Search

Abstract Escherichia coli F17 isolated from horse feces was studied in respect to the O antigen (O polysaccharide) structure and genetics. The lipopolysaccharide was isolated by phenol-water extraction of bacterial cells and cleaved by mild acid hydrolysis to yield the O polysaccharide, which was studied by sugar analysis and selective solvolysis with CF 3 CO 2 H along with one- and two-dimensional 1H and 13C NMR spectroscopy. The O polysaccharide was found to have a branched pentasaccharide repeat (O-unit) containing one residue each of d -galactose, d -mannose, l -rhamnose, d -glucuronic acid, and N -acetyl- d -glucosamine; about 2/3 units bear a side-chain glucose residue. To our knowledge, the F17 O-polysaccharide structure established is unique among known bacterial polysaccharide structures. The O-antigen gene cluster of E. coli F17 between the conserved genes galF and gnd was sequenced and found to be 99% identical to that of E. coli 102,755 assigned to a novel OgN8 genotype (A. Iguchi, S. Iyoda, K. Seto, H. Nishii, M. Ohnishi, H. Mekata, Y. Ogura, T. Hayashi, Front. Microbiol. 7 (2016) 765). Genes in the cluster were annotated taking into account the F17 O-polysaccharide structure. The data obtained confirm that E. coli F17 and E. coli strains belonging to the OgN8 genotype can be considered as a candidate to a new E. coli O-serogroup. The O antigen of this novel type was demonstrated to make for an effective shield protecting the intimate outer membrane surface of bacteria from direct interaction with bacteriophages. [ABSTRACT FROM AUTHOR]

The O-antigen is a part of the outer membrane of Gram-negative bacteria and is related to bacterial virulence. It is one of the most variable cell constituents, and its structural diversity is almost entirely due to genetic variation of the O-antigen gene cluster. In this study, the O-antigen structure of Escherichia coli O62 was elucidated by chemical analysis and nuclear magnetic resonance spectroscopy, but showing not consistent with the O-antigen gene cluster between conserved genes galF and gnd reported earlier. The complete genome of E. coli O62 was then sequenced and analyzed, and another O-antigen gene cluster was found and characterized that correlated perfectly with the established O-antigen structure. A deletion and complementation experiment confirmed the functionality of the novel gene cluster and demonstrated that the O62-antigen is synthesized by the ABC transporter-dependent system. To our knowledge, this is the first report that the O-antigen gene cluster is positioned at a novel locus in E. coli. Comparative analysis indicated that E. coli O62 likely originated from E. coli O68 via an IS event resulting in the repression of the O68-antigen synthesis, followed by the acquisition of a novel O-antigen gene cluster from Enterobacter aerogenes. [ABSTRACT FROM AUTHOR]

Fine structure of the O-polysaccharide chain of the lipopolysaccharide (O-antigen) defines the serospecificity of bacterial cells, which is the basis for O-serotyping of medically and agriculturally important gram-negative bacteria including Escherichia coli . In order to obtain the O-polysaccharide for structural analysis, the lipopolysaccharide was isolated from cells of E. coli O84a by phenol/water extraction and degraded with mild acid. However, the O-polysaccharide was cleaved at a highly acid-labile β- l -fucopyranosyl phosphate (β- l -Fuc p -1- P ) linkage to give mainly a pentasaccharide that corresponded to the O-polysaccharide repeat. Therefore, the lipopolysaccharide and the pentasaccharide as well as their O-deacylated derivatives were studied using sugar analysis, NMR spectroscopy, and (for oligosaccharides) ESI HR MS, and the O84-polysaccharide structure was established. The O-polysaccharide is distinguished by the presence of β- l -Fuc p -1- P and randomly di-O-acetylated 6-deoxy- d -talose, which are found for the first time in natural carbohydrates. The gene cluster for the O84-antigen biosynthesis was analysed and its content was found to be consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

Upon mild acid degradation of the lipopolysaccharide of Escherichia coli O165, the O-polysaccharide chain was cleaved at the glycosidic linkage of 5-N-acetyl-7-N-[(R)-3-hydroxybutanoyl]pseudaminic acid (Pse5Hb7Ac). Analysis of the resulting linear tetrasaccharide and alkali-treated lipopolysaccharide by ¹H/13C 1D and 2D nuclear magnetic resonance spectroscopy enabled elucidation of the following structure of the O-polysaccharide: →8)-α-Psep5Hb7Ac-(2→6)-β-D-Galp-(1→4)-β-D-Gl?p-(1→3)-α-DGl?pNAc-(1→. The β-D-Galp-(1→4)-β-D-Gl?p-(1→3)-D-Gl?pNAc structural element is also present in the O-polysaccharide of E. coli O82. The content of the O-antigen gene cluster of E. coli O165 was found to be consistent with the O-polysaccharide structure established. Functions of proteins encoded in the gene cluster, including enzymes involved in the Pse5Hb7Ac biosynthesis and glycosyltransferases, were putatively assigned by comparison with sequences in available databases. [ABSTRACT FROM AUTHOR]

Abstract: A polysaccharide was isolated by GPC after mild acid treatment of the lipopolysaccharide of Vibrio vulnificus CECT4602 and found to contain l-Rha, d-GlcpNAc and 2-acetamido-2,3,6-trideoxy-3-(3-hydroxybutanoylamino)-l-mannose (l-RhaNAc3NHb). GLC analysis of the trifluoroacetylated (S)-2-octyl esters derived by full acid hydrolysis of the polysaccharide showed that ∼80% of the 3-hydroxybutanoic acid has the S configuration and ∼20% the R configuration. The following structure of the polysaccharide was established by 1H and 13C NMR spectroscopies, including 2D ROESY and 1H/13C HMBC experiments: ▪ [Copyright &y& Elsevier]

