MOLECULAR ANALYSIS OF A STREPTOCOCCUS-MUTANS STRAIN EXHIBITING POLYMORPHISM IN THE TANDEM GTFB AND GTFC GENES

Streptococcus mutans UA101, which was previously demonstrated to be highly cariogenic in gnotobiotic rats, exhibited much lower water-insoluble glucan (IG) synthetic activity compared with that of S. mutans GS5 and was unable to express sucrose-dependent colonization of smooth surfaces in vitro. On the basis of Southern and Western blot (immunoblot) analyses, it was demonstrated that, unlike most S. mutans strains, strain UA101 contained a single copy of a gene coding for IG synthesis. The gene was isolated from a clone bank constructed with the plasmid pTH10 clone bank in Escherichia coli and had apparently evolved after homologous recombination of the gtfB and gtfC genes present on the chromosome of a recent ancestor of strain UA101. The enzyme expressed from the gene, gtfBC, was purified to near homogeneity by utilizing a single-step preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis system and was characterized. A derivative of strain UA101, UA101LBS, containing a chromosomal insertion of the GS5 gtfC gene was constructed after transformation. UA101LBS exhibited high IG synthetic activity and colonized smooth surfaces in vitro. By utilizing a conventional rat model system involving animals fed a high-sucrose diet, strain UA101 exhibited low levels of smooth surface caries activity relative to Streptococcus sobrinus 6715. By contrast, UA101LBS was as cariogenic as strain 6715. However, sulcal caries occurred equally well with all of the strains tested. These results are evaluated relative to the role of gtf gene products in cariogenicity.