Annual Project REEport 2019-20

PREFACE

This report serves as the summary of first year project updates for VitisGen2. The following is an excerpt from the annual report submitted to the USDA-NIFA Research, Extension, and Education Project Online Reporting system (REEport) for the project entitled “VitisGen2: Application of Next Generation Technologies to Accelerate Grapevine Cultivar Development”. The report is cumulative from all project team members and covers the period from September 1, 2019 through August 31, 20 and was submitted on November 30, 2020 The report has been reformatted for readability prior to distribution to project directors, collaborators, and the advisory panel.

PROJECT ACCOMPLISHMENTS

Summary

Recent analyses have emphasized the economic and environmental importance of developing new grape cultivars with high fruit quality (FQ) and resistance to powdery mildew (PM). An Advisory Panel of table, juice, raisin and wine grape industry members concurs that high-quality, PM-resistant cultivars are a top priority. The proposed project (VitisGen2) will i) expand on VitisGen progress in developing novel economic, phenotyping and genetics knowledge and tools related to new grape cultivars, and ii) translate these and previous VitisGen innovations into new applications for improving grape breeding programs and managing existing vineyard plantings. The Economics team will evaluate the consequences of introducing new grape traits, including impacts upon cost, yield, revenue, profit, pesticide use, and the environment. The Trait Evaluation team  will develop novel high-throughput methods, and apply these along with proven approaches to characterize a range of phenotypes, such as PM resistance durability and undesirable fruit qualities, as well as locally important traits. The Genetics team will couple phenotyping results with high-resolution genetic maps, which can be combined with genome assembly and RNA-Seq analyses to develop inexpensive, high-resolution markers spanning key genes. Finally, the Outreach team will communicate scientific opportunities and discoveries, and provide stakeholders with knowledge of the benefits of adopting new high-quality PM resistant cultivars along with new tools for characterizing PM and FQ in their existing plantings. The proposed work utilizes plant breeding and genomics approaches to improve grape characteristics and enhance the economic and environmental sustainability of grape production

Major Project Goals

Powdery mildew (PM) resistance consistently ranks as a top research priority facing the U.S. grape industry, based on more than a dozen stakeholder meetings focused on research priorities since 2005 and numerous industry surveys. PM is the most significant grape disease in California – and likely worldwide – in terms of expenses for control and losses in quality and yield. In an international survey of grape scientists, PM was statistically significant in being both the most important and the most genetically tractable of twelve key traits. However, breeding for PM resistant cultivars via traditional methods frequently results in negative fruit quality trait introgression from wild Vitis. Winemakers report that poor sensory attributes and color are major challenges to using interspecific hybrids, particularly the presence of high acidity, low tannin and color stability, and off aromas. To address the need for PM resistant cultivars with desirable fruit quality, the long-term objectives of the project are listed in the next section.

Accomplishments

This year, we completed and validated the development of the core rhAmpSeq set with >2000 amplicons, effective for genetic map construction and marker-assisted-selection (MAS) in Vitis. The flower sex marker reported last year was folded into the core rhAmpSeq panel and is now available for screening seedlings for sex alleles – M, f, or H. We discovered 2 different H (H1 and H2) alleles in the cultivated grapevine gene pool. Over 32,544 samples across 19 programs – public and private, domestic and international, etc., were processed using the new genotyping platform. A low-cost single marker KASP platform was explored to support QTL mapping. Validated markers were used to support MAS in several breeding programs. A user friendly software for MAS quality control and statistical interpretation was developed for breeders to use in selection. Also, the AmpSeq marker platform developed in VitisGen1 was used in the genetic studies and monitoring of PM Erysiphe necator fungus. Reference genome assemblies to promote PM resistance and gene-stacking were pursued. High-throughput and recent statistical approaches were adopted for efficient laboratory and field assessment of traits. A fee-for-service rhAmpSeq genotyping option was opened to the public and private breeders not funded by the project this year. The ‘Blackbird’ imaging robots for PM and DM assessments are now commercially available for purchase. We continued to pursue knowledge extension to other scientists, grape growers, and the public using various channels including but not limited to publications, conferences, and the project website (vitisgen2.org).

Objective 1: Technological Innovations Driven by Trait Economics

  • Integrate genome-wide data with innovations in phenotyping powdery mildew (PM) resistance, and table and wine grape quality for genetic characterization of high priority traits. Specific goals focus on gene discovery and trait assays: de novo annotation of relevant wild genomes; RNASeq of all VitisGen parents; low-cost AmpSeq marker implementation; automated PM quantitation without staining; multiple high throughput phenotyping screens for key fruit quality traits, and identification of candidate genes for PM resistance and fruit quality.
  • Complete and disseminate economic analysis of several key agronomic and quality traits to drive research and breeding.