The O-specific polysaccharide (OPS) isolated from the lipopolysaccharide of Proteus mirabilis O36 was found to have a pentasaccharide repeating unit of the following structure: →2)-β-d-Rib f-(1→4)-β-d-Gal p-(1→4)-α-d-Glc pNAc6Ac-(1→4)-β-d-Gal p-(1→3)-α-d-Glc pNAc-(1→. The structure is unique among Proteus OPS, which is in agreement with the classification of this strain into a separate Proteus O-serogroup. Remarkably, the P. mirabilis O36-polysaccharide has the same structure as the OPS of Escherichia coli O153, except that the latter is devoid of O-acetyl groups. The cross-reaction of anti-O36 antibodies with the O-part of E. coli O153 lipopolysaccharide is observed. In the present study, two steps of serotyping Proteus strains are proposed: screening of dry mass with enzyme-linked immunosorbent assay and immunoblot with the crude lipopolysaccharides. This method allowed serotyping of 99 P. mirabilis strains infecting the human urinary tract. Three strains were classified into serogroup O36. The migration pattern of these lipopolysaccharides fraction with long O-specific PSs was similar to the standard laboratory P. mirabilis O36 (Prk 62/57) lipopolysaccharide. The relatively low number of clinical strains belonging to serogroup O36 did not correspond to the presence of anti- P. mirabilis O36 antibodies in the blood donors' sera. Twenty-five percent of tested sera contained a statistically significant elevated level of antibodies reacting with thermostable surface antigens of P. mirabilis O36. The presence and amount of antibodies correlated with Thr399Ile TLR4 polymorphism types ( P=0.044). [ABSTRACT FROM AUTHOR]

O-Polysaccharides were obtained from the lipopolysaccharides of Proteus mirabilis CCUG 10704 (OE) and Proteus vulgaris TG 103 and studied by chemical analyses and one- and two-dimensional 1H and 13C nuclear magnetic resonance spectroscopy, including rotating-frame nuclear Overhauser effect spectroscopy, H-detected 1H,13C heteronuclear single-quantum spectroscopy and 1H,31P heteronuclear multiple-quantum spectroscopy experiments. The Proteus mirabilis OE polysaccharide was found to have a trisaccharide repeating unit with a lateral glycerol phosphate group. The Proteus vulgaris TG 103 produces a similar O-polysaccharide, which differs in incomplete substitution with glycerol phosphate ( c. 50% of the stoichiometric amount) and the presence of an O-acetyl group at position 6 of the 2-acetamido-2-deoxygalactose (GalNAc) residue. These structures are unique among the known bacterial polysaccharide structures. Based on the structural and serological data of the lipopolysaccharides, it is proposed to classify both strains studied into a new Proteus serogroup, O54, as two subgroups, O54a,54b and O54a,54c. The serological relatedness of the Proteus O54 and some other Proteus lipopolysaccharides is discussed. [ABSTRACT FROM AUTHOR]

Abstract: An alkali-treated lipopolysaccharide of Proteus penneri strain 60 was studied by chemical analyses and 1H, 13C and 31P NMR spectroscopy, and the following structure of the linear pentasaccharide–phosphate repeating unit of the O-polysaccharide was established:Rabbit polyclonal O-antiserum against P. penneri 60 reacted with both core and O-polysaccharide moieties of the homologous LPS. Based on the unique O-polysaccharide structure and serological data, we propose to classify P. penneri 60 into a new, separate Proteus serogroup O70. A weak cross-reactivity of P. penneri 60 O-antiserum with the lipopolysaccharide of Proteus vulgaris O8, O15 and O19 was observed and discussed in view of the chemical structures of the O-polysaccharides. [Copyright &y& Elsevier]

The O-polysaccharide of Providencia stuartii O33 was obtained by mild acid degradation of the lipopolysaccharide and the following structure of the tetrasaccharide repeating unit was established:→ 6)-α-d-GlcpNAc-(1 → 4)-α-d-GalpA-(1 → 3)-α-d-GlcpNAc-(1 → 3)-β-d-Quip4N(Ac-d-Asp)-(1 →, where d-Qui4N(Ac-d-Asp) is 4-(N-acetyl-d-aspart-4-yl)amino-4,6-dideoxy-d-glucose. Structural studies were performed using sugar and methylation analyses and NMR spectroscopy, including conventional 2D 1H, 1H COSY, TOCSY, NOESY and 1H, 13C HSQC experiments as well as COSY and NOESY experiments in an H2O–D2O mixture to reveal correlations for NH protons. The O-polysaccharide of P. stuartii O33 shares an α-d-Glcp NAc-(1 → 3)-β-d-Quip4N(Ac-d-Asp) epitope with that of Proteus mirabilis O38, which seems to be responsible for a marked serological cross-reactivity of anti-P. stuartii O33 serum with the lipopolysaccharide of the latter bacterium. P. stuartii O33 is serologically related also to P. stuartii O4, whose O-polysaccharide contains a lateral β-d-Qui4N(Ac-l-Asp) residue. [Copyright &y& Elsevier]

The O-polysaccharide of Aeromonas hydrophila O:34 was obtained by mild-acid degradation of the lipopolysaccharide and studied by chemical methods and NMR spectroscopy before and after O-deacetylation. The polysaccharide was found to contain d-Man, d-GalNAc and 6-deoxy-l-talose (l-6dTal), and the following structure of the tetrasaccharide repeating unit was established:where 6dTalI is O-acetylated stoichiometrically at position-2 and 6dTalII carries no, one or two O-acetyl groups at any positions. [Copyright &y& Elsevier]

