Beneficial soil bacteria confer immunity against a wide range of foliar diseases by activating plant defenses, thereby reducing a plant's susceptibility to pathogen attack. Although bacterial signals ...have been identified that activate these plant defenses, plant metabolites that elicit rhizobacterial responses have not been demonstrated. Here, we provide biochemical evidence that the tricarboxylic acid cycle intermediate L-malic acid (MA) secreted from roots of Arabidopsis (Arabidopsis thaliana) selectively signals and recruits the beneficial rhizobacterium Bacillus subtilis FB17 in a dose-dependent manner. Root secretions of L-MA are induced by the foliar pathogen Pseudomonas syringae pv tomato (Pst DC3000) and elevated levels of L-MA promote binding and biofilm formation of FB17 on Arabidopsis roots. The demonstration that roots selectively secrete L-MA and effectively signal beneficial rhizobacteria establishes a regulatory role of root metabolites in recruitment of beneficial microbes, as well as underscores the breadth and sophistication of plant-microbial interactions.
Plant growth-promoting rhizobacteria, in association with plant roots, can trigger induced systemic resistance (ISR). Considering that low-molecular weight volatile hormone analogues such as methyl ...jasmonate and methyl salicylate can trigger defense responses in plants, we examined whether volatile organic compounds (VOCs) associated with rhizobacteria can initiate ISR. In Arabidopsis seedlings exposed to bacterial volatile blends from Bacillus subtilis GB03 and Bacillus amyloliquefaciens IN937a, disease severity by the bacterial pathogen Erwinia carotovora subsp. carotovora was significantly reduced compared with seedlings not exposed to bacterial volatiles before pathogen inoculation. Exposure to VOCs from rhizobacteria for as little as 4 d was sufficient to activate ISR in Arabidopsis seedlings. Chemical analysis of the bacterial volatile emissions revealed the release of a series of low-molecular weight hydrocarbons including the growth promoting VOC (2R,3R)-(-)-butanediol. Exogenous application of racemic mixture of (RR) and (SS) isomers of 2,3-butanediol was found to trigger ISR and transgenic lines of B. subtilis that emitted reduced levels of 2,3-butanediol and acetoin conferred reduced Arabidopsis protection to pathogen infection compared with seedlings exposed to VOCs from wild-type bacterial lines. Using transgenic and mutant lines of Arabidopsis, we provide evidence that the signaling pathway activated by volatiles from GB03 is dependent on ethylene, albeit independent of the salicylic acid or jasmonic acid signaling pathways. This study provides new insight into the role of bacteria VOCs as initiators of defense responses in plants.
Elevated sodium (Na+) decreases plant growth and, thereby, agricultural productivity. The ion transporter high-affinity K+ transporter (HKT)1 controls Na+ import in roots, yet dysfunction or ...overexpression of HKT1 fails to increase salt tolerance, raising questions as to HKT1's role in regulating Na+ homeostasis. Here, we report that tissue-specific regulation of HKT1 by the soil bacterium Bacillus subtilis GB03 confers salt tolerance in Arabidopsis thaliana. Under salt stress (100 mM NaCl), GB03 concurrently down- and upregulates HKT1 expression in roots and shoots, respectively, resulting in lower Na+ accumulation throughout the plant compared with controls. Consistent with HKT1 participation in GB03-induced salt tolerance, GB03 fails to rescue salt-stressed athkt1 mutants from stunted foliar growth and elevated total Na+ whereas salt-stressed Na+ export mutants sos3 show GB03-induced salt tolerance with enhanced shoot and root growth as well as reduced total Na+. These results demonstrate that tissue-specific regulation of HKT1 is critical for managing Na+ homeostasis in salt-stressed plants, as well as underscore the breadth and sophistication of plant-microbe interactions.
Haloxylon ammodendron, a typical xerophyte, tolerates various abiotic stresses and is widely distributed in desert areas. Two PGPR strains, Bacillus sp. WM13-24 and Pseudomonas sp. M30-35, were ...previously isolated from the rhizosphere of H. ammodendron in Tengger Desert, Gansu province, northwest China. The aim of this study was to investigate the role of M30-35 and WM13-24 in drought stress alleviation of ryegrass (Lolium perenne L.). Under normal condition, both M30-35 and WM13-24 increased shoot fresh and dry weight, chlorophyll content, total nitrogen and phosphorus contents and altered phytohormone distribution compared to control. Moreover, after 7 days of drought stress, WM13–24 and M30-35 enhanced photosynthetic capacity, relative water content, the activities of catalase (CAT) and peroxidase (POD) and proline content, resulted in decreased malondialdehyde (MDA) content, relative membrane permeability (RMP) and H2O2 accumulation; interestingly, the two strains decreased ABA content in leaves. This study demonstrated that the two PGPR strains promoted ryegrass growth and root development via regulating plant hormone distribution and enhanced drought tolerance of ryegrass through improving the activities of antioxidant enzymes, regulating ABA signaling and maintaining plant growth. Our results indicated that PGPR strains from rhizosphere of the desert plant species could be considered as promising bioinoculants for grass plants.