The new marker platforms were used in MAS to screen >6000 seedlings. Target crosses to generate new populations that combine disease-resistance with high fruit quality wine, table or natural dry-on-vine raisin grape attributes were made across different locations. Overall, traits considered for MAS include PM resistance (Run1, Ren1, Ren2, Ren3/9, Ren4, Ren10), DM resistance (Rpv1, Rpv3, Rpv10, Rpv12, and new loci from amurensis), muscat, di- vs. mono-glucosides, seedlessness, and flower sex. Mapping family-customized convolutional neural network models that improved detection accuracies and downstream QTL analyses were developed for PM (hyphae and conidia) and DM quantification. The development and validation of vineyard-based imaging of diseases continued on mapping populations using the Bloomfield Robotics imaging platform developed during the SCRI Efficient Vineyard project. A low-cost QR code system attached to trellis wires was developed for vine identification. Laboratory ‘Blackbird’ imaging robots were upgraded to the latest mirrorless imaging technology from Nikon to improve image quality and eliminate inherent wear issues. A collaborative robotic arm for leaf disk sample processing and arraying was implemented. Over 1500 grape genotypes were phenotyped for the identification of candidate resistant genes, gene stacking, race specificity testing, etc., of PM and DM. Over 25,000 files (3.46 TB) of microscopy and 223,000 (2.33 TB) vineyard images were generated. A 2D cluster architecture software was trained using images collected between 2016 and 2018 for cluster related analyses. Also, we are adopting spectral measurements for a variety of traits at Cornell. In Minnesota, the use of SPME for field evaluations of floral compounds was explored. Multilocation evaluation of F2 population (180-200 genotypes, 3 reps) continued at J. Lohr vineyard, CA and Brookings, SD. Fruit samples (>820) from breeders were analyzed for quality related attributes. Detailed pre- and post-veraison data were collected to determine the contribution of malate accumulation, degradation, and dilution to final malate concentrations on the Horizon x Illinois 547-1 family at Cornell. The analysis of 5-year malate data on ripe fruit uncovered QTL associated with malate on chromosomes 17 and 7 explaining >40% variation in malate and are stable in >3 years of the study. Genotypes differ significantly in fruit malate concentrations and the differences between haplotypes in the detected loci were up to 6.9 g/L at ripeness. Twenty genotypes bulked into 4 malate segregating groups and will be validated with metabolomic and transcriptomic analyses. Other variations (e.g. potassium), pre-and post veraison were equally observed.

Several QTL studies were carried out on several mapping families for many fruit quality, agronomic, and disease traits including tannin, malate, flowering time, veraison time, berry weight, berry size, cluster weight, leaf trichomes, foliar phylloxera resistance, Japanese beetle feeding, leaf variegation, root architecture, bud break, veraison, wood acclimation, bud freezing resistance, etc., across institutions. Gene expression and abundance as well as ethanol, acetaldehyde, CO2 production, and internal O2 levels during fruit development and ripening on vinifera, riparia, and cinerea genotypes were monitored. Results of the analysis showed that samples are primarily differentiated by phenological stage, and to a lesser extent by species. Also, the onset of ripening is associated with hypoxic conditions in both vinifera and riparia species, and vinifera berries accumulated more pyruvate and lactate, while riparia berries accumulated up to 10-fold ethanol. Wild Vitis spp. generate more CO2 per g fresh tissue than vinifera berries, a sign of a higher rate of energy generating metabolic pathways like respiration.

Objective 2: Knowledge Extension and Application

  • Incorporate technological innovations and economics-oriented priorities in the generation of grapevine seedlings in breeding programs and the selection of elite breeding lines. Publicly release grapevines, pollen, and/or seed lots with various combinations of RUN1, REN1, REN2, REN3, REN4, REN6, REN7 and REN10 PM resistance.
  • Demonstrate the impact of VitisGen advances to grape growers, enologists, and specialty crop researchers.

To promote the release of grapevines with various combinations of PM resistant genes and quality traits, pollen samples with different levels of PM genes were shared among breeding institutions: Run1Ren1Ren4 (USDA-Parlier to Minnesota); Ren3Ren9, and Run1Ren6Ren7 (Cornell to USDA-Parlier), etc. Also, lines with stacked PM resistant genes will be sent to Foundation Plant Services for sharing with other U.S. breeding programs. Besides, pollens with stacked sources for PM and DM resistance were received from Oliver Trapp, Geilweilerhof, Germany, and used for crosses at Cornell. Reference genomes for characterizing the biological processes associated with PM resistant loci REN1, REN2, and REN3 using BSR-seq analysis is ongoing. Single Molecule Real-Time (SMRT) bell libraries were prepared for 4 parents: V. vinifera spp. vinifera Y315-43-04 (REN1+ ), 2 parents of Illinois 547-1 (REN2+ ): V. rupestris B38 and V. cinerea B9 (REN2+ ), and Horizon (REN3+ /REN9+ ). Thirty-six cDNA stranded libraries from the 2 breeding families: V. vinifera spp. vinifera Y315-43-04 (REN1+ ) x V. cinerea C81-227, and Horizon (REN3+ /REN9+ ) x Illinois 547-1 (REN2+ ), were sequenced in single-end 150-bp mode (NextSeq500 sequencer) resulting in ~11 million high-quality reads per sample. Also, the genome assembly of V. piasezkii DVIT2027 (REN6+ /REN7+ ) was further scaffolded using the BioNano optical maps generated last year. The target chromosome-scale assembly will serve as a reference for the characterization of the REN6 and REN7 loci. Eight F1 accessions from the breeding population 11373 (Pap et al., 2016), from V. vinifera spp. vinifera F2-35 (REN6- /REN7- ) x V. piazeskii DVIT2027 (REN6+ /REN7+ ) cross was selected for DNA sequencing: 2 REN6+ /REN7- , 2 REN6- /REN7+ , 2 REN6+ /REN7+, and 2 REN6- /REN7- using SMRT DNA sequencing (Pacific Biosciences or Illumina DNA sequencing (DNA-seq) due to problematic DNA extraction. To better define REN1 locus, 5 V. vinifera spp. vinifera accessions possessing a REN1-like locus (Amrine et al., 2015) were sequenced using PacBio SMRT DNA sequencing. Genome assembly was performed for 4 of them: Husseine, Karadzhandal, Khalchili, and Sochal. Genome assembly of Late Vavilov is ongoing. There are genotyping efforts to ascertain new sources of PM resistance from V. doaniana 588149, V. amurensis 588631, and B37-28.

