Powdery mildew resistance genes in vines: an opportunity to achieve a more sustainable viticulture

Grapevine (Vitis vinifera) is one of the main fruit crops worldwide. In 2020, the total surface area planted with vines was estimated at 7.3 million hectares. Diverse pathogens affect grapevine yield, fruit, and wine quality of which powdery mildew is the most important disease prior to harvest. Its causal agent is the biotrophic fungus Erysiphe necator, which generates a decrease in cluster weight, delays fruit ripening, and reduces photosynthetic and transpiration rates. In addition, powdery mildew induces metabolic reprogramming in its host, affecting primary metabolism. Most commercial grapevine cultivars are highly susceptible to powdery mildew; consequently, large quantities of fungicide are applied during the productive season. However, pesticides are associated with health problems, negative environmental impacts, and high costs for farmers. In paralleled, consumers are demanding more sustainable practices during food production. Therefore, new grapevine cultivars with genetic resistance to powdery mildew are needed for sustainable viticulture, while maintaining yield, fruit, and wine quality. Two main gene families confer resistance to powdery mildew in the Vitaceae, Run (Resistance to Uncinula necator) and Ren (Resistance to Erysiphe necator). This article reviews the powdery mildew resistance genes and loci and their use in grapevine breeding program

Genetic and molecular analysis of mildew disease resistance in grapevine

The thesis aimed the genetic and molecular analysis of mildew disease resistance in grapevine. The following investigations were performed: i) construction of a genetic map and localisation of quantitative traits conferring resistance to powdery and downy mildew; ii)Integration of functional and structural resistance candidate genes into the genetic map; iii)Transcriptional analysis of grapevine after infection with powdery mildew and iv) Isolation and characterization of resistance gene family

Gombarezisztencia gének térképezése szőlőben = Mapping resistance genes against fungi in grapevine

Lisztharmat (PM) és peronoszpóra (DM) rezisztencia génekkel kapcsolt markerek szelekcióra való alkalmasságát vizsgáltuk szőlő inter-és intraspecifikus térképezési populációiban. Az interspecifikus hibridek a Muscadina rotundifolia x Vitis vinfera BC4 Cardinal, Kismis moldavszkij és Kismis vatkana fajtákkal előállított BC5 nemzedékei voltak. A M. rotundifolia az ismert RUN1 (PM) és az RPV1 (DM) domináns rezisztencia géneket tartalmazza. A BC5 nemzedékekben 1 CAPS és 3 SSR markerrel hatékonyan szelektáltuk a rezisztens genotípusokat. A V. vinifera fajták általában fogékonyak a lisztharmatra, de a fogékonyságuk eltérő. A Dzsandzsal karát írták le először PM rezisztens fajtaként, később azonban többet is azonosítottak, köztük a Kismis vatkanát, rezisztencia génjüket azonban nem jellemezték. A Nimrang x Kismis vatkana hibrid család elemzése során bebizonyosodott, hogy a Kismis vatkana PM génje, amelyet REN1-nek neveztek el, nem azonos a RUN1-gyel. A 13-as kromoszómára térképeződött, míg a RUN1 a 12-re. A REN1 körül azonosított 3 SSR markerrel genotipizáltuk a Génuai zamatos x Kismis vatkana és BC4 x Kismis vatkana utódokat. Az utóbbi család egyedei közül RUN1/REN1 piramidált genotípusokat szelektáltunk. Az azonos fenotípust meghatározó piramidált géneket tartalmazó növények azonosítása csak DNS-szintű elemzéssel lehetséges. A MAS hatékonyságának növelésére multiplex PCR módszert dolgoztunk ki. A REN1-gyel kapcsolt marker SSR profil alapján a Dzsandzsal kara is REN1 gént hordoz. | For validating markers linked to powdery (PM) and downy (DM) mildew resistance genes, applying them in marker assisted selection (MAS) we analyzed mapping populations, deriving from interspecific crosses of Vitis vinifera with Muscadinia rotundifolia carrying the dominant RUN1 (PM) and RPV1 (DM) resistance genes. One CAPS and 3 SSR markers proved to be adequate for selecting RUN1/RPV1 genotypes in the (M. rotundifolia x V. vinifera) BC4 x Cardinal, BC4 x Kishmish moldavskij and BC4 x Kishmish vatkana families. Kishmish vatkana is a PM resistant V. vinifera cultivar such as Dzhandzhal kara. Involving V. vinifera resistance genes into breeding gives the chance to avoid interspecific crosses. Analysis of a Nimrang x Kishmis vatkana progeny proved that PM resistance gene of Kishmish vatkana, called REN1, is different from RUN1. REN1 mapped into linkage group/LG 13, while RUN1 is in LG12. Three SSR markers were identified around the REN1 locus and applied for MAS in Génuai zamatos x Kishmis vatkana and BC4 x Kishmish vatkana hybrids. In this latter cross we proved the presence of the pyramided PM resistance genes. Plants carrying both RUN1 and REN1 for the same phenotype can be identified only with DNA analysis. This is the first time when SSR markers linked to REN1 were used for MAS. We elaborated a multiplex PCR method suitable for agarose electrophoresis. SSR profiles in REN1 linked loci suggest that Kismish vatkana and Dzhandzhal kara possess the same REN1 PM resistance gene

