STRUCTURAL VARIATION IN THE O-SPECIFIC POLYSACCHARIDES OF KLEBSIELLA-PNEUMONIAE SEROTYPE-O1 AND SEROTYPE-O8 LIPOPOLYSACCHARIDE - EVIDENCE FOR CLONAL DIVERSITY IN RFB GENES

The O-polysaccharide fraction of the lipopolysaccharide from Klebsiell a pneumoniae serotype O8 was found to comprise two galactose-containin g homopolymers. Structural analysis, using chemical and high-field nuc lear magnetic resonance (NMR) techniques, established that the K. pneu moniae O8 polysaccharides are composed of the linear, disaccharide rep eating units [GRAPHICS] K. pneumoniae 08 mutant RFK-1 was isolated by resistance to phage KO1-2; strain RFK-1 expressed only D-galactan I-OA c. The H-1- and C-13-NMR resonances from this O-polysaccharide indicat e that all of the 0-acetyl groups within the K. pneumoniae 08 polysacc haride are carried on D-galactan I and O-acetylation occurs only on th e beta-D-galactofuranose residues; 60% of the available beta-D-galacto furanose residues are non-acetylated. The O-acetylation of the remaini ng residues is equally distributed between the O-2 and O-6 positions. The carbohydrate backbone structures in the O8 polysaccharide are iden tical to D-galactan I and II expressed by K. pneumoniae O1, accounting for the antigenic cross-reaction between strains belonging to serotyp es O1 and O8. However, the O1 polysaccharides are not acetylated and t he O-acetyl groups present in the K. pneumoniae serotype O8 polysaccha rides provide a structural basis for their recognition as distinct ser otypes. The rfb (O-polysaccharide biosynthesis) gene cluster of K. pne umoniae serotype O1 determines the synthesis of D-galactan 1. rfb(KpO1 )-specific gene probes were used to examine conservation in the rfb ge ne clusters of other K. pneumoniae serotypes which produce D-galactan I. Six O1 strains were examined and all showed hybridization with rfb( KPO1) probes under conditions of high stringency. Three serotype O2 st rains produce D-galactan I and these strains also contained DNA sequen ces recognized by rfb(KpO1) probes under high stringency. The physical maps of these homologous rfb chromosomal regions showed some polymorp hism. Surprisingly, the rfb(KpO8) region from K. pneumoniae serotype O 8 was only recognized by rfb(KpO1) probes under low-stringency hybridi zation conditions, providing evidence for two substantially different clonal groups of rfb genes from K. pneumoniae strains with structurall y related O-antigens.