MgO-induced defence against bacterial wilt disease in Arabidopsis thaliana

Bacterial wilt, caused by Ralstonia solanacearum, is a devastating soilborne disease in plants that limits the production of many crops worldwide. Although management of bacterial wilt has so far been unsuccessful, enhancing host resistance to the pathogen may be an effective control strategy. Recently, magnesium oxide (MgO) was found to induce defence responses against R. solanacearum in tomato plants. Here, the mechanisms underlying MgO-induced defence responses against R. solanacearum (MgO-i DARS) were investigated using Arabidopsis thaliana as a host plant. MgO-i DARS was confirmed in A. thaliana mutants deficient in jasmonic acid or ethylene signalling pathways as well as in the wildtype (Col-0) plants. In contrast, no MgO-i DARS was found in A. thaliana mutants deficient in the salicylic acid (SA) production (sid2-2) and signalling pathways (tga1-1 and npr-1). MgO treatment led to significant accumulation of SA in both roots and shoots of Col-0. The SA biosynthesis gene isochorismate synthase 1 (ICS1) was induced in roots and shoots of A. thaliana treated with MgO. An NADPH oxidase gene respiratory burst oxidase homolog D (AtRbohD) was up-regulated in both roots and shoots of Col-0 treated with MgO. No MgO-i DARS was observed in A. thaliana mutants deficient in AtRbohD. These results suggest that SA and RBOHD-mediated ROS are pivotal for MgO-i DARS in A. thaliana.     

芽胞杆菌BvR001对大丽轮枝菌抑制效果研究

为研发绿色、安全、高效的作物黄萎病生物防治产品,本研究以生防菌株BvR001为研究对象,通过对黄萎病菌的抑菌效果测定、可湿性粉剂研制、在寄主根际的定殖能力与防效测定,明确该菌株对黄萎病具有防控效果.gyrB序列检测和系统发育树构建分析表明,该菌株属于芽胞杆菌属,并且与贝莱斯芽胞杆菌Bacillus velezensis...     

Increased resistance to Verticillium dahliae in Arabidopsis plants defective in auxin signalling

Auxin signalling and transport participate in plant–microbe interactions as positive or negative regulators of disease resistance. The present study investigated the responses of Arabidopsis thaliana plants impaired in the auxin receptors TIR1, AFB1 and AFB3 and the auxin transporter AXR4, upon infection by the soilborne root pathogen Verticillium dahliae. Fewer symptoms were recorded in afb1, afb3 and axr4 plants compared to the wild type (wt). qPCR analysis revealed that the decrease in symptom severity in afb1, afb3 and axr4 was correlated with reduction in the growth of the pathogen in the plants. Therefore, afb1, afb3 and axr4 are partially resistant to V. dahliae. Upon V. dahliae infection, the expression of TIR1, AFB1, AFB3 and AXR4 was up‐regulated in roots, while indole‐3‐acetic acid levels were similar to mocks. The partial resistance of afb1, afb3 and axr4 against V. dahliae can be attributed in part to the up‐regulation of defence‐related genes, as it was observed that afb1 and axr4 had higher PR1 levels than wt, while afb3 had higher PDF1.2 levels than wt. The findings of the present study suggest that the auxin signalling defective mutants afb1, afb3 and axr4 show increased resistance against V. dahliae.     

Effects of Verticillium dahliae on tomato root morphology considering plant growth response and defence

The soilborne pathogen Verticillium dahliae invades its host via the root, and spreads systemically throughout the plant. Although a functional root system of appropriate size is essential for water and nutrient uptake, to date, effects of pathogens on root morphology have not been frequently investigated. Therefore, this study aims to improve knowledge of how V. dahliae infection impairs root morphological characteristics of tomato, considering plant growth and physiological responses, particularly those involved in defence in roots and leaves over a growing period of up to 28 days post‐inoculation. Verticillium dahliae infection suppressed the growth of both shoot and root. Diseased plants developed a smaller leaf area, and exhibited a reduction in the rate of photosynthesis and stomatal conductance. An early response to pathogen invasion in the host root was the up‐regulation of several defence‐related genes, such as proteinase inhibitor II (Pin2), β‐1,3‐glucanase A (GluA) and two pathogenesis‐related genes (PR‐1a, PR‐1b). However, this response did not prevent colonization of the roots by the pathogen. Although a high variability in pathogen density was found within the root system, a significant increase of both the specific root length and surface area was observed in response to pathogen invasion; these traits correlated with water use efficiency. Morphological changes of the root may represent an adaptive response evolved to sustain the supply of both water and nutrients in the presence of the pathogen.     

