Effects of Varying Environmental Conditions on Biological Control of Fusarium Wilt of Tomato by Nonpathogenic Fusarium spp

ABSTRACT The influence of varying environmental and cropping conditions including temperature, light, soil type, pathogen isolate and race, and cultivar of tomato on biological control of Fusarium wilt of tomato by isolates of nonpathogenic Fusarium oxysporum (CS-20 and CS-24) and F. solani (CS-1) was evaluated in greenhouse and growth chamber experiments. Liquid spore suspensions (10(6)/ml) of the biocontrol isolates were applied to soilless potting mix at the time of tomato seeding, and the seedlings were transplanted into pathogen-infested field soil 2 weeks later. Temperature regimes ranging from 22 to 32 degrees C significantly affected disease development and plant physiological parameters. Biocontrol isolate CS-20 significantly reduced disease at all temperature regimes tested, yielding reductions of disease incidence of 59 to 100% relative to pathogen control treatments. Isolates CS-24 and CS-1 reduced disease incidence in the greenhouse and at high temperatures, but were less effective at the optimum temperature for disease development (27 degrees C). Growing plants under shade (50% of full light) versus full light affected some plant growth parameters, but did not affect the efficacy of biocontrol of any of the three bio-control isolates. Isolate CS-20 effectively reduced disease incidence (56 to 79% reduction) in four different field soils varying in texture (sandy to clayey) and organic matter content (0 to 3.2%). Isolate CS-1 reduced disease in the sandy and loamy soils (49 to 66% reduction), but was not effective in a heavy clay soil. Both CS-1 and CS-20 were equally effective against all three races of the pathogen, as well as multiple isolates of each race (48 to 66% reduction in disease incidence). Both isolates, CS-1 and CS-20, were equally effective in reducing disease incidence (66 to 80% reduction) by pathogenic races 1, 2, and 3 on eight different tomato cultivars containing varying levels of inherent resistance to Fusarium wilt (susceptible, resistant to race 1, or resistant to races 1 and 2). These results demonstrate that both these Fusarium isolates, and particularly CS-20, can effectively reduce Fusarium wilt disease of tomato under a variety of environmental conditions and have potential for further development.     

Induction of systemic disease resistance and pathogen defence responses in Asparagus officinalis inoculated with nonpathogenic strains of Fusarium oxysporum

The ability of nonpathogenic isolates of Fusarium oxysporum (np Fo ) to induce systemic resistance and defence responses against subsequent challenge with a pathogenic strain of F. oxysporum f. sp. asparagi ( Foa ) was examined in Asparagus officinalis . In a split-root experiment, roots inoculated with np Fo exhibited a hypersensitive response and those subsequently inoculated with Foa displayed resistance. Induction of systemic resistance in np Fo -treated plants led to significantly fewer necrotic lesions ( P  = 0·05) and reduced Foa disease severity compared with plants not treated with np Fo . In hyphal-sandwich root inoculation experiments, activities of peroxidase and phenylalanine ammonia-lyase and lignin content were higher in np Fo -treated plants and increased more rapidly than in np Fo -untreated plants after Foa inoculation. Antifungal activity (inhibition of fungal spore germination and germ-tube growth) from exudates of roots inoculated with Foa were observed for np Fo -treated plants but not for np Fo -untreated plants. Thus, isolates of np Fo may function as inducers of systemic acquired resistance (SAR) and defence responses against Foa invasion in A. officinalis .     

Efficacy and dose–response relationship in biocontrol of Fusarium disease in maize by Streptomyces spp.

