Assessment of Parasitic Activity of Fusarium Strains Obtained from a Heterodera schachtii-Suppressive Soil

This study assessed the potential impact of various Fusarium strains on the population development of sugarbeet cyst nematodes. Fungi were isolated from cysts or eggs of Heterodera schachtii Schmidt that were obtained from a field suppressive to that nematode. Twenty-six strains of Fusarium spp. were subjected to a phylogenic analysis of their rRNA-ITS nucleotide sequences. Seven genetically distinct Fusarium strains were evaluated for their ability to influence population development of H. schachtii and crop performance in greenhouse trials. Swiss chard (Beta vulgaris) seedlings were transplanted into fumigated field soil amended with a single fungal strain at 1,000 propagules/g soil. One week later, the soil was infested with 250 H. schachtii J2/100 cm³ soil. Parasitized eggs were present in all seven Fusarium treatments at 1,180 degree-days after fungal infestation. The percentage of parasitism ranged from 17 to 34%. Although the most efficacious F. oxysporum strain 471 produced as many parasitized eggs as occurred in the original suppressive soil, none of the Fusarium strains reduced the population density of H. schachtii compared to the conducive check. This supports prior results that Fusarium spp. were not the primary cause of the population suppression of sugarbeet cyst nematodes at this location.     

Growth response ofTheobroma cacao L. seeldings to inoculation with vesicular-arbuscular mycorrhizal fungi

Summary The effects of vesicular-arbuscular mycorrhiza (VAM) on the growth and phosphorus uptake of cocoa seedlings (Theobroma cacao L.) grown for 100 days in polythene bags, were studied at five levels of phosphorus fertilization in both steamed and unsterile Bungor Series soil (a fine clayey, kaolinitic isohyperthermic Typic Paleudult). The cocoa seedlings responded well to phosphorus fertilization and mycorrhizal treatments. Plants inoculated with VAM fungi (Gigaspora spp.) gave the most vigorous growth and higher phosphorus in the leaf tissues in unsterile soil compared to plants grown in steamed soil. However, the mycorrhizal effect was significantly more pronounced (P<0.01) in plants grown in steamed than in unsterile soil. High levels of phosphorus application depressed mycorrhizal development. Phosphorus fertilizer applied at the rates of 250 and 500 ug g⁻¹ soil gave maximum root colonization and spore counts in both soil types used.     

Nitrate Reductase in Wheat Plants Grown Under Water Stress and Inoculated with Azospirillum spp.

The present investigation has been performed to evaluate nitrate reductase (NR) and nitrogenase activities as well as growth and mineral nutrition of wheat plants grown under drought stress and inoculated with different Azospirillum strains (NR⁻ and NR⁺). Fresh, dry mass and water content decreased with decreasing soil moisture content, which was accompanied with low soluble sugars and soluble protein content and increase in the total amino acids content. Azospirillum inoculation with either bacterial strain (NR⁻ and NR⁺) significantly increased the above characteristics even at 40 % moisture content. NR activity decreased in both the shoots and roots by decreasing soil moisture content. NR⁺ strain exhibited increased root NR activity compared with uninoculated plants or inoculated with NR⁻ strain. However, plants inoculated with NR⁻strain increased NR activity in the shoot more than in the root of the same plant and in the shoot of control plants. Inoculation with either NR⁻ and NR⁺ Azospirillum strains gave higher nitrogenase activity than uninoculated control plants. The low N supply (0.5 mM) did not affect nitrogenase activity. NR⁻strain was less effective than NR⁺strain in promoting total N-yield, spike numbers and their mass per pot. Azospirillum inoculation exhibited no significant changes in wheat Mg²⁺ content. However, K⁺ and Ca²⁺ have shown significantly increased values. Azospirillum beneficial effect on plant N balance and growth are most probably composed of multiple mechanisms and beneficial NR is one of them. The importance of Azospirillum NR⁺strains for increasing wheat resistance to water stress is also supported by the obtained data. This revised version was published online in July 2006 with corrections to the Cover Date.     

The involvement of Pseudomonas bacteria in induced systemic resistance in plants (Review)

This article reviews the most recent results of studies on the mechanism of induced systemic resistance (ISR) elicited in plants by non-pathogenic bacteria of the genus Pseudomonas. Several examples of Pseudomonas strains eliciting resistance against fungal phytopathogens in different species of crop plants are presented. Literature data dealing with bacterial elicitors and the effect of their interaction with plant receptors are quoted. Special focus is focused on the controversial issue of the correlation between the synthesis of pathogenesis-related proteins (PRs) and ISR.     

