Structural studies of the O polysaccharides from the lipopolysaccharides of Azospirillum thiophilum BV-ST and Azospirillum griseum L-25-5w-1T.

Diazotrophic bacteria of the genus Azospirillum are known widely, because they are ubiquitous in the rhizosphere and can promote the growth and performance of nonlegume plants. Recently, more Azospirillum species have been isolated from sources other than plants or soil. We report the structures of the O polysaccharides (OPSs) from the lipopolysaccharides of the type strains A. thiophilum BV-ST (1) and A. griseum L-25-5w-1T (2), isolated from aquatic environments. Both structures have a common tetrarhamnan in the repeating-unit, which is decorated with a side xylose in the OPS of A. thiophilum BV-ST.

Structure, Physicochemical Properties and Biological Activity of Lipopolysaccharide from the Rhizospheric Bacterium Ochrobactrum quorumnocens T1Kr02, Containing d-Fucose Residues.

Lipopolysaccharides (LPSs) are major components of the outer membranes of Gram-negative bacteria. In this work, the structure of the O-polysaccharide of Ochrobactrum quorumnocens T1Kr02 was identified by nuclear magnetic resonance (NMR), and the physical-chemical properties and biological activity of LPS were also investigated. The NMR analysis showed that the O-polysaccharide has the following structure: →2)-ß-d-Fucf-(1→3)-ß-d-Fucp-(1→. The structure of the periplasmic glucan coextracted with LPS was established by NMR spectroscopy and chemical methods: →2)-ß-d-Glcp-(1→. Non-stoichiometric modifications were identified in both polysaccharides: 50% of d-fucofuranose residues at position 3 were O-acetylated, and 15% of d-Glcp residues at position 6 were linked with succinate. This is the first report of a polysaccharide containing both d-fucopyranose and d-fucofuranose residues. The fatty acid analysis of the LPS showed the prevalence of 3-hydroxytetradecanoic, hexadecenoic, octadecenoic, lactobacillic, and 27-hydroxyoctacosanoic acids. The dynamic light scattering demonstrated that LPS (in an aqueous solution) formed supramolecular particles with a size of 72.2 nm and a zeta-potential of -21.5 mV. The LPS solution (10 mkg/mL) promoted the growth of potato microplants under in vitro conditions. Thus, LPS of O. quorumnocens T1Kr02 can be recommended as a promoter for plants and as a source of biotechnological production of d-fucose.

Structure of the O-specific polysaccharide of Ochrobactrum endophyticum KCTC 42485T containing 3-(3-hydroxy-2,3-dimethyl-5-oxoprolyl)amino-3,6-dideoxy-d-galactose.

Ochrobactrum endophyticum (syn. Brucella endophytica) is an aerobic species of Alphaproteobacteria isolated from healthy roots of Glycyrrhiza uralensis. Here we report the structure of the O-specific polysaccharide obtained by mild acid hydrolysis of the lipopolysaccharide of the type strain KCTC 42485:→3)-α-l-FucpNAc-(1→3)-ß-d-QuipNAc-(1→2)-ß-d-Fucp3NAcyl-(1→ where Acyl is 3-hydroxy-2,3-dimethyl-5-oxoprolyl. The structure was elucidated using chemical analyses along with 1H and 13C NMR spectroscopy (including 1H,1H COSY, TOCSY, ROESY and 1H,13C HSQC, HMBC, HSQC-TOCSY and HSQC-NOESY experiments). To our knowledge the OPS structure is novel and has not been previously published.

Epinephrine extensively changes the biofilm matrix composition in Micrococcus luteus C01 isolated from human skin.

The importance of the impact of human hormones on commensal microbiota and microbial biofilms is established in lots of studies. In the present investigation, we continued and extended the research of epinephrine effects on the skin commensal Micrococcus luteus C01 and its biofilms, and also the matrix changes during the biofilm growth. Epinephrine in concentration 4.9 × 10-9 M which is close to normal blood plasma level increased the amount of polysaccharides and extracellular DNA in the matrix, changed extensively its protein, lipid and polysaccharide composition. The Ef-Tu factor was one of the most abundant proteins in the matrix and its amount increased in the presence of the hormone. One of the glucose-mannose polysaccharide was absent in the matrix in presence of epinephrine after 24 h of incubation. The matrix phospholipids were also eradicated by the addition of the hormone. Hence, epinephrine has a great impact on the M. luteus biofilms and their matrix composition, and this fact opens wide perspectives for the future research.

Equine Intestinal O-Seroconverting Temperate Coliphage Hf4s: Genomic and Biological Characterization.

