Effect of moisture on thermal inactivation of soilborne pathogens under structural solarization

ABSTRACT Structural solarization of greenhouses for sanitation by closing them involves dry heating to 60 degrees C and higher with a consequent low relative humidity (RH) ( approximately 15%), thus requiring an extended period for thermal inactivation of pathogens. In an attempt to enhance pathogen control by increasing moisture during the hot hours of the day, various regimes of inoculum moistening were studied. However, wetting inoculum of Fusarium oxysporum f. sp. melonis and F. oxysporum f. sp. radicis-lycopersici resulted in less effective pathogen control compared with that of dry heating. Fifty percent effective dose (ED(50)) values of thermal inactivation of wetted and dry inoculum for the former pathogen were 18 and 7 days, respectively, and for the latter, a respective 9 and 4 days. This was because wetting resulted in inoculum cooling due to evaporation, which eventually led to its drying. A model describing the drying of wet inoculum in a wetted greenhouse, based on the fact that there was an approximately 10 degrees C difference between greenhouse and ambient temperatures, was proposed. A double-tent system reduced this difference to 1 to 2 degrees C, reduced moisture loss, and led to improved inoculum inactivation of F. oxysporum f. sp. radicis-lycopersici. Thus, the ED(50) value of thermal inactivation was reduced from 15 days to 1 day, because this system provided both high temperature ( approximately 60 degrees C) and high RH ( approximately 100%), resulting in effective wet heating.     

Combined soil treatments and sequence of application in improving the control of soilborne pathogens

ABSTRACT The effects of reduced doses of methyl bromide (MB) or metham sodium, heating, short solarization, and soil microbial activity, alone or in combination, on survival of soilborne fungal pathogens were tested in a controlled-environment system and field plots. Sublethal doses of heating or MB delayed germination of Sclerotium rolfsii sclerotia. Combining MB and heating treatments was more effective than either treatment alone in controlling S. rolfsii and Fusarium oxysporum f. sp. basilici. The application heating followed by fumigation with MB, was significantly more effective in delaying and reducing germination of S. rolfsii sclerotia and in controlling F. oxysporum f. sp. basilici than the opposite sequence. Further, incubation in soil and exposure to microbial activity of previously heated or MB-treated sclerotia increased the mortality rate, indicating a weakening effect. Similarly, incubation of chlamydospores of F. oxysporum f. sp. melonis and F. oxysporum f. sp. radicis-lycopersici in soil in the field after fumigation further reduced their survival, confirming the laboratory results. In field tests, combining MB or metham sodium at reduced doses with short solarization was more effective in controlling fungal pathogens than either treatment alone. Treatment sequence significantly affected pathogen control in the field, similar to its effect under controlled conditions. This study demonstrates a frequent synergistic effect of combining soil treatments and its potential for improving pathogen control and reducing pesticide dose, especially when an appropriate sequence was followed.     

Survival of plant pathogens under structural solarization

Structural solarization of greenhouses is a nonchemical sanitation procedure. The method involves dry heating, since maximal temperatures may exceed 60°C and consequent relative humidity (r.h.) is low (ca 15%), under fluctuating temperature and r.h. regimes. Thirty-five structural solarization experiments were performed over 7 years, testing one bacterial and five fungal plant pathogens. Various aspects of pathogen thermal inactivation under this method were studied. Thermal inactivation of the various pathogens differed according to the organism and inoculum form. Sensitivity to heat was highest withClavibacter michiganensis subsp.michiganensis and lowest withFusarium oxysporum f.sp.radicislycopersici inoculum in dry infected tomato stems, with ED₈₀ values of 7 and 47 days, respectively; intermediate values were obtained forPythium sp.,F. oxysporum f.sp.melonis, F. oxysporum f.sp.basilici andSclerotium rolfsii. The maximal ambient temperatures were in the range of 28.2° to 33.1°C. Structural solarization for sanitation can be a useful component of integrated pest management in greenhouses. http://www.phytoparasitica.org posting Sept. 28, 2004.     

