Grapevine Accessions Research Articles

Ampelographic Diversity Assessment of Local Grape Genotype Grown in India

Grapevine is well adapted to the agro climatic conditions of India, which makes it one of the major element of the Indian horticulture. However, there has been limited focus on assessing grapevine germplasm, and genetic resources have previously remained underutilized despite their potential for adapting to environmental changes. This study aims to evaluate the diversity of local (developed in India) grapevine accessions growing in different regions in India. A total of 20 local varieties were evaluated using 14 fruit traits previously developed by DUS. An important variability was revealed among the local varieties studied based on fruit characteristics. The most discriminant traits were bunch weight, berry diameter, bunch length, berry weight, berry shape, seed number per berry. Hierarchical clustering analysis showed three main clusters, each regrouping accessions of different named varieties. Mean values of cluster for various yield parameters revealed that cluster I possessed maximum value for yield related attributes. Maximum bunch weight was observed in Manjari Medika (270.0g) followed by Pusa Seedless (243.1 g), Arka Shyam (213.5 g) while higher 50 berry weight (201.0 g) and berry diameter (20.30 mm) in Arka Majestic genotype. Arka Majestic and Manjari Medika showed better quality traits. Our results indicate a potential of morphological diversity within the local genotypes that should be further investigated in order to understand their performance and to evaluate them in selection programs.

Seed morphometrics unravels the evolutionary history of grapevine in France.

The cultivation of grapevines has spanned millennia, leading to thousands of varieties through exchanges, mutations, and crosses between genotypes, as well probably as gene flow from wild populations. These varieties are typically categorized by regional origin and primary use, either for wine production or fruit consumption. France, within the Western European group, hosts many of the world's renowned wine grape varieties. However, the historical development of cultivated grapevines in France and in the world remains poorly understood. This study applies morphometry on 19,377 charred and waterlogged archaeological grape pips to investigate the evolutionary history of grapevine in France over the last 10,000 years. The study compares seed outlines and lengths, corrected for taphonomic distortions, with a reference collection of 80 wild and 466 modern domestic grapevine accessions. Findings reveal a shift from wild grapevine exploitation to the expansion of domestic varieties around 600-500 BCE, coinciding with Mediterranean cultural influences and the introduction of eastern grape types. The identification of the East-Table group, a group of varieties of eastern origin for fruit consumption, indicates that grapes were also grown for food, especially in Mediterranean regions and near urban areas, alongside wine production. Early French viticulture featured a notable presence of Western European wine-type grapevines. The abundance of pips with wild-like morphology suggests early cultivation involved plants at an initial domestication stage and gene flow between introduced and wild grapevines. As viticulture spread northward, wild and Eastern morphotypes declined, leading to the dominance of Western European wine types in inner France during the Middle Ages.

Integrative genomics reveals the polygenic basis of seedlessness in grapevine

Seedlessness is a crucial quality trait in table grape (Vitis vinifera L.) breeding. However, the development of seeds involved intricate regulations, and the polygenic basis of seed abortion remains unclear. Here, we combine comparative genomics, population genetics, quantitative genetics, and integrative genomics to unravel the evolution and polygenic basis of seedlessness in grapes. We generated the haplotype-resolved genomes for two seedless grape cultivars, "Thompson Seedless" (TS, syn. "Sultania") and "Black Monukka" (BM). Comparative genomics identified a ∼4.25 Mb hemizygous inversion on Chr10 specific in seedless cultivars, with seedless-associated genes VvTT16 and VvSUS2 located at breakpoints. Population genomic analyses of 548 grapevine accessions revealed two distinct clusters of seedless cultivars, and the identity-by-descent (IBD) results indicated that the origin of the seedlessness trait could be traced back to "Sultania." Introgression, rather than convergent selection, shaped the evolutionary history of seedlessness in grape improvement. Genome-wide association study (GWAS) analysis identified 110 quantitative trait loci (QTLs) associated with 634 candidate genes, including previously unidentified candidate genes, such as three 11S GLOBULIN SEED STORAGE PROTEIN and two CYTOCHROME P450 genes, and well-known genes like VviAGL11. Integrative genomic analyses resulted in 339 core candidate genes categorized into 13 functional categories related to seed development. Machine learning-based genomic selection achieved a remarkable prediction accuracy of 97% for seedlessness in grapevines. Our findings highlight the polygenic nature of seedlessness and provide candidate genes for molecular genetics and an effective prediction for seedlessness in grape genomic breeding.

