The Serotype of Type Ia and III Group B Streptococci Is Determined by the Polymerase Gene within the Polycistronic Capsule Operon

Abstract

Streptococcus agalactiae is a primary cause of neonatal morbidity and mortality. Essential to the virulence of this pathogen is the production of a type-specific capsular polysaccharide (CPS) that enables the bacteria to evade host immune defenses. The identification, cloning, sequencing, and functional characterization of seven genes involved in type III capsule production have been previously reported. Here, we describe the cloning and sequencing of nine additional adjacent genes, cps III FGHIJKL, neu III B, and neu III C. Sequence comparisons suggested that these genes are involved in sialic acid synthesis, pentasaccharide repeating unit formation, and oligosaccharide transport and polymerization. The type III CPS (cpsIII) locus was comprised of 16 genes within 15.5 kb of contiguous chromosomal DNA. Primer extension analysis and investigation of mRNA from mutants with polar insertions in their cpsIII loci supported the hypothesis that the operon is transcribed as a single polycistronic message. The translated cpsIII sequences were compared to those of the S. agalactiae cpsIa locus, and the primary difference between the operons was found to reside in cps III H, the putative CPS polymerase gene. Expression of cps III H in a type Ia strain resulted in suppression of CPS Ia synthesis and in production of a CPS which reacted with type III-specific polyclonal antibody. Likewise, expression of the putative type Ia polymerase gene in a type III strain reduced synthesis of type III CPS with production of a type Ia immunoreactive capsule. Based on the similar structures of the oligosaccharide repeating units of the type Ia and III capsules, our observations demonstrated that cps Ia H and cps III H encoded the type Ia and III CPS polymerases, respectively. Additionally, these findings suggested that a single gene can confer serotype specificity in organisms that produce complex polysaccharides.Group B streptococci (GBS) (Streptococcus agalactiae) are the leading cause of serious bacterial infections (bacteremia, pneumonia, and meningitis) in newborns, causing two to three cases per 1,000 live births (47). An indispensable GBS virulence determinant is the production of a type-specific capsular polysaccharide (CPS), which prevents the deposition of host complement factor C3b and inhibits opsonophagocytosis (45). Nine distinct capsular serotypes, Ia, Ib, and II to VIII, have been identified (54), and their chemical compositions and structures have been determined (16-19, 51, 55, 56, 58). Type Ia, Ib, II to V, and VII CPS consist of the monosaccharides glucose, galactose, N-acetylglucosamine, and Nacetylneuraminic acid. Serotypes VI and VIII lack N-acetylglucosamine, and type VIII contains rhamnose (19). Although serotypes Ia, III, and V are currently the most common isolates from the United States associated with early-onset disease (within 1 week of birth), comprising 82% of isolates (27), type III GBS are the most prevalent isolates associated with neonatal disease (5).We previously identified a region of the GBS chromo...

Structure of the Type IX Group B Streptococcus Capsular Polysaccharide and Its Evolutionary Relationship with Types V and VII

“…The above results are in agreement with previous data from Chaffin et al (36), who reported that a single gene can confer GBS serotype specificity. In that case, the authors reported that cpsH differences were solely responsible for the structural diversity between types Ia and III.…”

A Eukaryotic Type Serine/Threonine Kinase and Phosphatase inStreptococcus agalactiae Reversibly Phosphorylate an Inorganic Pyrophosphatase and Affect Growth, Cell Segregation, and Virulence

“…Selection for double recombination events using kanamycin or chloramphenicol selection and screening for loss of plasmid replication (erythromycin sensitivity) was carried out as described previously (39). At least four independent colonies were obtained from A909/pLR5 that were stk1 mutants (Stk1 Ϫ ).…”

“…A909 (WT) is a type Ia capsular polysaccharide clinical isolate of GBS (38). Recombinant DNA and other techniques were performed as described (39). Open reading frame (ORF) and BLAST homology searches were performed using the NCBI Internet server (www.ncbi.nlm.nih.gov).…”

Discovery and characterization of sialic acid O-acetylation in group BStreptococcus

“…The genetic basis of Sia biosynthesis in GBS has been localized to a cluster of genes at the transcriptional end of the 16-gene capsule operon (32). Similar to E. coli (33,34), GBS gene products of neuC, neuB, and neuA, respectively, catalyze the epimerization of UDP-GlcNAc to N-acetyl mannosamine (ManNAc) (35), the condensation of ManNAc and phosphoenolpyruvate to form Neu5Ac (36), and the activation of Neu5Ac with CTP, to generate CMP-Neu5Ac (37).…”