Wild grapevine: silvestris,hybrids or cultivars that escaped from vineyards? Molecular evidence in Sardinia

Vitis vinifera ssp. silvestris, the spontaneous subspecies of V. vinifera L., is believed to be the ancestor of present grapevine cultivars. In this work, polymorphism at 13 SSR loci was investigated to answer the following key question: are wild plants (i) true silvestris, (ii) hybrids between wild and cultivated plants or (iii) or ‘escapes’ from vineyards? In particular, the objective of the present study was to identify truly wild individuals and to search for possible hybridization events. The study was performed in Sardinia, the second largest island in the Mediterranean Sea, which is characterized by a large and well‐described number of both grape cultivars and wild populations. This region was ideal for the study because of its spatial isolation and, consequently, limited contamination from outside material. The results of this study show that domesticated and wild grapevine germplasms are genetically divergent and thus are real silvestris. Pure lineages (both domesticated and wild) show very high average posterior probabilities of assignment to their own clusters, with a low level of introgression.     

Genetic diversity and parentage of Tunisian wild and cultivated grapevines (Vitis vinifera L.) as revealed by single nucleotide polymorphism (SNP) markers

Based on 261 single nucleotide polymorphism (SNP) markers, we analyzed 57 grapevine genotypes, consisting of 29 wild grapevines (Vitis vinifera subsp. sylvestris) prospected from the northwest part of Tunisia and 28 cultivated accessions (V. vinifera subsp. vinifera) maintained in the repository of the Arid Land Institute of Medenine (Tunisia). Pair-wise multilocus comparison with the ICVV SNP database allowed the identification of 13 cultivated genotypes, including ten synonymous groups with known Mediterranean or international varieties, three cases of color sports, and two misnomers. Genotypic analysis showed a high level of genetic diversity for both wild and cultivated groups. Multivariate and structure analyses clearly differentiated wild from cultivated grapevines and showed high average posterior probabilities of assignment to their group of origin. The clustering results largely supported the perceived classification and reflect that most of the present Tunisian cultivated varieties do not derive directly from the local wild populations but could correspond to materials introduced from different locations during historical times. Parentage analysis allowed the determination of the genetic origin of four Tunisian cultivars, “Garai”, “Jerbi” (from Kerkennah), “Mahdoui”, and “Reine de Vignes faux”, and showed that “Heptakilo” and “Planta Fina”, two old and widely distributed varieties in the Mediterranean basin, had an important role in the origin of Tunisian grapevines. The present study demonstrates the efficacy of SNP makers for germplasm characterization and genetic studies in grapevine.     

Grapevine (<Emphasis Type="Italic">Vitis vinifera</Emphasis> L.): Old Varieties are Reflected in Works of Art

Grapevine (Vitis vinifera L.): Old Varieties Are Reflected in Works of Art. The northwest of the Iberian Peninsula is home to a number of ancient grapevine varieties now in danger of extinction, regarding which the literature contains only a few references, dating from the 19th century. In this region, baroque religious art, which is commonly ornamented with grapevine motifs, achieved great importance. This work reports the ampelographic comparison of the leaves of 19 old grapevine varieties from this region with those represented on 42 baroque altarpieces. Many of the latter were found to be ampelographically correct representations of grapevine leaves; in some cases they showed such similarity to these old varieties that their cultivation at the time when the corresponding sculptures were made can be confirmed. A larger study may therefore help determine when other varieties were cultivated in the past.     