棉花黄萎病拮抗细菌筛选与B-101菌株抗菌蛋白分离

 The antifungal proteins of antagonistic strains NCD-2, B-101, DHT-13, BDT-25, JJ-134, JJ-102 and 80b-2 isolated from the soil infected with Verticillium dahliae were obtained from their fermentation fil-trates through precipitation with 75% ammonium sulfate salting out. Antagonistic activities of the proteins were determined via cylinder-plate method. Results indicated that antifungal proteins from 80b-2, NCD-2, DHT-13 and B-101 had higher specific activity, of which B-101 and NCD-2 could produce clear inhibition zone. Antifungal protein of B-101 strain was obtained by using the salt fractionation. It was found that protein precipitated with 20%-30% saturation ammonium sulfate could produce transparent and bigger inhibition zone, thus it had obvious antifungal activity. Furthermore the antifungal protein was separated by DEAE-cellulose chromatography. An antifungal protein fraction named RFP-6 was obtained, which showed definite inhibition activity against Verticillium dahliae. Its molecular weight was approximately 14.4 kD as determined with SDS-PAGE.     

25种中草药提取物对棉花黄萎病菌的抑制作用

以棉花黄萎病菌Verticillium dahliae为供试菌种,采用菌丝生长速率法测定了25种中草药乙醇提取物的抑菌作用。结果表明,25种中草药乙醇提取物中,花椒、黄柏和木香的抑菌作用最好,抑制率分别为59.30%、48.29%和45.54%。同时,这3种中草药的乙醇提取物抑菌活性均显著高于石油醚提取物的抑菌活性。在室内毒力测定中,花椒、黄柏和木香的乙醇提取物对供试菌株的EC_(50)分别为3.74,13.98和17.02 mg/mL。     

Involvement of kaempferol in the defence response of virus infected Arabidopsis thaliana

The roles of several phenolic compounds in plant defence response have been extensively studied, yet little is known about the role of flavonoids in plant-virus interaction. Quantitative and qualitative changes of selected phenolics in Arabidopsis thaliana induced by Cucumber mosaic virus containing satellite RNA (CMVsat) infection were analysed accompanied by plant hormone, chalcone synthase and pathogenesis-related gene expression analysis. Lower leaves of infected plants had a lower concentration of total phenolics compared to control plants. The concentration of kaempferol in upper leaves of all infected plants was significantly lower compared to control plants, while the expression of the chalcone synthase gene in those leaves was in most cases upregulated. All infected plants had a higher concentration of indole-3-acetic acid in lower leaves, which was accompanied with a lower concentration of kaempferol in upper leaves. Our research demonstrates a correlation between kaempferol and indole-3-acetic acid in response to CMVsat infection in Arabidopsis. We demonstrated two different metabolic patterns in infected plants suggesting the activation of two different defence responses. We also propose kaempferol to be an important part of the auxin-dependent defence response which limits systemic movement of CMVsat and that this defence response is activated prior to the well-known salicylic acid dependent defence response. Further research on kaempferol and its role in Arabidopsis-CMVsat interaction will improve our understanding on the role of flavonoids in plant defence.     

棉花黄萎病防治策略

黄萎病是棉花生产中的最主要病害之一,广泛分布于世界各产棉国.由于该病是土壤传播的维管束病害,防治难度较大,至今尚无特效的防治药剂,只能依靠种植抗病品种为主的综合防治措施.但是,我国目前棉花品种的抗病性只能达到抗至耐病水平,致使该病在环境条件适宜时连续流行为害.控制该病的猖獗为害,已成为棉花生产可持续发展的主要问题之一.     

