Colonization of lettuce cultivars and rotation crops by Fusarium oxysporum f. sp. lactucae,the cause of fusarium wilt of lettuce

The severity of fusarium wilt is affected by inoculum density in soil, which is expected to decline during intervals when a non‐susceptible crop is grown. However, the anticipated benefits of crop rotation may not be realized if the pathogen can colonize and produce inoculum on a resistant cultivar or rotation crop. The present study documented colonization of roots of broccoli, cauliflower and spinach by Fusarium oxysporum f. sp. lactucae, the cause of fusarium wilt of lettuce. The frequency of infection was significantly lower on all three rotation crops than on a susceptible lettuce cultivar, and the pathogen was restricted to the cortex of roots of broccoli. However, F. oxysporum f. sp. lactucae was isolated from the root vascular stele of 7·4% of cauliflower plants and 50% of spinach plants that were sampled, indicating a greater potential for colonization and production of inoculum on these crops. The pathogen was also recovered from the root vascular stele of five fusarium wilt‐resistant lettuce cultivars. Thus, disease‐resistant plants may support growth of the pathogen and thereby contribute to an increase in soil inoculum density. Cultivars that were indistinguishable based on above‐ground symptoms, differed significantly in the extent to which they were colonized by F. oxysporum f. sp. lactucae. Less extensively colonized cultivars may prove to be superior sources of resistance to fusarium wilt for use in breeding programmes.     

A new race of Fusarium oxysporum f. sp. lactucae of lettuce

Fusarium oxysporum f. sp. lactucae, the causal agent of fusarium wilt of lettuce (Lactuca sativa), occurs in most countries in which lettuce is grown and causes serious economic losses. Three races (1, 2 and 3) of the pathogen have previously been identified on the basis of their ability to cause disease on differential lettuce cultivars, as well as by means of molecular tools developed to characterize different races of this pathogen. Only race 1 has been detected in Europe so far. In this study, two isolates of F. oxysporum, obtained from lettuce plants grown in the Netherlands showing symptoms of wilt, have been characterized by combining the study of pathogenicity with differential cultivars of lettuce and molecular assays to determine whether the isolates are different from the known races of F. oxysporum f. sp. lactucae. This study reports the presence of F. oxysporum f. sp. lactucae for the first time in the Netherlands. The causal pathogen has been identified, using the IRAP‐SCAR technique, as a new race of F. oxysporum f. sp. lactucae. Specific primers have been designed to identify this new race.     

Pathogenicity Groups in Fusarium oxysporum f. sp. lactucae on Horticultural Types of Lettuce Cultivars

Isolates of Fusarium oxysporum f. sp. lactucae obtained from six localities in Japan were divided into three patho-genicity groups. Group 1 was highly pathogenic to lettuce cultivars of crisphead and red leaf types and was less pathogenic to butterhead and green leaf type cultivars. Group 2 was highly pathogenic to butterhead type and less pathogenic to crisphead and leaf types. Group 3 was less pathogenic to all lettuce types than groups 1 and 2. These results indicated pathogenic differentiation in F. oxysporum f. sp. lactucae, roughly relating to horticultural types of host lettuce cultivars. Received 21 February 2001/ Accepted in revised form 28 May 2001     

Rapid detection of Fusarium oxysporum f. sp. lactucae on soil,lettuce seeds and plants using loop‐mediated isothermal amplification

Fusarium oxysporum f. sp. lactucae (FOL) is a soil‐ and seedborne pathogen and the causal agent of fusarium wilt on lettuce. Four races have been identified within FOL, with different worldwide distribution. Several molecular techniques have been used to detect and identify this pathogen; however, not all of them have the optimal characteristics in terms of sensitivity to perform FOL detection in plant and seed material. A loop‐mediated isothermal amplification (LAMP) assay was developed based on the sequence‐characterized amplified region (SCAR) obtained in a previous rapid amplification of polymorphic DNA (RAPD) study. The LAMP assay has been validated according to the EPPO standard PM7/98. The LAMP assay was tested with lettuce seeds, soil and plant material, and can be used successfully to amplify DNA from each of these matrices. In seed lots artificially inoculated with FOL, the detection limit of the LAMP test was 0.004% infected seed.     

Stem canker and wilt of delphinium caused by Fusarium oxysporum f. sp. delphinii in Japan

Stem canker and severe wilt were observed on delphinium plants (Delphinium elatum) in Aomori Prefecture, Japan, in 2008. The fungus isolated from the diseased crown was identified as Fusarium oxysporum f. sp. delphinii on the basis of morphological characteristics, nucleotide sequences, and host range. The isolate induced similar stem canker and wilt symptoms in inoculated delphinium plants. We propose the name “stem canker and wilt” for the disease.     

