Research Project: Improving Control of Stripe Rusts of Wheat and Barley through Characterization of Pathogen Populations and Enhancement of Host Resistance

Location: Wheat Health, Genetics, and Quality Research

2019 Annual Report

Objectives
Stripe rust is one of the most important diseases of wheat throughout the U.S. and stripe rust of barley causes significant yield losses in western U.S. Significant progress has been made in understanding biology of the pathogen, virulence compositions of the pathogen population, identification of new sources of resistance, disease forecasting, and control of the disease using fungicides. However, research is needed to develop more effective strategies for sustainable control of stripe rusts. Therefore, this project has the following objectives for the next five years: Objective 1: Monitor and characterize stripe rust pathogen populations for providing essential information to growers for implementing appropriate measures to reduce damage on wheat and barley. Subobjective 1A: Identify virulent races of stripe rust pathogens to determine effectiveness of resistance genes in wheat and barley. Subobjective 1B: Develop molecular markers and characterize stripe rust pathogen populations to identify factors and mechanisms of pathogen dynamics for developing new management strategies. Objective 2: Enhance resistance in wheat and barley cultivars for sustainable control of stripe rusts. Subobjective 2A: Identify and map new genes for stripe rust resistance, and develop new germplasm for use in breeding programs. Subobjective 2B: Screen breeding lines for supporting breeding programs to develop wheat and barley cultivars with adequate and durable resistance to stripe rust. Accomplishment of these objectives will lead to improved knowledge of the disease epidemiology for developing more effective control strategies, more resistance genes and resistant germplasm to be used by breeders to develop stripe rust resistant wheat and barley cultivars, and more effective technology to be used by wheat and barley growers to achieve sustainable control of stripe rust.

Approach
Monitoring the prevalence, severity, and distribution of stripe rust and identify races of the pathogens, commercial fields, monitoring nurseries, trap plots, and experimental plots of wheat and barley, as well as wild grasses, will be surveyed during the plant growing-season. Recommendations will be made based on stripe rust forecasts and survey data to control stripe rust on a yearly basis. Rust samples will be collected by collaborators and ourselves during surveys and data-recording of commercial fields, experimental plots, and monitoring nurseries. Stripe rust samples will be tested in our laboratory for race identification. New races will be tested on genetic stocks, commercial cultivars, and breeding lines to determine their impact on germplasm and production of wheat and barley. Isolates of stripe rust pathogens will also be characterized using various molecular markers to determine population structure changes. To identify new genes for resistance to stripe rust in wheat, the mapping populations, which have been and will be developed in our laboratory, will be phenotyped for response to stripe rust in fields and/or greenhouse and genotyped using different approaches and marker techniques such as simple sequence repeat (SSR), single-nucleotide polymorphism (SNP), and /or genotyping by sequencing (GBS) markers. Linkage maps will be constructed with the genotype data using software Mapmaker and JoinMap. Genes conferring qualitative resistance will be mapped using the phenotypic and genotypic data using JoinMap or MapDisto, and quantitative trait locus (QTL) mapping will be conducted using the composite interval mapping program in the WinQTL Cartographer software. Relationships of genes or QTL to previous reported stripe rust genes will be determined based on chromosomal locations of tightly linked markers, type of resistance and race spectra, and origins of the resistance gene donors. Allelism tests will be conducted with crosses made between lines with potentially new gene with previously reported genes in the same chromosomal arms to confirm the new genes. A homozygous line carrying a identified new gene with improved plant type and agronomic traits will be selected from the progeny population using both rust testing and molecular markers. Lines with combinations of two genes in a same chromosome will be selected and confirmed using phenotypic and marker data. The new germplasm lines will be provided to breeding programs for developing resistant cultivars. To support breeding programs for developing cultivars with stripe rust resistance, wheat and barley nurseries will be evaluated in two locations: Pullman (eastern Washington) and Mount Vernon (western Washington) under natural infection of the stripe rust pathogens. Variety trials and uniform regional nurseries will also be tested in seedling and adult-plant stages with selected races of the stripe rust pathogen under controlled greenhouse conditions. The data will be provided to breeding programs for releasing new cultivars with adequate level and potentially durable stripe rust resistance.

