Genetic variability among isolates of Fusarium oxysporum from sugar beet

Fusarium yellows, caused by the soil‐borne fungus Fusarium oxysporum f. sp. betae (Fob), can lead to significant yield losses in sugar beet. This fungus is variable in pathogenicity, morphology, host range and symptom production, and is not a well characterized pathogen on sugar beet. From 1998 to 2003, 86 isolates of F. oxysporum and 20 other Fusarium species from sugar beet, along with four F. oxysporum isolates from dry bean and five from spinach, were obtained from diseased plants and characterized for pathogenicity to sugar beet. A group of sugar beet Fusarium isolates from different geographic areas (including nonpathogenic and pathogenic F. oxysporum, F. solani, F. proliferatum and F. avenaceum), F. oxysporum from dry bean and spinach, and Fusarium DNA from Europe were chosen for phylogenetic analysis. Sequence data from β‐ tubulin, EF1α and ITS DNA were used to examine whether Fusarium diversity is related to geographic origin and pathogenicity. Parsimony and Bayesian MCMC analyses of individual and combined datasets revealed no clades based on geographic origin and a single clade consisting exclusively of pathogens. The presence of FOB and nonpathogenic isolates in clades predominately made up of Fusarium species from sugar beet and other hosts indicates that F. oxysporum f. sp. betae is not monophyletic.     

晋南地区黄瓜、黑籽南瓜死秧的病原菌鉴定及品种抗性研究

经分离、培养对不同菌种培养性状的观察,确定了侵染黄瓜、黑籽南瓜造成死秧的镰刀菌主要为尖镰孢菌黄瓜专化型、尖镰孢菌西瓜专化型、串珠镰刀菌和腐皮镰孢菌4种。经致病性测定,4种镰刀菌均能侵染黄瓜,引起发病造成死秧,可分为强致病类型和中强致病类型。经抗病性鉴定,黑籽南瓜种子只有南瓜4号为耐病品种;黄瓜种子也只有津优31号为耐病品种。     

Morphology,phylogeny and pathogenicity of Fusarium species from Sansevieria trifasciata in Malaysia

Leaf blight is a common disease affecting Sansevieria trifasciata in many countries, including Malaysia. In the present study, Fusarium isolates were consistently recovered from the diseased leaves collected from various locations throughout the country. Based on morphology and multigene phylogenetic analysis using mitochondrial small subunit (mtSSU), intergenic spacer region (IGS) and translation elongation factor 1-α (TEF1-α) gene sequences, seven Fusarium species were identified, with F. oxysporum being the most prevalent (67.6%) among 34 isolates. Pathogenicity tests resulted in the discovery of pathogenic isolates that belonged to F. oxysporum, F. proliferatum, and F. pseudocircinatum, whereas all isolates of F. brachygibbosum, F. concentricum, F. mangiferae, and F. solani were nonpathogenic. The results suggest that several Fusarium species are accountable for causing disease on S. trifasciata in Malaysia.     

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.     

Seasonal progression of leaf rust in ‘Niagara Rosada’ grapevine in a biannual crop system in Brazil

The soil-borne fungus Fusarium oxysporum can cause both Fusarium yellows and Fusarium root rot diseases with severe yield losses in cultivated sugar beet. These two diseases cause similar foliar symptoms but different root response and have been proposed to be caused by two distinct F. oxysporum formae speciales. Fusarium yellows, caused by F. oxysporum f. sp. betae, presents vascular discoloration, whereas Fusarium root rot, due to F. oxysporum f. sp. radicis-betae, appears as black rot visible on the root surface. The aim of this work was to study the host-pathogen interaction between sugar beet lines and isolates originally characterized as Fusarium oxysporum f. sp. betae. Eight susceptible sugar beet lines, selected by the USDA-ARS (US) and UNIPD (University of Padova, Italy) breeding programs, were inoculated with three different isolates of F. oxysporum f. sp. betae, the causal agent of Fusarium yellows, representing different genetic groups. All inoculated lines developed symptoms, but severity, expressed as area under the disease progress curve (AUDPC), differed significantly (P < 0.05) among lines. Two lines from UNIPD, 6 and 9, were the most susceptible to the disease, whereas the other lines showed similar levels. The three isolates of F. oxysporum f. sp. betae differed significantly (P < 0.05) in disease severity. Five weeks after inoculation the plants were harvested and roots examined. Surprisingly, severe root rot was observed in the susceptible UNIPD lines when inoculated with all three isolates, while this symptom was never observed in the USDA germplasm. The development of this disease symptom obviously depends on the plant genotype.     

