Structure of the K141 capsular polysaccharide produced by Acinetobacter baumannii isolate KZ1106 that carries KL141 at the chromosomal K locus.

The structure of the K141 type capsular polysaccharide (CPS) produced by Acinetobacter baumannii KZ1106, a clinical isolate recovered from Kazakhstan in 2016, was established by sugar analyses and one- and two-dimensional 1H and 13C NMR spectroscopy. The CPS was shown to consist of branched tetrasaccharide repeating units (K-units) with the following structure: This structure was found to be consistent with the genetic content of the KL141 CPS biosynthesis gene cluster at the chromosomal K locus in the KZ1106 whole genome sequence. Assignment of the encoded enzymes allowed the first sugar of the K unit to be identified, which revealed that the ß-d-GlcpNAc-(1→3)-d-GlcpNAc bond is the linkage between K-units formed by the WzyKL141 polymerase.

New Obolenskvirus Phages Brutus and Scipio: Biology, Evolution, and Phage-Host Interaction.

Two novel virulent phages of the genus Obolenskvirus infecting Acinetobacter baumannii, a significant nosocomial pathogen, have been isolated and studied. Phages Brutus and Scipio were able to infect A. baumannii strains belonging to the K116 and K82 capsular types, respectively. The biological properties and genomic organization of the phages were characterized. Comparative genomic, phylogenetic, and pangenomic analyses were performed to investigate the relationship of Brutus and Scipio to other bacterial viruses and to trace the possible origin and evolutionary history of these phages and other representatives of the genus Obolenskvirus. The investigation of enzymatic activity of the tailspike depolymerase encoded in the genome of phage Scipio, the first reported virus infecting A. baumannii of the K82 capsular type, was performed. The study of new representatives of the genus Obolenskvirus and mechanisms of action of depolymerases encoded in their genomes expands knowledge about the diversity of viruses within this taxonomic group and strategies of Obolenskvirus-host bacteria interaction.

Revised structure of the polysaccharide from Acinetobacter baumannii LUH5551 assigned as the K63 type capsular polysaccharide.

K63 capsular polysaccharide produced by Acinetobacter baumannii isolate LUH5551 (previously designated isolate O24) was re-examined using sugar analysis, Smith degradation, and one- and two-dimensional 1H and 13C NMR spectroscopy. Though previously reported as O24 consisting of linear tetrasaccharide units that include a 7-acetamido-5-acylamino form of 8-epilegionaminic acid [8eLeg5R7Ac, acylated at C5 with (S)-3-hydroxybutanoyl or acetyl (1:1)], the elucidated structure of the K63 type capsule was found to include a derivative of 5,7-diamino-3,5,7,9-tetradeoxy-d-glycero-d-galacto-non-2-ulosonic (legionaminic) acid, Leg5Ac7R, where R is either (S)-3-hydroxybutanoyl or an acetyl group (∼1:1 ratio). This finding is consistent with the presence of the lgaABCHIFG gene module for Leg5Ac7R biosynthesis in the KL63 gene cluster at the capsular polysaccharide (CPS) biosynthesis K locus in the LUH5551 genome. The glycosyltransferases (Gtrs) and Wzy polymerase encoded by KL63 were assigned to linkages in the linear K63 tetrasaccharide unit and linkage of the K63 units.

The Acinetobacter baumannii K70 and K9 capsular polysaccharides consist of related K-units linked by the same Wzy polymerase and cleaved by the same phage depolymerases.

IMPORTANCE: Bacteriophage show promise for the treatment of Acinetobacter baumannii infections that resist all therapeutically suitable antibiotics. Many tail-spike depolymerases encoded by phage that are able to degrade A. baumannii capsular polysaccharide (CPS) exhibit specificity for the linkage present between K-units that make up CPS polymers. This linkage is formed by a specific Wzy polymerase, and the ability to predict this linkage using sequence-based methods that identify the Wzy at the K locus could assist with the selection of phage for therapy. However, little is known about the specificity of Wzy polymerase enzymes. Here, we describe a Wzy polymerase that can accommodate two different but similar sugars as one of the residues it links and phage depolymerases that can cleave both types of bond that Wzy forms.

A highly branched novel galactofuranan in the cell wall of Clavibacter tesselarius VKM Ac-1406T.

