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Extensive genetic diversity among clinical isolates of Streptococcus pyogenes serotype M5
Meeta Desai1, Asha Tanna2, Androulla Efstratiou2
1Molecular Biology Unit, Virus Reference Division.
Abstract:
The genetic diversity of clinical isolates of Streptococcus pyogenes serotype M5 has been characterized. Strain genotypes were defined by macrorestriction profile, 16S ribotype, emm gene subtype, insertion element IS1239 profile, and exotoxin gene determinant. By these criteria, clinical isolates of M5 constituted a multiplicity of strain clusters rather than a homogeneous population as found for certain serotypes. Distance matrices and an unrooted tree were constructed from macrorestriction data with three rarely cutting endonucleases, determined by PFGE. A single IS1239 profile was common to 85% of isolates but there was great diversity of both ribotype and macrorestriction profile, and 18 different emm gene subtypes were detected by PCR-RFLP. DNA sequence analysis of the antigen-coding 5' (hypervariable) region of emm gene amplicons (about 240 bp) showed that 14/18 exhibited up to 6% divergence. Four amplicons had highly divergent sequences--corresponding to those previously determined for emm6, emm11, emm18 and emm77. Further serological and hybridization studies were used to analyse the discrepancy between the Lancefield serotype of these strains (M5) and their emm genotype. Overall, this study shows a high degree of genetic diversity in serotype M5, with implications for the Lancefield scheme itself, for the epidemiology of group A streptococci, and for recombinant DNA strategies for M protein-based vaccine development.
Insights
Genetic diversity was high in Streptococcus pyogenes M5 clinical isolates, challenging homogeneous population assumptions. This finding impacts group A streptococcus epidemiology and vaccine development strategies.
Area of Science:
- Microbiology
- Genetics
- Epidemiology
Background:
- Streptococcus pyogenes (group A Streptococcus) is a significant human pathogen.
- Serotyping, particularly M protein (emm) gene typing, is crucial for understanding GAS epidemiology.
- Previous studies suggested homogeneity in certain serotypes, but M5 diversity was less understood.
Purpose of the Study:
- To genetically characterize clinical isolates of Streptococcus pyogenes serotype M5.
- To assess the genetic diversity within the M5 serotype using multiple molecular markers.
- To evaluate the implications of observed genetic diversity on current classification schemes and vaccine development.
Main Methods:
- Macrorestriction profiling using pulsed-field gel electrophoresis (PFGE).
- 16S ribosomal RNA (16S rRNA) gene ribotyping.
- emm gene subtyping via polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP).
- Insertion sequence (IS) element profiling (IS1239).
- Exotoxin gene determinant analysis.
- DNA sequencing of emm gene hypervariable regions.
- Serological and hybridization studies.
Main Results:
- Clinical isolates of M5 displayed significant genetic diversity, forming multiple strain clusters rather than a homogeneous population.
- While a single IS1239 profile was common (85%), high diversity was observed in ribotype and macrorestriction profiles.
- Eighteen different emm gene subtypes were detected, with 14 showing up to 6% sequence divergence in the hypervariable region.
- Four emm gene amplicons exhibited highly divergent sequences, corresponding to previously identified emm types (emm6, emm11, emm18, emm77).
- Discrepancies were noted between Lancefield serotype (M5) and emm genotype in some isolates.
Conclusions:
- Serotype M5 of Streptococcus pyogenes is genetically diverse, contradicting assumptions of homogeneity.
- The observed genetic heterogeneity has implications for the accuracy of the Lancefield serotyping scheme.
- Understanding this diversity is critical for accurate epidemiology of group A streptococci.
- The genetic variability necessitates careful consideration for developing effective M protein-based vaccines.