Differentiation of Rhizoctonia AG-D Isolates from Turfgrass into Subgroups I and II Based on rDNA and RAPD Analyses

Binucleate Rhizoctonia anastomosis group (AG) D is the cause of rhizoctonia-patch and elephant-footprint diseases of zoysiagrass, and winter-patch disease of bentgrass. Rhizoctonia AG-D is also known as the causal pathogen of other diseases such as sharp-eye-spot of cereals, foot-rot of cereals and winter-stem-rot of mat rush. Isolates of AG-D have been divided into the two subgroups AG-D (I) and AG-D (II), based on the results of cultural characteristics and pathogenicity tests. Isolates obtained from zoysiagrass exhibiting symptoms of rhizoctonia-patch disease, from bentgrass with winter-patch disease, from wheat with foot-rot disease, and from mat rush with winter-stem-rot disease were reported to belong to subgroup AG-D (I). On the other hand, isolates obtained from zoysiagrass with elephant-footprint disease were assigned to subgroup AG-D (II). To confirm the existence of these two subgroups in AG-D, the genetic structure of AG-D isolates from turfgrass and other crops was compared. RFLP analysis of the ITS region from rDNA after digestion with the restriction enzymes EcoRI, HaeIII, HhaI, HinfI, and MboI separated AG-D isolates into two groups corresponding to AG-D (I) and AG-D (II). Furthermore, other AGs except AG-Q (AGs-A, Ba, Bb, C, E, F, G, I, K, L, O, P, and R. solani AG1-IC) did not have the same patterns that were seen for the two AG-D subgroups. AG-Q isolates from bentgrass showed the same patterns as AG-D (I). The results of the RAPD analysis also revealed the existence of two groups that corresponded to AG-D (I) and AG-D (II). These analyses revealed that Rhizoctonia AG-D isolates from turfgrass could be divided into two subgroups consistent with those based on cultural characteristics and pathogenicity. In addition, isolates of foot-rot disease of wheat and isolates of winter-stem-rot disease of mat rush whose cultural characteristics were the same as those of AG-D (I) also showed similar RFLP and RAPD patterns to those of AG-D (I) isolates from turfgrass.     

Biological Mode of Action of the Fungicide, Flusulfamide, Against Plasmodiophora brassicae (clubroot)

Flusulfamide (2, 4-dichloro-,,-trifluoro-4-nitro-m-toluenesulfonanilide) was investigated for its mode of action against Plasmodiophora brassicae Woronin. Seedlings of Chinese cabbage (Brassica rapa L. subsp. pekinensis) were grown for 14 and 21 days in soil infested with P. brassicae and then transplanted into soil containing flusulfamide (0.9µg a.i.g^–1 dry soil). Clubroot was not suppressed by this treatment, indicating that the fungicide is ineffective against P. brassicae established within cortical cells of the host root. Where seedlings were grown in soil infested with resting spores which had previously been treated with flusulfamide, root-hair infection and club formation were suppressed. This indicates that flusulfamide directly acts against resting spores. When placed in root exudates of Chinese cabbage, untreated resting spores germinated at a high frequency while flusulfamide-treated resting spores hardly germinated at all. Use of the Evan's blue staining assay indicated that flusulfamide-treated resting spores remained viable. Flusulfamide was detected by high performance liquid chromatography on resting spores treated with flusulfamide for 30min. This indicates that the chemical is adsorbed onto resting spores. These results suggest that flusulfamide suppresses clubroot disease by inhibiting germination of P. brassicae resting spores through adsorption onto their cell walls.     

Lytic activity of l-menthol derivatives against the snow blight disease fungus,Micronectriella nivalis