Opportunities for training and professional development

Many training and professional development opportunities were provided and include the training of nine summer students and technicians in a Computer Vision Academy, organized by the VitisGen2 powdery mildew team, and presented by Cornell University and USDA-ARS, Geneva scientists, technicians, and postdocs. Participants learned grapevine biology, growth and development, pathology, and remote hands-on image annotation for neural network analysis of a diverse array of traits. Their efforts resulted in an impressive database of annotated images for the project and insight into which traits are more amenable to computer vision analysis, and improved methods for vineyard imaging and analysis. Two postdoctoral researchers at UC, Davis, and Cornell were engaged to sustain trait economic evaluations and two Graduate Research Assistants at WSU and UCD were recruited to assist with data assembly and analysis, and general survey development. They were exposed to all aspects of research methods and provided with opportunities to acquire and enhance their research skills. A graduate student and another Post-doctoral Associate (Dr. Noam Reshef) at Cornell working on fruit quality analyses enhanced their knowledge on experimental design related to understanding differences in metabolite accumulation and degradation between wild Vitis and V. vinifera. Dr. Reshef’s work attracted the Vaadia-BARD post-doctoral grant for performing research on the regulation of malate in wild and domesticated grape species. The award is sponsored by the USA-Israel Binational Agricultural Research and Development (BARD) fund. Eleven undergraduate students were trained on berry sampling and analytical approaches at the fruit quality laboratory at Cornell. At NDSU, three graduate students and a Post-doc were trained on leaf material collection for DNA extraction, field phenotyping for disease (powdery mildew, downy mildew), phenology (budbreak, bloom, veraison, wood acclimation), and fruit composition. One graduate student at SDSU was trained in phenotyping and genetic analysis. At Parlier, two undergraduate students were trained on field evaluations for fruit quality, DNA extractions, and other rudimentary aspects of plant breeding. and a graduate student gained additional training on QTL mapping, especially on fruit quality traits.

Three scientists from the project participated at the International Table Grape Symposium, Santiago, Chile in February 2020. Cornell and USDA-Geneva scientists participants attended the Cornell Recent Advances in Viticulture and Enology meeting in Ithaca, New York, in fall 2019. Over 51 scientists took part in a QTL pipeline training including quality control assessment organized remotely using a slack channel in January 2020.

Dissemination of results to communities of interest

Various channels were used to communicate the research efforts and findings from the project. Conference and Summer tour events involving VG2 scientists were organized at NDSU for grape growers and enologists. The North Dakota Grape and Wine Association Annual Conference, Mandan, ND, held in February 2020 had ~60 attendees, and the use of genetic maps for localization of traits of interest, and marker-assisted selection for disease resistance were highlighted. Due to COVID-19 regulations, the North Dakota Grape and Winery Association Summer Farm Tour: Grapevine Breeding Research Update. Agriculture Experiment Station, NDSU, Fargo, ND. Sept. 2020 (Attendees: ~55) took place online. The use of genetic maps for the localization of traits of interest and marker assisted selection for disease resistance were presented. Attendees were also shown the experimental vineyards containing individuals selected with rhAmpSeq markers and observed the variation in fruit ripeness. Attendees were visually shown relative PM resistance of individuals from a target PM resistant cross (Rip.711 x NY 10.0934.01) containing RUN1 allele, compared to individuals with no known PM resistance. Results from VitisGen2 scientists were featured in trade publications (two articles), Wine Business Monthly (circulation 14,000), American Vineyard magazine (circulation 7,700), Good Fruit Grower, and Western Farm Press. Media Services at Cornell highlighted the rhAmpSeq DNA marker platform in a Cornell Chronicle article.

We use social media, webinars, and podcasts to reach our national and international audience. Our scientists – Dr. Clark and Dr. Reisch were recently featured on podcasts: https://www.organicwinepodcast.com/ and https://www.vineyardteam.org/resources/resource-library/pest-management.php id=861, respectively, where they highlighted the work of VitisGen2. The VitisGen2 website (https://www.vitisgen2.org/) is broadly visited by both public and private grape community and the project’s Twitter account (@VitisGen) is constantly used to communicate scientific news to the public and scientist alike.

FUTURE DIRECTIONS

Efforts will be intensified on the current genome sequencing project. Optical maps of V. piasezkii will be used to improve sequence contiguity. Structural and functional gene annotation of the three genome assemblies will be performed to serve as a reference for the full sibling transcriptomic analysis. Additional parental genomes will be sequenced: V. cinerea B9 (Ren2), Horizon (Ren3), Y315-43-04 (Ren1), and V. rupestris B38. A major goal is to focus on the biology of the key traits of powdery mildew and fruit quality.

We  plan to incorporate the rhAmpSeq data into a practical haplotype graph structure used by other breeding programs in the development of genomic selection tools. The plan for implementing this future aim is to partner with the Breeding Insight project to genotype the existing USDA germplasm. Incorporating this genome-wide data for the complete repository system will allow us to fully leverage rhAmpSeq technology to identify key germplasm for important traits. We will partner with Breeding Insight to make the newly developed user-friendly software for MAS widely available for other specialty crops, filling a void for crops where single SNPs are not useful markers for selection.

