Effectiveness of three Glomus species in protecting pepper (Capsicum annuum L.) against verticillium wilt

The objective of this work was to study the influence of three Glomus species—Glomus mosseae (Nicol. and Gerd.) Gerd. and Trappe, Glomus intraradices (Schenck and Smith) and Glomus deserticola (Trappe, Bloss, and Menge)—on the development of Verticillium-induced wilt in Capsicum annuum cv. Piquillo. Results showed that the effectiveness of arbuscular mycorrhizal fungi (AMF) as biocontrol agents varied among different Glomus species. In pepper colonized by G. intraradices the severity of the disease was even higher than that observed in non-mycorrhizal plants in terms of plant growth and pepper yield. On the other hand, the high effectiveness exhibited by G. mosseae in improving plant growth and the early beginning of the reproductive stage in these plants was not associated with great plant protection and high pepper yield in diseased plants. Only plants associated with G. deserticola had greater yield than non-mycorrhizal ones despite the lower P fertilization applied to the mycorrhizal treatment and this fact was observed in both healthy and diseased plants. It is suggested that the higher specific phosphorus uptake in Verticillium-inoculated plants associated with G. deserticola could contribute to diminish the deleterious effect of pathogen on yield. On the other hand, the possible influence of endogenous phenolics in roots on the tolerance or resistance of pepper against wilt induced by Verticillium dahliae remains unclear.     

Defence-related Enzymes in Pepper Roots During Interactions with Arbuscular Mycorrhizal Fungi and/or Verticillium dahliae

Previous studies have described that arbuscular mycorrhizal fungi (AMF) can reduce the deleterious effect of Verticillium dahliae Kleb. on pepper growth and yield. In mycorrhizal plants, the bioprotection against soil-borne pathogens can result from the preactivation of defence responses that include some structural modifications and the accumulation of Pathogenesis-Related (PR) proteins. Our first objective was to study if V. dahliae induced defence mechanisms in roots before infected pepper developed visible symptoms of disease. The second aim was to determine if AMF induced defence-related enzymatic activities in pepper roots before or after pathogen’s attack. Results showed that the colonization of pepper roots by Glomus deserticola (Trappe, Bloss and Menge) induced the appearance of new isoforms of acidic chitinases, superoxide dismutase (SOD) and, at early stages, peroxidases. In contrast, V. dahliae neither stimulated the phenylpropanoid pathway nor elicited hydrolytic activities in infected pepper roots. Only in mycorrhizal plants, the inoculation with V. dahliae slightly increased both phenylalanine ammonia-lyase (PAL) and peroxidase activities two weeks later. Mycorrhizal-specific induction of new isoforms of acidic chitinases and SOD together with enhanced peroxidase and PAL activities 2 weeks after pathogen inoculation could be involved in the biocontrol of Verticillium-induced wilt in pepper by AMF.     

First report of <Emphasis Type="Italic">Verticillium tricorpus</Emphasis> isolated from potato tubers in Japan

In 1998, Verticillium sp. (CE98Vt1 and CE98Vt2) were isolated from discolored vascular structures of potato tubers sold at a market in Chiba Prefecture. These isolates were identified as Verticillium tricorpus on the basis of cultural and morphological characteristics and PCR diagnosis. This observed vascular discoloration of the potato tuber was demonstrated in three cultivars (Touya, Toyoshiro, and Waseshiro) among eight cultivars by inoculation to seedlings. External and internal symptoms of these isolates were not distinct in potato plants. The virulence of these isolates to potato was very low as compared with Verticillium dahliae. These two isolates were not pathogenic to Chinese cabbage, eggplant, green pepper, larkspur, parsley, snapdragon, soybean, tobacco, and tomato. This is the first report of V. tricorpus from potato in Japan.     

