Influence of Soil Type,Cultivar and Verticillium dahliae on the Structure of the Root and Rhizosphere Soil Fungal Microbiome of Strawberry

Sustainable management of crop productivity and health necessitates improved understanding of the ways in which rhizosphere microbial populations interact with each other, with plant roots and their abiotic environment. In this study we examined the effects of different soils and cultivars, and the presence of a soil-borne fungal pathogen, Verticillium dahliae, on the fungal microbiome of the rhizosphere soil and roots of strawberry plants, using high-throughput pyrosequencing. Fungal communities of the roots of two cultivars, Honeoye and Florence, were statistically distinct from those in the rhizosphere soil of the same plants, with little overlap. Roots of plants growing in two contrasting field soils had high relative abundance of Leptodontidium sp. C2 BESC 319 g whereas rhizosphere soil was characterised by high relative abundance of Trichosporon dulcitum or Cryptococcus terreus, depending upon the soil type. Differences between different cultivars were not as clear. Inoculation with the pathogen V. dahliae had a significant influence on community structure, generally decreasing the number of rhizosphere soil- and root-inhabiting fungi. Leptodontidium sp. C2 BESC 319 g was the dominant fungus responding positively to inoculation with V. dahliae. The results suggest that 1) plant roots select microorganisms from the wider rhizosphere pool, 2) that both rhizosphere soil and root inhabiting fungal communities are influenced by V. dahliae and 3) that soil type has a stronger influence on both of these communities than cultivar.     

Influence of inoculum density of Verticillium dahliae, temperature and relative humidity on epidemics of verticillium wilt of cotton in northern Iran

During 1992--2003, frequency of Verticillium dahliae propagules, disease incidence and severity of verticillium wilt of cotton were determined in several cotton growing fields in Golestan province, northeastern Iran. Inoculum density varied among fields and different years ranging between 2-47 propagules/g of air-dried soil with an average of 18.96+/-0.73. In addition, the pattern of diseased plants varied with type of field and year. Simple regression analysis showed a linear relationship between inoculum density of V. dahliae at planting time on one hand, disease incidence and severity for all years on the other. The straight line model described the increase in disease intensity index over the accumulated physiological time from sowing. The number of days above 28 degrees C (T) and the area under relative humidity (RH) had significant effects on inoculum density in soil (MS) and final disease development (Y) and fitted the Y = 65.840 - 0.0034 RH + 0.57 MS - 1.7T model with R2 = 0.859 and significant F-function (p<0.0001).     

Effect of forest fire on number, viability and post-fire re-establishment of arbuscular mycorrhizae

 Forest fire can affect arbuscular mycorrhizal (AM) fungi by changing the soil conditions and by directly altering AM proliferation. We studied the effects of a severe forest fire at Margalla Hills near Islamabad on the number and viability of AM fungal propagules in the burnt soil and their role in the re-establishment of post-fire infection in colonized plants. Compared with a nearby control area, the burnt site had a similar number of total spores but a lower number of viable AM fungal propagules. The roots of the two most frequent species at the burnt site, Dodonaea viscosa and Aristida adscensionis, showed a gradual increase in percentage root length colonized by AM fungi in general and hyphal infection in particular. Our results indicate resumption of mycorrhizal activity following the fire, probably from AM hyphae in the roots of these dominant shrubs. Accepted: 18 July 1997     

Characterization of Moroccan Isolates of Verticillium dahliaeKleb Using RAPD Markers

Isolates of Verticillium dahliae were sampled from different olive tree orchards in Morocco. These olive trees were located in different commercial culture locations in southern, central and northern Morocco. The isolates were characterized using genetic markers obtained after their DNA PCR amplification with random amplified polymorphic DNA (RAPD) primers. Among the 40 primers tested, 10 generated a total of 66 polymorphic fragments. Among the 38 isolates of V. dahliae tested, RAPD markers were successful in the characterization of groups based on their geographic origin. With the exception of one specific isolate, no correlation could be established among the isolates, based on the morphological appearance of the colony in culture.     