Phylogeographical structure and conservation genetics of wild grapevine

The distribution of Vitis vinifera subsp. silvestris, the wild grapevine subspecies of Vitis vinifera L., has been dramatically reduced in its major sites of diffusion, at first by the spread, over the last 150 years, of pathogens from North America and, more recently, with fragmentation of habitat and disbranching by humans. In this work, 418 wild grapevine samples, belonging to 78 populations, were collected in their main Mediterranean distribution areas, including the Caucasus area, and the extent of their genetic variability evaluated by analysing plastid microsatellite DNA polymorphism. Results show low haplotype diversity value, with five haplotypes detected within the analysed populations. The highest within-population haplotypic diversity, with the presence of all five detected haplotypes, was found in the Caucasus regions and in the central regions of Italy. The distribution of all detected haplotypes suggests the Caucasian region as the possible center of origin of Vitis vinifera subsp. silvestris. A principal plastid lineage was found to be fixed in several locations, in the Northernmost European countries and in the Southern island of Sardinia. These results draw attention to two different refugium sites in the Mediterranean basin and suggest that conservation priority should be given to grapevine populations still preserved in hotspots of these regions.     

From the cradle of grapevine domestication: molecular overview and description of Georgian grapevine (Vitis vinifera L.) germplasm

Historical information and archaeological and palaeobotanical findings point Georgia, in the South Caucasus, as a cradle for grapevine (Vitis vinifera L.) domestication from its wild form (V. vinifera silvestris Beck.) and subsequent selection and development of varieties with characters suitable for human consumption. The hypothesis of Georgia being a center of domestication, combined with its distance from western countries and the importance of its viticulture and wine production, make Georgian grape germplasm particularly interesting to be investigated under the genetic point of view. Twenty nuclear microsatellite loci were used to genotype 112 Georgian grapevine accessions (V. vinifera sativa Beck.) from germplasm collections and 18 from spontaneous growing plants (V. vinifera silvestris Beck.) found in wild conditions and to compare them to a large international cultivar collection in France. Data analysis shows that Georgian grapevine germplasm has maintained distinctive traits despite arrival of international, foreign varieties and still conserve characteristics of local breeding linked to traditional wine production regions of the country. Results have identified alleles, overall loci, well represented in the Georgian germplasm (cultivated and wild) and absent or poorly represented in other countries, highlighting uniqueness and originality of traits of this viticulture. Moreover, the search for relationships between Georgian and foreign viticulture has evidenced few interesting cases linking the Georgian varieties with Western European ones and with neighboring Caucasian countries, helping to identify the real place of origin in some doubtful cases. In addition, populations or sparse individuals of wild grapevine still preserved in the Georgian natural environments present smaller genetic distances with local cultivars than in other European regions. Principal component analysis (PCA) has also identified special overlapping of the wild compartment with some cultivated varieties. This work provides a highly significant new contribution to applied aspects of Georgian grapevine genetic resources management and use. Uniqueness of the Georgian cultivated grapevine gene pool together with its close relatedness with the wild compartment makes this country a good candidate to address questions regarding domestication and grapevine genetic resource conservation.     

Functional Annotation,Genome Organization and Phylogeny of the Grapevine (<Emphasis Type="Italic">Vitis vinifera</Emphasis>) Terpene Synthase Gene Family Based on Genome Assembly,FLcDNA Cloning,and Enzyme Assays

Background  

Terpenoids are among the most important constituents of grape flavour and wine bouquet, and serve as useful metabolite markers in viticulture and enology. Based on the initial 8-fold sequencing of a nearly homozygous Pinot noir inbred line, 89 putative terpenoid synthase genes (VvTPS) were predicted by in silico analysis of the grapevine (Vitis vinifera) genome assembly [1]. The finding of this very large VvTPS family, combined with the importance of terpenoid metabolism for the organoleptic properties of grapevine berries and finished wines, prompted a detailed examination of this gene family at the genomic level as well as an investigation into VvTPS biochemical functions.     

