Identification of cpsD, a gene essential for type III capsule expression in group B streptococci.
We showed previously that a mutant strain of group B Streptococcus (GBS) defective in capsule production was avirulent. This study describes the derivation of an unencapsulated mutant from a highly encapsulated wild-type strain of type III GBS, COH1, by transposon mutagenesis with Tn916 delta E. The mutant, COH1-13, was sensitive to phagocytic killing by human leukocytes in vitro and was relatively avirulent in a neonatal rat sepsis model compared with the wild-type strain. No capsular polysaccharide was evident in the cytoplasm or on the cell surface of the mutant strain. The Tn916 delta E insertion site in COH1-13 was mapped to the same chromosomal location as the Tn916 insertion site in the unencapsulated type III mutant COH31-15 reported previously. Nucleotide sequencing of DNA flanking the insertion site in COH1-13 revealed an open reading frame, designated cpsD, with significant homology to the rfbP gene of Salmonella typhimurium. RfbP encodes a galactosyl transferase enzyme that catalyses the transfer of galactose to undecaprenol phosphate, the initial step in O-polysaccharide synthesis. A particulate fraction of a lysate of wild-type strain GBS COH1 mediated the transfer of galactose from UDP-galactose to an endogenous acceptor. The galactose-acceptor complex partitioned into organic solvents, suggesting it is lipid in nature or membrane-associated. Galactosyl transferase activity was significantly reduced in the unencapsulated mutant strain COH1-13. These results, together with the similarity in deduced amino acid sequence between cpsD and rfbP suggest that cpsD encodes a galactosyl transferase essential for assembly of the GBS type III capsular polysaccharide.
Rubens
C E
CE
Division of Infectious Disease, Children's Hospital and Medical Center, Seattle, Washington 98105.
Heggen
L M
LM
Haft
R F
RF
Wessels
M R
MR
D000595:Amino Acid Sequence; D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics*; D001483:Base Sequence; D002240:Carbohydrate Sequence; D005700:Galactosyltransferases / Q000235:genetics*; D006801:Humans; D008969:Molecular Sequence Data; D016254:Mutagenesis, Insertional; D016366:Open Reading Frames; D010587:Phagocytosis; D011135:Polysaccharides, Bacterial / Q000096:biosynthesis*; D051381:Rats; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid; D013292:Streptococcus agalactiae / Q000235:genetics* / Q000378:metabolism; D014774:Virulence / Q000235:genetics
D001426:Bacterial Proteins; D011135:Polysaccharides, Bacterial; C032263:streptococcal polysaccharide type III group B; C082664:CpsD protein, bacteria; D005700:Galactosyltransferases