Monitoring and Genotyping of Wild Grapevine (Vitis vinifera L. subsp. sylvestris) in Slovenia.

Vitis vinifera L. subsp. sylvestris (sylvestris) is the only native wild grapevine in Eurasia (Europe and western Asia) and is the existing ancestor of the grapevine varieties (for wine and table grape production) belonging to the subsp. sativa. In Slovenia, the prevailing opinion has been that there are no Slovenian sylvestris habitats. This study describes sylvestris in Slovenia for the first time and aims to present an overview of the locations of the wild grapevine in the country. In this project, a sample set of 89 accessions were examined using 24 SSR and 2 SSR markers plus APT3 markers to determine flower sex. The accessions were found in forests on the left bank of the Sava River in Slovenia, on the border between alluvial soils and limestone and dolomite soils, five different sites, some of which are described for the first time. The proportion of female to male accessions differed between sites. At two sites, female plants dominated; at others, the ratio was balanced. The plants' genetic diversity and structure were compared with autochthonous and unique varieties of subsp. sativa from old vineyards in Slovenia and with rootstocks escaped from nature from abandoned vineyards. Sylvestris was clearly distinguishable from vinifera and the rootstocks. Based on genetic analyses, it was confirmed that Slovenian sylvestris is closest to the Balkan and German sylvestris groups. Meanwhile, a safety duplication of the wild grapevine accessions has been established at the University Centre of Viticulture and Enology Meranovo, Faculty of Agriculture and Life Sciences at the University of Maribor.

Monitoring and Genetic Characterization of Historical Grapevine Varieties (V. vinifera ssp.) from Styria in Slovenia

The aim of this research work was to find historical varieties that existed in this area before phylloxera and to identify them on the basis of historical written sources and genetic analyses. At the beginning of the 19th century, around 300 grape varieties were cultivated in Styria. Between 2020 and 2022, old vineyards were monitored at 115 locations in Styria (between the Mura and Sava rivers) in Slovenia. The directly collected samples (340 grapevine accessions) were determined by molecular analysis with 24 SSR markers. A total of 66 different genotypes were detected. After comparison with the available databases, 29 historical varieties and 37 unknown historical genotypes were identified. Several parameters were calculated to evaluate the usefulness of the selected loci in this work, and a dendrogram representing the genetic similarities between the origins was created using the neighbor-joining method to investigate possible ancestry relationships in the sample set. The most common historical varieties were ‘Belina’ (‘Heunisch weiss’), ‘Vrbovec’ (‘Tantovina Eihenblaetrige’), ‘Ranfol’ (‘Ranfol beli’), and ‘Pelesovna’ (‘Vulpea’). Varieties from the current variety list were also frequently found, such as ‘Frankinja’ (‘Blaufraenkisch’) and ‘Žametovka’ (‘Kavčina črna’). In a few locations, one of the most important red varieties from the beginning of the 20th century was also found in this area (alongside ‘Frankinja’ and ‘Žametovka’), i.e., ‘Vranek’ (‘Zimmettraube balu’). At that time, this variety was planted in multi-variety vineyards and was preserved, but its importance in single-variety vineyards quickly declined due to female flower. In addition, genetic analyses have shown that 37 unknown historical genotypes have been found in this area. These genotypes need to be described ampelographically and technologically evaluated in the future. Most of the vegetative offsprings of these genotypes have already been transferred to the Meranova gene bank, where they can be accurately described ampelographically under the same pedoclimatic conditions.

In Vitro Collection for the Safe Storage of Grapevine Hybrids and Identification of the Presence of Plasmopara viticola Resistance Genes.

This paper focuses on the creation of an in vitro collection of grapevine hybrids from the breeding program of the Kazakh Scientific Research Institute of Fruit Growing and Viticulture and investigates the presence of Plasmopara viticola resistance mediated by Rpv3 and Rpv12 loci. We looked at the optimization of in vitro establishment using either shoots taken directly from field-grown plants or from budwood cuttings forced indoors. We further screened for the presence of endophyte contamination in the initiated explants and optimized the multiplication stage. Finally, the presence of the resistance loci against P. viticola was studied. The shoots initiated from the field-sourced explants were the more effective method of providing plant sources for in vitro initiation once all plant accessions met the goal of in vitro establishment. The concentration of phytohormones and the acidity of the culture medium have a great effect on the multiplication rate and the quality of in vitro stock cultures. Out of 17 grapevine accessions, 16 showed the presence of single or combined resistance loci against P. viticola. The grapevine accessions identified as carrying Rpv3 and Rpv12 alleles represent important genetic resources for disease resistance breeding programs. These accessions may further contribute to the creation of new elite cultivars of economic interest.