Genetic diversity of wild grapevine populations in Spain and their genetic relationships with cultivated grapevines

The wild grapevine, Vitis vinifera L. ssp. sylvestris (Gmelin) Hegi, considered as the ancestor of the cultivated grapevine, is native from Eurasia. In Spain, natural populations of V. vinifera ssp. sylvestris can still be found along river banks. In this work, we have performed a wide search of wild grapevine populations in Spain and characterized the amount and distribution of their genetic diversity using 25 nuclear SSR loci. We have also analysed the possible coexistence in the natural habitat of wild grapevines with naturalized grapevine cultivars and rootstocks. In this way, phenotypic and genetic analyses identified 19% of the collected samples as derived from cultivated genotypes, being either naturalized cultivars or hybrid genotypes derived from spontaneous crosses between wild and cultivated grapevines. The genetic diversity of wild grapevine populations was similar than that observed in the cultivated group. The molecular analysis showed that cultivated germplasm and wild germplasm are genetically divergent with low level of introgression. Using a model‐based approach implemented in the software structure , we identified four genetic groups, with two of them fundamentally represented among cultivated genotypes and two among wild accessions. The analyses of genetic relationships between wild and cultivated grapevines could suggest a genetic contribution of wild accessions from Spain to current Western cultivars.     

High incidence of arbuscular mycorrhizal fungi in rare and endangered wild grapevine

High levels of arbuscular mycorrhizal fungi colonisation of 90–100%, as well as high vesicular and arbuscular colonisation were found in the roots of the rare and endangered wild grapevine (Vitis vinifera subsp. sylvestris) along the Neretva River in southwestern Bosnia and Herzegovina, surpassing colonisation in region vineyads.     

Study of genetic relationships between wild and domesticated grapevine distributed from Middle East Regions to European countries

Archaeobotanical-archaeological, cultural and historical data indicate that grapevine domestication can be dated back from 6000 to 7000 years ago and that it took place in the Caucasian and Middle East Regions. However, events leading to the domestication of this crop species are still an open issue. In this paper, 6 chloroplast microsatellites have been used to assess genetic similarities among, and within, domesticated and wild grapevine accessions representative of 7 distinct geographical regions from the Middle-East to Western Europe. Results show that 2 out of the 6 analyzed chloroplast loci are polymorphic within the 193 domesticated individuals and the 387 samples of 69 wild populations. Allele variants of the Cp-SSR loci combine in a total of 6 different haplotypes. The data show that the haplotype distribution is not homogeneous: the 6 haplotypes are present in the domesticated varieties, but only 5 (haplotype VI is absent) are observed in wild populations. The analysis of haplotype distribution allows discussion of the relationships between the two grape subspecies. The contribution of the wild grape germplasm to the domesticated gene pool still growing in different geographical regions can be, in cases, made evident, suggesting that beside domestication, gene introgression has also played a role in shaping the current varietal landscape of the European viticulture.

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Cryptic diversity of <Emphasis Type="Italic">Plasmopara viticola</Emphasis> (Oomycota,Peronosporaceae) in North America

Plasmopara viticola is the causal agent of grapevine downy mildew and is among the most important diseases in viticulture. It originates from North America, where it coevolved with wild Vitis species. Beginning in the 1870s it turned into a global epidemic that has been causing severe yield losses. It is generally believed that a single species is causing downy mildew on a large variety of economically important cultivars. Here we report, based on one nuclear and two mitochondrial markers, that isolates from vineyards in the United States fall into three highly distinct phylogenetic lineages. One of these contains European strains and affects Vitis vinifera cultivars, while the other two lineages affect also other species of Vitis. The divergence between these lineages is high, and, judging from the genetic variation in other Plasmopara lineages, might reflect distinct species. Due to the potentially significant implications for quarantine regulations and resistance breeding, detailed studies will be necessary to clarify whether these genetically distinct lineages occur outside of North America or are still confined there.     

Vitis vinifera L.: Wild or cultivated? Study of the grape pips found at Petra, Jordan; 150 B.C. – A.D. 40

The identification of carbonised grape pips (Vitis vinifera ssp.) is problematic, and the morphological features generally used to distinguish the wild subspeciesV. vinifera ssp.sylvestris from the cultivated subspeciesV. vinifera ssp.vinifera are not satisfactory. Different biometric studies were carried out on Nabataean and Roman seeds found at Petra, Jordan, dated to 150 B.C. – A.D. 400, and the results were compared to known phytogeographical and climatic data. Depending on the identification method selected, the seeds were attributed either to wild grapevines (based on the ratio of breadth over length, and on discriminant analyses of size variables such as pip length, stalk length, and chalaza position), or to an archaic variety of vine with seeds morphologically close to those of wild grapevines (ratio of stalk length over total pip length). The methods used here were applied to European grape pips; they should be tested on Near Eastern material. Archaeological data did not clarify the situation, and the importance of viticulture, which was prohibited during the Nabataean period, remains difficult to evaluate at Petra.     