Modelling and Estimation of the Relative Potential for Infection of Winter Wheat by Inoculum of Gaeumannomyces graminis Derived from Propagules and Infected Roots

Abstract A model was developed to estimate the mean number of infections of seminal roots of wheat exposed to two sources of inoculum of the take-all fungus Gaeumannomyces graminis var. tritici , in an experimental system. The sources comprise discrete propagules of initial, soil inoculum and infected roots of volunteer plants that had been infected by the initial inoculum, prior to the growth of crop plants. The model was based on the probability of escape from infection by individual roots ofthe crop plants. Parameter estimation was by maximum likelihood. A model was first fitted to data for infection of roots from the soil inoculum. This yielded estimates for the efficiency of soil inoculum to cause infection in the absence of volunteer plants. The parameter for efficiency of infection by soil inoculum was resolved into components for inoculum density, survival of inoculum and the probability of success of individual propagules. The model was extended to include simultaneous exposure of crop roots to soil inoculum and to root inoculum on the volunteer plants. The presence of volunteer seedlings prior to sowing of crop plants resulted in an increase in the effectiveness of inoculum to cause disease. Sowing date and soil condition, as affected by addition of sand, were shown to have significant effects on the efficiency of both sources of inoculum.     

Efficacy of Talaromyces flavus Alone or in Combination with other Antagonists in Controlling Verticillium dahliae in Growth Chamber Experiments

Talaromyces flat-US reduced viability of microsclerotia of Verticitlium dahliae on senescent potato stems collected from the field when applied as ascospores in carboxy-methylcellulose or in talcum powder. Incorporating an alginate wheat-bran preparation of T. fiavus in soil at a rate of 0.5% (w/w) was followed by a decrease of > 90% of the population of V. dahliae in soil at both 15 and 25 C. Population densities of V. dahliae were negatively correlated (r = -0.50; P = 0.001) with those of T. flavus. However, the population of V. dahliae was also reduced in soil with alginate wheat-bran alone. When incorporated in soil in alginate wheat-bran and simultaneously coated on seeds in ta.lcum powder. T. flavus reduced colonization of roots and infection of eggplants by K. dahliae. Although to a lesser extent than with the antagonist, alginate wheat-bran without T. flavus also reduced infection by the pathogen. Treatment with combinations of T. flavus with other biocontrol agents, viz. Bacillus subtilis, Fusarium oxysporum or Gliocladium roseum , containing half of the inoculum of the single application of each antagonist, gave similar control of root colonization and stem infection by V. dahliae as application of the single antagonists. Population densities on the root of each antagonist were not or only slightly affected by the presence of the co-inoculated antagonist suggesting that the combinations were compatible.     

Wood Chip-Polyacrylamide Medium for Biocontrol Bacteria Decreases Verticillium dahliae Infection on Potato

The lack of consistent success of biological control of soilborne plant pathogens may be due to the introduction of the organism into a foreign environment. We hypothesized that wood chip-polyacrylamide (PAM) cores surrounding host plant roots could alter the soil environment to favour growth of introduced biocontrol microorganisms, thereby reducing Verticillium dahliae infection of potato ( Solanum tuberosum L.) in a greenhouse. A 7cm diameter ×15cm deep hole (core) was drilled in the center of a 20 ×30cm deep pot (1.9 kg) containing soil infested with V. dahliae inoculum. Cores were then filled with wood chip-PAM-biocontrol organism mixtures. Soils that had Streptomyces lydicus inoculated into wood chip-PAM cores had lower levels of V. dahliae symptoms ( V vis ) and V. dahliae isolations ( V iso ) than all other treatments in three soils. V vis and V iso on plants growing in soils amended with S. lydicus or Pseudomona corrugata inoculated into the soil itself (without wood chip-PAM cores) did not differ from soils that were unamended with these biocontrol organisms. V. dahliae biomass was lower in wood chip-PAM cores inoculated with S. lydicus than control or wood chip-PAM cores without biocontrol bacteria. Soils with wood chip-PAM cores inoculated with S. lydicus or P. corrugata generally had higher microbial biomass/ V. dahliae biomass (MB/VB) ratios than control soils, or soils with S. lydicus or P. corrugata inoculated into the soil. Wood chipPAM cores alone and wood chip-PAM cores inoculated with S. lydicus had higher MB/VB ratios than wood chip-PAM cores inoculated with P. corrugata . V vis and V iso were curvilinearly correlated with the MB/VB ratios in negative relationships, respectively (r 2 = 0.68, r 2 = 0.68). As the MB/VB ratio increased, V vis and V iso decreased. Although field studies and economic evaluations are necessary, amending soil with wood chips-PAM and a biocontrol bacterium may be a valuable method to increase the effectiveness of biocontrol organisms.     

