Duration: 10/01/2023 to 09/30/2028

Administrative Advisor(s):

NIFA Reps:

Non-Technical Summary

Statement of Issues and Justification

The Need: Agriculture in the United States contributes to both national and global food security, and supports many diverse industries (e.g., food, ornamental, textile, medicine). Sustainability and diversification of agricultural industries depend on the development of genetically enhanced cultivars to combat emerging pests and diseases, climate and environmental changes, and shifting consumer demands. Germplasm, or genetic resources (sources of genetic diversity), provides the foundation for crop improvement. However, diverse genetic resources are at risk due to reduced diversity in large-scale cultivation, changes in environmental conditions, degradation of native habitats, and international inaccessibility. These resources are difficult or can even be impossible to reconstitute if lost. This is especially true for fruit genetic resources, where preservation clonal identity is paramount.

The mission of the United States Department of Agriculture (USDA), Agricultural Research Service (ARS), National Plant Germplasm System (NPGS) is to acquire, safeguard, document, and distribute plant germplasm, which is accomplished through a cooperative effort with State, Federal, and non-profit partners. NPGS serves research, breeding, and higher education as a public source of plant genetic diversity. The composition of NPGS collections includes landraces, older commercial cultivars, pre-breeding, elite breeding material, and crop wild relatives. As of 2022, more than 16,000 plant species in the form of more than 605,000 accessions were actively held by NPGS. An average of 296,488 samples per year were distributed during the five-year period 2018 – 2022; typically, 70% of these were domestic and 30% were foreign distributions.

The NPGS Plant Genetic Resources Unit (PGRU) located on the campus of Cornell AgriTech at the New York State Agricultural Experiment Station (NYSAES) in Geneva, NY is comprised of the Northeast Regional PI Station for seed crop collections, the National Clonal Germplasm Repository-Geneva, as well as the Apple Rootstock Breeding Program. In 2021, PGRU initiated the new NPGS Hemp (Cannabis sativa) collection. Major collections conserved are tomato, onion, celery, winter squash, brassica vegetable crops, radish, hemp, apple, cold-hardy grape, and tart cherry, including crop wild relatives. The Seed, Hemp, and Clonal repositories hold 12,723, 301, and 6,504 active NPGS accessions, respectively.

Safeguarding these genetic resources is critical to meeting future stakeholder demands, including states in the Northeast US, where many of these crops are principal sources of economic activity and potential commercial growth. The Northeast Regional Multistate Research Project, NE9, brings together representatives from 12 states (CT, DE, ME, MD, MA, NH, NJ, NY, PA, RI, VT, WV) and Washington DC to address mutual interests in plant breeding, research, and extension/education. Members of the Regional Technical Advisory Committee (RTAC) represent state universities, State Agricultural Experiment Stations (SAESs), and the USDA-ARS; the duty of Project Administrative Advisor is assigned to the director of Cornell AgriTech at the New York SAES. Breeding, research, and extension within the NE9 region are supported and strengthened by services and activities performed by PGRU and NPGS. Funding from the NE9 Project has been critical for the realization and sustainability of PGRU germplasm activities. PGRU Seeds and Clonal germplasm projects rely heavily on collaborations for evaluation trials and cultivar development, which are largely beyond the scope of NPGS.

Proposed Objectives: Objectives of this project are directed towards providing the required germplasm to assure stable and sustainable production of vegetable, hemp, and fruit crops in the Northeast USA and worldwide:

1. 
   Efficiently and effectively acquire and maintain the safety, genetic integrity, health, and viability of priority genetic resources, and distribute them and their associated information worldwide. 

2. Develop more effective germplasm maintenance, evaluation, and characterization methods and apply them to priority genetic resources. Record and disseminate evaluation and characterization data via the Germplasm Resources Information Network (GRIN-Global) and other data sources. 

3. With other NPGS gene banks and Crop Germplasm Committees (CGCs) develop, update, document, and implement best management practices and Crop Vulnerability Statements (CVSs) for priority vegetable, hemp, and fruit genetic resources and information management. 

4. Actively engage in and support the development of novel priority vegetable, hemp, and fruit germplasm that integrates diverse, useful genes from various resources and breed, release, maintain, and evaluate improved and regulatory compliant germplasm and cultivars. Devise and apply research tools, knowledge of genetics, and of the genetic control of priority traits to broaden the diversity available for agricultural production systems. The role of the PGRU staff in the development of novel priority germplasm will vary across different crops and projects, and can range from providing germplasm resources to projects, advising project planning and implementation, to direct action on data collection and analysis.

Note: Objectives 3 and 4 require collaboration. Developing strong collaborative relationships among PGRU staff and reliable and productive cooperators are viewed as part of these objectives.

Importance of the Work:  The vegetable genetic resources, which includes tomato, onion, brassica, winter squash, celery, and asparagus, managed by this project represent approximately 36% of the combined dollar value of fresh and processing vegetables in the USA. The average production annual value from 2016-2021 in the US for PGRUs major vegetable collections was $5.145 billion (Appendix A, Table 1). During the past five years, PGRU maintained more than 12,600 accessions of tomato, onion, radish, winter squash, cabbage, cauliflower, broccoli, other brassica crops, celery, tomatillo, asparagus, other vegetables, and buckwheat, representing 29 genera, 151 species and 1,999 taxa. Approximately 150 – 200 seed crop accessions were regenerated per year to replenish stocks. In addition, 285 distinct inventories were acquired since 2017. 

In 2021, the hemp germplasm repository achieved all federal and state compliance to initiate germplasm acquisition, characterization, evaluation, and distribution of regulatory compliant genetic stocks. This collection has grown rapidly to include over 300 accessions and is now the largest international public hemp germplasm collection. The collection contains feral, fiber, grain/oilseed, secondary metabolite, landrace, breeding lines, and other classes of diverse germplasm. Distribution of regulatory compliant materials began in 2022. Work is planned for 2023 to genotype the entire collection and to conduct population structure analysis. These genotyping efforts will guide collection and conservation priorities, development of mapping populations, and provide higher stakeholder utility as inputs into breeding programs.

PGRU maintains 4,940 accessions of Malus, 1,415 accessions of Vitis, and 149 accessions of tart cherries. Through introductions, explorations, and exchanges, PGRU acquired 104 Malus, 1 Vitis, and 13 Prunus accessions since 2017. The fruit crops maintained by PGRU account for about 49% of the value of US fruit and vine crop production (Appendix A, Table 1). Apples, grapes, and tart cherries contribute significantly to the US economy, with average annual values of $3.129, $5.908, and $0.062 billion from 2016-2021, respectively (Appendix A, Table 1). 

Historically, the NE9 Project has made substantial contributions to the vegetable and fruit industries through distribution of germplasm and associated information for developing improved varieties with higher and more stable yield, disease and insect resistance, and improved quality. From 2017 – 2022, PGRU distributed 52,612 seed lots (47% domestic and 53% foreign) comprising 11,155 unique seed accessions. PGRU distributed 24,399 clonal crop samples (97% domestic and 3% foreign) comprising 3,790 unique accessions. In NE9 states, there were 4,930 seed samples from 3,788 accessions distributed and 8,778 clonal samples from 2,8236 unique accessions distributed (Appendix A, Tables 2-7). Close to 40% of distribution of PGRU fruit germplasm is directed to NE9 states. The collections have been extensively used worldwide to develop new cultivars and for other research purposes, such as genetic analysis of disease resistance, fruit quality, genetic diversity, and population structure. PGRU scientists characterize germplasm for priority traits to make the material more readily accessible. Much of this characterization and evaluation is performed in collaboration with scientists from the NE9 region and other regions in the USA and abroad. Research into quality and health-beneficial traits was initiated at the request of partners in various CGCs and has become increasingly emphasized.

Technical Feasibility and Value of a Multi-state Project:  

:  Acquisition, conservation, and characterization of germplasm collections are more efficient at a central location than through individual state organizations, which would result in unnecessary duplication of efforts. A cooperative approach among state partners and the PGRU allows for an efficient conservation of vegetable, hemp, and fruit germplasm while plant breeders and other researchers can take the lead in characterization and evaluation, especially for quantitative traits that require replicated field trials. Utilization of germplasm for crop improvement by geneticists and breeders at individual SAESs capitalizes on the genetic resources and the characterization/evaluation information maintained by the NPGS.

