Microsatellite marker-based detection of Fusarium wilt pathogens of Psidium guajava L.

Wilt of Psidium guajava L., incited by Fusarium oxysporum f. sp. psidii and Fusarium solani is a serious soil-borne disease of guava in India. Forty-two isolates each of F. oxysporum f. sp. psidii (Fop) and F. solani (Fs) collected from different agro climatic zones of India showing pathogenicity were subjected to estimate the genetic and molecular characterisation in terms of analysis of microsatellite marker studies. Out of eight microsatellite markers, only four microsatellite markers, viz. MB 13, MB 17, RE 102 and AY212027 were amplified with single band pattern showing the character of identical marker for molecular characterisation and genetic identification. Microsatellite marker MB 13 was amplified in F. oxysporum f. sp. psidii and F. solani isolates. Product size of 296 bps and 1018 bps were exactly amplified with a single banding pattern in all the isolates of F. oxysporum f. sp. psidii and F. solani, respectively. Microsatellite markers, viz. MB 17, RE 102 and AY212027 were also exactly amplified with a single banding pattern. MB 17 was amplified in F. oxysporum f. sp. psidii isolates with a product size of 300 bp. RE 102 and AY212027 were amplified in F. solani isolates with the product size of 153 bp and 300 bp, respectively. Therefore, amplified microsatellite marker may be used as identifying DNA marker.     

Studies on a species of Monosporascus isolated from triticum

Hyphal parasitic behaviour of Fusarium oxysporum on Rhizoctonia solani and consecutive changes during this phenomenon have been investigated and studied. The hyphal parasitism was very commonly recorded between the test fungi. During the course of parasitism coiling, penetration, lysis and formation of chlamydospores by F. oxysporum on R. solani were observed. R. solani is a new host record for the mycoparasite F. oxysporum.     

Prevalence of wilt-root rot and their associated pathogens at reproductive phase in lentil

A survey of 116 districts of nine lentil growing states covering 603 farmers' fields revealed a range of 0.7–9.3% mean plant mortality at reproductive stages in different lentil growing states of the country. The overall mean mortality was 6.3%. The main pathogens found associated with plant mortality at this stage were Fusarium oxysporum f. sp. lentis (62.0%), Rhizoctonia bataticola (25.2%) and Sclerotium rolfsii (9.8%). The minor involvement of 1.8% was that of F. solani, F. chlamydosporum. F. equisetii, and R. solani. For the first time a national scenario of lentil wilt-root rot incidence at the crucial reproductive stage and their associated pathogens is reported here.     

Mycoparasitism ofFusarium SPP. onRhizoctonia solani Kühn

Summary Experiments on nutrient and staled agar were carried out to investigate the mycoparasitic activity of some fusaria againstRhizoctonia solani Kühn. Penetration and coiling byFusarium oxysporum Sch.,F. semitectum Berk & Rav. andF. udum Butler in and around theR. solani hyphae was observed. Lysis ofF. udum hyphae was observed inside theR. solani hyphae showing the reverse of the normal direction of necrotrophic mycoparasitic relationships. The mycoparasitic activity ofFusarium spp. was much affected in staled agar plates.     

Biological Control ofMonilinia laxaandFusarium oxysporumf. sp.lycopersiciby a Lytic Enzyme-ProducingPenicillium purpurogenum

Biological control experiments were conducted with the lytic enzyme-producing fungusPenicillium purpurogenumagainst the plant pathogensMonilinia laxaandFusarium oxysporumf. sp.lycopersici.Applications ofP. purpurogenumto peach shoots previously inoculated withM. laxareduced lesion length and extent of pathogen colonization of shoots by 90 and 80% (P ≤ 0.05), respectively, comparable to the level of disease control obtained with the fungicide captan. Disease severity in tomato plants inoculated withF. oxysporumf. sp.lycopersiciwas decreased by 30% (P ≤ 0.05) with the biological treatment. The fungusP. purpurogenumproduced β-1,3-glucanase and chitinase activities in liquid culture that were inducible by cell walls and live mycelium ofM. laxabut not ofF. oxysporumf. sp.lycopersici.Crude filtrates or crude enzyme preparations ofP. purpurogenumcultures with lytic enzyme activities produced lysis of hyphae and spores ofM. laxaandF. oxysporumf. sp.lycopersici.These lytic effects were strong inM. laxaand ended in complete dissolution of mycelium. The induction of lytic enzymes byM. laxaand the effects of lytic enzymes on mycelia of the pathogens in relation to the different degrees of biological control obtained are discussed.     