Resistance gene stacks (vines with multiple resistance genes) will be distributed to the clean plant network for public distribution. Phenotypic effects and downstream RNA pathways in single-gene lines for stacks of known resistance loci as well as how those pathways are affected in stacked lines will be investigated. For unknown resistance loci, phenotypic data for genetic mapping/QTL discovery will be generated and effort will be put in place to phenotype recombinants to narrow resistance loci, improve markers, and identify candidate genes. The project will encourage the widespread distribution of Blackbird imaging robots to scientists at other institutions and locations while we continue to support central screening at Cornell. Immature leaves from at least the V. amurensis x Valley Pearl population will be provided for PM phenotyping in 2021 from USDA-Parlier. Also, we will develop a rhAmpSeq marker set for Erysiphe necator population genetics and fungicide resistance screening. For RNAseq analysis, we plan to expand our isolation, sequencing, and analysis efforts to explore differences in gene expression between stacked and unstacked resistant grapevine genotypes. This is to better understand how resistance genes network in stacked backgrounds and potentially identify optimal stacking designs to increase the resilience of disease resistance.

The project will continue to implement marker-assisted breeding with additional loci on mapping families using recently obtained data. New genetic maps for populations for fruit quality and Ren10 PM resistance segregation will be pursued in Minnesota. At USDA, Parlier, more backcrosses will be performed with a variety of different parents to increase the genetic base of our stacked PM resistant hybrids. Fruit (berries) from the color population will be photographed to provide records of skin color profile of individual vines. At South Dakota, an F2 population will be developed from selected individuals in the F1 population of white riparia x Alicante Bouschet. Seed from 2020 (multiple breeding programs) will be germinated; rhAmpSeq will be employed for marker-assisted selection, and questionable marker-trait associations will be validated. Seedlings are now fruiting in groups that were selected for mono- and di glucoside pigmentation based on AmpSeq markers. These selections will be validated in 2021 based on fruit samples.

Mapping populations from different breeding programs will be processed and analyzed for fruit chemistry. Also, volatiles will be measured by GC-MS on different populations from breeders. Efforts to understand the malic acid degradation difference between wild and cultivated varieties are also planned for the coming year. Metabolomic and transcriptomic data from the time-resolved sampling of Vinifera, Riparia, and Cinerea genotypes will be integrated to detect differentially expressed genes and accumulated metabolites that are associated with the dissimilation of malate in Vinifera berries and lack of in Riparia and Cinerea genotypes. Metabolite analysis of samples stored under normoxia and hypoxia will be completed to test the hypothesis of an association between a differential response to hypoxia and malate degradation. We will continue testing the previously published markers and develop new KASP markers for transferability and accuracy at predicting cluster architecture traits. Fruit texture data will be evaluated for population 11-3527 to detect putative QTL.

We will finalize the analysis of the data collected via the consumer survey and distribute the outcome of the survey on the economics of new varieties for table grapes to both the general and the scientific communities. Extension outputs will focus on the central role of mapping populations, the genetics of sex determination in grapes, etc. We will equally assess the economic impact and the effect of new variety releases in providing an alternative for table and wine grapes.

Target Audience

In 2019/20, the PM phenotyping laboratory (aka, the Robot Lab) remained a frequent site of outreach events focused on communicating the importance of multidisciplinary (biology, informatics, engineering) and multi-institutional research, with about 20 tours provided to scientists and the public, such as high school students interested in engineering, graduate students in plant breeding, stakeholders from grape and biocontrol industries, USDA national program leaders, congressional representatives, and economic development experts.

Through the project’s website (vitisgen2.org), webinars, scientific journals, trade publication articles (Wine Business Monthly, American Vineyard, Good Fruit Grower), a Twitter feed, and non-technical summaries of VitisGen2 journal articles, we reached out to a broad national and international audience of other scientists, growers, and industry leaders, as well as the public. There were notable outreaches to key industry, farmer, and consumer stakeholder groups within the host community and states of participating VitisGen2 institutions. For example, in Minnesota and North Dakota: the Minnesota Grape Growers Association; Minnesota Farm Winery Association; and the Southern Minnesota Wine Grower’s Alliance. Local grape growers and winemakers in North Dakota, Montana, and western Minnesota audiences were reached this year. The scientific and the academic community equally benefit from the numerous publications in top peer-reviewed journals, seminar presentations, workshops websites, and social media posts. We offered training and education to high school students, undergraduates, graduates, post-doctoral associates, and visiting scholars locally and internationally through various programs. Our online survey tools have been important in reaching out directly to grape end-users to generate important information such as trait preferences which are necessary for making consumer-informed improvement decisions.

