Identification of differential resistance to six Fusarium oxysporum f. sp. cepae isolates in commercial onion cultivars through the development of a rapid seedling assay

Fusarium oxysporum isolates collected from onions in the UK and other countries were characterized using sequences of the transfer elongation factor 1‐α (TEF) gene and compared with published sequence data for 10 other isolates. Isolates associated with diseased onion bulbs in the UK formed two clades. Isolates from both clades were selected for pathogenicity testing and to develop a rapid seedling assay to screen commercial onion cultivars for resistance to F. oxysporum f. sp. cepae (FOC), the cause of basal rot. Differences in the levels of aggressiveness between isolates were observed and isolates from both clades were pathogenic. Differences in resistance/susceptibility were also observed amongst 10 commercial onion cultivars, with cvs Ailsa Craig Prizewinner and White Lisbon showing the highest levels of resistance. The results from the seedling assay were supported by those from a subsequent onion bulb rot assay. Thus, this study reports the development of a rapid, simple and repeatable seedling assay that can be used to screen large numbers of onion cultivars for resistance to FOC and which is indicative of resistance at the bulb stage.     

Fusarium oxysporum,F. proliferatum and F. redolens associated with basal rot of onion in Finland

In recent years in Finland, Fusarium infections in onions have increased, both in the field and in storage, and Fusarium species have taken the place of Botrytis as the worst pathogens causing post‐harvest rot of onion. To study Fusarium occurrence, samples were taken from onion sets, harvested onions and also from other plants grown in the onion fields. Isolates of five Fusarium species found in the survey were tested for pathogenicity on onion. Fusarium oxysporum was frequently found in onions and other plants, and, of the isolates tested, 31% caused disease symptoms and 15% caused growth stunting in onion seedlings. Fusarium proliferatum, a species previously not reported in Finland, was also identified. Over 50% of the diseased onion crop samples were infected with F. proliferatum, and all the F. proliferatum isolates tested were pathogenic to onion. Thus, compared to F. oxysporum, F. proliferatum seems to be more aggressive on onion. Also some of the F. redolens isolates were highly virulent, killing onion seedlings. Comparison of the translation elongation factor 1α gene sequences revealed that the majority of the aggressive isolates of F. oxysporum f. sp. cepae group together and are distinct from the other isolates. Incidence and relative proportions of the different Fusarium species differed between the sets and the mature bulbs. More research is required to determine to what extent Fusarium infections spoiling onions originate from infected onion sets rather than the field soil.     

Characterization of <Emphasis Type="Italic">Trichoderma</Emphasis> species isolated in Ecuador and their antagonistic activities against phytopathogenic fungi from Ecuador and Japan

Native Trichoderma spp. were isolated from agricultural fields in several regions of Ecuador. These isolates were characterized via morphological observation as well as molecular phylogenetic analysis based on DNA sequences of the rDNA internal transcribed spacer region, elongation factor-1α gene and RNA polymerase subunit II gene. Fifteen native Trichoderma spp. were identified as T. harzianum, T. asperellum, T. virens and T. reesei. Some of these strains showed strong antagonistic activities against several important pathogens in Ecuador, such as Fusarium oxysporum f. sp. cubense (Panama disease) and Mycosphaerella fijiensis (black Sigatoka) on banana, as well as Moniliophthora roreri (frosty pod rot) and Moniliophthora perniciosa (witches’ broom disease) on cacao. The isolates also showed inhibitory effects on in vitro colony growth tests against Japanese isolates of Fusarium oxysporum f. sp. lycopersici, Alternaria alternata and Rosellinia necatrix. The native Trichoderma strains characterized here are potential biocontrol agents against important pathogens of banana and cacao in Ecuador.     

