Mechanisms of powdery mildew resistance in the Vitaceae family

The cultivated grapevine, Vitis vinifera, is a member of the Vitaceae family, which comprises over 700 species in 14 genera. Vitis vinifera is highly susceptible to the powdery mildew pathogen Erysiphe necator. However, other species within the Vitaceae family have been reported to show resistance to this fungal pathogen, but little is known about the mechanistic basis of this resistance. Therefore, the frequency of successful E. necator penetration events, in addition to programmed cell death (PCD) responses, were investigated in a representative genotype from a range of different species within the Vitaceae family. The results revealed that penetration resistance and PCD-associated responses, or combinations of both, are employed by the different Vitaceae genera to limit E. necator infection. In order to further characterize the cellular processes involved in the observed penetration resistance, specific inhibitors of the actin cytoskeleton and secretory/endocytic vesicle trafficking function were employed. These inhibitors were demonstrated to successfully break the penetration resistance in V. vinifera against the nonadapted powdery mildew E. cichoracearum. However, the use of these inhibitors with the adapted powdery mildew E. necator unexpectedly revealed that, although secretory and endocytic vesicle trafficking pathways play a crucial role in nonhost penetration resistance, the adapted powdery mildew species may actually require these pathways to successfully penetrate the plant host.     

Lack of trade-off between direct and indirect defence against grape powdery mildew in riverbank grape

Abstract.  1. The hypothesis that plants that invest more in an indirect defence will invest less in direct mechanisms was tested using genotypes of the riverbank grape Vitis riparia that varied in expression of acarodomatia, tufts of non-glandular trichomes on underside of leaves that are frequently occupied by mycophagous mites. Mycophagous mites in this system have been shown to protect vines from grape powdery mildew caused by the fungus Uncinula necator .
2. Multiple cane cuttings from 24 genotypes of V. riparia , a species native to North America, were obtained from the USDA grape genetics repository and planted out in a common garden.
3. Genotypes varied in size of acarodomatia by threefold and varied in density of mycophagous mites by over 10-fold. Genotypes originating from the western part of the distribution had smaller acarodomatia and fewer mycophagous mites than genotypes from the eastern part of the distribution. There was a positive genetic correlation between acarodomatia size and mite density.
4. In a separate experiment, where vines were kept free of mycophagous mites, genotypes showed 10-fold variation in percentage leaf surface colonised by mildew. There was, however, no relationship between mildew on leaves and size of acarodomatia. Mildew did not vary depending on origin of genotype, although female vines tended to have greater levels than males.
5. No evidence was found for a trade-off between investment in an indirect defence mediated through acarodomatia and investment in direct defence against grape powdery mildew. Mite/plant defence mutualisms are contrasted with ant/plant defence mutualisms where there is better evidence for trade-offs under field conditions.     

Synthesis and fungicidal activity of quinolin-6-yloxyacetamides,a novel class of tubulin polymerization inhibitors

A novel class of experimental fungicides has been discovered, which consists of special quinolin-6-yloxyacetamides. They are highly active against important phytopathogens, such as Phytophthora infestans (potato and tomato late blight), Mycosphaerella graminicola (wheat leaf blotch) and Uncinula necator (grape powdery mildew). Their fungicidal activity is due to their ability to inhibit fungal tubulin polymerization, leading to microtubule destabilization. An efficient synthesis route has been worked out, which allows the diverse substitution of four identified key positions across the molecular scaffold.     

A mutation in the 14 alpha-demethylase gene of Uncinula necator that correlates with resistance to a sterol biosynthesis inhibitor.

We investigated the molecular basis of resistance of the obligate biotrophic grape powdery mildew fungus Uncinula necator to sterol demethylation-inhibiting fungicides (DMIs). The sensitivity of 91 single-spore field isolates of U. necator to triadimenol was assessed by using a leaf disc assay. Resistance factors (RF) ranged from 1.8 to 26.0. The gene encoding the target of DMIs (eburicol 14 alpha-demethylase) from five sensitive and seven resistant isolates was cloned and sequenced. A single mutation, leading to the substitution of a phenylalanine residue for a tyrosine residue at position 136, was found in all isolates exhibiting an RF higher than 5. No mutation was found in sensitive or weakly resistant (RF, < 5) isolates. An allele-specific PCR assay was developed to detect the mutation. Among the 91 isolates tested, only isolates with RF higher than 5 carried the mutation. Three of the 19 resistant isolates and all sensitive and weakly resistant isolates did not possess the mutation. The mutation at codon 136 is thus clearly associated with high levels of resistance to triadimenol.     