Characterization of genome-wide long terminal repeat retrotransposons provide insights into trait evolution of four grapevine species

Grapevine is one of the most economically important crops in the world. Although long terminal repeat (LTR) retrotransposons are thought to have played an important role in plants, its distribution in grapevine is not clear. Here, we identified genome-wide intact LTR retrotransposons in a total of six high-quality grapevine genomes from Vitis vinifera L., Vitis sylvestris C.C. Gmel., Vitis riparia Michx. and Vitis amurensis Rupr. with an average of 2938 per genome. Among them, the Copia superfamily (particularly for Ale) is a major component of the LTR retrotransposon in grapevine. Insertion time and copy number analysis revealed that the expansion of 70% LTR retrotransposons concentrating on approximately 2.5 Ma was able to drive genome size variation. Phylogenetic tree and syntenic analyses showed that most LTR retrotransposons in these genomes formed and evolved after species divergence. Furthermore, the function and expression of genes inserted by LTR retrotransposons in V. vinifera (Pinot noir) and V. riparia were explored. The length and expression of genes related to starch metabolism and quinone synthesis pathway in Pinot noir and environmental adaptation pathway in V. riparia were significantly affected by LTR retrotransposon insertion. The results improve the understanding of LTR retrotransposons in grapevine genomes and provide insights for its potential contribution to grapevine trait evolution.     

Hybridization of cultivated Vitis vinifera with wild V. californica and V. girdiana in California

Hybridization of introduced domesticates and closely related natives is well documented in annual crops. The widespread introduction of the domesticated grapevine, Vitis vinifera, into California where it overlaps with two native congenerics, with which it is interfertile, provides opportunity to investigate hybridization between woody perennials. Although geographically widespread, the introduction over the past two centuries has been limited to a few elite clonal cultivars, providing a unique opportunity to study the effects of hybridization on the native species. The amount of hybridization with V. vinifera and the genetic diversity of wild‐growing Vitis californica and Vitis girdiana were examined using nineteen microsatellite markers. STRUCTURE analysis was used to define hybrid and introgressed individuals and to analyze genetic structure of the native species. FAMOZ software was used to identify which V. vinifera cultivars served as parents of F₁ hybrids. The three species were clearly distinguished by STRUCTURE analysis. Thirty percent of 119 V. californica vines were hybrids. The domesticated parent was identified for 16 F₁ hybrid vines; the original California cultivar, ‘Mission’, was the parent of eight. Backcrosses were also found, showing introgression into subsequent generations. Similar results were obtained for a small sample of V. girdiana. Removing hybrids greatly reduced the genetic variation of the presumed pure species, among which there was essentially no genetic structure. Limited genetic variability indicates the California natives may be threatened by genetic erosion. The discovery of F₁ hybrids of ‘Mission’, a cultivar not grown in the areas for ~100 years, suggests long generation times for wild vines that, often, grow into expansive liana and propagate by layering, all factors that limit recruitment in populations already disjunct by habitat lose. Hermaphroditic flowers and fruit that is more attractive to birds may favor the production of backcross seed and establishment of introgressed individuals.     