Lytic activity of l-menthol (1) derivatives [(-)-(1S,3R,4S,6S)-6-hydroxymenthol (2), (-)-(1S,3R,4S)-1-hydroxymenthol (3), and (+)-(1S,3R,4R,6S)-6,8-dihydroxymenthol (4)] against the snow blight disease fungus, Micronectriella nivalis was investigated. Compounds 2, 3, and 4 had 85.0, 63.9, and 81.9% lytic activity, respectively, at a concentration of 0.2 mg/mL. The activity of each of the three compounds increased in a dose-response manner. To study the structure-activity relationship, acetyl esters of 1-4 [(-)-menthyl acetate (1Ac), (-)-6-hydroxymenthyl diacetate (2Ac), (-)-1-hydroxymenthyl 3-monoacetate (3Ac), and (-)-6,8-dihydroxymenthyl 3,6-diacetate (4Ac)] were synthesized with yields of 80.2-99.8% and were also assayed. The acetyl esters of 1Ac, 2Ac, 3Ac, and 4Ac had 51.2, 91.5, 66.0, and 95.2% lytic activity, respectively, at a concentration of 0.2 mg/mL, and these compounds showed further high lytic activity compared with the alcohols of 1-4. These acetyl esters also showed higher lytic activity as their concentration was increased. Of particular interest is the fact that 2Ac and 4Ac both had higher lytic activity at 0.05-0.2 mg/mL compared with copper 8-hydroxyquinolate, a standard chemical widely used to control snow blight. This is the first report on lytic activity of l-menthol derivatives.     

<Emphasis Type="Italic">Talaromyces wortmannii</Emphasis> FS2 emits <Emphasis Type="Italic">β</Emphasis>-caryphyllene,which promotes plant growth and induces resistance

A plant-growth-promoting fungus (PGPF), Talaromyces sp. was isolated from an agricultural field in southwestern Japan. We found that this fungus emitted several terpenoid-like volatiles including β-caryophyllene. Then we investigated the effect of β-caryophyllene on promoting the growth and inducing resistance of Brassica campestris L. var. perviridis. The compound significantly enhanced the growth of seedlings and their resistance to Colletotrichum higginsianum. On the basis of these results, we discuss the role of β-caryophyllene in the activities of PGPF.     

Systemic resistance to bacterial leaf speck pathogen in Arabidopsis thaliana induced by the culture filtrate of a plant growth-promoting fungus (PGPF) Phoma sp. GS8-1

The culture filtrate (CF) from the plant growth-promoting fungus Phoma sp. GS8-1 was found to induce systemic resistance in Arabidopsis thaliana against the bacterial leaf speck pathogen Pseudomonas syringae pv. tomato DC3000 (Pst), and the underlying mechanism was studied. Roots of A. thaliana were treated with CF from GS8-1, and plants expressed a clear resistance to subsequent Pst infection; disease severity was reduced, and proliferation of pathogen was suppressed. Various mutants of A. thaliana were used to test whether the CF induced resistance through one of the known signaling pathways: salicylic acid (SA), jasmonic acid (JA), and ethylene (ET). The CF was fully protective against Pst in Arabidopsis mutants jar1 and ein2 similar to wild-type plants. However, its efficacy was reduced in plants containing transgene NahG. Examination of systemic gene expression revealed that CF modulates the expression of SA-inducible PR-1, PR-2 and PR-5 genes, the JA/ET-inducible ChitB gene, and the ET-inducible Hel gene. Moreover, the expression of these genes was further enhanced upon subsequent stimulation after attack by Pst. Our data suggest that in addition to a partial requirement for SA, the signals JA and ET may also play a role in defense signaling in Arabidopsis.     

Suppression of Fusarium wilt of spinach with hypovirulent binucleate <Emphasis Type="Italic">Rhizoctonia</Emphasis>

 Four isolates of hypovirulent binucleate Rhizoctonia (HBNR) were evaluated for their ability to control Fusarium wilt of spinach (FWS) caused by Fusarium oxysporum f. sp. spinaciae (FOS). Fourteen-day-old spinach seedlings grown in paper pots with HBNR-amended soil (1% w/w ground barley grain inoculum) were transferred to artificially pathogen-infested soil. Treatments with HBNR isolates significantly (P = 0.05) reduced disease and discoloration severity by 56%–100% and 52%–100%, respectively. The numbers of colony-forming units of FOS per gram fresh weight in petioles or roots were reduced significantly (P = 0.01) in the plants treated with HBNR. HBNR isolates were well reisolated from the roots inside paper pots where they were inoculated, whereas inconsistent colonization of HBNR was recorded from the roots outside paper pots where only pathogen was inoculated. Root extracts from HBNR-treated and pathogen-challenged plants significantly inhibited germination and germling length of FOS. The fresh weight of spinach leaves in the HBNR-treated plants increased significantly (P = 0.01), as much as 53%–63%, over the untreated and pathogen-challenged plants. This is the first report of biocontrol of FWS by HBNR. Received: July 18, 2002 / Accepted: October 22, 2002 Acknowledgments We are grateful to Dr. Komada for providing nonpathogenic Fusarium F13. The senior author (A.M.) thanks the Ministry of Education, Culture, Sports, Science, and Technology (Monbukagakusho) Japan, for financial assistance.     