Evaluation of Compost Amendments for Control of Vascular Wilt Diseases

Vascular wilt fungal pathogens cause heavy economic losses to a wide range of crops; amongst them are Fusarium oxysporum f. sp. melonis (FOM) and Verticillium dahliae Kleb. Several strategies for controlling these pathogens have been introduced, such as soil solarization, resistant rootstocks and biological control. In this study, the suppressive ability of seven different compost amendments and the plant growth-promoting rhizobacterium Paenibacillus alvei K165 (with proven activity against V. dahliae ) were tested against FOM in melon and V. dahliae in eggplant. It was shown that K165 had a suppressive effect against the pathogens in all experiments. On the contrary, the composts exhibited a narrow spectrum of effectiveness against the pathogens. Two composts were effective against V. dahliae and one against FOM. Moreover, we investigated the potential of the various compost samples and K165 to induce resistance in an Arabidopsis thaliana – V. dahliae or FOM model system. It was demonstrated that three composts and K165 were effective against V. dahliae ; whereas, one compost and K165 were effective against FOM. In a naturally V.dahliae infested field, the ability of K165 to enhance the suppressive effect of one of the compost amendments, was evaluated. It was demonstrated that fortification of the compost with strain K165 significantly reduced disease severity, whereas the single application of the compost was not sufficient to significantly protect the plants.     

Characterization and localization of prodiginines from Streptomyces lividans suppressing Verticillium dahliae in the absence or presence of Arabidopsis thaliana

The ascomycete Verticillium dahliae causes worldwide vascular wilt of many field and horticultural plants. The melanized resting structures of this fungus, so-called microsclerotia, survive for many years in soils and continuously re-infect plants. Due to the absence of known fungicides, Verticillium wilt causes immense crop losses. We discovered that the Gram-positive, spore-forming soil bacterium Streptomyces lividans expresses members of the prodiginine family during co-cultivation with V. dahliae. Using HPLC and LC-MS analysis of cultures containing S. lividans alone or grown together with V. dahliae, we found that undecylprodigiosin [394.4 M+H](+) is highly abundant, and streptorubin B [392.4 M+H](+) is present in smaller amounts. Within co-cultures, the quantity of undecylprodigiosin increased considerably and pigment concentrated at and within fungal hyphae. The addition of purified undecylprodigiosin to growing V. dahliae hyphae strongly reduced microsclerotia formation. Undecylprodigiosin was also produced when S. lividans grew on the roots of developing Arabidopsis thaliana plants. Furthermore, the presence of the undecylprodigiosin producer led to an efficient reduction of V. dahliae hyphae and microsclerotia on plant-roots. Based on these novel findings and previous knowledge, we deduce that the prodiginine investigated leads to multiple cellular effects, which ultimately impair specific pathways for signal transduction and apoptosis of the fungal plant pathogen.     

Fungal endophytes in potato roots studied by traditional isolation and cultivation-independent DNA-based methods

The composition and relative abundance of endophytic fungi in roots of field-grown transgenic T4-lysozyme producing potatoes and the parental line were assessed by classical isolation from root segments and cultivation-independent techniques to test the hypothesis that endophytic fungi are affected by T4-lysozyme. Fungi were isolated from the majority of root segments of both lines and at least 63 morphological groups were obtained with Verticillium dahliae, Cylindrocarpon destructans, Colletotrichum coccodes and Plectosporium tabacinum as the most frequently isolated species. Dominant bands in the fungal fingerprints obtained by denaturing gradient gel electrophoresis analysis of 18S rRNA gene fragments amplified from total community DNA corresponded to the electrophoretic mobility of the 18S rRNA gene fragments of the three most abundant fungal isolates, V. dahliae, C. destructans and Col. coccodes, but not to P. tabacinum. The assignment of the bands to these isolates was confirmed for V. dahliae and Col. coccodes by sequencing of clones. Verticillium dahliae was the most abundant endophytic fungus in the roots of healthy potato plants. Differences in the relative abundance of endophytic fungi colonizing the roots of T4-lysozyme producing potatoes and the parental line could be detected by both methods.     