Use of in vitro method to evaluate some grapevine varieties for tolerance and susceptibility to sodium bicarbonate-induced chlorosis

Nodal shoot segments of four grapevine genotypes well known for their Fe-chlorosis characteristic [Vitis vinifera × Vitis berlandieri Fercal, resistant; V. berlandieri × Vitis rupestris 1103 P, mid-resistant; Solonis (Vitis riparia × V. rupestris × Vitis candicans) × Othello (Vitis labrusca × V. riparia × V. vinifera) 1613 C, susceptible; V. vinifera L. cv. Perlette, resistant] were cultured in vitro. The effects of three levels of iron sodium ethylene-diaminotetraacetate (FeNaEDTA; 9, 18, 36 mg l⁻¹) and three mixtures of iron and 840 mg l⁻¹ NaHCO₃ (sodium bicarbonate) in the Murashige and Skoog (MS) medium supplemented 4.9 μM indole-3-butyric acid (IBA) were compared. We assayed the chlorosis rating of leaves, total chlorophyll of leaves, dry shoot weights of the plantlets, and active and total Fe content of the leaves. The most suitable concentration in determining the reaction of genotypes to iron chlorosis was 9 mg l⁻¹ FeNaEDTA. Bicarbonate addition had negative effects on the iron intake and development of plants. While all genotypes were affected by non-ferrous conditions, Fercal and Perlette were found to be the most resistant genotypes and 1613 C rootstock as the most susceptible. The chlorosis rating of the tested genotypes ranked according to their known degree of tolerance and susceptibility to lime-induced chlorosis. The results of this study showed that the in vitro technique could successfully be used in viticulture to get results in shorter times in the studies, aiming at breeding new rootstocks and varieties suitable to calcareous soil conditions and determining the reactions of existing genotypes to Fe chlorosis.     

Molecular characterization and evolutionary pattern of the 9- cis -epoxycarotenoid dioxygenase NCED1 gene in grapevine

This study provides the first analysis of the level and patterns of nucleotide polymorphism of the NCED1 gene in grapevine (Vitis vinifera L.). A total of 123 sequences of the gene were analyzed to give a sample of 50 wild accessions and 73 cultivars. A high single nucleotide polymorphism and haplotype diversity was revealed in the cultivars studied, especially Tunisian germplasms which present an important and diverse reservoir of genetic diversity for grape breeding and conservation. The haplotype distribution highlights two origins of the cultivars studied: one may be related to primary grapevine gene pool domestication while the second seems to be more recent. Thus, besides domestication, gene introgression has also played a role in shaping the current varietal landscape of grape cultivars. Higher nucleotide and haplotype polymorphism was recorded for cultivars. This was accompanied by a higher recombination rate in cultivated grapevines for this gene, a recent selective sweep in wild samples and a balancing selection in cultivars. The conservation of genetic diversity of the endangered wild germplasm is important to ensure that the wild population can be used in future breeding programs of the domesticated cultivars. The high number of alleles discovered can be used as a valuable source for association studies between allele frequencies and phenotypic variations in this gene. In addition to natural selection, molecular evidence shows that genetic variation in this locus appears to be shaped by a combination of mutation and recombination events.     

Cutting the Gordian Knot of abiotic stress in grapevine: From the test tube to climate change adaptation

In Mediterranean climate areas, the available scenarios for climate change suggest an increase in the frequency of heat waves and severe drought in summer. Grapevine (Vitis vinifera L.) is a traditional Mediterranean species and is the most valuable fruit crop in the world. Currently, viticulture must adjust to impending climate changes that are already pushing vine‐growers toward the use of irrigation, with the concomitant losses in wine quality, and researchers to study tolerance to stress in existing genotypes. The viticulture and winemaking worlds are in demand to understand the physiological potential of the available genotypes to respond to climate changes. In this review, we will focus on the cross‐talk between common abiotic stresses that currently affect grapevine productivity and that are prone to affect it deeper in the future. We will discuss results obtained under three experimental stress conditions and that call for specific responses: (1) acclimatization of in vitro plantlets, (2) stress combinations in controlled conditions for research purposes, (3) extreme events in the field that, driven by climate changes, are pushing Mediterranean species to the limit. The different levels of tolerance to stress put in evidence by the plasticity of phenotypic and genotypic response mechanisms, will be addressed. This information is relevant to understand varietal adaptation to impending climate changes and to assist vine growers in choosing genotypes and viticulture practices.     

