Group B streptococci (GBS; Streptococcus agalactiae) are the leading cause of neonatal invasive diseases and are also important pathogens for adults. Penicillins are the drugs of first choice for the treatment of GBS infections, since GBS have been regarded to be uniformly susceptible to penicillins so far. Here we characterize the first strains of GBS with reduced penicillin susceptibility (PRGBS) identified in Japan. Fourteen PRGBS strains were clinically isolated from the sputa of elderly patients from 1995 to 2005; and the MICs of penicillin, oxacillin, and ceftizoxime ranged from 0.25 to 1 g/ml, 2 to 8 g/ml, and 4 to 128 g/ml, respectively. Moreover, some strains were also insusceptible to ampicillin, cefazolin, cefepime, and cefotaxime. All the PRGBS isolates tested possessed a few amino acid substitutions adjacent to the conserved SSN and KSG motifs (amino acids 402 to 404 and 552 to 554, respectively) of PBP 2X, and the amino acid substitutions could be classified into two types, Q557E and V405A. Western blotting analysis of the 14 clinical isolates with anti-PBP 2X-specific serum suggested that the amount of PBP 2X among the 14 PRGBS isolates was reduced, although the 2 ATCC strains produced a significant amount of PBP 2X. The introduction of PRGBS-derived PBP 2X genes into penicillin-susceptible strains through allelic exchange elevated their penicillin insusceptibility, suggesting that these altered PBP 2X genes are responsible for the penicillin insusceptibility in PRGBS strains. In this study, we characterized for the first time PRGBS strains on a molecular basis, although several reports have so far mentioned the existence of ␤-lactam-insusceptible GBS from a phenotypic standpoint.

Type III group B streptococci (GBS) isolated from Tokyo and Salt Lake City were classified according to the similarity of HindIII and Sse83871 restriction digest patterns (RDPs) of bacterial DNA. The bacteria were clustered into three RDP types, with excellent correlation between subtyping based on the two enzymes. The majority (91%) of invasive isolates obtained from neonates were RDP type III-3. The mean sialic acid content of the III-3 strains was higher than that of other type III strains. Closely related isolates were concordant for expression of the bacterial enzyme C5a-ase, but invasive strains were no more likely to be C5a-ase positive than were strains isolated from the genitourinary tract of pregnant women. These data indicate that a group of genetically related organisms with increased capsule production causes the majority of invasive type III GBS disease. Group B streptococci (GBS) 3) causes most serotype III neonatal sepsis, suggesting the RDP typing. DNA was extracted from GBS suspended in agarose existence of a genetically related subgroup of serotype III GBS gel plugs (InCert; FMC BioProducts, Rockland, ME) according to that are intrinsically more virulent than other serotype III strains the manufacturer's instructions except that mutanolysin and SDS con- [4]. The purpose of these studies was to determine whether taining proteinase K were used for digestion of the bacteria. DNA in invasive serotype III isolates from Salt Lake City, as well as the agarose gel plugs was digested with HindIII, extracted from the more recently isolated clinical isolates from Japan, are also agarose with phenol, and redigested with HindIII. The DNA sample RDP III-3. We also validated the HindIII typing by typing the was then subjected to electrophoresis in a conventional ethidium bromide-agarose gel. The similarity between densitometric RDPs from individual strains was expressed as a Pearson product moment correlation coefficient (PPMCC) and clustered by the unweighted

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