Phenolic Compounds of Apple and their Relationship to Scab Resistance

The contents of 15 different phenolic compounds in apple leaves and fruit skin from various orchards were determined by HPLC. High concentrations of flavan-3-ols were found in those orchards which were not infected by Venturia inaequalis . This observation corroborates earlier findings on different cultivars. The hypothesis that these compounds could be involved in the resistance of apple to scab was tested by inhibiting the key enzyme of the phenol biosynthesis, the phenylalanine-ammonia-lyase, in young shoots of the resistant genotype 'Sir Prize'. After inoculation of the inhibitor-treated leaves, severe symptoms of the disease occurred. The fungal infection was confirmed by histological studies. No flavonal-accumulation could be observed at the infection site which was the case in the non-inhibited but inoculated controls.     

Effect of Inoculum Density on Verticillium Wilt Incidence in Commercial Olive Orchards

Verticillium wilt, caused by Verticillium dahliae Kleb., is presently the most destructive disease of olive, particularly in Andalucía (southern Spain). ‘Picual’ and ‘Arbequina’ are the dominant cultivars being planted in Spain. Both cultivars are highly susceptible to the defoliating pathotype of V. dahliae when artificially inoculated by root‐dipping or stem injection. Conversely, ‘Arbequina’ is considered more resistant than ‘Picual’ based on field observations and farmer's experience. In this study, the differential reaction between of cultivars was confirmed by surveys of naturally infested orchards with different inoculum densities of the pathogen. The average percentage of affected olive trees of ‘Picual’ was 60.2%, while only 13.1% of trees of ‘Arbequina’ showed disease symptoms. Overall, the pathogen caused extensive wilting of branches and defoliation on the trees of ‘Picual’, whereas ‘Arbequina’‐infected trees showed chlorotic symptoms and slight defoliation. The relationship between inoculum density and disease incidence fit a logarithmic function for both cultivars. The percentage of affected trees of ‘Arbequina’ per year increased linearly (y = 0.3559x, R² = 0.5652, and P = 0.0195) with the inoculum density in the soil, whereas this relationship was not observed for the ‘Picual’. Planting density had no effect on disease incidence for any of the two cultivars.     

Effect of fungal species involved in the olive fruit rot on the qualitative properties of olive oil

Olive oil is the most important product of olive fruits with worldwide consumption, particularly in Mediterranean countries. Olive oil is generally extracted mechanically from the olive fruits. Some biotic and abiotic factors may affect the quality of oil extracted from olive fruit. Contamination with fungi during growth period in the garden or during the conservation of the harvested crop under storage condition may leave negative effects on the quality of olive oil. However, there is no data available on the effects of fungal infections on qualitative properties of olive oil in Iran. In the present study effects of several fungal groups previously isolated from rotten olive fruit in olive orchards including Alternaria alternata, Fusarium nygamai, Aspergillus ochraceus, Arthrinium phaeospermum, Cladosporium cladosporioides, Aureobasidium pullulans, Epicoccum nigrum, Penicillium expansum, Truncatella angustata, Trichothecium roseum and Trichoderma harzianum were evaluated on some qualitative properties of olive oil, under laboratory condition on two olive cultivars (Zard & Roghani). For this purpose fresh and healthy fruits of olive, were surface sterilisation with 96% ethanol and rinsed with sterile water and then inoculated with each of the fungal groups separately using spore suspension (10^6?ml^?1). The experiment was carried out in two replicates for each treatment (fungal isolates). The results of this study revealed that fungal infection caused significant increase in the extracted oil acidity and peroxide values. However, there was no significant difference in the acidity and peroxide values among different treatments (fungal isolates).     

