Studies on plant-microbe interaction to improve stress tolerance in plants for sustainable agriculture

Biotic and abiotic stress factors have a major impact on plants and cause extensive losses to crop production. Bacteria that provide growth promotion and prime stress tolerance of plants have great potential to improve crop production and support durable and environmental friendly resource management. Priming refers here to when plants upon appropriate stimulation develop an enhanced capacity to express defense responses to a later stimulus. In this study strains of the beneficial bacterium Bacillus amyloliquefaciens were analysed for their effects on plants. Direct antagonistic effect of B. amyloliquefaciens on several Brassica phytopathogens (Botrytis cinerea, Alternaria brassicae, Alternaria brassicicola, Verticillium longisporum, and Sclerotinia sclerotiorum) was demonstrated by bacteria and exudates in vitro. A bacterial exudate fraction containing lipopeptide antibiotics was analysed and the strongest antagonistic activity was connected with a novel linear form of fengycin identified using mass spectrometry. Growth promotion of Arabidopsis thaliana Col-0, coi1-16, jar1 and npr1 but not in myb72 plants by B. amyloliquefaciens UCMB5113 was demonstrated with increased shoot and root biomass and increased number of lateral roots and root hairs while the primary root growth decreased. Bacillus inoculation resulted in profound effects on various plant hormones that will affect a variety of plant functions. Growth promotion was also demonstrated in split dish experiments where Bacillus strains were sequestered from Arabidopsis plants indicating a role for volatile organic compounds (VOCs). Bacillus VOCs also caused growth suppression of several phytopathogens. GC-MS analysis identified a large number of compounds in the VOC blend and the composition of VOCs was dependent on the medium used for cultivation and the effects on the plant also varied. Thus these Bacillus strains promote growth of plants and improve the survivability of plants exposed to biotic stress challenges by priming of stress tolerance. These findings can be extended to different crops to improve crop productivity under various environmental conditions

Analysis of plant growth-promoting properties of Bacillus amyloliquefaciens UCMB5113 using Arabidopsis thaliana as host plant

This study showed that Bacillus amyloliquefaciens UCMB5113 colonizing Arabidopsis roots changed root structure and promoted growth implying the usability of this strain as a novel tool to support sustainable crop production.Root architecture plays a crucial role for plants to ensure uptake of water, minerals and nutrients and to provide anchorage in the soil. The root is a dynamic structure with plastic growth and branching depending on the continuous integration of internal and environmental factors. The rhizosphere contains a complex microbiota, where some microbes can colonize plant roots and support growth and stress tolerance. Here, we report that the rhizobacterium Bacillus amyloliquefaciens subsp. plantarum UCMB5113 stimulated the growth of Arabidopsis thaliana Col-0 by increased lateral root outgrowth and elongation and root-hair formation, although primary root elongation was inhibited. In addition, the growth of the above ground tissues was stimulated by UCMB5113. Specific hormone reporter gene lines were tested which suggested a role for at least auxin and cytokinin signaling during rhizobacterial modulation of Arabidopsis root architecture. UCMB5113 produced cytokinins and indole-3-acetic acid, and the formation of the latter was stimulated by root exudates and tryptophan. The plant growth promotion effect by UCMB5113 did not appear to depend on jasmonic acid in contrast to the disease suppression effect in plants. UCMB5113 exudates inhibited primary root growth, while a semi-purified lipopeptide fraction did not and resulted in the overall growth promotion indicating an interplay of many different bacterial compounds that affect the root growth of the host plant. This study illustrates that beneficial microbes interact with plants in root development via classic and novel signals

Functional classification of protein-coding genes in UCMB5113.

<p>Distibution of UCMB5113 coding sequences (93.6%) in COG functional classes. Genes that did not have any inferred COG annotation were assigned to category X.</p

NRPS and PKS gene clusters involved in synthesis of secondary metabolites in <i>B. amyloliquefaciens</i>.

