Structures of the O-polysaccharide chains of the lipopolysaccharides of Xanthomonas campestris pv phaseoli var fuscans GSPB 271 and X campestris pv malvacearum GSPB 1386 and GSPB 2388

O-polysaccharides of phytopathogenic bacteria Xanthomonas campestris were isolated by mild acid degradation of the lipopolysaccharides and studied by sugar and methylation analysis, along with 1H and 13C NMR spectroscopy. The following structures of the repeating units of the polysaccharides of X. campestris pv. phaseoli var. fuscans GSPB 271 (1). and X. campestris pv. malvacearum GSPB 1386 and GSPB 2388 (2). were established:The O-polysaccharides of X. campestris are structurally similar to those of some Pseudomonas syringae strains.     

Structural and serological characterisation of the O-antigenic polysaccharide of the lipopolysaccharide from Acinetobacter baumannii strain 24

Extraction of dry bacteria of Acinetobacter baumannii strain 24 by phenol-water yielded a lipopolysaccharide (LPS) that was studied by serological methods and fatty acid analysis. After immunisation of BALB/c mice with this strain, monoclonal antibody S48-3-13 (IgG(3) isotype) was obtained, which reacted with the LPS in western blot and characterized it as S-form LPS. Degradation of the LPS in aqueous 1% acetic acid followed by GPC gave the O-antigenic polysaccharide, whose structure was determined by compositional analyses and NMR spectroscopy of the polysaccharide and O-deacylated polysaccharide as [carbohydrate structure: see text] where QuiN4N is 2,4-diamino-2,4,6-trideoxyglucose and GalNAcA 2-acetamido-2-deoxygalacturonic acid. The amino group at C-4 of the QuipN4N residues is acetylated in about 2/3 of LPS molecules and (S)-3-hydroxybutyrylated in the rest.     

Structure elucidation of the O-chain from the major lipopolysaccharide of the Xanthomonas campestris strain 642

A novel O-polysaccharide consisting of D-Xylp and L-Rhap in the molar ratio of 1:2.5 was identified as the major component in the lipopolysaccharide fraction of Xanthomonas campestris strain 642, which is responsible for a new bacterial disease of the strawberry plant. Its structure was mainly determined using chemical analysis, Smith degradation and 1D and 2D NMR spectroscopy experiments as: carbohydrate sequence [see text].     

Structure of the O-polysaccharide of Xanthomonas cassavae GSPB 2437

The following structure of the O-polysaccharide of the phytopathogenic bacterium Xanthomonas cassavae GSPB 2437 was determined by sugar analysis along with 1H and 13C NMR spectroscopy: [structure: see text].     

Structural analyses of novel glycerophosphorylated alpha-cyclosophorohexadecaoses isolated from X. campestris pv. campestris

Novel periplasmic anionic cyclic glucans produced by Xanthomonas campestris pv. campestris were isolated by trichloroacetic acid treatment and various chromatographic techniques. No report has been made on the presence of substituted cyclic glucans of the Xanthomonas species. We show, for the first time, that X. campestris pv. campestris produces the anionic cyclic glucans with phosphoglycerol residues, the presence of which can be predicted by analyzing the sequence database with the aid of the NCBI RefSeq database. To analyze the structure of isolated anionic cyclic glucans analyses, we used NMR spectroscopy, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOFMS) and electrospray-ionization mass spectrometry (ESIMS). The results suggest that the novel anionic forms of the cyclic glucans of X. campestris pv. campestris are glycerophosphorylated alpha-cyclosophorohexadecaose with one or two phosphoglycerol substituents at the C-6 positions of the glucose residues.     

Structural characterization of the O-chain polysaccharide of Aeromonas caviae ATCC 15468 lipopolysaccharide

The O-chain polysaccharide produced by a mild acid degradation of Aeromonas caviae ATCC 15468 lipopolysaccharide was found to be composed of L-rhamnose, 2-acetamido-2-deoxy-D-glucose, 2-acetamido-2-deoxy-D-galactose and phosphoglycerol. Subsequent methylation and CE-ESIMS analyses and 1D/2D NMR ((1)H, (13)C and (31)P) spectroscopy showed that the O-chain polysaccharide is a high-molecular-mass acidic branched polymer of tetrasaccharide repeating units with a phosphoglycerol substituent having the following structure: [structure: see text] where Gro represents glycerol and P represents a phosphate group.     