Tailed bacteriophages constitute the bulk of the intestinal viromes of vertebrate animals. However, the relationships between lytic and lysogenic lifestyles of phages in these ecosystems are not always clear and may vary between the species or even between the individuals. The human intestinal (fecal) viromes are dominated mostly by temperate phages, while in horse feces virulent phages are more prevalent. To our knowledge, all the previously reported isolates of horse fecal coliphages are virulent. Temperate coliphage Hf4s was isolated from horse feces, from the indigenous equine Escherichia coli 4s strain. It is a podovirus related to the Lederbergvirus genus (including the well-characterized Salmonella bacteriophage P22). Hf4s recognizes the host O antigen as its primary receptor and possesses a functional O antigen seroconversion cluster that renders the lysogens protected from superinfection by the same bacteriophage and also abolishes the adsorption of some indigenous equine virulent coliphages, such as DT57C, while other phages, such as G7C or phiKT, retain the ability to infect E. coli 4s (Hf4s) lysogens. IMPORTANCE The relationships between virulent and temperate bacteriophages and their impact on high-density symbiotic microbial ecosystems of animals are not always clear and may vary between species or even between individuals. The horse intestinal virome is dominated by virulent phages, and Hf4s is the first temperate equine intestinal coliphage characterized. It recognizes the host O antigen as its primary receptor and possesses a functional O antigen seroconversion cluster that renders the lysogens protected from superinfection by some indigenous equine virulent coliphages, such as DT57C, while other phages, such as G7C or phiKT, retain the ability to infect E. coli 4s (Hf4s) lysogens. These findings raise questions on the significance of bacteriophage-bacteriophage interactions within the ecology of microbial viruses in mammal intestinal ecosystems.

Structural studies on the O-specific polysaccharide of the lipopolysaccharide from Pseudomonas donghuensis strain SVBP6, with antifungal activity against the phytopathogenic fungus Macrophomina phaseolina.

An O-specific polysaccharide (OPS) was isolated from the lipopolysaccharide (LPS) of Pseudomonas donghuensis SVBP6, a bacterium with a broad-spectrum antifungal activity in vitro, particularly that against Macrophomina phaseolina. This latter is one of the most virulent and dangerous pathogens of plants, including soybean which is an economically important crop in Argentina today. The OPS was studied by sugar analysis and spectroscopy (1D and 2D 1H and 13C NMR) showing the following trisaccharide repeating unit: →6)-ɑ-D-ManpNAc-(1 â†’ 3)-ß-l-Rhap-(1 â†’ 4)-ß-D-Glcp-(1→. The crude LPS, the purified LPS and the O-chain were assayed for their antifungal activity against M. phaseolina at 25, 50, 100, and 200 µg plug-1. The results showed that the crude LPS best inhibition was at 200 µg plug-1, able to inhibit the fungus growth by about 45%, while purified LPS and the corresponding OPS, in the same condition, reduced fungus growth by 65%, and 75%, respectively. Furthermore, the purified LPS and OPS significantly reduced the growth of M. phaseolina already at 100 µg plug-1 compared to the crude LPS. The structure of the O-chain is unique among the bacterial LPS and this is the first time that both the antifungal activity of a bacterial LPS and its corresponding O-chain were described.

Structure of cell-wall glycopolymers of Micrococcus luteus C01.

Glycopolymers of two types were isolated from the cell wall of Micrococcus luteus C01 by stepwise extraction with cold and hot 10% aq CCl3CO2H. The following structures of the glycopolymers were established by compositional analysis and 1D and 2D NMR spectroscopy: where L-Glu indicates glutamic acid.

The O-specific polysaccharides structures of Pseudomonas strains isolated from the Ficus elastica.

Two Pseudomonas strains were isolated from the Ficus elastica leaves. The O-antigens were obtained using phenol-water method and mild acid degradation. The following structures of the O-polysaccharides were established by sugar analysis and 2D NMR spectroscopy: OPS of Pseudomonas psychrotolerans BIM B-1171 G -2)[aDGlcp(1-3)]bDRhap(1-3)aDManp(1-3)aDRhap(1- OPS of Pseudomonas sp. BIM B-1172 G -2)bDRhap(1-3)aDRhap(1-3)[aDGlcp(1-2)]aDRhap(1-.

Lipopolysaccharide of Pantoea agglomerans 7460: O-specific polysaccharide and lipid A structures and biological activity.

