Analysis of growth characteristics, multilocus enzyme electrophoresis, restriction digest pattern (RDP) typing and multilocus sequence typing have identified clonotypes of serotype III group B Streptococcus agalactiae (GBS) associated with invasive infection in neonates. This study sought to unify phenotypic and genotypic classifications of type III GBS strains associated with increased virulence in newborns. High-virulence clonotype (HVC) strains possessed the translation initiation factor 2 (infB) C allele, found in RDP type III-3 strains, and hybridized with the RDP type III-3-specific probe AA3.6, whereas non-HVC strains shared the infB A allele and genomic DNA from these strains did not hybridize with the AA3.6 probe. The characteristic growth lag of HVC GBS at 40 degrees C has been attributed to the presence of a heat-labile fructose-1,6-bisphosphate aldolase (Fba) enzyme in these strains. The deduced amino acid sequence of fba genes of both HVC and non-HVC strains, however, were identical. HVC and RDP type III-3 represent the same genetically related group of bacteria. The characteristic growth differences of virulent strains of type III GBS, however, are not directly attributable to differences in fba.