Two isolates of Streptomyces spp. DAUFPE 11470 and DAUFPE 14632 were evaluated to determine the antagonist–pathogen inoculum concentration relationship under greenhouse conditions. Pathogen and antagonist concentration, significantly (P < 0.05) affected development of Fusarium disease in maize with a significant interaction between pathogen and antagonist concentration. Dose–response relationship also differed significantly (P < 0.05) between the two isolates, but both isolates demonstrated effective control of Fusarium disease, regardless of pathogen concentration. The isolate DAUFPE 11470 provided the most effective control. The lowest value for disease incidence occurred at low pathogen (10³ chlamydospore g⁻¹ soil) and high antagonist concentration (10⁶ cfu ml⁻¹) for both isolates. The disease incidence for control plants ranged from 19% to 76%. However, in relation to control the lowest disease reduction occurred at low pathogen (10³ chlamydospore g⁻¹ soil) and high antagonist concentrations (10⁶ cfu ml⁻¹). These reductions were 10.6% and 13% for DAUFPE 14632 and DAUFPE 11470, respectively. The highest disease reductions, in relation to control plants, occurred at high pathogen (10⁶ chlamydospore g⁻¹ soil) and antagonist (10⁶ cfu ml⁻¹) concentrations for both isolates. These values were 55% and 62.2% for DAUFPE 14632 and DAUFPE 11470, respectively. The chlamydospore germination of Fusarium moniliforme was affected by glucose addition, antagonist isolates and type of inoculum. The lowest chlamydospore germination was observed with bacterial suspensions of the isolates, for all glucose additions. The results suggested that both Streptomyces spp. isolates were effective at different doses as biocontrol agents against F. moniliforme. Also, there was evidence for at least two mechanisms of biocontrol and that apparently, both isolates showed the same mechanisms of biocontrol action related to production of bioactive compounds and competition for carbon. Further studies will be developed to improve the level and effectiveness of control by these isolates.     

Biological Control Efficiency of Fusarium Wilt of Tomato by Nonpathogenic Fusarium oxysporum Fo-B2 in Different Environments

ABSTRACT Efficiency of nonpathogenic Fusarium oxysporum Fo-B2 for the biological control of Fusarium wilt of tomato, caused by F. oxysporum f. sp. lycopersici CU1, was examined in different environments: a growth chamber with sterile soil-less medium, a greenhouse with fumigated or nonfumigated soil, and nonfumigated field plots. Inoculation of Fo-B2 onto tomato roots significantly reduced the severity of disease, but the efficiency of disease suppression decreased as the experimental environment became less controlled. Relationships between the recovery of Fo-B2 from hypocotyls and the disease severity indicated that the biocontrol agent was most effective when it colonized vascular tissues intensively. Moreover, the degree of Fo-B2 colonization was greatly reduced when the seedlings were grown in nonfumigated soil. Dose-response models (negative exponential, hyperbolic saturation, and logistic) were fit to observed data collected over a range of inoculum densities of the pathogen and the antagonist; the logistic model provided the best fit in all environments. The ratios of an 50% effective dose parameter for Fo-B2 to that of CU1 increased as the environment became less controlled, suggesting that environmentally related efficiency reduction impacted the antagonist more than the pathogen. The results suggest that indigenous soil microbes were a primary factor negatively influencing the efficiency of Fo-B2. Therefore, early establishment of the antagonist in a noncompetitive environment prior to outplanting could improve the efficacy of biological control.     

Recovery of Fusarium oxysporum Fo47 Mutants Affected in Their Biocontrol Activity After Transposition of the Fot1 Element

ABSTRACT To investigate the biocontrol mechanisms by which the antagonistic Fusarium oxysporum strain Fo47 is active against Fusarium wilt, a Fot1 transposon-mediated insertional mutagenesis approach was adopted to generate mutants affected in their antagonistic activity. Ninety strains in which an active Fot1 copy had transposed were identified with a phenotypic assay for excision and tested for their biocontrol activity against F. oxysporum f. sp. lini on flax in greenhouse experiments. Sixteen strains were affected in their capacity to protect flax plants, either positively (more antagonistic than Fo47) or negatively (less antagonistic). The molecular characterization of these mutants confirms the excision of Fot1 and its reinsertion in most of the cases. Moreover, we demonstrate that other transposable elements such as Fot2, impala, and Hop have no transposition activity in the mutant genomes. The phenotypic characterization of these mutants shows that they are affected neither in their in vitro growth habit nor in their competitiveness in soil compared with wild-type strain Fo47. These results show that mutants are not impaired in their saprophytic phase and suggest that the altered biocontrol phenotype should likely be expressed during the interaction with the host plant.     