PGRU is primarily supported by appropriated funds authorized by Congress, which provides long term stability for performing basic genebank activities. It is located within a vibrant agricultural region at Cornell AgriTech on the NYSAES campus and is well suited to take maximum advantage of additional multi-state funds from the NE9 project for conservation and characterization/evaluation of vegetable, hemp, and fruit germplasm of important crops to the Northeast region.

Funding from NE9 provides critical resources for better management of the collections and quality service of germplasm distribution. It also supports major efforts in supplying germplasm to screen for high-priority traits, such as important disease and pest resistances and traits important to human health, much of which is done in collaboration with scientists from SAESs. The budget allocated to NE9 over the last 5 years was kept constant, even though salaries and expenses kept increasing. To retain skilled staff and maintain our current operational levels with the addition of a high-value crop, a 7% increase in the budget is requested, matching the 7% increase the Northeast region received during the last 5 years for Hatch and Multistate federal allocations. The budget will stay constant for the 5-year project. This increase will allow NE9 to maintain resources for a growing, valuable genetic resource.

Impact: Genetic resources in the PGRU repository will continue to prove useful in developing improved cultivars of vegetable, hemp, and clonal crops while supporting and stabilizing agricultural production. Expansion of global trade and complex food systems, increases the risk of introduction of exotic pests and diseases to the United States.  Agricultural research in the United States has intensified efforts to improve the sustainability of national food production while reducing its deleterious impacts. For example, crops with genetic resistance to pests and diseases reduces dependency on pesticides and preventative chemical sprays and reduces risks for agricultural workers and impact on the environment. In response to changing environmental conditions, PGRU collections will be used as sources of resistance to environmental stresses to increase the range of adaptation of crops. The Northeastern United States is particularly well suited to embrace emerging agricultural opportunities, to which PGRU can provide well documented germplasm for our growing conditions.

Finally, maximizing the use of available germplasm at PGRU will help domestic producers thrive in a competitive global marketplace. For example, within the current NE9 project, PGRU has provided accessions of tomatillo and cabbage as sources of natural pigments for breeding programs aimed at emerging markets; screened the tomato collection for biotic and abiotic resistances; identified a wild tomato accession used for late blight resistance in cultivated tomatoes. In 2021-2022 PGRU has assembled the world’s largest public hemp germplasm collection, collected approximately 100,000 data points for priority horticultural and agronomic traits, and these materials have been used to develop five long-read whole genome sequences to be deposited in the National Center for Biotechnology Information. The hemp germplasm collection has been used to develop fiber quality standards that will underpin an emerging domestic fiber industry. Apple germplasm has been utilized to develop new rootstocks with multiple disease resistances and improved plant architecture. Both apple and grape accessions have been screened for biotic and abiotic resistances, leading to new scion cultivars being developed in large, multi-state projects.

Germplasm from PGRU has proven useful in developing improved cultivars of vegetable and fruit in the Northeast region, the USA, and the world:

• Genes from wild tomatoes have been exploited to increase ease of harvesting, disease resistance, and for stress and drought tolerance.
• More than 20 genes from the PGRU tomato collection for bacterial speck, spotted wilt virus, tobacco mosaic virus, leaf mold, fusarium wilt, verticillium wilt, late blight, and nematode resistance have been bred into modern varieties.
• Phylloxera resistant grape rootstocks and hybrids derived from North American wild Vitis germplasm were instrumental in rescuing the European grape and wine industry.
• The recent spread of grape cultivation throughout the USA, especially in the northeast, has been made possible by use of the germplasm collection for breeding of new cultivars of Vitis vinifera and Vitis vinifera – Vitis species hybrids that are adapted to environments where vinifera could not previously be grown.
• Genetic resources for resistance to apple scab, fire blight, and wooly apple aphids maintained in the germplasm collection have been deployed in disease resistant apple rootstocks and cultivars. Insect and disease resistant apple cultivars can be traced back to the PGRU apple collection.
• PGRU apple germplasm, especially historic cultivars, has been used directly by growers to expand the US apple cider industry.

Germplasm maintained at PGRU is currently or will be used for crop improvement of fruits, hemp, and vegetables:

• Tomato accession are being tested for resistance to race 1 strains of Rs5 bacteria, ToBrFV virus, salt, and drought tolerance in growth chamber facilities. These phenotypic characterizations of the PGRU tomato germplasm collection will identify genomic regions controlling pathogen and stress tolerances and add significant value to our long-term breeding effort and cultivar development.
• Cucurbit accessions have been evaluated for resistance to the oomycete pathogen Phytophthora capsica, which result in outbreaks that are challenging to manage and can result in huge yield loss.
• Specialty cucurbit accessions from PGRU and other NPGS collections will be screened for powdery and downy mildew resistance, and critical agronomic traits as novel specialty crops to serve historically underserved communities in the Northeast.
• Hemp germplasm from PGRU has been used to create five of the highest-quality cannabis sativa reference genomes and will likely contribute to the development of a cannabis pan-genome.
• PGRU’s hemp collection has been screened for plant architectural traits, fiber and grain agronomic characteristics, and secondary plant metabolite composition and profile. This data will enhance subsequent curation, distribution, and evaluation efforts.
• PGRU’s vegetable, hemp, and fruit germplasm collections are being screened for medicinal and nutraceutical properties for development of cultivars that will improve the health benefits of consumption.
• Genomic resources developed for major apple progenitors from Central Asia and other wild Malus are key to accelerating breeding programs to introduce resistance and adaptability traits in modern apple cultivars.
• Grape germplasm will continue to be used in developing new grape cultivars for better resistance to disease and climate change.

Related, Current and Previous Work

PGRU is responsible for acquiring, conserving, distributing, and characterizing genetic diversity of an array of vegetable, hemp, and fruit crop taxa adapted to temperate regions, including crop wild relatives (CWRs). This reservoir of genetic diversity contributes to food security and meeting human nutritional requirements. Seed or clonal cuttings, pollen, and DNA of accessions plus associated information are distributed worldwide for purposes of breeding, research, and higher education. Gaps in collections must be filled to preserve traits that have evolved naturally or through human selection over thousands of years. Germplasm is systematically regenerated, quality tested, and securely backed-up to ensure long-term availability. Regeneration and cultivation protocols must be assessed and refined to overcome dynamically changing pest, disease, and weather conditions. Descriptions of traits including growth, morphology, phenology, production, disease resistance, and health beneficial components are collected to provide for targeted requests and efficient utilization of accessions. Community-wide expertise in best management practices is leveraged to improve efficiency of operations and to document the skills and knowledge required to effectively execute this complex project. Crop vulnerability statements (CVSs) are consulted and updated in collaboration with respective CGCs, ensuring long-term protection of major components of the vegetable and fruit industries. 

Some of the on-going research cooperation includes: 

