Genotyping of the capsule gene cluster (cps) in nontypeable group B streptococci reveals two major cps allelic variants of serotypes III and VII

Forty group B Streptococcus (GBS) isolates obtained from Europe and the Uni ted States previously reported to be nontypeable (NT) by capsule serotype d etermination were subjected to buoyant density gradient centrifugation, Fro m nearly half of the isolates capsule-expressing variants could be selected . For characterization of the remaining NT-GBS isolates, the capsule operon (cps) was amplified by the long-fragment PCR technique and compared by res triction fragment length polymorphism (RFLP) analysis, The patterns from se rotype reference isolates (n = 32) were first determined and used as a comp arison matrix for the NT-GBS isolates. Using two restriction enzymes, SduI and AvaII, cluster analysis revealed a high degree of similarity within ser otypes but less than 88% similarity between serotypes, However, serotypes I II and VII were each split in two distant RFLP clusters, which were designa ted III1 and III2 and VII1 and VII2, respectively. Among the isolates that remained NT after repeated Percoll gradient selections, two insertional mut ants were revealed. Both were found in blood isolates and harbored insertio n sequence (IS) elements within cpsD: one harbored IS1548, and the other ha rbored IS861, All other NT-GBS isolates could, by cluster analysis, be refe rred to different serotypes by comparison to the RFLP reference matrix. In pulsed-field gel electrophoresis of SmaI-restricted chromosomal DNA, patter ns from allelic type 1 and 2 isolates were essentially distributed in separ ate clusters in serotypes III and VII, A covariation with insertion sequenc e IS1548 in the hylB gene was suggested for serotype III, since allelic typ e III1 harboring IS1548 in hylB, clustered separately. The variation in ser otype VII was not dependent on the presence of IS1548, which was not detect ed at any position in the type VII chromosome.