Research Project: Plant Genetic Resource Management and Information System Development

Location: Plant Introduction Research

2021 Annual Report

Objectives
Objective 1: Efficiently and effectively acquire and maintain the safety, genetic integrity, health, and viability of priority maize, vegetable, oilseed, minor grain crop, medicinal, and ornamental genetic resources, and distribute them and associated information worldwide. Sub-objectives: 1A. Expand, regenerate and conserve collections of priority maize, vegetable, oilseed, minor grain crop, medicinal, and ornamental genetic resources. 1B. Enhance genetic integrity of genetic resources maintained. 1C. Backup accessions at second sites, increasing the proportion of accessions backed up to more than 83% of 2016 holdings. 1D. Distribute germplasm and associated information to support research objectives. 1E. Monitor accessions for viability and phytosanitary health to ensure availability of healthy propagules and to preserve their genetic integrity. 1F. Develop and/or refine diagnostic methods to detect seed-borne pathogens. Objective 2: Develop more effective genetic resource maintenance, evaluation, or characterization methods and apply them to maize, vegetable, oilseed, minor grain crop, medicinal, and ornamental genetic resources. Record and disseminate evaluation, characterization, and adaptation data via GRIN-Global (GG) and other data sources. Sub-objectives: 2A. Enhance capacities to increase collection availability. 2B. Evaluate crop collections for phenotypic, morphological, composition, and productivity related traits such as biotic/abiotic stress resistance and yield. 2C. Apply and utilize genetic marker technologies to better characterize key collections. 2D. Provide information and findings that facilitate germplasm utilization via GG, other data sources, and publications. Objective 3: With other National Plant Germplasm System (NPGS) genebanks and Crop Germplasm Committees (CGCs), develop, update, document, and implement best management practices and Crop Vulnerability Statements (CVSs) for priority maize, vegetable, oilseed, minor grain crop, medicinal, and ornamental genetic resources and information management. Sub-objectives: 3A. Modify, document and implement best management practices, and share them publicly. 3B. Provide input for annual/periodic updates of CVSs. Objective 4: Continue to expand the capacity and capabilities of the GRIN-Global (GG) plant genetic resource (PGR) information management system to meet the needs of crop curators and genetic resource users, and to ensure smooth integration of its data with information from other sources, such as model organism databases (MODs). Sub-objectives: 4A. Cooperate with GG users and developer communities to identify and develop wizards and applications to improve system utility, information delivery, and genebank workflow efficiency. 4B. Provide technical leadership to the NPGS for GG evolution to meet genebank information and workflow needs.

Approach
Obj 1: Plant genetic resource (PGR) regeneration/management priorities are determined in response to PGR demand, external pressures such as stakeholder needs, or the status of resources, i.e. threatened or endangered, and inventory quantity, viability, and quality status. Genetic integrity will be assured through use of best management practices and monitoring to detect contamination. Seed lots will be backed up at the NCGRP in Ft. Collins, CO, and an international seed vault. Periodic testing to assess collection quality and health will assure their security, and new protocols will be developed. Quality PGR and associated information are distributed to support research objectives based on criteria which may govern their distribution or use. Seeds/plantings will be monitored for disease and assayed for pathogen identity, with phytosanitary precautions implemented at several points to preclude seedborne pathogens. New methods for detection of seed-borne pathogens will be evaluated and implemented. Obj 2: Collaborations with other NPGS site personnel or tropical nursery providers are required to grow certain accessions where they are adapted, including facilities that support quarantine grow-outs of maize originating from some countries. In-house facilities will be improved to handle accessions with challenging growth requirements. Phenology and basic morphological descriptors are captured during seasonal activities. High priority evaluation traits will be determined. Woody ornamental germplasm will be evaluated via the NC7 Ornamental Trial system to determine adaptation and survival across the North Central US. Evaluations will be conducted using established protocols, and findings published on the GRIN-Global website or suitable fora. Genetic marker technologies will be utilized to assess maize accession relationships. Sunflower genetic marker information will be assessed to determine whether valid associations can be made with habitat/geographic information, and used to determine how well crop wild relatives are represented in the collection. Obj 3: Best management practices for collection management and standard operating procedures that govern genebank workflows will be reviewed, modified, and documented periodically. Crop Germplasm Committee chairs and members will be provided information on the status of the collections and specific issues that threaten the security and/or availability and backup status of the collections, and opportunities to mitigate threats. Personnel will assist in development of Crop Vulnerability Statements. Obj 4: The capabilities of the GRIN-Global PGR information management system will be expanded to meet crop curator and genetic resource user needs. Wizards and applications to improve system utility, information delivery, and genebank workflow efficiency will be identified and developed. Improvements necessary to support system adoption will be made. Use of improved communications technologies will be facilitated. Interoperability of GRIN-Global with multiple information providers will be supported.