Resistance of Sorghum Line CS 621 to Rhizoctonia solani AG1-IA and Other Sorghum Pathogens

Sorghum line CS 621 was evaluated along with other lines and cultivars for resistance to Rhizoctonia sheath blight, tar spot and gray leaf spot for 3 years. CS 621 was consistently resistant to these diseases even under a heavy natural outbreak of tar spot in the breeding nursery. It was also found to be more resistant to Rhizoctonia sheath blight than the resistant lines from Japan during the 1993 to 1994 screening tests. Resistance to Rhizoctonia solani AG1-IA was expressed in the form of reduction in the severity of disease and rate of infection. Evaluation of progenies from crosses involving CS 621 and a susceptible variety, UPL Sg5, indicated that additive and dominant gene effects are important in the expression of quantitative resistance to R. solani. CS 621 could therefore serve as a source of multiple resistance genes in a breeding program for high yield and stability against sorghum diseases. Received 6 August 1999/ Accepted in revised form 18 October 1999     

Characterization of acetylated wood decayed by brown-rot and white-rot fungi

The objective of this study was to characterize the decay of acetylated wood due to brown-rot and white-rot fungi by analysis of chemical composition, X-ray measurements, and¹³C-NMR spectroscopy. The decay by brown-rot fungus became inhibited at a weight percent gain (WPG) due to acetylation of more than 10%, and the mass loss (LOSS) due to decay became zero at a WPG of about 20%. The LOSS due to white-rot fungus decreased slowly with the increase in WPG, reaching zero at a WPG of about 12%. The losses of lignin by brown-rot decay increased initially with the decrease in LOSS owing to the progressing acetylation and then decreased at a LOSS of less than 60%. Polysaccharides were more easily decomposed than lignin during the decay of acetylated wood due to brown-rot fungus. The losses of both components due to white-rot decay decreased as the LOSS decreased with progressing acetylation. The white-rot fungus tended to preferentially decompose the lignin during the decay of acetylated wood. The brown-rot fungus decomposed the cellulose in the crystalline region to a large degree when the LOSS was more than 40%, whereas the white-rot fungus decomposed the crystalline region and the noncrystalline region in acetylated wood to the same degree. The brown-rot fungus preferentially decomposed unsubstituted xylose units in acetylated wood and partly decomposed the mono-substituted xylose units. It was suggested that the mono- and disubstituted cellulose were partly decomposed by brown-rot fungus.This paper was presented at the 46th and 47th annual meetings of the Japan Wood Research Society at Kumamoto and Kochi in April 1996 and April 1997, respectively     

Population structure of <Emphasis Type="Italic">Fusarium asiaticum</Emphasis> from two Japanese regions and eastern China

Genetic subdivision of Fusarium asiaticum was investigated using a collection of 478 isolates originating from the Kyushu area and Aichi Prefecture, Japan and Zhejiang Province in China. Trichothecene-type determination by a multiplex PCR-test indicated that all isolates were either of a nivalenol (NIV) or a 3-acetyl deoxynivalenol (3ADON) type. The 15-acetyl deoxynivalenol (15ADON) type was not detected in this collection. Based on a Bayesian model-based clustering method using allele data obtained with 11 variable number of tandem repeats (VNTR) markers, we detected three genetic clusters. The majority of isolates in the clusters were NIV isolates from both Japan and China, Japanese 3ADON and Chinese 3ADON isolates, respectively. High levels of fixation indices and low levels of effective number of migrants were observed between the genetic clusters. Data was re-analyzed by classifying the isolates into six groups according to trichothecene type and geographic location. Population analyses of these re-classified groups indicated that the genetic subdivisions of F. asiaticum were correlated with both trichothecene type and geographic differences. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.