• M. Mazourek, Horticulture, Cornell University and Dr. Z. Fei, Boyce Thompson Institute: germplasm diversity in winter squash.
• A. Shi, University of Arkansas: screening and evaluation of tomato genetic resources for biotic and abiotic stressors.
• S. Kousik and Dr. K. Ling, USDA-ARS USVL: evaluation of vegetable genetic resources for pathogen and viral resistance.
• S. Branham, Clemson University: development of a leafy brassica genome-wide association panel for priority trait evaluation.
• C. Brummer, UC-Davis: development, evaluation, and contribution of germplasm to the hemp collection.
• S. Ellison, UW-Madison: feral hemp germplasm collection, evaluation, and contribution of germplasm to the hemp collection.
• L. Smart, Cornell University: phenotyping and genotyping of the hemp germplasm collection.
• C. Delhom, USDA-ARS SRRC: germplasm evaluation and development of hemp fiber quality standards.
• M. Berhow, USDA-ARS NCUAR: germplasm evaluation and protocol development of hemp secondary metabolites.
• E. Cebert, Alabama A&M University: germplasm evaluation and regeneration.
• C. Smart, Cornell University: pathogen screening of PGRU hemp germplasm.
• P. Bates, Washington State University: hemp seed fatty acid and protein method development and germplasm screening of PGRU collection.
• S. Brown, Horticulture, Cornell University: (1) apple scion breeding and (2) characterization of fruit quality traits of the Malus collection.
• K. Xu, Horticulture, Cornell University: genetic and genomics of fruit quality and tree architecture traits.
• L. Cheng, Horticulture, Cornell University: evaluation of bitter pit using Malus germplasm.
• G. Peck, Horticulture, Cornell University: (1) importation of new cider apple varieties from England, (2) evaluation of Malus collection for hard cider production potential, and (3) establishing new hard cider varietal trials in NY.
• O. Hurtado-Gonzales, USDA APHIS: import/quarantine of apples, genetic characterization of incoming Malus germplasm, and virus testing of the apple collection.
• A. Khan, Plant Pathology and Plant-Microbe Biology, Cornell University: (1) screening Malus collection for new fire blight and scab resistance, (2) importation of international apple varietal set for apple scab screening and monitoring, (3) genetic mapping of fire blight resistance genes and leaf spot resistance gene, (4) allele mining of disease resistance genes in Malus and Vitis.
• M. Fuchs, Plant Pathology and Plant-Microbe Biology, Cornell University, study of viruses in Malus and Vitis.
• M. Rivera, Entomology, Cornell University: trapping and monitoring of black stem borer of Malus.
• B. Reisch, Horticulture, Cornell University: (1) grape scion breeding and (2) VitisGen project.
• L. Cadle-Davidson, USDA-ARS GGRU, Geneva, NY: (1) resistance gene mining for powdery and downy mildews, (2) high-throughput phenotype resistance screening (including AI data mining), and (3) VitisGen project.
• G.Y. Zhong, USDA-ARS GGRU, Geneva, NY: aromatic compounds in Vitis germplasm.
• C. Heinitz, USDA-ARS: fruit quality traits of Vitis cultivars.
• K. Gold, Plant Pathology, Cornell University: Resistance to various fungal pathogens in Vitis germplasm.
• R. Hestrin, Plant Biology, University of Massachusetts Amherst: Soil microbiome composition in Vitis germplasm soils, by species and root structure.
• J. Londo, Horticulture, Cornell University: climate change adaptations of Malus, Vitis, and Prunus genetic resources.
• K. Gasic, Plant and Environmental Sciences: Clemson University, cold-hardy evaluation of Prunus genetic resources.
• C. Gottschalk, USDA-ARS, Kearneysville, WV: (1) apple breeding, (2) genomic characterization of apple germplasm, (3) exploration of wild apple germplasm.
• Gennaro Fazio, USDA-ARS PGRU, genetic characterization and rootstock evaluation of apple genetic resources.
• Apple and Grape Vulnerability Statements updates, with members from Apple and Grape CGCs, including many scientists from Cornell University. 

Acquisition: 

Collecting and conserving geographic and ecological diversity of CWRs in situ and ex situ is of high priority, as many wild populations face extinction due to various factors such as pests and diseases, competition with invasive species, and permanent alteration or loss of habitats. With respect to acquiring novel germplasm, obtaining precise estimates of gaps in cultivars, breeding material, and landrace or weedy material presents challenges because of the predominant influence of human activities. Germplasm acquired to fill gaps in PGRU collections is based on enhancing diversity of collections to reduce risks to agriculture.  

Significant accomplishments in the period 2017-2022 include:

• 285 new accessions or mapping populations were acquired of onion, celery, brassica, winter squash, buckwheat, tomatillo, radish, tomato, and miscellaneous taxa.
• 301 new accessions of hemp were acquired.
• 104 Malus, 1 Vitis, and 26 Prunus accessions were added. 

Maintenance and regeneration: 

For the PGRU seed repository, accessions are routinely generated as seed stocks are depleted through distribution and viability is reduced during storage. Regeneration and maintenance procedures must minimize genetic changes within accessions. Loss of genetic diversity through genetic drift and through unintentional selection is avoided to the extent possible by managing the optimal health and size of regeneration populations. Long term safety of collections is ensured by back-ups of 2,000 seed with a minimum of 85% viability per accession at National Seed Storage Laboratory (NSSL) in Ft. Collins, CO. Additional backup is at the Svalbard Global Seed Vault. For clonal crops, cryopreservation at NSSL serves as backup storage for Malus and tart cherry accessions. Wild Malus and Vitis seed accessions are maintained in cold storage by the PGRU and NSSL. A research strategy for long term storage of Vitis is being investigated by NSSL with cooperation from the National Clonal Germplasm Repository in Davis, CA, and PGRU. 

Significant accomplishments in the period 2017-2022 include:

• Regenerations and germplasm rescues were completed for over 1,000 vegetable crop accessions in Geneva, NY and 192 accessions were regenerated via collaborator increases.
• 1,651 backup samples for seed crops were added or replaced at the NLGRP.
• A Cooperative Agreement with University of California, Davis was used each year to support the CM Rick Tomato Genetics Resources Center (TGRC) of unique wild and genetic stock tomato accessions.
• Hemp collaborative regens were initiated at Oregon State University, University of California-Davis, and Alabama A&M University.
• 250 Malus and 150 Prunus accessions submitted for cryopreservation. Section and propagation of 250 wild apple seedlings for permanent maintenance. Seedling blocks of >2,000 trees removed following propagation.
• Seeds of wild and hybrid grapes were collected to backup allelic diversity. 

Characterization: 

Grant proposals funded by USDA upon recommendation from CGCs serve as a unique and valuable resource with which to study high priority traits in NPGS germplasm collections. NPGS germplasm and associated information are provided to the investigative team and resultant data are deposited into GRIN-Global and/or other public data bases. As ex-officio members of any CGC that includes PGRU crops conserved, the PGRU curators have firsthand knowledge of proposals submitted for screening NPGS germplasm and whether they are funded. Criteria for funding include a current Crop Vulnerability Statement, scientific merit, the national need for evaluation data, the likelihood of success, and the likelihood that the evaluation data will be entered into GRIN-Global. 

Significant accomplishments in the period 2017-2022 include: 

• PGRU scientists in collaboration with Dr. Shi at the University of Arkansas are evaluating and conducting genome-wide association study and genomic prediction for bacterial wilt and salt and drought tolerance in the PGRU tomato germplasm collection.
• In collaboration with Dr. Mazourek and PhD student, Marlie Luckach, PGRU is evaluating exotic cucurbits (luffa/ridge gourd, bitter gourd, pepinillo, and calabaza) for biotic pathogen resistances and suitability for emerging New York markets.
• In collaboration with PGRU researchers, two large hemp Supplemental and Alternative Crop germplasm acquisition and characterization grants have been awarded. These grants will add an estimated 500 hemp accessions to the PGRU collection and collect phenotypic data for approximate 75 priority traits.
• PGRU researchers drafted the UDSA Hemp Phenotyping and Descriptor Manual. This manual is being implemented by multiple groups to standardize phenotyping protocols across diverse hemp germplasm.
• PGRU researchers are collaboration with other USDA-ARS and Cornell University researchers to develop a hemp Breeding Insights phenotyping and breeding platform to characterize and breed enhanced hemp genetic resources.
• Following a fire blight outbreak, PGRU completed a disease resistance evaluation of the entire Apple Collection, one of the largest evaluations of its kind (Dougherty et al. 2021).
• PGRU with its collaborators explored the genomic impact of apple domestication and developed genome and pangenome resources for wild Malus species focusing on domesticated apple’s primary progenitors (Sun et al. 2020; Migicovsky et al. 2021).
• Additional genetic markers (20K SNP array) ~300 apple accessions were generated and will be used to evaluate the pedigree and taxonomic gaps of the Apple Collection.
• Summarized an evaluation of the Vitis collection for phenological diversity. PGRU identified unique germplasm that may be suited for cold-climate breeding objectives (Gutierrez et al. 2021).
• PGRU completed a five-year fruit quality evaluation of 135 tart cherry accessions, identifying key cultivars which could improve the nutritional value of the US tart cherry industry. 