Grapevine powdery mildew (Erysiphe necator): a fascinating system for the study of the biology, ecology and epidemiology of an obligate biotroph

Few plant pathogens have had a more profound effect on the evolution of disease management than Erysiphe necator, which causes grapevine powdery mildew. When the pathogen first spread from North America to England in 1845, and onwards to France in 1847, 'germ theory' was neither understood among the general populace nor even generally accepted within the scientific community. Louis Pasteur had only recently reported the microbial nature of fermentation, and it would be another 30 years before Robert Koch would publish his proofs of the microbial nature of certain animal diseases. However, within 6 years after the arrival of the pathogen, nearly 6 million grape growers in France were routinely applying sulphur to suppress powdery mildew on nearly 2.5 million hectares of vineyards (Campbell, 2006). The pathogen has remained a focus for disease management efforts ever since. Because of the worldwide importance of the crop and its susceptibility to the disease, and because conventional management with modern, organic fungicides has been compromised on several occasions since 1980 by the evolution of fungicide resistance, there has also been a renewed effort worldwide to explore the pathogen's biology and ecology, its genetics and molecular interactions with host plants, and to refine current and suggest new management strategies. These latter aspects are the subject of our review. Taxonomy: The most widely accepted classification follows. Family Erysiphaceae, Erysiphe necator Schw. [syn. Uncinula necator (Schw.) Burr., E. tuckeri Berk., U. americana Howe and U. spiralis Berk. & Curt; anamorph Oidium tuckeri Berk.]. Erysiphe necator var. ampelopsidis was found on Parthenocissus spp. in North America according to Braun (1987), although later studies revealed isolates whose host range spanned genera, making the application of this taxon somewhat imprecise (Gadoury and Pearson, 1991). The classification of the genera before 1980 was based on features of the mature ascocarp: (i) numbers of asci; and (ii) morphology of the appendages, in particular the appendage tips. The foregoing has been supplanted by phylogeny inferred from the internal transcribed spacer (ITS) of ribosomal DNA sequences (Saenz and Taylor, 1999), which correlates with conidial ontogeny and morphology (Braun et al., 2002). Host range: The pathogen is obligately parasitic on genera within the Vitaceae, including Vitis, Cissus, Parthenocissus and Ampelopsis (Pearson and Gadoury, 1992). The most economically important host is grapevine (Vitis), particularly the European grape, V. vinifera, which is highly susceptible to powdery mildew. Disease symptoms and signs: In the strictest sense, macroscopically visible mildew colonies are signs of the pathogen rather than symptoms resulting from its infection, but, for convenience, we describe the symptoms and signs together as the collective appearance of colonized host tissues. All green tissues of the host may be infected. Ascospore colonies are most commonly found on the lower surface of the first-formed leaves near the bark of the vine, and may be accompanied by a similarly shaped chlorotic spot on the upper surface. Young colonies appear whitish and those that have not yet sporulated show a metallic sheen. They are roughly circular, ranging in size from a few millimetres to a centimetre or more in diameter, and can occur singly or in groups that coalesce to cover much of the leaf. Senescent colonies are greyish, and may bear cleistothecia in various stages of development. Dead epidermal cells often subtend the colonized area, as natural mortality in the mildew colony, the use of fungicides, mycoparasites or resistance responses in the leaf result in the deaths of segments of the mildew colony and infected epidermal cells. Severely affected leaves usually senesce, develop necrotic blotches and fall prematurely. Infection of stems initially produces symptoms similar to those on leaves, but colonies on shoots are eventually killed as periderm forms, producing a dark, web-like scar on the cane (Gadoury et al., 2011). Inflorescences and berries are most susceptible when young, and can become completely coated with whitish mildew. The growth of the berry epidermal tissue stops when severely infected, which may result in splitting as young fruit expand. Berries in a transitional stage between susceptible and resistant (generally between 3 and 4 weeks after anthesis) develop diffuse, nonsporulating mildew colonies only visible under magnification. Diffuse colonies die as berries continue to mature, leaving behind a network of necrotic epidermal cells (Gadoury et al., 2007). Survival over winter as mycelium in buds results in a distinctive foliar symptom. Shoots arising from these buds may be heavily coated with fungal growth, stark white in colour and stand out like white flags in the vine, resulting in the term 'flag shoots'. More commonly, colonization of a flag shoot is less extensive, and infection of a single leaf, or of leaves on one side of the shoot only, is observed (Gadoury et al., 2011).     