Response of different intermediate-day onion hybrids to natural infestation by Phoma terrestris and Fusarium oxysporum f. sp. cepae in Ciudad Real, Spain with assessment of different soil disinfestation methods

Incidence and severity of Pink root caused by Phoma terrestris and Fusarium basal rot caused by Fusarium oxysporum f. sp. cepae were field-tested on factorial combinations of different soil fumigation treatments in five different intermediate-day onion hybrids widely adapted at the Bola?os de Calatrava area (Ciudad Real, Spain). The effect of Dazomet and virulence of the local isolates were then tested in trays under greenhouse conditions. When compared to Electric, the other four onion hybrids showed a certain degree of tolerance to Pink root and Fusarium basal rot with little difference among them. The field trial carried out in 2006 showed significant but negligible beneficial effects in medium bulb weight and Pink root severity due to Dazomet. The field trial carried out in 2007 showed significant beneficial effects of Metham-sodium on plant growth and Pink root and Fusarium basal rot severities. Sodium tetrathiocarbonate performed worse than the control. Greenhouse test showed that Dazomet performed nearly as well as sterilization under controlled conditions. Virulence of local isolates was lower than that of reference isolates.     

香蕉枯萎病拮抗菌筛选及其抑菌活性

为获得对香蕉枯萎病菌具有稳定拮抗作用的微生物菌株,分别从海南省临高县南宝镇新营农场、皇桐村及美台镇美梅村采集感病土壤和健康土壤进行微生物分离,并以香蕉枯萎病菌4号小种(Fusarium oxysporum f.sp.cubense Race 4)等9种病原菌为靶标菌株,采用平板对峙法和含药介质法对拮抗菌的抑菌活性进行评价,同时根据形态特征、培养特征及生理生化特征和16S rDNA序列分析对其进行鉴定。结果表明:6个土壤样品中共分离得到具有拮抗作用的细菌93株和放线菌133株,其中T3-G-59菌株对9种病原菌具有广谱抗性,菌丝生长抑制率为43.58%~86.43%,孢子萌发抑制率为42.22%~81.11%,其中对香蕉枯萎病菌4号小种的菌丝生长和孢子萌发抑制率分别为86.43%和81.00%,经鉴定该菌株为多产色链霉菌Streptomyces polychromogenes,表明该多产色链霉菌T3-G-59菌株具有应用于香蕉枯萎病生物防治的潜力。     

Response of transgenic cucumber expressing a rice class I chitinase gene to two fungal pathogens with different infectivities

A transgenic cucumber line (CR32) over-expressing the rice class I chitinase gene exhibited resistance to Phytophthora rot (Phytophthora nicotianae var. parasitica) but not to Fusarium wilt (Fusarium oxysporum f. sp. cucumerinum). The infection behavior of these fungi on CR32 and nontransgenic plants was examined with an optical microscope. In zoosporangia of P. nicotianae var. parasitica, the rates of germination and penetration on leaves of both CR32 and nontransgenic plants were almost equal. After infection, however, the growth of infection hyphae was markedly suppressed in CR32 compared with their growth in the nontransgenic plants. In F. oxysporum f. sp. cucumerinum, the infection hyphae localized in petiole vessels of both CR32 and nontransgenic plants, and growth did not differ in the two plants. We investigated the antifungal activity of a high-molecular-weight fraction (HF) and a low-molecular-weight fraction (LF) of crude leaf extracts from CR32 and from the nontransgenic line. CR32 HF, which included the rice chitinase, had antifungal activity only against F. oxysporum f. sp. cucumerinum. In contrast, CR32 LF, which did not have the rice chitinase, had strong antifungal activity against the two fungi. These results suggested that a low-molecular-weight antifungal substance(s) was induced in CR32 and might function as a factor of resistance to P. nicotianae var. parasitica, which has cell walls that almost never contain chitin. Because rice chitinase has already been demonstrated not to localize in vessels of CR32, the infection localization of F. oxysporum f. sp. cucumerinum in vessels might enable the fungus to avoid antifungal substance(s), resulting in Fusarium wilt in transgenic cucumber.     