Sporulation of Fusarium oxysporum f. sp. lycopersici on Stem Surfaces of Tomato Plants and Aerial Dissemination of Inoculum

ABSTRACT Plants exhibiting symptoms of wilt and xylem discoloration typical of Fusarium wilt caused by Fusarium oxysporum f. sp. lycopersici were observed in greenhouses of cherry tomatoes at various sites in Israel. However, the lower stems of some of these plants were covered with a pink layer of macroconidia of F. oxysporum. This sign resembles the sporulating layer on stems of tomato plants infected with F. oxysporum f. sp. radicis-lycopersici, which causes the crown and root rot disease. Monoconidial isolates of F. oxysporum from diseased plants were assigned to vegetative compatibility group 0030 of F. oxysporum f. sp. lycopersici and identified as belonging to race 1 of F. oxysporum f. sp. lycopersici. The possibility of coinfection with F. oxysporum f. sp. lycopersici and F. oxysporum f. sp. radicis-lycopersici was excluded by testing several macroconidia from each plant. Airborne propagules of F. oxysporum f. sp. lycopersici were trapped on selective medium in greenhouses in which plants with a sporulating layer had been growing. Sporulation on stems was reproduced by inoculating tomato plants with races 1 and 2 of F. oxysporum f. sp. lycopersici. This phenomenon has not been reported previously with F. oxysporum f. sp. lycopersici and might be connected to specific environmental conditions, e.g., high humidity. The sporulation of F. oxysporum f. sp. lycopersici on plant stems and the resultant aerial dissemination of macroconidia may have serious epidemiological consequences. Sanitation of the greenhouse structure, as part of a holistic disease management approach, is necessary to ensure effective disease control.     

Spatial distribution and temporal development of fusarium crown and root rot of tomato and pathogen dissemination in field soil

ABSTRACT The spatial distribution and temporal development of tomato crown and root rot, caused by Fusarium oxysporum f. sp. radicis-lycopersici, were studied in naturally infested fields in 1996 and 1997. Disease progression fit a logistic model better than a monomolecular one. Geostatistical analyses and semivariogram calculations revealed that the disease spreads from infected plants to a distance of 1.1 to 4.4 m during the growing season. By using a chlorate-resistant nitrate nonutilizing (nit) mutant of F. oxysporum f. sp. radicis-lycopersici as a "tagged" inoculum, the pathogen was found to spread from one plant to the next via infection of the roots. The pathogen spread to up to four plants (2.0 m) on either side of the inoculated focus plant. Root colonization by the nit mutant showed a decreasing gradient from the site of inoculation to both sides of the inoculated plant. Simulation experiments in the greenhouse further established that this soilborne pathogen can spread from root to root during the growing season. These findings suggest a polycyclic nature of F. oxysporum f. sp. radicis-lycopersici, a deviation from the monocyclic nature of many nonzoosporic soilborne pathogens.     

Controlled laboratory system to study soil solarization and organic amendment effects on plant pathogens

ABSTRACT A controlled laboratory system for simulating soil solarization, with and without organic amendment, was developed and validated using physical, chemical, and biological parameters. The system consists of soil containers that are exposed to controlled and constant aeration, and to temperature fluctuations that resemble those occurring during solarization at various depths. This system enables a separate analysis of volatiles and other components. We recorded a sharp decrease in oxygen concentration in the soil atmosphere followed by a gradual increase to the original concentration during solarization in the field and heating in the simulation system of soil amended with wild rocket (Diplotaxis tenuifolia) or thyme (Thymus vulgaris). The combined treatment of organic amendment and solarization (or heating in the controlled system) was highly effective at controlling populations of Fusarium oxysporum f. sp. radicis-lycopersici. Changes in soil pH, enzymatic activities, and microbial populations followed, in most cases, trends which were similar under both solarization and the heating system, when exposed to controlled aerobic conditions. The reliability and validity of the system in simulating physical, chemical, and biological processes taking place during solarization is demonstrated.     

The infestation of tomato seed by Fusarium oxysporum f.sp. radicis-lycopersici

The infestation of seed by Fusarium oxysporum f.sp. radicis-lycopersici occurred at rates of 0-1 to 0-01 % in fruit on stem-infected plants. Direct infection of fruit in the flower or young developing fruit stage resulted in a grayish-brown lesion on the stylar end of the fruit or mummification. F. oxysporum f.sp. radicis-lycopersici was isolated from all seeds in such fruit. Picked fruit inoculated on the stem scar also became infected but 96 h after inoculation of the fruit, the seed was not infested or infected. The contact of clean seed with hands that had previously handled F. oxysporum f.sp. radicis-lycoperisid-infesied sawdust resulted in a high level of seed infestation. The fungus survived on seed sent across Canada and stored for up to 12 weeks. Treatment with NaOCl or 0·1 N HCl did not completely disinfest infested seed.     