PUBLICATIONS

Peer-reviewed journal articles

  • Lior Gur, Moshe Reuveni, Yigal Cohen, Lance Cadle-Davidson, Breanne Kisselstein, Shmuel Ovadia, Omer Frenkel (2020). Population structure of Erysiphe necator on domesticated and wild vines in the Middle East shed new light on origins of the grapevine powdery mildew pathogen. Molecular Ecology.
  • Cheng Zou, Avinash Karn, Bruce Reisch, Allen Nguyen, Yongming Sun, Yun Bao, Michael S. Campbell, Deanna Church, Stephen Williams, Xia Xu, Craig A Ledbetter, Sagar Patel, Anne Fennell, Jeffrey C. Glaubitz, Matthew Clark, Doreen Ware, Jason Londo, Qi Sun and Lance Cadle-Davidson (2019. Haplotyping the Vitis collinear core genome with rhAmpSeq improves marker transferability in a diverse genus. Nature Communications. https://doi.org/10.1038/s41467- 019-14280-1.
  • Lu Yin, Avinash Karn, Cheng Zou, Lance Cadle-Davidson, Anna Underhill, Paul Atkins, Dan Voytas, Erin Treiber, Matthew Clark (2020) Genetic mapping and fine mapping of leaf trichome density in cold-hardy hybrid wine grape populations. Frontiers in Plant Science.
  • Teh, S.L., Rostandy, B., Awale, M. et al. Genetic analysis of stilbenoid profiles in grapevine stems reveals a major mQTL hotspot on chromosome 18 associated with disease-resistance motifs. Hortic Res 6, 121 (2019). https://doi.org/10.1038/s41438-019-0203-x
  • The Grape Genome, Cantu and Walker (Eds). Springer International Publishing. Number of Pages: XXVII, 367; DOI: 10.1007/978-3-030-18601-2.
  • Maher, M. F., Nasti, R. A., Vollbrecht, M., Starker, C. G., Clark, M. D., & Voytas, D. F. (2019). Plant gene editing through de novo induction of meristems. Nature Biotech. doi: https://doi.org/10.1038/s41587 019-0337-2
  • Underhill, A. N., Clark, M. D., & Hirsch, C. D. (2020). Evaluating and mapping grape color using image-based phenotyping. Plant Phenomics: 8086309. https://doi.org/10.34133/2020/8086309.
  • Yin, L., Clark, M., Burkness, E., & Hutchison, W. (2019). Grape Phylloxera, Daktulosphaira vitifoliae Fitch (Hemiptera: Phylloxeridae), on Cold-hardy Hybrid Wine Grapes (Vitis spp.): A Review of Pest Biology, Damage, and Management Practices. J. Integrated P Genetic understanding of resistance to foliar phylloxera, Daktulosphaira vitifoliae Fitch, in coldhardy hybrid grapesest Management. doi: https://doi.org/10.1093/jipm/pmz011.
  • Elizabeth M. Demmings, Brigette R. Williams, Cheng-Ruei Lee, Paola Barba, Shanshan. Yang, Chin-Feng Hwang, Bruce I. Reisch, Daniel H. Chitwood and Jason P. Londo (2019). Quantitative Trait Locus Analysis of Leaf Morphology Indicates Conserved Shape Loci in Grapevine. Frontiers in Plant Science 10: 1373.
  • Barba, P., Loughner, R., Wentworth, K., Nyrop, J.P., Loeb, G.M. and Reisch, B.I. 2019. A QTL associated with leaf trichome traits has a major influence on the abundance of the predatory mite Typhlodromus pyri in a hybrid grapevine population. Horticulture Research 6, 87 https://doi.org/10.1038/s41438-019-0169-8.
  • Fresnedo-Ramírez, J., S. Yang, Q. Sun, A. Karn, B.I. Reisch, and L. Cadle-Davidson. 2019. Computational analysis of AmpSeq data for targeted high throughput genotyping of amplicons. Front. Plant Sci. 14 May 2019 / doi: 10.3389/fpls.2019.00599.
  • Demmings, E.M., B. Williams, C.R. Lee, P. Barba Burgos, S. Yang, C.F. Hwang, B.I. Reisch, D.H. Chitwood, and J.P. Londo. 2019. QTL analysis of leaf morphology indicates conserved shape loci in grapevine. Front. Plant Sci. doi: 10.3389/fpls.2019.01373
  • Zou, C., A. Karn, B. Reisch, A. Nguyen, Y. Sun, Y. Bao, M.S. Campbell, D. Church, S. Williams, X. Xu, C.A. Ledbetter, S. Patel, A. Fennell, J. Glaubitz, M. Clark, D. Ware, J.P. Londo, Q. Sun, and L. Cadle Davidson. 2020. Haplotyping the Vitis collinear core genome improves marker transferability in a diverse genus. Nature Communications doi: 10.1038/s41467- 019-14280-1.
  • Julian M. Alston, Torey Arvik, Jarrett Hart, and James T. Lapsley. 2020. “Brettanomics I: The Cost of Brettanomyces in California Wine Production.” Journal of Wine Economics 1–28. doi:10.1017/jwe.2020.20
  • Jarrett Hart, and Julian M. Alston. 2020. “Evolving Consumption Patterns in the U.S. Alcohol Market: Disaggregated Spatial Analysis.” Journal of Wine Economics 15(1), 5–41. doi:10.1017/jwe.2019.14
  • Hart, Jarrett, and Julian M. Alston. 2019. “Under the (Ancestral) Influence: Demographic Drivers of Diverging Demands in the U.S. Alcohol Market“. ARE Update 23(2): 5–8. University of California Giannini Foundation of Agricultural Economics.
  • Jarrett Hart, and Julian M. Alston. 2019. “Persistent Patterns in the U.S. Alcohol Market: Looking at the Link between Demographics and Drinking.” Journal of Wine Economics 14(4), 356–364. doi:10.1017/jwe.2019.26

Peer-reviewed journal article (submitted)

  • Cheng Zou, Melanie Massonnet, Andrea Minio, Sagar Patel, Victor Llaca, Avinash Karn, Fred Gouker, Lance CadleDavidson, Bruce Reisch, Anne Fennell, Dario Cantu, Qi Sun, Jason Londo (2020). Multiple independent recombinations led to hermaphroditism in grapevine. PNAS (Submitted).
  • Underhill, A. N., Clark, M. D., & Hirsch, C. D. Image-based phenotyping identifies QTL for cluster compactness in grape. J ASHS. [Accepted 6/14/20].
  • Yin, L., Karn, A., Zhou, C., Cadle-Davidson, & Clark, M.D. Fine mapping of a foliar phylloxera resistance locus revealed candidate genes in a large population of hybrid grape. [Submitted to Frontiers in Plant Science]