Integration of machine learning models with microsatellite markers: New avenue in world grapevine germplasm characterization

Development of efficient analytical techniques is required for effective interpretation of biological data to take novel hypotheses and finding the critical predictive patterns. Machine Learning algorithms provide a novel opportunity for development of low-cost and practical solutions in biology. In this study, we proposed a new integrated analytical approach using supervised machine learning algorithms and microsatellites data of worldwide vitis populations. A total of 1378 wild (V. vinifera spp. sylvestris) and cultivated (V. vinifera spp. sativa) accessions of grapevine were investigated using 20 microsatellite markers. Data cleaning, feature selection, and supervised machine learning classification models vis, Naive Bayes, Support Vector Machine (SVM) and Tree Induction methods were implied to find most indicative and diagnostic alleles to represent wild/cultivated and originated geography of each population. Our combined approaches showed microsatellite markers with the highest differentiating capacity and proved efficiency for our pipeline of classification and prediction of vitis accessions. Moreover, our study proposed the best combination of markers for better distinguishing of populations, which can be exploited in future germplasm conservation and breeding programs.

Allelic diversity of the SSR locus VVIB23 linked to the flower sex in wild-growing grapevine genotypes of Krasnodar Territory

Background. Studying wild-growing grapevines is of interest because it expands the knowledge on the genetic diversity within the gene pool and helps to search for new resistance sources. One of the main differences between wild vines and cultivars is the type of their reproductive system: wild grapes are represented by cross-pollinated dioecious vines, while cultivated ones are, with rare exceptions, hermaphrodites capable of self-pollination. A DNA marker applicable to determine the grape flower sex has been identified.Materials and methods. The VVIB23 DNA marker to the Sex gene determining the flower sex was used to study 36 genotypes of wild-growing grapes selected in 2019–2021 within the Utrish Nature Reserve near Gelendzhik and the Krasny Les State Nature Reserve on the right bank of the Kuban River. PCR technique was applied, and the results were evaluated on a Nanofor 05 genetic analyzer.Results. The studied grapevine accessions were found to be variable for the VVIB23 SSR locus: 7 types of alleles were identified. Alleles corresponding to the male flower type were present in 14 genotypes, and those corresponding to the female type in 21. A PCR fragment whose size, according to the published data, correlated with hermaphroditism was not found, which is typical for wild genotypes. The DNA marker analysis failed to determine the flower sex of one accession.Conclusion. The results of the DNA marker analysis showed that the studied wild-growing grapevine genotypes were dioecious forms

Expression of the genes (CBF2, CBF4, NCED1, MYB14, PLD) during acclimation and de-acclimation in dormant buds of Chambourcin and 3309 C grapevine accessions

Expression of the genes (CBF2, CBF4, NCED1, MYB14, PLD) during acclimation and de-acclimation in dormant buds of Chambourcin and 3309 C grapevine accessions

Incidence and genetic diversity of raspberry bushy dwarf virus (RBDV) in Rubus spp. in Turkey

AbstractRaspberry bushy dwarf virus (RBDV), recently renamed to Idaeovirus rubi, is one of the most common viruses infecting Rubus species worldwide but there is still a limited number of genome sequences available in the GenBank database and the majority of the sequences include partial sequences of RNA‐1 and RNA‐2. The distribution and incidence of RBDV in main raspberry and blackberry growing provinces in Turkey were monitored during 2015–2019 and 537 Rubus spp. samples were tested by both DAS‐ELISA and RT‐PCR. Among the tested samples, 36 samples tested positive for RBDV by DAS‐ELISA and 67 samples by RT‐PCR. There was relatively low nucleotide diversity among the Turkish isolates. Turkish isolates shared 93%–97.7%, 84.3%–98.9%, and 85%–99.2% nucleotide sequence identities with available sequences in the GenBank, in partial RNA‐1, movement protein (MP) and coat protein (CP) genes, respectively. In the phylogenetic tree constructed for RNA‐1, MP, and CP sequences, all Turkish raspberry isolates were clustered in a distinct clade. However, the blackberry isolates showed considerable variation in nucleotide sequences and were placed in three distinct groups. The divergent blackberry isolates showed high variability in MP (84.5%–89.3%) and CP (85.5%–89.7%) regions and were placed in a distinct group. The rest of blackberry isolates clustered together with sweet cherry RBDV isolates adjacent to the grapevine clade or together with raspberry isolates. The comparative analysis conducted on three RNA segments of RBDV highlighted the high sequence diversity of Turkish RBDV isolates. This study also emphasizes the importance of regular monitoring of RBDV infections in Turkey, with special regard to those Rubus spp. and grapevine accessions employed in conservation and selection programmes. In particular, the presence of new RBDV genetic variants and infection of Rubus species must be taken into account to choose a correct detection protocol and management strategy.

