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ARS Home » Midwest Area » St. Paul, Minnesota » Cereal Disease Lab » Research » Research Project #423040

Research Project: CEREAL RUST FUNGI: GENETICS, POPULATION BIOLOGY, AND HOST-PATHOGEN INTERACTIONS

Location: Cereal Disease Lab

2015 Annual Report


Objectives
Objective 1: Monitor, collect, and characterize U.S. cereal rust pathogen populations. Sub-objective 1.A. Monitor, collect and characterize cereal rust pathogen populations in the U.S. for virulence phenotypes to rust resistance genes in cereal cultivars. Sub-objective 1.B. Determine levels of genetic variation in P. triticina and P. graminis populations. Sub-objective 1.C. Refine phylogenetics and systematics of P. graminis and P. triticina. Objective 2: Discover and characterize fungal genes that are involved in pathogenesis and the obligate biotrophic interactions of cereal rust pathogens and their hosts. Objective 3: Identify and characterize rust resistance genes in novel and elite germplasm to assist in the development of resistant cereal cultivars. Sub-objective 3.A. Evaluate wheat, oat and barley germplasm from U.S. breeding programs for rust resistance. Sub-objective 3.B. Identify and characterize new sources of rust resistance in wheat, barley, and oat. The proposed research objectives are central to the mission of the USDA ARS Cereal Disease Laboratory (CDL): to reduce losses in wheat, oat, and barley to major diseases using host resistance. Research is focused on genetic variation in both the host cereals and their rust pathogens that determine the resistance/susceptible phenotype of the interaction. Isolates of rust fungi obtained from annual surveys of the wheat, barley, and oat crops are used to inform the breeding process. Successful control of cereal rusts with host resistance cannot be achieved without knowledge of variation in cereal rust populations. Studies of virulence and molecular variation in cereal rust populations can answer questions that range from the applied, such as which host resistance genes are effective against the current rust population and what resistance genes are in current cereal cultivars, to more basic questions like what are the origins of new races and how do they spread. Discovery of the molecular determinants of pathogenesis and obligate biotrophy in cereal rust fungi via genomic approaches offers intriguing leads in the development of novel resistance mechanisms. Identification, characterization, and introgression of new host resistance to cereal rusts are key to increasing the diversity of resistance genes in our cereals and staying ahead of these "shifty" pathogens.


Approach
Cereal rust fungi are dynamic leading to constant changes in the U.S. population which leads to the erosion of effective rust resistance in cereal crops. In addition, the introduction of foreign isolates, such as Ug99, further threaten cereal production. Development of cereal cultivars with effective rust resistance and management strategies of these diseases will depend on the monitoring and characterization, virulence phenotypes and molecular genotypes, of cereal rust pathogen populations. Rust resistant cereal germplasm will be selected by testing wheat, oat, and barley lines from breeding programs throughout the United States for resistance to Puccinia coronata, P. graminis, and P. triticina, using the prevalent races, and races that have high virulence to rust resistance genes common in released cultivars and breeding lines. Testing with selected isolates of the cereal rust pathogens and host genetics studies will identify the rust resistance genes in breeding lines and germplasm. Advanced germplasm lines with combinations of rust resistance genes will be selected. Rust fungi produce a large arsenal of effector proteins in order to infect and colonize the plant host. Genetic and genomic approaches will be used to identify and characterize effector genes from P. graminis.