<Emphasis Type="Italic">Pseudomonas chlororaphis</Emphasis>: a salt-tolerant bacterial inoculant for plant growth stimulation under saline soil conditions

Saline soils constitute a serious production problem for vegetable crops as they are known to suppress plant growth. One of the possible measures to improve crop health in such conditions is to use salt-tolerant bacterial inoculants which can control diseases and promote plant growth. In the present work the ability of Pseudomonas chlororaphis isolate TSAU13 to promote cucumber and tomato plant growth and to improve fruit yield by protecting these plants against soil-borne pathogens in salinated soil were investigated. The bacterial strain stimulated shoot growth (up to 32%), dry matter (up to 43%), and the fruit yield of tomato and cucumber (up to 16%) compared to the uninoculated control plants under saline conditions. The strain was able to survive on the root of 2-month-old plants. 29% of the cucumber and 27% of the tomato plants which had grown in soil to which no Fusarium solani spores had been added were diseased, whereas in the presence of the pathogenic fungus 58% of the cucumber and 52% tomato plants had disease symptoms. P. chlororaphis TSAU13 showed statistically significant disease reduction in comparison to the Fusarium-uninfected and infected control plants. Those results showed that P. chlororaphis TSAU13 has a great biotechnological potential in improvement of vegetable production in commercial greenhouses under saline conditions.     

Somaclonal variation in celery: screening for resistance to Fusarium oxysporum f. sp. apii

Summary Two methods were used to screen putative Fusarium-resistant celery (Apium graveolens L.) plantlets from cell culture: placing plantlets on a mycelial mat for one month or planting them directly in Fusarium-infested soil. Resistant phenotypes were identified with both methods, but the plants grown on the mycelial mat died before they reached reproductive maturity. Four plants, K, T-2, T-3, and R-R₁ from the soil screen, survived and produced viable seed. Tests of self-pollinated progeny, in field and greenhouse conditions, showed that T-2, T-3, and R-R₁ were superior to the original cultivar, 5270R, with respect to disease resistance, as measured by vascular discoloration and plant height. Chi-square analysis of progeny scores for root and crown decay showed that the new variation was heritable and appeared to be conditioned by more than one locus.     

Evaluation of endophytic actinobacteria as antagonists of Gaeumannomyces graminis var. tritici in wheat

Endophytic actinobacteria isolated from healthy cereal plants were assessed for their ability to control fungal root pathogens of cereal crops both in vitro and in planta. Thirty eight strains belonging to the genera Streptomyces, Microbispora, Micromonospora, and Nocardioidies were assayed for their ability to produce antifungal compounds in vitro against Gaeumannomyces graminis var. tritici (Ggt), the causal agent of take-all disease in wheat, Rhizoctonia solani and Pythium spp. Spores of these strains were applied as coatings to wheat seed, with five replicates (25 plants), and assayed for the control of take-all disease in planta in steamed soil. The biocontrol activity of the 17 most active actinobacterial strains was tested further in a field soil naturally infested with take-all and Rhizoctonia. Sixty-four percent of this group of microorganisms exhibited antifungal activity in vitro, which is not unexpected as actinobacteria are recognized as prolific producers of bioactive secondary metabolites. Seventeen of the actinobacteria displayed statistically significant activity in planta against Ggt in the steamed soil bioassay. The active endophytes included a number of Streptomyces, as well as Microbispora and Nocardioides spp. and were also able to control the development of disease symptoms in treated plants exposed to Ggt and Rhizoctonia in the field soil. The results of this study indicate that endophytic actinobacteria may provide an advantage as biological control agents for use in the field, where others have failed, due to their ability to colonize the internal tissues of the host plant.     

Sources of fungal linamarases

Forty-four strains of Aspergillus, Penicillium, Fusarium, Trichoderma and Rhizopus were grown on a liquid medium containing glucose and cassava-root extract. All of the Aspergillus and Fusarium strains, eight out of 10 Penicillium strains and three of seven Trichoderma strains showed linamarase activity. No such activity was detected in any Rhizopus strain. The crude enzyme preparation from F. oxysporum had the highest affinity for linamarin whereas that from A. nidulans was the most heat-stable.The authors are with the Department of Botany, National University of Singapore. Kent Ridge. Singapore 0511     

Bacillus halotolerans strain LYSX1-induced systemic resistance against the root-knot nematode Meloidogyne javanica in tomato

Purpose

Determination of the nematicidal potential and mode of action of bacteria isolated from tobacco rhizosphere soil against the root-knot nematode Meloidogyne javanica in tomato plants.