Resistance to fusarium basal rot of onion in greenhouse and field and associated expression of antifungal compounds

Greenhouse and field evaluations of onion for resistance to Fusarium basal rot caused byFusarium oxysporum f.sp.cepae were conducted on cultivars ‘Akgün 12’ and ‘Rossa Savonese’ previously described as resistant at the seedling stage. In the greenhouse experiments inoculations were carried out on seeds or soil; in the field experiments evaluation was performed on onion sets from plants grown in naturally infested soils. Akgün 12 and to a lesser extent Rossa Savonese were resistant to the disease at the bulb stage in all experiments. Results were also consistent with those obtained from a previous screening at the seedling stage. Onion sets were also extracted and fractionated by thin layer chromatography to determine their content of antifungal compounds. Extracts were characterized by the expression of distinct antifungal components, which may be involved in resistance to the pathogen. http://www.phytoparasitica.org posting July 14, 2004.     

Unraveling the multilevel aspects of least explored plant beneficial <Emphasis Type="Italic">Trichoderma saturnisporum</Emphasis> isolate GITX-Panog (C)

Species of Trichoderma offer an eco-friendly and economical solution to the management of plant pathogens in agricultural crops. The secretion and spectrum of different bioactive molecules by Trichoderma spp. is known to follow adaptive and species/strain behavior, depending upon the edaphic conditions. In present study, GITX-Panog (C), an antagonistic isolate of Trichoderma saturnisporum against Fusarium oxysporum was screened for the production of different glycosyl hydrolases and secondary metabolites. The evaluation of antagonistic isolates for lytic enzymes and secondary metabolites revealed their significance as plant probiotics. Profiling of lytic enzymes such as chitinases and β-glucanases secreted by T. saturnisporum GITX-Panog (C) in the presence of autoclaved mycelium of F. oxysporum and colloidal chitin or laminarin showed increased enzyme activities. Production of bioactive secondary metabolites such as siderophore, peptaibols, 6-pentyl-α-pyrone, and the known potential of these compounds in mineral acquisition and suppression of plant F. oxysporum, revealed the potential significance of Trichoderma saturnisporum isolate GITX-Panog C in agricultural application against biotic and abiotic stresses. The antagonistic isolate also showed significant increases in fresh biomass and seed production in Arabidopsis thaliana in the greenhouse compared to controls. Thus, the current study highlights the multiple attributes of an antagonistic isolate of T. saturnisporum for future agricultural applications as a plant probiotic agent.     

Auftreten und Nachweis von Fusarium oxysporum und F. proliferatum an Steck- und Saatzwiebeln

The pathogen Fusarium oxysporum f. sp. cepae inducing the Fusarium basal rot mainly spreads in warmer cultivation regions due to its adaptibility to high temperature. Meanwhile the pathogen occurs in Germany as well, especially in years with relatively high average temperature during the growing season. Phytopathological investigations of 300 symptomless onion bulbs showed a contamination rate of approximately 10% with regard to Fusarium spp, with F.?oxysporum proving to be the predominant species. Onion sets planted in these fields were latently infected with F.?oxysporum at rates of 19?C98%. Unexpectedly, the contaminated sets did not indispensably lead to a high occurrence of plants exhibiting characteristic symptoms of Fusarium basal rot such as wet and dry rot. Presumably, the development of symptoms is particularly affected by given climatic conditions. The results of pathogenicity tests of isolated Fusarium spp. isolates under controlled conditions support this assumption. The inoculation of the substrate with selected Fusarium spp. isolates resulted in a reduction of emergence by up to 70% under controlled conditions, which are suboptimal with regard to the cultivation of onions. The emergence of plants was not affected by Fusarium spp. under optimal cultivation of onions. However, under optimal cultivation conditions a reduction of plant growth occurred in a subsequent growth stage. Beside F.?oxysporum, F.?proliferatum could be detected in onion bulbs as well as seeds. The proportion of contaminated seeds accounted to 62%. Both species F.?oxysporum and F.?proliferatum proved to be pathogenic in onion although their isolates varied much in their virulence.     