Potential role for salicylic acid in induced resistance of asparagus roots to Fusarium oxysporum f.sp. asparagi

Pre-inoculation of asparagus ( Asparagus officinalis ) roots with selected nonpathogenic isolates of Fusarium oxysporum (np Fo ) has previously been shown to induce systemic resistance against infection by F. oxysporum f.sp. asparagi ( Foa ) through activation of plant-defence mechanisms. To elucidate the putative np Fo -mediated defence pathways, the effect of salicylic acid (SA) was examined in a split-root system of asparagus where one half of the seedling root system was drenched with SA and the activation of defence responses was measured subsequently on the remaining roots. SA-treated plants exhibited enhanced systemic resistance, with a significant reduction in disease severity of the roots inoculated with Foa , compared with untreated plants. SA activated peroxidase and phenylalanine ammonia-lyase, as well as lignification, upon Foa attack, in a manner similar to that observed with np Fo pretreatment. In addition, application of diphenyleneiodonium, an SA biosynthesis inhibitor, led to failure of np Fo to induce lignin deposition and systemic resistance. Treatment of fungal spores with SA did not affect germination and growth of either np Fo or Foa in in vitro antifungal assays. Production of SA at the site of np Fo infection may be involved in the induction of Foa resistance in asparagus roots.     

Remodelling of actin cytoskeleton in tomato cells in response to inoculation with a biocontrol strain of Fusarium oxysporum in comparison to a pathogenic strain

Knowing that actin microfilaments play a key role in the mobilization of the defence‐associated cellular responses, the aim of this study was to compare changes affecting the actin cytoskeleton in tomato cells after inoculation with germinated microconidia of a biocontrol (Fo47, Fom24) or a pathogenic (Fol8) strain of Fusarium oxysporum. Actin microfilaments were observed by labelling with TRITC‐phalloidin combined with fluorescence microscopy. Results showed that only tenuous changes in the actin cytoskeleton architecture occurred after inoculation with the biocontrol strains whereas the actin cytoskeleton was significantly altered after inoculation with the pathogenic one. In the two types of interaction, cell death occurs and can be considered as one key component of cell defence responses. A pharmacological approach using cytochalasins was chosen to determine whether the inhibition of actin polymerization differently affects the kinetics of tomato cell death. Data showed that cytochalasins reduced cell death induced by the biocontrol strain Fo47, and in contrast, increased cell death induced by the pathogenic strain Fol8, suggesting that the pathway leading to cell death differs in the protective and compatible interactions.     

Implication of Systemic Induced Resistance in the Suppression of Fusarium Wilt of Tomato by Pseudomonas fluorescens WCS417r and by Nonpathogenic Fusarium oxysporum Fo47

Fluorescent pseudomonads and nonpathogenic Fusarium oxysporum have been shown to suppress fusarium wilts. This suppression has been related to both microbial antagonism and induced resistance.The aim of the present study was to assess the relative importance of systemic induced resistance in the suppression of fusarium wilt of tomato in commercial-like conditions by a reference strain of each type of microorganism (P. fluorescens WCS417r and nonpathogenic F. oxysporum Fo47). The spatial separation of the pathogen and the biocontrol strains excluded any possible microbial antagonism and implicated the involvement of the systemic induced resistance; whereas the absence of any separation between these microorganisms allowed the expression of both mechanisms. Since systemic induced resistance has often been associated with the synthesis of PR-proteins, their accumulation in tomato plants inoculated with WCS417r or with Fo47 was determined.The analysis of the results indicates that the suppression of fusarium wilt by P. fluorescens WCS417r was ascribed to systemic induced resistance without any detection of the PR-proteins tested (PR-1 and chitinases). In contrast, the suppression achieved by nonpathogenic F. oxysporum Fo47 appeared to be mainly ascribed to microbial antagonism but also to a lesser extent to systemic induced resistance. This induced resistance could be related to the accumulation of PR-1 and chitinases.The possible relationship between the ability of Fo47 to suppress fusarium wilt more efficiently than WCS417r and its ability to show both mechanisms is discussed.     