Structural characterization of the O-chain polysaccharide from an environmentally beneficial bacterium Pseudomonas chlororaphis subsp. aureofaciens strain M71

Pseudomonas chlororaphis subsp. aureofaciens strain M71 was isolated from the root of a tomato plant and it was able to control in vivo Fusarium oxysporum f. sp. radicis-lycopersici responsible for the tomato crown and root rot. Recently, strain M71 was evaluated even for its efficacy in controlling Seiridium cardinale, the causal agent of bark canker of common cypress (Cupressus sempervirens L.). Strain M71 ability to persist on the tomato rhizosphere and on the aerial part of cypress plants could be related to the nature of the lipopolysaccharides (LPS) present on the outer membrane and in particular to the O-specific polysaccharide.A neutral O-specific polysaccharide was obtained by mild acid hydrolysis of the lipopolysaccharide from P. chlororaphis subsp. aureofaciens strain M71. By means of compositional analyses and NMR spectroscopy, the chemical repeating unit of the polymer was identified as the following linear trisaccharide.     

The O-specific polysaccharide structure from the lipopolysaccharide of the Gram-negative bacterium Raoultella terrigena

The structure of the repeating unit of the O-specific polysaccharide from the lipopolysaccharide of the enterobacterium Raoultella terrigena was determined by means of chemical and spectroscopical methods and was found to be a linear tetrasaccharide containing a cyclic acetal of pyruvic acid (Pyr) as depicted below.[Carbohydrate structure: see text].     

Structural studies on the exopolysaccharide from Erwinia persicina

The Gram-negative bacterial strain HKI 0380 was isolated from biofilms located on palaeolithic rock paintings in the Cave of Bats in Zuheros, southern Spain. It was identified as the phytopathogenic Erwinia persicina and attracted attention due to the production of considerable quantities of slime. The acidic exopolysaccharide produced by the E. persicina was studied after O-deacylation by sugar and methylation analyses, along with (1)H and (13)C NMR spectroscopy. The following structure of the branched pentasaccharide repeating unit of the O-deacylated exopolysaccharide was established: [carbohydrate structure: see text].     

Structural characterization of the O-specific polysaccharide from the lipopolysaccharide of the fish pathogen Aeromonas bestiarum strain P1S

The O-specific polysaccharide obtained by mild-acid degradation of lipopolysaccharide of Aeromonas bestiarum P1S was studied by sugar and methylation analyses along with ¹H and ¹³C NMR spectroscopy. The sequence of the sugar residues was determined using ¹H,¹H NOESY and ¹H,¹³C HMBC experiments. The O-specific polysaccharide was found to be a high-molecular-mass polysaccharide composed of tetrasaccharide repeating units of the structureSince small amounts of a terminal Quip3N residue were identified in methylation analysis, it was assumed that the elucidated structure also represented the biological repeating unit of the O-specific polysaccharide.     

The Extracellular Proteases of the Phytopathogenic Bacterium Xanthomonas campestris

The culture liquids of three Xanthomonas campestris pv. campestris strains were found to possess proteolytic activity. The culture liquid of strain B611 with the highest proteolytic activity was fractionated by salting-out with ammonium sulfate, gel filtration, and ion-exchange chromatography. The electrophoretic analysis of active fractions showed the presence of two proteases in the culture liquid of strain B611, the major of which was serine protease. The treatment of cabbage seedlings with the proteases augmented the activity of peroxidase in the cabbage roots by 28%.     

野油菜黄单胞菌野油菜致病变种8004菌株wxcA基因与EPS的产量有关

在以前的工作中,采用转座子Tn5 gusA5对野油菜黄单胞菌野油菜致病变种(Xcc)8004菌株进行诱变,获得一批胞外多糖(EPS)合成减少的突变体,对这些突变体的Tn5 gusA5的插入位点进行分析后,发现有两株突变体是wxcA基因不同插入位点的突变体。以前认为wxcA基因与脂多糖(LPS)的O-抗原合成有关而与EPS的合成无关。为明确wxc4基因的功能,对8004菌株的wxcA基因进行缺失,获得的△wxcA突变体的EPS产量与野生型菌株相比,减少了50％,并且一段PCR合成的包含wxcA基因的DNA片段能反式互补△wxcA突变体,恢复突变体的EPS产量。这证实了8004菌株的wxcA基因与EPS的合成产量有关。     

Structural determination of the O-chain polysaccharide from the haloalkaliphilic Halomonas alkaliantarctica bacterium strain CRSS

In hypersaline environments there are plenty of microorganisms belonging to both Bacteria and Archaea domains. These extremophiles have developed biochemical adaptations which comprise the accumulation of molar concentrations of potassium and chloride and the biosynthesis and/or the accumulation of organic osmotic solutes (osmolytes) within the cytoplasm. Moreover, to maintain the turgor of the cells halophiles enhance the production of anionic phospholipids and alter the fatty acid composition of the membrane lipids, but very little is known about adaptational structural changes of the lipopolysaccharides (LPS), the main constituent of the outer leaflet of the outer membrane of Gram-negative bacteria. The aim of this work is to investigate the chemical structure of these LPS in order to provide insight into the adaptation mechanism of halophiles to live at high salt concentration. For this, Halomonas alkaliantarctica, a haloalkaliphilic Gram-negative bacterium isolated from salt sediments of a saline lake in Cape Russell in the Antarctic continent, was cultivated and the LPS were extracted and analysed. The structure of the O-chain of the LPS from H. alkaliantarctica was determined by chemical analysis, 1-D and 2-D NMR spectroscopy. The polysaccharide was constituted of a linear trisaccharidic repeating unit as follows:→3)-β-l-Rhap-(1→4)-α-l-Rhap-(1→3)-α-l-Rhap-(1→A comparison among the O-chain structures of H. alkaliantarctica and other Halomonas species is also reported.     

