植物生育促進菌類による全身抵抗性誘導に関わる植物の転写応答解析

博士(農学)岐阜大学(Gifu University

Systemic resistance induced by volatile organic compounds emitted by plant growth-promoting fungi in Arabidopsis thaliana.

Volatile organic compounds (VOC) were extracted and identified from plant growth-promoting fungi (PGPF), Phoma sp., Cladosporium sp. and Ampelomyces sp., using gas chromatography-mass spectrometry (GC-MS). Among the three VOC extracted, two VOC blends (emitted from Ampelomyces sp. and Cladosporium sp.) significantly reduced disease severity in Arabidopsis plants against Pseudomonas syringae pv. tomato DC3000 (Pst). Subsequently, m-cresol and methyl benzoate (MeBA) were identified as major active volatile compounds from Ampelomyces sp. and Cladosporium sp., respectively, and found to elicit induced systemic resistance (ISR) against the pathogen. Molecular signaling for disease suppression by the VOC were investigated by treating different mutants and transgenic Arabidopsis plants impaired in salicylic acid (SA) or Jasmonic acid (JA)/ethylene (ET) signaling pathways with m-cresol and MeBA followed by challenge inoculation with Pst. Results show that the level of protection was significantly lower when JA/ET-impaired mutants were treated with MeBA, and in SA-, and JA/ET-disrupted mutants after m-cresol treatment, indicating the involvement of these signal transduction pathways in the ISR primed by the volatiles. Analysis of defense-related genes by real-time qRT-PCR showed that both the SA-and JA-signaling pathways combine in the m-cresol signaling of ISR, whereas MeBA is mainly involved in the JA-signaling pathway with partial recruitment of SA-signals. The ET-signaling pathway was not employed in ISR by the volatiles. Therefore, this study identified two novel volatile components capable of eliciting ISR that may be promising candidates in biological control strategy to protect plants from diseases

Specific detection of Type 1 and Type 2 isolates of <i>Pyrenochaeta lycopersici</i> by loop-mediated isothermal amplification reaction

<p><i>Pyrenochaeta lycopersici</i> is the causal agent of corky root, an important soilborne disease of tomato (<i>Lycopersicon esculentum</i>) and other solanaceous crops. <i>P. lycopersici</i> isolates are classified into Types 1 and 2 on the basis of several physiological and molecular features. In this study, we aimed to establish a loop-mediated isothermal amplification (LAMP) method to identify Types 1 and 2 isolates. Using specific LAMP primer sets for Types 1 and 2, both types were easily identified within 35 min. The LAMP method demonstrated equal sensitivity to the polymerase chain reaction in molecular identification of the pathogen in cultured mycelia, infested plant roots, and their surrounding soil. The LAMP technology will contribute to the detection and identification of the pathogen, crop protection, and thorough understanding about the ecology of corky root disease of tomato.</p

Relative expression of defense-related genes on leaves of <i>A. thaliana</i> treated with <i>m</i>-cresol and MeBA.

<p>Amplification of JA−/ET-responsive genes <i>PR3</i> and <i>PDF1.2</i>, JA-responsive genes <i>AtVSP2</i> and <i>MYC2</i>, ET-responsive gene <i>PR4</i>, and SA-inducible genes <i>PR1</i>, <i>PR2</i> and <i>PR5</i> were analyzed with real-time qRT-PCR. Leaves from 15 representative plants were sampled 5 days after inoculation. Asterisks indicate statistically significant differences (Student’s <i>t</i>-test, <i>P</i> = 0.01) compared with the control treatment.</p

Expression of of defense-related genes.

<p>A. SA-responsive gene <i>PR</i>-1 and B. JA-responsive gene <i>PDF</i> 1.2 in leaves of <i>Arabidopsis thaliana</i> treated with volatile blends emitted from <i>Phoma</i> sp. and <i>Ampelomyces</i> sp. and MeBA from <i>Cladosporium</i> sp. in semi-quantitative RT-PCR analysis. Asterisks indicate statistically significant differences (Student’s <i>t</i>-test, <i>P</i> = 0.01) compared with the control. Data are from representative experiments that were repeated at least 3 times with similar results.</p

Systemic resistance induced by <i>m</i>-cresol and methyl benzoate (MeBA) at different concentrations.

<p>A. Reduction in disease severity and B. Growth of <i>Pst</i> DC3000 in leaves after pretreatment of plants with <i>m</i>-cresol and MeBA at different concentrations followed by challenge inoculation with <i>Pst</i> DC3000. Disease severity was scored for each plant as the percentage of total leaf surface with symptoms, from 0 = no symptoms to 100 = most severe, with necrotic symptoms. Different letters indicate significant differences between treatments according to Bonferroni multiple comparison test (P = 0.05). Data are from representative experiments that were repeated at least 3 times with similar results.</p

Retention index (RI) and peak areas for volatile organic compounds (VOC) extracted from 14-d-old cultures of the plant-growth-promoting fungi <i>Phoma</i> sp. (GS8-3), <i>Ampelomyces</i> sp. (F-a-3) and <i>Cladosporium</i> sp. (D-c-4) using SPME-based GC-MS analysis.

<p>Note: Compounds were identified by comparing the RI and mass spectra with data in the NIST database.</p