Chemical composition and immunochemical characteristics of the lipopolysaccharide of nitrogen-fixing rhizobacterium Azospirillum brasilense CD

Konnova, O. N.; Burygin, Gennady L.; Fedonenko, Yu. P.; Matora, L. Yu.; Pankin, K. E.; Коннова, С. А.; Игнатов, В. В.

doi:10.1134/s0026261706030143

Cited by 7 publications

(4 citation statements)

References 12 publications

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“…Th e peculiarities of the fatty acid composition of lipid A are oft en used as additional chemotaxonomic criteria in the taxonomy of Gramnegative bacteria. Our results for the presence of saturated, monounsaturated, and hydroxylated acids with a chain length of C14-C18 in the composition of LPS of the studied A. brasilense strains with the dominance of 3-hydroxytetradecanoic, hexadecanoic, 3-hydroxyhexadecanoic, and octadecenoic acids, as well as minor amounts of tetradecanoic, pentadecanoic, and octadecanoic acids, are consistent with data on the fatty acid composition of LPS of another Azospirillum [21,24].…”

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confidence: 90%

Characterization of Azospirillum brasilense Lipopolysaccharides

Bulyhina¹,

Varbanest²

2022

Mikrobiol. Z.

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Azospirillum brasilense is a gram-negative, nitrogen-fixing bacterium that colonizes the rhizosphere of various types of grasses and cereals. Lipopolysaccharides (LPS) are a class of complex glycolipids present in the cell membrane of gramnegative bacteria and mediate plant-bacteria interactions. Although the effects of LPS of pathogenic plant bacteria on the induction of plant defense mechanisms have been characterized, the role of LPS of beneficial rhizobacteria on plant growth is less clear. Therefore, a very important point is the study of the chemical, biological, and functional activities of A. brasilense LPS, which was the aim of this work. Methods. A. brasilense LPSs were isolated from dry bacterial mass by the phenol-water method. The carbohydrates were analyzed by the Dubois method, nucleic acids — by Spirin, protein content — by Lowry and 2-keto-3-deoxyoctonic acid (KDO) — by Osborn. Pyrogenicity of LPS was tested observing the rules of bioethics in rabbits. Serological studies were performed by the Ouchterlony method. The identification of monosaccharides and fatty acids in LPS preparations was carried out on an Agilent 6890N/5973 inert chromatomass spectrometry system. Polyacrylamide gel electrophoresis in the presence of sodium dodecyl sulfate (SDS-PAAG electrophoresis) was performed according to Laemmli. Results. LPS of 3 strains of A. brasilense were isolated from dry bacterial mass and purifi ed from nucleic acids by ultracentrifugation. The purified LPSs were characterized by different relative yields from 2.44% to 4.75%, which is slightly higher than other strains of the A. brasilense (1—3%). The studied preparations were characterized by a rather high content of carbohydrates from 50.1% to 72.1%. All LPS contained up to 0.17% KDO, which is a specific component of the LPS of gram-negative bacteria. Analysis of the monosaccharide composition indicates that the LPSs of the studied A. brasilense strains turned out to be heterogeneous. At the same time, such monosaccharides as mannose, galactose, glucose, and heptose were recorded in the LPS of all tested strains. The study of the fatty acid composition of LPS showed the presence of fatty acids containing from 14 to 18 carbon atoms. Нydroxylated, saturated, and monounsaturated acids and their cis isomers were found. In the investigated LPS, the dominant fatty acids were 16:0, 18:1, 14:0(3-OH), and 16:0(3-OH), which coincides with the literature data. The research of the pyrogenic effect of LPS of A. brasilense studied strains showed that LPS solutions are apyrogenic. The double immunodiffusion reaction in Ouchterlon agar showed that all tested LPS in homologous systems exhibited ancultitigenic activity. Serological cross-reactions can be used as an approach in classifying different bacteria. Thus, we found that antisera to A. brasilense 18-2 and 61 react with all LPSs of the studied strains, which may indicate the presence of common antigenic determinants in them and that these strains belong to the same serogroup. The electrophoretic distribution data indicate that A. brasilense produces S-forms of LPS which differ in the length of O-specifi c polysaccharide chains. Conclusions. For the first time, LPS were isolated from cells of A. brasilense 10/1, 18-2 and 61. A characteristic feature of these LPS is their heterogeneity in monosaccharide and fatty acid composition, all of them were apyrogenic. The results obtained during biological-functional studies of three strains of A. brasilense LPS contribute to the biological characteristics of this species.

