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Genome-wide dissection of globally emergent multi-drug resistant serotype 19A Streptococcus pneumoniae.

Pillai DR, Shahinas D, Buzina A, Pollock RA, Lau R, Khairnar K, Wong A, Farrell DJ, Green K, McGeer A, Low DE - BMC Genomics (2009)

Bottom Line: DNA sequencing revealed that alleles at key drug resistance loci pbp2a, pbp2x, pbp2b, ermB, mefA/E, and tetM were conserved between pre-PCV7 ST 320 19F and post-PCV7 ST 320 19A most likely due to a capsule switch recombination event.A genome wide comparison of MDR 19A ST320 with MDR 19F ST320 identified 822 unique SNPs in 19A, 61 of which were present in antimicrobial resistance genes and 100 in virulence factors.Our results suggest a complex genetic picture where high-level drug resistance, vaccine selection pressure, and SPN mutational events have created a "perfect storm" for the emergence of MDR 19A.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Laboratory Medicine and Pathobiology, University of Toronto, ON, Canada. dylan.pillai@oahpp.ca

ABSTRACT

Background: Emergence of multi-drug resistant (MDR) serotype 19A Streptococcus pneumoniae (SPN) is well-documented but causal factors remain unclear. Canadian SPN isolates (1993-2008, n = 11,083) were serotyped and in vitro susceptibility tested. A subset of MDR 19A were multi-locus sequence typed (MLST) and representative isolates' whole genomes sequenced.

Results: MDR 19A increased in the post-PCV7 era while 19F, 6B, and 23F concurrently declined. MLST of MDR 19A (n = 97) revealed that sequence type (ST) 320 predominated. ST320 was unique amongst MDR 19A in that its minimum inhibitory concentration (MIC) values for penicillin, amoxicillin, ceftriaxone, and erythromycin were higher than for other ST present amongst post-PCV7 MDR 19A. DNA sequencing revealed that alleles at key drug resistance loci pbp2a, pbp2x, pbp2b, ermB, mefA/E, and tetM were conserved between pre-PCV7 ST 320 19F and post-PCV7 ST 320 19A most likely due to a capsule switch recombination event. A genome wide comparison of MDR 19A ST320 with MDR 19F ST320 identified 822 unique SNPs in 19A, 61 of which were present in antimicrobial resistance genes and 100 in virulence factors.

Conclusions: Our results suggest a complex genetic picture where high-level drug resistance, vaccine selection pressure, and SPN mutational events have created a "perfect storm" for the emergence of MDR 19A.

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Minimum spanning tree of multi-drug resistant MDR 19F isolates (n = 29) from the pre-PCV7 era using BioNumerics software. A categorical clustering was performed based on multi-locus sequence type (MLST). Sequence types sharing the maximum number of single-locus variants were connected first. Each circle represents a sequence type (ST) the size of which is proportional to the number of isolates within that particular ST. Colors within circles indicate the minimum inhibitory concentration (MIC) ranges for penicillin. Relationships between the STs are depicted by the lines connecting the STs and the relative lengths of the branches linking them. Distance coding enumerates the number of differences at a given MLST locus. A distance coding of greater than 2 implies a different clonal complex. Angles of the line connections and the overlapping circles have no significance.
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Figure 5: Minimum spanning tree of multi-drug resistant MDR 19F isolates (n = 29) from the pre-PCV7 era using BioNumerics software. A categorical clustering was performed based on multi-locus sequence type (MLST). Sequence types sharing the maximum number of single-locus variants were connected first. Each circle represents a sequence type (ST) the size of which is proportional to the number of isolates within that particular ST. Colors within circles indicate the minimum inhibitory concentration (MIC) ranges for penicillin. Relationships between the STs are depicted by the lines connecting the STs and the relative lengths of the branches linking them. Distance coding enumerates the number of differences at a given MLST locus. A distance coding of greater than 2 implies a different clonal complex. Angles of the line connections and the overlapping circles have no significance.

Mentions: Due to the rising absolute number of MDR 19A isolates, MLST was performed to establish the genetic background of these strains. Figure 2 demonstrates that sequence type (ST) 320, part of CC271, accounts for the majority of MDR 19A following the introduction of PCV7 (post-PCV7) in Canada. Prior to PCV7 introduction, ST320 was most significantly associated with serotype 19F in this study (our unpublished data). ST320 is a single-locus variant (different at one gene in the MLST schema comprising seven genes) of Taiwan 19F-14 (ST236) which spread globally in the pre-PCV7 era [17]. Categorical clustering of MDR 19A based on MLST demonstrated that ST320 was associated with high-level penicillin resistance (minimum inhibitory concentration/MIC ≥ 4 μg/mL) (Figure 3). In contrast, non-MDR 19A control isolates taken from CBSN were associated with different STs (Figure 4). Pre-PCV7 MDR 19F high-level penicillin resistance was also strongly associated with ST320 in our Canadian database (Figure 5). Table 1 to 5 summarize actual MIC values for individual MDR 19A isolates by ST. MIC values for penicillin (Table 1), ceftriaxone (Table 2), amoxicillin (Table 3), erythromycin (Table 4), and ciprofloxacin (Table 5) are generally higher for ST320 when compared to other STs amongst MDR 19A. Although not statistically significant, the most notable trends among these antibiotics were penicillin and amoxicillin where ST320 was high-level resistant, while other STs amongst MDR 19A were less resistant or susceptible. Additional file 1http://www.pillailab.com/suppdata/index.html shows eBURST results and summary statistics for all MLST carried out in this study. Novel MLST are detailed in Additional file 2http://www.pillailab.com/suppdata/index.html.


