Screening for resistance of Turkish olive cultivars and clonal rootstocks to Verticillium wilt

Verticillium wilt resistance of 77 olive cultivars including 71 domestic and six foreign ones, and four clonal rootstocks available in the olive gene bank, were tested using a highly virulent isolate (D pathotype) of Verticillium dahliae. The pathogen was stem-inoculated into the own-rooted saplings. Most cultivars and rootstocks were found to be extremely susceptible to the disease. ‘Sinop No. 1’, ‘E?riburun Nizip’, ‘Erkence’, ‘E?riburun Tatayn’, ‘Girit Zeytini’ and ‘Marantelli’ were highly resistant, as their disease severities did not exceed 10%. Additionally, 11 domestic cultivars (‘Sar? Habe?i’, ‘Ya?l?k Çelebi’, ‘Zoncuk’, ‘Dilmit’, ‘?am’, ‘Hurma Karaca’, ‘Erdek Ya?l?k’, ‘Melkabaz?’, ‘Yün Çelebi’, ‘Kan Çelebi’ and ‘Siyah Salamural?k’), two foreign cultivars (‘Arbequina’ and ‘Frantoio’) and one wild clonal rootstock (‘D36’) were found to be resistant, with disease severities less than 30%. On the other hand, the moderately susceptible group comprised ten domestic cultivars (‘Ak Zeytin’, ‘Ya? Çelebi’, ‘Saurani’, ‘Butko’, ‘Gemlik’, ‘Otur’, ‘Ya? Zeytini’, ‘Belluti’, ‘Sinop No. 2’ and ‘Samanl?’), three foreign cultivars (‘Leccino’, ‘Chemlali’ and ‘Ascolana’) and one wild clonal rootstock (‘D9’). The number of cultivars within highly resistant and resistant groups was 17 out of the 71 domestic cultivars from all regions (four from Aegean, seven from southeastern Anatolia, two from Black Sea and three from Marmara).     

番茄黄萎病病原菌研究

近年在黑龙江省保护地番茄上发现一种番茄植株黄萎病害,从其病茎上分离得到10个菌株。将其接种番茄幼苗,发病症状与自然发病症状完全一致,并从接种病株上重新分离到该菌。经鉴定,病原菌为大丽轮枝菌(Ver-tillium dahliae Kleb)。对其生物学特性研究表明:该菌生长最低温度为5℃,最适范围20-25℃,以22.5℃生长最适。孢子萌发温度为5-30℃,最适温度为25℃。病菌菌丝在pH4-8都能生长,以pH 5生长最好。     

Insights into the role of ethylene perception in tomato resistance to vascular infection by Verticillium dahliae

A Tobacco rattle virus (TRV)-based virus-induced gene silencing (VIGS) system was employed to investigate the role of the tomato ethylene receptor ETR4. By comparing wilting symptoms of verticillium wilt in wild-type, ethylene-insensitive Never ripe ( Nr ) mutant tomato plants and ETR4 -silenced plants, it was demonstrated that disease severity in the Nr and ETR4 -silenced plants was statistically reduced compared to wild-type plants. Disease incidence and severity were reduced by 11 and 20%, respectively, in the Nr plants compared to the wild-type plants, at 33 days post-inoculation (d.p.i.). In the ETR4 -silenced plants, disease incidence and severity were reduced by 14 and 15%, respectively, compared to the TRV -only-inoculated plants, at 37 d.p.i. Quantification of Verticillium dahliae by qPCR revealed that the reduction in symptom severity in the Nr plants was associated with significant reduction of growth of the pathogen in the vascular tissues of the Nr plants compared to that in the wild-type plants, suggesting that impaired perception of ethylene via the Never-ripe receptor results in increased disease resistance. Fungal reduction was evident at each sampling day in the Nr plants, ranging from 1·5 to 1·75 times less than that in the wild-type plants. Fungal quantification in the ETR4- silenced and TRV -only-inoculated plants showed similar levels of fungal biomass.     