Methods

Antagonistic bacteria were isolated from rhizosphere soil of tobacco infested with root-knot nematodes. Culture filtrate was used to examine nematicidal activity and ovicidal action of bacterial strains. Biocontrol of M. javanica and growth of treated tomato plants were assessed in pot experiments. To clarify whether secondary metabolites of bacteria in tomato roots induced systemic resistance to M. javanica, bacterial culture supernatants and second-stage juvenile nematodes were applied to spatially separated tomato roots using a split-root system. Bacterial strains were identified by 16S rDNA and gyrB gene sequencing and phylogenetic analysis.

Results

Of the 15 bacterial strains isolated, four (LYSX1, LYSX2, LYSX3, and LYSX4) demonstrated nematicidal activity against second-stage juveniles of M. javanica, and strain LYSX1 showed the greatest antagonistic activity; there was dose-dependent variability in nematicidal activity and inhibition of egg mass hatching by strain LYSX1. In vivo application of LYSX1 to tomato seedlings decreased the number of egg masses and galls and increased the root and shoot fresh weight. Treatment of half of the split-root system with LYSX1 reduced nematode penetration to the other half by 41.64%. Strain LYSX1 was identified as Bacillus halotolerans.

Conclusion

Bacillus halotolerans LYSX1 is a potential microbe for the sustainable biocontrol of root-knot nematodes through induced systemic resistance in tomato.

     

Effect of organic amendment on phenolic content of soil and plant and response ofMeloidogyne javanica and its host to related compounds

Summary Amendment of soil with margosa cake or sawdust supplemented with NPK fertilizers increased its phenolic content. The concentration of total phenols was related to the amount of amendment used and varied with the length of decomposition period. Total phenols estimated in ether extract were more in margosa cake amended soil than in sawdust amended soil. Roots of tomato plants grown in amended soil showed presence of higher quantity of total phenols than those grown in non-amended soil. Exposure of females ofMeloidogyne javanica to benzoic, phenyl butyric, phenyl acetic and cinnamic acids significantly reduced their egg laying capacity. Suppression of larval motility was one of the main direct effects of these acids on the nematode. Exposure of tomato roots to different concentrations of phenyl acetic, benzoic, phenyl butyric and cinnamic acids imparted some resistance to invasion by the nematode. In such treated plants fewer larvae could penetrate the roots and develop into mature females and fewer eggs were produced. Research paper No.1455 through the Experiment Station G.B.P.U,A, & T., Pantnagar     

Suppressive soil against Sclerotinia sclerotiorum as a source of potential biocontrol agents: selection and evaluation of Clonostachys rosea BAFC1646

The fungal diversity structures of soils that are suppressive and non-suppressive to Sclerotinia sclerotiorum were characterised and screened for fungal strains antagonistic to the S. sclerotiorum pathogen. Soil suppressiveness was associated with a particular fungal diversity structure. Principal component analysis showed that antagonism by fungal species in suppressive soils was associated with the occurrence of Fusarium oxysporum, Fusarium solani, Talaromyces flavus var. flavus and Clonostachys rosea f. rosea. In particular, C. rosea f. rosea occurred exclusively in suppressive soil samples, suggesting that this morpho-species plays an important role in suppression of S. sclerotiorum diseases. One strain of C. rosea f. rosea (BAFC1646) was selected for further experiments. Dual-culture assays confirmed the antagonistic behaviour of C. rosea f. rosea BAFC1646 against three different S. sclerotiorum strains. Antifungal activity was corroborated by diffusion assays with metabolite extracts. Greenhouse assays with soybean plants showed that the selected C. rosea f. rosea strain reduced the percentage of dead plants when co-inoculated with S. sclerotiorum. In addition, inclusion of C. rosea f. rosea alone increased shoot lengths significantly. In this work, we established the involvement of fungal species in soil suppressiveness and in further assays confirmed that C. rosea f. rosea BAFC1646 exhibits a bioprotective effect against S. sclerotiorum in soybean plants.     

Use of a Colonization-Deficient Strain of Escherichia coli in Strain Combinations for Enhanced Biocontrol of Cucumber Seedling Diseases

Seed treatments containing combinations of Escherichia coli S17R1 and Burkholderia cepacia Bc-B provided significantly greater (P ≤ 0.05) suppression of cucumber seedling pathogens in a field soil naturally infested with Pythium and Fusarium spp. than seeds treated individually with strains Bc-B, S17R1, or Enterobacter cloacae 501R3. Although strain S17R1 had no effect on disease severity when applied alone and did not colonize cucumber rhizosphere, it enhanced the biocontrol effectiveness of strain Bc-B.     

