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Evidence for a purifying selection acting on the β-lactamase locus in epidemic clones of methicillin-resistant Staphylococcus aureus.

Milheiriço C, Portelinha A, Krippahl L, de Lencastre H, Oliveira DC - BMC Microbiol. (2011)

Bottom Line: Thirteen allotypes for blaZ, nine for blaI and 12 for blaR1 were found.In a total of 121 unique single-nucleotide polymorphisms (SNP) detected, no frameshift mutations were identified and only one nonsense mutation within blaZ was found in a MRSA strain.In addition, the data shows that the sensor-inducer blaR1 is the primary target for the accumulation of mutations in the bla locus, presumably to modulate the response to the presence of β-lactam antibiotic.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Molecular Genetics, Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa (ITQB/UNL), Oeiras, Portugal.

ABSTRACT

Background: The β-lactamase (bla) locus, which confers resistance to penicillins only, may control the transcription of mecA, the central element of methicillin resistance, which is embedded in a polymorphic heterelogous chromosomal cassette (the SCCmec element). In order to assess the eventual correlation between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes, the allelic variation on the bla locus was evaluated in a representative collection of 54 international epidemic methicillin-resistant Staphylococcus aureus (MRSA) clinical strains and, for comparative purposes, also in 24 diverse methicillin-susceptible S. aureus (MSSA) strains.

Results: Internal fragments of blaZ (the β-lactamase structural gene) were sequenced for all strains. A subset of strains, representative of blaZ allotypes, was further characterized by sequencing of internal fragments of the blaZ transcriptional regulators, blaI and blaR1. Thirteen allotypes for blaZ, nine for blaI and 12 for blaR1 were found. In a total of 121 unique single-nucleotide polymorphisms (SNP) detected, no frameshift mutations were identified and only one nonsense mutation within blaZ was found in a MRSA strain. On average, blaZ alleles were more polymorphic among MSSA than in MRSA (14.7 vs 11.4 SNP/allele). Overall, blaR1 was the most polymorphic gene with an average of 24.8 SNP/allele. No correlation could be established between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes. In order to estimate the selection pressure acting on the bla locus, the average dN/dS values were computed. In the three genes and in both collections dN/dS ratios were significantly below 1.

Conclusions: The data strongly suggests the existence of a purifying selection to maintain the bla locus fully functional even on MRSA strains. Although, this is in agreement with the notion that in most clinical MRSA strains mecA gene is under the control of the bla regulatory genes, these findings also suggest that the apparently redundant function of blaZ gene for the MRSA resistant phenotype is still important for these strains. In addition, the data shows that the sensor-inducer blaR1 is the primary target for the accumulation of mutations in the bla locus, presumably to modulate the response to the presence of β-lactam antibiotic.

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blaZ allotype frequency per MRSA lineage as defined by MLST clonal cluster.
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Figure 1: blaZ allotype frequency per MRSA lineage as defined by MLST clonal cluster.

Mentions: Thirteen different blaZ allotypes were identified within our collection, which comprised 54 MRSA and 24 MSSA (Tables 1 and 2, respectively). Although seven alleles were common to MRSA and MSSA strains, we found four alleles present in MRSA strains only and two present in MSSA strains only. Moreover, the relative frequencies of each allele were different among MRSA and MSSA strains (Table 3); for instance, blaZ allotype 1 was dominant in MRSA strains accounting for 43% (23 out of 54) of the isolates whereas in MSSA it accounted for 21% (5 out of 24) of the isolates, and blaZ allotype 6 was present in 11% (6 out 54) of MRSA but was dominant among MSSA accounting for 46% (11 out 24) of the isolates. The diversity of blaZ gene as measured by the Simpson index of diversity (SID) was higher for the MRSA collection than for MSSA, although not statistically significant due to the partial overlapping of the confidence intervals (SID = 79.18, 95%CI 69.6-88.8 vs SID = 76.09, 95%CI 61.3-90.9, respectively) - see Table 4. Within the length of blaZ region analyzed (492 nucleotides), we detected 43 unique single-nucleotide polymorphisms (SNP) and on average, each blaZ allele has 12.4 SNP comparing to the prototype blaZ sequence of Tn552 (allele 1) - see Tables 3 and 4. Overall, blaZ alleles were more variable in MSSA than in MRSA (14.7 and 11.4 SNP/allele, respectively). As illustrated by the allelic frequency distribution per MRSA lineage (Figure 1) or the cluster tree of the thirteen blaZ alleles found in our collections (Figure 2), there is no clustering according to genetic lineages, as defined by MLST sequence type and SCCmec type, or MSSA/MRSA phenotype; i.e. the same allele could be detected in different genetic lineages or among MRSA and MSSA, and the same lineage could be characterized by several alleles. In addition, there was also no clear clustering of blaZ allotypes according to geographic origin or isolation date of the MRSA isolates (see Table 1).


Evidence for a purifying selection acting on the β-lactamase locus in epidemic clones of methicillin-resistant Staphylococcus aureus.