Newsletters, trade and grower magazines

  • Martinson T and Sacks G. “Flavor Challenges of Breeding Disease-Resistant Grapes Using North American Vitis spp.” Appellation Cornell. Issue 39, Nov 2019.
  • Naegele RP, Clark M, Martinson T. “Getting the perfect cluster shape: defining traits and developing DNA markers” American Vineyard magazine. Vol. 29. June 2020.
  • Carlos Martinez-Meija, Breanne Kisselstein and David Gadoury. 2020. Host-Microbe Interactions: a preliminary study of an obligate biotroph, Erysiphe necator, and how it is influenced by its grape host types. Poster presented at the National Summer Undergraduate Research Project (NSURP).
  • R. Naegele, M. Clark, and T. Martinson (2020). Getting the Perfect Cluster Shape: Defining Table & Wine Grape Traits with DNA Markers. American Vineyard Magazine, 79:22-25. June 2020. Malcolm Media, Fresno, CA. Online at: https://cpb-us e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2020/06/AV620ClusterShape pdf
  • T. Martinson, Q. Sun, C. Zou, and L. Cadle-Davidson (2019). Grape Breeders Search for Reliable DNA Markers: Why the Pinot noir PN40024 Reference Genome is Not Enough. Wine Business Monthly. December 2019. 92-97. Online at: https://cpb-us e1.wpmucdn.com/blogs.cornell.edu/dist/c/7890/files/2019/12/Grape-Breeders Search-forReliable-Markers-WBM-December-2019.pdf
  • T. Martinson, B. Reisch, and R. Wiepz. (2020). The central role of mapping populations in marker-assisted grape breeding: A tale of three related mapping populations at Cornell Agritech. Wine Business Monthly, Submitted July 2020. Scheduled for Feb 2021 issue.
  • T. Martinson, A. Kovaleski, and B. Reisch. (2020) Grape mapping populations reveal genetic variation in bloom and fruit development. Blog post at www.vitisgen2.org
  • T. Martinson and B. Reisch. (2020) The Core Grape Genome and Cheap DNA Sequencing: A New Roadmap for Grape Breeders. Appellation Cornell Issue 42, August 2020.
  • T. Martinson and G. Sacks. (2019) Flavor Challenges of Breeding Disease-Resistant Grapes Using North American Vitis spp. Research Focus Article, Appellation Cornell Issue 39, November 2019.
  • J. Van Zoeren, T. Martinson, C. Ledbetter, M. Clark, and B. Reisch 2019. Grape Selections from the VitisGen and VitisGen2 Projects
  • Lee Allan. 2020. Genetic Markers are the Future of Viticulture Western Farm Press, September 2020
  • Sarah Thompson. 2020. Genetic marking discovery could ease plant breeders’ work. Cornell Chronicle, January 2020.
  • A. Underhill (2020). Quantifying color in hybrid wine grapes. RIPE (Research in Plain English) summary of Anna Underhill, Cory Hirsch, Matthew Clark (2020) Evaluating and mapping grape color using image-based phenotyping. Plant Phenomics. 2020, Article ID 8086309 DOI:10.34133/2020/8086309.
  • R. Weipz (2020) What is the economic value of breeding grapes that are resistant to powdery mildew? RIPE summary of Olena Sambucci, Julian M. Alston, Kate B. Fuller, and Jayson Lusk (2020) The pecuniary and non-pecuniary costs of powdery mildew and the potential value of resistant varieties in California grapes. American Journal of Enology and Viticulture, 70 (2), pages 177-187. DOI: 10.5344/ajev.2018.18032.
  • M. Clark (2020) New resistance genes mapped for an important foliar insect pest of some hybrid grape cultivars. RIPE summary of Matthew D. Clark, Soon L. Teh, Eric Burkness, Laise Moreira, Grace Watson, Lu Yin, William D. Hutchison and James J. Luby (2020) Quantitative trait loci identified for foliar phylloxera resistance in a hybrid grape population. Australian Journal of Grape and Wine Research, 24 (3), pages 292-300. https://doi.org/10.1111/ajgw.12341. February 2018.
  • Raising a glass to grapes’ surprising genetic diversity Fruit Growers News, September 2019

Book Chapters

  • James T. Lapsley, Julian M. Alston, and Olena Sambucci. 2019. “The U.S. Wine Industry.” Chapter in Adeline Alonso Ugaglia, Jean-Marie Cardebat, and Alessandro Corsi (eds) the Palgrave Handbook of Wine Industry Economics.

Dissertations/Theses

  • Olson, J. Genetic analysis and characterization of variegation in hybrid grape populations (Vitis spp.)
  • Yin, L. Genetic understanding of resistance to foliar phylloxera, Daktulosphaira vitifoliae Fitch, in cold-hardy hybrid grapes
  • Alahakoon, Dilmini. Exploring Phenotypic Diversity and Quantitative Trait Loci Mapping for Root Architecture, Freezing Tolerance, Chilling Fulfillment, and Photoperiod Traits in Grapevine Populations.

PROJECT OUTPUTS

Webinars

  • NA for 2019-20

Databases and software

  • A database of historical table grape acreage, prices, and volume, by major variety was assembled by the traits economics team.

