2005 Annual Report 1.What major problem or issue is being resolved and how are you resolving it (summarize project aims and objectives)? How serious is the problem? What does it matter? The primary goals for this project, the North Central Regional Plant Introduction Station (NCRPIS) or PIRU are to. 1) acquire, document, conserve and make available for distribution a broad spectrum of plant genetic resources (PGR), also known as plant germplasm, and associated information and to encourage its use in research and crop improvement;. 2)characterize and evaluate PGR;. 3)elucidate plant-environment interactions, emphasizing those key for invasiveness;. 4)transfer technology in the form of genetic resources and associated information to researchers and breeders worldwide. Conservation, management and utilization of PGR form the means to preserve genetic diversity, harness its potential, and strengthen agricultural production systems and their sustainability. Crop improvement depends on utilizing genes found in germplasm collections, such as those of the NCRPIS. The function of a PGR collection is analogous to that of a library; a researcher who ‘borrows’ its resources uses the material to develop and provide solutions for a wide array of basic plant research objectives, dietary and nutritional needs, biotic and abiotic production issues, phytoremediation and rehabilitation of disturbed environments. The library is as useful as the breadth and quality of its holdings, the information association with the holdings, and their organization and presentation. As more research is completed and knowledge transferred, the value of the collections increases; investigators build upon previous discoveries. Genetic diversity and the PGR in which it is encoded provides the raw material upon which genetic improvement is based, and allows us options to develop new crops and new products which create value in either a traditional or entrepreneurial sense, new uses for existing crops, to develop varieties with enhanced compositional profiles to serve nutritional or industrial applications, and genetic solution for issues which threaten production. The centers of origin for many of the food and fiber species, upon which the US depends, continue to suffer severe environmental damage from a variety of man-made and natural causes. Much of the germplasm residing in our collections is no longer collectible; many are difficult to regenerate and maintain. As world population increases, the pressures on sustainable, increasing global food production increase, and thus the importance of the role of germplasm conservation centers (such as the NCRPIS) as a primary source of potential genetic solutions increases. Reliable and timely access to PGR to support our objects is critical. This is reflected by continuing high demand for our collections, and our ability to track and assess the impact our germplasm plays in advancing research and product development. The US has developed the most extensive national system for the preservation of agricultural biodiversity in the world. Because of its stability, it is able to consistently serve international germplasm resource needs better than any other nation. Returns on the investments made in our National Plant Germplasm System (NPGS) are maximized when the collections are actively, carefully, and systematically documented and managed. Collection managers work closely with a dynamic community of PGR users to ensure that we have high quality germplasm and associated information to serve their current and future needs. Germplasm must be acquired via legitimate access channels and propagated via best available methods to preserve the genetic profile. Maintaining these materials in a viable condition true to their original genetic profile is a key objective. Understanding how to do this effectively and efficiently is crucial to managing available resources. The diversity of appropriate methodology reflects the diversity of the collections and their unique properties and issues. The germplasm maintained by the NCRPIS generally consists of outcrossing, heterogeneous, heterozygous crops and their wild relatives. Our collections include maize and its wild relative, teosinte; oilseed crops such as sunflower, oilseed Brassica (canola) and miscellaneous crucifers, wild and domestic flax; vegetables such as cantaloupe, cucumber, pumpkin, squash, umbels such as carrot, dill, coriander; amaranth, quinoa, and warm-season grasses; medicinal/nutriceutical plants such as Echinacea, Hypericum (St. John’s wort) and black cohosh; and many woody and herbaceous ornamentals. Activities supporting our objectives and PGR users’ needs include: acquisition of both foreign and domestic germplasm; maintenance of germplasm for the medium term under controlled conditions at NCRPIS with backup for long-term security at the National Center for Germplasm Resource Preservation (NCGRP) at Ft. Collins, Colorado; monitoring of germplasm health and viability in storage, and development of appropriate storage regimens which support germplasm and genetic diversity preservation objectives; characterization of germplasm by assaying highly heritable morphological, phenotypic and molecular traits; evaluation of germplasm to provide data to facilitate well-targeted utilization and enhancement programs for traits such as adaptation, host plant resistance, identification of genes and/or gene blocks via molecular methods, high-value traits, and yield. Collaborations with researchers whose goals are to enhance existing crops, develop viable new crops systems, and to better understand the factors that determine adaptation or invasive potential are ongoing. 