Detection of penicillin-binding protein 2b gene alteration in Streptococcus mitis by polymerase chain reaction

Three isolates of β-lactam-resistant streptococci from the saliva of healthy adults were identified as Streptococcus mitis. Minimum inhibitory concentrations (MICs) were 2 to 4μg/ml for ampicillin (ABPC) and 64 to more than 128μg/ml for cefaclor (CCL). To determine the position of base alterations of the penicillin-binding protein 2b (pbp2b) gene, upstream primers containing possible mutation points were designed, and used for polymerase chain reaction (PCR), together with a downstream primer. Alterations adjacent to the conserved motifs of the pbp2b gene were apparent. DNA sequencing data indicated replacements in deduced amino acid sequences in all resistant isolates: from threonine to alanine just after the serineserine- asparagine (SSN) motif, and from alanine to glycine two residues downstream of the lysine-threonine-glycine (KTG) motif. These changes were the same as those in penicillin-resistant Streptococcus pneumoniae (PRSP), suggesting importance for the enzymatic activity of the protein. Thus, β-lactam susceptibility of S. mitis may be partially predicted by PCR using our primer set for pbp2b.