Investigation of in-vitro methods for measuring eggplant resistance to Verticillium dahliae

An assay based on electrolyte leakage (LEA) from callus tissue of Solanum spp. treated with culture filtrates of Verticillium dahliae was useful for screening eggplant genotypes for resistance to Verticillium wilt. Loss of electrolytes from the most resistant genotypes was significantly lower than that from the susceptible genotypes when callus tissue was incubated with a 10% filtrate solution of the virulent isolate 122. No difference was detected with filtrate solutions of the avirulent isolate 809 of the fungus. Among the tested genotypes, the S. melongena lines CCR3 and SM19/14 and the wild species S. torvum were the most resistant in the glasshouse tests and released the smallest amounts of electrolytes. The most susceptible cultivars, Lunga Violetta and Giant of China, produced the greatest leakage of electrolytes. LEA performed on stem segments failed to distinguish between susceptible and resistant genotypes of eggplant.     

Sulphur supply impairs spread of Verticillium dahliae in tomato

Vascular wilt caused by the soil-borne fungus Verticillium dahliae is a major yield and quality-limiting disease across a broad spectrum of crop plants worldwide. Sulphur-enhanced plant defence mechanisms provide an opportunity to effectively and environmentally safely constrain the wilt disease levels in planta. To evaluate the influence of sulphur nutrition on the protective potential of these mechanisms, two near-isogenic tomato genotypes differing in fungal susceptibility, were treated with low or supra-optimal sulphur supply. Microscopic analysis revealed a significant sulphur-induced decrease in the amount of infected vascular cells in both genotypes. However, plant shoot and severely pathogen-affected root growth did not display this distinct alleviating influence of sulphur nutrition. Rates of leaf photosynthesis were impeded by Verticillium dahliae infection in both genotypes especially under low sulphur nutrition. However, assimilate transport rates in the phloem sap were enhanced by fungal infection more in the resistant genotype and under high sulphur nutrition suggesting a stronger sink for assimilates in infected plant tissues possibly involved in sugar-induced defence. A SYBR Green-based absolute quantitative Real-Time assay using a species-specific primer was developed which sensitively reflected sulphur nutrition-dependent changes in fungal colonization patterns. High sulphur nutrition significantly reduced fungal spread in the stem in both tomato genotypes. Concentrations of selected sulphur-containing metabolites revealed an increase of the major anti-oxidative redox buffer glutathione under high sulphur nutrition in response to fungal colonization. Our study demonstrates the existence of sulphur nutrition-enhanced resistance of tomato against Verticillium dahliae mediated by sulphur-containing defence compounds.     

Variation for pathogenicity on tomato among parasexual recombinants of Verticillium dahliae

Ten haploid prototrophic recombinant isolates (HPRs) were obtained from each of 15 parasexual crosses between complementary autotrophs derived from nine tomato isolates and one eggplant isolate of V. dahliae , including those identified as race 1 and race 2. These HPRs were tested for pathogenicity to the tomato cultivar Roma which is susceptible to both races. HPRs from a 'selfed' race 2 × race 2 cross were as pathogenic as the wild-type parent. The pathogenicity of HPRs derived from all other crosses was variable, and generally lower than the parental mean. A particularly marked reduction in pathogenicity, compared with the parental mean, was observed for HPRs recovered from two crosses between isolates belonging to different heterokaryon compatibility groups. The results suggest that pathogenicity to cv. Roma is controlled by complex interactions between genes at numerous loci.     