Enzymes Associated with Defence against Toxic Oxygen Species in PCNB-Tolerant and Sensitive Soil Fungi

The specific activities of enzymes such as superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPOX) and glutathione reductase (GR), which are involved in protection against toxic species of oxygen, were determined in mycelia extracts of pentachloronitrobenzene (PCNB)-tolerant and susceptible soil fungi. The organisms assayed were the highly PCNB-sensitive Rhizoctonia solani and Rhizopus arrhizus; Sclerotium rolfsii and Trichoderma harzianum, which are moderately susceptible to PCNB, and the fungicide-tolerant Fusarium oxysporum f. sp. melonis and Pythium aphanidermatum. No GPOX activity was detected in the six examined fungi. Significant differences in the specific activities of the other enzyme systems among the fungi were evident. Remarkably low levels of CAT activities were measured in R. solani. Except for T. harzianum, no meaningful differences regarding SOD, CAT and GR activities with age of the fungi cultures were observed. The electrophoretic patterns of SOD and CAT displayed dissimilarities among the fungi under study. P. aphanidermatum is more polymorphic with respect to both SOD and CAT enzyme systems as compared to the other fungi. The SOD of F. oxysporum f. sp. melonis, R. arrhizus and T. harzianum is a cuprozinc enzyme, while the mangano-SOD species was detected in S. rolfsii, R. solani and T. harzianum.     

Cross-pathogenicity between Formae Speciales of Fusarium oxysporum, the Pathogens of Cucumber and Melon

The population structure of Fusarium oxysporum f. sp. cucumerinum was studied using the vegetative compatibility grouping (VCG) approach. All 37 of the examined isolates from Israel were assigned to VCG 0180, the major VCG found in North America and the Mediterranean region. Approximately two‐thirds of the tested isolates were pathogenic to both cucumber and melon, but cumulatively they were more aggressive on cucumber, their major host, than on melon. Disease symptoms on melon plants were less destructive and often expressed as growth retardation. Melon cultivars differing in Fom genes for resistance to F. oxysporum f. sp. melonis were inoculated with three isolates of F. oxysporum f. sp. cucumerinum. Results showed that Fom genes do not confer resistance to F. oxysporum f. sp. cucumerinum, although different horticultural types may respond differently to this pathogen. The reciprocal inoculation of F. oxysporum f. sp. melonis on cucumber, using four physiological races, did not result in disease symptoms or growth retardation. It is concluded that cucumerinum and melonis should remain two distinct formae speciales.     

Effect of the high polycyclic aromatic hydrocarbon, benzo[a]pyrene, on the lipid content of Fusarium solani

The purpose of the present paper was to study the effect of the high polycyclic aromatic hydrocarbon (PAH), benzo[a]pyrene, on the lipid [fatty acid (FA) and sterol] composition and content of the fungi Fusarium solani and F. oxysporum, respectively recognized as good and poor PAH degraders. The major FAs and the major sterol that characterized the tested Fusarium strains were C16:0, C18:1, C18:2, and ergosterol. Lipid profiles of F. solani remained unchanged with the addition of benzo[a]pyrene in the culture media at all concentrations and duration of treatment. However, in the presence of benzo[a]pyrene, significant decreases in FA content, which reached 18 % in young cultures and 28 % in mature colonies, were registered. Similarly, the sterol content of F. solani was reduced by 27 % in the presence of benzo[a]pyrene. In contrast, no modification in lipid profile and lipid content were observed with F. oxysporum, a strain recognized as a low benzo[a]pyrene degrader.     