The following structure of the O‐specific polysaccharide (OPS) of Proteus vulgaris O45 lipopolysaccharide (LPS) was established using 1H‐ and 13C‐NMR spectroscopy, including two‐dimensional NOESY and H‐detected 1H, 13C heteronuclear multiple‐quantum coherence (HMQC) experiments: &figfu1; Immunochemical studies, using rabbit polyclonal anti‐P. vulgaris O45 serum and LPS, OPS and Smith‐degraded OPS of P. vulgaris O45, showed the importance of β‐d‐GlcA in manifesting the serological specificity of the O‐antigen studied. [ABSTRACT FROM AUTHOR]

The following structure of the O-specific polysaccharide chain (O-antigen) of the Proteus vulgaris O32 lipopolysaccharide (LPS) was established by 1H-NMR and 13C-NMR spectroscopy, including two-dimensional NOESY and H-detected 1H,13C heteronuclear multiple-quantum coherence (HMQC) experiments : ←2)-α-L-RhapI-(1←2)-α-L-RhapII-(1←4)-β-D-GalpAI-(1←3)-β-D-GlcpNAc-(1←4)-α-D-GalpAII-(1←. In addition, an O-acetyl group was detected, which, most probably, is located at position 3 of a part of RhapI residues. Serological studies, using rabbit polyclonal anti-(P. vulgaris O32) serum, homologous and heterologous Proteus O-antigens and related artificial antigens, revealed the importance of an α-D-GalA-associated epitope in manifesting the immunospecificity of P. vulgaris O32 and substantiated serological relationships between the O-antigen studied and those of some other Proteus strains. [ABSTRACT FROM AUTHOR]

Abstract: Cronobacter sakazakii G2706 and G2704 are the reference strains of serotypes O5 and O6 in the serological classification of this species proposed recently. Mild acid degradation of the lipopolysaccharides of both strains resulted in cleavage of the O-polysaccharide chains at the acid-labile linkage of 3-deoxy-d-manno-oct-2-ulosonic acid (Kdo) to yield oligosaccharides representing repeating units of the O-polysaccharides. The oligosaccharides and alkali-degraded lipopolysaccharides were studied by sugar analysis along with 1D and 2D 1H and 13C NMR spectroscopy, and the following O-polysaccharide structures were established:▪ The structure of strain G2706 is unique among the known bacterial polysaccharide structures, whereas that of strain G2704 is identical to the structure of Cronobacter malonaticus 3267 [MacLean, L. L.; Vinogradov, E.; Pagotto, F.; Farber, J. M.; Perry, M. B. Biochem. Cell Biol. 2009, 87, 927–932], except for that the latter lacks O-acetylation. Putative functions of the genes in the O-antigen gene clusters of C. sakazakii strains studied are in agreement with the O-polysaccharide structures. [Copyright &y& Elsevier]

Abstract: The O-polysaccharide of Vibrio cholerae O43 was studied using chemical analyses, triflic acid solvolysis and 2D NMR spectroscopy, including 1H/1H COSY, TOCSY, NOESY and 1H/13C gradient-selected HSQC experiments. The following structure of the tetrasaccharide repeating unit of the polysaccharide was established: →3)-β-d-Quip4NAcyl-(1→3)-α-d-GalpNAcA-(1→4)-α-d-GalpNAc-(1→3)-α-d-QuipNAc-(1→ where d-QuiNAc stands for 2-acetamido-2,6-dideoxy-d-glucose, d-Qui4NAcyl for 4-(N-acetyl-l-allothreonyl)amino-4,6-dideoxy-d-glucose and d-GalNAcA for 2-acetamido-2-deoxy-d-galacturonic acid. [Copyright &y& Elsevier]

Abstract: A polysaccharide isolated after mild acid degradation of the lipopolysaccharide of Vibrio vulnificus clinical isolate YJ016 was found to contain l-Fuc, d-GlcpNAc, 2,4-diacetamido-2,4,6-trideoxy-d-glucose (di-N-acetylbacillosamine, d-QuiNAc4NAc), and 2-acetimidoylamino-2-deoxy-l-galacturonic acid (l-GalNAmA). The last sugar derivative was confirmed by correlations for nitrogen-linked protons in 2D TOCSY and ROESY spectra measured in a H2O–D2O mixture. The following structure of the polysaccharide was established by 1H and 13C NMR spectroscopy, including 2D ROESY and 1H,13C HMBC experiments: Display Omitted where the degree of 6-O-acetylation of the lateral β-GlcNAc residue is ∼70%. [Copyright &y& Elsevier]

The serological classification scheme of the opportunistic Proteus bacilli includes a number of Proteus penneri strains. The tested P. penneri 4034-85 strain turned out to be serologically distinguished in ELISA and Western blotting. The O-polysaccharide was obtained by mild acid degradation of the lipopolysaccharide of this strain and studied by sugar and methylation analyses and dephosphorylation along with 1H and 13C NMR spectroscopy, including 2D 1H,1H COSY, TOCSY, ROESY, 1H,13C HSQC, HMBC, and HSQC-TOCSY experiments, The O-polysaccharide was found to have a linear repeating unit containing glycerol 1-phosphate and two residues each of Gal and GlcNAc. The following O-polysaccharide structure was established, which, to our knowledge, is unique among known bacterial polysaccharide structures: →4)-β- d -Glc p NAc-(1→3)-α- d -Gal p -(1→3)-β- d -Glc p NAc-(1→2)-β- d -Gal p -(1→3)-Gro-1- P -(O→. We identified a unique O-specific polysaccharide from a clinical Proteus penneri strain, creating a new O83 serogroup in the genus Proteus →4)-β- d -Glc p NAc-(1→3)-α- d -Gal p -(1→3)-β- d -Glc p NAc-(1→2)-β- d -Gal p -(1→3)-Gro-1- P -(O→. Unlabelled Image [ABSTRACT FROM AUTHOR]