Effect of Relative Humidity on Sporulation of Fusarium oxysporum in Various Formulations and Effect of Water on Spore Movement Through Soil

ABSTRACT Fusarium oxysporum f. sp. erythroxyli is being investigated as a mycoherbicide for the narcotic plant coca. Sporulation of the fungus in seven formulations containing different organic substrates and movement of its propagules through soil were studied. The formulations were a granular wheat flour/kaolin (pesta); an extruded wheat and rice flour (C-6); and five alginate pellet products containing corn cobs, soybean hull fiber, canola meal, rice flour, or rice flour plus canola oil. Formulations were incubated at 25 degrees C for 6 weeks in desiccators with various salt solutions to provide nine relative humidities (RH), ranging from 100% (pure deionized water) to 0% (anhydrous (CaSO(4)). Hyphae of F. oxysporum f. sp. erythroxyli grew out of alginate pellets with canola meal, rice, and rice plus canola oil as early as 24 h at 100% constant RH. Alginate pellets of rice plus canola oil and granular C-6 and pesta formulations consistently produced more microconidia, macroconidia, and CFU than the other four formulations at all RH tested. The C-6 formulation produced more propagules than the other formulations at low RH (<53%). Canola meal pellets produced more spores than three other formulations when exposed to fluctuating RH (100 to 75%). The effect of percolating water on spore movement through soil was studied using a plant-pathogenic isolate of F. oxysporum f. sp. niveum. To determine the effect of water percolation on propagule movement, formulations were placed on soil columns and artificial rain was applied. In general, 10-fold fewer CFU were recovered at a 8- to 10-cm depth compared with a 0- to 2-cm depth.     

The effects of soil solarization and soil fumigation on Fusarium wilt of watermelon grown in plastic house in south-eastern Spain

The effects of pre-planting solarization or fumigation with metham-sodium of sand-mulched soil on fusarium wilt of watermelon in plastic house culture were investigated at Almeria, south-eastern Spain. In two trials, 2 months' solarization increased the average maximum soil temperature by c. 5°C to 44-48° C at 10 cm depth and by 4-5° C to 40-42° C at 20-30 cm. The amount of Fusarium oxysporum in the upper 15 cm of a naturally infested soil was reduced by solarization and by fumigation. During the 9 months following treatment, the F. oxysporum population stabilized at a low level in soil solarized for 2 months, but fluctuated in soil solarized for 1 month and increased in fumigated soil. The amount of wilt in watermelon sown into this soil after treatment was generally low; plants growing in solarized or fumigated soil suffered less wilt than plants in untreated soil but the differences were not significant. In a soil artificially infested with the highly pathogenic race 2 of F. oxysporum f. sp. niveum, F. oxysporum populations were greatly reduced following solarization or fumigation, and fluctuated erratically thereafter. Solarization for 2 months completely controlled wilt in watermelon and gave a fruit yield almost five times that of plants in untreated soil. Solarization for 1 month only slowed disease development slightly but gave a yield more than twice that in untreated soil. Fumigation with metham-sodium retarded disease development considerably and tripled fruit yield. Plant performance was significantly better in soil solarized for 2 months than in uninfested control soil, suggesting beneficial effects of this treatment additional to wilt control.     

Disease Development Following Infection of Tomato and Basil Foliage by Airborne Conidia of the Soilborne Pathogens Fusarium oxysporum f. sp. radicis-lycopersici and F. oxysporum f. sp. basilici