Cytological Analysis of Defense-Related Mechanisms Induced in Pea Root Tissues in Response to Colonization by Nonpathogenic Fusarium oxysporum Fo47

The ability of nonpathogenic Fusarium oxysporum, strain Fo47, to trigger plant defense reactions was investigated using Ri T-DNA-transformed pea roots. Cytological investigations of strain Fo47-inoculated roots showed that the fungus grew actively at the root surface and colonized a number of epidermal and cortical cells, inducing marked host cell metabolic changes. In roots inoculated with pathogenic F. oxysporum f. sp. pisi, the pathogen multiplied abundantly through much of the tissues, whereas in Fo47-inoculated roots, fungal growth was restricted to the epidermis and the outer cortex. Invading cells of strain Fo47 suffered from serious alterations, a phenomenon that was not observed in control roots in which F. oxysporum f. sp. pisi grew so actively that the vascular stele was invaded within a few days. Strain Fo47 establishment in the root tissues resulted in a massive elaboration of hemispherical wall appositions and in the deposition of an electron-opaque material frequently encircling pathogen hyphae and accumulating in the noninfected xylem vessels. This suggests that the host roots were signaled to defend themselves through the rapid stimulation of a general cascade of nonspecific defense responses. The specific relationship established between strain Fo47 and the root tissues is discussed in relation to other types of plant-fungus interactions, including pathogenic and symbiotic associations.     

The potential of nonpathogenic Fusarium oxysporum and other biological control organisms for suppressing fusarium wilt of banana

The aim of this study was to evaluate the ability of nonpathogenic F. oxysporum and Trichoderma isolates from suppressive soils in South Africa to suppress fusarium wilt of banana in the glasshouse. Several biological control agents and commercial biological control products were included in the study. The isolates were first screened in vitro on potato dextrose agar. In glasshouse evaluations, the fungal and bacterial isolates were established on banana roots before they were replanted in pathogen-infested soil, while the commercial biocontrol agents were applied as directed by the supplier. Banana plantlets were evaluated for disease development after 7 weeks. In vitro tests showed none of the nonpathogenic isolates suppressed Fusarium oxysporum f.sp. cubense ( Foc ), while slight suppression was observed with the two Trichoderma isolates. Results of the glasshouse evaluations revealed that two of the nonpathogenic F. oxysporum isolates, CAV 255 and CAV 241, reduced fusarium wilt incidence by 87·4 and 75·0%, respectively. The known biological control agent Fo47 did not suppress Foc significantly. Pseudomonas fluorescens strain WCS 417, known for its ability to suppress other fusarium wilt diseases (WCS 417), reduced disease incidence by 87·4%. These isolates should be further evaluated for potential application in the field, independently and in combination.     

产抗菌素2,4-二乙酰基藤黄酚假单胞杆菌的分离及其防治沙打旺根腐病的室内测定

从山东、内蒙古、北京等地土壤中分离筛选出荧光菌 50 0 0余株 ,其中 1 2 0 0余株为抑制性荧光菌。经PCR检测 ,获得 73株 2 ,4 二乙酰基藤黄酚 (2 ,4 DAPG)产生菌。平板筛选结果表明 ,2 ,4 DAPG产生菌CPF 1 0和 2P8对沙打旺根腐病菌Sad1和Sad2均有较好的抑制效果 ,其中CPF 1 0的抑菌带宽分别为 5.0和 1 2 .0mm ;2P8抑菌带宽分别为 3 .5和 7.0mm。温室试验两次调查表明 ,CPF 1 0对沙打旺根腐病防治效果最好 ,达 63 .5%和 67.8% ;2P8防效也在 40 %左右 ,且均达到极显著水平。 2 ,4 DAPG产生菌可以显著促进沙打旺植株根系发育 ,CPF 1 0处理后地上部株高差异不显著 ,但鲜重和干重与对照相比有极显著的增加 ,说明两菌株菌剂处理可以促进植株生长。根部定殖结果表明 ,两菌株在沙打旺根部都有一定的定殖能力 ,在根表种群数量比较稳定 ,根内细菌数量在调查时间内呈逐渐上升的趋势     