Fusarium oxysporum Fo47 confers protection to pepper plants against Verticillium dahliae and Phytophthora capsici,and induces the expression of defence genes

The application of the nonpathogenic isolate Fusarium oxysporum 47 (Fo47) reduced the symptoms of verticillium wilt, phytophthora root rot and phytophthora blight in pepper plants. Botrytis cinerea was also tested on the leaves of plants treated with Fo47, but no protection was observed. Verticillium dahliae colonies cultured in the presence of Fo47 grew slower than control cultures, but Phytophthora capsici growth was unaffected by Fo47. At least part of the protection effect observed against V. dahliae could therefore be due to antagonism or competition. In order to search for induced resistance mechanisms, three defence genes previously related to pepper resistance were monitored over time. These genes encode a basic PR‐1 protein (CABPR1), a class II chitinase (CACHI2) and a sesquiterpene cyclase (CASC1) involved in the synthesis of capsidiol, a phytoalexin. These three genes were transiently up‐regulated in the roots by Fo47 in the absence of inoculation with the pathogen, but in the stem only CABPR1 was up‐regulated. In plants that were inoculated with V. dahliae after the Fo47 treatment, the three genes had a higher relative expression level than the control in both the roots and the stem.     

Vegetative-compatibility groups in Verticillium dahliae from Israel

Nitrate-nonutilizing (nit) mutants were used to determine vegetative compatibility among 34 isolates of Verticillium dahliae from cotton, potato, olive, eggplant, chrysanthemum and tomato from 12 sites in Israel. Based on the formation of complementary heterokaryons, 33 isolates were assigned to two vegetative- compatibility groups (VCGs): one VCG contained 15 isolates from cotton, eggplant, chrysanthemum and olive; and the other VCG contained 18 isolates from potato, olive and cotton. The status of an additional isolate from tomato, which was compatible with both VCGs, remained unclear. In a limited pathogenicity test with 10 isolates, two (from tomato and eggplant) were pathogenic on tomato, eggplant and cotton; most isolates from cotton were pathogenic on cotton and eggplant only; and one from cotton was non-pathogenic. Fewer isolates were pathogenic on tomato than on cotton or eggplant. The diversity of vegetative compatibility found in our V. dahliae collection is comparable to that found in studies of American populations.     

Genetic variability of tolerance to Verticillium albo-atrum and Verticillium dahliae in Medicago truncatula

Verticillium wilt caused by Verticillium albo-atrum and Verticillium dahliae is responsible for yield losses in many economically important crops. The capacity of pathogenic fungi to adapt to new hosts is a well-known threat to the durability of resistant crop varieties. In the present work, 25 Medicago truncatula genotypes from a core collection and six V. albo-atrum and V. dahliae strains were used to study the potential of non-host Verticillium strains isolated from different plant species to infect this legume plant and the plant’s susceptibility to the pathogens. The experiment was designed as factorial with randomised complete blocks and with three repetitions. The wilt symptoms caused by V. albo-atrum and V. dahliae were scored on a disease index scale from 0 to 4, during 30 days after inoculation of 10-day-old plantlets. Disease severity was quantified by maximum symptom scores (MSS) and areas under the disease progress curves (AUDPC), which integrate the time course of symptom development. Highly significant differences were observed among plant genotypes and fungal strains, and their interaction was also significant. The correlation between MSS and AUDPC was 0.86. The most severe symptoms were caused by the alfalfa strain V. albo-atrum V31-2 and the least severe by V. dahliae JR2, as shown by mean values obtained on the 25 genotypes. M. truncatula genotype TN8.3 was the most susceptible genotype by mean values obtained with the six fungal strains, whereas F11013-3, F83005.9 and DZA45.6 were highly resistant to all strains studied. The results can be used to choose parents for studying the genetics of resistance in Medicago truncatula to Verticillium strains with different levels of aggressiveness.     

Spatial Patterns of Microsclerotia of Verticillium dahliae in Soil and Verticillium Wilt of Cauliflower

ABSTRACT The spatial patterns of microsclerotia of Verticillium dahliae in soil and wilt symptoms on cauliflower were determined at three sites in each of two fields in 1994 and 1995. Each site was an 8 x 8 grid divided into 64 contiguous quadrats (2 by 2 m each). Soil samples were collected to a depth of 15 cm with a probe (2.5 cm in diameter), and samples from four sites in each quadrat were bulked. Plants in each quadrat were cut transversely, and the number of plants with vascular discoloration and the number without discoloration were recorded. The soil was assayed for microsclerotia by the modified Anderson sampler technique. Lloyd's index of patchiness (LIP) was used as an indicator to evaluate the aggregation of microsclerotia in the field. Spatial autocorrelation and geostatistical analyses were also used to assess the autocorrelation of microsclerotia among quadrats. The LIP for microsclerotia was greater than 1, indicating aggregation of propagules; however, the degree of aggregation at most sites was not high. Significant autocorrelation within or across rows was detected in some spatial autocorrelograms of propagules, and anisotropic patterns were also detected in some oriented semivariograms from geostatistical analyses for microsclerotia, indicating the influence of bed preparation in the fields on pathogen distribution. The parameter estimates p and theta in the beta-binomial distribution and the index of dispersion (D) associated with the distribution were used to assess the aggregation of diseased plants at each site. A random pattern of wilt incidence was detected at 7 of 12 sites, and an aggregated pattern was detected at 5 of 12 sites. The degree of aggregation was not high. A regular pattern of wilt severity was detected at all sites. The high disease incidence (77 to 98%) observed at 11 of the 12 sites could be explained by high inoculum density.     