Genetic diversity of Fusarium oxysporum and related species pathogenic on tomato in Algeria and other Mediterranean countries

In order to characterize the pathogen(s) responsible for the outbreak of fusarium diseases in Algeria, 48 Fusarium spp. isolates were collected from diseased tomato in Algeria and compared with 58 isolates of Fusarium oxysporum originating from seven other Mediterranean countries and 24 reference strains. Partial sequences of the translation elongation factor EF‐1α gene enabled identification of 27 isolates as F. oxysporum, 18 as F. commune and three as F. redolens among the Algerian isolates. Pathogenicity tests confirmed that all isolates were pathogenic on tomato, with disease incidence greater at 28°C than at 24°C. All isolates were characterized using intergenic spacer (IGS) DNA typing, vegetative compatibility group (VCG) and PCR detection of the SIX1 (secreted in xylem 1) gene specific to F. oxysporum f. sp. lycopersici (FOL). No DNA polymorphisms were detected in the isolates of F. redolens or F. commune. In contrast, the 27 Algerian isolates of F. oxysporum were shown to comprise nine IGS types and 13 VCGs, including several potentially new VCGs. As none of the isolates was scored as SIX1+, the 27 isolates could be assigned to F. oxysporum f. sp. radicis‐lycopersici (FORL). Isolates from Tunisia were also highly diverse but genetically distinct from the Algerian isolates. Several Tunisian isolates were identified as FOL by a PCR that detected the presence of SIX1. The results show that isolates from European countries were less diverse than those from Tunisia. Given the difference between Algerian populations and populations in other Mediterranean countries, newly emergent pathogenic forms could have evolved from local non‐pathogenic populations in Algeria.     

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.     

The potential for Fusarium oxysporum f. sp. fragariae,cause of fusarium wilt of strawberry,to colonize organic matter in soil and persist through anaerobic soil disinfestation

Fusarium wilt of strawberry, caused by Fusarium oxysporum f. sp. fragariae, is a disease of primary concern for strawberry production in many countries. Crop rotation and anaerobic soil disinfestation (ASD) have gained recent interest for their potential to contribute to management of this disease. Both techniques involve incorporation of organic matter into soil, which may be utilized by strains of Fusarium that are competitive saprophytes. We show that F. oxysporum f. sp. fragariae can colonize strawberry, lettuce, raspberry, and broccoli leaf tissues, which are sources of organic matter generated during crop rotation. This pathogen increased in soil population density during ASD treatments that did not become anaerobic, possibly as a result of growth on the organic amendment. However, significant population decreases were observed after ASD treatment when at least 100,000 cumulative reduced mV hours occurred in a 14-day experiment. Post-ASD abundance of F. oxysporum f. sp. fragariae in soil was negatively correlated with cumulative reduced mV hours. The only treatment that consistently caused disinfestation was exposed to a maximum temperature of 22 °C, which indicates there is potential for developing effective ASD treatments in the cool climates where strawberries are grown. Awareness that F. oxysporum f. sp. fragariae can act as a competitive soil saprophyte should be further investigated for its potential to alter disease outcomes where organic amendments are applied.     

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.     