Progress Report
Obj 1 Related: New germplasm acquisitions (841) included transfer of 46 accessions from the National Laboratory for Genetic Resources Preservation (NLGRP) in Ft. Collins, Colorado of many of the species we curate, as well as from exploration and transfer. These include a group 196 accessions of Chenopodium from Brigham Young University, 46 accessions from the Germplasm Enhancement of Maize (GEM) Project, 57 medicinal, herbaceous, and woody landscape accessions, maize accessions with expired Plant Variety Protection certificates, and cultivated sunflower accessions. The standard germination testing using 200 seed samples to monitor condition and viability of 2,310 accessions was conducted in 2020. Germination tests were also conducted on 5,464 accessions using reduced seed amounts of 20-100 seeds to decrease the backlog of accessions needing to be tested and to help save germplasm and other resources. Lower seed quantity germination samples that were determined to be below species specific collection guidelines were submitted for additional testing at the standard 200 seed sample size. Accessions found to be of poor germination or viability condition were flagged for regeneration. Regeneration plans must address accessions with declining viability. Viability was tested for collection holdings, focused on Brassica, Crucifers, maize, Cucurbita, and Amaranthus. Regeneration of portions of the Brassica collection and the flax collection is needed due to declining viability. Pandemic restrictions and concerns about availability of labor in summer 2020 delayed regeneration plans for accessions of several crops until 2021. Accessions identified with declining viability are placed in the crop priority regeneration list for 2022. ARS personnel in Parlier, California supported regeneration of specific wild Helianthus taxa that need a longer growing season than Ames, Iowa can provide, and mountain and desert species that do not thrive in midwestern humidity and heavy soils. ARS personnel in Salinas, California provided support for spinach seed increases, which require use of positive pressure growth chambers. Private sector collaborators provided support for regeneration of Daucus relative and maize. Obj 2 Related: Beneficial insects are used both in the field cages and greenhouse rooms to control pest insects. These include rove beetles, ladybug beetles, green lacewings, beneficial nematodes, and Encarsia formosa, a beneficial wasp. Our entomology team provided six species of pollinator insects to pollinate 809 accessions which require insect pollination for reproduction. These include honeybees, bumblebees, Osmia (mason) bees, Alfalfa Leafcutter bees, and two species of flies. Field operations and equipment changes have enhanced productivity and reduced labor costs. Establishment of "permanent" cage fields where cage frames remain year-round have reduced labor costs associated with field griding, measuring, installing, and removing cage frames each year. Specialized equipment is now used to prepare soil within the cage area which allows establishment of turf alleys between cages. The change allows for reduced soil compaction and erosion, improved weed control, and better access for accession harvest. Focus was placed on uploading of images to GRIN-Global (GG) for genebank accessions. Image mass-loading continued in 2020 to enhance collection characterization. Over 1,730 images of 1,028 accessions and 36,697 characterization datapoints were loaded into GG. The images and characterization data are important for researchers who search the public GG database to verify and/or select materials for research based on visual seed or plant images, as well as phenotypic and genetic information. Crop accession evaluation information (milestone 2B) was provided for 7,379 accessions, 84% of which was associated with maize inbreds, populations, and expired Plant Variety Protected (PVP) inbred lines. The backlog of accession images awaiting release was reduced with 4,906 images of 2,844 accessions provided. Passport information was updated with 167 new documents filed as well as 34 Re-Identification requests forms. All 902 legacy PI number assignments forms were uploaded documenting requests dating back to 1997. Soil sampling protocols were fully implemented for all station fields. This information will facilitate actions to improve soil fertility and enhance field management. No new pathogens were noted in Ames, Iowa in scouting of field and greenhouse seed increases. Effort to identify methods to reduce seedborne Acidovorax citrulli, the causal agent of bacterial fruit blotch, in Cucumis melo seeds focused on dry heat and antibiotic seed treatment. Heat treatment of seeds degraded germination of the seeds but failed to eliminate the pathogen. Research into use of antibiotics is ongoing but shows promise. Antibiotic seed treatment trials with 12 different compounds showed pathogen sensitivity to Doxycycline, Trimethoprim, Sulfamethoxazole, and Ceftazidime. Further investigation coupled with use of ELISA tests is required to verify whether bacteria are consistently detected in seeds. Field and greenhouse plantings of 879 seed increases were inspected for plant pathogens. No diseases of phytosanitary concern were observed Seed health testing and treatment programs ensure seed health and support utilization. In addition, nearly 3,900 plots were inspected for the GEM breeding project. Additional declarations were written in support of phytosanitary certificates for 96 orders. Stem lesions characteristic of anthracnose disease were observed on plants of two quinoa accessions PI 510547 and PI 596293 grown in the field during summer 2019. Fungi were isolated from both accessions and isolates examined morphologically. Sequencing and phylogenetic analyses based on five loci (ACT, GAPDH, CHS-1, TUB2 and ITS) identified the isolates as Colletotrichum nigrum and Colletotrichum truncatum. Koch’s postulates were used to confirm that both strains were pathogenic in quinoa. The postulates are as follows: 1) the microorganism must be found in diseased but not healthy individuals, 2) the microorganism must be cultured from the diseased individual, 3) inoculation of a healthy individuals, and 4) the microorganism must be reisolated from the inoculated, diseased individual and identified as identical to the original causative agent. This study is the first report of quinoa anthracnose caused by Colletotrichum nigrum and C. truncatum in the United States. Obj 3 Related: Crop Vulnerability Statement update was completed for the Proso Millet Crop Germplasm Committee, in collaboration with the members. Documentation of best management practices and standard operating protocols are modified frequently. Virus testing protocols were updated. Plant/seed protection practices were reviewed and re-evaluated. A pilot study was conducted with three sunflower accessions to determine if low germination is the result of seed deterioration or the presence of seed-borne pathogens. Results showed no difference in the microflora present on seed lots with low or high germination. Whereas the electrical conductivity was significantly higher in the seed lots with low germination indicating electrolyte leakage and possible seed deterioration from membrane damage. Obj 4 Related: Protocols were developed to detail how applications can appropriately be shared and vetted between U.S. and international GRIN-Global (GG) collaborating institutions. This will advance evolution of the system, facilitate genebank workflows and information objectives, and extend the lifecycle of the system. Following security code scanning, the Office of the Chief Information Officer approves all new versions for release. New Order, Cooperator and Viability wizard versions of GG were completed in response to evolving genebank personnel users desired enhancements. Curator Tool (CT) build version 1.9.9.9 was released. GG interoperability with the Maize Genetics and Genomics Database (MaizeGDB), Soybean Genetics and Genomics Database (SoyBase), Peanut Genetics and Genomics Database (PeanutBase), and Legume Information System (LIS) information providers was inspected; routine information updating is currently limited to ETL (extract, transform and load) processing. To address this, a software prototype using a RESTFUL interface has been developed to facilitate pulling data from GG to use in conjunction with data from other providers. Ames, Iowa ARS personnel participated in virtual weekly development team meetings with ARS personnel in Beltsville, Maryland, twice monthly with the GG Advisory Committee to work on priorities for developers and the database administrator, and monthly with the International Developer's Advisory Committee.