根系土壤假丝酵母菌Candida sp. YIN9对大丽轮枝菌和全齿复活线虫的抑制作用

[目的] 研究黄萎病抗性棉(海 7124)根际土壤中酵母菌株对棉花黄萎病病原真菌大丽轮枝菌和全齿复活线虫的拮抗效果,为生物防治棉花黄萎病和全齿复活线虫提供理论依据。[方法] 通过镜检、糖发酵实验、碳源同化实验、26S rRNA测序对菌株的形态、生理生化特征及其系统发育关系进行鉴定,并利用七叶苷筛选、刚果红染色、平皿对峙实验、盆栽实验、平板生测实验测试其产酶活性以及抑制大丽轮枝菌和杀线虫活性。[结果] 从大批黄萎病抗性棉(海 7124)根际土壤中筛选出编号为YIN9的酵母菌菌株,分类鉴定结果表明:YIN9菌株属于假丝酵母属Candida。平皿对峙实验结果表明:菌株YIN9对大丽轮枝菌的抑菌率达59%;将菌株YIN9的无菌发酵滤液与大丽轮枝菌孢子共培养12 h后镜检发现,用菌株YIN9处理的实验组,大部分棉花黄萎病病菌孢子不能正常萌发。盆栽实验结果表明:菌株YIN9对棉花黄萎病的平均防治效果为60.02%,可以显著降低感病棉棉花黄萎病的发病率和病情指数。此外,与从黄萎病抗性棉根际土壤中筛选获得的其他酵母菌株相比,菌株YIN9具备较高的杀线虫活性:菌株作用全齿复活线虫48 和60 h后,线虫死亡率分别为90%和100%。将菌株YIN9发酵液煮沸后,其抑制大丽轮枝菌和杀线虫活性均急剧下降,进一步测试发现,该菌株拥有较高的蛋白酶和纤维素酶活性。[结论] YIN9中的生防因子可能是热不稳定性物质,具备较高的杀线虫活性,可以显著提高感病棉对黄萎病的抗性。     

Mapping of Ve in tomato: a gene conferring resistance to the broad-spectrum pathogen, Verticillium dahliae race 1

 The soil-borne fungi Verticillium spp. cause vascular wilt disease in a wide range of crop plants. In tomato, resistance to Verticillium dahliae race 1 is conferred by a single dominant gene, Ve. Previous efforts to map Ve in tomato have yielded confusing results, locating it on different chromosomes, which subsequently raised the possibility that Verticillium resistance may be controlled by a number of loci. We used three different mapping populations to obtain an unambiguous map location of Ve: a recombinant inbred (RI) line population; an F₂ population segregating for Verticillium resistance; and a population of 50 introgression lines (IL). In all of the mapping populations Ve was positioned on the short arm of chromosome 9 tightly linked to the RFLP marker GP39. This linkage was confirmed by screening for GP39 in different breeding lines with known resistance or susceptibility to Verticillium. A perfect match was found between GP39 and the Verticillium response of the lines, indicating the potential of GP39 in the rapid detection of Verticillium resistance and as a starting point for map-based cloning of Ve. This approach is particularly relevant for Verticillium dahliae race 1, since in the present work we also show that the isolate that infects tomato is responsible for wilt disease in other important crop plants. Received: 5 July 1998 / Accepted: 28 July 1998     

Moderate drought influences the effect of arbuscular mycorrhizal fungi as biocontrol agents against Verticillium-induced wilt in pepper

Previous studies have shown that the arbuscular mycorrhizal fungus (AMF) Glomus deserticola (Trappe, Bloss and Menge) can diminish the negative effect of Verticillium dahliae Kleb. on pepper yield. On the other hand, it is known that AMF can be more beneficial for plant growth and physiology under dry conditions than when soil moisture is plentiful. Therefore, our objective was to assess if a moderate water deficit imposed on pepper plants before their inoculation with V. dahliae could improve the effectiveness of G. deserticola as biocontrol agent. In the present experiment, the delay in disease development in Verticillium-inoculated plants associated with AMF did not occur under well watered conditions. In addition, the establishment of mycorrhizal symbiosis and the development of structures by AMF were delayed when both symbiotic and pathogenic fungi infected the same root. Therefore, it is suggested that the equilibrium between pepper plant, G. deserticola and V. dahliae is so complex that small changes in competition between symbiotic and pathogenic fungi for host resources can modify the efficiency of AMF as a biocontrol agent. On the other hand, water deficit enhanced the deleterious effect of V. dahliae on fruit set and yield only when pepper plants were not associated with G. deserticola, which reinforces the idea that AMF may be more important for host plants subjected to stressful conditions. However, comparing well watered non-mycorrhizal and predroughted mycorrhizal plants, we found that moderate water deficit imposed before inoculation with V. dahliae did not improve the effectiveness of G. deserticola as a biocontrol agent.     