棉花抗黄萎病品种选育方法探讨

 在棉花抗病育种中,通过病圃或重病地选择抗病单株是常规的选育方法,但其在抗黄萎病育种中的效果并不理想。如何提高抗黄萎病育种效率,促进育种进程是目前急需解决的问题之一。为此,19982002年我们在北京人工病圃,对不同抗病杂交组合和选育方法的选育效果进行了研究。结果表明,在双亲本均为抗(耐)品种的杂交后代群体中,以9月中旬是抗病还是感病并没有多少关联,对黄萎病抗性的选育应选群体抗病性强的株系或组合,以其丰产性为主要选育目标;只有抗与抗(或高耐)的组合中才能选出黄萎病抗性更高的株系;在抗(耐)品种中,后期无论是抗或感的单株其后代的抗病性差异不明显     

Race 2 of Verticillium dahliae infecting tomato in Japan can be split into two races with differential pathogenicity on resistant rootstocks

Verticillium dahliae infecting tomato can be differentiated into races 1 and 2 based on differential pathogenicity on tomato cultivars carrying resistance gene Ve1. Although no commercial cultivars resistant to race 2 are available, race 2‐resistant rootstock cultivars Aibou and Ganbarune‐Karis have been bred in Japan. Nevertheless, the resistance of these rootstocks appears to be unstable in commercial tomato fields. Pathogenicity assays conducted under controlled conditions revealed that these rootstock cultivars are resistant to some isolates of race 2; this resistance is controlled by a single dominant locus, denoted by V2, based on segregation of resistance in F₂ populations from selfed rootstock cultivars. However, some other isolates of race 2 can overcome this resistance. Therefore it is proposed that the current race 2 of V. dahliae should be divided into two races, i.e. ‘race 2’ (nonpathogenic on Aibou) and ‘race 3’ (pathogenic on Aibou). The distribution of these races was surveyed in 70 commercial tomato fields in Hida, Gifu Prefecture, Japan. Race 3 was found in 45 fields, indicating that race 3 had already spread throughout the region. On the other hand, 25 fields had only race 2, and thus race 2‐resistant rootstocks would be effective for disease management in these fields. Races 2 and 3 could not be identified by genomic Southern hybridization probed with a telomere sequence, nor with previously reported race‐specific PCR assays. Elucidation of race‐determining mechanisms and development of methods for quick race identification should be made in future studies.     

Screening for resistance to Didymella bryoniae in rootstocks of melon

Glasshouse evaluations of rootstocks of melon for resistance to Didymella bryoniae were conducted on different cucurbits previously selected as resistant or partially resistant to race 1,2 of Fusarium oxysporum f. sp. melonis. Cucumis anguria, C. ficifolius, C. figarei, C. metuliferus, C. zeyheri and Benincasa hispida showed a very high degree of resistance to D. bryoniae both on leaves and stems. Among the commercial rootstocks, Cucurbita hybrids ELSI, ES 99-13, RS 841, displayed a similar level of resistance. Stem inoculation of three cucurbit species grafted with susceptible melon cv. Proteo determined the occurrence of limited symptoms, but the lesions remained confined in the rootstock not affecting the grafted plant. Cultivation of susceptible melon cultivars grafted on resistant rootstocks may represent an efficient method for controlling Didymella crown rot and Fusarium wilt.     

Aggressiveness of Verticillium dahliae isolates from different vegetative compatibility groups to potato and tomato

Aggressiveness of Verticillium dahliae isolates from three vegetative compatibility groups (VCGs) was tested on potato and tomato. VCG4B was the most aggressive to potato and VCG2A was the most aggressive to tomato; VCG2B was the least aggressive to both potato and tomato. In potato, disease incidence, symptom severity and colonization index of stem segments were significantly higher in plants inoculated with VCG4B isolates than in those inoculated with VCG2B and VCG2A isolates. Inoculation with VCG4B and VCG2A decreased plant height and fresh weight more than inoculation with VCG2B. In tomato, VCG2A caused significantly more severe symptoms than either VCG4B or VCG2B. The colonization index in tomato plants inoculated with VCG2A was also significantly higher than in those inoculated with VCG4B and VCG2B. Similar patterns of relative aggressiveness were observed in potato and tomato when the pathogenicity of isolates of various VCGs, each originating from a specific host (cotton, potato or eggplant), was compared.     

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.     