Identification of Vitis vinifera L. Local Cultivars Recovered in Andalusia (Spain) by Using Microsatellite Markers

In Andalusia (Spain), there are different wine regions that have a great recognized tradition. In these regions, the cultivation of the vine is ancient and there are still vineyards planted with local varieties of Vitis vinifera L. that have not yet been identified. The aim of this research study was to identify 49 accessions of grapevine collected in the districts of four provinces in Andalusia (Spain). All samples were genotyped with 20 microsatellite markers in order to ascertain the identity and analyze the genetic diversity of the collected material. In total, 30 different genotypes were obtained, 22 of them which were identified with named, known varieties by comparison to the Spanish or European microsatellite databases, and eight which are referred to as new genotypes. All loci were polymorphic, and a total of 159 alleles were detected, ranging from 4 to 12 alleles per locus, with an average allele number of 7.95. The overall observed heterozygosity was 0.763 and was slightly higher than expected (0.715), while the gene diversity per locus varied between 0.167 (VVIN73) and 0.967 (VVMD5). A dendrogram representing the genetic similarities among cultivars was depicted using the UPGMA method to investigate their relationships. The eight new genotypes identified in this research work could represent ancient local varieties in danger of extinction. These new cultivars may be used to determine original wines.

Potential Implications and Management of Grapevine Viruses in Mexico: A Review

Worldwide, virus infections in grapevines are of concern due to the potential for economic loss. Although the grape industry in Mexico is relatively small and focused mainly on the local market, production dates back to the time of the Spanish colonization. This manuscript discusses the findings on grapevine viruses in Mexico. Nine viruses have been identified in the last fifty years, including grapevine red blotch virus (GRBV), grapevine leafroll-associated virus 3 (GLRaV-3), grapevine fanleaf virus (GFLV), and grapevine virus A (GVA). Important information is provided about these viruses and viral pathogens that have not yet been reported in Mexico, but represent an ongoing threat to plant health and grapevine production in other viticultural regions of the world. Strategies for virus control in vineyards are described. The information discussed here should be shared with growers and stakeholders to prevent future negative impacts on the Mexican grapevine industry and to save ancient grapevine accessions.

Genetic diversity of some autochthonous white grape varieties from Romanian germplasm collections

We investigated the phenotypic and genetic diversity among 25 grapevine accessions of white grape maintained in a germplasm collection ex situ. The selected varieties were characterised using standardised ampelographic descriptors and microsatellite markers to obtaintheir complete description. The ampelographic analysis based on 24 OIV descriptors and the obtained first dendrogram based on common features, revealed the extent of the phenotypic diversity ranging from 0.29 (between Coarnă albă and Chardonnay) to 0.92 (between Crâmpoşie and Cioinic), without any connection with their supposed origin or the current geographic area of distribution. The SSR allelic analysis with nine standard markers proved and confirmed its high discrimination potential for genotype identification and diversity. The second cluster analysis aimed to evaluate the genetic relationships among analysed accessions,based on Dice’s similarity coefficient, revealed two groups of varieties: the first group (A) included twelve accessions with a degree of similarity between 0.37 and 0.5. The second (B) included eleven accessions, with a similarity degree between 0.33 and 0.67. The genotypic cluster analysis showed a distinct lower similarity between the Majarcă albă variety and all the varieties from the first group, and between Busuioacă de Bohotin and all the analysed varieties. The phenotypic and genetic matrices and dendrograms, although not correlated, were complimentary and provided valuable information about the diversity of the grapevine genotypes and the identification of the synonyms.