Host perception of jasmonates promotes infection by Fusarium oxysporum formae speciales that produce isoleucine‐ and leucine‐conjugated jasmonates

Three pathogenic forms, or formae speciales (f. spp.), of Fusarium oxysporum infect the roots of Arabidopsis thaliana below ground, instigating symptoms of wilt disease in leaves above ground. In previous reports, Arabidopsis mutants that are deficient in the biosynthesis of abscisic acid or salicylic acid or insensitive to ethylene or jasmonates exhibited either more or less wilt disease, than the wild‐type, implicating the involvement of hormones in the normal host response to F. oxysporum. Our analysis of hormone‐related mutants finds no evidence that endogenous hormones contribute to infection in roots. Mutants that are deficient in abscisic acid and insensitive to ethylene show no less infection than the wild‐type, although they exhibit less disease. Whether a mutant that is insensitive to jasmonates affects infection depends on which forma specialis (f. sp.) is infecting the roots. Insensitivity to jasmonates suppresses infection by F. oxysporum f. sp. conglutinans and F. oxysporum f. sp. matthioli, which produce isoleucine‐ and leucine‐conjugated jasmonate (JA‐Ile/Leu), respectively, in culture filtrates, whereas insensitivity to jasmonates has no effect on infection by F. oxysporum f. sp. raphani, which produces no detectable JA‐Ile/Leu. Furthermore, insensitivity to jasmonates has no effect on wilt disease of tomato, and the tomato pathogen F. oxysporum f. sp. lycopersici produces no detectable jasmonates. Thus, some, but not all, F. oxysporum pathogens appear to utilize jasmonates as effectors, promoting infection in roots and/or the development of symptoms in shoots. Only when the infection of roots is promoted by jasmonates is wilt disease enhanced in a mutant deficient in salicylic acid biosynthesis.     

Phytopathogenic Fusarium oxysporum Schlecht, as a Necrotrophic Mycoparasite

Fusarium oxysporum f. sp. dianthi, f. sp. lycopersici, f. sp. cepae, f. sp. niveum and one unidentified F. oxysporum isolate proved to be active necrotrophic mycoparasites. In dual cultures hyphae of Trichoderma hamatum, T. longibrachiatum, T. pseudokoningii, T. harzianum, Botrytis cinerea and Rhizoctonia solani were parasitized and destroyed by F. oxysporum. One isolate of Phytophthora sp. was not affected. Mutual parasitism between F. oxysporum and T. pseudokoningii and T. longibrachiatum has been observed, too. Details of parasitic hyphal interactions: hyphal coiling, penetration sites, resistance sheat formation, hyphal invasion and internal growing are described. The mycoparasitic feature as well as antimicrobial metabolic production of F. oxysporum is probably a common phenomenon to ensure this important plant pathogenic species to compete successfully against other soil-borne fungal pathogens and saprophytes.     

Identification and evaluation of a potential biocontrol agent,Bacillus subtilis,against Fusarium sp. in apple seedlings

To find a potential biocontrol agent against Fusarium sp. in apple seedlings, an endophytic bacterium strain was isolated from apple tree tissues. The inhibitive efficiency of the isolated strain against the hyphal growth of Fusarium sp. and Rhizoctonia solani was tested. Strain Y-1 showed significant inhibitory effects against Fusarium oxysporum, F. moniliforme, F. proliferatum, F. solani and R. solani. Its antifungal activity against F. oxysporum was the highest, reaching up to 64.90 %. In vivo tests indicated that strain Y-1 effectively protects apple from F. oxysporum infections. The control effect reached 92.26 % when bacterial inoculation was performed 3 days prior to pathogen inoculation. Strain Y-1 could colonize the rhizosphere and tissues within 30 days. It was also able to induce systemic resistance in apple seedlings as shown by the activities of SOD and POD. Strain Y-1 significantly increased the root length, root wet and dry weights, and plant height of the apple seedlings compared with the control group. The homology analysis of the 16S rRNA sequence, together with morphological, physical, and biochemical analyses, revealed that strain Y-1 is Bacillus subtilis.     

Effect of mycorrhiza and biofertilisers on reducing the incidence of Fusarium root and pod rot diseases of peanut

Pathogenicity tests of twenty-six fungal isolates were tested on peanut plants (Giza 5 cv.) and the results revealed that, Fusarium oxysporum isolate (No. I) followed by F. solani (No. II) then F. moniliforme (No III) significantly caused highest incidence of root rot disease. Also, F. moniliforme (No III) followed by F. solani (No II) then F. oxysporum (No I) gave the highest incidence of pod rot disease. The effectiveness of vescular arbuscular-mycorrhiza (VAM) at different application rates on the incidence of root rot, pod rot diseases and plant growth parameters of peanut was studied. All soil treatments with each rate of VAM significantly reduced root and pod rot diseases compared with control (rate 0%). The best reduction in the severity of both diseases with VAM was found at the rate of 3%. Application of rhizobacterin, microbin and cerialin biofertilisers at the different concentrations decreased the severity of both root rot and pod rot severity diseases compared with non-treated seeds. The greatest reduction in both diseases was achieved at a concentration of 8/100?g seeds. The highest number of pods and fresh weight (g) was achieved in seed supplemented with each biofertiliser at concentration of 8/100?g seed.     