Variation in potential effector genes distinguishing Australian and non‐Australian isolates of the cotton wilt pathogen Fusarium oxysporum f.sp. vasinfectum

This study identified genes that distinguish Australian Fusarium oxysporum f.sp. vasinfectum (Fov) isolates from related co‐localized non‐pathogenic F. oxysporum isolates and from non‐Australian Fov isolates. One gene is a homologue of the F. oxysporum f.sp. lycopersici (Fol) effector gene SIX6, encoding a 215‐residue cysteine‐rich secreted protein. The Six6 proteins from Fol and Fov contained eight conserved cysteine residues, five of which occurred in the highly diverged 48‐amino‐acid region where FovSix6 differs from FolSix6 at 32 residues. Two other potential effector genes, PEP1 and PEP2, were identified in a cDNA library of Fov genes expressed during infection of cotton. The presence of FovSIX6 and other differences in DNA fingerprints clearly distinguished Australian Fov isolates from non‐Australian Fov isolates and these differences further support the hypothesis based on earlier phylogenetic analysis that Australian Fov is different from Fov in other cotton‐growing areas. A specific diagnostic for Fov based on FovSIX6 is described.     

Fusarium commune associated with wilt and root rot disease in rice

Wilt and root rot disease in plants has been caused mainly by Fusarium species. Previous studies reported that members of the Fusarium oxysporum species complex (FOSC) were usually associated with this disease, but there has been no report of it being caused in rice by specific Fusarium species. However, in this study, Fusarium commune was identified and characterized as a causal agent of wilt and root rot disease of rice. Four Fusarium isolates (BD005R, BD014R, BD019R, and BD020R) were obtained from different parts (root, stem, and seeds) of diseased rice plants. In morphological studies, these isolates produced key characteristics of F. commune, such as long and slender monophialides, polyphialides, and abundant chlamydospores. In molecular studies, the isolates were identified as F. commune based on sequences of the translation elongation factor 1-α (TEF1) gene that had 99.7%–100% sequence identity with the reference strain F. commune NRRL 28058. The phylogenetic tree showed that all four isolates belonged to the F. commune clade. A mating type test determined that three isolates carried MAT1-2. Their teleomorph stage was still unknown. Pathogenicity assays showed that all the isolates produced wilt and root rot symptoms and the isolate BD019R was observed as the most virulent among the isolates. To our knowledge, this is the first report of F. commune causing wilt and root rot disease on rice.     

苦瓜枯萎病菌的鉴定及其遗传多样性分析

 对分离获得的32株苦瓜枯萎病菌菌株进行形态学特征和寄主专化型测定, 结果表明, 测试的苦瓜枯萎病菌株均为尖孢镰刀菌苦瓜专化型 (Fusarium oxysporum f. sp. momordicae), 这些菌株可以侵染苦瓜和瓠瓜幼苗, 但不侵染其他葫芦科瓜类作物。对苦瓜枯萎病菌菌株的rDNA-ITS区 (ITS1、5.8S和ITS2)序列进行扩增测序, 结果显示其序列长度均为456 bp;聚类分析表明测序菌株与镰刀菌属中尖孢镰刀菌不同专化型的菌株聚为一群。利用RAPD标记技术分析苦瓜枯萎病菌的遗传多样性, 结果显示苦瓜枯萎病菌株与其他葫芦科瓜类作物枯萎病菌株间的遗传相似系数范围为0.59~0.99, 当遗传相似系数为0.85时, 供试的48个菌株分成10个类群 (G_1~10)。在RAPD聚类树中所有苦瓜枯萎病菌株聚在一个分支上 (G₁群), 菌株间的遗传相似系数范围为0.92~1.00, 具有较高的遗传相似性, 且菌株的聚群与地理来源存在一定的相关性。     

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.     