Intracellular mycoparasites in action: interactions between powdery mildew fungi and Ampelomyces

Pycnidial fungi of the genus Ampelomyces are widespread intracellular mycoparasites of powdery mildew fungi worldwide. Their pycnidia are produced in hyphae, conidiophores and immature ascomata of their mycohosts. Thus, they suppress both the asexual and the sexual reproduction of the invaded powdery mildew mycelia, and then destroy them completely. Conidia of Ampelomyces are released from the intracellular pycnidia by the rupture of the pycnidial wall; conidia then germinate on the host plant surfaces, penetrate the intact hyphae of powdery mildew mycelia found in their vicinity and invade them internally growing from cell to cell through the septal pores of the mycohost. The early stage of mycoparasitism is apparently biotrophic, but the invaded cytoplasm then begins to die and a necrotrophic interaction results. Toxin production has not been detected in Ampelomyces, so it might act directly by invasion and destruction of the host cytoplasm. Experimental data showed that parasitized powdery mildew colonies can continue their growth, but their sporulation is stopped soon after Ampelomyces penetrated their mycelia. It is concluded that these mycoparasites represent a stress factor in the life cycle of their mycohosts but their role in the natural control of powdery mildew infections requires further investigations

Results from new fungus-tolerant grapevine varieties for Organic Viticulture

Two red and three white new fungus-tolerant grape varieties were tested within a period of five years. REGENT, RONDO, JOHANNITER and Gf 48-12 show a better wine quality than PINOT NOIR or SILVANER and can be recommended for Organic Viticulture as well as for the conventional viticulture to reduce copper and fungicide applications

Fungicides and the grapevine wood mycobiome: a case study on Tracheomycotic Ascomycete Phaemoniella chlamydospora reveals potential for two novel control strategies

Original ResearchPhaeomoniella chlamydospora is a tracheomycotic fungus that colonizes the xylem of grapevines (Vitis vinifera L.), causing wood discoloration, brown wood streaking, gummosis, and wood necrosis, which negatively affect the overall health, productivity, and life span of vines. Current control strategies to prevent or cope with P. chlamydospora infections are frequently ineffective. Moreover, it is unclear how fungicides commonly applied in vineyards against downy and powdery mildew agents affect the wood mycobiome, including wood pathogens such as P. chlamydospora. In this study, we used next-generation sequencing to assess the effects of foliar spray of grapevines with inorganic (copper oxychloride and sulfur), synthetic (penconazole and fosetyl-aluminum), and natural (Blad) fungicides currently used against the downy and powdery mildews. The subjects of our investigation were (i) the resident wood mycobiome, (ii) the early colonization by a consortium of fungal wood endophytes (ACEA1), (iii) the wood colonization success of P. chlamydospora, and (iv) the in planta interaction between P. chlamydospora and ACEA1, under greenhouse conditions, in rooted grapevine cuttings of cv. Cabernet Sauvignon. The data obtained suggest that the resident mycobiome is affected by different fungicide treatments. In addition, the early colonization success of the endophytes composing ACEA1 varied in response to fungicides, with relative abundances of some taxa being overrepresented or underrepresented when compared with the control. The wood colonization by P. chlamydospora comported significant changes in the mycobiome composition, and in addition, it was greatly affected by the foliar spray with Blad, which decreased the relative abundance of this pathogen 12-fold (4.9%) when compared with the control (60.7%) and other treatments. The presence of the pathogen also decreased considerably when co-inoculated into the plant with ACEA1, reaching relative abundances between 13.9% and 2.0%, depending on the fungicide treatment applied. This study shows that fungicides sprayed to prevent infections of powdery and downy mildews have an control strategies to fight P. chlamydospora, namely, the foliar spray with Blad and the use of ACEA1. Further studies to confirm these results are requiredinfo:eu-repo/semantics/publishedVersio

Adaptive genomic structural variation in the grape powdery mildew pathogen, Erysiphe necator.