ABSTRACT Fusarium oxysporum f. sp. radicis-lycopersici, the causal agent of Fusarium crown and root rot of tomato, and F. oxysporum f. sp. basilici, the causal agent of Fusarium wilt in basil, are soilborne pathogens capable of producing conspicuous masses of macroconidia along the stem. The role of the airborne propagules in the epidemics of the disease in tomato plants was studied. In the field, airborne propagules of F. oxysporum f. sp. radicis-lycopersici were trapped with a selective medium and their prevalence was determined. Plants grown in both covered and uncovered pots, detached from the field soil, and exposed to natural aerial inoculum developed typical symptoms (82 to 87% diseased plants). The distribution of inoculum in the growth medium in the pots also indicated the occurrence of foliage infection. In greenhouse, foliage and root inoculations were carried out with both tomato and basil and their respective pathogens. Temperature and duration of high relative humidity affected rate of colonization of tomato, but not of basil, by the respective pathogens. Disease incidence in foliage-inoculated plants reached 75 to 100%. In these plants, downward movement of the pathogens from the foliage to the crown and roots was observed. Wounding enhanced pathogen invasion and establishment in the foliage-inoculated plants. The sporulation of the two pathogens on stems, aerial dissemination, and foliage infection raise the need for foliage protection in addition to soil disinfestation, in the framework of an integrated disease management program.     

Treatment with the Mycoparasite Pythium oligandrum Triggers Induction of Defense-Related Reactions in Tomato Roots When Challenged with Fusarium oxysporum f. sp. radicis-lycopersici

ABSTRACT The influence exerted by the mycoparasite Pythium oligandrum in triggering plant defense reactions was investigated using an experimental system in which tomato plants were infected with the crown and root rot pathogen Fusarium oxysporum f. sp. radicis-lycopersici. To assess the antagonistic potential of P. oligandrum against F. oxysporum f. sp. radicis-lycopersici, the interaction between the two fungi was studied by scanning and transmission electron microscopy (SEM and TEM, respectively). SEM investigations of the interaction region between the fungi demonstrated that collapse and loss of turgor of F. oxysporum f. sp. radicis-lycopersici hyphae began soon after close contact was established with P. oligandrum. Ultrastructural observations confirmed that intimate contact between hyphae of P. oligandrum and cells of the pathogen resulted in a series of disturbances, including generalized disorganization of the host cytoplasm, retraction of the plasmalemma, and, finally, complete loss of the protoplasm. Cytochemical labeling of chitin with wheat germ agglutinin (WGA)/ovomucoid-gold complex showed that, except in the area of hyphal penetration, the chitin component of the host cell walls was structurally preserved at a time when the host cytoplasm had undergone complete disorganization. Interestingly, the same antagonistic process was observed in planta. The specific labeling patterns obtained with the exoglucanase-gold and WGA-ovomucoid-gold complexes confirmed that P. oligandrum successfully penetrated invading cells of the pathogen without causing substantial cell wall alterations, shown by the intense labeling of chitin. Cytological investigations of samples from P. oligandrum-inoculated tomato roots revealed that the fungus was able to colonize root tissues without inducing extensive cell damage. However, there was a novel finding concerning the structural alteration of the invading hyphae, evidenced by the frequent occurrence of empty fungal shells in root tissues. Pythium ingress in root tissues was associated with host metabolic changes, culminating in the elaboration of structural barriers at sites of potential fungal penetration. Striking differences in the extent of F. oxysporum f. sp. radicis-lycopersici colonization were observed between P. oligandrum-inoculated and control tomato plants. In control roots, the pathogen multiplied abundantly through much of the tissues, whereas in P. oligandrum-colonized roots pathogen growth was restricted to the outermost root tissues. This restricted pattern of pathogen colonization was accompanied by deposition of newly formed barriers beyond the infection sites. These host reactions appeared to be amplified compared to those seen in nonchallenged P. oligandrum-infected plants. Most hyphae of the pathogen that penetrated the epidermis exhibited considerable changes. Wall appositions contained large amounts of callose, in addition to be infiltrated with phenolic compounds. The labeling pattern obtained with gold-complexed laccase showed that phenolics were widely distributed in Fusarium-challenged P. oligandrum-inoculated tomato roots. Such compounds accumulated in the host cell walls and intercellular spaces. The wall-bound chitin component in Fusarium hyphae colonizing P. oligandrum-inoculated roots was preserved at a time when hyphae had undergone substantial degradation. These observations provide the first convincing evidence that P. oligandrum has the potential to induce plant defense reactions in addition to acting as a mycoparasite.     