Mild acid degradation of the lipopolysaccharide of Yersinia kristensenii C-134 afforded a glycerol teichoic acid-like O-polysaccharide, which was studied by sugar analysis, O-deacetylation and dephosphorylation along with 1D and 2D NMR spectroscopy. The following structure of the O-polysaccharide was established: This structure is related to those of other Y. kristensenii O-polysaccharides studied earlier. The O-antigen gene cluster of Y. kristensenii С-134 was analyzed and found to be consistent with the O-polysaccharide structure established. Image 1 • Structure of the O-polysaccharide of Yersinia kristensenii C-134 was established. • The O-polysaccharide has a glycerol teichoic acid-like structure. • Dephosphorylation of the O-polysaccharide afforded a glycerol-linked hexasaccharide. • The O-antigen gene cluster was found to be consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

A 3,6-dideoxy- l - xylo -hexose (colitose)-containing partially O-acetylated branched polysaccharide was obtained by mild acid hydrolysis (2% HOAc, 100 °C, 2 h) of the lipopolysaccharide of Escherichia albertii HK18069 followed by gel-permeation chromatography on Sephadex G-50 Superfine. Part of colitose residues (~40%) was cleaved upon hydrolysis, and the full cleavage was achieved by prolonged hydrolysis (8 h) under the same conditions and resulted in a modified linear polysaccharide. Structure of the O-polysaccharide of E. albertii HK18069 was established by 1D and 2D 1H and 13C NMR spectroscopy applied to both initial and modified O-deacetylated and colitose-free polysaccharides: Image 2 where β- d -Gal p is mono-O-acetylated at position either 3 (~50%) or 4 (~30%). The O-antigen gene cluster of E. albertii HK18069 between conserved galF and gnd genes together with flanking regions was sequenced, and predicted functions of the genes were found to be consistent with the O-polysaccharide structure established. The O-polysaccharide structure and the O-antigen gene cluster of E. albertii HK18069 are related to those of Esherichia coli O55 and E. coli O128 reported earlier. It is proposed to create for strain HK18069 a new E. albertii O-serogroup, O8. Image 1 • A colitose-containing O-polysaccharide is harbored by E. albertii HK18069 (serotype O8). • Structure of the O-polysaccharide was established by sugar analysis and NMR spectroscopy. • O-antigen gene cluster of E. albertii HK18069 was sequenced and the genes were annotated. • Predicted gene functions are consistent with the O-polysaccharide established. • O-antigen structures and gene clusters of E. albertii O8 and E. coli O55 and O128 are related. [ABSTRACT FROM AUTHOR]

O-specific polysaccharide was obtained by mild acid hydrolysis of the lipopolysaccharide of plant-growth-promoting rhizobacteria Azospirillum doebereinerae GSF71T and studied by sugar analysis along with 1H and 13C NMR spectroscopy, including 2D 1H,1H COSY, TOCSY, ROESY, 1H,13C HSQC, and HMBC experiments. It was established that the polysaccharide is linear and consists of tetrasaccharide repeating units with the following structure: Image 1 • OPS was isolated from Azospirillum doebereinerae GSF71T lipopolysaccharide. • Structure of the OPS was established by chemical analyses and NMR spectroscopy. • The OPS is built up of linear tetrasaccharide repeating units. • The OPS contains fully O-acetylated Rha and partially O-acetylated GlcNAc. [ABSTRACT FROM AUTHOR]

The current serological classification scheme of the medically important bacteria from the genus Proteus consists of 80 O serogroups, the last four of which (O77–O80) were created from clinical strains from Łódź, Poland. There are more serologically unique strains isolated from patient that do not fit into the existing scheme, such as Proteus mirabilis strain Sm 99 isolated from urine of a 74-year-old woman in Łódź. Serological investigation involving ELISA and Western blotting failed to classify the Proteus mirabilis strain Sm 99 into any of the 80 Proteus O serogroups. Sugar analysis along with two-dimensional NMR spectroscopy showed that the O-polysaccharide is composed of branched pentasaccharide repeating units containing one residue each of d -Glc, d -GlcNAc, d -GalNAc, d -glucuronic acid, and 4-[( R )-3-hydroxybutanoylamino]-4,6-dideoxy- d -glucose. The chemical and serological data show that the O antigen of P . mirabilis Sm 99 is unique among the known Proteus O antigens. Based on this finding, it is proposed to extend the current serological classification scheme of Proteus by adding a new serogroup, O81, which at present consists of P . mirabilis strain Sm 99 only. [ABSTRACT FROM AUTHOR]

A capsular polysaccharide (CPS) was isolated from strain MAR13-1452 of an emerging pathogen Acinetobacter baumannii and assigned type K125. The following structure of the CPS was established by sugar analysis, Smith degradation, and 1D and 2D 1 H and 13 C NMR spectroscopy: Unlabelled Image Proteins encoded by the KL125 gene cluster in the genome of MAR13-1452, including three glycosyltransferases, were assigned roles in the biosynthesis of the K125 CPS. [ABSTRACT FROM AUTHOR]

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 1 H and 13 C NMR spectroscopy: →3)-α- l -Rha p -(1 → 2)-α- l -Rha p -(1 → 3)-β- l -Rha p -(1 → 4)-β- d -Glc p NAc-(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 -Fuc p 3NAc 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 -Fuc p 3NAc from the O50 polysaccharide is evidently a point mutation in the aminotransferase gene fdtB of the d -Fuc p 3NAc synthesis pathway resulting in a single amino acid change from histidine in O2 to arginine in O50. [ABSTRACT FROM AUTHOR]