陆地棉对黄萎病抗性的分子标记研究

 利用陆地棉标准系TM-1和常抗棉2个陆地棉品种杂交并自交,获得109个F₂单株及F_2:3家系为作图群体,以SSR、RAPD和SRAP 3种分子标记进行抗黄萎病性状的分子标记筛选。结果从1611对(条)引物中仅筛选到70对(条)多态性引物,获得75个多态性位点并进行标记间的连锁性分析。75个标记构建了一个包括15个连锁群,全长535 cM的陆地棉品种间分子标记遗传连锁图,标记间平均距离为11.15 cM,有27个标记不能进入任何连锁群。连锁群的标记数最少2个,最多6个;长度从1.0 cM到92.7 cM不等。对其F_2:3家系的成株期抗黄萎病性状即平均病情指数的分布进行分析,显示其呈正态分布,进一步说明陆地棉对黄萎病的抗性为数量遗传;单标记分析及复合区间作图,检测出与抗黄萎病性相关的3个QTL,分别位于第3、5、6连锁群上,贡献率分别为14.15%、3.45%和18.78%。另外,对该群体生长过程中黄萎病不同发病高峰期的病情也进行了分析。     

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.     

Evaluation of Olive Cultivars for Resistance to Verticillium dahliae

Resistance of 23 important olive cultivars to Verticillium dahliae has been evaluated in four experiments under controlled conditions. Nine-month-old nursery olive plants were inoculated with a cotton non-defoliating (ND) (V4) or a cotton defoliating (D) (V117) isolate of V. dahliae. Resistance was evaluated by assessing symptom severity using a 0–4 rating scale and estimating the area under disease progress curves. The percentage of plants killed and of those which recovered from the disease were used as additional parameters for including a particular cultivar into a defined category. Most of the evaluated cultivars were susceptible, although at different levels, to both isolates of V. dahliae. All cultivars were more susceptible to the D pathotype than to the ND one. A group of 11 cultivars, including several important Spanish cultivars, were susceptible or extremely susceptible to both pathotypes of V. dahliae. A second group showed differences of resistance depending on the pathotype used. They were susceptible or extremely susceptible to the D pathotype but resistant or moderately susceptible to the ND one. Finally, 'Frantoio', 'Oblonga' and 'Empeltre' were moderately susceptible to the D isolate of V. dahliae and resistant to the ND one. The resistance of 'Empeltre' was evident by the plant ability to recover from infection with either isolates. 'Empeltre' is considered to be a valuable cultivar for inclusion in breeding programmes for resistance to Verticillium wilt.     

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.     

Screening tomato genotypes for resistance and tolerance to Tomato chlorosis virus

Tomato chlorosis virus (ToCV) is an emerging crinivirus in Brazil that causes an economically important disease in tomato (Solanum lycopersicum) and other solanaceous species. ToCV is transmitted predominantly by the whitefly Bemisia tabaci Middle East‐Asia Minor 1 (MEAM1, formerly biotype B), in a semipersistent manner. As all cultivated tomato varieties and hybrids are susceptible to this crinivirus, the main alternatives for the control of the disease are the use of healthy seedlings for transplanting and the chemical control of the insect vector. The objective of this work was to evaluate the responses of tomato genotypes to infection with this crinivirus and their tolerance to the disease in order to support the development of other alternatives for disease control. Resistance to infection was evaluated by ToCV inoculation with viruliferous B. tabaciMEAM1 followed by virus detection by RT‐PCR and RT‐qPCR. To measure tolerance to the disease, plant development and fruit yield of ToCV‐infected and healthy plants were compared. Among 56 genotypes, only the lineage IAC‐CN‐RT (S. lycopersicum ‘Angela Gigante’ × S. peruvianum ‘LA 444‐1’) was highly resistant to infection with ToCV. Tolerance to the disease over two trials with different genotypes showed variable results. The effect of ToCV on plant development varied from 2.9% to 71.9% reduction, while yield loss varied from 0.2% to 51.8%. The highly ToCV‐resistant lineage IAC‐CN‐RT, which is also resistant to a Spanish isolate of ToCV, might be useful for tomato breeding programmes.     