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confidence: 90%

Characterization of Azospirillum brasilense Lipopolysaccharides

Bulyhina¹,

Varbanest²

2022

Mikrobiol. Z.

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“…Recently, nonadecanoic acid (C19:0), octadecanoic acid (C18:0), hexadecanoic (C16:0) acid were identified among the main components of the hydrophobic part of the lipopolysaccharide of the associative diazotrophic rhizobacterium Azospirillum brasilense CD. 17) Therefore it can not be ruled out that nonadecanoic acid (C19:0) (and related fatty acids such as C19:1) should occur naturally in traces amounts and coimmigrated with polysaccharides during the ethanolic extraction used in this study.…”

Section: Resultsmentioning

confidence: 96%

Possible Involvement of Long Chain Fatty Acids in the Spores of <i>Ganoderma lucidum</i> (Reishi Houshi) to Its Anti-tumor Activity

Fukuzawa

Yamaguchi

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et al. 2008

Biological & Pharmaceutical Bulletin

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Ganoderma lucidum (G. lucidum Reishi), a traditional Chinese medical fungus, has been used as a health-promoting supplement due to its anti-tumor and immuno-modulating effects. The major bioactive components in G. lucidum are reported to be polysaccharide [1][2][3] and gonoderic acid. 4)Traditionally, the fruiting body has been the only part of G. lucidum that is used for medical purposes. Recently, Sliva et al. have reported the extract of G. lucidum using boiling water suppresses the mobility of breast and prostate cancers by inhibiting nuclear factor (NF)-kB. 5,6) As G. lucidum is available as a dietary supplement in various forms (mushroom powder, purified whole spores, broken spores, or mixtures), they compared their biological activity on cell migration and inhibition of NF-kB in human cancer cells and concluded the spores also show strong activity against human cancer cell.6) The possible role of mushrooms to prevent cancer has been summarized in several reviews. 7,8) Cancer inhibition by mushrooms or mushrooms polysaccharides was generally achieved through activating the immune response of the host rather than by directly inhibiting tumor cell growth. However, in medical mushroom research, polysaccharide and polysaccharide-peptides are known active anticancer compounds isolated from medicinal mushrooms such as maitake (Grifola frondosa), shiitake (Lentinula edodes), turkey tail (Coriolus versicolor) and reishi (G. lucidum). 9,10)The polysaccharides and polysaccharide-peptides are generally thought to exert their anti-tumor effects via a systemic effect on immunity. There is an evidence that the glucanprotein complex induces biological responses by binding to membrane compartment receptor three (CR3, or CD11b/CD18) on immune effectors cells such as macrophages.11) Furthermore, recent research has demonstrated that some of these mushroom-derived materials can produce direct cytotoxic effects on cancer cells. 12,13) A purified fraction by HPLC from the spores of G. lucidum guided by the inhibitory activity on HL-60 cell proliferation showed biological activity contains a mixture of long chain fatty acids with molecular weights in the range of m/z 250.0 to m/z 296.4, although as yet these could not be unambiguously authenticated. Therefore, in this study, using several authentic fatty acids predicted from our analytical data, we have compared the effect of the spore extract of G. lucidum with those of several authentic fatty acids on the inhibitory activity of HL-60 cell proliferation and the production of tumor necrosis factor (TNF) production induced by lipopolysaccharide (LPS) in a mouse macrophage preparation. MATERIALS AND METHODSThe spores (100 g) of G. lucidum, cultured at the area of Dabieshan, Anhui, China from October 2003 to July 2004 (Lot AFA), kindly supplied from Isukura Co., Ltd. (Tokyo) were dissolved in five volumes of 95% ethanol in a sealed bag, shaken gently at room temperature for 24 h and centrifuged at 10000ϫg for 20 min. The solution was evaporated to dryness on a rotary evaporator. T...