•Both strains Bacillus sp. WM13-24 and Pseudomonas sp. M30-35 had plant growth promoting traits.•Both strains improved root system architecture and promoted the growth of ryegrass seedlings.•Both strains enhanced drought tolerance of ryegrass via enhancing antioxidant enzyme activities and regulating ABA signal.
Photosynthesis is regulated by environmental factors as well as endogenous sugar signals. Whereas light-driven sugar biosynthesis is essential for terrestrial organisms, as well as belowground ...microflora, whether and how soil symbionts regulate photosynthesis has yet to be reported. Here, we show that the plant growth-promoting soil bacterium Bacillus subtilis GB03 augments photosynthetic capacity by increasing photosynthetic efficiency and chlorophyll content in Arabidopsis. Mechanistic studies reveal an elevation of sugar accumulation as well as the suppression of classic glucose signaling responses, including hypocotyl elongation and seed germination, with exposure to GB03. Compared with wild-type plants, two Arabidopsis mutants defective in hexokinase-dependent sugar signaling exhibit increased photosynthetic capacity, which is not further enhanced with GB03 exposure. Overlap in sugar/ABA sensing is observed in GB03-exposed plants, with a reduction of ABA-biosynthetic transcripts as well as downstream metabolite levels in leaves. Moreover, exogenous ABA abrogates GB03-triggered increases in photosynthetic efficiency and chlorophyll content. These results demonstrate that certain rhizobacteria elevate photosynthesis through the modulation of endogenous sugar/ABA signaling, and establish a regulatory role for soil symbionts in plant acquisition of energy.
Background and Aims Soil salinization limits conventional agriculture since most food-based plant cultivars require low soil-sodium (Na⁺) levels for robust growth. Moreover, modern agricultural ...practices, especially in arid environments, can exacerbate soil salinization as belowground water sources utilized in irrigation are frequently tainted with salt. While salt tolerance has previously been shown to be augmented in several glycophyte species by the soil bacterium Bacillus subtilis (GB03), here we reported that this beneficial rhizobacterium promotes growth and augments higher salt-tolerance in halophyte grass Puccinellia tenuiflora. Methods The optimal Bacillus subtilis strain for P. tenuiflora was screened. P. tenuiflora was grown from seeds with NaCl (0, 100, 200 and 300 mM) for salt treatments with or without inoculation of B. subtilis GB03. Growth parameters, chlorophyll content and endogenous Na⁺ and K⁺ contents were determined at the time of harvest. Seedlings were grown in medium with 0 or 200 mM NaCl, then were harvested to extract total RNA after 48 h of exposure to GB03 VOCs. Semiquantitative RT-PCR was used to investigate the relative amount of PtHKT1; 5, PtHKT2; 1 and PtSOS1 in P. tenuiflora regulated by GB03. Results The optimal Bacillus subtilis strain for P. tenuiflora was GB03. GB03 significantly improved shoot and root growth at two, three, four and five weeks after inoculation. Under various salinity stresses, GB03 significantly promoted growth of Pʒ tenuiflora seedlings. Na⁺ accumulation was reduced with K⁺ accumulation unaffected by GB03 exposure. Therefore, GB03 enhanced selective absorption capacity of P. tenuiflora for K⁺ over Na⁺ (SA) from media. Gene expression analysis demonstrated that GB03 up-regulated PtHKT1; 5 and PtSOS1, but down-regulated PtHKT2;1 expression, specifically in roots when plants are grown under greatly-elevated salt conditions (200 mM NaCl). Conclusions Our results presented here established that B. subtilis GB03 promoted the growth and improved the salt tolerance and the selective absorption capacity for K⁺ over Na⁺ in the monocotyledonous halophyte P. tenuiflora to a higher level. Interestingly, GB03-triggered up-regulation of PtHKT1; 5 and PtSOS1 and down-regulation of PtHKT2; 1 in roots reduced Na⁺ transport from root to shoot as well as Na⁺ uptake in roots. This study provides the physiological and molecular evidence that application of selected bacteria to salttolerant Monocots can ameliorate deleterious effects of high soil saline toxicity.
Certain plant growth-promoting rhizobacteria (PGPR), in the absence of physical contact with a plant stimulate growth via volatile organic compound (VOC) emissions, through largely unknown ...mechanisms. To probe how PGPR VOCs trigger growth in plants, RNA transcript levels of Arabidopsis seedlings exposed to Bacillus subtilus (strain GB03) were examined using oligonucleotide microarrays. In screening over 26,000 protein-coded transcripts, a group of approximately 600 differentially expressed genes related to cell wall modifications, primary and secondary metabolism, stress responses, hormone regulation and other expressed proteins were identified. Transcriptional and histochemical data indicate that VOCs from the PGPR strain GB03 trigger growth promotion in Arabidopsis by regulating auxin homeostasis. Specifically, gene expression for auxin synthesis was up regulated in aerial regions of GB03-exposed plants; auxin accumulation decreased in leaves and increased in roots with GB03 exposure as revealed in a transgenic DR5::GUS Arabidopsis line, suggesting activation of basipetal auxin transport. Application of the auxin transport inhibitor 1-naphthylphthalamic acid (NPA) restricted auxin accumulation to sites of synthesis thereby preventing GB03-mediated decreases in shoot auxin levels as well as thwarting GB03-mediated growth promotion. In addition, microarray data revealed coordinated regulation of cell wall loosening enzymes that implicated cell expansion with GB03 exposure, which was confirmed by comparative cytological measurements. The discovery that bacterial VOCs, devoid of auxin or other known plant hormones regulate auxin homeostasis and cell expansion provides a new paradigm as to how rhizobacteria promote plant growth.