Gene genealogies support Fusarium oxysporum f. sp. ciceris as a monophyletic group

Fusarium oxysporum f. sp. ciceris (Foc), the causal agent of fusarium wilt of chickpea, consists of two pathotypes (yellowing and wilting) and eight races (races 0, 1B/C, 1A and 2–6) of diverse geographical distribution. Six Foc isolates, one each of races 0, 1B/C, 1A, 4, 5 and 6, representing the two pathotypes and the geographical range of the pathogen, showed identical sequences in introns of the genes for translation elongation factor 1α ( EF1 α), β-tubulin, histone 3, actin and calmodulin. Eleven additional Foc isolates representative of all races, pathotypes and geographical range, and three isolates of F. oxysporum (Fo) nonpathogenic to chickpea were further analysed for sequence variation in the EF1 α gene. All isolates pathogenic to chickpeas shared an identical EF1 α gene sequence, which differed from that shared by the three Fo isolates nonpathogenic to chickpea. EF1 α gene sequences from the 17 Foc isolates and the three Fo isolates were compared with 24 EF1 α gene sequences in GenBank from isolates of 11 formae speciales of F. oxysporum by parsimony analysis. Foc isolates formed a grouping distinct from other formae speciales and nonpathogenic isolates. These results indicate that F. oxysporum f. sp. ciceris is monophyletic.     

Modes of action of the protective strain Fo47 in controlling verticillium wilt of pepper

The protective fungus Fusarium oxysporum Fo47 reduces the severity of wilt caused by the soilborne pathogen Verticillium dahliae in pepper. Modes of action responsible for the biocontrol activity were studied. Microscopic observations of fluorescent protein‐transformed strains colonizing the root surface show that the colonization patterns of Fo47 and V. dahliae were similar. Pixel counting of the images obtained by confocal microscopy showed that Fo47 reduces colonization of the root surface by V. dahliae, suggesting a possible role of competition for nutrients at the root surface. Besides these effects on surface colonization, the hormonal pathways activated during priming of plant defence responses were identified by measuring the amount of some phytohormones and their derivatives in roots and stems of pepper. Results showed an early, slight increase of jasmonyl isoleucine, followed by a transient increase of salicylic acid during the pre‐challenged phase of priming and an increase of 12‐oxo‐phytodienoic acid during the challenge phase of priming. The caffeic, ferulic and chlorogenic acids, known to play a role in plant defence reactions, showed a strong antimicrobial activity against V. dahliae in vitro. In pepper roots, Fo47 stimulated the biosynthesis of caffeic acid and primed that of chlorogenic acid. These results demonstrated that the effective control of V. dahliae provided by Fo47 is based on different but complementary mechanisms.     

Differential Colonization of Tomato Roots by Nonpathogenic and Pathogenic Fusarium oxysporum Strains May Influence Fusarium Wilt Control