Mild acid hydrolysis of the lipopolysaccharide of Escherichia coli O54 afforded an O-polysaccharide, which was studied by sugar analysis, solvolysis with anhydrous trifluoroacetic acid, and 1 H and 13 C NMR spectroscopy. Solvolysis cleaved predominantly the linkage of β- d -Rib f and, to a lesser extent, that of β- d -Glc p NAc, whereas the other linkages, including the linkage of α- l -Rha p, were stable under selected conditions (40 °C, 5 h). The following structure of the O-polysaccharide was established: →4)-α- d -Gal p A-(1 → 2)-α- l -Rha p -(1 → 2)-β- d -Rib f -(1 → 4)-β- d -Gal p -(1 → 3)-β- d -Glc p NAc-(1→ The O-antigen gene cluster of E. coli O54 was analyzed and found to be consistent in general with the O-polysaccharide structure established but there were two exceptions: i) in the cluster, there were genes for phosphoserine phosphatase and serine transferase, which have no apparent role in the O-polysaccharide synthesis, and ii) no ribofuranosyltransferase gene was present in the cluster. Both uncommon features are shared by some other enteric bacteria. [ABSTRACT FROM AUTHOR]

An O-polysaccharide was isolated from the lipopolysaccharide of Escherichia albertii O2 and studied by chemical methods and 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure of the O-polysaccharide was established: Image 2 . The O-polysaccharide is characterized by masked regularity owing to a non-stoichiometric O-acetylation of an l -fucose residue in the main chain and a non-stoichiometric side-chain l -fucosylation of a β-GlcNAc residue. A regular linear polysaccharide was obtained by sequential Smith degradation and alkaline O-deacetylation of the O-polysaccharide. The content of the O-antigen gene cluster of E. albertii O2 was found to be essentially consistent with the O-polysaccharide structure established. [ABSTRACT FROM AUTHOR]

Lipopolysaccharide was isolated from bacteria Yersinia intermedia H9-36/83 (O:17) and degraded with mild acid to give an O-specific polysaccharide, which was isolated by GPC on Sephadex G-50 and studied by sugar analysis and 1D and 2D NMR spectroscopy. The polysaccharide was found to contain 3-deoxy-3-[( R )-3-hydroxybutanoylamino]- d -fucose ( d -Fuc3N R 3Hb) and the following structure of the heptasaccharide repeating unit was established: Image 2 The structure established is consistent with the gene content of the O-antigen gene cluster. The O-polysaccharide structure and gene cluster of Y. intermedia are related to those of Hafnia alvei 1211 and Escherichia coli O:103. [ABSTRACT FROM AUTHOR]

An O-specific polysaccharide was isolated by mild acid degradation of the lipopolysaccharide of Escherichia coli O33 followed by gel-permeation chromatography on Sephadex G-50. The polysaccharide was found to contain glycerol 2-phosphate (Gro-2- P ), and the following structure of its tetrasaccharide repeat was established by sugar analysis, dephosphorylation, and 1D and 2D 1 H and 13 C NMR spectroscopy: Image 2 The O33-antigen gene cluster was analyzed and found to be essentially consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

The O-specific polysaccharide (O-antigen) was obtained by mild acid degradation of the lipopolysaccharide of Escherichia albertii O5 (strain T150248) and studied by sugar analysis, selective cleavages of glycosidic linkages, and 1D and 2D 1 H and 13 C NMR spectroscopy. Partial solvolysis with anh (anhydrous) CF 3 CO 2 H and hydrolysis with 0.05 M CF 3 CO 2 H cleaved predominantly the glycosidic linkage of β-Gal p NAc or β-Gal f, respectively, whereas the linkages of α-Glc p NAc and β-Gal p were stable. Mixtures of the corresponding tri- and tetra-saccharides thus obtained were studied by NMR spectroscopy and high-resolution ESI MS. The following new structure was established for the tetrasaccharide repeat (O-unit) of the O-polysaccharide: →4)-α- d -Glc p NAc-(1 → 4)-β- d -Gal p 6Ac-(1 → 6)-β- d -Gal f -(1 → 3)-β- d -Gal p NAc-(1→where the degree of O-acetylation of d -Gal p is ∼70%. The O-polysaccharide studied has a β- d -Gal p -(1 → 6)-β- d -Gal f -(1 → 3)-β- d -Gal p NAc trisaccharide fragment in common with the O-polysaccharides of E. albertii O7, Escherichia coli O124 and O164, and Shigella dysenteriae type 3 studied earlier. The orf5-7 in the O-antigen gene cluster of E. albertii O5 are 47%, 78%, and 75% identical on the amino acid level to genes for predicted enzymes of E. albertii O7, including Gal p -transferase wfeS, UDP- d -Gal p mutase glf, and Gal f -transferase wfeT , respectively, which are putatively involved with the synthesis of the shared trisaccharide fragment of the O-polysaccharides. The occurrence upstream of the O-antigen gene cluster of a 4-epimerase gene gnu for conversion of undecaprenyl diphosphate-linked d -GlcNAc (UndPP- d -GlcNAc) into UndPP- d -GalNAc indicates that d -GalNAc is the first monosaccharide of the O-unit, and hence the O-units are interlinked in the O-polysaccharide of E. albertii O5 by the β- d -Gal p NAc-(1 → 4)-α- d -Glc p NAc linkage. [ABSTRACT FROM AUTHOR]

Two specific polysaccharides, together with an →4)-α- d -Glc p -(1→ glucan (bacterial glycogen), were obtained from a lipopolysaccharide preparation isolated from the bacterium Pseudomonas putida BIM B-1100 by phenol/water extraction. The following structures of the polysaccharides were established by composition analysis, Smith degradation, ESI-MS, and 1D and 2D NMR spectroscopy: Image 2 [ABSTRACT FROM AUTHOR]

