Studies on hemolysin of Anginosus group streptococci isolated from the human oral cavity

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Total 15 β hemolytic streptococcal strains were isolated from subgingival plaque samples (5 from patients with adult periodontal disease and 10 from healthy adults). Of these strains, 13 strains exhibited streptogram code of 0110434, indicating that they have close properties of reference strain of Streptococcus anginosus NCTC 10713. Furthermore, 12 strains out of the 13 ones reacted with anti G group streptococcal serum in slide latex agglutination test of Strept LA.These observations suggest that the isolated β hemolytic microorganisms are identical to Streptococcus anginosus which are considered to distribute widely in the oral cavity. The hemolysin of this strain was detected in the extracellular culture fluids and the much more production was observed in general anaerobic medium (GAM) than in brain heart infusion or Todd Hewitt broth.Purification of the hemolysin in the culture supernatant was carried out by concentration by ammonium sulfate (60% saturation), ion-exchange chromatography, gel filtration, and hydroxyapatite chromatography by 625 fold with the recovery of 33.6%. The purified hemolysin formed diffuse bands between 50 to 60 kDa in SDS-PAGE.Isoelectric point was 4.2 and it was inactivated by heating at 70℃ for 10 min. The optimum pH for hemolysis was 7.5. The activity was enhanced 2-3 fold by 2 mercaptoethanol or dithiothreitol. The hemolytic spectrum of erythrocytes was found to be wide ; human, bovine, horse, sheep, guinea pig, and rabbit erythrocytes were lyzed at almost the same degree by this hemolysin. The other properties were quite similar to those of S.anginosus NCTC 10713.Both purified hemolysin samples from the isolated strain and reference strain (NCTC 10713) induced the production of TNF-α and IL-6 by peritoneal macrophages of BALB/c mice.