Genotyping wine and table grape cultivars from Apulia (Southern Italy) using microsatellite markers

Thirty-eight typical grapevine varieties of the Apulia region, Southern Italy, were genotyped at 6 microsatellite loci (VVS2, VVMD5, VVMD7, VrZAG47, VrZAG62, VrZAG79) with the aim to find synonymy and to confirm some pedigrees reported in literature. Microsatellites were amplified by PCR with 33P-ATP labelled primers and alleles separated by electrophoresis using 6 % acrylamide sequencing gels. The results confirm the high information level of the selected microsatellites. The number of alleles ranged from 7 to 11, producing up to 23 different combination patterns. The observed heterozygosity varied between 81.6 and 94.7 %, the discrimination power between 0.888 and 0.939, and the probability of identity was as low as 0.06-0.12. All cultivars of the study were discriminated from each other, except Regina (syn. Afuz Ali) and Mennavacca, which had the same profile. Finally, we were able to confirm the parentage of Victoria (Cardinal x Afuz Ali) and Matilde (Italia x Cardinal).

Characterisation of the grapevine cultivar Picolit by means of morphological descriptors and molecular markers

The phenotypic and genotypic variability of cv. Picolit, an ancient, female-flower cultivar from north-eastern Italy was investigated by means of ampelographic and ampelometric descriptors and by molecular markers, such as microsatellites and AFLPs. Thirty nine samples were collected from old plants (30-100 years old), which showed some differences in morphology and growth. In two samples (P6 and P7) morphological differences were found. These samples showed a different allelic profile at 18 out of the 21 SSRs analysed and were therefore considered not to belong to the cv. Picolit. Of the remaining samples, 35 gave the same allelic pattern at all SSRs and they were therefore considered ‘true-to-type’ Picolit, whereas two of them (P4 and P8) showed several variations, including extra alleles. One of the possible causes of such differences is chimerism. The AFLP analysis, from which samples P6 and P7 were excluded, enabled screening of a larger portion of the genome and confirmed the differences of the P4 and P8 samples from the remaining ones. P4 and P8 were different from the majority of samples at 13 and 37 AFLP loci respectively. A few further polymorphic bands were recorded in the remaining samples, but they were disregarded since they were not always reproducible. This research confirmed the appreciable somatic stability of SSR markers even in long-lived, vegetatively propagated plants, and the occasional occurrence of solid mutations and chimerisms.

Genetics-assisted breeding for downy/powdery mildew and phylloxera resistance at fem

The genetics-assisted program for resistance to biotic stresses began at the Edmund Mach Foundation in 2007 and has developed on two lines. The first line was based on obtaining resistant materials with parents the historical varieties of Trentino. During the three-year period 2015–2017, 3 Teroldego X Merzling genotypes, 3 Marzemino X Merzling genotypes, 1 Nosiola X Bianca genotype and 1 Nosiola X Kulneany genotype were collected the data related to resistance to various fungal diseases both on the leaf and on the bunch. At harvest the plant production data and the must characteristics were recorded; the grapes were vinified and for each year the wines were subjected to sensorial analysis. 5 resistant selections were considered interesting for all the characteristics found and therefore the data for the registration to the National Register of Grapevine Varieties will be collected. The second line of research was based on the retrieval and both genotypic and phenotypic characterization of potentially parental lines acquired from foreign breeding programs and of wild materials. Once the crossings have been planned and obtained, the evaluation of the progeny takes place following a process of Marker-Assisted Selection optimized in order to maintain a compromise between efficiency and cost containment

Secondary and primary metabolites reveal putative resistance-associated biomarkers against Erysiphe necator in resistant grapevine genotypes