Use of potassium bicarbonate (Armicarb) on the control of powdery mildew (Sphaerotheca mors-uvae) of gooseberry (Ribes uva-crispa)

Powdery mildew (Sphaerotheca mors-uvae) severely infects young shoots, stems and fruits of gooseberry (Ribes uva-crispa). Environmental friendly and biological control measures are being sought throughout the world. Especially in organic gooseberry growing effective control measures are needed, because powdery mildew infections may result in a total loss of the crop. In organic currant growing the number of adequate control methods is very limited. Sulphur as a fungicide against powdery mildew in e.g. gooseberry or table grape growing is not recommendable due to possible bleaching of berries and scorching of tender shoots. Various bicarbonate salts are suggested as a good option to control powdery mildew. In a field trial the effect of potassium bicarbonate (Armicarb) on the on the control of powdery mildew of gooseberry was evaluated. Four treatments; i.e. two preventive strategies and two curative strategies, were applied. PLants were sprayed until runoff. The percent of infected fruits and disease severity were assessed. In the unsprayed control plots very high disease incidences were observed; on average more than 90% of the berries were infected with powdery mildew. The weekly (preventive) spray applications with potassium carbonate reduced the disease incidences on fruits significantly. On average approximately 10% of the fruits were affected by powdery mildew. However, the number of spray applications was high. In conclusion, our results indicate that applications of potassium bicarbonate (as Armicarb) are effective in reducing the incidence and severity of American powdery mildew in gooseberry. Early spray applications are necessary to protect berries against powdery mildew infections. Future research will focus on reducing the number of applications, e.g. warning models based on powdery mildew of rose (Sphaerotheca pannosa).     

Preventative and Curative Applications of Verticillium lecanii for Biological Control of Cucumber Powdery Mildew

The effect of timing of the application of the mycoparasite , Verticillium lecanii, on cucumber powdery mildew , Sphaerotheca fuliginea, was studied in a rooted cucumber leaf bioassay . The mycoparasite was applied at different times before and after mildew inoculation . At near - to maximum humidity ( 95% relative humidity) , early preventative (9 and 5 days before mildew inoculation) and early curative control treatments (2 days after mildew inoculation) gave considerable reduction in mildewed leaf areas , while late curative treatments resulted in greater mildewed leaf area but ultimately a reduced amount of healthy mildewed leaf area ( 20 % ) . Appropriate timing of biocontrol treatment application of V. lecanii is important to achieve good control .     

The integration of host resistance with fungicides in the control of oat powdery mildew

Treatment of seed with the systemic fungicides triadimenol plus fuberidazole as Baytan significantly decreased powdery mildew in field experiments involving eight oat cultivars differing widely in their resistance to the disease. This was so up to growth stage G.S. 61 (Zadoks) even in a year of high mildew incidence, although the effect diminished after flowering. Significant (P≥ 0.001) interactions between fungicide treatments and cultivars occurred in each of three years at nearly all assessment times. In years of high mildew incidence (1983 and 1984), susceptible cultivars developed similar levels of mildew in seed-treated and untreated plots by about flowering time, although cultivars with adult plant resistance (APR) had significantly less mildew when seed-treated than when left untreated. Later, as the APR was fully expressed and the fungicide effectiveness was declining, similar mildew levels were recorded on treated and untreated plots. Untreated APR cultivars generally had less mildew than treated susceptible cultivars and in a year of late and light mildew, APR alone provided good protection. Seed treatments combined with foliar sprays of tridemorph as Calixin almost completely controlled mildew except on very susceptible cultivars late in the season in high mildew years. Over all cultivars, seed treatment gave yield advantages of 5.3% (1983) and 6.6% (1984) but in 1982, a year of low mildew, the response was small. The possible influence of integrated host resistance and fungicides in stabilising the pathogen population at economically unimportant levels and the environmental benefit of using host resistance to minimise fungicide usage is discussed.     