The structures of two cell wall glycopolymers were studied in the plant pathogenic bacterium Clavibacter tesselarius VKM Ac-1406T (family Microbacteriaceae, order Micrococcales, class Actinomycetes). The predominant polymer was a novel (1 â†’ 6)-linked ß-d-galactofuranan with a highly branched repeating unit, α-L-Rhap-(1 â†’ 3)-α-D-Galp-(1 â†’ 2)-[α-L-Rhap-(1 â†’ 3)]-α-D-Fucp-(1 →, at O-2 on every second galactofuranose residue. The second polymer present in small amounts was acidic with the repeating unit, →3)-α-D-Galp-(1 â†’ 3)-α-D-[4,6-S-Pyr]-Manp-(1 â†’ 3)-α-D-Manp-[2OAc]0.2-(1→, and was reported in all Clavibacter species investigated to date. The presented results expand our knowledges of structural diversity of phosphate-free cell wall glycopolymers and provide evidence in support of their taxonomic specificity for bacterial species and genera.

A novel cell wall galactofuranan in Clavibacter phaseoli VKM Ac-2641T.

A glycopolymer of novel structure was found in the cell wall of plant pathogen Clavibacter phaseoli VKM Ac-2641T (family Microbacteriaceae, class Actinomycetes). The glycopolymer was (1 â†’ 6)-linked ß-d-galactofuranan with side branched trisaccharide, α-D-Manp-(1 â†’ 2)-[α-D-Manp-(1 â†’ 3)]-α-D-Ribf-(1→ at O-2 on every second galactofuranose residue. The galactofuranan structure was established by chemical and NMR spectroscopic methods using one- and two-dimensional techniques 1H,1H COSY, TOCSY, ROESY and 1H,13C HSQC, HMBC. The results of this study provide new data on diversity of bacterial glycopolymers, may prove useful for bacterial taxonomy and contribute to the understanding of the host plant-microbiota interaction mechanisms.

Structural and genetic relatedness of the O-antigens of Escherichia coli O50 and O2.

An O-specific polysaccharide (O-antigen) was isolated by mild acid degradation of the lipopolysaccharide of Escherichia coli O50 followed by gel chromatography on Sephadex G-50. The following structure of the tetrasaccharide repeat was established by sugar analysis and 1D and 2D 1H and 13C NMR spectroscopy: →3)-α-l-Rhap-(1 → 2)-α-l-Rhap-(1 → 3)-ß-l-Rhap-(1 → 4)-ß-d-GlcpNAc-(1→ The linear O50 polysaccharide has the same structure as the main chain of the branched O polysaccharide of E. coli O2 studied earlier [Jansson et al., Carbohydr. Res. 161 (1987) 273-279], which differs in the presence of a side-chain α-d-Fucp3NAc residue. In spite of the difference between the O-polysaccharides, the corresponding genes in the O2- and O50-antigen gene cluster are 99-100% identical. The genetic basis for the lack of d-Fucp3NAc from the O50 polysaccharide is evidently a point mutation in the aminotransferase gene fdtB of the d-Fucp3NAc synthesis pathway resulting in a single amino acid change from histidine in O2 to arginine in O50.

Serological classification and epitope specificity of Proteus vulgaris TG 251 from Proteus serogroup O65.

INTRODUCTION: Proteus rods are currently subdivided into five named species, i.e. Proteus mirabilis, P. vulgaris, P. penneri, P. hauseri, and P. myxofaciens, and three unnamed Proteus genomospecies 4 to 6. Based on the serospecificity of the lipopolysaccharide (LPS; O-antigen), strains of P. mirabilis and P. vulgaris were divided into 49 O-serogroups and 11 additional O-serogroups were proposed later. About 15 further O-serogroups have been proposed for the third medically important species, P. penneri. Here the serological classification of P. vulgaris strain TG 251, which does not belong to these serogroups, is reported. Serological investigations also allowed characterization of the epitope specificity of its LPS. MATERIALS AND METHODS: Purified LPSs from five Proteus strains were used as antigens in enzyme immunosorbent assay (EIA), SDS/PAGE, and Western blot and alkali-treated LPSs in the passive immunohemolysis (PIH) test, inhibition of PIH and EIA, and absorption of the rabbit polyclonal O-antisera with the respective LPS. RESULTS: The serological studies of P. vulgaris TG 251 LPS indicated the identity of its O-polysaccharide with that of P. penneri O65. The antibody specificities of P. vulgaris TG 251 and P. penneri O65 O-antisera, were described. CONCLUSIONS: P. vulgaris TG 251 was classified to the Proteus O65 serogroup. Two disaccharide-associated epitopes present in P. vulgaris TG 251 and P. penneri O65 LPSs are suggested to be responsible for cross-reactions with three heterologous Proteus strains.