Lipopolysaccharide (LPS) was isolated from Pantoea agglomerans 7460 cells by phenol-water extraction. Mild acid degradation allowed to separate OPS and lipid A. Lipid A was analyzed by negative-ion mode ESI MS and found to consist mainly of hexaacylated derivative containing biphosphorylated GlcN disaccharide, four 14:0 (3-OH), 18:0 and 12:0 fatty acids. The structure of the O-specific polysaccharide was established by chemical, NMR and computational methods: The LPS of Р. agglomerans 7460 showed low level of toxicity and pyrogenicity to compare with LPS of E. coli O55:B5 and pyrogenal, respectively. The ability of the modified (succinylated) LPS, which have lost its toxicity, to block the toxic effects of native LPS has been shown.

Structure of the O-specific polysaccharide from Azospirillum formosense CC-Nfb-7(T).

The lipopolysaccharide was obtained from the cells of Azospirillum formosense CC-Nfb-7(T), a diazotrophic bacterium isolated from agricultural soil. The O-specific polysaccharide (OPS) was released by mild acid hydrolysis of the lipopolysaccharide and was studied by sugar analysis along with 1H and 13C NMR spectroscopy, including 1H,1H COSY, TOCSY, ROESY, 1H,13C HSQC, and HMBC experiments, and Smith degradation. The following structure of partially methylated OPS composed of trisaccharide repeating units was established.

Structure of O-Polysaccharide and Lipid A of Pantoea Agglomerans 8488.

The Pantoea agglomerans 8488 lipopolysaccharide (LPS) was isolated, purified and characterized by monosaccharide and fatty acid analysis. The O-polysaccharide and lipid A components of the LPS were separated by mild acid degradation. Lipid A was studied by electrospray ionization mass spectrometry (ESI-MS) and found to consist of hexa-, penta-, tetra- and tri-acylated species. Two-dimensional nuclear magnetic resonance (NMR) spectroscopy revealed the following structure of the O-polysaccharide repeating unit →3)-α-L-Rhap-(1→6)-α-D-Manp-(1→3)-α-L-Fucp-(1→3)-ß-D-GlcNAcp-(1→. The LPS showed a low level of toxicity, was not pyrogenic, and reduced the adhesiveness index of microorganisms to 2.12, which was twofold less than the control. LPS modified by complex compounds of germanium (IV) and tin (IV) were obtained. It was found that six LPS samples modified by Sn compounds and two LPS samples modified by Ge compounds lost their toxic activity when administered to mice in a dose of LD50 (105 µg/mice or 5 mg/kg). However, none of the modified LPS samples changed their serological activity in an Ouchterlony double immunodiffusion test in agar.

O-specific polysaccharides structures of Pseudomonas strains isolated from the strawberry leaves.

Two Pseudomonas strains were isolated from the strawberry leaves. The O-antigens were obtained using phenol-water method and mild acid degradation. The following structures of the O-polysaccharides were established by sugar analysis and 2D NMR spectroscopy: OPS of Pseudomonas koreensis BIM B-970G →3)-α-D-FucNAcp-(1 â†’ 2)-ß-D-Quip3NAc-(1 â†’ 3)-α-L-6dTalp4OAc-(1→ OPS of Pseudomonas oryzihabitans BIM B-1072G →4)-α-L-FucpNAm3OAc-(1 â†’ 3)-α-D-QuipNAc-(1 â†’ 4)-ß-D-GlcpNAc3NAcA-(1→ Where Am - acetimidoyl.

Structure, gene cluster of the O antigen and biological activity of the lipopolysaccharide from the rhizospheric bacterium Ochrobactrum cytisi IPA7.2.

Lipopolysaccharide (LPS) of Ochrobactrum cytisi IPA7.2, a bacterium isolated from the roots of Solanum tuberosum L., was extracted from dry bacterial cells and chemically characterized. The O-specific polysaccharide was obtained by mild acid hydrolysis of the LPS and studied by sugar analysis and 1H and 13C NMR spectroscopy, including 1H,1H COSY, 1H,1H TOCSY, 1H,1H ROESY, 1H,13C HSQC, and 1H,13C HMBC experiments. The polysaccharide was linear and consisted of trisaccharide repeating units of the following structure: A putative O-antigen gene cluster of O. cytisi IPA7.2 was identified and found to be consistent with the O-specific polysaccharide structure. The LPS of Ochrobactrum cytisi IPA7.2 promoted the growth of potato microplants in vitro.

Pantoea agglomerans P1a lipopolysaccharide: Structure of the O-specific polysaccharide and lipid A and biological activity.