Response of carnation cultivars to Fusarium oxysporum f.sp. dianthi in the field

The response of carnation (Dianthus caryophyllus L.) cultivars toFusarium oxysporum f.sp.dianthi (F.o.d.) was evaluated in an artificially infested field from 1988/9 to 1991/2. Disease incidence was highly correlated with disease severity, indicating that disease incidence may be used to estimate the impact ofF.o.d. on the host. Based on the results, the following stepwise procedure was developed for characterizing the response of carnation cultivars toF.o.d. First, the general response of the tested cultivar was classified as resistant or susceptible on the basis of disease incidence values recorded 180 days after planting. Empirical analysis of the data revealed that a disease incidence level of 75% may be taken as a reliable cut-off point for separation of cultivars into the two groups. Within each group, cultivars were then subjected to a more explicit classification: in the resistant group the records of actual disease incidence were used for classification, while in the susceptible group the linear rearrangement of the disease progress curve was calculated according to the Gompertz function, and the value of the intercept was used for classification.Contribution No. 3539-E series, from the Agricultural Research Organization.     

Physiological races and vegetative compatibility groups of butterhead lettuce isolates of Fusarium oxysporum f. sp. lactucae in Japan

Races were identified among butterhead lettuce isolates of Fusarium oxysporum f. sp. lactucae collected from three geographical areas of Hokkaido, Shizuoka, and Fukuoka in Japan by inoculation tests using Fujinagas race differential cultivars of lettuce (i.e., Patriot, Costa Rica No. 4, and Banchu Red Fire). Eighteen isolates from Shizuoka and Fukuoka were designated race 3, with two unknown vegetative compatibility groups (VCGs) that differed from Ogisos VCG 1 and 2. These two new VCGs were obtained from both Shizuoka and Fukuoka. On the other hand, three isolates from Hokkaido were classified as race 1 and identified as VCG 1, which represents a VCG of crisphead isolates from Nagano.     

Induced resistance in tomato plants against Fusarium wilt invoked by Fusarium oxysporum f.sp. dianthi

Tomato plants, susceptible toFusarium oxysporum f. sp.lycopersici, were inoculated by immersing the roots in a conidial suspension ofF. oxysporum f. sp.lycopersici race 1,F. oxysporum f. sp.dianthi race 2 or a mixture of both fungi. Plants inoculated withF. oxysporum f. sp.lycopersici showed disease symptoms after 2 weeks, whereas plants inoculated withF. oxysporum f. sp.dianthi or a mixture of both fungi remained symptomless for over 7 weeks, the duration of the experiment. In another experiment root systems of plants were split and each half was separately inoculated. One half was firstly inoculated withF. oxysporum f. sp.dianthi or treated with water, followed after a week by a second inoculation of the other half withF. oxysporum f. sp.lycopersici or by a water treatment. The disease symptoms in the half firstly inoculated withF. oxysporum f. sp.dianthi were significantly delayed, compared to plants of which that half had been treated with water. BecauseF. oxysporum f. sp.dianthi reduced disease symptoms caused byF. oxysporum f. sp.lycopersici without any direct interaction with this pathogen, it is concluded thatF. oxysporum f. sp.dianthi is able to induce resistance againstF. oxysporum f. sp.lycopersici in tomato plants.     

Development of sequence tagged site markers to identify races of Fusarium oxysporum f. sp. lactucae

By random amplified polymorphic DNA (RAPD) analysis of the representative isolates of each race of Fusarium oxysporum f. sp. lactucae, RAPD fragments of 0.6, 1.6, and 2.9kb were obtained. The 0.6-kb RAPD fragment was common to the representative isolates of all three races. Amplification of the 1.6- and 2.9-kb fragments were unique to the isolates of races 1 and 2, respectively. Sequence tagged site (STS) marker FLA0001, FLA0101, and FLA0201 were generated from the 0.6-, 1.6-, and 2.9-kb RAPD fragments, respectively. Polymerase chain reaction (PCR) analysis showed that FLA0001 was common to all 49 isolates of F. oxysporum f. sp. lactucae. FLA0101 was specifically generated from all 23 isolates of race 1 but not from races 2 or 3. FLA0201 was specifically amplified from all 12 isolates of race 2 but not from races 1 or 3. In two isolates of F. oxysporum f. sp. lactucum, PCR amplified FLA0001 and FLA0101 but not FLA0201. On the other hand, these STS markers were not detected from isolates of five other formae speciales. Because these STS markers were not generated from isolates of other plant pathogenic fungi, bacteria, or plant materials examined in this study, PCR analysis combined with the three STS markers should be a useful means for rapid identification of races of F. oxysporum f. sp. lactucae.     