Progress Report
Progress was made on the two objectives and four subobjectives, all of which fall under National Program 303 Plant Diseases. Under Sub-objective 1A, ARS scientists in Pullman, Washington, completed all tests for identifying races from the stripe rust collections in the U.S. in 2018, and reported the results to the cereal community of researchers, developers, and growers. During the 2019 growing season, they conducted field surveys, made recommendations for control of stripe rust, and collected stripe rust samples in the Pacific Northwest. Through collaborators in other regions, they received more than three hundred stripe rust samples from fifteen states. These samples were used to inoculate susceptible wheat plants for recovering and increasing spores. Each isolate was tested on the set of wheat or barley differentials for identifying races of the wheat or barley stripe rust pathogen. From the 2018 samples, they identified 23 races (including 1 new race) of the wheat stripe rust pathogen and 12 races (including 1 new race) of the barley stripe rust pathogen from the 2018 collection and determined the distributions and frequencies of the races and virulence factors. From the 2019 collection, nine races of the wheat stripe rust pathogen have been identified. Based on the virulence data of the races, resistance genes, such as Yr5, Yr15, and others, have been determined to be effective against all populations of the wheat stripe rust pathogen in the U.S., and these genes can be used in breeding programs for developing stripe rust resistant wheat cultivars. Similar information has also been obtained for the barley stripe rust pathogen and barley resistance genes. Based on virulence patterns, distributions, and frequencies, they selected races for use in their wheat and barley germplasm screening research. This progress is on schedule. Under Sub-objective 1B, ARS scientists in Pullman, Washington, sequenced the genomes of more than 150 stripe rust isolates including 30 isolates developed by chemical mutagenesis and 117 isolates produced through sexual reproduction on alternate host barberry plants under controlled greenhouse conditions. They established high-quality genome sequences for both wheat and barley stripe rust pathogens using the PacBio long-read sequencing technology for use as references for various genome comparison studies. By comparing sequences of wheat and barley stripe rust pathogens, they determined that these two forms of stripe rust fungus has been diverged several million years ago mainly through gene losses due to transposon activities, and identified genes involved in plant host adaptation. They published another high-quality genome sequence of a wheat stripe rust isolate collected from grass, which allow them to study a wide range of avirulence genes. Through analyzing the sequence data of the mutant and sexually produced populations together with virulence data, they mapped virulence loci and identified candidate genes for different avirulence factors. They used the genome sequences of previously sequenced isolates to develop molecular markers and characterize the stripe rust population. They have selected 18 simple sequence repeat (SSR) markers as a core set of markers and used them to characterize stripe rust collections from the U.S. and fifteen other countries. They completed the marker tests for the collections up to 2017. Based on the marker data of the international collections, they identified two major genetic groups and 19 subgroups of the wheat stripe rust pathogen and their distributions and frequencies in different countries. They identified similar genetic groups present in countries of different continents. Through molecular characterization of the U.S. stripe rust collections from 1968 to 2017, they identified several events of stripe rust pathogen introduction and different population structures in various epidemiological regions. These results greatly improve the knowledge of the pathogen biology and genetics and understanding of stripe rust epidemiology, which support the needs for developing wheat and barley varieties with non-race specific and durable resistance against the ever changing and spreading stripe rust pathogen populations at the global scale. Under Sub-objective 2A, ARS scientists in Pullman, Washington, planted more than 40 wheat mapping populations at Pullman and Mount Vernon, Washington, for phenotyping stripe rust responses to map resistance genes in these crosses. They have recorded stripe rust responses. The rust response data will be used to identify and map new and effective resistance genes. In addition, they planted more than 900 spring U.S. wheat varieties for stripe rust phenotyping in the fields in 2019 to get one more year data in the fields. The spring wheat panel is parallel to their study of over 800 U.S. winter wheat accessions, for which they completed the phenotype experiments in 2018. These two panels of wheat germplasm have also been tested with selected stripe rust races in the greenhouse. The entries in both winter and spring wheat panels have been genotyped with single-nucleotide polymorphism (SNP) markers. They are currently conducting genome-wide association studies (GWAS) using the rust response data and molecular marker data, and their preliminary results identified more than 20 different genes in each panel. They have completed a study to identify and map genes for stripe rust resistance in the winter wheat variety ‘Eltan’, which has been widely grown and used in breeding programs in the Pacific Northwest since the early 1990s. In this study, they identified five quantitative trait loci (QTL) controlling different types of resistance and with different levels of effectiveness. Their results show that the resistance reduction of Eltan was caused by changes of the pathogen population from avirulent to virulent, overcoming the race-specific all-stage resistance and the low level of high-temperature adult-plant (HTAP) resistance. Based on the results, they pointed out a better strategy for developing wheat varieties with durable, high level resistance through combining high-level HTAP resistance genes with effective all-stage resistance. Under Sub-objective 2B, ARS scientists in Pullman, Washington, planted more than 35,000 wheat and barley entries in the fields near Pullman and Mount Vernon, Washington for screening the materials for resistance to stripe rust. Some of the nurseries were also planted in three additional locations. They have completed stripe rust response data for the nurseries planted in different locations. They also have tested the wheat and barley variety trial nurseries and uniform nurseries from various wheat and barley production regions in the U.S. in both seedling and adult-plant stages with selected races of the stripe rust pathogen in the greenhouse. The thorough tests in both field and greenhouse allow identifying different types of resistance to stripe rust as well as different levels of resistance. They have provided the data to various breeding programs throughout the country for selecting resistant lines for releasing as new varieties. The test results of currently grown varieties are used to update stripe rust reactions for the varieties listed in Seed-Buyers’ Guide to be selected by growers to grow. Growing resistant varieties will reduce the potential risk of stripe rust damage.