Effects of elevated atmospheric CO2 and temperature on the management of powdery mildew of zucchini

The impact of combined environmental factors, such as temperature and CO₂, on the control of the powdery mildew of zucchini, caused by Podosphaera xanthii, and of different control measures has been studied on plants grown in phytotrons. Five experimental trials were conducted, and the powdery mildew severity of both treated and untreated zucchini plants was found to be significantly affected by the interaction between temperature (three different regimes: 16–18; 18–22; 22–26°C), CO₂ (two concentrations: 400–450 and 800–850 ppm) and the treatments. However, at the end of the trials, the efficacy of all the products was not affected by the different, tested environmental conditions. Sulphur consistently provided the highest disease control (75%–85% efficacy). Among the resistant inducers that were tested, calcium oxide was the most effective, in terms of powdery mildew control under all the conditions tested in phytotrons, reducing disease severity from 46% to 61%. Foliar applications of phosphite (14%–28% efficacy), Ampelomyces quisqualis (12%–23% efficacy) and potassium silicate (13%–24% efficacy) only slightly reduced the disease severity for all the tested temperature regimes and CO₂ concentrations, compared to the untreated control. The results obtained under our experimental conditions show that a possible increase in CO₂ concentration and temperature, which is expected for the next few years, should not influence the efficacy of the tested resistance inducers or of sulphur against powdery mildew on zucchini. Moreover, the suppressive effect of calcium oxide is in light of its possible use in greenhouses for zucchini powdery mildew control under 400–450 ppm of CO₂ and under enriched condition of 800–850 ppm of CO₂.     

Genetic similarity of flag shoot and ascospore subpopulations of Erysiphe necator in Italy

The overwintering mode of the grape powdery mildew fungus, Erysiphe necator (syn. Uncinula necator), as mycelium in dormant buds (resulting in symptoms known as flag shoots) or as ascospores in cleistothecia, affects the temporal dynamics of epidemics early in the growing season. We tested whether distinct genetic groups (I and III) identified previously in E. necator correlate to overwintering modes in two vineyards in Tuscany, Italy, to determine whether diagnostic genetic markers could be used to predict overwintering. Samples from one vineyard were collected from flag shoots; the other vineyard, 60 km away, had no flag shoots, and mildew colonies were assumed to be derived from ascospores. Genetic markers putatively diagnostic for groups I and III showed that both types were common in the flag shoot subpopulation. Both genetic types were found in the ascospore population, although group III was dominant. We did not find strong genetic differentiation between the two subpopulations based on inter-simple sequence repeat markers. Although there was significant (P < 0.001) genetic differentiation between these subpopulations in 1997 and when 1997 and 1998 subpopulations were pooled (theta = 0.214 and 0.150, respectively), no differentiation was evident between vineyards in 1998 (theta = 0.138, P = 0.872). Moreover, we did not observe distinct lineages corresponding to overwintering modes, as observed in previous studies. We could not determine if differentiation resulted from biological differences or restricted gene flow between the two vineyards. Our samples were taken from both subpopulations early in the epidemic, while previous studies confounded overwintering mode and sampling time. These results do not support a strong correlation between overwintering and genetic groups, highlighting the need to base population biology studies on sound biological and epidemiological knowledge.     

Detection and quantification of Erysiphe necator DNA in wine grapes and resultant must and juice

Powdery mildew of grapevines is difficult to assess visually at the weighbridge, particularly in large consignments of machine-harvested fruit. To facilitate accurate methods for the detection and quantification of the disease in grape samples obtained from both the vineyard and winery, we developed a DNA probe for the pathogen Erysiphe necator. The E. necator-specific 450 bp DNA fragment pEnA1, targets highly repetitive sequences and was isolated from a partial genomic library. In screening for species specificity, clone pEnA1 was used in slot-blot hybridization and detected E. necator DNA from grapes and resultant must and juice, but not from clarified juice and wine. The detection threshold was approximately 50 pg ofE. necator DNA per 100 ng total DNA of grape sample and was equivalent to 1–5 % of a grape bunch visually affected by powdery mildew. Disease severity, expressed as the percentage of surface area of a bunch with powdery mildew, and E. necator DNA content were highly correlated, r² = 0.955, P < 0.001. The DNA-based hybridization assay has the potential to predict the severity of powdery mildew in grape samples from the vineyard and in must and juice samples at the winery. The DNA sequence of clone pEnA1 was used to design species-specific primers, the results maintaining the same specificity patterns observed in the initial hybridization assays. The PCR-based assay was sensitive enough to detect approximately 1 pg DNA, being equivalent to 1 conidium per sample. This is the first report to date of the detection of all known phenetic groups of E. necator DNA and of the quantification of DNA from grape samples at the winery. Accurate information on the extent of powdery mildew contamination of grape lots would enable wineries to make more informed decisions about the use of fruit and must.     