Progress Report
Progress was made on all three objectives. Annual survey of wheat stem and leaf rust, oat stem and crown rust, and barley leaf rust in the United States were completed. Survey samples were characterized for virulence phenotypes. Survey and race identification results were disseminated through an interactive map hosted at the ARS website located in St. Paul, Minnesota and through bi-weekly Rust Bulletins. Representative isolates from individual states were accessioned into ARS culture collection held in St. Paul. Stem rust sentinel plots were established in Texas, California and Arizona for the purpose of detecting potential Ug99 incursion into North America. Sentinel plots were also established in Ecuador. Close surveillance of the region is important to predict potential rust epidemics in North America. Drought in southern United States limited the number of samples collected from surveys and by collaborators. As a part of the effort contributing to global surveillance on Ug99, wheat stem rust samples from international collaborators were analyzed at the ARS. Three hundred and eighty isolates of leaf rust collected in 2014 were tested for virulence and race identity was determined. Fifty isolates collected in 2015 in Texas and Louisiana have been tested and race identity determined. Goal for 2015 will be completed in December 2015-January 2016, with additional testing of ca. 300 isolates from the Ohio Valley, Great Plains, and northeastern United States. Over 550 samples of the wheat stem rust pathogen was genotyped from 20 countries with a focus on samples from Africa, Middle East, and Europe. Genotyping of isolates from Ethiopian collections of wheat stem rust pathogen (race TKTTF) is part of a larger lineage found throughout the Middle East and Central Asia and Europe. In contrast to the Ug99 lineage, which appears to be formed rather recently based on low levels of genetic variation; the lineage containing isolates from the Ethiopian samples is complex and composed of multiple distinct subgroups indicating a much older lineage. A DNA based diagnostic assay was developed for rapid monitoring of wheat stem rust pathogen (race TKTTF) lineage. Twenty-one isolates of wheat leaf rust from Israel, 73 isolates from Chile, 27 isolates from Mexico, Spain, Tunisia, and Morocco, and 12 isolates from Pakistan were tested for virulence on differential set of Thatcher near isogenic lines. Isolates from Israel, Pakistan, Spain, and Chile had virulence to genes Lr17, Lr3bg and were avirulent to genes Lr2a and Lr28. Isolates with the identical virulence were previously found in the United States, South America, Europe, and the Middle East. These races were initially found in the United States in the mid 1990s and have since spread to many wheat growing regions worldwide. P. triticina isolates with high virulence to durum wheat that are avirulent to many genes in hexaploid wheat were collected and characterized from Chile, Mexico, Spain, Morocco, and Tunisia. Isolates with virulence to Lr14a were found in Chile, Spain, and Tunisia. Many durum wheat cultivars have Lr14a as the most important gene for leaf rust resistance. Increase of durum type isolates with virulence to Lr14a will erode the effective resistance of many cultivars worldwide. DNA will be extracted from a total of 80 selected isolates and SSR genotypes determined in the summer of 2015. A total of 759 isolates collected worldwide have been genotyped at 23 SSR loci and phenotyped with 20 Thatcher differential lines. The total population was analyzed using standard population genetic analysis. Rust infections on Berberis vulgaris were surveyed and collected from Minnesota and Wisconsin, and Puccinia graminis f. sp. secalis was identified and isolated. Native B. fendleri was surveyed in southern Colorado. Aecial infections on this species were not surveyed due to time constraint. The role of native barberry and Mahonia plants as potential sources for pathogen variations (generating new races) remains unknown except in northeastern Washington where Puccinia graminis undergoes active sexual cycle. Berberis x ottawensis, a naturally occurring hybrid between B. vulgaris and B. thunbergii in the New England states, was investigated for susceptibility to stem rust through inoculation experiments of 150 plants. Investigations in the role of barberry for stem rust variation in eastern Africa established that B. holstii is a functional alternate host for P. graminis in Ethiopia. Functional testing of P. graminis candidate effector gene alleles corresponding to Sr28 is underway. Progress continues to be made on developing a genetic map for P. graminis. Wheat breeding lines from the Northern Regional Performance Nursery (33 entries), Southern Regional Performance Nursery (42 entries), Uniform Southern Soft Red Winter Wheat (30 entries) Nursery and Uniform Eastern Soft Red Winter Wheat Nursery (31 entries), Uniform Hard Red Spring Wheat Nursery (31 entries), Gulf Atlantic (44 entries), Mason-Dixon Nursery (79 entries), Southern Uniform Wheat Nursery (79 entries) and 30 entries from the Kansas performance test, and 19 soft red winter wheat cultivars were evaluated for seedling leaf rust resistance with 10 different rust races. A total of 1600 wheat breeding lines from the uniform regional nurseries, preliminary regional nurseries, and individual breeding programs were tested with multiple stem rust races in greenhouse and in the field nursery. Data were provided to nursery coordinators and posted at ARS website. Oat crown rust nursery with close to 200 entries from across the United States were planted and disease screening data provided to coordinators and posted at the ARS website. Barley breeding lines were not tested as our collaborator at the University of Minnesota did the stem rust testing. Advanced and preliminary wheat breeding lines from the University of Minnesota were evaluated for leaf rust resistance in field plots. Hard red spring wheat entries in the Uniform spring wheat nursery were evaluated for seedling resistance with 10 leaf rust races and the leaf rust resistance genes were postulated for each entry. The wheat alloplasmic collections have seven different nuclear donor species ("euplasmic parents"): five T. aestivum cultivars (Chris, Selkirk, Chinese Spring, TVE, and Salmon), T. turgidum (durum), and T. timopheevii. All seven euplasmic parents were screened with the foreign stem rust races TTKSK (Ug99), TTTSK, JRCQC, and TRTTF to establish their susceptibilities. Seven domestic stem rust races were also tested. Selkirk was susceptible to nearly all races tested (except for JRCQC and one domestic race), while Chris was resistant to most races tested. Therefore, a subset of 40 alloplasmic lines, all with Selkirk nucleus, was screened with TTKSK, TTTSK, and TRTTF in replicates. No significant improvements to resistance were found in these alloplasmic lines. Seed for all of the alloplasmic lines were bulked in the greenhouse this winter (many must be maintained by crossing) to prepare for further disease screening efforts. Screening with the domestic races, followed by a more comprehensive screening with the foreign races continues. Substantial progress was made in understanding the genetics of resistance to Ug99 stem rust in the United States Spring wheat lines. A number of publications detail identified resistance loci and linked molecular markers. In addition to understanding the genetics of plant resistance, we coordinated the screening of 364 and 304 spring wheat breeding lines for screening at Ug99 field nurseries in Ethiopia and Kenya, respectively. These breeding lines were submitted by 8 public and private spring wheat breeders. Data on the reaction of advanced breeding lines was communicated to the breeders to allow for the selection of the United States wheat cultivars with Ug99 resistance. Also, 7 mapping populations were evaluated in Africa that segregate for quantitative resistance to Ug99. In collaboration with Kobe University in Japan, we also began working with an advanced T. dicoccum x T. dicoccoides population (BC2F11). Screening on the parental lines demonstrated that T. dicoccoides is highly susceptible to Ug99 (TTKSK) as well as three other foreign races (TTTSK, JRCQC, and TRTTF). On the other hand, T. dicoccum is susceptible only to JRCQC, but resistant to TTKSK, TTTSK, and TRTTF. The fact that T. dicoccum is susceptible to JRCQC rules out the possibility that the differences between the parental lines is solely conferred by the tetraploid wheat gene Sr13. Therefore, the population was screened with the three races for which T. dicoccum was resistant, while T. dicoccoides was susceptible. In all cases, segregation for resistance was found in the population. The identified gene(S) in this population can be introgressed into cultivated durum and bread wheat germplasm. F6 populations of Tc*3/Duster, Tc*3/Santa Fe, Tc/AC Taber, and Tc*3/CI13227 have been derived and tested for segregation of adult plant leaf rust resistance in greenhouse and field plot tests. The F6 populations have been genotyped with a 90K SNP chip. Genotype data is currently being processed to construct genetic a map of each population. The rust severity and response data from the field and greenhouse tests will be used with the maps to identify important genomic regions in each population. A RIL population of Tc*3/Bill Brown is being advance from F2 to F3 generation. The durable leaf rust resistance derived from the winter wheat cultivars Caldwell, CI13227, Santa Fe and Duster will be mapped, along with the resistance in the spring wheat AC Taber. RIL populations of Tc*2/Bill Brown F3 and Tc*2/Deliver F3 were advance from F2 to F3 generations.