Genetic diversity in Fusarium oxysporum f.sp. dianthi and Fusarium redolens f.sp. dianthi

Pathogenic isolates were selected representing all known vegetative compatibility groups (VCGs) and races of Fusarium oxysporum sensu lato from Dianthus spp. On basis of differences in the internal transcribed spacer region of the ribosomal DNA, six VCGs were classified as F. oxysporum f.sp. dianthi and four as F. redolens f.sp. dianthi. All VCGs of F. oxysporum f.sp. dianthi were characterized by unique restriction fragment length polymorphisms (RFLPs), unique overall esterase profiles, and unique virulence spectra, supporting a clonal lineage concept. Two VCGs of F. oxysporum f.sp. dianthi nevertheless comprised more than one race, but races within the same VCG shared the same distinct overall virulence spectrum. VCGs belonging to F. redolens f.sp. dianthi also had unique RFLPs and unique virulence spectra, but had grossly identical esterase profiles. Three new races (9, 10 and 11) are described for F. oxysporum f.sp. dianthi, and four for F. redolens f.sp. dianthi. Two races previously considered lost were recovered; race 7 was identified as a member of VCG 0021 of F. oxysporum f.sp. dianthi while race 3 was identified as a distinct VCG and race of F. redolens f.sp. dianthi. A summary of races and VCGs in F. oxysporum f.sp. dianthi and F. redolens f.sp. dianthi is presented.     

Plant defence reactions against fusarium wilt in chickpea induced by incompatible race 0 of Fusarium oxysporum f.sp. ciceris and nonhost isolates of F. oxysporum

Germinated seeds of 'kabuli' chickpea cv. ICCV 4 were inoculated with a conidial suspension of the incompatible race 0 of Fusarium oxysporum f.sp. ciceris (Foc) or of nonhost F. oxysporum resistance 'inducers', and 3 days later were challenged by root dip with a conidial suspension of highly virulent Foc race 5. Prior inoculation with inducers delayed the onset of symptoms and/or significantly reduced the final amount of fusarium wilt caused by race 5. However, the extent of disease suppression varied with the nature of the inducing agent; the nonhost isolates of F. oxysporum were more effective at disease suppression than the incompatible Foc race 0. Inoculation with the inducers gave rise to synthesis of maackiain and medicarpin phytoalexins in inoculated seedlings; these did not accumulate in plant tissues but were released into the inoculum suspension. Inoculation with inducers also resulted in accumulation of chitinase, β-1,3-glucanase and peroxidase activities in plant roots. These defence-related responses were induced more consistently and intensely by nonhost isolates of F. oxysporum than by incompatible Foc race 0. The phytoalexins and, to a lesser extent, the antifungal hydrolases, were also induced after challenge inoculation with Foc race 5. However, in this case the defence responses were induced in both preinduced and noninduced plants infected by the pathogen. It is concluded that the suppression of fusarium wilt in this study possibly involved an inhibitory effect on the pathogen of preinduced plant defences, rather than an increase in the expression of defence mechanisms of preinduced plants following a subsequent challenge inoculation.     

Selection methods for determining resistance of carnation cultivars to Fusarium oxysporum f.sp. dianthi

The level of resistance of carnation ( Dianthus caryophyllus ) cultivars to wilt caused by Fusarium oxysporum f.sp. dianthi was compared in root-dip-inoculated plants grown in pots (filled with tuff or sandy soil) in a greenhouse and plants grown in a field where the soil was artificially infested with the fungus. In the field, wilt symptoms appeared first in susceptible and subsequently in resistant cultivars; none was immune. Variations in the level of resistance were expressed either by different percentages of wilted plants (i.e. disease incidence) or by delayed disease progress as compared to a susceptible cultivar. The range of disease severity in the field, ranked on a scale from 0 to 4, was highly and significantly correlated with the percentage of diseased plants. The greenhouse test was unreliable as a predictor of the degree of resistance observed in the field. Similar wilt levels in the greenhouse and the field were found only in susceptible cultivars.     