The O-specific polysaccharides (OPSs) called O-antigens were obtained by mild acid degradation of the lipopolysaccharides of Escherichia albertii serotypes O3, O4, O6, and O7 and studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure was established for the OPS of E. albertii O4, which, to our knowledge, is unique among known bacterial polysaccharide structures: →2)-α- l -Rha p -(1 → 2)-α- l -Fuc p- (1 → 2)-β- d -Gal p- (1 → 3)-α- d -Gal p NAc - (1 → 3)-β- d -Glc p NAc - (1→ The OPS structure of the strain of E. albertii O7 studied was identical to that of strain LMG 20973 (= Albert 10457), whose structure has been reported earlier (R. Eserstam et al. Eur. J. Biochem. 269 (2002) 3289–3295). E. albertii O3 and O6 shared the OPS structures with Escherichia coli O181 and O3, respectively, except for the lack of O-acetylation in E. albertii O3, which is present in E. coli O181. The gene clusters driving the O-antigen biosynthesis of the E. albertii strains were sequenced, the genes were annotated by comparison with sequences in the available databases, and the predicted functions of the encoded proteins were found to be consistent with the OPS structures established. In accordance with the relatedness of the OPS structures, the O-antigen gene clusters of E. albertii O3 and O6 contain the same genes and have the same organization as those of E. coli O181 and O3, the entire gene clusters being 83% and 98% identical, respectively. [ABSTRACT FROM AUTHOR]

An O-specific polysaccharide was obtained by mild acid degradation of the lipopolysaccharide of bacteria Pseudomonas putida TSh-18, capable of degrading non-ionogenic technical detergents. The polysaccharide was found to contain a rarely occurring sugar derivative 4,6-dideoxy-4-[( R )-3-hydroxybutanoylamino]- d -galactose [ d -Fuc p 4N( R Hb)]. Sugar and methylation analyses, Smith degradation, solvolysis with CF 3 CO 2 H, and 1 H and 13 C NMR spectroscopy enabled elucidation of the following structure of the branched trisaccharide repeating units of the polysaccharide: Image 2 [ABSTRACT FROM AUTHOR]

An O-polysaccharide was isolated by mild acid degradation of the lipopolysaccharide of Enterobacter cloacae C4115 and studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure of the linear pentasaccharide repeating unit of the O-polysaccharide was established: →2)-α- l -Rha p -(1 → 2)-α- l -Rha p -(1 → 2)-α- l -Rha p -(1 → 2)-α- d -Gal p -(1 → 3)-α- d -Fuc p NAc-(1→ The O-antigen gene cluster of E. cloacae C4115 was sequenced. The gene functions were tentatively assigned by comparison with sequences in the available databases and found to be in consistence with the O-polysaccharide structure. The O-antigen structure and gene cluster of E. cloacae C4115 are similar to those of E. cloacae G3421 studied by us earlier (Perepelov A.V. et al. Carbohydr. Res. 427 (2016) 55–59). [ABSTRACT FROM AUTHOR]

The O-specific polysaccharide (O-antigen) was obtained by mild acid degradation of the lipopolysaccharide of Escherichia albertii serotype O1 strain SP20140089 and studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure was established for the trisaccharide repeating unit of the O-polysaccharide: →4)-β- d -Man p NAc3NAcA-(1 → 4)-β- d -Glc p NAm3NAcA-(1 → 3)-α -d -Glc p NAc-(1→ where ManNAc3NAcA and GlcNAm3NAcA indicate 2,3-diacetamido-2,3-dideoxymannuronic acid and 2-acetimidoylamino-3-acetamido-2,3-dideoxyglucuronic acid, respectively. While showing some similarity with O-polysaccharide structures of a group of Pseudomonas aeruginosa serotypes (O2, O5, O16, O18, and O20), that of E. albertii O1 is unique among known bacterial polysaccharide structures. The gene cluster for biosynthesis of the O1-antigen was sequenced and functions of the genes were predicted by comparison with sequences in the available databases, including those involved in the synthesis of nucleotide precursors of 2,3-diamino-2,3-dideoxyhexuronic acid derivatives in P. aeruginosa O5. [ABSTRACT FROM AUTHOR]

On mild acid degradation of the lipopolysaccharide of Enterobacter cloacae C5529, the O-polysaccharide chain was cleaved at the linkages of 5,7-diacetamido-3,5,7,9-tetradeoxy- l - glycero - l - manno -non-2-ulosonic acid (di- N -acetylpseudaminic acid, Pse p 5Ac7Ac). The resultant oligosaccharide and an alkali-treated lipopolysaccharide were studied by sugar analysis along with 1 H and 13 C NMR spectroscopy, and the following structure of the tetrasaccharide repeating unit of the O-polysaccharide was established: →4)-β-Pse p 5Ac7Ac-(2 → 3)-β- d -Gal p -(1 → 6)-β- d -Gal f -(1 → 3)-α- d -Gal p -(1→ It differs from a structurally related O-polysaccharide of E. cloacae G3045 studied early (Perepelov, A. V.; Wang, M.; Filatov, A. V.; Guo, X.; Shashkov, A. S.; Wang, L.; Knirel, Y. A. Carbohydr. Res. 2015; 407:59–62) in positions of substitution of β-Pse p 5Ac7Ac (O-4 vs. O-8) and β-Gal p (O-3 vs. O-6) and the absence of a side-chain α-Gal p residue. The O-antigen gene clusters of E. cloacae C5529 and G3045 are organized identically and include genes with the same putative functions in the O-polysaccharide synthesis. Based on these and serological data, it is suggested to combine E. cloacae C5529 and G3054 in one O-serogroup as two subgroups. [ABSTRACT FROM AUTHOR]

The O-polysaccharide (O-antigen) of Escherichia coli O131 was studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure of the linear tetrasaccharide repeating unit of the polysaccharide was established: →8)-α-Neu p 5Ac-(2 → 6)-β- D -Gal p -(1 → 6)-β-D-Gal p -(1 → 3)-β-D-Gal p NAc-(1→ The gene functions were tentatively assigned by comparison with sequences in the available databases and found to be in agreement with the E. coli O131-antigen structure. [ABSTRACT FROM AUTHOR]

