A method based on long PCR amplification and restriction endonuclease analysis of the virulence regulon was developed for a rapid (2 days), simple differentiation of group A streptococci. The PCR product size varied from 12.3 kb for serotypes M1 (NCTC 8198) and M12 (NCTC 10085) to 7.8 kb for serotype M6 (NCTC 8302). The fragment patterns formed on HaeIII digestion of the products were unique and this allowed the differentiation of each of the M-type strains (M1, M3, M4, M5, M6, M11, M12, M28, M76 and M78) studied. Contemporary M1 isolates all gave the same fragment pattern but differed from the prototype strain (NCTC 8198) in not having a 1.25-kb fragment. Isolates of serotypes M1 and M3 each had similar patterns, an indication of their clonality and global dispersion. In contrast, more than one restriction fragment length polymorphism (RFLP) pattern was detected among clinical isolates of serotypes M5, M6, M12, M4, M(R)28 and M78. Two strains that were M-protein non-typable by serological means were provisionally classified as M6 by comparisons of Hae III long PCR fragment patterns.
StevensD. L.,
TannerM. H.,
WinshipJ. Severe group A streptococcal infections associated with a toxic shock-like syndrome and scarlet fever toxin A. N Engl J Med1989; 321:1–7
RottaJ.,
KrauseR. M.,
LancefieldR. C.,
EverlyW.,
LacklandH. New approaches for the laboratory recognition of M types of group A streptococci. J Exp Med1971; 134:1298–1315
TopF. H.,
WannamakerL. W. The serum opacity reaction of Streptococcus pyogenes: frequency of production of streptococcal lipoproteinase by strains of different serological types and the relationship to M protein production. J Hyg1968; 66:49–58
TopF. H.,
WannamakerL. W. The serum opacity reaction of Streptococcus pyogenes. The demonstration of multiple, strain-specific lipoproteinase antigens. J Exp Med1968; 127:1013–1034
WiddowsonJ. P.,
MaxtedW. R.,
GrantD. L. The production of opacity in serum by group A streptococci and its relationship with the presence of M antigen. J Gen Microbiol1970; 61:343–353
TaggJ. R.,
BannisterL. V. “Fingerprinting” beta-haemolytic streptococci by their production of and sensitivity to bacteriocine-like inhibitors. J Med Microbiol1979; 12:397–411
MusserJ. M.,
GrayB. M.,
SchlievertP. M.,
PichicheroM. E.Streptococcus pyogenes pharyngitis: characterization of strains by multilocus enzyme genotype, M and T protein serotype, and pyrogenic exotoxin gene probing. J Clin Microbiol1992; 30:600–603
MylvaganamH.,
BjorvatnB.,
HofstadT.,
HjetlandR.,
Arne HoibyE.,
HolmS.E. Small-fragment restriction endonuclease analysis in epidemiological mapping of group A streptococci. J Med Microbiol1994; 40:256–260
BruneauS.,
de MontelosH.,
DrouetE.,
DenoyelG. A. rRNA gene restriction patterns of Streptococcus pyogenes: epidemiological applications and relation to serotypes. J Clin Microbiol1994; 32:2953–2958
WhatmoreA. M.,
KehoeM. A. Horizontal gene transfer in the evolution of group A streptococcal emm-like genes: gene mosaics and variation in Vir regulons. Mol Microbiol1994; 11:363–374
PodbielskiA.,
Krebs, B, KaufholdA. Genetic variability of the ewm-related gene of the large vir regulon of group A streptococci: potential intra- and intergenomic recombination events. Mol Gen Genet1994; 243:691–698
ChenC. C.,
BormannN.,
ClearyP. R. VirR and Mry are homologous trans-acting regulators of M protein and C5a peptidase expression in group A streptococci. Mol Gen Genet1993; 241:685–693
ChenC. C.,
ClearyP. P. Complete nucleotide sequence of the streptococcal C5a peptidase gene from Streptococcus pyogenes. J Biol Chem1990; 265:3161–3167
WhatmoreA. M.,
KapurV.,
MusserJ. M.,
KehoeM. A. Molecular population genetic analysis of the enn subdivision of group A streptococcal emm-liks genes: horizontal gene transfer and restricted variation among enn genes. Mol Microbiol1995; 15:1039–1048
BarnesW. M. PCR amplification of up to 35-kb DNA with high fidelity and high yield from λ bacteriophage templates. Proc Natl Acad Sci USA1994; 91:2216–2220
Perez-CasalJ.,
CaparonM. G.,
ScottJ. R. Mry, a trans-acting positive regulator of the M protein gene of Streptococcus pyogenes with similarity to the receptor proteins of two-component regulatory systems. J Bacteriol1991; 173:2617–2624
Perez-CasalJ. F.,
DillonH. F.,
HusmannL. K.,
GrahamB.,
ScottJ. R. Virulence of two Streptococcus pyogenes strains (types Ml and M3) associated with toxic-shock-like syndrome depends on an intact myr-like gene. Infect Immun1993; 61:5426–5430
McLandsboroughL. A.,
ClearyP. P. Insertional inactivation of vfrR in Streptococcus pyogenes M49 demonstrates that VirR functions as a positive regulator of Sep A, FcRA, OF, and M protein. FEMS Microbiol Lett1995; 128:45–52
GardinerD.,
HartasJ.,
CurrieB.,
MathewsJ. D.,
KempD. J.,
SriprakashK. S. Vir typing: a long-PCR method for Group A streptococci. PCR Methods Appl1995; 4:288–293
UptonM.,
CarterP. E.,
MorganM.,
EdwardsG. F.,
PenningtonT. H. Clonal structure of invasive Streptococcus pyogenes in Northclonal Scotland. Epidemiol Infect1995; 115:231–241