Accomplishments
1. Mapped stripe rust resistance genes in winter wheat variety Eltan. Stripe rust is best controlled through growing resistant varieties, and Eltan has been one of the most widely grown winter wheat variety and used intensively in breeding programs in the Pacific Northwest, but its genes for stripe rust resistance was unknown. ARS scientists in Pullman, Washington, mapped a total of five genes for resistance to stripe rust, of which two major genes and a minor gene conferred race-specific all-stage resistance and two minor genes conferred non-race specific high-temperature adult-plant (HTAP) resistance in Eltan. They determined that the reduction of stripe rust resistance in Eltan in the recent years was due to the fact that the recent predominant races overcome the race-specific all-stage resistance and the level of HTAP resistance is low. Their results point out the importance of combining high-level HTAP resistance and effective all-stage resistance for developing wheat varieties with high-level, durable resistance to stripe rust. The markers identified for HTAP resistance genes in Eltan are useful for combining with other effective resistance genes for developing new wheat varieties.

2. Screened more than 35,000 wheat and barley germplasm for resistance to stripe rust. Developing resistant varieties is the most effective, easy-to use, and environmental-friendly approach to control stripe rust. ARS scientists in Pullman, Washington, screened more than 35,000 wheat and barley germplasm from breeding programs throughout the U.S. in various field locations and in the greenhouse for response to stripe rust in 2019. They provided the data to various breeding programs for releasing new resistant varieties and to growers for selecting released resistant varieties to grow. Based on their stripe rust data in the recent years, they collaborated with various breeding programs in releasing and registering 15 wheat and 2 barley varieties with resistance to stripe rust. Growing these new varieties will reduce potential risk of stripe rust.

4. Established high-quality genome sequences and identified genetic groups of the stripe rust pathogen. The genomic relationship of wheat and barley stripe rust pathogens is not clear. ARS scientists in Pullman, Washington, completed studies to establish high-quality genome sequences for both wheat and barley stripe rust pathogens using advanced sequencing technologies. They determined that the divergence of the two different forms of the stripe rust fungus started several million years ago and gene loss is one of the major evolutionary mechanisms in the formation of two highly specialized pathogens. Through comparing with other plant pathogenic fungi, they identified unique effectors in the wheat and barley stripe rust pathogens. The results significantly improve the understanding of the pathogen evolution, disease epidemiology, and mechanisms of plant-pathogen interactions. The sequence data are being used by ARS and Australian scientists for identifying avirulence genes and developing molecular markers for monitoring the wheat and barley stripe rust pathogens and improving stripe rust management.

5. Tested fungicides for control of stripe rust and wheat varieties for response to fungicide application. Although stripe rust can be effectively controlled by growing resistant varieties, fungicides are still needed for reducing damage in fields grown with varieties without an adequate level of resistance. During the 2019 growing season, ARS scientists from Pullman, Washington, tested 33 fungicide treatments, including new chemicals, for control of stripe rust on both winter and spring wheat crops. They also tested 23 winter wheat and 23 spring wheat varieties, which were selected based on their acreage grown in previous years in the Pacific Northwest, plus a highly susceptible variety as a check for each crop, to determine their yield losses caused by stripe rust and responses to fungicide application. The efficacies, rates, and timing of the fungicides for stripe rust control and potential yield loss to stripe rust and yield increase to fungicide application for each variety were determined. The data of fungicide efficacy and variety potential yield loss are the essential components for the integrated stripe rust management strategy based on individual varieties, and the results can be used for the chemical developers to register new fungicides and for growers to have more choices of chemicals.