Decomposition of cellulose strips in relation to climate,litterfall nitrogen,phosphorus and C/N ratio in natural boreal forests

Isolates of Alternaria alternata, Botrytis cinerea, Fusarium oxysporum, Penicillium sp., Rhizoctonia solani, Stemphylium sp., Thielaviopsis basicola, and Verticillium dahliae were cultured on potato–dextrose agar (PDA), barley-sand and alfalfa-sand substrates in petri-dish or in column microcosms. N-mineralization by fungi and fungal-feeding nematodes in combination or fungi alone was assessed. Numbers of Aphelenchus avenae or Aphelenchoides composticola supported by the fungi were measured every 7 days. Times for full colonization of the substrates by fungi ranged from 5 to 15 days. Rhizoctonia solani and B. cinerea on PDA supported the largest A. avenae and A. composticola populations, respectively. Penicillium sp. was a nonhost for A. composticola and A. avenae. Rhizoctonia solani, B. cinerea, V. dahliae, and F. oxysporum supported significantly more nematodes than the other four fungal species. The ranked order of fungi based on the amount of N mineralized in columns free of nematodes was A. alternata (with a rate of 0.052 μg N/g-sand per day), Stemphylium sp., V. dahliae, T. basicola, B. cinerea, F. oxysporum, R. solani, and Penicillium sp. (with a rate of 0.0045 μg N/g-sand perday). The presence of A. avenae resulted in significant increases in mineral N, compared to nematode-free columns colonized by F. oxysporum, R. solani, and T. basicola alone. The presence of A. composticola resulted in significant increases in mineral N, compared to nematode-free columns colonized by A. alternata, B. cinerea, F. oxysporum, and R. solani alone. There was more mineral N incolumns in the presence of A. composticola than A. avenae in most cases. This revised version was published online in August 2006 with corrections to the Cover Date.     

The Effect of Pseudomonas fluorescens and Fusarium oxysporum f.sp. cubense on Induction of Defense Enzymes and Phenolics in Banana

The effect of Pseudomonas fluorescens treatment and Fusarium oxysporum f. sp. cubense inoculation on induction of phenylalanine ammonia-lyase (PAL), peroxidase (POX), chitinase, -1,3-glucanase and accumulation of phenolics in banana (Musa sp.) was studied. When banana roots were treated with P. fluorescens strain Pf10, a two-fold increase in phenolic content in leaf tissues was recorded 3 – 6 d after treatment. Challenge inoculation with F. oxysporum, the wilt pathogen, steeply increased the phenolic content in P. fluorescens-treated banana plants. Significant increase in POX activity was detected 6 – 9 d after P. fluorescens treatment. PAL, chitinase and -1,3-glucanase activities increased significantly from 3 d after P. fluorescens treatment and reached the maximum 6 d after treatment. Challenge inoculation with F. oxysporum further increased the enzyme activities. These results suggest that the enhanced activities of defense enzymes and elevated content of phenolics may contribute to bioprotection of banana plants against F. oxysporum.     