Application of RAPD-PCR for Virulence Type Analysis within Verticillium dahliae and V. longisporum

A collection of 24 isolates of Verticillium dahliae, 11 isolates of V. longisporum and one isolate of V. albo‐atrum originating from different host plants and geographical regions was tested for genetic variability by random amplified polymorphic DNA‐polymerase chain reaction (RAPD–PCR). Based on nine primers, the three Verticillium species could be clearly differentiated. Likewise, this analysis provided a distinct separation of vegetative compatibility groups (VCG) 2B, 4A and 4B of V. dahliae by specific DNA banding patterns. Additionally, V. longisporum was found to segregate into two subgroups with only 88% similarity. This molecular‐genetic approach was used for the analysis of randomly selected Verticillium isolates from a field with high intensity of oilseed rape cultivation (33% in crop rotation). RAPD‐PCR analysis revealed that 95 of 100 isolates tested belonged to V. longisporum and 5 to VCG 2B of V. dahliae. This study demonstrates an adaptation of Verticillium soil populations to a specific cropping history.     

Transfer of fungal tolerance through interspecific somatic hybridisation between Solanum melongena and S. torvum

Tolerance to Verticillium wilt is found in a wild relative of eggplant, Solanum torvum. To transfer this tolerance to eggplant (S. melongena), protoplast fusions between eggplant and irradiated S. torvum protoplasts were performed. Putative hybrids were regenerated and evaluated for Verticillium tolerance and field value. A total of 12 plants were eventually selected to be incorporated into the breeding program, as they showed a high degree of Verticillium tolerance under field conditions combined with the morphological characters of eggplant, including normal seed set. DNA analysis showed one of these tolerant plants to have a changed pattern, with some changes corresponding to specific pattern characteristics of S. torvum. Received: 12 April 1998 / Revision received: 9 September 1998 / Accepted: 29 November 1998     

Transfer of tomato immune receptor Ve1 confers Ave1‐dependent Verticillium resistance in tobacco and cotton

Verticillium wilts caused by soilborne fungal species of the Verticillium genus are economically important plant diseases that affect a wide range of host plants and are notoriously difficult to combat. Perception of pathogen(‐induced) ligands by plant immune receptors is a key component of plant innate immunity. In tomato, race‐specific resistance to Verticillium wilt is governed by the cell surface‐localized immune receptor Ve1 through recognition of the effector protein Ave1 that is secreted by race 1 strains of Verticillium spp. It was previously demonstrated that transgenic expression of tomato Ve1 in the model plant Arabidopsis thaliana leads to Verticillium wilt resistance. Here, we investigated whether tomato Ve1 can confer Verticillium resistance when expressed in the crop species tobacco (Nicotiana tabcum) and cotton (Gossypium hirsutum). We show that transgenic tobacco and cotton plants constitutively expressing tomato Ve1 exhibit enhanced resistance against Verticillium wilt in an Ave1‐dependent manner. Thus, we demonstrate that the functionality of tomato Ve1 in Verticillium wilt resistance through recognition of the Verticillium effector Ave1 is retained after transfer to tobacco and cotton, implying that the Ve1‐mediated immune signalling pathway is evolutionary conserved across these plant species. Moreover, our results suggest that transfer of tomato Ve1 across sexually incompatible plant species can be exploited in breeding programmes to engineer Verticillium wilt resistance.     

Effects of earthworms, nematodes, cultivations and host plants on Verticillium wilt of peach and cherry

Verticillium dahliae but not V. albo-atrum Berth was isolated from eight out of twenty-one stone fruit orchards surveyed for Verticillium wilt disease in western New York. Wilt incidence was related to the cultivation of tomato or legumes as previous or inter-crop with stone fruit trees. A limited cross species inoculation using isolates of V. albo-atrum and V. dahliae from woody and herbaceous plants showed that peach and cherry were susceptible to both species. The effect of V. dahliae on growth of cherry seedlings in the presence of Tylenchorynchus claytoni, Pratylenchus penetrans and Meloidogyne hapla was compared. P. penetrans and M. hapla produced more severe growth reduction than T. claytoni. The adverse effect of Verticillium on the growth of cherry seedlings was greater acting together with any one of the three nematodes than acting alone. V. dahliae was shown to be capable of passage through earthworms without loss of infectivity.     