Evaluation of organic,conventional and intensive beef farm systems: health,management and animal production

The overall aim of the present study was to analyse and compare organic beef cattle farming in Spain with intensive and conventional systems. An on-farm study comparing farm management practices and animal health was carried out. The study also focussed on a slaughterhouse analysis by comparing impacts on the safety and quality of the cattle products. Twenty-four organic and 26 conventional farms were inspected, and farmers responded to a questionnaire that covered all basic data on their husbandry practices, farm management, veterinary treatments and reproductive performance during 2007. Furthermore, data on the hygiene and quality of 244, 2596 and 3021 carcasses of calves from organic, intensive and conventional farms, respectively, were retrieved from the official yearbook (2007) of a slaughterhouse. Differences found between organic and conventional farms across the farm analysis did not substantially reflect differences between both farm types in the predominant diseases that usually occur on beef cattle farms. However, calves reared organically presented fewer condemnations at slaughter compared with intensive and to a lesser extent with conventionally reared calves. Carcass performance also reflected differences between farm type and breed and was not necessarily better in organic farms.     

Effects of Sunflower Oil on Tomato Powdery Mildew Caused by Oidium neolycopersici

When tomato leaves were sprayed with 0.1% emulsified canola oil, corn oil, grape seed oil, peanut oil, safflower oil, soya bean oil or sunflower oil, the severity of powdery mildew caused by Oidium neolycopersici was greatly reduced. Among these edible oils tested, sunflower oil was the most effective in the control of powdery mildew. When sprayed with 0.5% sunflower oil, powdery mildew on tomato leaves was reduced to a negligible level. Sunflower oil applied to halves of upper leaf surface did not induce resistance against the pathogen in the non‐treated halves. When applied to halves of lower leaf surface, it also failed to reduce the severity of powdery mildew on the upper leaf surface right above the treated area indicating that control of the powdery mildew by sunflower oil did not result from activation of host defence mechanisms. Scanning electron microscopy showed that control of powdery mildew with sunflower oil resulted mainly from the inhibition of conidial germination and suppression of mycelial growth of the pathogen.     

Identification and characterization of a fruit-specific, thaumatin-like protein that accumulates at very high levels in conjunction with the onset of sugar accumulation and berry softening in grapes.

The protein composition of the grape (Vitis vinifera cv Muscat of Alexandria) berry was examined from flowering to ripeness by gel electrophoresis. A protein with an apparent molecular mass of 24 kD, which was one of the most abundant proteins in extracts of mature berries, was purified and identified by amino acid sequence to be a thaumatin-like protein. Combined cDNA sequence analysis and electrospray mass spectrometry revealed that this protein, VVTL1 (for V. vinifera thaumatin-like protein 1), is synthesized with a transient signal peptide as seen for apoplastic preproteins. Apart from the removal of the targeting signal and the formation of eight disulfide bonds, VVTL1 undergoes no other posttranslational modification. Southern, northern, and western analyses revealed that VVTL1 is found in the berry only and is encoded by a single gene that is expressed in conjunction with the onset of sugar accumulation and softening. The exact role of VVTL1 is unknown, but the timing of its accumulation correlates with the inability of the fungal pathogen powdery mildew (Uncinula necator) to initiate new infections of the berry. Western analysis revealed that the presence of thaumatin-like proteins in ripening fruit might be a widespread phenomenon.     