Biological control of fusarium wilt of carnations with different Fusarium oxysporum strains1

Three strains of Fusarium oxysporum were tested for their antagonism to Fusarium oxysporum f.sp. dianthi, two from alluvial soils of the Durance valley (Provence, FR) known to be suppressive to F.o. f. sp. melonis (strains 47 and 20 of Alabouvette) and a third from carnation growing medium, in which it spontaneously appeared (TF3). Strains 47 and 20 were introduced into perlite before planting carnations and found to have variable efficacy against fusarium wilt, according to the formulation of the inoculum. With a talc preparation, these strains gave only 15% witted plants, compared with 49% in the control. With a bran-perlite inoculum, only strain TF3 gave better results than the control. Repeated introduction of TF3 through the season increased the incidence of disease, whereas a single treatment before planting in perlite was effective whatever the formulation (humid bran-perlite, bran-perlite powder, talc, liquid). However, the antagonistic activity of TF3 was not sufficient to control the disease in a naturally infested soil.     

Changes induced by Trichoderma harzianum in suppressive compost controlling Fusarium wilt

The addition of species of Trichoderma to compost is a widespread technique used to control different plant diseases. The biological control activity of these species is mainly attributable to a combination of several mechanisms of action, which may affect the microbiota involved in the suppressiveness of compost. This study was therefore performed to determine the effect of inoculation of Trichoderma harzianum (T. harzianum) on compost, focusing on bacterial community structure (16S rRNA) and chitinase gene diversity. In addition, the ability of vineyard pruning waste compost, amended (GCTh) or not (GC) with T. harzianum, to suppress Fusarium wilt was evaluated. The addition of T. harzianum resulted in a high relative abundance of certain chitinolytic bacteria as well as in remarkable protection against Fusarium oxysporum comparable to that induced by compost GC. Moreover, variations in the abiotic characteristics of the media, such as pH, C, N and iron levels, were observed. Despite the lower diversity of chitinolytic bacteria found in GCTh, the high relative abundance of Streptomyces spp. may be involved in the suppressiveness of this growing media. The higher degree of compost suppressiveness achieved after the addition of T. harzianum may be due not only to its biocontrol ability, but also to changes promoted in both abiotic and biotic characteristics of the growing media.     

Effect of some organic amendments on onion bulb rot

The effect of soil amended with stalks of sunflower (Helianthus annuus L.), alfalfa (Medicago sativa L.) and Hungarian vetch (Vicia pannonica Crantz) on onion bulb rot caused byFusarium oxysporum Schlecht. f.sp.cepae (Hanz.) Snyder Hansen andAspergillus niger v. Tieghem was tested in a soil naturally infested with the pathogens under field conditions. All treatments significantly reduced bulb rot incidence. The fungal and antibacterial populations varied among all of the amended tested soils. Heterotrophic fungal populations were more effective than heterotrophic bacterial populations in the suppression of disease. The highest population of fungal microflora was found in plots amended with sunflower stalks. The carbon-to-nitrogen ratio and available moisture content of amended soils have an indirect suppressive role on disease, by increasing the heterotrophic fungal population. http://www.phytoparasitica.org posting June 18, 2002.     

Physiological races and vegetative compatibility groups within Fusarium oxysporum f.sp. gladioli

The pathogenicity and vegetative compatibility of mainly Dutch isolates ofFusarium oxysporum collected from diseased gladioli and other Iridaceae were investigated. Based on their pathogenicity to two differential gladiolus cultivars, the isolates could tentatively be divided into two races. All self-compatible isolates ofFusarium oxysporum f.sp.gladioli belonged to one of three distinct vegetative compatibility groups, VCG 0340, 0341 or 0342, and were incompatible with isolates that were not pathogenic to gladiolus. Isolates of one of the two races were restricted to one VCG while isolates of the other race were present in all three VCGs.     