Dimorphism in inflorescence scent of dioecious wild grapevine

Wild grapevine (Vitis vinifera subsp. sylvestris) is the dioecious ancestral form of grapevine, from which the domesticated cultivars have derived (V. vinifera subsp. vinifera). Little is known about the floral scent compounds of wild grapevine that is considered as being partly insect pollinated. The knowledge of volatiles released by male and female inflorescence may contribute to the understanding of the pollination biology of this endangered taxon. Inflorescence scents of male and female individuals were collected by dynamic headspace and analysed by thermal desorption-GC/MS. A total of 17 compounds of C5-branched chain alcohols, aliphatics, aromatics, and terpenoids were identified with benzyl alcohol being most abundant in both sexes. Eight of the compounds were sex-specific and differences in 1,2-dimethoxybenzene were most obvious. This aromatic compound was a main constituent in the scent of females (30%), but it did not occur in males. Some of the main compounds of the scent samples are known to be detected by beetles (Cerambycidae) or attract sweat bees (Halictidae) and honey bees (A. mellifera), all well-known inflorescence visitors in wild grapevine. The data presented here are an important step in understanding the chemical communication between wild grapevine and its inflorescence visitors/potential pollinators.     

Chloroplast Genome Diversity in Portuguese Grapevine (Vitis vinifera L.) Cultivars

Grapevine chloroplast (cp) DNA diversity was analysed for the first time through amplification and digestion of fragments of the large single copy (LSC) region by polymerase chain reaction–restriction fragment length polymorphism methodology and also by amplification of three microsatellite loci, previously described as polymorphic in grapevine. Thirty-eight grapevine cultivars collected mainly in the North of Portugal, including some neglected cultivars, four international cultivars (Chasselas, Muscat of Alexandria, Muscat of Hamburg and Pinot) and Vitis riparia and Vitis rupestris, were used in this study with the main goal of finding out their cpDNA diversity and compare the obtained results with previously published data on cultivars from other regions to ascertain their possible origin. Two different alleles were found in each of the three cpSSR loci. Allele variants of the three loci combined in a total of three different haplotypes (A, B and D). The most frequent haplotype, A, was previously reported as the most frequent in Iberian Peninsula and Occidental Europe. Haplotype B was unique to Rabigato, Muscat of Alexandria, V. riparia and V. rupestris. This haplotype was previously proposed to be an ancestral haplotype. Twenty-seven fragments of the LSC region of Vitis vinifera cpDNA were amplified and then digested with HinfI and TaqI restriction enzymes. Polymorphisms were found in the trnT-psbC (TC) and orf184-petA (OA) fragments. In the TC fragment, the polymorphism corresponds to a point mutation in a restriction site of TaqI and is only present in all cultivars with cpSSR haplotype D. In the OA fragment, a short deletion exclusive to the Rabigato cultivar was found. In this case, one sequence tagged site-based marker was developed and will be very useful in future phylogenetic and fingerprinting studies in a broader number of cultivars and in wild grapevine populations. Inference about the progenitors of the Touriga Franca cultivar is done. The present work supports and completes its origin as a descendent of the female and male parents, Marufo and Touriga Nacional.     

Genetic Analysis of East Asian Grape Cultivars Suggests Hybridization with Wild Vitis

Koshu is a grape cultivar native to Japan and is one of the country’s most important cultivars for wine making. Koshu and other oriental grape cultivars are widely believed to belong to the European domesticated grape species Vitis vinifera. To verify the domesticated origin of Koshu and four other cultivars widely grown in China and Japan, we genotyped 48 ancestry informative single nucleotide polymorphisms (SNPs) and estimated wild and domesticated ancestry proportions. Our principal components analysis (PCA) based ancestry estimation revealed that Koshu is 70% V. vinifera, and that the remaining 30% of its ancestry is most likely derived from wild East Asian Vitis species. Partial sequencing of chloroplast DNA suggests that Koshu’s maternal line is derived from the Chinese wild species V. davidii or a closely related species. Our results suggest that many traditional East Asian grape cultivars such as Koshu were generated from hybridization events with wild grape species.