BackgroundPowdery mildew, caused by the obligate biotrophic fungus Erysiphe necator, is an economically important disease of grapevines worldwide. Large quantities of fungicides are used for its control, accelerating the incidence of fungicide-resistance. Copy number variations (CNVs) are unbalanced changes in the structure of the genome that have been associated with complex traits. In addition to providing the first description of the large and highly repetitive genome of E. necator, this study describes the impact of genomic structural variation on fungicide resistance in Erysiphe necator.ResultsA shotgun approach was applied to sequence and assemble the genome of five E. necator isolates, and RNA-seq and comparative genomics were used to predict and annotate protein-coding genes. Our results show that the E. necator genome is exceptionally large and repetitive and suggest that transposable elements are responsible for genome expansion. Frequent structural variations were found between isolates and included copy number variation in EnCYP51, the target of the commonly used sterol demethylase inhibitor (DMI) fungicides. A panel of 89 additional E. necator isolates collected from diverse vineyard sites was screened for copy number variation in the EnCYP51 gene and for presence/absence of a point mutation (Y136F) known to result in higher fungicide tolerance. We show that an increase in EnCYP51 copy number is significantly more likely to be detected in isolates collected from fungicide-treated vineyards. Increased EnCYP51 copy numbers were detected with the Y136F allele, suggesting that an increase in copy number becomes advantageous only after the fungicide-tolerant allele is acquired. We also show that EnCYP51 copy number influences expression in a gene-dose dependent manner and correlates with fungal growth in the presence of a DMI fungicide.ConclusionsTaken together our results show that CNV can be adaptive in the development of resistance to fungicides by providing increasing quantitative protection in a gene-dosage dependent manner. The results of this work not only demonstrate the effectiveness of using genomics to dissect complex traits in organisms with very limited molecular information, but also may have broader implications for understanding genomic dynamics in response to strong selective pressure in other pathogens with similar genome architectures

Identification and expression analysis of Fragaria vesca MLO genes involved in interaction with powdery mildew (Podosphaera aphanis)

Strawberry powdery mildew, caused by Podosphaera aphanis is a major fungal disease that affects strawberry yield and quality. In the model plant species Arabidopsis and the crop plants barley, tomato and pea, the Mildew resistance locus O (MLO) proteins have been found to be required for powdery mildew susceptibility. The present study, based on the sequence of a wild plum (Prunus americana) MLO protein, identified 16 MLO genes within the genome of woodland strawberry, Fragaria vesca and examined their expression pattern in response to powdery mildew infection in three diploid strawberry cultivars. Phylogenetic analysis showed that the FvMLO genes can be classified into six clades. Four FvMLO genes were grouped into clade III, which comprises MLO genes from Arabidopsis, tomato and grapevine that mediate powdery mildew susceptibility. A RNA-seq analysis of two diploid strawberry cultivars, F. vesca ssp. vesca accession Hawaii 4 (HW) and F. vesca f. semperflorens line “Yellow Wonder 5AF7” (YW) at 1 d (1 DAI) and 8 d (8 DAI) after infection showed the expression of 12 out of the 16 FvMLO genes. The comparison of Fragments Per Kilobase of transcript per Million mapped reads (FPKM values) detected by RNA-seq and expression values of qRT-PCR for FvMLO genes showed substantial agreement. The FvMLO3 gene, which was grouped in clade III and orthologous to the Arabidopsis, tomato and grapevine genes, was highly expressed in YW compared to other FvMLO genes across varieties. The results showed that FvMLO genes can be used as potential candidates to engineer powdery mildew resistance in strawberry based on MLO suppression or genome editing

Investigations on the influence of N fertilizer on resistance to powdery mildew (Oidium tuckeri) downy mildew (Plasmopara viticola) and on phytoalexin synthesis in different grapevine varieties

1-year-old grapevine cuttings (cvs. Gf Ga-58-30, Fr 993-60, Riesling, Kerner) were grown in a greenhouse in pots at four levels of nitrogen supply, in order to test their resistance to powdery mildew, downy mildew, and their possible stilbene synthesis.Our most significant findings are that: increased rates of nitrogen supply decrease the degree of resistance against powdery and downy mildew and phytoalexin (stilbene) synthesis; the hybrids (Gf Ga-58-30 and Fr 993-60) show a higher powdery and downy mildew resistance and a higher stiloene synthesis than the V. 11inifera varieties (Riesling and Kerner); the amount of stilbene (resveratrol and E-viniferin) increases during the growing cycle