Composition of<Emphasis Type="Italic">Casuarina</Emphasis> leaf litter and its influence on<Emphasis Type="Italic">Frankia-Casuarina</Emphasis> symbiosis in soil

Plant needles ofCasuarina equisetifolia were collected and analyzed in parallel with soil analysis. In three strains ofFrankia—symbionts ofCasuarina—their infectivity and plant performance was determinedin vitro after soil amendment with different leaf litter concentrations. Only one strain was able to nodulate the plant at all litter concentrations (0.5, 3 and 5%) although the nodules were very small. However, all treated plants grew poorly; their growth was reduced by approximately 90% (for 5% litter concentration) compared to plants grown on untreated soil, on the basis of total dry mass. Inhibition of nodulation can be attributed to high concentrations of some elements and compounds that were either found inC. equisetifolia litter or originally found in soil (i.e. chloride, cyanide, copper, manganese and phenols). In general, plant growth decreased as more litter was added. Plant total nitrogen content was also reduced after increasing the litter concentration. The inhibitory effect of high litter concentrations was mainly on plant growth and to a lesser extent on plant nodulation byFrankia strains.     

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.     

Biological detoxification of the mycotoxin deoxynivalenol and its use in genetically engineered crops and feed additives

Deoxynivalenol (DON) is the major mycotoxin produced by Fusarium fungi in grains. Food and feed contaminated with DON pose a health risk to humans and livestock. The risk can be reduced by enzymatic detoxification. Complete mineralization of DON by microbial cultures has rarely been observed and the activities turned out to be unstable. The detoxification of DON by reactions targeting its epoxide group or hydroxyl on carbon 3 is more feasible. Microbial strains that de-epoxidize DON under anaerobic conditions have been isolated from animal digestive system. Feed additives claimed to de-epoxidize trichothecenes enzymatically are on the market but their efficacy has been disputed. A new detoxification pathway leading to 3-oxo-DON and 3-epi-DON was discovered in taxonomically unrelated soil bacteria from three continents; the enzymes involved remain to be identified. Arabidopsis, tobacco, wheat, barley, and rice were engineered to acetylate DON on carbon 3. In wheat expressing DON acetylation activity, the increase in resistance against Fusarium head blight was only moderate. The Tri101 gene from Fusarium sporotrichioides was used; Fusarium graminearum enzyme which possesses higher activity towards DON would presumably be a better choice. Glycosylation of trichothecenes occurs in plants, contributing to the resistance of wheat to F. graminearum infection. Marker-assisted selection based on the trichothecene-3-O-glucosyltransferase gene can be used in breeding for resistance. Fungal acetyltransferases and plant glucosyltransferases targeting carbon 3 of trichothecenes remain promising candidates for engineering resistance against Fusarium head blight. Bacterial enzymes catalyzing oxidation, epimerization, and less likely de-epoxidation of DON may extend this list in future.     

Effects of soil water level on the development of adult plant resistance to powdery mildew in barley

Barley grown in dry soil developed greater adult plant resistance (APR) to powdery mildew (Erysiphe graminis DC. f. sp. hordei Mérat) than barley grown in wet soil. Conidial germination and appressorium formation were less, and fungal development between formation of appressoria and elongating secondary hyphae on upper leaves was inhibited, when adult plants were grown in dry soil. Mildew colonies expanded more slowly on leaves of adult plants than on leaves of seedlings, especially if adult plants had grown in dry soil. APR was reduced if plants, previously grown in dry soil, were well watered more than 32 h before inoculation. Conidia originating from plants grown in dry soil had a lower solute potential and greater ability to infect plants grown in dry but not wet soil than conidia originating from plants grown in wet soil. APR could not be attributed simply to increased cell wall or cuticle thickness, nor to lowered leaf solute potentials, as has sometimes been suggested for powdery mildew diseases. Increasing plant age and water stress induced increases in cell wall and cuticle thickness, but these changes did not always coincide with changes in disease resistance. Increasing plant age and water stress also lowered leaf solute potentials but fungal solute potentials were lower than leaf solute potentials and, more importantly, were lower than leaf water potentials. Thus, fungal growth was not limited by the availability of water from the host during penetration and hyphal establishment. It is suggested that resistance levels may be determined not by the thickness of epidermal structures, nor by lowering of solute potential per se, but by specific substances harmful to the fungus which accumulate in either cell wall, cuticle or sap, and whose concentration is dependent on the age and water stress of leaves.     

