2006 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? Why does it matter? The most critical problem facing the U.S. hop industry is the need for hop varieties that provide superior brewing quality and that contain resistance to two serious diseases, powdery (PM) and downy mildew (DM). Although DM has been a problem in U.S. hop producing regions for many years, hop PM was first identified in the Pacific Northwest growing regions in 1997. Downy mildew breeding has not progressed significantly over the past half century due to the highly variable nature of disease expression in hops. The increased costs associated with controlling these two diseases threaten the economic viability of the entire U.S. hop industry. Presently, the only way to control PM and DM is by growing resistant varieties or applying expensive fungicides, as many as eight times per season. The costs of these applications can easily exceed the gross receipts for the crop. When PM first emerged in U.S. hop production fields, only a few resistant varieties were in production and we had no understanding of genetic mechanisms that would contribute to disease resistance. Furthermore, we know even less about the epidemiology and genetic mechanisms for resistance to hop DM. Without significant and rapid progress in developing more economic means to control the disease, U.S. hop production cannot remain competitive in global hop markets, farm profitability will severely decline, and reliance of brewers on foreign hop imports will increase. At the same time, it also is important to identify specific compounds in hop cones that contribute superior flavor attributes, and develop varieties that produce enhanced levels of these compounds for commercial production. The objectives of this research are:. 1)Identify, sequence, map and characterize hop genes that confer specific phenotypic functions of economic importance.. 2)Characterize genetic diversity of hop accessions maintained in the USDA hop germplasm collection.. 3)Identify and develop hop germplasm possessing new and unique traits that have economic impact on the American hop industry. To solve these problems, both cultivated and wild hop germplasm are being evaluated for their potential use in development of new varieties, traditional plant breeding is being used to incorporate traits that provide disease resistance and superior brewing characteristics into enhanced germplasm, and molecular markers are being developed to decrease the length of time required to develop new varieties. This research addresses objectives of National Programs #301- Plant Genetic Resources, Genomics, and Genetic Improvement; National Program #303 - Plant Diseases; and National Program #306 -Quality and Utilization of Agricultural Products. This work is relevant to U.S. breweries, hop marketing companies, and hop producers and critical to maintain U.S. leadership in hop production and export is critical with more than 60% of the U.S. hop acreage planted in USDA-ARS varieties. 2.List by year the currently approved milestones (indicators of research progress) Year 1 (FY2003) Make Genomic Libraries Estimate Genetic diversity/ Publish results Make year 2003 Crosses Year 2 (FY2004) Run micro-satellites/Take field data-1st yr Collect second year data estimating heritability of specific traits Year 3 (FY2005) Finish sequencing genes/ analyze data for Objective 1 Finish micro-satellites/ 2nd year data collection Collect 1st year data and release superior germplasm from year 2001 crosses Year 4 (FY2006) Collect 2nd year data for Objective 2 Collect 2nd year data for Objective 3 Year 5 (FY2007) Release superior germplasm from year 2003 crosses 4a.List the single most significant research accomplishment during FY 2006. Molecular Detection of Airborne Spores of the Hop DM Pathogen. The USDA-ARS Forage Seed and Cereal Research Unit in collaboration with Washington State University developed molecular assays to detect the hop DM pathogen in near real-time from the air of hop yards. This is significant because fungicides commonly are over-used for management of hop DM because rapid and sensitive methods for detecting the pathogen are not available. The assay and a novel spore trapping method were developed and validated under field conditions, and were able to detect airborne spores of the pathogen 10 days before symptoms development. This discovery may allow growers to minimize pesticide use by eliminating applications when the pathogen is not detected, and result in optimized fungicide use, reduced input costs for producers, and enhanced environmental protection. This accomplishment addresses National Program 303 – Plant Diseases; Component I - Disease Diagnosis Detection, Identification and Characterization of Plant Pathogens; Problem Statement b) Detection, Identification, Characterization, and Classification of Pathogens. 4b.List other significant research accomplishment(s), if any. Plant Resistance Genes and General Stress-related Genes Cloned and Identified in Hop. The USDA-ARS Forage Seed and Cereal Research Unit identified 221 gene sequences (EST’s) expressed by hop during initial PM infection phase. Hop PM is a recently introduced fungal pathogen causing significant damage on susceptible varieties with virtually no information available on host-pathogen resistance mechanisms. To accomplish this, we used suppressive subtraction hybridization technique that effectively eliminated non-disease resistance genes and purified and amplified plant resistance genes expressed during PM infection. This is the first study of such magnitude identifying genes involved in plant resistance in hop and will ultimately provide geneticists and breeders molecular tools and genetic knowledge to select for resistance to new races of PM as they arise in hop production. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component II -Crop Informatics, Genomics, and Genetic Analyses; Problem Statement c) Genetic Analyses and Mapping of Important Traits. 4c.List significant activities that support special target populations. None. 4d.Progress report. Opportunities to submit additional programmatic information: A CRADA is in the process of being established with Roy Farms Inc (Yakima, WA) working towards the mutual development of dwarf hop lines. In addition, a joint USA-Czech Republic research project is being pursued to work on improving genetic diversity of hop using plant collection and evaluation of new material. 5.Describe the major accomplishments to date and their predicted or actual impact. Genetic diversity of USA hop germplasm estimated and genetic bottlenecks identified. AFLP Fingerprinting and estimation of genetic diversity of the USDA-ARS germplasm pool identified specific genetically diverse male-female pairs for germplasm development and helped delineate a significant genetic bottleneck present in current USA hop cultivars. Hop is a clonally propagated perennial crop with virtually no information regarding the genetic relatedness of cultivated and wild accessions present in the USDA-ARS hop germplasm collection. We analyzed the AFLP fingerprints of 154 cultivated and wild North American hop accessions and determined the genetic relationship amongst male-female pairs of accessions. Use of genetic diversity information has enabled better decisions regarding choice of parents for germplasm development and should result in less dependence upon a genetically similar group of cultivars all of which are potentially susceptible to any newly introduced pathogen. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component I – Plant and Microbial Genetic Resource Management; Problem Statement b) Assess the Systematic Relationships and Genetic Diversity of Crop Genetic Resources. Molecular markers to select for resistance to PM identified. PCR techniques including AFLP and differential display analysis were used to identify 24 different potential molecular markers for PM resistance. Conventional breeding techniques for resistance to PM is both time-consuming and prone to errors while selection using molecular techniques helps overcome these shortcomings. We used both differential display on treated and untreated hop lines and AFLP on seven near-isogenic hop accessions (bred for the presence of the seven different R-genes for PM) to identify potential molecular markers for selection. These studies were the first preliminary work identifying genetic sequences in hop involved in plant resistance mechanisms. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component II - Crop Informatics, Genomics, and Genetic Analyses; Problem Statement c)Genetic Analyses and Mapping of Important Traits. Molecular markers to select for DM resistance identified. Five AFLP molecular markers linked significantly to DM resistance in hop were identified by researchers at the Forage Seed and Cereal Research Unit in Corvallis, OR. Selection for hop DM resistance has defied conventional breeding techniques due to a combination of complex inheritance and environmental influences, and use of molecular breeding techniques may help overcome these difficulties. Representative accessions of the USDA-ARS hop germplasm collection were scored for resistance to DM and molecular markers from AFLP analysis on these same individuals were statistically associated with disease scores using logic regression. These findings will potentially enable the use of molecular selection procedures and increase the efficiency and accuracy of selection for plant resistance to this economically important plant pathogen. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component II - Crop Informatics, Genomics, and Genetic Analyses; Problem Statement c)Genetic Analyses and Mapping of Important Traits. New public hop varieties developed and released. Hop varieties ‘Newport’, ‘Santiam’, ‘Sterling’, ‘Horizon’, ‘Bittergold’ and ‘Teamaker’ were released in direct response to requests from industry for aroma and bittering hop varieties possessing high yields, better brewing characteristics and/or superior disease resistance. Hop industry representatives requested development of superior, higher-yielding, disease resistant public varieties that would enable growers to reduce costs for production and simultaneously reduce environmental impact by decreasing pesticide usage. Hop varieties were developed in cooperation between USDA ARS Forage Seed and Cereal Research Unit and Washington State University using traditional breeding techniques and additional funding from public and private stakeholders. The development and release of these varieties should result in significantly higher farm profits due to increased yields coupled with lower costs associated with reduced pesticide applications and will ultimately reduce the impact of pesticides upon our environment. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component III - Genetic Improvement of Crops; Problem Statement c) Germplasm Enhancement/Release of Improved Genetic Resources and Varieties. Hop PM resistance genes “cascaded” into a single genetic background. The hop PM resistance genes from three different sources of resistance were incorporated into a single genetic background. Hop PM is an obligate fungal pathogen capable of quickly overcoming single-gene based resistance mechanisms and experience in other crops demonstrated extended resistance to similar pathogens using “gene cascading techniques.” We utilized traditional breeding methods to incorporate genes from three different sources of resistance into a single genetic background. Our germplasm development efforts should result in extended resistance before the pathogen overcomes plant resistance based upon presence of all three resistance genes. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component III - Genetic Improvement of Crops; Problem Statement c) Germplasm Enhancement/Release of Improved Genetic Resources and Varieties. Fungal resistance gene isolated and inserted into susceptible hop line. In collaboration with Huell Hop Research Center, Germany, we successfully inserted the chitinase gene isolated and identified by our lab in 1999 into a PM-susceptible hop variety resulting in increased resistance to PM. Breweries desired a genetic solution to develop new hop varieties that differ from currently used varieties solely by possession of superior resistance to fungal pathogens. Agrobacterium tumefaciens constructs were used to insert a continuous-expression-promoter coupled with the hop chitinase gene into a PM susceptible hop variety. The over-expression of this hop chitinase gene appears to result in increased resistance to PM and if proven, offers a solution for developing brewery acceptable hop varieties differing only in the presence of pathogen resistance genes. This accomplishment addresses National Program 301 – Plant Genetic Resources, Genomics, and Genetic Improvement; Component III - Genetic Improvement of Crops; Problem Statement c) Germplasm Enhancement/Release of Improved Genetic Resources and Varieties. Leaf wetness reduces PM severity. In collaboration with USDA-ARS Horticultural Crops Research Laboratory, we demonstrated that leaf wetness greatly reduces the severity of infection of hop by its PM pathogen. Infection risk models for hop PM are not accurate in regions with high rainfall because knowledge of the role of leaf wetness on disease is incomplete. Controlled environment experiments simulated field conditions and found that short durations of surface wetness suppressed the disease. Knowledge of this environmental factor should lead to better prediction of PM diseases since leaf wetness is not considered in current risk assessment models. This accomplishment addresses National Program 303 – Plant Diseases; Component II - Biology, Ecology, Epidemiology, and Spread of Plant Pathogens and Their Relationships with Hosts and Vectors; Problem Statement c) Population Dynamics, Spread, and Epidemiology of Pathogens. Fungal resistance to phenylamide persists after 20 years. In collaboration with Washington State University, discovered resistance to phenylamide fungicides has persisted in the DM pathogen for nearly 20 years in the near absence of their use. Hop DM management relies heavily on chemical inputs, but indiscriminate use of fungicides has resulted in the rapid development of pathogen resistance. Laboratory evaluations of hop DM isolates from the Pacific Northwest measured pathogen sensitivity to certain fungicides, revealing that insensitive phenotypes are present at a high frequency within the population. This discovery will allow for the elimination of chemical controls to which the pathogen remains insensitive. This accomplishment addresses National Program 303 – Plant Diseases, Component I - Disease Diagnosis Detection, Identification and Characterization of Plant Pathogens; Problem Statement b) Detection, Identification, Characterization, and Classification of Pathogens. 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? 1) Hop research program tours were presented to representatives of Anheuser Busch and Oregon Hop Commission, Washington Hop Commission and to individual hop producers. Most of the science and technology presented in these tours were focused towards growers and are adaptable immediately, and hold long-term relevance to production. 2) Presented research reports to the "Hop Research Council" discussing recent advances in hop genetics and plant pathology. The science and technology presented are relevant towards the future and are focused primarily upon brewery as well as grower adaptation and usage. 3) Collaborated with the Oregon State University Integrated Plant Protection Center to make the hop powdery mildew risk index available publicly in near-real time for the entire U.S. This technology is available immediately and is only constrained by grower access to internet. 4) Led USA hop breeders (both private and public) in discussions focused upon systematizing reporting of hop varietal release data and delineating the requirements for germplasm and varietal release. The science and technology presented were applicable to geneticists and breeders and is available for usage immediately in their programs. 7.List your most important publications in the popular press and presentations to organizations and articles written about your work. (NOTE: List your peer reviewed publications below). Oral or poster presentations to the following organizations: -Hop Research Council (January, 2006 and August, 2006) -Oregon Hop Research Council (July 2006) -Washington Hop Commission Scientific Meeting (January, 2006)