Numerous fungicide applications are required to control Erysiphe necator, the causative agent of powdery mildew. This increased demand for cultivars with strong and long-lasting field resistance to diseases and pests. In comparison to the susceptible cultivar ‘Teroldego’, the current study provides information on some promising disease-resistant varieties (mono-locus) carrying one E. necator-resistant locus: BC4 and ‘Kishmish vatkana’, as well as resistant genotypes carrying several E. necator resistant loci (pyramided): ‘Bianca’, F26P92, F13P71, and NY42. A clear picture of the metabolites’ alterations in response to the pathogen is shown by profiling the main and secondary metabolism: primary compounds and lipids; volatile organic compounds and phenolic compounds at 0, 12, and 48 hours after pathogen inoculation. We identified several compounds whose metabolic modulation indicated that resistant plants initiate defense upon pathogen inoculation, which, while similar to the susceptible genotype in some cases, did not imply that the plants were not resistant, but rather that their resistance was modulated at different percentages of metabolite accumulation and with different effect sizes. As a result, we discovered ten up-accumulated metabolites that distinguished resistant from susceptible varieties in response to powdery mildew inoculation, three of which have already been proposed as resistance biomarkers due to their role in activating the plant defense response

Semi-Targeted Profiling of the Lipidome Changes Induced by Erysiphe Necator in Disease-Resistant and Vitis vinifera L. Varieties

The ascomycete Erysiphe necator is a serious pathogen in viticulture. Despite the fact that some grapevine genotypes exhibit mono-locus or pyramided resistance to this fungus, the lipidomics basis of these genotypes’ defense mechanisms remains unknown. Lipid molecules have critical functions in plant defenses, acting as structural barriers in the cell wall that limit pathogen access or as signaling molecules after stress responses that may regulate innate plant immunity. To unravel and better understand their involvement in plant defense, we used a novel approach of ultra-high performance liquid chromatography (UHPLC)-MS/MS to study how E. necator infection changes the lipid profile of genotypes with different sources of resistance, including BC4 (Run1), “Kishmish vatkhana” (Ren1), F26P92 (Ren3; Ren9), and “Teroldego” (a susceptible genotype), at 0, 24, and 48 hpi. The lipidome alterations were most visible at 24 hpi for BC4 and F26P92, and at 48 hpi for “Kishmish vatkhana”. Among the most abundant lipids in grapevine leaves were the extra-plastidial lipids: glycerophosphocholine (PCs), glycerophosphoethanolamine (PEs) and the signaling lipids: glycerophosphates (Pas) and glycerophosphoinositols (PIs), followed by the plastid lipids: glycerophosphoglycerols (PGs), monogalactosyldiacylglycerols (MGDGs), and digalactosyldiacylglycerols (DGDGs) and, in lower amounts lyso-glycerophosphocholines (LPCs), lyso-glycerophosphoglycerols (LPGs), lyso-glycerophosphoinositols (LPIs), and lyso-glycerophosphoethanolamine (LPEs). Furthermore, the three resistant genotypes had the most prevalent down-accumulated lipid classes, while the susceptible genotype had the most prevalent up-accumulated lipid classes

Two-omics data revealed commonalities and differences between Rpv12- and Rpv3-mediated resistance in grapevine

Plasmopara viticola is the causal agent of grapevine downy mildew (DM). DM resistant varieties deploy effector-triggered immunity (ETI) to inhibit pathogen growth, which is activated by major resistance loci, the most common of which are Rpv3 and Rpv12. We previously showed that a quick metabolome response lies behind the ETI conferred by Rpv3 TIR-NB-LRR genes. Here we used a grape variety operating Rpv12-mediated ETI, which is conferred by an independent locus containing CC-NB-LRR genes, to investigate the defence response using GC/MS, UPLC, UHPLC and RNA-Seq analyses. Eighty-eight metabolites showed significantly different concentration and 432 genes showed differential expression between inoculated resistant leaves and controls. Most metabolite changes in sugars, fatty acids and phenols were similar in timing and direction to those observed in Rpv3-mediated ETI but some of them were stronger or more persistent. Activators, elicitors and signal transducers for the formation of reactive oxygen species were early observed in samples undergoing Rpv12-mediated ETI and were paralleled and followed by the upregulation of genes belonging to ontology categories associated with salicylic acid signalling, signal transduction, WRKY transcription factors and synthesis of PR-1, PR-2, PR-5 pathogenesis-related proteins