Protocols

  • NA for 2019-20

Patents

  • NA for 2019-20

Survey instruments

  • The traits economics team developed and implemented an online survey for consumers of table grapes targeted at evaluating consumer acceptance of gene-edited table grapes and specific varietal traits.

Models

  • NA for 2019-20

Educational aids, curricula, training

  • M. Clark. August 28, 2020 Interview on grape breeding with Organic Wine Podcast. https://www.organicwinepodcast.com/?s=clark
  • Facebook Live Podcast, hosted by Briede Family Vineyards, Hybrid Grapes with Bruce Reisch and Nate Walsh. April 2020 Accessed at: https://www.facebook.com/watch/live/ v=234078887699564&ref=watch_permalink
  • J. Alston and A. Sambucci. Feb 20, 2020. (Some of) the Economics of Grape Varietal Innovations. https://www.youtube.com/watch?v=zLH0PDZU7oU&feature=youtu.be 30 live audience, 52 recorded views.
  • Q. Sun, C. Zou, and A. Karn. Apr 14th 2020. A low-cost core genome marker platform that works well across the diverse Vitis genus (https://youtu.be/Y0XpYgkCFIs). 92 live audience, 118 recorded views.
  • Karn A. Organized a virtual workshop over Slack to guide Vitisgen associated scientists and students in genetic map construction using LepMap3 software. 31 attendees. Jan 27-31, 2020.
  • Reisch, B.I. 2020. “Hybrids” with Bruce Reisch and Nate Walsh. Facebook live conference, 22 April 2020 sponsored by Briede Family Vineyards, Virginia.

Websites

  • Website – www.vitisgen2.org: Staff Spotlights featuring Profiles of VitisGen2 staff members by R. Wiepz and J. Van Zoeren for Jack Olson, MS candidate, UMN; Noam Reshef, Postdoc, Cornell; Dilmini Alhakoon, PhD candidate South Dakota State University; Heather Scott; technician, Cornell University; Anna Underhill, USDA ARS, Cornell AgriTech. Others include featured posts (23) and pages in 2019-2020 by Martinson, T. and R. Wiepz, 9472 page views in US, 897 international
  • Social Media: @Vitisgen twitter account established to provide brief updates on project accomplishments. 36 ‘tweets’ posted, resulting in 72,823 ‘impressions’.

Workshops and project meetings

  • Lance

Presentations (including seminars, lectures, and conference talks/posters)

  • Lance Cadle-Davidson. 2020. Computer vision quantification of foliar disease severity. Northeast Division American Phytopathological Society Annual Meeting, in Northampton, Massachusetts, March 11 13, 2020.
  • Cantu, D. “The role of domestication, breeding, and clonal propagation on genomic diversity in grapevines”. Max Planck Institute for Developmental Biology. February 12, 2019, Tübingen, Germany.
  • Cantu, D. “Leveraging plant and microbial genomics to study grapevine diseases”. Penn State University. February 25, 2019, State College, PA.
  • Cantu, D. “Challenges Impacting the California Wine Industry”. UC Davis Chancellor’s Board of Advisors. April 19 2019. Napa, CA.
  • Cantu, D. “The wild side of grapevine genomics”. Plant and Animal Genome XXVIII Conference. January 12, 2020, San Diego, CA.
  • Anne Fennell. Cold Genetics. North Dakota Grape and Wine Association Conference, February 29, 2020, Mandan, ND.
  • Roberto Villegas-Diaz, Dilmini Alahakoon, Jason Londo, Anne Fennell. MetaPipe: A High-Performance Computing pipeline for QTL mapping of large metabolic datasets. XXII Simposio Internacional de Métodos Matemáticos Aplicados a las Ciencias. February 25-28, 2020, Costa Rica.
  • Zou C, Gouker F, Karn A, Fennell A, Cantu D, Xu X, Clark M, Reisch B, Sun Q, Cadle-Davidson L, Londo J. Molecular Characterization of the Sex Loci in Wild and Domesticated Grapes. Plant and Animal Genome Meeting. San Diego, CA. January 12, 2020.
  • Roberto Villegas-Diaz. Parallel computing using Rmpi. XXII Simposio Internacional de Métodos Matemáticos Aplicados a
    las Ciencias February 25-28, 2020, Costa Rica. Git hub repository: https://github.com/villegar/xxii-simmac.
  • Moreira, L., Wannemuehler, S.D., Treiber, E., Suresh, J., Brockman, S., Clark, M.D., & Hegeman, A. Sensory and metabolomic analyses link attributes of flavor and aroma in North American cold hardy grapes. American Society for Horticultural Science, Virtual Conference. (August 10-13, 2020)
  • Clark, M.D., Treiber, E., Karn, A., Zou, C., Cadle-Davidson, L., & Reisch, B. Seedling selection with AmpSeq Enriches Populations for Seedlessness, Muscat Aroma, and Powdery Mildew Resistance. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Moreira, L., & Clark, M.D.¸Characterization of flavor and aroma compounds in Minnesota cold-hardy grapes. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Treiber, E., Moreira, L., & Clark, M. D. Cold-hardy table grapes in Minnesota. 9th International Table Grape Symposium, Santiago, Chile. (February 17-21, 2020).
  • Chin-Feng Hwang, Surya D. Sapkota, Li-Ling Chen and Karlene Negus (2020). QTL Mapping of Botrytis Bunch Rot Resistance in a Vitis aestivalis-derived ‘Norton’-based Population. Abstract for the 2020 Show Me Grape and Wine Conference and Symposium
  • Chin-Feng Hwang and Li-Ling Chen (2019). Optimizing Grape Breeding with Marker Assisted Selection. Abstract for the 2019 International Conference on Grape Genomics and Genetic Breeding.
  • Sapkota, S. D. Martinez, B. Reisch, D. Gadoury, and L. Cadle-Davidson. 2019. Do resistance genes act synergistically against grapevine powdery mildew (Erysiphe necator)?. Plant Health 2019, APS Annual Meeting, 3-7 August 2019, Cleveland, OH, USA (abstr.)
  • Reisch, B.I. and L. Cadle-Davidson. 2019. Perspectives from the VitisGen2 project; Update from Geneva’s grape breeding program. 2019 North American Grape Breeders Conference. 15-16 August 2019. Missouri State University, Columbia, MO, USA. (abstr.)
  • Fennell, A., D. Alahakoon, A. Karn, C. Zou, Q. Sun, J. Londo, B. Reisch, L. Cadle Davidson, and J. Luby. 2019. Genetic analysis of grapevine root system architecture. 2019 North American Grape Breeders Conference. 15-16 August 2019. Missouri State University, Columbia, MO, USA. (abstr.)
  • Reshef, N., E.A. Burzynski-Chang, A. Karn, J. Londo, B. Reisch, G.L. Sacks. 2019. The Real Sour Grapes: Integrated QTL Mapping & Omics to Elucidate Malic Acid Regulation Across Grapevine Species. Society for Experimental Biology Annual Meeting. September 2019. (abstr.)
  • Reshef, N., E.A. Burzynski-Chang, A. Karn, J. Londo, B. Reisch, G.L. Sacks. 2020. The real sour grapes – elucidating malate regulation across grapevine species. 71st American Society for Enology and Viticulture National Conference. 15- 18 June 2020. Portland, OR.
  • Reisch, B.I. 2020. Cornell’s grape breeding program and the VitisGen2 project. 2020. Annual Meeting of the Virginia Vineyards Association. 20 Feb. 2020. Charlottesville, VA.