2.List the milestones (indicators of progress) from your Project Plan. OBJECTIVE 1 : Acquisition and conservation of genetic resources and associated information for a broad spectrum of heterozygous, heterogeneous, generally outcrossing crops. MILESTONES AND EXPECTED OUTCOMES: Based on projected growth, the size of the collections in 2007 is estimated to be 50,000 accessions. An achievable target for the end of 2007, given existing resources and assuming continual application of useful technologies, would be 75% collection availability and 80% of the collection backed up at the NCGRP. OBJECTIVE 2: Characterization and Evaluation of Plant Genetic Resources. Genetically characterize the preceding PGR, and evaluate them for useful traits. MILESTONES AND EXPECTED OUTCOMES: Increased information resulting from continued characterization and evaluation of the collections. Currently, 20,000+ observation points are added to the GRIN database annually; the nature of distributions reflects availability of new information. Making these data available through GRIN also supports Objective 4. A reasonable expectation would be to maintain at least this rate of increase in datapoints to GRIN for the duration of the project. OBJECTIVE 3: Elucidate plant-environment interactions, emphasizing those key for invasiveness. MILESTONES AND EXPECTED OUTCOMES: New research findings are expected to illuminate the nature of plant adaptation processes, and provide useful guidance as to the worthiness and appropriateness of new plant introductions under a wide range of environmental conditions. Understanding of the factors affecting invasive potential also supports Objectives 1 and 2. A better understanding of patterns of genetic diversity in relation to the control of life cycle and the timing of flowering can be applied to the development of more efficient regeneration protocols in support of Objective 1. We expect such improvements to be implemented by 2007, based on the work of Cruz and the Buckler-Goodman collaborations. OBJECTIVE 4: Transfer technology in the form of genetic resources and associated information to researchers and breeders world wide. MILESTONES AND EXPECTED OUTCOMES: An average annual distribution of 10,000 accessions is expected; the makeup of these distributions fluctuates with production issues, new crop opportunities, and progress in research and development. Analysis of past distributions and current demand will be used to improve customer service and create more efficient regeneration and seed handling systems. 4a.What was the single most significant accomplishment this past year? Successful regeneration of 1363 accessions, deposit of 615 samples for backup storage at the NCGRP, resulting in 76% of our collection currently backed up in long-term storage; distribution of 17,983 seed packets of 10,481 accessions; collection and transfer of over 18,600 information items to the GRIN database relating to flowering, adaptation, quality traits, etc. This is important to accomplish our mission of conserving and transferring germplasm and associated information to the research public. Alfalfa leaf cutter bee use was investigated for improving efficiency of regeneration for some key species; cost-effective, non-stinging pollinators which demonstrate excellent pollinating efficacy with NCRPIS species are needed to facilitate our objectives. Our Pathologist and Vegetable Curator published comprehensive information on powdery mildew resistance in the NPGS cucumber collection. 4c.List any significant activities that support special target populations. None. 5.Describe the major accomplishments over the life of the project, including their predicted or actual impact. Obj 1 Progress: 2005: Total new acquisitions numbered 450 accessions, including wild Helianthus from the southwestern US; Hypericum (St. John’s wort), Brassica from other collections; vegetable germplasm; maize and its relatives from existing collections and other researchers; Brassica species with phytoremediation applications. Conserve: Over 1600 total accessions were grown for regeneration with 1,363 harvested. Approximately 1,197 accessions were made available to the public. Over 3,000 accessions were tested for viability, approximately 6% of our holdings. About 615 accessions were backed up at the NCGRP; 76% of our holdings are currently backed up, and 71% are available to requestors. A Helianthus collection trip in the High Plains is currently underway. Our medicinal/nutriceutical curator, J. McCoy, acquired 138 new accessions of medicinal species, representing 44% of the current collection. Nineteen accessions of black cohosh (Actaea racemosa) from its native range were acquired through a research donor. Black cohosh, goldenseal (Hydrastic candensis), blood