Fusarium Redolens f.sp asparagi, Causal Agent of Asparagus Root Rot, Crown Rot and Spear Rot

Two Fusarium species, F. oxysporum f.sp. asparagi and F. proliferatum, are known to be involved in the root and crown rot complex of asparagus. We have investigated reports on the involvement of F. redolens, a third species, which until recently was considered conspecific with F. oxysporum because of morphological similarities. RFLP analysis of the rDNA internal transcribed spacer region and AFLP fingerprinting identified eight strains from asparagus unambiguously as F. redolens. Four of these were tested and found to be pathogenic to asparagus either in this study (two strains) or in a previous one in which they were classified as F. oxysporum (three strains). Disease symptoms and disease development were the same as with F. oxysporum f.sp. asparagi and F. proliferatum. Present data and literature reports identify F. redolens as a host-specific pathogen involved in root, crown and spear rot of asparagus. The pathogen is formally classified as F. redolens Wollenw. f.sp. asparagi Baayen.     

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.     

Pathogenic variation and molecular characterization of <Emphasis Type="Italic">Fusarium</Emphasis> species isolated from wilted Welsh onion in Japan

Thirty-two isolates of Fusarium species were obtained from wilted Welsh onion (Allium fistulosum) grown on nine farms from six regions in Japan and identified as F. oxysporum (18 isolates), F. verticillioides (7 isolates), and F. solani (7 isolates). The pathogenicity of 32 isolates was tested on five commercial cultivars of Welsh onion and two cultivars of bulb onion in a seedling assay in a greenhouse. The Fusarium isolates varied in the degree of disease severity on the cultivars. Five F. oxysporum isolates (08, 15, 17, 22, and 30) had a higher virulence on the cultivars than the other isolates. The host range of these five isolates was limited to Allium species. Molecular characterization of Fusarium isolates was performed using polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) analysis of the internal transcribed spacer (ITS) regions of ribosomal DNA. The 32 isolates were grouped into eight types (four types for F. oxysporum, one for F. verticillioides, and three for F. solani). Restriction patterns of the ITS region were not related to pathogenicity. However, the haplotypes obtained with five enzymes (RsaI, HinfI, HaeIII, ScrFI, and MspI) and the phylogenetic analysis permitted the discernment of the three Fusarium species. The PCR-RFLP analysis should provide a rapid, simple method for differentiating Fusaruim species isolated from wilted Welsh onion in Japan.     

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.     

Genetic diversity and pathogenicity of <Emphasis Type="Italic">Fusarium oxysporum</Emphasis> isolated from wilted rocket plants in Italy

Thirty-six isolates of Fusarium oxysporum originated from Eruca vesicaria and Diplotaxis tenuifolia together with eight reference strains belonging to the formae speciales raphani, matthioli and conglutinans, typical on the Brassicaceae family, were tested for pathogenicity on two species of rocket plants (E. vesicaria L., syn. E. sativa, cv. ‘Rucola coltivata’; and D. tenuifolia cv. ‘Winter’) cultivated in the glasshouse. The results showed that different isolates were slightly, moderately or highly virulent. The strains were examined for differences in the nucleotide sequence of the ribosomal DNA (rDNA) intergenic spacer (IGS) region, about 2.5 kb long. The phylogenetic (neighbor-joining) analysis performed on the isolates enabled identification of four different groups, named I, II, III and IV. Thirty-one isolates out of 36 clustered in group I and were genetically similar to F. oxysporum f.sp. raphani. By considering the pathogenicity of the strains included in Group I, a partial host specialization could be observed: the average disease index of the isolates from D. tenuifolia was higher on wild rocket, whereas the average disease index of the isolates from E. vesicaria was higher on cultivated rocket. Moreover, isolates from cultivated rocket showed, on average, a higher degree of aggressiveness than the isolates from wild rocket. Concerning Group I, the sequence analysis confirmed the homogeneity of the population, with only five parsimony-informative SNPs and five haplotypes. Twenty-six out of 31 isolates belonged to haplotype 1. Groups II and III were genetically similar to strains of F. oxysporum f.sp. matthioli. Three other strains, not pathogenic or with a medium level of virulence, clustered together in Group 4, but their sequence was distant from that of other formae speciales. The pathogenicity and IGS analysis confirmed the presence of virulence variation and genetic diversity among the F. oxysporum isolates studied. To our knowledge, this is the first report of differentiation of formae speciales of F. oxysporum on rocket plants by IGS analysis.