Hosts of Verticillium dahliae in Kriti (Greece)*

Naturally wilted cultivated and weed species were collected from several fields in Kriti (GR) over the years 1992/1997, and tested by isolation for the presence of Verticillium dahliae. The fungus was most frequently recovered from cultivated Solanaceae and Cucurbitaceae. Infection of many known Verticillium hosts was confirmed, and the fungus was found for the first time in Kriti in pistachio and grapevine and in several common weeds.     

Induction of resistance in cocoa against Crinipellis perniciosa and Verticillium dahliae by acibenzolar- S-methyl (ASM)

The benzothiadiazole compound acibenzolar- S -methyl (ASM) was assessed as an inducer of resistance against Crinipellis perniciosa , agent of witches' broom, and Verticillium dahliae , agent of vascular wilt, both on cocoa. ASM induced a reduction in incidence of witches' broom of up to 84·5% when sprayed 30 days before inoculation on cocoa seedlings of cv. Catongo. ASM also induced a reduction in severity of Verticillium wilt to 55·4% on cv. Theobahia. For both pathosystems, effects of dose on disease were not clearly observed. The efficacy of the inducer increased with the interval between sprayings and the respective inoculations with the pathogens. In another experiment, the effect of ASM on the control of witches' broom on cocoa seedlings was compared with that of cuprous oxide and tebuconazole, all sprayed 15 days before inoculation. ASM reduced disease incidence by 60·1% compared with the inoculated control. ASM was superior to tebuconazole, and there was also a tendency for ASM to be better than cuprous oxide. To understand the mechanism of action of ASM as an inducer of resistance, alterations in the levels of total phenolics, polyphenol oxidases and peroxidases were evaluated 3, 15 and 30 days after spraying of seedlings of cv. Catongo. Enzyme activities from seedlings of cv. Theobahia were evaluated 30 days after spraying. On cv. Catongo, no significant differences in total phenolic content and polyphenol oxidase activity were detected after spraying. However, an increase in peroxidase activity was detected at all times of evaluation. On cv. Theobahia, significant increases in activities of peroxidase and polyphenol oxidase were detected, indicating that defence responses due to ASM were dependent on host genotype.     

山东棉花黄萎菌致病力分化的研究

选用山东省有代表性的棉黄萎菌１６个菌株（系）各配制成一定浓度的孢子悬浮液，于棉苗２～３叶期蘸根接种，观察发病的反应型。结果表明：各菌株致病力强弱差异明显，可分为致病力强的Ⅰ型（即与落叶型相似）、致病力中等的Ⅱ型和致病力弱的Ⅲ型，并发现山东省有落叶型菌系存在。     

土壤大丽轮枝菌微菌核的快速定量检测

 微菌核是大丽轮枝菌在土壤中的主要存活结构和黄萎病的初侵染来源。对土壤中大丽轮枝菌微菌核进行定量是黄萎病监测和预警的基础。本研究以大丽轮枝菌Internal Transcribed Spacer (ITS)区特异性引物对P1/P2扩增产物的重组质粒为标准品,构建SYBR Green I实时荧光定量PCR反应的标准曲线,结合土样水筛法建立了土壤大丽轮枝菌微菌核定量检测体系。同时,建立了土壤中微菌核数量与棉花黄萎病发病率的关系模型。结果表明,实时定量PCR检测灵敏度比常规PCR高10倍,检测下限为1个微菌核/克土,在5.54×10²～5.54×10⁷copies范围内,DNA拷贝数的对数值与Ct值具有良好的线性关系。建立的土壤中微菌核个数n与Ct值之间的关系为n=e^7.3-Ct/3.905。温室人工接种微菌核数量与棉花黄萎病发病率间的线性关系为y=2.710n+0.251。