The Effect of Sulfur on the Composition of Arbuscular Mycorrhizal Fungal Communities During the Pod-Setting Stage of Different Soybean Cultivars

This study sought to investigate the effect of sulfur levels on changes in the fungal community composition of arbuscular mycorrhizae (AM) at the pod-setting stage and the relationship between the amount of applied sulfur and AM fungal diversity in different soybean cultivars. The objective of the research was to determine the optimal sulfur application level for different soybean cultivars and to improve soybean yield and quality from the perspective of AM fungal diversity. Three soybean cultivars, Heinong 44, Heinong 48, and Heinong 37, were selected as study materials. In addition to 0.033?g each of N, P₂O₅ and K₂O per kg of soil, 0, 0.02, 0.04, or 0.06?g of elemental sulfur was applied to each kg of soil for the four treatment groups, S1, S2, S3, and S4, respectively. The AM fungal community structure was analyzed in the soil and root of different soybean cultivars using the PCR-DGGE technology. The results indicated a significant effect of sulfur on the AM fungal community structure in the roots and rhizospheric soil of different soybean cultivars. The three soybean cultivars in group S2 exhibited the highest diversity in AM fungus. Significant changes in the dominant fungal species were observed in the DGGE fingerprints of each sample, and Glomus, Funneliformis, Rhizophagus, and Claroideoglomus fungi were the dominant species of AM fungus in the roots and soil of soybean. The application of an appropriate amount of sulfur improved the diversity of AM fungi in roots and rhizospheric soil of different soybean cultivars.     

Patterns of arbuscular mycorrhiza down the profile of a heavy textured soil do not reflect associated colonization potential

Colonization of roots by arbuscular mycorrhizal (AM) fungi in several annual crops in two consecutive seasons was compared with, in the second season, the density of fungal propagules in the soil with the use of a bioassay. Root density decreased down the soil profile in both years in all crops, and a high proportion of roots were mycorrhizal throughout the profile. AM colonization decreased down the profile in cotton and lablab in the second season only. The bioassay indicated that most propagules of AM fungi in soils under cotton were located near the surface, with virtually no propagules at 1 m. The absence of propagules at depth indicates a lack of mycelium deep in the soil, and suggests that mycorrhizas are primarily initiated in the surface soil and that the fungi colonize the root system mostly through secondary spread down the profile. The use of AM colonization in the field as an indicator of propagule density and symbiotic function should be qualified by an understanding of the depth in the soil from which roots were extracted.     

基因沉默技术在抗真菌病害中的应用和展望

真菌病害严重威胁作物的产量和品质,给国家和人民造成巨大的经济损失。尤其是引起维管束病害的土传真菌,化学农药的作用效果很不理想。利用抗性基因进行遗传育种是目前生物防治的重要手段之一,但对于缺乏抗性资源的物种,面对强大的土壤真菌病害,研究者也时常束手无策。近年来,利用RNA干扰技术发展而来的宿主诱导的基因沉默(Host induced gene silencing,HIGS)策略,在抗病虫害领域逐渐崭露头角,但由于真菌侵染的复杂多样性及土壤传播的特性,HIGS在土壤真菌病害中的应用充满神秘和挑战。本研究室近期揭示了棉花黄萎病(一种严重的土壤真菌病害)的"罪魁祸首"——大丽轮枝菌的侵染结构和侵染过程;并首次证明了宿主植物内源小RNA能够跨界进入病原菌细胞中降解致病基因表达的抗病作用;在此基础上,本研究室利用HIGS在棉花上获得了对黄萎病抗性较高的品系,成功地开辟了抗土壤黄萎真菌病害的新天地,研究结果显示出基因沉默技术在这一领域强大的应用潜力和前景。     

Functional Analysis of Autophagy Genes via Agrobacterium-Mediated Transformation in the Vascular Wilt Fungus Verticillium dahliae

Autophagy is a widely conserved intracellular process for degradation and recycling of proteins,organelles and cytoplasm in eukaryotic organisms and is now emerging as an important process in foliar in...