Phylogenetic position and nucleotide diversity of Fusarium oxysporum f. sp. vanillae worldwide based on translation elongation factor 1α sequences

Fusarium oxysporum f. sp. vanillae is considered the most important fungus affecting vanilla crops around the world, causing rot on vanilla roots and stems. Previous studies showed that the ability to infect vanilla plants is a polyphyletic trait among strains of the Fusarium oxysporum species complex (FOSC). The same studies proposed a single origin for F. oxysporum f. sp. vanillae isolates sampled from Mexico, the centre of origin and distribution of vanilla. The aim of this work was to test the hypothesis of the monophyletic origin of a wider sample of isolates of F. oxysporum f. sp. vanillae infecting Mexican vanilla and estimate nucleotide diversity of pathogen isolates from the main vanilla‐producing countries. Sequence data for the TEF1α gene from 106 isolates was assembled. The phylogenetic analyses suggest that some Mexican isolates of F. oxysporum f. sp. vanillae belong in two well‐supported clades, mixed with isolates from Madagascar, Indonesia, Réunion and Comoros. The phylogenetic position of other Indonesian and Mexican isolates is unresolved. Estimations of nucleotide diversity showed that the population from Mexico is genetically more diverse than the other three populations from Madagascar, Indonesia and Réunion. The results support a polyphyletic origin of vanilla‐infecting isolates of F. oxysporum worldwide, and also reject the proposition that Mexican isolates have a single origin. The phylogenetic optimizations over the strict consensus tree of the ability to infect vanilla plants suggest that pathogenic strains around the world are the product of multiple shifts of pathogenesis and dispersion events.     

Genetic diversity and pathogenicity of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> isolated from wilted Welsh onion in Japan

Thirty isolates of Fusarium oxysporum from wilted Welsh onion plants were examined for their diversity in nucleotide sequences of the ribosomal DNA (rDNA) intergenic spacer (IGS) region and for pathogenicity with regard to five Welsh onion cultivars. Phylogenetic analysis based on the IGS sequences revealed polyphyletic origins of the isolates and a relationship between phylogeny and pathogenicity; low virulence isolates differed genetically from those with high and moderate virulence. Mating type analysis revealed that all F. oxysporum isolates were MAT1-1 idiomorphs, suggesting that the pathogens may be clonal in the fields examined.     

四季豆枯萎病病原鉴定及防治

从南宁市郊11个病区采集的四季豆枯萎病株标样,经分离培养鉴定和致病性测定,证明其病原菌为尖孢镰刀菌菜豆专化型(Fusarium oxysporum f. sp. phaseoli Kend & Syd)。此病在南宁于4月上中旬四季豆初花期开始发生,5月中下旬盛花至结荚期为发病高峰期。用滤纸碟法进行药效试验的结果,40%灭病威300-500倍液的抑菌圈最大,田间灌根防治也有一定效果。可用种子重量的0.5%多菌灵可湿性粉拌种。品种间抗病性有显著差异,秋抗19号和秋抗6号较抗病。     

Seed transmission of<Emphasis Type="Italic">Fusarium oxysporum</Emphasis> f.sp.<Emphasis Type="Italic">lactucae</Emphasis>