O-chain structure from the lipopolysaccharide of the human pathogen Halomonas stevensii strain S18214

Halomonas stevensii is a Gram-negative, pathogenic, moderately halophilic bacterium isolated from the blood of a renal care patient. It optimally grows at 30–35 °C at pH 8–9 and at a sea salt concentration ranging from 3.0% to 7.5%. Gram-negative bacterial infections are closely associated with the presence of the lipopolysaccharides (LPSs) on the outer membrane. These molecules consist of three regions covalently linked: the glycolipid (lipid A), the oligosaccharide region (core region), and the O-specific polysaccharide (O-chain, O-antigen). O-antigen seems to play an important role in the colonization step (adherence) and the ability to bypass host defense mechanisms. For this reason the structure elucidation of the O-chain repeating unit is important to improve knowledge about the role of LPS in the host-pathogen interaction. In this paper, we report the complete structure of the O-chain from the LPS of H. stevensii. The bacterial cells were cultivated and LPS was extracted by the PCP (phenol–chloroform–petroleum ether) method. After mild acid hydrolysis, the lipid A was removed by centrifugation and the obtained polysaccharide was analyzed by means of chemical analysis and one- and two-dimensional NMR spectroscopy giving the following structure:     

Structural elucidation of a novel core oligosaccharide backbone of the lipopolysaccharide from the new bacterial species Agrobacterium larrymoorei

Agrobacterium larrymoorei is a Gram-negative phytopathogenic bacterium, which produces tumours on Ficus benjamina plants and differs from other Agrobacteria both genetically and biochemically. The lipopolysaccharide (LPS) plays an important role in the pathogenesis of Agrobacteria. The present paper is the first report on the molecular primary structure of the core region of an Agrobacterium LPS. The following structure of the core and lipid A carbohydrate backbone of an R-form LPS of A. larrymoorei was determined by chemical degradations and 1D and 2D NMR spectroscopy: [carbohydrate structure: see text] All sugars are alpha-D-pyranoses if not stated otherwise, Kdo is 3-deoxy-D-manno-oct-2-ulosonic acid, Qui3NAcyl is 3,6-dideoxy-3-(3-hydroxy-2,3-dimethyl-5-oxoprolylamino)glucose, GlcAN and GalAN are amides of GlcA and GalA.     

Influence of Acidic Exopolysaccharide of Xanthomonas campestris IBPM 124 on the Kinetic Parameters of Extracellular Bacteriolytic Enzymes

Interactions of a negatively charged exopolysaccharide of Xanthomonas campestris IBPM 124 with its extracellular enzymes (muramidase, endopeptidase, and neutral phosphatase) and also with egg lysozyme, lysostaphin, muramidase of Streptomyces globisporus, and a bacteriolytic enzyme complex of Streptomyces albus were studied. All these enzymes were positively charged under the conditions of their maximal activity. It was shown that interaction of the acidic exopolysaccharide from X. campestris with these enzymes changed their kinetic parameters. The change was either positive (increase in reaction rate) or negative (decrease in reaction rate) and depended on the enzyme and type of substrate cleaved. Due to such interactions, the acidic exopolysaccharide secreted by X. campestris into the environment not only retained and transported positively charged exoenzymes into the near-cellular space, but also regulated their activity.     

Elucidation of the O-chain structure from the lipopolysaccharide of Agrobacterium tumefaciens strain C58

A linear homopolysaccharide built of 3-alpha-L-6dTalp residues, randomly acetylated at position C-4, is described for the O-specific polysaccharide of Agrobacterium tumefaciens strain C58. This structure, determined by spectroscopical and chemical methods, is strictly correlated to that of Rhizobium loti strain NZP2213, which differs for the degree and the position of O-acetylation.     

The structure of the O-specific polysaccharide of the lipopolysaccharide from Burkholderia gladioli pv. agaricicola

A neutral O-specific polysaccharide containing d-mannose, d-rhamnose and d-galactose was obtained by mild acid hydrolysis of the lipopolysaccharide of the plant pathogenic bacterium Burkholderia gladioli pv. agaricicola. By means of compositional analyses and NMR spectroscopy, the chemical repeating unit of the polymer was identified as a linear trisaccharide of the structure shown below, in which the mannose residue was quantitatively acetylated at C-2. [carbohydrate structure: see text]