Choline (Cho) is an essential nutrient for humans as well as the precursor of glycine betaine (GlyBet), an important compatible solute in eukaryotes that protects cells from osmotic stress caused by ...dehydrating conditions. The key enzyme for plant Cho synthesis is phosphoethanolamine N-methyltransferase (PEAMT), which catalyzes all three methylation steps, including the rate-limiting N-methylation of phosphoethanolamine. Herein, we report that the beneficial soil bacterium Bacillus subtilis (strain GB03) enhances Arabidopsis Cho and GlyBet synthesis associated with enhanced plant tolerance to osmotic stress. When stressed with 100 mM exogenous mannitol, GB03-exposed plants exhibit increased transcript level of PEAMT compared with stressed plants without bacterial exposure. Endogenous Cho and GlyBet metabolite pools were elevated by more than two- and fivefold, respectively, by GB03 treatment, consistent with increased stress tolerance. Moreover, in the xipotl mutant line with reduced Cho production, a loss of GB03-induced drought tolerance is observed. Osmotic-stressed plants with or without GB03 exposure show similar levels of abscsisic acid (ABA) accumulation in both shoots and roots, suggesting that GB03-induced osmoprotection is ABA independent. GB03 treatment also improves drought tolerance in soil-grown plants as characterized by phenotypic comparisons, supported by an elevated accumulation of osmoprotectants. These results provide a biological strategy to enhance Cho biosynthesis in plants and, in turn, increase plant tolerance to osmotic stress by elevating osmoprotectant accumulation.
SPME utilized for profiling of volatiles in
Bacillus subtilis (GB03) and
B. amyliquefaciens (IN937a) revealed significant differences in volatile composition among both strains, most notably in ...3-methyl-butanol levels when inoculated on MS media or potato roots.
Chemical and plant growth studies of Bacilli strains GB03 and IN937a revealed that the volatile components 2,3-butanediol and acetoin trigger plant growth promotion in Arabidopsis. Differences in growth promotion when cytokinin-signaling mutants are exposed to GB03 versus IN937a volatiles suggest a divergence in chemical signaling for these two bacterial strains. To provide a comprehensive chemical profile of bacterial volatiles emitted from these biologically active strains, headspace solid phase microextraction (SPME) coupled with software extraction of overlapping GC-separated components was employed. Ten volatile metabolites already reported from GB03 and IN937a were identified as well as 28 compounds not previously characterized. Most of the newly identified compounds were branched-chain alcohols released from IN937a, at much higher levels than in GB03. Principal component analysis clearly separated GB03 from IN937a, with GB03 producing higher amounts of 3-methyl-1-butanol, 2-methyl-1-butanol and butane-1-methoxy-3-methyl. The branched-chain alcohols share a similar functional motif to that of 2,3-butanediol and may afford alternative structural patterns for elicitors from bacterial sources.
Soil salinity is an increasingly serious problem worldwide that reduces agricultural output potential. Selected beneficial soil bacteria can promote plant growth and augment tolerance to biotic and ...abiotic stresses. Bacillus subtilis strain GB03 has been shown to confer growth promotion and abiotic stress tolerance in the model plant Arabidopsis thaliana. Here we examined the effect of this beneficial soil bacterium on salt tolerance in the legume forage crop, white clover. Plants of white clover (Trifolium repens L. cultivar Huia) were grown from seeds with or without soil inoculation of the beneficial soil bacterium Bacillus subtilis GB03 supplemented with 0, 50, 100, or 150 mM NaCl water into soil. Growth parameters, chlorophyll content, malondialdehyde (MDA) content and osmotic potential were monitored during the growth cycle. Endogenous Na(+) and K(+) contents were determined at the time of harvest. White clover plants grown in GB03-inoculated soil were significantly larger than non-inoculated controls with respect to shoot height, root length, plant biomass, leaf area and chlorophyll content; leaf MDA content under saline condition and leaf osmotic potential under severe salinity condition (150 mM NaCl) were significantly decreased. Furthermore, GB03 significantly decreased shoot and root Na(+) accumulation and thereby improved K(+)/Na(+) ratio when GB03-inoculated plants were grown under elevated salt conditions. The results indicate that soil inoculation with GB03 promotes white clover growth under both non-saline and saline conditions by directly or indirectly regulating plant chlorophyll content, leaf osmotic potential, cell membrane integrity and ion accumulation.
PDF