ABSTRACT Histochemical staining, beta-glucuronidase (GUS) activity, or placing roots on agar were methods used to characterize interactions between the pathogenic fungus, Fusarium oxysporum f. sp. lycopersici, and the nonpathogenic biocontrol F. oxysporum strain 70T01 with respect to colonization behaviors, interaction sites, and population densities on tomato roots. Mycelia of strain 70T01, a genetic transformant expressing stable GUS activity, hygromycin B resistance, and effective disease control, were localized in epidermal and cortex cell layers of tomato roots in a discontinuous and uneven pattern. In contrast, mycelia of F. oxysporum f. sp. lycopersici were found in the vascular bundles. Thus, direct interactions between the two fungi likely happen in the root surface cell layers. Colonization density of strain 70T01 was related to the inoculation density but decreased with distance from the inoculation site. Host defense reactions, including increased cell wall thickness or papilla deposits, were adjacent to 70T01 hyphae. Experiments done in soil showed that strain 70T01 densities in roots were highest at inoculation zones and barely detectable for root segments more than 2 cm away from the inoculation sites. F. oxysporum f. sp. lycopersici densities were lowest at 70T01 inoculation zones and highest (>10 times) where strain 70T01 was not directly applied. Newly elongating roots where strain 70T01 did not reach were available for infection by the pathogen. The higher strain 70T01 density was always found when the plants were simultaneously infected by F. oxysporum f. sp. lycopersici, suggesting that F. oxysporum f. sp. lycopersici has as much influence in predisposing the plant to colonization by strain 70T01 as strain 70T01 has on providing disease protection against the pathogen.     

Fusarium oxysporum Fo47 confers protection to pepper plants against Verticillium dahliae and Phytophthora capsici,and induces the expression of defence genes

The application of the nonpathogenic isolate Fusarium oxysporum 47 (Fo47) reduced the symptoms of verticillium wilt, phytophthora root rot and phytophthora blight in pepper plants. Botrytis cinerea was also tested on the leaves of plants treated with Fo47, but no protection was observed. Verticillium dahliae colonies cultured in the presence of Fo47 grew slower than control cultures, but Phytophthora capsici growth was unaffected by Fo47. At least part of the protection effect observed against V. dahliae could therefore be due to antagonism or competition. In order to search for induced resistance mechanisms, three defence genes previously related to pepper resistance were monitored over time. These genes encode a basic PR‐1 protein (CABPR1), a class II chitinase (CACHI2) and a sesquiterpene cyclase (CASC1) involved in the synthesis of capsidiol, a phytoalexin. These three genes were transiently up‐regulated in the roots by Fo47 in the absence of inoculation with the pathogen, but in the stem only CABPR1 was up‐regulated. In plants that were inoculated with V. dahliae after the Fo47 treatment, the three genes had a higher relative expression level than the control in both the roots and the stem.     

Systemically induced resistance and microbial competitive exclusion: implications on biological control

The root-knot nematode, Meloidogyne incognita, is among the most damaging agricultural pests, particularly to tomato. The mutualistic endophytes Fusarium oxysporum strain Fo162 (Fo162) and Rhizobium etli strain G12 (G12) have been shown to systemically induce resistance toward M. incognita. By using triple-split-root tomato plants, spatially separated but simultaneous inoculation of both endophytes did not lead to additive reductions in M. incognita infection. More importantly, spatially separated inoculation of Fo162 and G12 led to a reduction in Fo162 root colonization of 35 and 39% when G12 was inoculated on a separate root section of the same plant in two independent experiments. In an additional split-root experiment, spatial separation of Fo162 and G12 resulted in a reduction of Fo162 root colonization of approximately 50% over the water controls in two independent experiments. The results suggested that the suppressive activity of G12 on Fo162 and M. incognita is possibly related to the induction of specific plant defense mechanisms. Thus, although Fo162 and G12 have the ability to systemically repress M. incognita infection in tomato, they can be considered incompatible biocontrol agents when both organisms are present simultaneously on the same root system.     