Interactions of Pratylenchus thornei and Fusarium oxysporum f. sp. ciceris on Chickpea

ABSTRACT Fusarium oxysporum f. sp. ciceris and the root-lesion nematode Pratylenchus thornei coinfect chickpeas in southern Spain. The influence of root infection by P. thornei on the reaction of Fusarium wilt-susceptible (CPS 1 and PV 61) and wilt-resistant (UC 27) chickpea cultivars to F. oxysporum f. sp. ciceris race 5 was investigated under controlled and field conditions. Severity of Fusarium wilt was not modified by coinfection of chickpeas by P. thornei and F. oxysporum f. sp. ciceris, in simultaneous or sequential inoculations with the pathogens. Root infection with five nematodes per cm(3) of soil and 5,000 chlamydospores per g of soil of the fungus resulted in significantly higher numbers of propagules of F. oxysporum f. sp. ciceris with the wilt-susceptible cultivar CPS 1, but not with the wilt-resistant one. However, infection with 10 nematodes per cm(3) of soil significantly increased root infection by F. oxysporum f. sp. ciceris in both cultivars, irrespective of fungal inoculum densities (250 to 2,000 chlamydospores per g of soil). Plant growth was significantly reduced by P. thornei infection on wilt-susceptible and wilt-resistant chickpeas in controlled and field conditions, except when shorter periods of incubation (45 days after inoculation) were used under controlled conditions. Severity of root necrosis was greater in wilt-susceptible and wilt-resistant cultivars when nematodes were present in the root, irrespective of length of incubation time (45 to 90 days), densities of nematodes (5 and 10 nematodes per cm(3) of soil), fungal inocula, and experimental conditions. Nematode reproduction on the wilt-susceptible cultivars, but not on the wilt-resistant one, was significantly increased by F. oxysporum f. sp. ciceris infections under controlled and field conditions.     

Control of Fusarium oxysporum f. sp. cucumerinum of cucumbers by soil solarization with impermeable plastics and/or reduced doses of methyl bromide*

Soil solarization is not broadly adopted as a soil deinfestation method mainly because of its long duration (4–6 weeks). We present evidence showing that the duration of solarization can be reduced to nearly half using impermeable plastics and/or low doses of methyl bromide, while still ensuring effective control of Fusarium oxysporum f. sp. cucumerinum. Chlamydospores of a pathogenic isolate of F. o. cucumerinum, formed in sterile soil, were inserted into nylon mesh envelopes and incorporated into the soil prior to treatment at 20‐ and 30‐cm soil depths. Soil treatments included untreated control, soil solarization with polyethylene or impermeable plastics (LMG), and soil solarization with polyethylene or impermeable plastics plus 20 g m^?2 methyl bromide. According to the effects on artificial inocula of F. o. cucumerinum checked at weekly intervals for 4 weeks, soil solarization with impermeable plastics was most effective in destroying pathogen populations even two weeks after soil covering.     

Vegetative compatibility of Fusarium oxysporum from sweet basil in Israel

Fusarium wilt and crown rot of sweet basil, caused by Fusarium oxysporum f.sp. basilici (F.o.ba.), is widespread in Israel. Affected plants show a variety of symptoms, including vascular wilt as well as crown rot, and masses of macroconidia on stem surfaces. We used vegetative compatibility to determine whether F.o.ba. isolates associated with various symptoms and sources are genetically related. All 119 isolates previously described as F.o.ba., and 42 additional F. oxysporum isolates which had not been tested for pathogenicity, belonged to a single vegetative compatibility group (VCG). The various symptoms are therefore induced by a single pathogenic form which appears to be a specific clone of F. oxysporum. The isolates of F.o.ba. from Israel were vegetatively compatible with eight isolates of F.o.ba. from Italy and the USA, but not with nonpathogenic isolates of F. oxysporum from basil, or with F.o. lycopersici or F.o. radicis-lycopersici from tomato. We conclude that the population of F.o.ba. in Israel belongs to the common VCG of this pathogen described in the USA, and which includes American and Italian isolates.     

