ANALYSIS OF THE ROLE OF M24 PROTEIN IN GROUP-A STREPTOCOCCAL ADHESIONAND COLONIZATION BY USE OF OMEGA-INTERPOSON MUTAGENESIS

We recently concluded that M protein mediates adherence of group A str eptococci to HEp-2 tissue culture cells, because the N-terminal half o f M protein blocked adherence and M(+) strains attached in greater num bers than M(-) streptococci. To further assess the role of M protein i n adhesion, an M(-), isogenic mutant of M type 24 group A streptococci was constructed by insertional inactivation of the emm24 gene with th e Omega-interposon flanked by emm24 gene sequences. Southern blot anal ysis confirmed that the Omega-element inserted only into emm24. The M( -) isogenic mutant M24-Omega 3 did not react with antiserum to M24 pro tein, nor did it survive in whole human blood. Electron micrographs of M24-Omega 3 showed a diminution of surface fibrillae and reduced bind ing of plasma components compared with the parent strain. The adhesion of the M(+) parent to HEp-2 cells and to mouse oral epithelial cells was dramatically greater than the adhesion of the M24-Omega 3 mutant, although there was no difference between the two in adhesion to human buccal cells. In addition, the parent strain was dramatically more eff ective than the M24-Omega 3 mutant in colonizing the oral cavity of mi ce. These results indicate that the M24 protein can serve as an adhesi n in streptococcal attachment to human cells in tissue culture and is important in the colonization of mouse mucosal surfaces.