root (Sanguinaria canadensis) and fairywand (Chamaelirium luteum) were determined to be able to survive the winter and successfully reproduce in Iowa. Maize acquisitions included 35 accessions, and included 12 newly expired PVP lines; six CSR-registered inbred lines; one Onaveno landrace collection from Mexico; five sweet corn populations from Dr. W. Tracy, Univ. of Wisconsin; two Arizona landrace collections via Dr. M. Krakowsky, the USDA breeding collection at Tifton, GA; an apomictic maize line from Dr. B. Kindiger, USDA-ARS, El Reno, OK. Ornamental and mint family acquisitions consisted of 58 accessions including 16collected from wild populations in MN by Dr. M. Widrlechner and J. Carstens; 10 from the Ozarks and Carolinas made by Dr. J.McCoy; 8 from the Republic of Georgia, and 10 which were re-obtained from original sources., Helianthus collection expanded by 13 H. californicus from Drs. T. Gulya and G. Seiler, USDA-ARS, Fargo, ND; 14 accessions from the SE US; three H. niveus ssp. tephroides from CA collected by BLM collaborator C. Knauf; 19 from four species collected in southern CA by Drs. T. Gulya and L. Marek. These new accessions include 10 species which have not previously been available for distribution. Increased interest by the sunflower research community in wild Helianthus germplasm has led to a CGC approved goal of acquiring representation of the geographic distribution of all 66 Helianthus species. The Eruca (arugula) collection received 62 new accessions from Dr. J. Janick, Purdue Univ., IN, which were collected in the Mediterranean area of Europe; one Alyssum murale accession was received from Dr. R. Cheney, USDA, Beltsville, MD, which is capable of Ni hyper-accumulation and may be useful for phytoremediation. Nine accessions of Linum usitatissimum (flax) were received from an ICARDA/NPGS collection trip in Tajikistan; one L. lewisii accessions received from the NRCS Plant Materials Center in Aberdeen, ID through Shawn Belt, NCCS Beltsville, as a selected ecotype for restoration of disturbed sites in the central intermountain west. One Cuphea viscosissima accession was collected in MO by Dr. J. McCoy. L. Towil of ARS-NCGRP successfully collaborated with M. Widrlechner to develop and evaluate methods of storing Salix (willow) clones via cryogenic conditions. Germination methods developed by Crane & Walters (NCGRP) in collaboration with NCRPIS staff members for Cuphea sp. were successfully implemented by the NCRPIS oilseeds curation team. This enabled regeneration of some accessions which previously could not be germinated adequately. Methods developed by D. Kovach, (ARS – PIR) and ISU Oilseeds Curator Laura Marek to break dormancy in wild Helianthus, Cuphea and Linum were tested and implemented to promote germination of these taxa. Other acquisitions included 22 accessions of Melilotus from a joint ICARD/USDA-ARS plant exploration project in Tajikistan; five millet accessions transferred from NCGRP; a small number of amaranth, quinoa and umbel accessions. Forty new vegetable accessions were received, including seven Daucus varieties transferred from the NCGRP in Ft. Collins, CO and incorporated into the collection following regeneration by cooperators; four Cucumis melo varieties and one Greek Cucumis landrace through California researchers; one Cucumis melo and 22 Daucus collected from Uzbekistan by Dr. P, Simon, USDA-ARS, Madison, WI; one Cucumis melo and two Cucurbita pepo from NCGRP as expired PVP lines. Obj 2: 2004-2005: Over 4,500 accessions were observed and characterized for a wide array of descriptor information; over 18,600 observations were transferred to the GRIN database in 2004. Digital image capture continued to be a major area of focus, resulting in capture of over 3,120 images of almost 1700 accessions. An obstacle to transfer of digital images to GRIN was development of an image mass-loading protocol. ARS IT Specialist P. Cyr and ISU Maize Curator Mark Millard completed software development in May, 2005, and various crop curation teams have implemented its use at various NPGS genebanks. Maize GDB now links molecular genetic information to accession information located in the GRIN database; its order module enables genomic researchers to order maize accessions referenced in the GRIN database. DNA extraction of coriander, isozyme and SSR analyses were conducted, and the information will be analyzed together with previously collected morphological, phenotypic and biochemical traits to establish phylogenetic associations. Initial analyses based on all data were completed in 2004, and will be repeated prior to December, 2005. DNA capture on paper storage media has been investigated; development will continue this winter in cooperation with the ISU Seed Science Center. Hypericum and Echinacea are being evaluated and characterized as they are regenerated. Data analysis was completed on cultivated sunflower accessions for resistance to sunflower moth feeding. Insect pollinator investigations: our entomologists are developing methods to better utilize alfalfa leaf cutter bees in our regeneration programs. Use of alfalfa leaf cutter bees (ALC) for sunflower, Brassica and vegetable curation programs was evaluated in the field