Genetic Diversity of Fusarium oxysporum Isolates from Cucumber: Differentiation by Pathogenicity, Vegetative Compatibility, and RAPD Fingerprinting

ABSTRACT A total of 106 isolates of Fusarium oxysporum obtained from diseased cucumber plants showing typical root and stem rot or Fusarium wilt symptoms were characterized by pathogenicity, vegetative compatibility, and random amplified polymorphic DNA (RAPD). Twelve isolates of other formae speciales and races of F. oxysporum from cucurbit hosts, three avirulent isolates of F. oxysporum, and four isolates of Fusarium spp. obtained from cucumber were included for comparison. Of the 106 isolates of F. oxysporum from cucumber, 68 were identified by pathogenicity as F. oxysporum f. sp. radicis-cucumerinum, 32 as F. oxysporum f. sp. cucumerinum, and 6 were avirulent on cucumber. Isolates of F. oxysporum f. sp. radicis-cucumerinum were vegetatively incompatible with F. oxysporum f. sp. cucumerinum and the other Fusarium isolates tested. A total of 60 isolates of F. oxysporum f. sp. radicis-cucumerinum was assigned to vegetative compatibility group (VCG) 0260 and 5 to VCG 0261, while 3 were vegetatively compatible with isolates in both VCGs 0260 and 0261 (bridging isolates). All 68 isolates of F. oxysporum f. sp. radicis-cucumerinum belonged to a single RAPD group. A total of 32 isolates of F. oxysporum f. sp. cucumerinum was assigned to eight different VCGs and two different RAPD groups, while 2 isolates were vegetatively self-incompatible. Pathogenicity, vegetative compatibility, and RAPD were effective in distinguishing isolates of F. oxysporum f. sp. radicis-cucumerinum from those of F. oxysporum f. sp. cucumerinum. Parsimony and bootstrap analysis of the RAPD data placed each of the two formae speciales into a different phylogenetic branch.     

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.     

Pathogenic, Genetic and Molecular Characterisation of Fusarium oxysporum f.sp. lilii

Isolates of Fusarium oxysporum from lily were screened for pathogenicity, vegetative compatibility and DNA restriction fragment length polymorphisms, and compared to reference isolates of F. oxysporum f.sp. gladioli and F. oxysporum f.sp. tulipae to justify the distinction of F. oxysporum f.sp. lilii. Twenty-four isolates from different locations in The Netherlands (18 isolates), Italy (4 isolates), Poland and the United States (1 isolate each) shared unique RFLP patterns with probes D4 and pFOM7, while hybridization did not occur with a third probe (F9). Except for a self-incompatible isolate, these 24 isolates all belonged to a single vegetative compatibility group (VCG 0190). Isolates belonging to VCG 0190 were highly pathogenic to lily, but not to gladiolus or tulip, except for a single nonpathogenic isolate. Six saprophytic isolates of F. oxysporum from lily were nonpathogenic or only slightly aggressive to lily, gladiolus and tulip, belonged to unique VCGs and had distinct RFLP patterns. Three pathogenic isolates previously considered to belong to F. oxysporum f.sp. lilii were identified as F. proliferatum var. minus; all three belonged to the same VCG and shared unique RFLP patterns. These three isolates were moderately pathogenic to lily and nonpathogenic to gladiolus and tulip. The reference isolates of F. oxysporum f.sp. tulipae were pathogenic to tulip, but not to lily and gladiolus; they shared a distinct RFLP pattern, different from those encountered among pathogenic and saprophytic isolates from lily, and formed a separate new VCG (VCG 0230). Reference isolates of F. oxysporum f.sp. gladioli belonging to VCG 0340 proved pathogenic to both gladiolus and lily, but not to tulip. These isolates, as well as isolates belonging to VCGs 0341, 0342 and 0343 of F. oxysporum f.sp. gladioli, had RFLP patterns different from those encountered among the isolates from lily or tulip. These findings identify F. oxysporum f.sp. lilii as a single clonal lineage, distinct from F. oxysporum f.sp. gladioli and f.sp. tulipae.     