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.     

Control of Verticillium dahliae on tomato in Tunisia by resistant cultivars and soil disinfestation1

Verticillium dahliae is an important soil-borne pathogen of vegetables in the central coastal region of Tunisia. Cultivation of resistant tomato cultivars does not always give satisfaction as they do not fulfil agronomic requirements (earliness, fruit size and yield). Soil treatment with dichloropropene/methyl isothiocyanate mixture (36.12 g + 16.45 g m^-2) gave excellent control of verticillium wilt and nearly tripled the yield of the susceptible cultivar H63–5. Dazomet (60 g m^-2) caused only slight disease depression and less yield increase. Parmi les pathogènes édaphiques des cultures maraîchères, Verticillium dahliae occupe une place importante dans le Sahel tunisien. Bien que les cultivars résistants donnent entière satisfaction sur le plan phytopathologique, leur utilisation est limitée à cause de leurs faiblesses agronomiques (précocité, calibre des fruits et rendement). La désinfection du sol avec le mélange dichloropropène/méthyl-isothiocyanate (36, 12 g + 16,45 g m^-2) réduit considérablement la verticilliose et triple presque le rendement de l'hybride sensible H63-5. L'incidence du dazomet (60 g m^-2) sur la trachéomycose et le rendement était moindre.     

Biofumigation potential of Brassicaceae cultivars to Verticillium dahliae

The biofumigation potential of brassicaceous green manures to Verticillium dahliae was systematically assessed. In a toxicological study, five different isothiocyanates (ITCs) were tested using a bioassay with sterile quartz sand artificially infested with microsclerotia. For 2-propenyl ITC, a LD₉₀ value of 88.7 nmol g^?1 was determined. Furthermore, 2-propenyl ITC with a dose of 150 nmol g^?1 soil was tested in 22 naturally infested soils. The efficiency varied from 9 % to 92 % and was negatively correlated with the organic carbon content of the soils, indicating that in many soils much higher concentrations will be necessary to achieve sufficient control. To evaluate the biofumigation potential of different Brassicaceae, 19 cultivars of Brassica juncea, Rhaphanus sativus and Sinapis alba were grown in the field. Biomass production was measured and glucosinolate concentrations were analyzed. Simulating the field situation, the biofumigation effect of cultivars was assessed in a standardized laboratory bioassay with microsclerotia-infested sterile quartz sand amended with freeze-dried ground plant tissues. Amendments of B. juncea shoot tissue reduced the number of viable microsclerotia significantly with efficiencies from 69.3 to 81.3 %. Total potentially released amounts of 2-propenyl ITC ranged between 50.6–78.1 nmol g^?1sand and indicate a clear ITC-related suppression. However these are considered too low for effective control in practice as low release efficiencies mean that effective levels fall well short of the estimated 150 nmol g^?1 of soil required. In comparison with B. juncea, the R. sativus and S. alba were less effective due to lower concentrations and/or toxicity of the ITC released. In summary, the biofumigation potential of the cultivars tested appears insufficient alone for effective control, especially on soils with higher (>1.0 %) organic carbon content.     

棉花黄萎病菌提取物对棉花的抗病效果

近年来棉花黄萎病发生严重 ,重病田病株率高达 95 %以上。目前生产上缺少抗病品种 ,防治方法不健全 ,田间防治效果不理想 ,生产上急需新的防治措施。随着环境和生态问题的日益突出 ,诱导抗病性研究日益被人们重视。本研究从诱导抗病性原理出发 ,旨在探索能够有效激发、诱导棉花抗黄萎病的因子 ,并取得一些结果 ,现报道如下。1　材料与方法1 .1　供试材料　棉种为中棉 1 6 ;棉花黄萎病菌 (Verticillium dahliae Kleb)由本室分离鉴定。1 .2　激发子的制备　将棉花黄萎病菌在马铃薯葡萄糖液体培养基中 ,5℃下恒温振荡培养两周后 ,滤纸过滤 ,滤…