RAPD analysis of local bell pepper genotypes in relation with powdery mildew incidence and fruit yield: Managing disease by single chemical molecule

Abstract

A field study was conducted to investigate powdery mildew disease incidence and fruit yield of locally available bell pepper genotypes by Kharif 2004 and Rabi 2005. The study includes a single chemical molecule in managing the powdery mildew disease during both the seasons. Field studies include five chemical treatments (T₁ to T₅) plus one water spray control and an untreated control in five replications each. The result shows that out all treatments T₅ was best in controlling the powdery mildew disease in genotype Indra by 77.1 and 73.7% of disease decrease over control and 85.3 and 95.7% increase in yield over control respectively during Kharif and Rabi. The next best performing genotypes were Arka Basant and California Wonder in terms of less disease incidence and high yield. Further, the study was extended to relate the selected varieties for its genetic diversity using Random Amplified Polymorphic DNAs, with respect to the disease incidence and yield. The highest genetic dissimilarity of 49% was noticed between the genotypes Indra and Arka Gaurav having contrasting characters such as high and low fruit yield, respectively. The lowest genetic diversity of 21% was noticed between the genotypes Arka Mohini and California Wonder that were similar in most of the morphological characters, fruit yield and susceptibility to the powdery mildew disease. Based on the molecular data, a remarkable difference was exhibited by the commercially grown bell pepper genotypes.     

烟草白粉病抗性基因的遗传分析

以烟草抗白粉病品种台烟7号为母本,感病品种NC89为父本,构建6个世代的群体,利用主基因 多基因混合遗传模型的分离分析方法,研究烟草白粉病的抗性遗传规律。结果表明,烟草白粉病抗性的遗传是由两对加性-显性-上位性主基因 加性-显性-上位性多基因（E-0模型）控制的。B1、B2和F2世代主基因的遗传率分别为88.05%、32.62%、84.43%,主基因遗传率很大,说明可以在抗病育种早期进行选择;B1、F2世代多基因遗传率均为0.00%,说明烟草白粉病的发生受一定环境影响。     

More years of field trials against Plasmopara viticola in organic viticolture

The organic viticulture assumes a decisive role in the national agricultural sector. More impelling problems in the management of organic vineyards are represented from the plants pathology defence and particularly of Plasmopara viticola containment. Copper represents one of the few usable fungicides in the organic farming and the only effective against downy mildew. With Regulation EC n. 473/2002, fixed maximum quantity usable of copper compounds, owing to the environment problems due to the copper accumulation in the soil. To reduce quantity of metal copper or replace it with natural products, are conducted field trials with copper compounds at a low rate or alternative to copper products. Besides, we are estimating possibility to reduce the operations against P. viticola optimizing fungicidal treatments. Field trials in the organic farms located near Rome, have been carried out. Guidelines EPPO/OEPP PP 1/31 (3) have been carried out. The results of the trials have showed that, using cupric products with low metallic content, to reduce copper quantities used, always allowed to respect the limits established by Regulation EC. The alternative products that were investigated have not guaranteed, instead, an adequate protection in high pressure of grapevine downy mildew. It was possible to reduce treatments against P. viticola through control of different environmental parameters. The trials confirm that the copper is indispensable for plant protection in organic farming as it is not possible to replace it with natural extracts substances. We can reduce, instead, the copper quantities used trough the use of new products with low quantity metal copper or through the evaluation of climatic and pedologic data that allow to rationalize the fungicidal treatments.     

Using aerated compost tea in comparison with a chemical pesticide for controlling rose powdery mildew

Rose powdery mildew (Sphaerotheca pannosa var. rosae) is one of the most common foliar diseases of roses worldwide. Application of chemical products on the plant or in the soil kills a range of the beneficial micro-organisms thereby disturbing ecosystem. Compost tea helps to restore and increase the populations of those beneficial micro-organisms. The objective of the present study was to evaluate the comparison of biopesticide (compost tea) and a chemical pesticide. The experiment was performed in three treatments, which were compost tea, fungicide (Topaz) and no treatment in three replications. After foliar applications of biopesticide and fungicide, the control percentage was estimated based on the number of infected flowers with powdery mildew. The results indicated that there was a significant difference between these treatments on rose in controlling powdery mildew (F?=?23.25, p?=?0.0015, df?=?2), at a probability level of 1% (p???0.01). So, that control percentage of compost tea treatment was the most.