Documentation:

CGCs maintain and update descriptor lists for crop traits that are highly heritable and therefore can be evaluated without costly, replicated experimental trials (http://www.ars-grin.gov/npgs/cgclist.html). There are typically dozens or more traits for a crop which are grouped into the major categories of disease resistance, growth habit, fruit morphology, phenology, and production. We continue to test and implement new methods and technologies for high-throughput data collection, such as FieldBook to facilitate data collection. In addition, digital images of plants and their parts (e.g., fruit, flowers, bulbs) are collected and deposited into GRIN-Global. 

Significant accomplishments in the period 2017-2022 include: 

• Phenotypic data, including digital images, were collected for routine regenerations of tomato, onion, brassica and winter squash accessions using CGC descriptors for highly heritable traits.
• Over 11,000 original seed lot passport data were digitized and associated with accessions in GRIN.
• Over 100,000 data points were collected for hemp genetic resources and will be associated with GRIN.
• Protocols for in-house seed viability tests were adopted and implemented in 2016. Several–hundred tests on regeneration samples were successfully completed.
• Documentation and routine evaluation of apple was limited due to fire blight outbreaks in the collection. However, fruit weights and images of 367 accessions of wild and hybrid Malus were documented and will be used for taxonomic evaluations in the Apple Collection. 

Distribution: 

New cultivars developed from germplasm contribute to diversity in crops, expand variety in diets, and provide benefits to human health and nutrition. Progress in crop genetics, genomics, genetic improvement, and horticultural production are accelerated by the information and genetic resources supported by NE9. Information associated with collections is distributed through GRIN-Global and other public databases such as National Center for Biotechnology Information (NCBI) or Sol Genomics Network (SGN). 

Significant accomplishments in the period 2017-2022 include: 

• The GRIN-Global public website interface for ordering accessions and accessing information has been substantially improved to make it more user-friendly and compatible with a wide range of electronic devices.
• PGRU distributed 33,466 samples (49% domestic and 51% foreign) of 29,092 seed crop accessions. On average, 5,818 unique accessions were distributed per year, i.e., approximately 46% of the active seed collections each year (Appendix A, Tables 2 and 3).
• International and domestic hemp seed distribution has begun in 2022.
• PGRU distributed 22,097 clonal samples (Appendix A, Tables 4 and 5) in the form of scions, seed, fruit, pollen, DNA, leaves, or use of trees for controlled This included an increase in request for “Botany of Desire” seeds (open pollinated seeds of M. sieversii) from the USA and particularly from European countries such as Austria, England, Finland, France, and Germany 

Research: 

NE9 is a cooperative research project among the State Agricultural Experiment Stations within the Northeast region of the USA. PGRU strives to provide vigorous, pathogen-inspected, and quarantine-acceptable germplasm samples to researchers and breeders. PGRU curators continue to serve as co-PIs or close collaborators on experimental studies, to ensure success of the research and follow-up on depositing data into GRIN-Global or other public databases. Genotypic research focusses on the analyses of diversity within and among accessions and taxa, as well as identifying functional alleles. Phenotypic research prioritizes traits for genetic improvement, such as tolerance of environmental stresses and extremes, nutritional content, flavor, color, horticultural traits, and host-plant resistance to diseases and pests. 

Significant accomplishments in the period 2017-2022 include: 

• Significant progress was made in digitizing early accession correspondence and passport information. For example, over 13,000 paper forms were digitally scanned, associated with their respective accession, and linked digitally within GRIN-Global.
• PGRU began a long-term regeneration of the asparagus collection using 56 unavailable or jeopardized seed stocks. In addition, we initiated a genotype-by-sequencing study evaluating 116 unique asparagus cultivars from 29 countries. This work evaluates overall genetic diversity and population structure within the broader pool of asparagus germplasm, as well as putative genetic bottlenecks that occurred during domestication processes. PGRU has identified several critical sub-clusters of unique genetic diversity within asparagus germplasm using approximately 41K high-quality DNA markers.
• PGRU completed a genetic diversity study on tomatoes (Labate, 2021) including partial genome sequencing on 190 tomato stocks from the PGRU collection and will provide gene discovery tools and other genetic information to increase the efficiency of selection and breeding.
• The PGRU radish collection was evaluated for genetic diversity (Arro and Labate, 2022) using 152 diverse accessions. These efforts will support ongoing radish conservation and improvement efforts.
• PGRU completed an evaluation of Brassica oleracea diversity using a diversity panel of 225 accessions from 19 different morphotypes and eight crop wild relative species and examined patterns of relationships among them (Mabry et al., 2019). These analyses point to the closest living relatives of oleracea as Brassica incana and Brassica cretica, indicating support for origin of cultivation in the Mediterranean region, and recover evidence for multiple origins of kales.
• The Hemp Germplasm Repository has met USDA-ARS, USDA-AMS, DEA, and NY State Hemp Program regulatory compliance for required germplasm collection, conservation, distribution, evaluation objectives. Hemp germplasm collection, regeneration, characterization, and evaluation efforts are already underway. At time of writing, the PGRU hemp germplasm holds over 300 unique accessions spanning all usage classes. To our knowledge, this is the largest publicly available hemp germplasm collection. Hemp has been added as a new crop within the New Crops Crop Germplasm Committee.
• PGRU has developed the first hemp phenotyping and descriptor handbook to assist breeders and researchers in identifying accessions with specific traits to facilitate germplasm selection within hemp (Cannabis sativa L.) improvement programs (Stansell and Osatuke, 2021). These efforts will help to identify gaps in the existing hemp collections and help formulate strategies for future collection and conservation efforts, to designate and maintain a core collection of critical materials, to increase NPGS user utility and accessibility to hemp germplasm and associated data, and to identify duplicate accessions and reduce costs of hemp genetic resource conservation.
• Major apple phenolics were evaluated and characterized genetically. In particular, research was done on apple dihydrochalcones, a group of apple-specific phenolics associated with human nutrition and Malus evolution and physiology. These compounds could be critical in elucidating the natural histories of apple species and a way to enrich the nutritional quality of modern apple cultivars (Gutierrez et al. 2018).
• PGRUs tart cherry collection was evaluated for viruses, from which collaborators identified a novel trichovirus which infects sweet cherries (Brewer et al. 2020).
• Phased diploid and pan-genomes were developed for cultivated apple and its wild progenitors. These resources help to elucidate the domestication history of apple and allow future research to leverage genomic resources to characterize traits in apple relatives (Sun et al. 2020).
• The apple collection was evaluated for fire blight resistance during a severe outbreak. The evaluation covered over 2,500 accessions and 48 taxa, representing one of the largest of its kind. Genome mapping of historic fire blight evaluation data identified novel loci related to resistance (Thapa et al. 2021; Dougherty et al. 2021).
• The apple collection was further characterized for kinship using GBS markers. Of the 1,900 cultivars evaluated, close to 900 of them had at least one first degree relationship, signaling substantial redundancy within the collection (Migicovsky et al. 2021).
• Foxy aromas in grapes are predominantly from methyl anthranilate, originating from Vitis labrusca and its hybrids. PGRU germplasm was evaluated for foxy aromas, and a major gene controlling for methyl anthranilate was described. Though foxiness may contribute positively to table grapes (Concord as an example), for wine these aromas are unfavorable, limiting the utilization of labrusca germplasm for breeding. The genetic control described will allow breeders to select against foxy aromas using marker assisted breeding. Over 1,300 accessions were evaluated for this allele (Yang et al. 2020).
• Over 5 years the Vitis collection was evaluated for phenological traits, including budbreak, bloom, and veraison. These three developmental traits set the limits for grape production. PGRU evaluated over 1,500 accession across 20 taxa. Increasing heat requirements for bloom was strongly associated with increased percentage of Vitis vinifera the European grape (Gutierrez et al. 2021).
• Profiling of epigallocatechins in grape species identified a phylogenetic linkage across diverse Vitis species (Brillouet et al. 2022).

Objectives

1. Efficiently and effectively acquire and maintain the safety, genetic integrity, health, and viability of priority genetic resources, and distribute them and their associated information worldwide.
   
2. Develop more effective germplasm maintenance, evaluation, and characterization methods and apply them to priority genetic resources. Record and disseminate evaluation and characterization data via the Germplasm Resources Information Network (GRIN-Global) and other data sources.
   