and greenhouse in 2004-2005, and for some ornamental species in the field in 2005. Seed production using ALC’s is being evaluated to compare their efficacy with honeybee, fly and bumblebee use in seed regeneration efforts. More extensive use of non-stinging, solitary ALC bees could improve worker safety and decrease program costs. In-house fly rearing was terminated in 2004; fly pupae are now purchased, resulting in cost-savings and enabling our researchers to spend their time more effectively. Field observations for disease resistance were made for sunflowers resistant to Septoria leaf blight; maize for artificially infested Stewart’s wilt and naturally occurring common rust and gray leaf spot; vegetables for viruses and bacterial diseases. Monitoring for naturally occurring pathogens to preclude accidental distribution of infested seed is an ongoing part of germplasm maintenance and distribution activities. Real-time PCR analysis was adopted by Pathologist C. Block’s team for routine detection and identification of Acidovorax avenae subsp. citrulli (Aac), the bacterial fruit blotch pathogen of melons. This allows monitoring and rapid identification of the organism directly from field or greenhouse leaves in one day. Cooperative research with Dr. R. Walcott, Univ. of Georgia, is focused on developing real-time PCR methods for the simultaneous detection of multiple pathogens from seeds. Block’s team developed real-time PCR primers for Pantoea stewartii, the causal agent of Stewart’s bacterial wilt of maize. Starting from a pure culture of the pathogen, the detection threshold for real-time PCR was about 10 colony forming units (CFU)/ml, compared with 103 CFU/ml for conventional (agarose gel) PCR and 105 CFU/ml for ELISA. Real-time PCR is not as sensitive when working with infected seeds because of inhibitors in the seeds, but work continues on magnetic bead capture methodology, which shows promise for rapid and specific extraction of the pathogen from corn seeds. Helianthus core collection analysis was temporarily stopped due to personnel changes and has been re-initiated in 2005. Nearly 197 plants of five accessions were distributed to approximately 20 sites for evaluation in the Regional NC-7 woody ornamental trial evaluations across the Midwestern U.S. Cooperator data was summarized for 1,5, and 10 year periods. These included tree seedlings grown from the seeds collected during the 1999 Ukraine exploration. The NC-7 Trials marked their 50th anniversary in 2004. Obj 3: 2005: Research to classify Brassica phylogenetically based on flowering date groups, sequencing and molecular profiles is in progress. Morphological and phenological data collected is complete; DNA sequencing is complete; SSR profiling is in process and will be analyzed in 2005-06. Evaluation is complete of the geographic distribution of Septoria resistant wild Helianthus; a manuscript is in process. C. Block is trying to determine if cucumber and rootworm beetles are capable of spreading plant viruses by feeding on Cucumis and Cucurbita plant parts pressed against field cage screens; beetles are being tested for presence of viruses via ELISA. Horticulturist M. Widrlechner continues to monitor changes to local woody ornamental flora. From 2001-2004, tree seedlings which were collected during a 1999 Ukraine exploration were distributed to NC-7 ornamental trial collaborators for long-term evaluation in order to identify well-adapted, new landscape plants for the NCR and to test scientific hypotheses about relationships among soils, climates, vegetation patterns and woody-plant adaptation. One, five and ten-year performance ornamental evaluations are in progress; data is posted in public GRIN utilizing a streamlined process to transfer the information from an Access database. Obj 4: 2005: Distribute germplasm; incorporate research findings on germplasm through databases, websites, publications, lectures and other outlets as appropriate. Distributions included 17,983 seed packets or plants in 809 orders to external germplasm requestors. These represented 10,481 accessions, consistent with an annual 20-25% distribution of all holdings. Approximately 69% of these were to U.S. and 31% to international requestors. Approximately 20-25% of the collection continues to be distributed each year. Seed order numbers have greatly increased (15%) due to interest from home gardeners; although our resources cannot support maintaining and distributing the collections to home gardeners, we have tried to use this development to educate the public about plant genetic resource conservation and encourage interested individuals to save seeds, conserve and share germplasm and associated information. Two multiple disease-resistant sunflower germplasm populations, SAM-1 and SAM-2, released in 2004, have been widely distributed. Other heavily demanded crops for research purposes include oilseed brassica and crucifers for disease screening (black stem rot and white mold), crop improvement, and phytoremediation; sunflower for disease resistance, oil quality traits, and adaptation; maize for many purposes; vegetables for disease traits, flavor and aesthetic traits, and general crop improvement; millets for crop improvement; prairie