VIRULENCE REGULON POLYMORPHISM IN GROUP-A STREPTOCOCCI REVEALED BY LONG PCR AND IMPLICATIONS FOR EPIDEMIOLOGIC AND EVOLUTIONARY STUDIES

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 v aried from 12.3 kb for serotypes M1 (NCTC 8198) and M12 (NCTC 10085) t o 7.8 kb for serotype M6 (NCTC 8302), The fragment patterns formed on HaeIII digestion of the products were unique and this allowed the diff erentiation of each of the M-type strains (M1, M3, M4, M5, M6, M11, M1 2, M28, M76 and M78) studied, Contemporary M1 isolates all gave the sa me fragment pattern but differed from the prototype strain (NCTC 8198) in not having a 1.25-kb fragment, Isolates of serotypes M1 and M3 eac h had similar patterns, an indication of their clonality and global di spersion, In contrast, more than one restriction fragment length polym orphism (RFLP) pattern was detected among clinical isolates of serotyp es M5, M6, M12, M4, M(R)28 and M78. Two strains that were M-protein no n-typable by serological means were provisionally classified as M6 by comparisons of HaeIII long PCR fragment patterns.