O-polysaccharides (OPSs) were obtained by mild acid degradation of the lipopolysaccharides of Escherichia coli O182-O187, and their structures were established by sugar analysis, Smith degradation, and 1 H and 13 C NMR spectroscopy. In addition to the monosaccharides that occur often in E. coli OPSs ( d -Glc, d -Gal, d -Man, d -GlcNAc, d -GalNAc, d -GlcA, l -Fuc, d -Rib), a number of less common components were identified as the OPS constituents, including 2-acetamido-2-deoxy- l -quinovose and 4-deoxy-4-[( S )-3-hydroxybutanoyl- l -alanyl]- d -quinovose (O186), 3-acetamido-3-deoxy- d -fucose (O187), 3-deoxy-3-[( R )-3-hydroxybutanoyl]- d -fucose (O184), and 2,3-diacetamido-2,3-dideoxy- l -rhamnose (O182). The OPS structures of E. coli O183 and O182 are identical to those of the OPS of Shigella boydii type 10 and the capsular polysaccharide of E. coli K48, respectively. The OPSs of E. coli O186 and O123 are closely related differing in the presence of a Glc residue in the former in place of a GlcNAc residue in the latter. The O-antigen gene clusters of the bacteria studied were analyzed and their contents were found to be consistent with the OPS structures. Predicted glycosyltransferases encoded in the gene clusters were tentatively assigned to glycosidic linkages based on similarities to sequences of other E. coli O-serogroups available from GenBank and taking into account the OPS structures established. [ABSTRACT FROM AUTHOR]

Mild alkaline degradation of the lipopolysaccharide of Escherichia coli O80 afforded a polysaccharide, which was studied by sugar analysis, selective cleavage of glycosidic linkages, and 1 H and 13 C NMR spectroscopy. Solvolysis of the polysaccharide with CF 3 CO 2 H cleaved the linkages of α-Fuc and β-linked GlcNAc and GalNAc residues to give two disaccharides. The following structure of the hexasaccharide repeating unit of the O-polysaccharide was established: The polysaccharide repeat also contains a minor O-acetyl group but its position was not determined. The O-antigen gene cluster of E. coli O80 between the conserved galF and gnd genes was analyzed and found to be consistent with the O-polysaccharide structure established. [ABSTRACT FROM AUTHOR]

The lipopolysaccharide of Escherichia coli O156 was degraded under mild acidic and alkaline conditions and the resulting polysaccharides were studied by sugar analysis and 1 H and 13 C NMR spectroscopy. The following structure of the pentasaccharide repeating unit of the O-polysaccharide was established: where R pyr indicates R -configurated pyruvic acid acetal. Minor O-acetyl groups also were present and tentatively localized on the Gal residues. The gene cluster for biosynthesis of the O-antigen of E. coli O156 was analyzed and shown to be consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

The O-polysaccharide was isolated by mild acid degradation of the lipopolysaccharide of Enterobacter cloacae G3421 and studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. In addition, partial solvolysis with anhydrous trifluoroacetic acid was applied, which cleaved selectively the α- l -rhamnopyranosidic linkages. The following structure of the branched hexasaccharide repeating unit was established. The O-polysaccharide studied shares the β- l -Rha p -(1→4)-α- l -Rha p -(1→2)-α- l -Rha p trisaccharide fragment with the O-polysaccharide of Shigella boydii type 18. The O-antigen gene cluster of E. cloacae G3421 was sequenced. Functions of genes in the cluster, including those for glycosyltransferases, were tentatively assigned by a comparison with sequences in the available databases and found to be consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

Mild acid degradation of the lipopolysaccharide of Escherichia coli O132 released its O-polysaccharide. Analysis by 1D and 2D 1 H and 13 C NMR spectroscopy prior and subsequent to O-deacetylation, in conjunction with sugar analysis, revealed a linear pentasaccharide repeating unit of the O-polysaccharide having the following structure: →2)-α- d -Gal f -(1→3)-α- l -Rha p 2Ac-(1→4)-α- d -Glc p -(1→2)-α- l -Rha p -(1→3)-β- d -Glc p NAc-(1→ Putative functions of genes in the O-antigen gene cluster of E. coli O132 are consistent with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

O-polysaccharides of E. coli O28ab, O37, and O100 were found to contain glycerol 1–phosphate and the following structures of their oligosaccharide repeats were established by sugar analysis, Smith degradation (for O28ab), 1D and 2D 1 H, 13 C, and 13 P NMR spectroscopy: Functions of putative glycosyltransferases genes in the O-antigen gene clusters of the strains studied were tentatively assigned based on similarities to genes of other E. coli O-serogroups available from GenBank and taking into account the O-polysaccharide structures established. [ABSTRACT FROM AUTHOR]

The O-polysaccharide (O-antigen) was isolated from the lipopolysaccharide of Escherichia coli O137 and studied by sugar analysis and NMR spectroscopy. The following structure of the branched tetrasaccharide repeating unit was established: Both structure and gene cluster of the E. coli O137 polysaccharide are related to those of the E. coli K40 polysaccharide (Amor et al., 1999), which lacks the side-chain glucosylation but contains serine that is amide-linked to GlcA. Functions of genes in the O137-antigen gene cluster were assigned by a comparison with those in K40 and sequences in the available databases. Particularly, predicted glycosyltransferases encoded in the gene cluster were assigned to the formation of three glycosidic linkages in the O-polysaccharide repeating unit. [ABSTRACT FROM AUTHOR]