Su Un Verticillium Parassita Del Cronartium Asclepiadeum

Summary

AA. observed that many aecia of Cronartium asclepiadeum on branches of Pinus nigra were covered by a snow-white, felt-like mycelium of a Verticillium which girled and entered the aecioconidia.

Comparative studies whit several species of Verticillium grown in identical conditions, confirmed by the analysis of variance of the lenght of the conidia, showed that the species isolated from Cronartium asclepiadeum was identical to V. coccorum. To the same species AA. refer V. Hemileiae Bouriq.

Inoculations experiments carried in watch-glasses showed that V. coccorum can also parasitize the uredoconidia of some other rustes.     

MORPHOGENESIS IN VERTICILLIUM: A SELF-INDUCED,NON-HEREDITARY VARIATION IN COLONY FORM

Some homokaryotic isolates of Verticillium albo-atrum (microsclerotial form) vary radically in colony appearance even when temperature, lighting, humidity and medium content are held constant. This variation is so great that it may easily be mistaken for hereditary variation. The environmental factors which control the variation are differences in shape of medium on which the colonies grow and presence or absence of other Verticillium colonies. The cause of the variation appears to be a diffusible substance which Verticillium itself produces.     

Genetic transformation of cotton with a harpin-encoding gene <Emphasis Type="Italic">hpa</Emphasis><Subscript><Emphasis Type="Italic">Xoo</Emphasis></Subscript> confers an enhanced defense response against different pathogens through a priming mechanism

Background  

The soil-borne fungal pathogen Verticillium dahliae Kleb causes Verticillium wilt in a wide range of crops including cotton (Gossypium hirsutum). To date, most upland cotton varieties are susceptible to V. dahliae and the breeding for cotton varieties with the resistance to Verticillium wilt has not been successful.     

Identification of Cauliflower Cultivars that Differ in Susceptibility to Verticillium longisporum using Different Inoculation Methods

The response of 13 European cauliflower cultivars to Verticillium longisporum was evaluated using two greenhouse tests and one in vitro inoculation test. The greenhouse tests involved dipping roots of 3‐week‐old seedlings in a conidial suspension or inoculating the soil of 3‐week‐old seedlings with Verticillium microsclerotia. The in vitro test involved the inoculation of 9‐day‐old seedlings with Verticillium conidia. Useful disease parameters were the area under disease progress curve and plant growth reduction for the greenhouse tests and fresh weight reduction for the in vitro test. Significant correlations were found among the three inoculation methods. Irrespective of the inoculation method used, cultivar ‘Sernio’ was most resistant to V. longisporum, while ‘Minaret’ was the most susceptible cultivar. The pathogen could be re‐isolated from the hypocotyls and from the stem of ‘Minaret’ 4 and 49 days after inoculation respectively, whereas V. longisporum could never be re‐isolated from ‘Sernio’. These results suggest that the more resistant cauliflower cultivar ‘Sernio’ can suppress the ascent and the proliferation of V. longisporum into the plant.     

Genetic and environmental control of the <Emphasis Type="Italic">Verticillium</Emphasis> syndrome in <Emphasis Type="Italic">Arabidopsis thaliana</Emphasis>

Background  

Verticillium spp. are major pathogens of dicotyledonous plants such as cotton, tomato, olive or oilseed rape. Verticillium symptoms are often ambiguous and influenced by development and environment. The aim of the present study was to define disease and resistance traits of the complex Verticillium longisporum syndrome in Arabidopsis thaliana (L.) Heynh. A genetic approach was used to determine genetic, developmental and environmental factors controlling specific disease and resistance traits and to study their interrelations.     