Real-time RT-PCR high-resolution melting curve analysis to detect and differentiate Brazilian variants of grapevine viruses

Detecting and identifying viral infections in perennial plants, such as grapevines, can be challenging. Therefore, the aim of this study was to perform a real-time RT-PCR (RT-qPCR) high-resolution melting (HRM) curve analysis to detect and differentiate Brazilian variants of grapevine leafroll-associated virus 3 (GLRaV-3) and grapevine fanleaf virus (GFLV) in 74 and 10 infected plants, respectively, maintained in a collection block of grapevines. A single amplification curve was generated for each sample by RT-qPCR. Considering the amplified region of genomes of these two viruses, it was possible to identify and distinguish different variants of GLRaV-3 and of GFLV, which showed significantly different melting temperature (Tm) values between themselves, reflecting differences in the nucleotide sequences of the respective amplicons, and allowing discriminating variants and assess the viral diversity in grapevine accessions. The HRM analysis was validated by sequencing and nucleotide comparisons among Brazilian isolates of GLRaV-3 and GFLV.

Genetic and ampelographic characterization of grapevine accessions maintained in the Lebanese national collection

Safeguarding grapevine biodiversity is one of the main concerns in viticulture today. Management of ex situ collections requires a comprehensive characterization of the conserved germplasm to provide basic material for selection and breeding programs. In this study, the local grapevine germplasm conserved at the national collection of the Lebanese Agricultural Research Institute and composed of 43 accessions was submitted to a genetic and ampelographic study. Nine ISSR primers, previously developed in grapevine, were used in this study. These primers generated a total of 51 bands, of which 77.7% were polymorphic allowing the differentiation of 41 genetic profiles vs. one case of synonymy that was recorded among three accessions. Ampelographic characterization was conducted using a set of 33 descriptors established by the International Office of Vine and Wine and related to leaf, bunch, and berry. Principal component analysis identified 12 descriptors i.e. veraison date, maturity date, berry length, upper and lower vein pigmentation, bunch density, bunch weight, sugar content at harvesting, flesh of juiciness, berry weight, flesh firmness, and color skin, as being the most discriminating descriptors. The correlation between the ISSR clustering and the ampelographic one was not significant (r=0.26) because of the divergence of accessions groups, except for the three accessions synonymy case which was confirmed in both dendrograms. Finally, this comprehensive evaluation of the existing local gene pool of grapevine revealed a substantial diversity. It would further allow the promotion of the valuable accessions directly through multiplication schemes, and their sustainable utilization in genetic improvement programs.

Virus and Virus-like Pathogens in the Grapevine Virus Collection of Croatian Autochthonous Grapevine Cultivars.

Grapevine collections play an important role, especially in the study of viruses and virus-like pathogens. In 2009, after an initial ELISA screening for eight viruses (arabis mosaic virus, grapevine fanleaf virus, grapevine fleck virus, grapevine leafroll-associated viruses 1, 2, and 3, and grapevine viruses A and B), a collection of 368 grapevine accessions representing 14 different Croatian autochthonous cultivars and containing single or mixed infection of viruses was established to further characterize the viral pathogens. Subsequently, Western blot, RT-PCR, cloning, and sequencing revealed that grapevine rupestris stem pitting-associated virus was frequently found in accessions of the collection, with isolates showing substantial genetic diversity in the helicase and coat protein regions. High-throughput sequencing of 22 grapevine accessions provides additional insight into the viruses and viroids present in the collection and confirms the fact that Croatian autochthonous grapevine cultivars have high infection rates and high virome diversity. The recent spread of “flavescence dorée” phytoplasma in Europe has not spared the collection. After the first symptoms observed in 2020 and 2021, the presence of phytoplasma was confirmed by LAMP in six grapevine accessions and some of them were lost. Single or multiple viruses and viroids, as well as own rooted grapevines in the collection, make the plants susceptible to various abiotic factors, which, together with the recent occurrence of “flavescence dorée”, makes the maintenance of the collection a challenge. Future efforts will be directed towards renewing the collection, as 56% of the original collection has been lost in the last 13 years.

First Report of Grapevine Kizil Sapak Virus in Grapevine in China.