O-specific polysaccharide and lipid A were obtained from the lipopolysaccharide from new strain of Рantoea agglomerans P1a by mild acid hydrolysis. It was found that the major form of lipid A presented by tetraacylated derivative containing biphosphorylated GlcN disaccharide, three 14:0 (3-OH) and 12:0 residues. The structure of the O-specific polysaccharide was established by chemical, NMR and computational methods: →3)-α-D-Manp-(1 → 4)-ß-D-Fucp-(1 → 4)-ɑ-D-Fucp-(1→The LPS of Р. agglomerans P1a showed low level of toxicity and pyrogenicity to compare with LPS of E. coli O55:B5 and pyrogenal (respectively).

Composition of the Biofilm Matrix of Cutibacterium acnes Acneic Strain RT5.

In skin, Cutibacterium acnes (former Propionibacterium acnes) can behave as an opportunistic pathogen, depending on the strain and environmental conditions. Acneic strains of C. acnes form biofilms inside skin-gland hollows, inducing inflammation and skin disorders. The essential exogenous products of C. acnes accumulate in the extracellular matrix of the biofilm, conferring essential bacterial functions to this structure. However, little is known about the actual composition of the biofilm matrix of C. acnes. Here, we developed a new technique for the extraction of the biofilm matrix of Gram-positive bacteria without the use of chemical or enzymatic digestion, known to be a source of artifacts. Our method is based on the physical separation of the cells and matrix of sonicated biofilms by ultracentrifugation through a CsCl gradient. Biofilms were grown on the surface of cellulose acetate filters, and the biomass was collected without contamination by the growth medium. The biofilm matrix of the acneic C. acnes RT5 strain appears to consist mainly of polysaccharides. The following is the ratio of the main matrix components: 62.6% polysaccharides, 9.6% proteins, 4.0% DNA, and 23.8% other compounds (porphyrins precursors and other). The chemical structure of the major polysaccharide was determined using a nuclear magnetic resonance technique, the formula being →6)-α-D-Galp-(1→4)-ß-D-ManpNAc3NAcA-(1→6)-α-D-Glcp-(1→4)-ß-D-ManpNAc3NAcA-(1→3)-ß-GalpNAc-(1→. We detected 447 proteins in the matrix, of which the most abundant were the chaperonin GroL, the elongation factors EF-Tu and EF-G, several enzymes of glycolysis, and proteins of unknown function. The matrix also contained more than 20 hydrolases of various substrata, pathogenicity factors, and many intracellular proteins and enzymes. We also performed surface-enhanced Raman spectroscopy analysis of the C. acnes RT5 matrix for the first time, providing the surface-enhanced Raman scattering (SERS) profiles of the C. acnes RT5 biofilm matrix and biofilm biomass. The difference between the matrix and biofilm biomass spectra showed successful matrix extraction rather than simply the presence of cell debris after sonication. These data show the complexity of the biofilm matrix composition and should be essential for the development of new anti-C. acnes biofilms and potential antibiofilm drugs.

Structure of the O-specific polysaccharide of Azospirillum doebereinerae type strain GSF71T.

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.

Structures of cell-wall glycopolymers of Lactobacillus rhamnosus BIM B-1039.

Glycopolymers of two types were isolated from the cell wall of Lactobacillus rhamnosus BIM B-1039 by stepwise extraction with cold and hot 10% aq CCl3CO2H followed by anion-exchange gel chromatography. The following structures of the glycopolymers were established by sugar analysis, Smith degradation and 1D and 2D NMR spectroscopy.

Investigation of O-polysaccharides from bacterial strains of Pseudomonas genus as potential receptors of bacteriophage BIM BV-45.

The structure of potential bacteriophage receptors located on cell walls of Gram-negative bacteria deposited at Belarusian collection of microorganisms was investigated. Studies by 1D and 2D 1H and 13C NMR spectroscopy enabled to elucidate the structure of the O-specific polysaccharides (OPS) constituting lipopolysaccharide (LPS) of some Pseudomonas species. The capacity of bacteriophage to adsorb to LPS molecules was tested.

Structures of O-specific polysaccharides of Pseudomonas psyhrotolerans BIM B-1158G.

Strain of Pseudomonas psychrotolerans was cultured on the nutrient agar and in a liquid nutrient broth. Bacterial cells were phage-typed with bacteriophages specific to Pseudomonas. O-antigen was isolated from cells using phenol-water method and mild acid degradation. The following structures of the polysaccharides extracted were established by sugar analysis and 1D, 2D NMR spectroscopy: PSI→3)-α-D-Manp-(1→2)-α-D-Manp-(1→; PSII→3)-α-D-Rhap-(1→2)-ß-D-Rhap-(1→3)-α-D-Rhap-(1→; α-D-Glcp-(1Ë©; 2.