Identification of genetic differences between two clinical isolates of Streptococcus mutans by suppression subtractive hybridization.

Streptococcus mutans is generally considered to be the principal aetiological agent for dental caries. Phenotypic variation in strains is often associated with differences in gene content, so the isolation of DNA fragments from these genes or associated regions is illuminating. The S. mutans strains 9-1 and 9-2, which both colonized the same oral cavity, were selected after screening for the possession of suspected virulence traits. Genomic DNA of strain 9-1 was used as the tester, and that of 9-2 was used as the driver. Suppression subtractive hybridization (SSH) was applied between the tester and the driver DNAs. The subtractive products were cloned into a pCR2.1 vector. Clone libraries representing sequence differences were obtained. The subtractive fragments that were found specifically in strain 9-1 but not in strain 9-2 were identified by dot blotting and then sequenced. BLASTn and BLASTx sequence homology analyses were subsequently performed. Twenty-seven sequences were found in the genome of strain 9-1 that were not in 9-2. Among them, three revealed no homology to published nucleotide sequences while the remaining sequences showed 81-100% homology to known genes of S. mutans strain UA159. These sequences are involved in competence development, signal transduction and transcriptional regulation, repairing stress damage, transport, carbohydrate catabolism, biochemical synthesis, or unknown functions. Differences exist in the genomes of different S. mutans isolates. SSH is effective in screening for S. mutans strain specific DNA sequences.