CHANGES/PROBLEMS

While we did not experience any problem that warranted a change in the proposed objectives, it is important to mention the impact of the COVID-19 pandemic on the project this reporting year. Due to major shutdowns and closures of facilities across the globe in response to the pandemic, we experienced a reduction in executing planned experiments, data collection, and analyses. The closing of DNA extraction labs, greenhouses, and sequencing centers resulted in a long delay in sample collection and submission. The delays at the sequencing centers in the regular rhAmpSeq marker processing and to provide low-cost alternatives to a single-marker analysis led to the testing of KASP and SkimSeq genotyping platforms to support quality data and results to breeders for several powdery mildew resistance loci, fruit chemistry (muscat and diglucosides), seedlessness and seed trace size, and male sterile and female-sterile alleles. These KASP markers were evaluated at Intertek-Sweden where DNA plates were extracted. DNA extraction services and additional commercially available KASP and SSR markers were evaluated at Agbiotech in California. Due to the COVID pandemic, it was not possible this year to set up the precise experiments needed to conduct such sensitive experiments as RNAseq studies. Rather, we decided to use existing DNA to pursue a strategy for candidate gene discovery based on the alignment of shotgun Illumina sequence data to our high-quality PacBio assemblies, for seedlings with recombinations under QTL and with existing phenotype data. This will enable us to narrow in on the specific location of candidate genes, considering structural variants in the source of each trait. At Cornell, a departure of a core transcriptomic and metabolomic analyses facility staff member led to slower processing time. However, we identified an external collaborator to perform these analyses.

Some breeding groups were unable to extract DNA for analysis, but others found ways to do so, though the number of genotypes analyzed was lower and the results of DNA analyses came much later. Leaf sample collections for powdery mildew analysis were delayed, reduced, or eliminated. A proposed cluster architecture experiment was delayed at USDA Parlier. From an ambitious plan of 28 large-scale PM laboratory experiments for the 2020 growing season, we were only able to execute 5. Breeding programs could not access the vineyards to sample leaf tissue for centralized phenotyping from approximately March through June 2020, and laboratory operations remain in reduced capacity with social distancing requirements greatly restricting our abilities to conduct large experiments. In previous years we had up to seven people in one lab to set up each experiment, and now many labs are restricted to one or two people. The robotic arm would help alleviate this, but for this robot, we only have enough trays to hold 3000 samples at a time and could not have moremanufactured due to the long-term COVID-19 closure of our manufacturing partner in Spain. We are developing a partnership with an injection molding manufacturer to produce hundreds of trays to alleviate this bottleneck and provide a low-cost solution for future Blackbird customers.

The shutdown and de-densification requirements after re-opening in many institutions also affected the number of personnel including interns, undergraduate assistants, and contract field labor involved in sample processing and evaluation. Five international scientists who were scheduled to come for research stays had their trips canceled or delayed. Two summer undergrad students were not able to research with us in summer 2020, two new technicians had their starting dates delayed until labs reopen, and two technician positions were abolished. Employment was affected by the pandemic. Travel and contact restrictions associated with the COVID-19 pandemic prevented in-person presentations and field days that were anticipated and scheduled; rather we reverted in most cases to online presentations and electronic publications. Also, we encountered a challenge in mapping 4 newly sequenced reference genomes associated with PM resistant loci (REN1, REN2 and REN3) onto the PN40024 genome (V1 version). The BSR-seq analysis and SMRT bell libraries had nonuniform read coverage over gene bodies, especially for long CDS (> 1kbp) and a lack of coverage of the 3′. Repeating the sequencing will likely solve this issue.