Mitsuro Hyakumachi

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.

Analysis of volatile organic compounds emitted by plant growth-promoting fungus Phoma sp. GS8-3 for growth promotion effects on tobacco.

We extracted volatile organic compounds (VOCs) emitted by a plant growth-promoting fungus (PGPF) Phoma sp. GS8-3 by gas chromatography and identified them by mass spectrometry. All of the identified compounds belonged to C4-C8 hydrocarbons. Volatiles varied in number and quantity by the culture period of the fungus (in days). 2-Methyl-propanol and 3-methyl-butanol formed the main components of the volatile blends for all the culture periods of fungus. Growth-promoting effects of the identified synthetic compounds were analyzed individually and in blends using tobacco plants. We found that the mixture of volatiles extracted from 3-day-old culture showed significant growth promotion in tobacco in vitro. The volatile blend showed better growth promotion at lower than higher concentrations. Our results confirm the potential role of volatile organic compounds in the mechanism of growth enhancement by GS8-3.

Induction of Systemic Resistance against Cucumber mosaic virus in Arabidopsis thaliana by Trichoderma asperellum SKT 1

Trichoderma asperellum SKT-1 is a microbial pesticide that is very effective against various diseases. Our study was undertaken to evaluate T. asperellum SKT-1 for induction of resistance against yellow strain of Cucumber mosaic virus (CMV-Y) in Arabidopsis plants. Disease severity was rated at 2 weeks post inoculation (WPI). CMV titre in Arabidopsis leaves was determined by indirect enzyme-linked immunosorbent assay (ELISA) at 2 WPI. Our results demonstrated that among all Arabidopsis plants treated with barley grain inoculum (BGI) of SKT-1 NahG and npr1 plants showed no significant reduction in disease severity and CMV titre as compared with control plants. In contrast, disease severity and CMV titre were significantly reduced in all Arabidopsis plants treated with culture filtrate (CF) of SKT-1 as compared with control plants. RT-PCR results showed increased expression levels of SA-inducible genes, but not JA/ET-inducible genes, in leaves of BGI treated plants. Moreover, expression levels of SA- and JA/ET-inducible genes were increased in leaves of CF treated plants. In conclusion, BGI treatment induced systemic resistance against CMV through SA signaling cascade in Arabidopsis plants. While, treatment with CF of SKT-1 mediated the expression of a majority of the various pathogen related genes, which led to the increased defense mechanism against CMV infection.

Bacillus thuringiensis Suppresses Bacterial wilt Disease Caused by Ralstonia solanacearum with Systemic Induction of Defense-Related Gene Expression in Tomato

Bacillus thuringiensis is a naturally abundant Gram-positive bacterium and a well-known, effective bio-insecticide. Recently, B. thuringiensis has attracted considerable attention as a potential biological control agent for the suppression of plant diseases. In this study, the bacterial wilt disease-suppressing activity of B. thuringiensis was examined in tomato plants. Treatment of tomato roots with B. thuringiensis culture followed by challenge inoculation with Ralstonia solanacearum suppressed the development of wilt symptoms to less than one third of the control. This disease suppression in tomato plants was reproduced by pretreating their roots with a cell-free filtrate (CF) that had been fractionated from B. thuringiensis culture by centrifugation and filtration. In tomato plants challenge-inoculated with R. solanacearum after pretreatment with CF, the growth of R. solanacearum in stem tissues clearly decreased, and expression of defense-related genes such as PR-1, acidic chitinase, and β-1,3-glucanase was induced in stem and leaf tissues. Furthermore, the stem tissues of tomato plants with their roots were pretreated with CF exhibited resistance against direct inoculation with R. solanacearum. Taken together, these results suggest that treatment of tomato roots with the CF of B. thuringiensis systemically suppresses bacterial wilt through systemic activation of the plant defense system.

Fusarium azukicola sp. nov., an exotic azuki bean root-rot pathogen in Hokkaido, Japan