coneflower for nutriceutical and ornamental traits; woody and herbaceous ornamentals for ornamental research and varietal improvement. Our scientific staff published presented information in scholarly and popular press journals, and delivered presentations to a wide array of audiences. A highlight was release of a digitally captured, searchable version of the now out-of-print volumes of the Races of Maize; this series of books was written by international researchers over a period of decades and remains invaluable as a resource for maize researchers. Over 320 visitors toured the NCRPIS during 2004, including students from K to postdoctoral, visiting scientists, researchers, educators, and interested members of the public. Valuable training and knowledge was transferred to over 70 college students who were employed by the ARS at the PI station. Implementation of an Access Security System has greatly enhanced the security of the germplasm collections and NCRPIS and PIR personnel. In addition to progress in acquiring, characterizing and evaluating PGR, our project has contributed to the development of the Germplasm Resource Information Network (GRIN), has transferred over 83,000 items related to evaluation and characterization of PGR since 2003, which have increased PGR research users to better target their efforts and improved the efficiency and efficacy of their work. The posting of digital images associated with the accessions increases germplasm requestors’ ability to better target traits and characteristics of interest. Our curators, scientists and staff have increased the awareness of the ability of the collections, done extensive research to support PGR user’s requests by identifying accessions which best support requestors’ objectives, and focused on development of methods that optimize resource use with respect to effective germplasm regeneration, conservation, evaluation and characterization, and enable us to move forward on many of our mission’s objectives simultaneously. Their impact is realized through development of new varieties and cultivars, and new or improved plant products as a result of their use in research and development programs. Their impact is also being realized in the field of genomic discovery, as the collections provide much genetic variation which genomic researchers and bioinformatics can utilize to understand plant development, evolutionary relationships, enzymatic and other biochemical processes, which in turn can result in new applications. 6.What science and/or technologies have been transferred and to whom? When is the science and/or technology likely to become available to the end-user (industry, farmer, other scientists)? What are the constraints, if known, to the adoption and durability of the technology products? The project conserves and transfers plant genetic resources and associated information to the research and educator public for development of new varieties, new uses, and solutions to agricultural production problems. These varieties also contribute to health and nutritional research, and to biobased industrial applications which contribute to the security and stability of our nation’s economy. Over the next three years, our horticulturist, plant pathologist, geneticists and curatorial staff will work to provide quality germplasm and associated information to germplasm researchers worldwide. Appropriate actions continue to be implemented to ensure the physical and electronic security of our collections, facilities and information resources. By increasing our regeneration and curatorial staff capacity and integrating new technology, our ability to meet our objectives is enhanced. Constraints to the adoption and durability of new technologies derived from plant genetic resources are those commonly associated with development of new agricultural and horticultural products. Review Publications Block, C.C., Reitsma, K.R. 2005. Powdery mildew resistance in the U.S. National Plant Germplasm System cucumber collection. Hortscience. 40:416-420. Walcott, R.R., Sanders, F.H., Block, C.C. 2004. A lognormal distribution of phytopathogenic bacteria in corn, cowpea, tomato and watermelon seeds. American Phytopathological Society. 94:S34. Widrlechner, M.P. 2005. Germplasm acquisition from conception to products. (Introduction to Workshop Proceddings). HortScience. 40:2096 Lopez, P., Widrlechner, M.P. 2004. Morphological and chemical variability in coriander germplasm. Association for the Advancement of Industrial Crops Conference, September 19-23, Minneapolis, MN. AAIC Abstracts p. 30. Palmer, R.G., Ortiz-Perez, E., Cervantez-Martinez, I.G., Horner, H.T., Hanlin, S.J., Wiley, H., Davis, W.H. 2005. Hybrid soybean:a positive progress report. American Farm Bureau Federation Publication. Paper No. 24. Block, C.C. 2005. Evaluation of wild helianthus annuus for resistance to septoria leaf blight. Proceedings Sunflower Research Workshop. Available: http://www.sunflowernsa.com/research/research-workshop/documents/Block_Septoria_05.PDF McCoy, J.H., Widrlechner, M.P., Carstens, J.D. 2005. A comprehensive Echinacea germplasm collection located at the North Central Regional Plant Introduction Station, Ames, Iowa. HortScience. 40:1063.