Frequency and diversity of Fusarium spp. colonizing roots of Egyptian cottons

Abstract

Fusarium species are known to play a role in several diseases of cotton including the seedling disease complex, wilt, and boll rot. Therefore, a mycoflora study was conducted in 1998 in order to identify Fusarium species found in association with cotton roots. A total of 109 samples of cotton seedlings infected with post-emergence damping-off or rotted roots of adult plants were obtained from different cotton-growing areas in Egypt. Forty-six isolates were recovered and were identified as follows: F. oxysporum (28 isolates), F. moniliforme (9), F. solani (6), F. avenaceum (2), F. chlamydosporum (1). F. oxysporum, F. moniliforme and F. solani, the dominant species, accounted for 60.9%, 19.6% and 13% of the total isolates, respectively in 1998. F. oxysporum showed the highest isolation frequency in Beharia and Minufiya while F. moniliforme showed the most isolation frequency in Minufiya and Gharbiya. F. oxysporum was one of the major taxa of the Fusarium assemblage from Giza 70. F. oxysporum showed the most frequently isolated fungus in May while F. moniliforme and F. solani were the most frequently isolated fungi in August. Isolation frequency of Fusarium spp. during July and August was significantly greater than that of April or June. This implies that cotton roots are subjected more to colonization by Fusarium spp. as plants mature. Regarding pathogenicity, of the 46 isolates of Fusarium spp. tested under greenhouse conditions, 38 isolates (82.4%) were pathogenic to seedlings of Giza 89. This study indicates that F. oxysporum and F. moniliforme are important pathogens in the etiology of cotton damping-off in Egypt.     

Radiometric assessment of tillage and seed treatment effect on soybean root rot caused by Fusarium spp. in central Minnesota

Soybean root rot, caused primarily by Fusarium solani f. sp. phaseoli in a complex with F. oxysporum and Rhizoctonia solani, has become an increasing problem for soybeans, dry beans, and other rotation crops in central Minnesota due to soil conditions associated with reduced tillage. This study was conducted, in two field sites in central Minnesota located near Staples and Verndale, to develop methods for nondestructive assessment of root rot severity using plant radiometric properties. Soybean canopy reflectance was measured with a hand-held multi-spectral radiometer. Prior to the radiometer measurements, attempts were made to create differing root rot situations with moldboard or chisel tillage, and with or without a biological seed treatment. Root rot severity was estimated using a visual disease severity scale. Colony-forming units (CFU) were determined to estimate soil populations of pathogenic F. solani and F. oxysporum. Results from the Verndale site consistently showed significant treatment effects in the measured canopy radiometric parameters, and in the visual disease rating and yield (significant for seed treatment). Values of a simple ratio vegetation index from this site exhibited negative relationships with disease rating and F. oxysporum CFU, and a positive linear relationship with yield. Treatment effects were generally not significant at the Staples site because of low initial F. oxysporum populations. The results indicate that remote sensing is potentially a rapid, nondestructive means for assessment of root rot diseases in soybean.     

Characterization of chitinase from Streptomyces luridiscabiei U05 and its antagonist potential against fungal plant pathogens

Streptomyces luridiscabiei U05 was isolated from wheat rhizosphere. It produced chitinase, which showed in vitro antifungal properties. The crude enzyme inhibited the growth of Alternaria alternata, Fusarium oxysporum, F. solani, Botrytis cinerea, F. culmorum and Penicillium verrucosum. The chitinase enzyme of the molecular weight of 45 kDa was purified using affinity chromatography of chitin. Streptomyces luridiscabiei U05 produced different chitinolytic enzymes. The highest enzyme activity was observed with the use of 4‐MU‐(GlcNAc)_, which points to the presence of an β‐N‐acetylhexosaminidase. The optimum activity was obtained at 35–40°C and pH 7–8. The enzyme showed thermostability at 35–40°C during 240 min of preincubation and lost its activity at 50°C and 60°C in 60 min. The chitinase activity from S. luridiscabei U05 was strongly inhibited by Hg²⁺ and Pb²⁺ ions, and sodium dodecyl sulphate (SDS). The Ca²⁺, Cu²⁺ and Mg²⁺ ions stimulated the activity of the enzyme.     

Induced systemic resistance in tomato by non-pathogenic Fusarium species for the management of Fusarium wilt

Isolates of non-pathogenic Fusarium moniliforme (Fu3, Fu7 and Fu24), F. oxysporum (Fu2, Fu4), F. solani (Fu25) and F. merismoides (Fu1) that were found to be effective in reducing wilt incidence in tomato were tested for their potential to elicit induced systemic resistance (ISR) in tomato. Talc formulations of these isolates derived from liquid fermentation as well as cell elicitors of these cultures were tested. Changes in the phenol and total protein contents and activities of peroxidase and polyphenol oxidase were studied. Isolate Fu3 induced more phenol and total protein contents as well as activities of peroxidase and polyphenol oxidase. Elicitors of Fu2 induced more of these compounds and enzymes. Although Fu1, Fu4 and Fu24 were found to give good control against Fusarium wilt incidence in an earlier study, they were less effective in inducing these defense related compounds. Peroxidase activity was increased when plants were treated with Fu3, Fu4, Fu7, Fu24 and Fu25, whereas polyphenol oxidase activity was increased only with the isolate Fu3 and elicitor of Fu2. It is suggested that ISR was the mode of action for the isolates Fu2 and Fu3, whereas for the other isolates, the mode of action may be root colonisation, competition for nutrition and so on. The role of ISR with non-pathogenic isolates of Fusarium spp. is discussed.     

