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Callus tissue is a popular tool in modern plant breeding and biotechnology. Macromolecules of plant-growth-promoting rhizobacteria can be beneficial for callus morphogenesis. We compared the effects of the lipopolysaccharides (LPSs) of three Azospirillum strains (A. brasilense SR55, A. brasilense SR75, and A. lipoferum SR65) on the calluses of two wheat (Triticum aestivum L. cv. Saratovskaya 29) lines (LRht-B1c and LRht-B1a) that differ in their morphogenic activity. The LPSs differed in the chemical structure of their O polysaccharides and in their physicochemical and serological properties. The LPS of A. lipoferum SR65 significantly promoted callus morphogenesis and regenerant development in both wheat lines. The yield of regenerated plants in terms of the total number of explants was significantly increased—2.15-fold in the highly morphogenic line LRht-B1c and 3.75-fold in the weakly morphogenic line LRht-B1a. In both lines, the LPSs of A. brasilense SR55 and SR75 increased either only the yield of morphogenic calluses or only the yield of regenerated plants, respectively. Overall, the Azospirillum LPSs affected the weakly morphogenic line LRht-B1a stronger than they did the highly morphogenic line LRht-B1c, and this resulted in a leveling of differences between the activities of the LRht-B1c and LRht-B1a morphogenic calluses. The LPSs of some Azospirillum strains are promising promoters of plant morphogenesis and may, in the future, find frequent use in plant breeding and genetic engineering experiments with callus tissue. Lipopolysaccharides isolated from the outer membranes of various strains of plant-growth-promoting rhizobacteria of the genus Azospirillum increase the morphogenic activity of soft wheat calluses with different efficiency.

In vitro somatic callus culturing is used widely in plant biotechnology, but its effectiveness depends largely on the donor plant genotype. Bacteria or components of their cells are rarely used to activate morphogenesis. In this work, inoculation of explants from immature wheat (Triticum aestivum L.) embryos with a suspension of living cells of the bacterium Azospirillum brasilense Sp7 resulted in callus death after 7 days of growth, in contrast to explant treatment with a suspension of heat-killed whole cells of Sp7. The experiments used two wheat lines, LRht-B1a and LRht-B1c, which differ in morphogenic activity. Growing calluses with the lipopolysaccharide of A. brasilense Sp7 increased the yield of regenerated plants 2- to 3.5-fold in both lines. This increase was through the activation of regenerant formation from morphogenic calluses. We have demonstrated for the first time the effects of bacterial flagellin on plant tissue culture. The polar-flagellum flagellin of A. brasilense Sp7 leveled the genotypic differences in the morphogenic ability of callus tissue. Specifically, it increased the yield of morphogenic calluses in the weakly morphogenic line LRht-B1a to the yield value in the highly morphogenic line LRht-B1c but lowered the yield of regenerants in the highly morphogenic line LRht-B1c to the yield value in the weakly morphogenic line LRht-B1a. Thus, bacterial lipopolysaccharides and flagellins can be used to regulate the formation of morphogenic calluses and regenerants in plant tissue culturing in vitro.

We evaluated the effect of lipopolysaccharides from the plant-growth-promoting associative bacterium Azospirillum brasilense Sp245 and from the enteric bacterium Escherichia coli K12 on the morphogenic potential of in vitro-growing somatic calluses of soft spring wheat (Triticum aestivum L. cv. Saratovskaya 29). A genetic model was used that included two near-isogenic lines of T. aestivum L. cv. Saratovskaya 29 with different embryogenic capacities; one of these lines carries the Rht-B1 dwarfing gene, whereas the other lacks it. When added to the nutrient medium, the lipopolysaccharide of A. brasilense Sp245 promoted the formation of calluses with meristematic centers and stimulated the regeneration ability of the cultured tissues in both lines. By contrast, the lipopolysaccharide of the enteric bacterium E. coli K12 barely affected the morphogenetic activity of callus cells and the yield of morphogenic calluses and regenerated plants. These findings indicate that the lipopolysaccharide of the plant-growth-promoting associative bacterium A. brasilense Sp245 specifically enhances the morphogenetic activity of wheat somatic tissues, which increases the efficacy of culturing of genotypes with a relatively low morphogenic potential. The results of the study may contribute to the improvement of the efficacy of plant cell selection and gene engineering and to a better understanding of the mechanisms responsible for plant recognition of lipopolysaccharides of associative bacteria.

International audience; AbstractMicroclonal propagation in vitro is being actively used in the production of healthy planting material of food and ornamental plants. However, it needs further improvement to increase the growth rates of microclones in vitro and enhance regenerant survivability ex vitro. A promising approach to this end could be inoculating in vitro-micropropagated plants with plant growth-promoting rhizobacteria, specifically Azospirillum. However, the influence of Azospirillum inoculation on microclone behavior throughout the production process, including plant adaptation ex vitro and food crop productivity, has been underinvestigated. In this study, in vitro-growing potato (Solanum tuberosum L.) microclones were inoculated with Azospirillum brasilense strain Sp245. The microclones were then grown on in soil in the greenhouse and field, with the experiment lasting for 120 days. Root-associated bacteria were identified immunochemically, and the mitotic index of root meristematic cells was determined by a cytological method. The plant morphological parameters determined were shoot length, number of nodes per shoot, number of roots per plant, maximal root length, leaf area, percentage of surviving plants in the soil, and tuber yield and weight. Our results show that bacterial inoculation of potato microclones in vitro enhances plant adaptive capacity ex vitro and increases minituber yield. The percent survival index of field-grown inoculated plants was 1.5-fold greater than that of uninoculated plants. The overall tuber weight per plant was more than 30 % greater in the inoculated plants than it was in the control ones. For all cultivars on average, tuber yield per square meter increased by more than 45 % as a result of inoculation in vitro. This study is the first to report that Azospirillum inoculation of potato microclones not only improves the quality of planting material produced in vitro but also significantly increases minituber yield through enhancing plant adaptive capacity in the field.