Piramidazione di loci di resistenza a peronospora e oidio mediante un processo ottimizzato di selezione assistita da marcatori molecolari

Oggi la transizione verso un'agricoltura sostenibile è di primaria importanza per la comunità nazionale ed internazionale. Una strategia promettente per ridurre l'uso di sostanze chimiche è rappresentata dall’incrocio tra varietà con elevato tenore organolettico e varietà resistenti a stress biotici. L'allevamento tradizionale di piante legnose è tuttavia estremamente impegnativo a causa di alcune limitazioni: il lungo ciclo riproduttivo e le grandi dimensioni delle piante rendono il periodo di valutazione molto lungo e costoso. Nell'ultimo decennio l'impiego della selezione assistita da marcatori molecolari (MAS) ha cambiato il paradigma, migliorando l'efficienza delle fasi di screening nel periodo iniziale e riducendo il numero di piantine che raggiungono il campo aperto. Con queste consapevolezza e visione, nel 2010 è iniziato il programma di miglioramento genetico per la resistenza allo stress biotico della vite presso la Fondazione Edmund Mach. Dopo una prima fase di esplorazione di un pool genetico vario e complesso di caratteri/loci di resistenza (R) a peronospora e oidio, è stato selezionato come donatore un gruppo di accessioni. Quindi, attraverso la selezione assistita dei parentali (MAPS), vari genotipi con loci piramidati hanno raggiunto il campo aperto e sono stati utilizzati per scopi di riproduzione. Successivamente, è stata condotta l'ottimizzazione dei protocolli di fenotipizzazione e genotipizzazione per la selezione dei semenzali assistita da marcatori (MASS) altamente efficiente. Sono stati sviluppati due flussi di lavoro di fenotipizzazione per selezionare prima preferenzialmente le resistenze a peronospora o oidio nel germoplasma riproduttivo, combinate possibilmente con la seconda resistenza. Alla fine, i costi di genotipizzazione sono stati ridotti di oltre un quarto (cinque loci selezionati in media per individuo). Nel 2018 circa 650 individui di progenie sono stati analizzati e caratterizzati a nove loci R, rivelando un massimo di sette loci combinati in totale di cui quattro piramidati contro la stessa malattia. Il processo ottimizzato è stato applicato in più di 20 popolazioni segreganti e i diversi assetti R-loci rilevati hanno permesso di svelare effetti sia inter-popolazione che intra-popolazione: il primo riguarda l'effetto di un diverso background genetico sulla stessa risorsa di R-loci, mentre il secondo rappresenta l'impatto di diversi assetti R-loci all'interno della stessa popolazione. Grazie a questi risultati, a partire dal 2019 è stato stabilito un processo di MAS forward ̶ senza screening fenotipico ̶ sulle progenie derivate da quelle linee parentali con un comportamento noto all'interno del programma di miglioramento genetico della FEM

The European Vitis Database (www.eu-vitis.de) – a technical innovation through an online uploading and interactive modification system

The objective of the European Vitis Database is to safeguard and enhance germplasm by monitoring its preservation. Two issues are strongly related to that purpose: (1) participation of collections covering almost all grape biodiversity and (2) assessment of accessions trueness to type. In the scope of the European project GrapeGen06 efforts have been made towards both objectives. The 35 participating grape germplasm repositories are found between the Iberian Peninsula and Transcaucasia, thus covering a broad range of grape diversity. Altogether they maintain 32,410 accessions. However with respect to biodiversity, gaps are still evident and further collections need to be included and trueness to type assessment absolutely needs to be pursued to organize duplication of endangered genotypes. Within the GrapeGen06 project focus was laid on the establishment of a database conferring the collection holders a high degree of responsibility and independence. Hence for the first time in a European Central Crop Database an on-line uploading application and an interactive modification system for data administration was implemented. These innovations disburden the database manager and offer the curators of collections more flexibility. Prerequisites for data import, descriptors applied, access levels, database contents, uploading, export and search functions are described