Inoculum Density and Population Dynamics of Suppressive and PathogenicStreptomycesStrains and Their Relationship to Biological Control of Potato Scab

SeveralStreptomycesstrains are capable of suppressing potato scab caused byStreptomyces scabies.Although these strains have been successful in the biocontrol of potato scab in the field, little is known about how populations of pathogenicStreptomycesin the potato rhizosphere are influenced by inoculation of the suppressive strains. The effects of inoculum densities of pathogenic and suppressiveStreptomycesstrains on their respective populations on roots and in rhizosphere soil were examined during the growing season. The relationships between inoculum density or rhizosphere population densities and disease severity were also investigated. Populations of suppressiveStreptomycesstrain 93 increased significantly on roots with increasing inoculum dose. At its highest inoculum dose, the suppressive strain reached a population density greater than 10⁶CFU/g root 14 weeks after planting. The ability of the suppressive strain to increase its populations with increasing inoculum density was hindered at high inoculum doses of the pathogen, suggesting that density-dependent competitive interactions may be occurring between the two antagonists. Strain 93 was most effective at preventing scab early in the growing season (8 weeks after planting), when tubers were most susceptible to the scab disease. Population densities of the suppressive strain in soil were more highly negatively correlated with scab severity than were populations on roots, suggesting that rhizosphere soil rather than potato roots may be the primary source of inoculum of the suppressive strain for tubers.     

Soil Suppressiveness to Fusarium Wilt of Melon, Induced by Repeated Croppings of Resistant Varieties of Melons

A field soil, artificially infested with pathogenic isolates of Fusarium oxysporum f. sp. melonis was continuously used for screening resistant varieties of melon to Fusarium wilt. After 9–10 years of continuous cropping with resistant varieties, the soil had developed induced suppressiveness. Seven to 9 experimental replantings of the induced suppressive soil with the susceptible cultivar of melon, ‘Ein-Dor', nullified its suppressiveness. This was expressed by 90 % disease incidence. Only 2 replantings were required to obtain the same disease incidence in an adjacent field of a conducive soil. Nonpathogenic isolates of F. oxysporum, isolated from the rhizospheres of melon seedlings, induced various degrees of soil suppressiveness when added to soil at various ratios to the pathogenic isolate.     

Alleviating salt stress in tomato seedlings using Arthrobacter and Bacillus megaterium isolated from the rhizosphere of wild plants grown on saline–alkaline lands

Salt-induced soil degradation is common in farmlands and limits the growth and development of numerous crop plants in the world. In this study, we isolated salt-tolerant bacteria from the rhizosphere of Tamarix chinensis, Suaeda salsa and Zoysia sinica, which are common wild plants grown on a saline–alkaline land, to test these bacteria's efficiency in alleviating salt stress in tomato plants. We screened out seven strains (TF1–7) that are efficient in reducing salt stress in tomato seedlings. The sequence data of 16S rRNA genes showed that these strains belong to Arthrobacter and Bacillus megaterium. All strains could hydrolyze casein and solubilize phosphate, and showed at least one plant growth promotion (PGP)-related gene, indicating their potential in promoting plant growth. The Arthrobacter strains TF1 and TF7 and the Bacillus megaterium strain TF2 and TF3 could produce indole acetic acid under salt stress, further demonstrating their PGP potential. Tomato seed germination, seedling length, vigor index, and plant fresh and dry weight were enhanced by inoculation of Arthrobacter and B. megaterium strains under salt stress. Our results demonstrated that salt-tolerant bacteria isolated from the rhizosphere of wild plants grown on saline–alkaline lands could be used for alleviating salt stress in crop plants.     

Effect of the VAM fungus Glomus sp. on the growth and yield of soybean inoculated with Bradyrhizobium japonicum

The effect of two Bradyrhizobium japonicum strains (D344 and Urbana), on the frequency and intensity of infection by a VAM fungal Glomus sp. and the effect of VAM on biomass production by nodulating plants were tested in soybean growing in a soil containing low levels of accessible P and N. During the initial stage of vegetative growth, mycorrhiza frequency in roots inoculated with the two rhizobial strains did not differ. However, during flowering it was 178% higher in roots with the strain D344 than in the presence of the strain Ubrana. At final harvest (green pods) the VAM frequency did not differ in the presence of either strain. VAM positively affected biomass production, foliar concentrations of P, Zn and Cu, and number and dry matter yield of pods, but did not increase concentrations of total N and K. In nonmycorrhizal plants total nitrogenase activity (not nodule mass) and growth were higher with the rhizobial strain Urbana. The greatest nitrogenase activity, growth and yield occurred in the presence of the VAM fungus, and did not differ for plants with different strains of rhizobia.