Rhizobacteria Inoculation Effects on Phytohormone Status of Potato Microclones Cultivated In Vitro under Osmotic Stress.

Water deficits inhibit plant growth and decrease crop productivity. Remedies are needed to counter this increasingly urgent problem in practical farming. One possible approach is to utilize rhizobacteria known to increase plant resistance to abiotic and other stresses. We therefore studied the effects of inoculating the culture medium of potato microplants grown in vitro with Azospirillum brasilense Sp245 or Ochrobactrum cytisi IPA7.2. Growth and hormone content of the plants were evaluated under stress-free conditions and under a water deficit imposed with polyethylene glycol (PEG 6000). Inoculation with either bacterium promoted the growth in terms of leaf mass accumulation. The effects were associated with increased concentrations of auxin and cytokinin hormones in the leaves and stems and with suppression of an increase in the leaf abscisic acid that PEG treatment otherwise promoted in the potato microplants. O. cytisi IPA7.2 had a greater growth-stimulating effect than A. brasilense Sp245 on stressed plants, while A. brasilense Sp245 was more effective in unstressed plants. The effects were likely to be the result of changes to the plant's hormonal balance brought about by the bacteria.

Structural investigation and comparative cytotoxic activity of water-soluble polysaccharides from fruit bodies of the medicinal fungus quinine conk.

The structures and cytotoxic activities of water-soluble polysaccharides were investigated to search for biologically active polysaccharides from the fruit bodies of quinine conks (Fomitopsis officinalis). The decoctions of this medical fungus are actively used in folk medicine in many countries and traditional Chinese medicine. From the fungal extract we prepared, only branched ß-glucan had cytotoxic activity among all the water-soluble polysaccharides. This glucan is characterized by a regular structure. Its backbone is formed by 1,3-linked ß-D-Glcp residues, of which every third residue is substituted at O-6 by a single ß-D-Glcp residue. It has a triple helix conformation according to the data obtained from a colorimetric assay with Congo red dye and is characterized by a high-weight average molar mass (Mw > 800 kDa). ß-Glucan possessed cytotoxic activity against HeLa cells (IC50 = 318 ±â€¯47 µg/mL) and induced the formation of apoptotic bodies around most cancer cells at a concentration of 200 µg/mL. It should be noted that extraction with boiling water, which is usually used to obtain extracts and decoctions, is unable to isolate active ß-glucan. Active ß-glucan can be obtained in an individual state by cold alkali extraction after dehydration of the fruit bodies and removal of the components extractable by boiling water.

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.

Functioning of plant-bacterial associations under osmotic stress in vitro.

The search for effective plant-growth-promoting strains of rhizospheric bacteria that would ensure the resistance of plant-microbial associations to environmental stressors is essential for the design of environmentally friendly agrobiotechnologies. We investigated the interaction of potato (cv. Nevsky) microplants with the plant-growth-promoting bacteria Azospirillum brasilense Sp245 and Ochrobactrum cytisi IPA7.2 under osmotic stress in vitro. The bacteria improved the physiological and biochemical variables of the microplants, significantly increasing shoot length and root number (1.3-fold, on average). Inoculation also led a more effective recovery of the plants after stress. During repair, inoculation contributed to a decreased leaf content of malonic dialdehyde. With A. brasilense Sp245, the decrease was 1.75-fold; with O. cytisi IPA7.2, it was 1.4-fold. During repair, the shoot length, node number, and root number of the inoculated plants were greater than the control values by an average of 1.3-fold with A. brasilense Sp245 and by an average of 1.6-fold with O. cytisi IPA7.2. O. cytisi IPA7.2, previously isolated from the potato rhizosphere, protected the physiological and biochemical processes in the plants under stress and repair better than did A. brasilense Sp245. Specifically, root weight increased fivefold during repair, as compared to the noninoculated plants, while chlorophyll a content remained at the level found in the nonstressed controls. The results indicate that these bacteria can be used as components of biofertilizers. A. brasilense Sp245 has favorable prospects for use in temperate latitudes, whereas O. cytisi IPA7.2 can be successfully used in saline and drought-stressed environments.

Ochrobactrum cytisi IPA7.2 promotes growth of potato microplants and is resistant to abiotic stress.

Bacteria in natural associations with agricultural crops are promising for use in the improvement of clonal micropropagation of plants. We clarified the taxonomic position of Ochrobactrum cytisi strain IPA7.2 and investigated its tolerance for salinity, high temperature, and glyphosate pollution. We also tested the strain's potential to promote the growth of potato (Solanum tuberosum L.) microplants. Using the IPA7.2 draft genome (no. NZ_MOEC00000000), we searched for housekeeping genes and also for the target genes encoding glyphosate tolerance and plant-growth-promoting ability. A multilocus sequence analysis of the gap, rpoB, dnaK, trpE, aroC, and recA housekeeping genes led us to identify isolate IPA7.2 as O. cytisi. The strain tolerated temperatures up to 50 °C and NaCl concentrations up to 3-4%, and it produced 8 µg ml-1 of indole-3-acetic acid. It also tolerated 6 mM glyphosate owing to the presence of type II 5-enolpyruvylshikimate-3-phosphate synthase. Finally, it was able to colonize the roots and tissues of potato microplants, an ability preserved by several generations after subculturing. We identified the development phase of potato microplants that was optimal for inoculation with O. cytisi IPA7.2. Inoculation of in vitro-grown 15-day-old microplants increased the mitotic index of root meristem cells (by 50%), the length of shoots (by 34%), the number of leaves (by 7%), and the number of roots (by 16%). Under ex vitro conditions, the inoculated plants had a greater leaf area (by 77%) and greater shoot and root dry weight (by 84 and 61%, respectively) than did the control plants. We recommend O. cytisi IPA 7.2 for use in the growing of potato microplants to improve the production of elite seed material.

Isolation and characterization of a glyphosate-degrading rhizosphere strain, Enterobacter cloacae K7.

Plant-growth-promoting rhizobacteria exert beneficial effects on plants through their capacity for nitrogen fixation, phytohormone production, phosphate solubilization, and improvement of the water and mineral status of plants. We suggested that these bacteria may also have the potential to express degradative activity toward glyphosate, a commonly used organophosphorus herbicide. In this study, 10 strains resistant to a 10 mM concentration of glyphosate were isolated from the rhizoplane of various plants. Five of these strains--Alcaligenes sp. K1, Comamonas sp. K4, Azomonas sp. K5, Pseudomonas sp. K3, and Enterobacter cloacae K7--possessed a number of associative traits, including fixation of atmospheric nitrogen, solubilization of phosphates, and synthesis of the phytohormone indole-3-acetic acid. One strain, E. cloacae K7, could utilize glyphosate as a source of P. Gas-liquid chromatography showed that E. cloacae growth correlated with a decline in herbicide content in the culture medium (40% of the initial 5mM content), with no glyphosate accumulating inside the cells. Thin-layer chromatography analysis of the intermediate metabolites of glyphosate degradation found that E. cloacae K7 had a C-P lyase activity and degraded glyphosate to give sarcosine, which was then oxidized to glycine. In addition, strain K7 colonized the roots of common sunflower (Helianthus annuus L.) and sugar sorghum (Sorghum saccharatum Pers.), promoting the growth and development of sunflower seedlings. Our findings extend current knowledge of glyphosate-degrading rhizosphere bacteria and may be useful for developing a biotechnology for the cleanup and restoration of glyphosate-polluted soils.