Review Publications
Cobo, N., Plfuger, L., Chen, X., Dubcovsky, J. 2018. Mapping QTL for resistance to new virulent races of wheat stripe rust from two Argentinean wheat varieties. Crop Science. 58(6):2470-2483. https://doi.org/10.2135/cropsci2018.04.0286.
Qie, Y., Liu, Y., Wang, M., Li, X., See, D.R., An, D., Chen, X. 2018. Development, validation, and re-selection of wheat lines with pyramided genes Yr64 and Yr15 linked on the short arm of chromosome 1B for resistance to stripe rust. Plant Disease. 103(1):51-58. https://doi.org/10.1094/PDIS-03-18-0470-RE.
Yang, Y., Basnet, B.R., Ibrahim, A.M., Rudd, J.C., Chen, X., Bowden, R.L., Xue, Q., Wang, S., Johnson, C., Metz, R., Mason, R.E., Hays, D.B., Liu, S. 2018. Developing KASP markers on a major stripe rust resistance QTL in TAM 111 using 90K array and genotyping-by-sequencing SNPs. Crop Science. 59(1):165-175. https://doi.org/10.2135/cropsci2018.05.0349.
Chen, X., Sprott, J.A., Evans, C.K., Liu, Y. 2019. Evaluation of foliar fungicides for control of stripe rust on winter wheat in 2018. Plant Disease Management Reports. 13:CF068.
Chen, X., Sprott, J.A., Evans, C.K., Liu, Y. 2019. Evaluation of foliar fungicides for control of stripe rust on spring wheat in 2018. Plant Disease Management Reports. 13:CF067.
Chen, X., Sprott, J.A., Evans, C.K., Liu, Y. 2019. Evaluation of Pacific Northwest winter wheat cultivars to fungicide application for control of stripe rust in 2018. Plant Disease Management Reports. 13:CF066.
Chen, X., Sprott, J.A., Evans, C.K., Liu, Y. 2019. Evaluation of Pacific Northwest spring wheat cultivars to fungicide application for control of stripe rust in 2018. Plant Disease Management Reports. 13:CF065.
Zen, Q., Wu, J., Liu, S., Chen, X., Yuan, F., Su, P., Wang, Q., Huang, S., Mu, J., Han, D., Kang, Z. 2019. Genome-wide mapping for stripe rust resistance loci in common wheat cultivar Qinnong 142. Plant Disease. 103(3):439-447. https://doi.org/10.1094/PDIS-05-18-0846-RE.
Kiszonas, A., Higgenbotham, R., Chen, X., Garland-Campbell, K.A., Bosque-Perez, N.A., Pumphrey, M., Rouse, M.N., Hole, D., Wen, N., Morris, C.F. 2019. Agronomic traits in durum wheat germplasm possessing puroindoline genes. Agronomy Journal. 111(3):1254-1265. https://doi.org/10.2134/agronj2018.08.0534.
Wan, A., Wang, M., Chen, X. 2019. Variation in telial formation of Puccinia striiformis in the United States. American Journal of Plant Sciences. 10:826-849. https://doi.org/10.4236/ajps.2019.105060.
Liu, L., Wang, M.N., Feng, J.Y., See, D.R., Chen, X. 2019. Whole genome mapping of stripe rust resistance QTL and race-specificity related to resistance reduction in winter wheat cultivar Eltan. Phytopathology. 109(7):1226-1235. https://doi.org/10.1094/PHYTO-10-18-0385-R.
Kosman, E., Chen, X., Dreiseitl, A., McCallum, B., Lebeda, A., Ben-Yehuda, P., Gultyaeva, E., Manisterski, J. 2019. Functional variation of plant-pathogen interactions: new concept and methods for virulence data analyses. Phytopathology. 109(8):1324-1330. https://doi.org/10.1094/PHYTO-02-19-0041-LE.
Siyoum, G.Z., Zeng, Q., Zhao, J., Chen, X., Badebo, A., Tian, Y., Huang, L., Kang, Z., Zhan, G. 2019. Inheritance of virulence and linkages of virulence genes in an Ethiopian isolate of the wheat stripe rust pathogen (Puccinia striiformis f. sp. tritici) determined through sexual recombination on Berberis holstii. Plant Disease. 103(9):2451-2459. https://doi.org/10.1094/PDIS-02-19-0269-RE.
Li, Y., Xia, C., Wang, M., Yin, C., Chen, X. 2019. Genome sequence resource of a Puccinia striiformis isolate infecting wheatgrass. Phytopathology. 109(9):1509-1512. https://doi.org/10.1094/PHYTO-02-19-0054-A.
Wang, J., Wang, J., Shang, H., Chen, X., Xu, X., Hu, X. 2019. TaXa21, a LRR-rich receptor like kinase associated with TaWRKY76 and TaWRKY62, plays positive roles in wheat high-temperature seedling plant resistance to Puccinia striiformis f. sp. tritici. Molecular Plant-Microbe Interactions. https://doi.org/10.1094/MPMI-05-19-0137-R.