We report on the phenotypic, molecular phylogenetic and pathogenic characterization of a novel azuki bean (Vigna angularis) root-rot (BRR) pathogen from Hokkaido, Japan, which formally is described herein as Fusarium azukicola. This species can be distinguished phenotypically from the other Phaseolus/Vigna BRR and soybean sudden-death syndrome (SDS) pathogens by the production of wider and longer four-septate conidia cultured on SNA. Molecular phylogenetic analyses of four anonymous intergenic loci, a portion of the translation elongation factor (EF-1α) gene and the nuclear ribosomal intergenic spacer region (IGS rDNA) strongly support the genealogical exclusivity of F. azukicola with respect to the other soybean SDS and BRR pathogens within Clade 2 of the F. solani species complex (FSSC). Evolutionary relationships of F. azukicola to other members of the SDS–BRR clade, however, are unresolved by phylogenetic analyses of the individual and combined datasets, with the exception of the IGS rDNA partition, which strongly supports it as a sister of the soybean SDS pathogen F. brasiliense. A multilocus genotyping assay is updated to include primer probes that successfully distinguish F. azukicola from the other soybean SDS and BRR pathogens. Results of a pathogenicity experiment reveal that the F. azukicola isolates are able to induce root-rot symptoms on azuki bean, mung bean (Vigna radiata), kidney bean (Phaseolus vulgaris) and soybean (Glycine max), as well as typical SDS foliar symptoms on soybean. Our hypothesis is that F. azukicola evolved in South America and was introduced to Hokkaido, Japan, on azuki bean but its possible route of introduction remains unknown.

Biological control of Fusarium wilt of tomato with Fusarium equiseti GF191 in both rock wool and soil systems

The plant growth-promoting fungus (PGPF) Fusarium equiseti GF191 was tested for its ability to control Fusarium wilt of tomato (FWT) caused by Fusarium oxysporum f. sp. lycopersici (FOL) in both a hydroponic rock wool and soil system. F. equiseti effectively controlled FWT, with protective effects based on disease severity of 66.7–88.6% in four experiments. The numbers of colony-forming units of FOL per gram fresh weight of stems were significantly reduced (Pxa0<xa00.05) in plants treated with F. equiseti. Stem extracts from F. equiseti-treated and pathogen-challenged plants significantly inhibited the germination and germ-tube length of FOL microconidia and the production of FOL budding-cells. Tomatine content in tomato stems treated with F. equiseti was significantly increased compared with the non-treated control.

Enhanced Resistance of Plants to Disease Using Trichoderma spp.

Abstract Trichoderma is a ubiquitous fungal genus composed of some of the most versatile biocontrol agents against a wide array of plant diseases. The biocontrol of Trichoderma is achieved through several mechanisms with a combination of two or more mechanisms acting together, probably responsible for the versatility of its biocontrol. A well-known mycoparasite, it secretes cell wall-degrading enzymes and other compounds that can directly kill the target pathogen. It also produces antibiotics, peptaibols, and other bioactive compounds that have antibiosis effect. A competent rhizosphere colonizer, it can compete for space and nutrients with other microorganisms in the rhizosphere. Its most recent discovered property, however, for which a few critical reviews already exist, is its ability to induce local and systemic resistance to a wide variety of plants. In this review we first summarize the multiple beneficial effects of Trichoderma spp. on plants and discuss some rudiments of systemic acquired resistance and induced systemic resistance followed by the recent advances on the enhanced resistance of plants against plant pathogens elicited by the application of several effective biocontrol strains of Trichoderma spp. and finally relating these results to the biological control of plant diseases.

Interaction between the plant growth-promoting fungus Phoma sp. GS8-2 and the arbuscular mycorrhizal fungus Glomus mosseae: impact on biocontrol of soil-borne diseases, microbial population, and plant growth

The influence of the interaction between Phoma sp. GS8-2 (GS8-2) and the arbuscular mycorrhizal fungus Glomus mosseae (Gm) on the development of soil-borne diseases, microbial population, and plant growth in cucumber, bentgrass and tomato plants was investigated under controlled conditions. The composite inoculation of GS8-2 and Gm yielded varied effects on suppression of symptoms of soil-borne pathogens depending on the host-pathogen combination involved. The GS8-2u2009+u2009Gm had neutral effect in reducing damping-off disease of cucumber caused by Rhizoctonia solani AG-4 and the crown and root rot disease of tomato caused by Fusarium oxysporum f. sp. radicis-lycopersici (Forl), but exacerbated the brown patch disease caused by R. solani AG2-2 in bentgrass. In tomato plants, Forl populations in the roots and rhizosphere were significantly reduced by GS8-2u2009+u2009Gm. Results also show that the composite inoculation of GS8-2 and Gm resulted to a synergistic effect on the reduction of fungal and bacterial populations in roots and rhizosphere of the plants. Plant growth enhancement was due to the individual effect of GS8-2 or Gm but not their interaction. GS8-2 root colonization of cucumber and tomato plants decreased significantly in dual inoculated plants compared to plants inoculated with GS8-2 alone; while no significant differences were found in the Gm root colonization of the three plant species indicating that GS8-2 had no effect on Gm.