Streptococcus pneumoniae CS109 and CS111 were isolated in the United States in 1991 and have high levels
of resistance to expanded-spectrum cephalosporins (MICs of 8 and 32 mg of cefotaxime per ml, respectively).
CS109, but not CS111, also showed high-level resistance to penicillin. As both strains expressed the serotype
23F capsule, were very closely related in overall genotype, and possessed identical or closely related mosaic
pbp1a, pbp2x, and pbp2b genes, it is likely that they have arisen from a recent common ancestor. High-level
resistance to expanded-spectrum cephalosporins was entirely due to alterations of penicillin-binding proteins
(PBPs) 1a and 2x, since a mixture of the cloned pbp1a and pbp2x genes from the resistant strains could
transform the susceptible strain R6 to the full level of cephalosporin resistance of the clinical isolates. Both
PBP1a and PBP2x of these strains were more resistant to inhibition by cephalosporins than those of typical
highly penicillin-resistant isolates. The pbp1a genes of CS109 and CS111 were identical in sequence, and the
fourfold difference in their levels of resistance to cephalosporins was due to a Thr-5503Ala substitution at the
residue following the conserved Lys-Ser-Gly motif of PBP2x. This substitution was also the major cause of the
16-fold-lower resistance of CS111 to penicillin. The pbp2x gene of CS111, in an appropriate genetic background,
could provide resistance to 16 mg of cefotaxime per ml but only to 0.12 mg of benzylpenicillin per ml. Removal
of the codon 550 mutation resulted in a pbp2x gene that provided resistance to 4 mg of cefotaxime per ml and
4 mg of benzylpenicillin per ml. The Thr-5503Ala substitution in CS111 therefore appears to provide increased
resistance to expanded-spectrum cephalosporins but a loss of resistance to penicillin.