Grapevine Kizil Sapak virus (GKSV) is a novel member of the family Betaflexiviridae classified into the proposed genus Fivivirus within the subfamily Trivirinae. It was first discovered in USA from a grapevine originating from Turkmenistan (Al Rwahnih et al. 2019) and later in France from a grapevine accession from Iran (Marais et al. 2020). In October 2019, an asymptomatic grapevine cv. 'Crimson Seedless' (native to USA) was collected from Xinjiang province in China and analyzed by high-throughput sequencing (HTS). Ribosome-depleted RNA preparations were used for library synthesis followed by HTS on an Illumina HiSeq X-ten platform. A total of 29,141,024 cleaned reads were obtained, and 7,878 contigs were generated using CLC Genomics Workbench 9.5 (QIAGEN). One long contig (7,328 bp) showed 88.2% nucleotide (nt) identity with the sequence of GKSV-127 (MN172165) via Blastx, with an average coverage of 284-X. Bioinformatic analysis of the remaining contigs showed the presence of Grapevine leafroll-associated virus 4, Grapevine rupestris vein feathering virus, Grapevine fabavirus, grapevine yellow speckle viroid-1 (GYSVd-1), GYSVd-2 and Hop stunt viroid in the sample. The presence of GKSV was checked by RT-PCR using the primer GKSV-F/R (Al Rwahnih et al. 2019); the 1,240 bp PCR product was cloned using a pTOPO-T vector (Aidlab, China) and sequenced. In pairwise comparison, the obtained nt sequences shared 92.6 to 95.2% identity to the corresponding HTS sequence, confirming the presence of GKSV in the sample. The complete GKSV genome sequence was obtained as two pieces of overlapping DNA sequence using primers GKSV-20A/20B (5'-TAGTCTGGATTTCCCTACCT/5'-CTCCCTAAACTGATTTGATG) and GKSV-25A/25B (5'-GCCACTGGTGAATGAAAAGA/5'-CTAAATGAATGGGCAGGTAT) designed based on the HTS-generated sequence. The 5' and 3' termini were determined by rapid amplification of cDNA ends using SMARTer RACE 5'/3' Kit (Takara, Dalian, China). The complete genome of GKSV isolate CS (MW582898) comprised 7,604 nt (without the polyA tail) and shared 77.8 to 89.2% identities with the other nine reported GKSV isolates, among which it shared the highest nt identity (89.2%) with GKSV-127. In phylogenetic analysis based on complete or nearly complete genome sequences of representative members of Betaflexiviridae, GKSV-CS clustered with the nine known GKSV isolates, forming a subclade with GKSV-127 (Supplementary Fig. 1). To determine the incidence and distribution of GKSV in China, 476 grapevine samples of 75 cultivars were collected from 20 provinces and tested by RT-PCR using primers GKSV-F/R (Al Rwahnih et al. 2019) and Vini-F1/R1 (Marais et al. 2020). The results showed that 0.42% (2 of 476) of the samples tested positive with both primers, including samples GKSV-CS and a 'Black Monukka' grape (native to India) also sampled from Xinjiang. Both PCR products of 'Black Monukka' were cloned and sequenced (MZ311588 to MZ311602) and they showed 85.1 to 88.9% nt identities to the GKSV-CS sequence. This is the first report of GKSV infecting grapevine in China. Although the pathogenicity of GKSV is yet to be determined, it has been found in several countries such as USA (Al Rwahnih et al. 2019), France (Marais et al. 2020) and China (this study). Both positive samples in this study were collected from Nanjiang region in Xinjiang province, indicating the sporadic occurrence of GKSV in this area.

Screening for adaptation to resistant grapevine accessions in Plasmopara viticola population of north-eastern Italy

Screening for adaptation to resistant grapevine accessions in <i>Plasmopara viticola</i> population of north-eastern Italy

First report of grapevine rupestris vein feathering virus in Vitis vinifera L. from Japan.