Accomplishments
1. Distribution of plant genetic resources for breeding and genetic research. Plant Genetic Resources (germplasm) were distributed for research and educational objectives from Ames researchers in Ames, Iowa. In 2020, more than 800 public and private sector researchers received 46,627 packets of seeds of 21, 089 accessions. Domestic recipients (70% of requestors) received 45% of the packets, and 55% of the packets were distributed to international recipients (30% of requestors). Demand remains strong for our collections of maize, vegetables, oilseeds (Brassica, crucifers, and sunflower), woody and herbaceous ornamentals, and miscellaneous crops such as quinoa, amaranth, spinach, panicum, and several others. These resources are utilized in plant breedings programs to enhance agricultural productivity and to expand our understanding of genetic diversity and inherent value of the germplasm.

2. Development of a seed increase protocol for Teosinte, a tropical wild relative of corn. Tropical corn wild relative accessions such as teosinte cannot be easily increased in temperate climate areas. ARS scientists in Ames, Iowa, collaborated with scientists at Texas A&M University to develop seed increase protocols utilizing available greenhouse resources. The process was documented and evaluated at each step from seed preparation, through germination, plant management, and seed harvest to optimize resource utilization, seed quality, and maintain accession purity. The developed system has expanded regeneration capacity and increase accession availability. Expanded availability of these accessions will enable and encourage research on these wild relatives and increase our understanding of the evolution of modern corn and its genetics.

Review Publications
Bernau, V.M., Jardón Barbolla, L., Mchale, L.K., Mercer, K.L. 2020. Germination response of diverse wild and landrace chile peppers (Capsicum spp.) under drought stress simulated with polyethylene glycol. PLoS ONE. 15(11). Article e0236001. https://doi.org/10.1371/journal.pone.0236001.
Pal, N., Testen, A.L. 2021. First report of quinoa anthracnose caused by Colletotrichum nigrum and C. truncatum in the United States. Plant Disease. 105(3):705. https://doi.org/10.1094/PDIS-07-20-1568-PDN.