<p>NRPS and PKS gene clusters involved in synthesis of secondary metabolites in <i>B. amyloliquefaciens</i>.</p

Genome Analysis of <i>Bacillus amyloliquefaciens</i> Subsp. <i>plantarum</i> UCMB5113: A Rhizobacterium That Improves Plant Growth and Stress Management

<div><p>The <i>Bacillus amyloliquefaciens</i> subsp. <i>plantarum</i> strain UCMB5113 is a Gram-positive rhizobacterium that can colonize plant roots and stimulate plant growth and defense based on unknown mechanisms. This reinforcement of plants may provide protection to various forms of biotic and abiotic stress. To determine the genetic traits involved in the mechanism of plant-bacteria association, the genome sequence of UCMB5113 was obtained by assembling paired-end Illumina reads. The assembled chromosome of 3,889,532 bp was predicted to encode 3,656 proteins. Genes that potentially contribute to plant growth promotion such as indole-3-acetic acid (IAA) biosynthesis, acetoin synthesis and siderophore production were identified. Moreover, annotation identified putative genes responsible for non-ribosomal synthesis of secondary metabolites and genes supporting environment fitness of UCMB5113 including drug and metal resistance. A large number of genes encoding a diverse set of secretory proteins, enzymes of primary and secondary metabolism and carbohydrate active enzymes were found which reflect a high capacity to degrade various rhizosphere macromolecules. Additionally, many predicted membrane transporters provides the bacterium with efficient uptake capabilities of several nutrients. Although, UCMB5113 has the possibility to produce antibiotics and biosurfactants, the protective effect of plants to pathogens seems to be indirect and due to priming of plant induced systemic resistance. The availability of the genome enables identification of genes and their function underpinning beneficial interactions of UCMB5113 with plants.</p></div

Blast comparison of NRPS/PKS clusters in UCMB5113 (above) and FZB42^T(below).

<p>Arrows indicate gene clusters; Macrolactin (light green), Bacilllaene (purple), BacillomycinD (orange), Difficidin (blue), Bacillibactin (turquoise), Surfactin (green), Bacilysin (grey), Fengycin (pink). Genes highlighted in red represent the differences where as black represent other genes flanking in each cluster.</p

<i>B. amyloliquefaciens</i> UCMB5113 related activity on different substrates.

<p>A branch pattern with massive groups of bacteria observed after 4 days of incubation period indicated swarming motility; bright pink color indicated the hydrolysis of urea to carbon dioxide and ammonia; siderophore biosynthetic cluster produced a clear zone on CAS agar; chitin degradation and utilization as carbon source; expressed hemolytic activity on blood agar; phosphate solubilization around bacteria apparent as a transparent zone; amylase activity on starch medium; aqueous drop collapse to assess production of biosurfactants.</p

Graphical representation of genomic features of <i>B. amyloliquefaciens</i> subsp. <i>plantarum</i> UCMB5113.

<p>Circles display (from the outside): (1) Sites of genome plasticity. (2) Predicted CDSs transcribed in the clockwise direction. (3) Predicted CDSs transcribed in the counterclockwise direction. (4) rRNA (blue), tRNA (orange), non-coding RNA (green), and NRPS/PKS gene clusters (grey). (5,6,7) Blast comparison of UCMB5113 genome with type strain FZB42^T and <i>B. subtilis</i> 168, respectively. (8) GC percent deviation (GC window - mean GC) in a 1000-bp window. (9) GC skew (G+C/G-C) in a 1000-bp window.</p

Plant growth promotion by UCMB5113 on <i>Arabidopsis thaliana</i> Col-0.

<p>(A). Plants grown on MS agar and treated with UCMB5113 display bigger leaves and increased root branching. (B) Plants grown on soil and treated with UCMB5113 have bigger leaves compared to control plants. The experiment was performed at least three times, and similar results were obtained in each case.</p