The production of wine grapes is gaining widespread popularity and being carried out on approximately 2,200 hectares of land in Japan. Scions grafted onto rootstocks generally have been imported from the EU, USA, New Zealand, and Australia into Japan. Unfortunately, viruses have spread in Japanese vineyards by slipping through the net of plant quarantine. Grapevine rupestris vein feathering virus (GRVFV), which was detected in a Greek grapevine accessions, is a member of genus Marafivirus in family Tymoviridae (El Beaino et al. 2001). GRVFV has been detected in many countries such as USA, Canada, Australia, New Zealand, Italy, Spain, Switzerland, Czech Republic, Uruguay, and Pakistan (Jo et al. 2015; Eichmeier et al. 2016; Xiao and Meng 2016; Blouin and MacDiarmid 2017; Reynard et al. 2017; Cho et al. 2018; Mahmood et al. 2019; Wu et al. 2020). Herein we report GRVFV infection in Vitis vinifera L. grapevines from Japan. In February 2021, dormant canes from 18 V. vinifera cv. Cabernet Sauvignon with leafroll-like disease symptoms, growing in a vineyard located in Kanagawa Prefecture, were collected. No typical vein banding symptom by GRVFV were observed in the grapevines during the growing season. Total RNA was isolated from the canes using an RNeasy Plant Mini Kit and QIAshredder (Qiagen, Valencia, CA), and subjected to cDNA synthesis using a PrimeScript 1st Strand cDNA Synthesis Kit (Takara Bio, Shiga, Japan). RT-PCR was performed with GRVFV_6156F and GRVFV_6600R primers for GRVFV detection (Reynard et al. 2017). The expected 445 nucleotides (nt) amplification product was obtained from four of 18 grapevines. Sequence analysis of the products revealed 91% identities to corresponding sequences of GRVFV isolates CHASS (KY513702) and Mauzac (KY513701) from Switzerland. Genome walking to determine the whole-genome sequence of the GRVFV isolates from the four grapevines was performed. Briefly, the upstream and downstream of the 445 nt amplification product were amplified from first-strand cDNA using gene-specific primers designed from the product and CHASS-specific primers. Each amplified fragment was Sanger sequenced. Next, gene-specific primers were designed to obtain the complete genome of GRVFV as 13 overlapping DNA fragments from each of the four grapevine samples. An identical complete genome of 6,704 bp was assembled from the overlapping DNA fragments using MEGA 10 software and named as NA1 isolate (DDBJ accession no. LC619667). Phylogenetic analysis of the NA1 genome and corresponding sequences of GRVFV from other countries showed that NA1 formed a cluster with isolate NZ ChTK0004 from New Zealand (MF000326; Supplementary Figure 1). In pairwise comparisons, the complete NA1 genome was most identical at 88% and 87%, respectively to isolates NZ ChTK0004 and Mauzac. The predicted amino acid sequences of NA1 polyprotein shared high homologies (96%) to the corresponding polyprotein sequences of NZ ChTK0004 and Mauzac, suggesting that NA1 is genetically similar to GRVFV isolates from New Zealand and Switzerland. The NA1-infected Cabernet Sauvignon was co-infected with Grapevine leafroll-associated virus 3, Grapevine virus A, and Grapevine rupestris stem pitting-associated virus according to RT-PCR assay for grapevine virus detection (Nakaune and Nakano 2006). The results underscore the importance of intensifying quarantine measures to prevent introduction of exotic viruses via contaminated wine grape vegetative cuttings.

Molecular and ampelographic characterizationof Pisco grapevine accessions conserved in a germplasm collection center of the Ica, Peru

Pisco is a distillate of cultural and economic importance for Peru, made from heterogeneous vine varieties called "pisqueras", covered by a Pisco Denomination of Origin legislation. The aim was to confirm the varietal identity of eight accessions conserved in the Germplasm Collection of the Centre for Productive Innovation and Technology Transfer Agroindustrial Ica (CITEagroindustrial), using 9 standardized microsatellite loci (SSR) and 14 morphological descriptors according to the International Organisation of Vine and Wine (OIV) to identify grapevine varieties. The SSR molecular profiles of 7 accessions were identified as varieties registered in the Vitis International Variety Catalog (VIVC) and only one had no association. Of these, Mollar, Albilla, Italia, Quebranta and Uvina accessions, were associated with Mollar Cano (VIVC 7901), Palomino Fino (VIVC 8888), Muscat of Alexandria (VIVC 8241), Quebranta (VIVC 9840) and Jacquez (VIVC 5627), respectively. The SSR profile of the Torontel accession was not associated with any variety registered in the VIVC and the Negra Criolla and Moscatel accessions were associated with the Spanish variety Listan Prieto (VIVC 6860). According to the ampelographic observations, it was determined that Moscatel presented phenotypic variations with respect to Negra Criolla, although at the molecular level it was perfectly associated with Listan Prieto, indicating that it would be a clonal variation. In addition, 7 variables were discriminant to identify the accessions evaluated. The molecular and ampelographic analysis allowed to clarify the varietal identity of the accessions conserved at CITEagroindustrial, Ica, which plays a preponderant role as a reference regarding the identification of the varieties used for the elaboration of Pisco in Peru.