterising divergence of strains that share identical or similar MLST profiles. For example, when we compared the patterns of SNPs of the sub-grouped β1a-β1e strains to their respective MLSTs, we found that strains RM7578 and DC800 shared similar blocks of SNPs when compared to strain 10810, in a pattern indicative of a common vertical inheritance. Strains RM7578 and DC800 had differed by two MLST alleles (Figure 2). Strains RM7122 and Eagan also differed by two MLST alleles but differed by 4,853 SNPs in comparison to strain 10810.

Bottom Line: Haemophilus influenzae is an important human commensal pathogen associated with significant levels of disease.When compared by Mauve alignment these indicated groupings of H. influenzae that were consistent with previously published analyses; capsule expressing strains fell into two distinct groups and those of serotype b (Hib) were found in two closely positioned lineages.For 18 Hib strains representing both lineages we found many discrete regions (up to 40% of the total genome) displaying sequence variation when compared to a common reference strain.

View Article: PubMed Central - HTML - PubMed

Affiliation: University of Oxford Department of Paediatrics, Medical Sciences Division, John Radcliffe Hospital, Headington, Oxford, UK. .

ABSTRACT

Background: Haemophilus influenzae is an important human commensal pathogen associated with significant levels of disease. High-throughput DNA sequencing was used to investigate differences in genome content within this species.

Results: Genomic DNA sequence was obtained from 85 strains of H. influenzae and from other related species, selected based on geographical site of isolation, disease association and documented genotypic and phenotypic differences. When compared by Mauve alignment these indicated groupings of H. influenzae that were consistent with previously published analyses; capsule expressing strains fell into two distinct groups and those of serotype b (Hib) were found in two closely positioned lineages. For 18 Hib strains representing both lineages we found many discrete regions (up to 40% of the total genome) displaying sequence variation when compared to a common reference strain. Evidence that this naturally occurring pattern of inter-strain variation in H. influenzae can be mediated by transformation was obtained through sequencing DNA obtained from a pool of 200 independent transformants of a recipient (strain Rd) using donor DNA from a heterologous Hib strain (Eagan).

Conclusion: Much of the inter-strain variation in genome sequence in H. influenzae is likely the result of inter-strain exchanges of DNA, most plausibly through transformation.

Show MeSH
Related in: MedlinePlus