Milheiriço C, Portelinha A, Krippahl L, de Lencastre H, Oliveira DC - BMC Microbiol. (2011)

blaZ allotype frequency per MRSA lineage as defined by MLST clonal cluster.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC3102608&req=5

Figure 1: blaZ allotype frequency per MRSA lineage as defined by MLST clonal cluster.
Mentions: Thirteen different blaZ allotypes were identified within our collection, which comprised 54 MRSA and 24 MSSA (Tables 1 and 2, respectively). Although seven alleles were common to MRSA and MSSA strains, we found four alleles present in MRSA strains only and two present in MSSA strains only. Moreover, the relative frequencies of each allele were different among MRSA and MSSA strains (Table 3); for instance, blaZ allotype 1 was dominant in MRSA strains accounting for 43% (23 out of 54) of the isolates whereas in MSSA it accounted for 21% (5 out of 24) of the isolates, and blaZ allotype 6 was present in 11% (6 out 54) of MRSA but was dominant among MSSA accounting for 46% (11 out 24) of the isolates. The diversity of blaZ gene as measured by the Simpson index of diversity (SID) was higher for the MRSA collection than for MSSA, although not statistically significant due to the partial overlapping of the confidence intervals (SID = 79.18, 95%CI 69.6-88.8 vs SID = 76.09, 95%CI 61.3-90.9, respectively) - see Table 4. Within the length of blaZ region analyzed (492 nucleotides), we detected 43 unique single-nucleotide polymorphisms (SNP) and on average, each blaZ allele has 12.4 SNP comparing to the prototype blaZ sequence of Tn552 (allele 1) - see Tables 3 and 4. Overall, blaZ alleles were more variable in MSSA than in MRSA (14.7 and 11.4 SNP/allele, respectively). As illustrated by the allelic frequency distribution per MRSA lineage (Figure 1) or the cluster tree of the thirteen blaZ alleles found in our collections (Figure 2), there is no clustering according to genetic lineages, as defined by MLST sequence type and SCCmec type, or MSSA/MRSA phenotype; i.e. the same allele could be detected in different genetic lineages or among MRSA and MSSA, and the same lineage could be characterized by several alleles. In addition, there was also no clear clustering of blaZ allotypes according to geographic origin or isolation date of the MRSA isolates (see Table 1).

Bottom Line: Thirteen allotypes for blaZ, nine for blaI and 12 for blaR1 were found.In a total of 121 unique single-nucleotide polymorphisms (SNP) detected, no frameshift mutations were identified and only one nonsense mutation within blaZ was found in a MRSA strain.In addition, the data shows that the sensor-inducer blaR1 is the primary target for the accumulation of mutations in the bla locus, presumably to modulate the response to the presence of β-lactam antibiotic.

View Article: PubMed Central - HTML - PubMed

Affiliation: Laboratory of Molecular Genetics, Instituto de Tecnologia Química e Biológica, Universidade Nova de Lisboa (ITQB/UNL), Oeiras, Portugal.

ABSTRACT

Background: The β-lactamase (bla) locus, which confers resistance to penicillins only, may control the transcription of mecA, the central element of methicillin resistance, which is embedded in a polymorphic heterelogous chromosomal cassette (the SCCmec element). In order to assess the eventual correlation between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes, the allelic variation on the bla locus was evaluated in a representative collection of 54 international epidemic methicillin-resistant Staphylococcus aureus (MRSA) clinical strains and, for comparative purposes, also in 24 diverse methicillin-susceptible S. aureus (MSSA) strains.

Results: Internal fragments of blaZ (the β-lactamase structural gene) were sequenced for all strains. A subset of strains, representative of blaZ allotypes, was further characterized by sequencing of internal fragments of the blaZ transcriptional regulators, blaI and blaR1. Thirteen allotypes for blaZ, nine for blaI and 12 for blaR1 were found. In a total of 121 unique single-nucleotide polymorphisms (SNP) detected, no frameshift mutations were identified and only one nonsense mutation within blaZ was found in a MRSA strain. On average, blaZ alleles were more polymorphic among MSSA than in MRSA (14.7 vs 11.4 SNP/allele). Overall, blaR1 was the most polymorphic gene with an average of 24.8 SNP/allele. No correlation could be established between bla allotypes and genetic lineages, SCCmec types and/or β-lactam resistance phenotypes. In order to estimate the selection pressure acting on the bla locus, the average dN/dS values were computed. In the three genes and in both collections dN/dS ratios were significantly below 1.

Conclusions: The data strongly suggests the existence of a purifying selection to maintain the bla locus fully functional even on MRSA strains. Although, this is in agreement with the notion that in most clinical MRSA strains mecA gene is under the control of the bla regulatory genes, these findings also suggest that the apparently redundant function of blaZ gene for the MRSA resistant phenotype is still important for these strains. In addition, the data shows that the sensor-inducer blaR1 is the primary target for the accumulation of mutations in the bla locus, presumably to modulate the response to the presence of β-lactam antibiotic.

Show MeSH
Related in: MedlinePlus