香蕉枯萎病菌生理分化研究摘要本研究

本研究对香蕉枯萎病菌菌株FOCAAA9(来自香蕉)和FOCABB1(来自粉蕉)进行培养试验和接种试验;在含粉蕉和香蕉组织浸提液的培养基上2个菌株的培养性状、菌丝生长速度、孢子形态、大小型孢子比率和产孢量显示出差异;接种结果FOCAAA9能侵染香蕉(MusaAAA)品种巴西蕉、红香蕉和台蕉引起枯萎病,而FOCABB1对3个香蕉品种无致病性。研究结果表明侵染香蕉和粉蕉的古巴尖镰孢[Fusariumoxysporumf.sp.cubense(E.F.Smith)Snyder]存在生理分化现象。     

Identification of Race 1 of Fusarium oxysporum f. sp. lactucae on Lettuce by Inter-Retrotransposon Sequence-Characterized Amplified Region Technique

ABSTRACT Fusarium wilt of lettuce, caused worldwide by Fusarium oxysporum f. sp. lactucae, is an emerging seed-transmitted disease on Lactuca sativa. In order to develop a molecular diagnostic tool for identifying race 1 (VCG0300) of the pathogen on vegetable samples, an effective technique is presented. Inter-retrotransposon amplified polymorphism polymerase chain reaction (PCR), a technique based on the amplification of genomic regions between long terminal repeats, was applied. It was shown to be useful for grouping F. oxysporum f. sp. lactucae race 1 isolates. Inter-retrotransposon sequence-characterized amplified regions (IR-SCAR) was used to develop a specific set of PCR primers to be utilized for differentiating F. oxysporum f. sp. lactucae isolates from other F. oxysporum isolates. The specific primers were able to uniquely amplify fungal genomic DNA from race 1 isolates obtained in Italy, Portugal, the United States, Japan, and Taiwan. The primers also were specific to pathogen DNA obtained from artificially infected lettuce seed and naturally and artificially infected plants.     

Fungal polygalacturonase activity reflects susceptibility of carnation cultivars to fusarium wilt

Carnation cultivars with different levels of partial resistance were inoculated with race 2 of Fusarium oxysporum f.sp. dianthi and monitored for accumulation of host phytoalexins, fungal escape from compartmentalization, production of fungal pectin-degrading enzymes and development of external disease symptoms. Accumulation of phytoalexins, assessed after 10 days in the first 5 cm above the inoculation site, was weakly (methoxydianthramide S) or not (hydroxydianthalexin B) correlated with resistance levels after 12 weeks. Fungal escape from compartmentalization, assessed after 3 weeks as percentages colonized plants at 8 cm above the inoculation site, was highly correlated with expression of susceptibility after 12 weeks. Polygalacturonase (PG) activity, assessed after 4 weeks in the first 5 cm above the inoculation site, was highly correlated to final disease development. Linear increases in disease severity were accompanied by quadratic increases in PG activity. In contrast to water-treated plants, that lacked any PG activity, inoculated plants contained two main groups of fungal PGs, the dominant forms of which had estimated pI values of 7.0 and minimally 9.5, respectively. Compared to those of the first group, enzymes of the second group were produced only in trace amounts in liquid media containing pectin or polygalacturonate as sole source of carbon. On these media, the fungus also produced a pectin methyl esterase (PME) with an estimated pI of 9.3. Besides PMEs of host origin, inoculated plants of susceptible cultivars contained the fungal PME while no more than traces were found in resistant ones.Assessment of phytoalexin production by the host during defense responses cannot replace monitoring of external symptoms as a resistance test. Assessment of fungal growth, whether by reisolations above the compartmentalization area or by measurement of PG activity, provides a both rapid and reliable prediction of disease development.     

香蕉枯萎病菌生理分化研究

本研究对香蕉枯萎病菌菌株FOCAAA9（来自香蕉）和FOCABB1（来自粉蕉）进行培养试验和接种试验；在含粉蕉和香蕉组织漫提液的培养基上2个菌株的培养性状、菌丝生长速度、孢子形态、大小型孢子比率和产孢量显示出差异；接种结果FOCAAA9能侵染香蕉（Musa AAA）品种巴西蕉、红香蕉和台蕉引起枯萎病，而FOCABB1对3个香蕉品种无致病性。研究结果表明侵染香蕉和粉蕉的古巴尖镰孢[Fusarium oxysporum f．sp．cubeilse（E．F．Smith）Snyder]存在生理分化现象。