The symptoms and causal agents of early-dying disease (Verticillium wilt) of potatoes

The only characteristic symptom produced by Verticillium albo-atrum and V. dahliae in infected potato plants is unilateral chlorosis and necrosis: this was not shown until the approach of host maturity, and was distinguishable from symptoms of natural senescence only in its slightly earlier expression. Of six species of Verticillium tested against potato (King Edward), V. albo-atrum, V. dahliae, V. nigrescens and V. nubilum were pathogenic (all produced ‘wilt’ symptoms and relative virulence was in that decreasing order) but V. tricorpus and V. lateritium did not induce disease. Isolates of V. albo-atrum and V. dahliae, obtained from a number of other host plants, were also pathogenic to potato. Possible reasons are given for the fewness of records of ‘Early dying’ disease (Verticillium wilt) of potatoes in the field.     

Verticase： a Fibrinolytic Enzyme Produced by Verticillium sp, Tj33, an Endophyte of Trachelospermum jasminoides

Plant endophytes are among the most important resources of biologically active metabolites. Twenty-three endophyte strains residing in Trachelospermum jasminoides were cultivated in vitro with the cultures assayed for the fibrinolytic substance production. As a result, the culture of VerticUlium sp. Tj33 was shown to be the most active. A fibrinolytic enzyme designated as verticase was subsequently purified from the supernatant of Verticillium sp. culture broth by a combination of DEAE-52, Sephadex G-75 and hydrophobic column chromatographies. Verticase, with its molecular mass of 31 kDa and pl of 8.5, was demonstrated to be homogeneous by sodium dodecyl suIfate-polyacrylamide gel electrophoresis and isoelectric focusing electrophoresis. Verticase is an enzyme that hydrolyzes fibrin directly without activation of plaminogen. It was stable in a broad pH range from 4 through to 11 with the optimal reaction pH value and temperature shown to be around 9-10 and 50-60℃, respectively. The fibrinolytic activity of verticase was severely inhibited by phenylmethylsulfony fluoride, indicating that verticase was a serine protease.     

黄萎病不同发生程度棉田中土壤微生物多样性

作物根际土壤微生物群落对土壤生态及作物健康至关重要。以棉花黄萎病不同发生程度棉田的土壤为研究对象,采用理化分析、微生物纯培养及高通量测序技术对土壤理化性质及微生物数量、细菌丰度多样性进行综合分析。结果表明:在纯培养条件下,大丽轮枝菌无菌发酵滤液对细菌生长有明显的抑制作用;棉田接种大丽轮枝菌对土壤中细菌、真菌、放线菌的数量及细菌菌群丰度多样性未产生明显影响,不同采样时间的土壤中细菌菌群结构差异更大。土壤中大丽轮枝菌微菌核数量与棉花黄萎病的发生程度呈显著正相关。土壤肥力对土壤中微生物数量起主导作用,而水稻-棉花轮作能够使棉田有效降盐、减病、改善土壤肥力。通过生物防治、作物轮作、深翻等调控措施增加土壤中有益菌群数量、改善土壤生态、降低棉田土壤中大丽轮枝菌菌源数量是减轻棉花黄萎病危害的基础。     

Screening and Identification of Endomannanase-Producing Microfungi from Hypersaline Environments

A culture-dependent enrichment technique was used to isolate endo-1,4-β-mannanase–producing fungi from a hypersaline environment. Galactomannan was used as carbon source and resulted in isolation of strains of Scopulariopsis brevicaulis, S. candida, and Verticillium dahliae. The Scopulariopsis isolates were found to be more dominant and could be isolated from consecutive evaporation ponds, whereas Verticillium was only isolated from one pond. The Scopulariopsis strains exhibited only endomannanase activity, whereas Verticillium displayed broad-activity spectrum by secreting endoxylanases and cellulases in addition to endomannanases. S. candida LMK004 and LMK008 produced 7420 and 14750 nkat g⁻¹ biomass, respectively. Endomannanase production in these strains increased with an increase in NaCl concentration up to 10% (w/v), after which both growth and enzyme production was decreased. V. dahliae LMK006 grew and produced up to 5000 nkat g⁻¹ biomass endomannanase in the absence of NaCl. Increased NaCl concentration had a negative effect on this strain. S. brevicaulis LMK002 showed poor endomannanase production but a similar growth trend as the other Scopulariopsis strains. In general, the Scopulariopsis strains exhibited better halotolerance than V. dahliae and could grow in the presence of 20% NaCl on solid medium.