Twenty-seven seed samples belonging to the lettuce cultivars most frequently grown in Lombardy (northwestern Italy), in an area severely affected by Fusarium wilt of lettuce, were assayed for the presence ofFusarium oxysporum on a Fusarium-selective medium. Isolations were carried out on subsamples of seeds (500 to 1500) belonging to the same seed lots used for sowing, and either unwashed or disinfected in 1% sodium hypochloride. The pathogenicity of the isolates ofF. oxysporum obtained was tested in four trials carried out on lettuce cultivars of the butterhead type, very susceptible to Fusarium wilt. Nine of the 27 samples of seeds obtained from commercial seed lots used for sowing in fields affected by Fusarium wilt were contaminated byF. oxysporum. Among the 16 isolates ofF. oxysporum obtained, only one was isolated from disinfected seeds. Three of the isolates were pathogenic on the tested cultivars of lettuce, exhibiting a level of pathogenicity similar to that of the isolates ofF. oxysporum f.sp.lactucae obtained from infected wilted plants in Italy, USA and Taiwan, used as comparison. The results obtained indicate that lettuce seeds are a potential source of inoculum for Fusarium wilt of lettuce. The possibility of isolatingF. oxysporum f.sp.lactucae, although from a low percent of seeds, supports the hypothesis that the rapid spread of Fusarium wilt of lettuce observed recently in Italy is due to the use of infected propagation material. Measures for prevention and control of the disease are discussed. http://www.phytoparasitica.org posting Dec. 16, 2003.     

Molecular cloning of two chitinase genes from Serratia marcescens and their expression in Pseudomonas species

Two chitinase encoding EcoRI fragments from the enteric soil bacterium Serratia marcescens were cloned. From a genomic library of 5686 transductants, 21 expressed chitinase activity as indicated by clearing of a chitin-containing medium. The chitinase encoding clones could be divided into two groups. Four had an 18kb EcoRI fragment and 17 had a 9·4 kb EcoRI fragment. In Southern hybridization experiments the 18kb fragment showed no homology to the 9·4 kb fragment and restriction enzyme maps indicated no similarity. Triparental mating with fluorescent Pseudomonas spp. yielded transconjugants that expressed chitinase activity, inhibited growth of Fusarium oxysporum f. sp. redolens germ tubes and reduced disease of radish caused by the same fungus.     

Vegetative Compatibility Grouping of Fusarium oxysporum f. sp. gladioli from Saffron

Fusarium corm rot of saffron (Crocus sativus L.), incited by Fusarium oxysporum f. sp. gladioli, causes severe yield losses in Italy. Major symptoms during flowering (October–November) include yellowing and wilting of shoots, basal stem rot and corm rot. Sixty-four isolates of F. oxysporum f. sp. gladioli, obtained from infected saffron crops located in Italy (Abruzzi, Tuscany and Umbria) and in Spain, were characterized by pathogenicity and vegetative compatibility. Chlorate-resistant, nitrate-nonutilizing (nit) mutants were used to determine vegetative compatibility among the isolates of the pathogen with the aim of examining the genetic relatedness among populations from different locations. All the isolates belonged to vegetative compatibility group 0340. Since saffron shares susceptibility to F. oxysporum f. sp. gladioli with other ornamental plants of the Iridaceae (Crocus, Gladiolus, Iris and Ixia), it is likely that a clone of the pathogen (VCG 0340) was introduced with other hosts and is responsible for the disease outbreak observed on saffron in Italy. Alternatively, or additionally, the clone of F. oxysporum f. sp. gladioli causing disease on saffron in other countries may have spread to the saffron fields in Italy through the import and dispersal of infested propagation material.     

Sequence variation in the putative effector gene SIX8 facilitates molecular differentiation of Fusarium oxysporum f. sp. cubense

Fusarium oxysporum f. sp. cubense (Foc), causal agent of fusarium wilt of banana, is among the most destructive pathogens of banana and plantain. The development of a molecular diagnostic capable of reliably distinguishing between the various races of the pathogen is of key importance to disease management. However, attempts to distinguish isolates using the standard molecular loci typically used for fungal phylogenetics have been complicated by a poor correlation between phylogeny and pathogenicity. Among the available alternative loci are several putative effector genes, known as SIX genes, which have been successfully used to differentiate the three races of F. oxysporum f. sp. lycopersici. In this study, an international collection of Foc isolates was screened for the presence of the putative effector SIX8. Using a PCR and sequencing approach, variation in Foc‐SIX8 was identified which allowed race 4 to be differentiated from race 1 and 2 isolates, and tropical and subtropical race 4 isolates to be distinguished from one another.