3. With other NPGS gene banks and Crop Germplasm Committees (CGCs) develop, update, document, and implement best management practices and Crop Vulnerability Statements (CVSs) for priority vegetable, hemp, and fruit genetic resources and information management.
   
4. Develop novel priority vegetable, hemp, and fruit germplasm that integrates diverse, useful genes from various resources and breed, release, maintain, and evaluate improved and regulatory compliant germplasm and cultivars. Devise and apply research tools, knowledge of genetics, and of the genetic control of priority traits to broaden the diversity available for agricultural production systems.
   

Methods

1. Efficiently and effectively acquire and maintain the safety, genetic integrity, health, and viability of priority genetic resources, and distribute them and their associated information worldwide.

NE9 will continue to serve as a conduit for movement and exploration of valuable plant genetic resources from worldwide origins to the northeastern states and beyond (Appendix A, Tables 3-7). The PGRU is equipped with facilities and equipment for conducting its service, research, education, and outreach activities (Appendix D). 

Acquisition

PGRU will identify collection gaps by reviewing taxonomic representation, geographic distribution, or missing allelic or phenotypic variation. These gaps will be filled through germplasm exchanges, cooperator donations, expired Plant Variety Protection materials, and explorations. Genetic diversity of tomato, B. oleracea, onion, winter squash, radish, and celery collections will be restored and enhanced by identifying gaps and sources of germplasm to fill the gaps. Genesys free online portal (https://www.genesys-pgr.org/welcome) will be used to explore sources of seed. Genesys is a plant genetic resources accession database that contains 3.6 million accession records from 481 institutes: the three largest being US-NPGS, Consortium of International Agricultural Research Centers (CGIAR), and European Cooperative Programme for Crop Genetic Resources Networks-European Internet Search Catalog (ECPGR-EURISCO). The USA became a Party to the Food and Agricultural Organization of the United Nations (FAO) International Treaty on Plant Genetic Resources for Food and Agriculture in March, 2017. 

Taxonomic gaps in apple are especially important, as recent genetic studies identified incorrect classification due to genetic admixture in hundreds of PGRU accessions (Volk et al. 2022). We will pursue plant exchanges with international collections for specific cultivars and wild germplasm inaccessible within the United States. For apple, cider cultivars are still of particular interest to stakeholders and a target for acquisition. Duke cherry cultivars (sweet cherry × tart cherry hybrids) are underrepresented in the NGPS and are targets for acquisition to expand the Prunus collection. Explorations for wild germplasm will target key species underrepresented in the NPGS, particularly domestic explorations for North American apple, grape, and Prunus species. 

Cannabis germplasm exploration and collection efforts are challenging due to a suite of complex international relationships. However, there are two hemp germplasm collection efforts in the early stages of planning, in Northwestern Vietnam and Uzbekistan. These regions have been identified by stakeholders as sources of locally adapted germplasm to abiotic stress tolerances and photoperiod insensitivity.

A recently funded USDA Postdoc collection and evaluation fellowship was awarded titled “Investigating the utility of feral populations for Brassica crop breeding" which describes a highly valuable and unique approach to address many important questions within the broader domain of crop genetic resource conservation and plant breeding. Specifically, this proposed work will investigate feralization within Brassica crops as a mechanism to both enhance conservation efforts while providing new insight into genetic control of priority traits (e.g., plant architecture, phenological development, yield, and domestication). This work would provide great value to the National Plant Germplasm System by identifying, collecting, and safeguarding vital genetic resources and support of breeding and other improvement efforts. NPGS curators will receive, accession, conserve, and help with characterization of genetic materials gathered during this work and ensure that phenotypic or other associated information generated from this research is associated with relevant accessions within GRIN-Global.

Preservation

We will use best management practices (BMP) to preserve and safeguard the vegetable, hemp, and fruit collections. We will ensure the long-term safety of collection by systematically completing backups of accessions at NLGRP, Fort Collins, CO. To meet distribution needs of NPGS customers and stakeholders, any accession that drops below minimum seed quantity (e.g., 1,000) or viability (80%) is routinely regenerated. Typically, 150 to 200 accessions are regenerated annually in-house or via collaborative increases. Regeneration planning and execution are being streamlined via prepopulated forms held in shared cloud storage. This approach mitigates numerous hand-off issues experienced between teams. Dr. Stansell has developed an automated, reproducible computational algorithm for identifying and assigning priorities for the approximately 60,000 unique inventories in the seed crop collection by evaluating inventories in greatest need of regeneration, rescue, and backup. Inventories are computationally processed within the R statistical environment, scored for seed age, quantity, viability, backup status, and stakeholder usage, and then assigned an aggregated regeneration/rescue/backup score. These reports are checked by hand against existing inventories and are then slated for future regenerations. 

The PGRU Clonal fruit collections are primarily maintained as field collections and vulnerable to abiotic and biotic stress. To better safeguard field collections, PGRU will enhance orchard and vineyard maintenance practices. The largest threat to the apple collection is fire blight, a bacterial disease which spreads quickly through an orchard and can kill susceptible trees within a season (Dougherty et al. 2021). Apple rootstocks can mediate scion qualities (Marini and Fazio 2018; Singh et al. 2019), which could improve PGRU maintenance practices through enhanced resistance to fire blight and other biotic and abiotic stresses, improve grafting success rate and nursery viability, reduced root suckering, and reduced tree size to accommodate higher density plantings. To evaluate new rootstocks, we will propagate a diverse panel of 100 PGRU Malus accessions onto four Geneva series rootstocks: G.890, G.210, G.30, and CG.6006, with M7 as a control. Each is a semi-dwarf rootstock that produces a tree 10-20% smaller than M.7 and has improved disease resistance, with reduced suckering relative to M.7 and a low tendency for biennial bearing compared to M.7. M. domestica and wild Malus accessions will be selected for evaluation based on genetic diversity using genetic data, ploidy variation, and phenotypic diversity. 

The grape collection is currently grown on an Umbrella Kniffin (UK) system, which requires yearly regrowth of canes and does not facilitate mechanical pruning. Vines will be shifted to a high wire cordon (HWC) to allow better canopy control through mechanical pruning, pesticide penetration, budwood growth, and air circulation around fruit. Highly vigorous accessions (primarily wild Vitis) will be repropagated to T-trellis systems over the next 5 years, and trained to a Munson system or Geneva Double Curtain (GDC) to better control shoot growth by inducing shorter internodes and fewer lateral shoots (Atucha and Wimmer 2018). 

Long-term storage of biological material in liquid nitrogen through cryopreservation provides a way to preserve clonal material free from environmental pressures and serves as a backup to replace field accessions lost to disease or catastrophic events. Currently, 70% of the permanent apple accessions are successfully cryopreserved as budwood. We coordinate closely with NLGRP (G. Volk) for cryopreservation of apple and tart cherry genetic resources. As of 2021, 2,088 apple accessions were considered backed up in NLGRP. The goal is to ensure that >95% of permanent Malus and tart cherry accessions are cryogenically backed up. Currently, 43 tart cherries and 460 M. domestica accessions remain to back up. Cherries will be completed in 2 years, and we will target 75-100 apples each year over the five-year period. Vitis cryopreservation methods are too laborious to apply to the PGRU collection. As such, seeds of select wild and hybrid Vitis accessions will be collected to preserve allelic diversity in the collection. Other potential backup methods are also being investigated and considered for viability within larger germplasm collections. 

Distribution

The distribution of genetic resources is our most direct and meaningful contribution to customers. Delivery of germplasm to stakeholders is managed through GRIN-Global, including customer information, germplasm use, and types of materials requested. Over the past 10 years, requests have significantly increased, particularly from individuals apparently not conducting research, breeding, or educational activities. Excessive requests hinder timely delivery and requires substantial input from PGRU staff. In response to increased public demand for germplasm, all requests are filtered through GRIN-Global administrators to restrict requests for personal use and target genuine research, conservation, commercial development, and genetic enhancement programs. PGRU provides germplasm openly to legitimate requestors and does not require Material Transfer Agreements for distribution. PGRU distributes germplasm in multiple forms including seeds, winter dormant cuttings, leaves, pollen, summer green cuttings, and DNA. Record keeping for order processing is maintained through GRIN-Global. Distribution of vegetable crops is directed towards research and crop improvement needs. The normal number of seed distributed is 50 seed per accession. Whenever seed is requested for an accession with low seed supply, it is given priority for regeneration. PGRU also extensively provides germplasm to researchers and extension personnel in fruit crop related projects including many cooperations within the NE9 region. 