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“…For example, genes pRhico046 to pRhico050 are involved in the biosynthesis of dTDP-rhamnose. In A. brasilense Cd, rhamnose, as well as galactose and mannose, have been found to be components of the LPS (Konnova et al, 2006). Fedonenko et al (2002) found an O-specific pentasaccharide composed of repeating units of D-rhamnose in A. brasilense Sp245.…”

Section: Discussionmentioning

confidence: 99%

The wzm gene located on the pRhico plasmid of Azospirillum brasilense Sp7 is involved in lipopolysaccharide synthesis

2009

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Several genes involved in the interaction between Azospirillum brasilense Sp7 and plants are located on the pRhico plasmid. Here we report the characterization of an Sp7 mutant strain with impairment of the pRhico-located gene wzm. This gene encodes an inner-membrane component of an ATP-binding cassette (ABC) transporter with similarity to transporters involved in surface polysaccharide export. Indeed, SDS-PAGE revealed that LPS synthesis is affected in the wzm mutant. No significant differences were observed between wild-type and mutant strains in exopolysaccharide (EPS) amount; however, several differences were observed between them in EPS monosaccharide composition, and only wild-type colonies stained positively with Congo red. Microscopy revealed that wzm mutant cells are longer and thinner, and exhibit several differences in their cell surface relative to the wild-type. The wzm mutant was more resistant to oxidative stress, starvation, desiccation, heat and osmotic shock than the wild-type. In contrast, the mutant was more susceptible than the wild-type to UV radiation and saline stress. The strains also differed in their susceptibility to different antibiotics. Differences between the strains were also observed in their outer-membrane protein composition. No differences were observed between strains in their ability to attach to sweet corn roots and seeds, and to promote growth under the tested conditions. As LPS plays an important role in cell envelope structural integrity, we propose that the pleiotropic phenotypic changes observed in the wzm mutant are due to its altered LPS relative to the wild-type. INTRODUCTIONThe Azospirillum genus belongs to the alpha-proteobacteria and comprises free-living, nitrogen-fixing, vibrio-or spirillum-shaped rods that exert beneficial effects on plant growth and yield of many crops of agronomic importance (Dobbelaere et al., 2001). Plant growth promotion by Azospirillum is attributed to morphological and physiological changes in inoculated plant roots, which enhance water and mineral uptake. Plant growth substances, such as auxins, cytokinins and gibberellins (Steenhoudt & Vanderleyden, 2000;Dobbelaere et al., 2001), as well as nitric oxide (Creus et al., 2005) produced by the bacteria are at least partially responsible for the plant growth promotion.Within the Azospirillum genus, one of the most studied species is Azospirillum brasilense (Tarrand et al., 1978). A. brasilense cells are surrounded by a dense, tightly cellbound layer of capsular polysaccharides (CPSs), and by outer exopolysaccharides (EPSs), which are loosely bound to the cell and easily detached by centrifugation (Burdman et al., 2000c). EPSs and CPSs, as well as extracellular proteins, have been shown to play important roles in bacterial aggregation and in the establishment of the bacterium-plant association (Burdman et al., 1999(Burdman et al., , 2000cSteenhoudt & Vanderleyden, 2000). It has been suggested that EPSs and CPSs are involved in anchoring the bacteria to the surface of wheat roots , and in t...

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Chemical composition and immunochemical characteristics of the lipopolysaccharide of nitrogen-fixing rhizobacterium Azospirillum brasilense CD

Cited by 7 publications

References 12 publications

Characterization of Azospirillum brasilense Lipopolysaccharides

Characterization of Azospirillum brasilense Lipopolysaccharides

Possible Involvement of Long Chain Fatty Acids in the Spores of <i>Ganoderma lucidum</i> (Reishi Houshi) to Its Anti-tumor Activity

The wzm gene located on the pRhico plasmid of Azospirillum brasilense Sp7 is involved in lipopolysaccharide synthesis

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