Characterization of Fusarium oxysporum f. sp. radicis-cucumerinum attacking melon under natural conditions in Greece

A severe root and stem rot disease of melon was observed during the 2001 growing season on four glasshouse crops in Heraklio, Greece. A total of 43 isolates of F. oxysporum , obtained in Crete from glasshouse-grown melon and showing fusarium wilt or root and stem rot symptoms, were characterized by pathogenicity and vegetative compatibility. The majority of these isolates was also fingerprinted via amplified fragment length polymorphic (AFLP) analysis. Of the total number of isolates, 22 were identified by pathogenicity tests as F. oxysporum f. sp. melonis , 20 as F. oxysporum f. sp. radicis-cucumerinum , while one isolate was nonpathogenic on cucumber, melon, sponge gourd and pumpkin. All 22 isolates of F. oxysporum f. sp. melonis were assigned to vegetative compatibility group (VCG) 0134, and all 20 isolates of F. oxysporum f. sp. radicis-cucumerinum to VCG 0260. Isolates of F. oxysporum f. sp. radicis-cucumerinum were incompatible with isolates of F. oxysporum f. sp. melonis. AFLP fingerprinting allowed for the clustering of the isolates of the two formae speciales of F. oxysporum along two separate phenetic groups: f. sp. melonis to AFLP major haplotype I, and f. sp. radicis-cucumerinum to AFLP major haplotype II. Overall, pathogenicity, vegetative compatibility grouping and AFLP analysis were correlated and effectively distinguished isolates of F. oxysporum from melon. This appears to be the first report of natural infection of melon by F. oxysporum f. sp. radicis-cucumerinum worldwide.     

Rapid Detection of the Fusarium oxysporum Lineage Containing the Canary Island Date Palm Wilt Pathogen

ABSTRACT Fusarium oxysporum f. sp. canariensis causes Fusarium wilt disease on the Canary Island date palm (Phoenix canariensis). To facilitate disease management, a polymerase chain reaction diagnostic method has been developed to rapidly detect the pathogen. A partial genomic library of F. oxysporum f. sp. canariensis isolate 95-913 was used to identify a DNA sequence diagnostic for a lineage containing all tested isolates of F. oxysporum f. sp. canariensis. Two oligonucleotide primers were designed and used to amplify a 567-bp fragment with F. oxysporum f. sp. canariensis DNAs. DNA from 61 outgroup isolates did not amplify using these primers. Once the primers were shown to amplify a 0.567-kb fragment from DNA of all the F. oxysporum f. sp. canariensis isolates tested, a rapid DNA extraction procedure was developed that led to the correct identification of 98% of the tested F. oxysporum f. sp. canariensis isolates.     

Development of a Specific Polymerase Chain Reaction-Based Assay for the Identification of Fusarium oxysporum f. sp. ciceris and Its Pathogenic Races 0, 1A, 5, and 6

ABSTRACT Specific primers and polymerase chain reaction (PCR) assays that identify Fusarium oxysporum f. sp. ciceris and each of the F. oxysporum f. sp. ciceris pathogenic races 0, 1A, 5, and 6 were developed. F. oxysporum f. sp. ciceris- and race-specific random amplified polymorphic DNA (RAPD) markers identified in a previous study were cloned and sequenced, and sequence characterized amplified region (SCAR) primers for specific PCR were developed. Each cloned RAPD marker was characterized by Southern hybridization analysis of Eco RI-digested genomic DNA of a subset of F. oxysporum f. sp. ciceris and nonpathogenic F. oxysporum isolates. All except two cloned RAPD markers consisted of DNA sequences that were found highly repetitive in the genome of all F. oxysporum f. sp. ciceris races. F. oxysporum f. sp. ciceris isolates representing eight reported races from a wide geographic range, nonpathogenic F. oxysporum isolates, isolates of F. oxysporum f. spp. lycopersici, melonis, niveum, phaseoli, and pisi, and isolates of 47 different Fusarium spp. were tested using the SCAR markers developed. The specific primer pairs amplified a single 1,503-bp product from all F. oxysporum f. sp. ciceris isolates; and single 900- and 1,000-bp products were selectively amplified from race 0 and race 6 isolates, respectively. The specificity of these amplifications was confirmed by hybridization analysis of the PCR products. A race 5-specific identification assay was developed using a touchdown-PCR procedure. A joint use of race 0- and race 6-specific SCAR primers in a single-PCR reaction together with a PCR assay using the race 6-specific primer pair correctly identified race 1A isolates for which no RAPD marker had been found previously. All the PCR assays described herein detected up to 0.1 ng of fungal genomic DNA. The specific SCAR primers and PCR assays developed in this study clearly identify and differentiate isolates of F. oxysporum f. sp. ciceris and of each of its pathogenic races 0, 1A, 5, and 6.