2. Develop more effective germplasm maintenance, evaluation, and characterization methods and apply them to priority genetic resources. Record and disseminate evaluation and characterization data via the Germplasm Resources Information Network (GRIN-Global) and other data sources.

We will cooperate with scientists from ARS and other public and private sectors to characterize priority traits in vegetable, hemp, and fruit collections. PGRU will carry out the characterization and evaluation of key morphological, horticultural, genetic, and biochemical attributes of accessions. We will optimize the protocols and develop/adapt new methods for data collection for collections. Characterization and evaluation data are distributed via GRIN-Global and other databases. 

Local regenerations will continue to be the primary method of maintaining the genetic resource collections PGRU. The viable lifetime of regenerated seed varies between different crop species, ranging from a few years to ~50 years, and is a major factor in deciding which accessions get regenerated in a given year. Total regenerations will be increased substantially by adopting more efficient approaches to field regeneration via method development and application to optimize seed production. For example, running greater numbers of regenerations of fewer taxa per production cycle will enable efficient scaling and throughput. Regeneration planning and execution are being streamlined via prepopulated forms held in shared cloud storage. This approach mitigates numerous hand-off issues experienced between teams with phenotyping integration in FieldBook. Typically, 150 to 200 accessions are regenerated annually in-house or via collaborative increases.

High-throughput hemp germplasm regenerations will be a critical early stage of supporting NE9 efforts. Pollination must be extremely carefully controlled in hemp. Hemp, unlike other annual crops maintained at PGRU, is anemophilous (wind pollinated) and the anthers typically produce many very small (~ 25 μm) pollen grains. Prevention of genetic drift and contamination requires more resources to be invested in regeneration on a per accession basis compared to self-pollinated crops such as tomato. Of note, outdoor pollination is not possible due to pollen drift. Currently, hemp pollen drift is not well understood, but there is an expectation that pollen can travel up to five miles by wind. It is almost certain that contaminating pollen sources exist within this radius of PGRU’s outdoor vegetable seed production facilities. We will work closely with Cornell researchers to coordinate pollen control within any cannabinoid trials. Several methods have been developed for PGRU’s hemp production systems: isolating a single population within a controlled environment growth chamber, isolating a single population (> 25 plants) within a greenhouse, isolating a single population in hemp pollination cages, isolating individual inflorescences in pollination bags with the addition of pollen, and collaborator increases using the above methods. PGRU has three greenhouses with the capacity for regeneration of one population at a time, four shared 10 x 10’ controlled environment growth chambers, two 6 x 12’ customized pollination cages, and customized hemp pollination bags and is developing relationships with collaborators capable of preforming routine regenerations. Based on these current resources, we estimate that between 15 to 50 populations can be regenerated per year. Cultural practices will follow PGRU’s in-house controlled environment protocols to produce healthy, virus and pathogen-tested plants and seed. Fertility and pest control will be controlled in accordance with Cornell Cooperative Extension recommendations for indoor hemp production. PGRU is exploring distributing hemp pollen as an alternate or complementary approach to seed distribution. Specific questions regarding storage longevity, scaling, implementation, and other technical elements are currently being investigated. PGRU is developing large-scale capture and storage solutions for hemp pollen, including developing microfiltration vacuum systems, running pollen longevity experiments in 20, 4, -20, and -80 °C conditions, and evaluating viability using tetrazolium testing.  

In apple, fire blight can be present in asymptomatic tissues (Tancos et al. 2017) and carried over during propagation which later proliferates in regenerated tissues, impacting PGRU propagation, distribution, and potentially cryopreserved germplasm. We hypothesize that cryopreservation techniques will significantly reduce fire blight inoculum in apple propagules. Currently, it is uncertain whether E. amylovora can survive cryopreservation, although other microbes have been reported to survive cryotemperatures (Bajerski et al. 2020). To determine the impact of cryotreatments on fire blight-infected apple scions, dormant buds will be inoculated with fire blight (Bell and Van Der Zwet 1987) and processed following NLGRP protocols (Volk et al. 2020). Cryotreatments will occur early to mid-January in Geneva, NY, following at least three successive days at or below 0℃. Scions will be cut into 2 cm nodal sections and:

1. Desiccated to 25-30% moisture content at -5℃.
2. Frozen to -30℃ at the rate of 1℃/hour.
3. Exposed to liquid nitrogen vapor for at least 1 hour.

Sample subsets will be removed during each progressive step and stored at 5 °C prior to E. amylovora culturing, including lab inoculated and non-inoculated controls. Each treatment will include five biological replicate samples, each with 10 to 20 buds. Five buds from each treatment replication will be grafted to determine bud viability. Depending on results, second-year samples will expand to cover more genetic diversity, including apple cultivars and wild progenitors amenable to cryopreservation. ANOVA will be used to determine statistical variation between treatments. Linear mixed models will be used to determine the effects of species and genotype diversity assay in the second year. If successful, methods for dormant bud cryopreservation could develop into a SOP for regeneration of priority and diseased accessions or international germplasm distribution. In a pilot study, we observed up to a 96% reduction of fire blight colony forming units (CFUs) in ‘Gala’ samples exposed to all three cryopreservation stages, compared to untreated, inoculated samples, but additional sampling is needed to determine significance. 

Evaluation of fruit quality traits are of high importance to stakeholders to identify accessions with desirable or undesirable fruit qualities within the apple, grape, and tart cherry collections. All data collected, including genotypic and phenotypic data will be deposited in GRIN-Global under the respective crop descriptors page. For grapes, metabolite profiles will be developed for aroma, color, and flavor compounds in accessions from approximately 130 grape hybrids from PGRU and approximately 340 Vitis vinifera cultivar accessions from the NPGS NCGR-Davis germplasm collection of wine grapes and warm season adapted grapes. PGRU germplasm will be selected based on GRIN-Global descriptors, with an emphasis on V. labrusca hybrids with foxy aromas (Yang et al. 2020). NCGR-Davis accessions were selected through analyses of genetic marker data to represent the diversity of V. vinifera, and excludes clones and first-degree relatives (Myles et al. 2011) and include accessions previously evaluated for phenolic diversity (Liang et al. 2011). Surveying broad genetic variation is challenging because each accession matures at a different time; both sites contend with personnel constraints for repeated BRIX measurements to determine maturity. Our strategy will be to collect samples of approximately 200 grams of berries from each of the selected accessions, from at least four clusters across each vine, at the end of the season, during which most of the selected accessions have reached maturity. Sampling will repeated be over three years. 

In apple, the following traits related to fruit quality will be evaluated: fruit size, fruit weight, soluble solids concentrations, tannin content, fruit texture, titratable acidity and pH, and juice volume (Kumar et al. 2021). Additionally, digital images of fruit at maturity will be collected. Fruit maturity will be determined using the cortex starch pattern index (Blanpied and Silsby 1992). Major fruit sweetness and acidity genes will be genotyped (Zhen et al. 2018; Kumar et al. 2021). Sets of 300 accessions will be evaluated for a two-year period for fruit quality traits, evaluating up to 900 accessions over the five-year period, representing about 63% (900/1,432) of the M. domestica cultivars in the collection. Sampling strategies will include biological replicates where possible, including repeated sets of commercial standards in each subset to make statistical inferences of the differences among the cultivars. 

Phenological characterization of perennial germplasm is critical for determining suitability of cultivars for an environment and identifying targets to breed for climate adaptations. Phenology studies the relationship between seasonal climate changes and the timing of biological events, including loss of dormancy leading to budbreak, onset of flowering, and fruit ripening. Phenological events are driven by environmental conditions, genetic factors, and the interaction of the two. Certain events, such as bloom in grapevine, may have a significant genetic component making it a target for genetic enhancement (Gutierrez et al. 2021). Timing of budbreak and flowering is particularly important due to the threat of frost damage resulting in crop loss. Acclimation and de-acclimation of cold hardiness are related to dormancy and budbreak, a key trait that limits crop production in certain areas where climate-adapted cultivars are not available. 