Influence of Temperature and Inoculum Density of Fusarium oxysporum f. sp. ciceris on Suppression of Fusarium Wilt of Chickpea by Rhizosphere Bacteria

The effects of temperature and inoculum density of Fusarium oxysporum f. sp. ciceris race 5 on suppression of Fusarium wilt in chickpea (Cicer arietinum) cv. PV 61 by seed and soil treatments with rhizobacteria isolated from the chickpea rhizosphere were studied in a model system. Disease development over a range of temperatures (20, 25, and 30 degrees C) and inoculum densities (25 to 1,000 chlamydospores per gram of soil) was described by the Gompertz model. The Gompertz relative rate of disease progress and final amount of disease increased exponentially and monomolecularly, respectively, with increasing inoculum densities. Disease development was greater at 25 degrees C compared with 20 and 30 degrees C. At 20 and 30 degrees C, disease development was greater at 250 to 1,000 chlamydospores per gram of soil compared with 25 to 100 chlamydospores per gram of soil. At 25 degrees C, increasing inoculum densities of the pathogen did not influence disease. Nineteen Bacillus, Paenibacillus, Pseudomonas, and Stenotrophomonas spp. out of 23 bacterial isolates tested inhibited F. oxysporum f. sp. ciceris in vitro. Pseudomonas fluorescens RGAF 19 and RG 26, which did not inhibit the pathogen, showed the greatest Fusarium wilt suppression. Disease was suppressed only at 20 or 30 degrees C and at inoculum densities below 250 chlamydospores per gram of soil. Bacterial treatments increased the time to initial symptoms, reduced the Gompertz relative rate of disease progress, and reduced the overall amount of disease developed.     

Stem rot of jewel orchids caused by a new forma specialis,Fusarium oxysporum f. sp. anoectochili in Taiwan

Stem rot of Anoectochilus formosanus (Af) caused by Fusarium oxysporum (Fo) is a major limiting factor to jewel orchid production in Taiwan. Fo causes discoloration in vascular tissues. However, some newly collected Fo isolates from Af stem rot do not cause vascular discoloration, suggesting changes may have occurred in the pathogen. Among recent Fo isolates from Af there are two colony types, the cottony alba (CA) and the sporodochial (S). In order to confirm that both colony types cause Af stem rot, 200 isolates were obtained from diseased stems in Nantou County and characterized by colony type and whether or not the infected plants had vascular discoloration. Isolates of both the CA and S types caused stem rot of Af; some isolates in each colony type caused vascular discoloration whilst others did not. Pathogenicity tests with 22 isolates resulted in stem rot disease severity ratings on Af of 3·1–4·0 and 2·1–4·0 with CA and S type colonies, respectively. The same isolates failed to cause disease on Cattleya, Dendrobium or Phalaenopsis plants. Phylogenetic analysis of partial intergenic spacer sequences showed that these isolates were distinguishable from other formae speciales of Fo and could be separated into two groups correlated with the CA or S type colonies with high bootstrap. Based on pathogenic, morphological and molecular characterizations, the Fo that causes stem rot of Af is proposed to be a new forma specialis, F. oxysporum f. sp. anoectochili, with different pathotypes.     

Isolation and characterization of nonpathogenic Fusarium oxysporum isolates from the rhizosphere of healthy banana plants

One of the most serious diseases of banana is fusarium wilt, caused by Fusarium oxysporum f.sp. cubense ( Foc ). The objectives of this study were to isolate and identify nonpathogenic F. oxysporum strains from soils suppressive to banana wilt, and to determine the diversity of these isolates. More than 100 Fusarium strains were isolated from the rhizosphere of banana plants and identified to species level. Pathogenicity testing was carried out to confirm that these isolates were nonpathogens of banana. A PCR-based RFLP analysis of the intergenic spacer region of the ribosomal RNA operon was used to characterize the nonpathogens. The isolates were also compared with isolates of Foc from South Africa and the known biological control isolate of F. oxysporum , Fo47. The species-specific primers FOF1 and FOR1, in addition to morphological features, were used to confirm the identity of F. oxysporum isolates included in the PCR-RFLP analysis. Twelve different genotypes could be distinguished, identified by a six-letter code allocated to each isolate following digestion with the restriction enzymes Hae III, Hha I, Hin fI, Msp I, Rsa I and Scrf I. Eleven of these included nonpathogenic F. oxysporum isolates, and these groups could all be distinguished from the genotype that included Foc . Fo47 was included in one of the genotype groups consisting of nonpathogenic F. oxysporum isolates from South Africa.