Genome-wide dissection of globally emergent multi-drug resistant serotype 19A Streptococcus pneumoniae.

Pillai DR, Shahinas D, Buzina A, Pollock RA, Lau R, Khairnar K, Wong A, Farrell DJ, Green K, McGeer A, Low DE - BMC Genomics (2009)

Minimum spanning tree of multi-drug resistant MDR 19F isolates (n = 29) from the pre-PCV7 era using BioNumerics software. A categorical clustering was performed based on multi-locus sequence type (MLST). Sequence types sharing the maximum number of single-locus variants were connected first. Each circle represents a sequence type (ST) the size of which is proportional to the number of isolates within that particular ST. Colors within circles indicate the minimum inhibitory concentration (MIC) ranges for penicillin. Relationships between the STs are depicted by the lines connecting the STs and the relative lengths of the branches linking them. Distance coding enumerates the number of differences at a given MLST locus. A distance coding of greater than 2 implies a different clonal complex. Angles of the line connections and the overlapping circles have no significance.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC2807444&req=5

Figure 5: Minimum spanning tree of multi-drug resistant MDR 19F isolates (n = 29) from the pre-PCV7 era using BioNumerics software. A categorical clustering was performed based on multi-locus sequence type (MLST). Sequence types sharing the maximum number of single-locus variants were connected first. Each circle represents a sequence type (ST) the size of which is proportional to the number of isolates within that particular ST. Colors within circles indicate the minimum inhibitory concentration (MIC) ranges for penicillin. Relationships between the STs are depicted by the lines connecting the STs and the relative lengths of the branches linking them. Distance coding enumerates the number of differences at a given MLST locus. A distance coding of greater than 2 implies a different clonal complex. Angles of the line connections and the overlapping circles have no significance.
Mentions: Due to the rising absolute number of MDR 19A isolates, MLST was performed to establish the genetic background of these strains. Figure 2 demonstrates that sequence type (ST) 320, part of CC271, accounts for the majority of MDR 19A following the introduction of PCV7 (post-PCV7) in Canada. Prior to PCV7 introduction, ST320 was most significantly associated with serotype 19F in this study (our unpublished data). ST320 is a single-locus variant (different at one gene in the MLST schema comprising seven genes) of Taiwan 19F-14 (ST236) which spread globally in the pre-PCV7 era [17]. Categorical clustering of MDR 19A based on MLST demonstrated that ST320 was associated with high-level penicillin resistance (minimum inhibitory concentration/MIC ≥ 4 μg/mL) (Figure 3). In contrast, non-MDR 19A control isolates taken from CBSN were associated with different STs (Figure 4). Pre-PCV7 MDR 19F high-level penicillin resistance was also strongly associated with ST320 in our Canadian database (Figure 5). Table 1 to 5 summarize actual MIC values for individual MDR 19A isolates by ST. MIC values for penicillin (Table 1), ceftriaxone (Table 2), amoxicillin (Table 3), erythromycin (Table 4), and ciprofloxacin (Table 5) are generally higher for ST320 when compared to other STs amongst MDR 19A. Although not statistically significant, the most notable trends among these antibiotics were penicillin and amoxicillin where ST320 was high-level resistant, while other STs amongst MDR 19A were less resistant or susceptible. Additional file 1http://www.pillailab.com/suppdata/index.html shows eBURST results and summary statistics for all MLST carried out in this study. Novel MLST are detailed in Additional file 2http://www.pillailab.com/suppdata/index.html.

Bottom Line: DNA sequencing revealed that alleles at key drug resistance loci pbp2a, pbp2x, pbp2b, ermB, mefA/E, and tetM were conserved between pre-PCV7 ST 320 19F and post-PCV7 ST 320 19A most likely due to a capsule switch recombination event.A genome wide comparison of MDR 19A ST320 with MDR 19F ST320 identified 822 unique SNPs in 19A, 61 of which were present in antimicrobial resistance genes and 100 in virulence factors.Our results suggest a complex genetic picture where high-level drug resistance, vaccine selection pressure, and SPN mutational events have created a "perfect storm" for the emergence of MDR 19A.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Laboratory Medicine and Pathobiology, University of Toronto, ON, Canada. dylan.pillai@oahpp.ca

ABSTRACT

Background: Emergence of multi-drug resistant (MDR) serotype 19A Streptococcus pneumoniae (SPN) is well-documented but causal factors remain unclear. Canadian SPN isolates (1993-2008, n = 11,083) were serotyped and in vitro susceptibility tested. A subset of MDR 19A were multi-locus sequence typed (MLST) and representative isolates' whole genomes sequenced.

Results: MDR 19A increased in the post-PCV7 era while 19F, 6B, and 23F concurrently declined. MLST of MDR 19A (n = 97) revealed that sequence type (ST) 320 predominated. ST320 was unique amongst MDR 19A in that its minimum inhibitory concentration (MIC) values for penicillin, amoxicillin, ceftriaxone, and erythromycin were higher than for other ST present amongst post-PCV7 MDR 19A. DNA sequencing revealed that alleles at key drug resistance loci pbp2a, pbp2x, pbp2b, ermB, mefA/E, and tetM were conserved between pre-PCV7 ST 320 19F and post-PCV7 ST 320 19A most likely due to a capsule switch recombination event. A genome wide comparison of MDR 19A ST320 with MDR 19F ST320 identified 822 unique SNPs in 19A, 61 of which were present in antimicrobial resistance genes and 100 in virulence factors.

Conclusions: Our results suggest a complex genetic picture where high-level drug resistance, vaccine selection pressure, and SPN mutational events have created a "perfect storm" for the emergence of MDR 19A.

Show MeSH
Related in: MedlinePlus