Cold-hardiness and bloom phenology are intrinsically connected to productivity as late-season frosts are among the leading reasons for cherry crop failure in the US. Acclimation and de-acclimation of cold-hardiness will be measured in the cherry collection. Weekly fine-scale monitoring of bud sensitivity to seasonal temperature changes will be completed using four species Prunus cerasus (‘Montmorency’), P. avium (‘Black Gold’) and P. fruticosa (‘Dwarfrich’), and the early-blooming Japanese species P. nipponica. A larger set (64) representing the diversity of the collection will be evaluated each month. Initial cold-hardiness will be determined at each timepoint, and the remaining samples will be maintained at 20°C and measured in 3 to 7-day intervals for up to four weeks to determine cold deacclimation; the interval of deacclimation testing decreases as accessions lose dormancy towards spring. Sampling will begin following the first frost (typically October) and repeated each week until budbreak (late March or early April), across three consecutive seasons. Budbreak and bloom phenology will be scored weekly in the field using the Biologische Bundesanstalt, Bundessortenamt und Chemische Industrie (BBCH) scale for cherries (Fadón et al. 2015). Cumulative growing degree days (GDD) will be determined for Prunus and analyzed following approaches described in grape germplasm to determine heritability and species effects (Gutierrez et al. 2021). 

For the cold-hardy Vitis collection, a test of bud hardiness is a critical missing descriptor from the collection. Previous research studies have evaluated 61 of the accessions for low temperature exotherm (LTE) (Londo and Kovaleski 2017; Londo and Kovaleski 2019), 43 of which were one species, Vitis riparia. Current evaluations of the Vitis core collection were successful, which includes 90 accessions (15 species). Testing will increase to 330 accessions per year to complete bud testing of the full germplasm collection within five years. The sampling frequency will be once a month from December through February. Each year, the evaluations will include cultivars ‘Concord’, ‘Riesling’, and ‘Merlot’, key industry standards in cold climate regions, as hardiness controls, for between-freezer and between-year controls. Nine buds per accession will be collected and processed at time of analysis. 

Differential thermal analysis (DTA) will be used to determine the LTE or lethality point for Prunus and Vitis germplasm. LTE is measured as the release of heat during ice formation when temperatures exceed the supercooling point, indicating the bud lethality point. Bud LTE will be assayed on thermoelectric modules in a programmable freezer from 4℃ to -40℃ with a rate of 3.4℃/hour. Methods in grapevine are described by Londo and Kovaleski (2017) and will be used here for grape and cherry testing. Detailed weather data within 5 km of the evaluation site is available at Network for Environment and Weather Applications (www.newa.cornell.edu). Data analysis includes analysis of covariance to determine significant differences between accessions LTE curves. Significant explanatory variables were determined previously in grapevine germplasm and will be reevaluated in these studies (Londo and Kovaleski 2017). These variables include species and a linear and quadratic function of the time covariate, and a statistically determined temperature index to correct for temperature shifts before sample collection. 

3. With other NPGS gene banks and Crop Germplasm Committees (CGCs) develop, update, document, and implement best management practices and Crop Vulnerability Statements (CVSs) for priority fruit and vegetable genetic resources and information management.

The knowledge, expertise, and experience of CGC members, and staff at other NPGS and international germplasm repositories will be leveraged to strengthen and improve germplasm conservation through best management practices (Rao, Dulloo et al. 2016). Other NPGS sites with shared taxa include NC7 (Brassica, Cucurbita), S9 (Cucurbita), and W6 (Allium, Grape, Prunus). Curators and other scientists meet on a regular basis at scientific conferences, CGC meetings, Regional Technical Advisory Committee meetings, and Plant Germplasm Operations Committee meetings. This provides many opportunities for mutually beneficial consultation, exchange of information, formulation of new ideas, and soliciting recommendations. All components of PGRU operations will be reviewed and documented as SOPs with sufficient detail to reduce risk of any lapse in operations. Thereafter, the finalized PGRU Operations Manual will be reviewed and updated annually. 

4. Develop novel germplasm that integrates diverse, useful genes from various resources and breed, release, maintain, and evaluate improved germplasm and cultivars. 

This objective is primarily met through collaborations among members of the SAESs of the Northeast (Appendix E) and the PGRU and other ARS scientists (Appendix F). 

We are working with breeders in the vegetables through collaborative characterizations/evaluations to provide the germplasm necessary for improvement of disease and insect resistance. Additionally, we provide germplasm to cooperators to improve traits such pathogen, drought, and salt cold tolerance resistance in tomato, locally adapted germplasm to southern Africa, and novel cucurbit germplasm to support historically underserved communities in the Northeastern US. 

Hemp germplasm is being evaluated at multiple testing locations including Geneva, NY and in collaborator nurseries in Normal, AL, Madison, WI, Corvallis, OR, Pullman, WA and Davis, CA. The same group of accessions will be tested at these locations for a least two years. Multiple environment testing will support development of unique hemp germplasm through routine total floral THC and CBD testing, dissemination of pertinent phenotypic data, and through characterization and elucidation of priority traits. Planting design will be consistent across locations with accessions replicated across three blocks in a RCBD. High quality photos of the plants growing in the field, stems, and seeds will be deposited in GRIN-Global. Genotype-by-environment effects and interactions will be calculated for each measured trait. Traits with high broad-sense heritability values will be selected as breeding targets. Accessions will be selected for hybridations based on trait performance within and across locations. 

Through discussions with researchers, breeders, and CGCs, we continue to update phenotyping and genotyping efforts of the PGRU fruit collections to provide information useful for crop improvement and cultivar development. In collaboration with the GGRU, Cornell University, Appalachian Fruit Research Station (ARS) in West Virginia, Washington State University, University of Minnesota, and other institutions in the USA and worldwide, we will continue to identify new sources of disease resistance and other useful traits. Where possible, pre-breeding populations will be developed to advance stakeholder objectives.

Measurement of Progress and Results

Outputs

• Identified priority germplasm to enhance collections and addition of new accessions with targeted characteristics.
• Improved field conditions for better maintenance and improved regeneration protocols
• Germplasm security, viability and availability secured through germination testing and back-ups.
• Collected data for growth, morphology, phenology, and production of priority accessions.
• An expanding GRIN-Global database with passport, characterization, and evaluation data including digital images.
• Organized germplasm and increased efficiency of filling orders allowing timely distribution of germplasm and associated information.
• Improved usability of germplasm for genetic research and cultivar development.

Outcomes or Projected Impacts

• Gaps in PGRU collections filled.
• Optimized regeneration protocols and cultivation conditions.
• Healthy germplasm with genetic integrity.
• Optimized characterization and documentation methods.
• Publicly available data and images for accessions.
• Updated PGRU Operations Manual and updated Crop Vulnerability Statements.
• Improved germplasm and cultivars.

Milestones

(2023):• Regenerate seed for 150 vegetable accessions. • Backup up 100 vegetable and hemp accessions and fully backup tomato core collection in NLGRP. • Improve barcoding and tracking of regeneration to manage vegetable crop regeneration efforts more effectively in real-time. • Rescue 50 jeopardized vegetable accessions in greenhouse regeneration. • Consult with plant pathologists to identify chronic diseases during seed regenerations, identify winter squash accessions with no seed set or production of low viability seed. • Apply FieldBook application to increase throughput of vegetable and hemp phenotyping efforts. • Develop effective tissue-culture protocols for hemp genetic resource conservation. • Regenerate 25 hemp accessions in controlled-environment conditions. • Upload 10,000 phenotypic data points to describe hemp germplasm collection. • Acquire over 20 novel hemp genetic resources. • Backup clonal accessions and test viability of cryo treated buds. • Collect descriptor data and digital images of clonal accessions. • Determine protocols and treatments for fire blight cryotherapy of apple. • Evaluate genetic data currently available for apple and grape to determine genetic gaps. • Evaluate fruit quality traits of apple and grape. • Improve maintenance and grapevine quality in the vineyard via trellis system changes. • Evaluate abiotic and biotic resistances in Vitis accessions. • Consult with CGC members to prioritize traits for evaluation of collections. • Draft outline for PGRU Operations Manual. Participate in CGC meetings and consult with members on status and plans for CVS.