Combination of Pseudomonas aeruginosa and Pochonia chlamydosporia for Control of Root-Infecting Fungi in Tomato

A plant growth‐promoting rhizobacterium, Pseudomonas aeruginosa strain IE‐6, and a fungal antagonist, Pochonia chlamydosporia, were tested for their ability to inhibit mycelial growth of root‐infecting fungi under laboratory conditions including Macrophomina phaseolina, Fusarium oxysporum, F. solani and Rhizoctonia solani. Biocontrol effectiveness of the bacterium and the fungus alone or in combination was also determined for the control of root‐infecting fungi under field conditions. In a dual‐culture plate assay, the colonies of P. chlamydosporia and P. aeruginosa met each other and no further growth of either organism occurred. Against M. phaseolina, F. solani and R. solani, an ethyl acetate extract of the culture filtrates of P. aeruginosa inhibited fungal growth greater than the hexane extract, but against F. oxysporum the hexane extract caused greater inhibition of fungal growth. By contrast, against M. phaseolina, F. oxysporum and F. solani, the hexane extract of P. chlamydosporia was more effective in the inhibition of fungal growth than the ethyl acetate fraction. Ethyl acetate extracts of P. aeruginosa at 1.0 mg/ml not only inhibited the radial colony growth of R. solani but also lysed the fungal mycelium. P. aeruginosa produced siderophores and hydrogen cyanide under laboratory conditions. Field experiments conducted in 1997 and repeated in 1998 revealed that Pochonia chlamydosporia and P. aeruginosa significantly suppressed the root‐infecting fungi M. phaseolina, F. oxysporum, F. solani and R. solani and that the combination of the two caused greater inhibition of the fungal pathogens than either alone. Application of P. chlamydosporia and P. aeruginosa as a soil drench also resulted in enhanced growth of tomato plants.     

Biocontrol of wilt disease complex of pea using Pseudomonas fluorescens and Rhizobium sp.

Biocontrol of wilt disease complex of pea caused by the root-knot nematode Meloidogyne incognita and Fusarium oxysporum f. sp. pisi was studied on pea (Pisum sativum L.) using plant growth-promoting rhizobacterium Pseudomonas fluorescens and root nodule bacterium Rhizobium sp. Inoculation of M. incognita and F.oxysporum alone caused significant reductions in plant growth over un-inoculated control. Reduction in plant growth caused by M. incognita was statistically equal to that caused by F. oxysporum. Inoculation of M. incognita plus F. oxysporum together caused a greater reduction in plant growth than the sum of damage caused by these pathogens singly. Inoculation of P. fluorescens and Rhizobium sp. individually or both together increased plant growth in pathogen inoculated and un-inoculated plants. Inoculation of P. fluorescens to pathogen-inoculated plants caused a greater increase in plant growth than caused by Rhizobium sp. Application of Rhizobium plus P. fluorescens caused a greater increase in plant growth than caused by each of them singly. Inoculation of P.fluorescens caused higher reduction in galling and nematode multiplication than caused by Rhizobium sp. Use of Rhizobium plus P. fluorescens caused higher reduction in galling and nematode multiplication than their individual inoculation. Plants inoculated with both pathogens plus Rhizobium showed less nodulation than plants inoculated with single pathogen plus Rhizobium. Inoculation of Rhizobium plus P. fluorescens resulted in higher root-nodulation than inoculated only with Rhizobium. Wilting indices were 4 and 5, respectively, when plants were inoculated with F. oxysporum and F. oxysporum plus M. incognita. Wilting indices were reduced maximum to 1 and 2, respectively, when plants inoculated with F.oxysporum and plants with both pathogens were treated with P. fluorescens plus Rhizobium.