Mild acid degradation of the lipopolysaccharide of Escherichia coli O96 afforded a mixture of two polysaccharides. The following structure of the pentasaccharide repeating unit of the major polymer was established by sugar analysis, Smith degradation, and 1 H and 13 C NMR spectroscopy: The O-antigen gene cluster of E. coli O96 between conserved galF and gnd genes was found to be consistent with this structure, and hence, the major polysaccharide represents the O96-antigen. The O96-antigen structure and gene cluster are similar to those of E. coli O170, and two proteins encoded in the gene clusters of both bacteria were putatively assigned a function of galactofuranosyltransferases. The minor polymer has the same structure as a peptidoglycan-related polysaccharide reported earlier in Providencia alcalifeciens O45 and several other O-serogoups of this species (Ovchinnikova OG, Liu B, Kocharova NA, Shashkov AS, Kondakova AN, Siwinska M, Feng L, Rozalski A, Wang L, Knirel YA. Biochemistry (Moscow) 2012; 77 :609-15) →4)-β- d -Glc p NAc-(1→4)-β- d -Glc p NAc3( R lac- l Ala)-(1→ where R lac- l Ala indicates ( R )-1-[( S )-1-carboxyethylaminocarbonyl]ethyl. [ABSTRACT FROM AUTHOR]

On mild acid degradation of the lipopolysaccharide of Escherichia coli O160, the O-polysaccharide was cleaved by acid-labile glycosyl phosphate linkages in the main chain. The resultant oligosaccharide and the alkali-treated lipopolysaccharide were studied by sugar analysis along with 1 H and 13 C NMR spectroscopies, and the following structure of the branched pentasaccharide repeating unit of the O-polysaccharide was established: The O-antigen gene cluster of E. coli O160 was found to be consistent with the O-polysaccharide structure established. [ABSTRACT FROM AUTHOR]

Enterotoxigenic Escherichia coli are causative agents of diarrhea in humans as well as animals, and E.   coli O170 belongs to this virotype. Upon mild acid degradation of the lipopolysaccharide of E.   coli O170, the branched O-polysaccharide chain was partially cleaved at β- d -glactofuranosidic linkages to give multiple products, including a linear tetrasaccharide and oligomers thereof. Studies of the acid degradation products and O-deacylated lipopolysaccharide by 1D and 2D 1 H and 13 C NMR spectroscopy enabled elucidation of the following O-polysaccharide structure: Functions of genes in the O-antigen biosynthesis gene cluster were tentatively assigned and found to be in agreement with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

The O-polysaccharide (O-antigen) of Escherichia coli O43 was isolated from the lipopolysaccharide and studied by chemical methods, including sugar analyses, Smith degradation, and solvolysis with anhydrous trifluoroacetic acid, along with 1 H and 13 C NMR spectroscopy. The following structure of the pentasaccharide repeating unit of the O-polysaccharide was established: Functions of genes in the O-antigen gene cluster of E. coli O43 were assigned by a comparison with sequences in the available databases and found to be in agreement with the O-polysaccharide structure. [ABSTRACT FROM AUTHOR]

Two polysaccharides were obtained by mild acid degradation of the lipopolysaccharide of associative nitrogen-fixing bacteria Azospirillum brasilense Jm125A2 isolated from the rhizosphere of a pearl millet. The following structures of the polysaccharides were established by sugar and methylation analyses, Smith degradation, and 1 H and 13 C NMR spectroscopy: Structure 1 has been reported earlier for a polysaccharide from A. brasilense S17 (Fedonenko YP, Konnova ON, Zdorovenko EL, Konnova SA, Zatonsky GV, Shaskov AS, Ignatov VV, Knirel YA. Carbohydr Res 2008;343:810–6), whereas to our knowledge structure 2 has not been hitherto found in bacterial polysaccharides. [ABSTRACT FROM AUTHOR]

The O-polysaccharide (O-antigen) of Escherichia coli O169 was studied by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy. The following structure of the branched hexasaccharide repeating unit was established: The O-polysaccharide of E. coli O169 differs from that of Shigella boydii type 6 only in the presence of a side-chain glucose residue. A comparison of the O-antigen biosynthesis gene clusters between the galF to gnd genes in the genomes of the two bacteria revealed their close relationship. The glycosyltransferase gene responsible for the formation of the β- d -Glc p -(1→6)-α- d -Gal p linkage in the O-antigen was identified in the gene cluster. [ABSTRACT FROM AUTHOR]

Mild acid hydrolysis of the lipopolysaccharide of Escherichia coli O57 afforded an O-polysaccharide, which was isolated by gel permeation chromatography (GPC) and studied by sugar analysis, Smith degradation and solvolysis with trifluoroacetic acid, along with 2D 1 H and 13 C NMR spectroscopy. The O-polysaccharide was found to contain d -Glc, d -Gal, d -GalA, d -GlcNAc, and l -FucNAc, as well as O-acetyl groups. Smith degradation of the O-deacetylated polysaccharide destroyed side-branch β-Glс p and α-Gal p A to give a modified linear polysaccharide. Solvolysis cleaved selectively the linkage of α- l -Fuc p NAc to give a pentasaccharide corresponding to the O-polysaccharide repeat. A comparison of the NMR spectra of the initial and O-deacetylated polysaccharides showed that α-Gal p A is non-stoichiometrically O-acetylated at position either 2 (∼30%) or 3 (∼40%). The following structure of the O-polysaccharide was established, which is unique among known bacterial polysaccharide structures: Image 2 [ABSTRACT FROM AUTHOR]

The following structure of the O-polysaccharide of Escherichia coli HS1/2 serving as a primary receptor for bacteriophage DT57-12 was elucidated by sugar analysis along with 1D and 2D 1 H and 13 C NMR spectroscopy: This structure is shared by E. coli O87 type strain. Putatively assigned functions of genes in the O-antigen gene cluster of E. coli O87 are consistent with the O-polysaccharide structure established. [ABSTRACT FROM AUTHOR]