(2024):• Regenerate seed for 150 vegetable accessions. • Backup up 100 vegetable and hemp accessions at NLGRP. • Rescue 50 jeopardized vegetable accessions in greenhouse regeneration. • Apply cucurbit phenotyping protocols developed with Cornell collaborators to critical genetic resources. • Apply treatments to reduce chronic diseases during seed regenerations, consult with experts for recommendations on regenerating winter squash accessions with no seed set or production of low viability seed. • Develop pre-breeding hemp populations for priority traits. • Regenerate 25 hemp accessions in controlled-environment conditions. • Upload 10,000 phenotypic data points to describe hemp germplasm collection. • Acquire over 20 novel hemp genetic resources. • Backup clonal accessions and test viability of cryopreserved buds if resources are available. • Evaluate 300 of 900 (set 1) apple cultivars for fruit quality. Year 1 evaluation of 470 accessions for grape juice metabolites. • Cryotherapy technique applied to diverse set of apple germplasm. • Identify apple germplasm and suitable rootstocks for the orchard system trial. Establish an apple nursery for rootstock evaluation. Retrain vines from the UK to TWC system. Identify accessions that could grow better on a T-trellis system. • Evaluate abiotic and biotic resistances in Vitis accessions. • Identify sources to fill genetic gaps through exchanges and explorations. • Compile and edit content of Operations Manual and contribute to updated CVS.

(2025):• Regenerate seed for 150 vegetable accessions. • Backup up 100 vegetable and hemp accessions at NLGRP. • Rescue 50 jeopardized accession in greenhouse regeneration. • Continue to refine regeneration protocols of seed crops. • Improve soil and assess onion bulb production. • Develop parents for hemp MAGIC populations with collaborators. • Regenerate 25 hemp accessions in controlled-environment conditions. • Upload 10,000 phenotypic data points to describe hemp germplasm collection. • Acquire over 20 novel hemp genetic resources. • Backup clonal accessions and test viability of cryo treated buds. • Evaluate apple nursery for rootstock compatibility. • Re-evaluate set 1 apple cultivars for fruit quality. Re-evaluate 470 accessions for grape juice metabolites (year 2). • Evaluate abiotic and biotic resistances in Vitis accessions. • Incorporate cryotherapy treatments, if successful, into standard practices. • Fill genetic gaps through exchanges and explorations. • Compile and edit content of Operations Manual, check for deficiencies in information and finalize content.

(2026):• Regenerate seed for 150 vegetable accessions. • Backup up 100 vegetable and hemp accessions at NLGRP. • Rescue 50 jeopardized accession in greenhouse regeneration. • Continue to refine regeneration protocols of seed crops. • Map critical genes underpinning priority traits in hemp. • Regenerate 25 hemp accessions in controlled-environment conditions. • Upload 10,000 phenotypic data points to describe hemp germplasm collection. • Acquire over 20 novel hemp genetic resources. • Backup clonal accessions and test viability of cryo treated buds. • Plant apple nursery trees in high-density blocks. • Evaluate new set (set 2) of 300 apple cultivars for fruit quality. Re-evaluate 470 accessions for grape juice metabolites (year 3). • Evaluate abiotic and biotic resistances in Vitis accessions. • Fill genetic gaps through exchanges and explorations.

(2027):• Regenerate seed for 150 vegetable accessions. • Backup up 100 vegetable and hemp accessions at NLGRP. • Rescue 50 jeopardized accession in greenhouse regeneration. • Begin development of hemp MAGIC populations with collaborators. • Regenerate 25 hemp accessions in controlled-environment conditions. • Upload 10,000 phenotypic data points to describe hemp germplasm collection. • Acquire over 20 novel hemp genetic resources. • Continue to refine regeneration protocols of seed crops. • Backup seed and clonal accessions and test viability of cryo treated buds. • Re-evaluate set 2 apple cultivars for fruit quality. Summarize grape evaluation. • Update the CVSs.

Projected Participation

View Appendix E: Participation

Outreach Plan

Improved documentation and utilization of PGRU germplasm will better support breeding and research objectives, and subsequently benefit consumers in the US and abroad with stable production, nutritionally enhanced crops with reduced risk of susceptibility to pests, diseases, and changing environments. Customers looking for sources of rare or exotic fruit or vegetable varieties, for example, small growers serving local niche markets, are also frequent requestors and recipients of our germplasm. This project provides a secured supply of germplasm, ensuring the availability of genetic diversity required by US fruit, hemp, and vegetable producers to remain successful and competitive in global markets. 

Primary stakeholder groups include international horticultural communities, including farmers; government and academic scientists including plant pathologists, physiologists, food scientists, entomologists; and scientists at private companies are major users of the germplasm and data from this project. Outreach objectives include publication of germplasm evaluation data in peer-reviewed journals; presentations at academic and industry meetings; and deposition of data into GRIN. To better engage stakeholders, we host several tours of our collections, including an annual open house to inform the general public about our program. Additionally, we engage with Cornell AgriTech and contribute to their outreach events aimed at serving underrepresented populations. Educational tours serve various groups including: primary, secondary, college and graduate students, growers, commodity groups, researchers, and botanical gardens, arboreta, and garden clubs. Additionally, PGRU hosts several undergraduate interns each year in an effort to train the next generation of germplasm researchers https://www.ars.usda.gov/northeast-area/geneva-ny/plant-genetic-resources-unit-pgru/docs/intern-corner/).

Organization/Governance

Regional Research Project NE9 can be effective only through federal, state, and private cooperation. The federal agency ARS, through acquisition, maintenance, characterization, documentation, and distribution activities, will make plant genetic resources available for evaluation and utilization research. ARS will provide support, staff, facilities, equipment, and specialized technical assistance at both the regional and national levels. The SAESs provide facilities, additional support staff, equipment, utilities, and local assistance. The NE9 Regional Technical Advisory Committee (RTAC) will provide technical guidance in this effort. This committee is composed of an Administrative Advisor, Regional Coordinator, plus technical representatives invited to participate from each of the Northeastern SAESs plus the District of Columbia. ARS representatives from the National Program Staff, the National Germplasm Resources Laboratory, and the PAGRP are also included on the committee as ex officio members. The names, affiliations, and areas of specialization of these individuals are presented in Appendices E and F. This committee has annual meetings with the PGRU staff at locations throughout the NE9 region which provides yearly review of genetic resources research in the region and provides technical advice to PGRU scientists.

Other committees contribute to the planning and management and are active participants in the NPGS. These include:

1. The ARS Plant Germplasm Operations Committee (PGOC) evaluates and recommends foreign/domestic exploration proposals, and assists the NPGS, ARS National Program Staff and other officials with plans needed to manage the NPGS.
2. CGCs have been established for 42 crops (or crop groups) to help advise the NPGS with regard to genetic vulnerability, gaps in current collections, operational procedures, evaluation needs, and current enhancement and utilization research associated with their specific commodity. 

Project scientists have monthly meetings to discuss progress in meeting milestones and to modify activities in order to obtain goals. Scientists have regular meetings with support staff to ensure all activities are coordinated and directed towards project milestones. Scientists within the USDA-ARS Geneva, NY Location meet on a regular basis to discuss new technologies and methodologies and their potential application to project research. An annual Plant Germplasm Operations Committee (PGOC) meeting provides a forum for NPGS scientists to discuss germplasm issues with each other and National Program leaders. Shared network folders are used for storage and use of common files, both documents and data. Our major source of stakeholder feedback is from the Crop Germplasm Committee (CGCs) members who meet annually to discuss germplasm user needs and concerns pertaining to the various crop collections. We also regularly discuss with our collaborators the research and regeneration activities through emails, conference calls, discussions at scientific meetings, and site visits. Progress towards meeting project milestones is reported annually in the project annual report (AD 421). Any changes in activities necessitated by unforeseen circumstances that will affect progress towards meeting project milestones are also documented in the project annual report.

Literature Cited

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