Accomplishments
1. Wheat stem rust is a fungal disease of wheat that can significantly impact crop yield. A strain of the wheat stem rust fungus known as Ug99 emerged in Uganda in 1999, and threatens global wheat production for its ability to infect nearly all wheat varieties. Inadvertent introduction can severely impact wheat production in the United States. Recently new virulent races have been identified in Kenya by ARS scientists in St. Paul, Minnesota. A stem rust race belonging to the Ug99 race group, TTKSK, was detected for the first time in Egypt, and two new races (TTKTK and TTKTT) in this group were identified from samples collected in Kenya. These new races attacked the resistance gene SrTmp, an important stem rust resistance gene carried by several newly released cultivars in eastern Africa. These new virulence races explained continued stem rust epidemics in Kenya on newly released Ug99 resistant cultivars, and posed unrelenting threat to wheat production. Constant vigilance by ARS scientists is necessary to assure continued development of resistant wheat varieties for growers in the United States.

2. Wheat leaf rust is a fungal disease of wheat that can significantly impact crop yields, as documented in several outbreaks over recent years. New leaf rust races identified in the Great Plains region of the United States can lead to significant yield losses. In 2014 and 2015, races of P. triticina with virulence to major plant resistance genes, Lr21 and Lr39, were found by ARS scientists in St. Paul, Minnesota. Lr21 is found in many hard red spring wheat varieties grown in Minnesota and North Dakota and Lr39 is found in many hard red winter wheat varieties grown in Texas, Oklahoma and Kansas. Races with virulence to both genes will be able to attack and infect wheat over the entire length of the Great Plains region with the potential to cause significant yield loss in the winter and spring wheat regions. ARS scientists are working toward identifying more resistance genes in various wheat germplasm for introduction and use in development of newer wheat varieties that can withstand this devastating disease.

3. Wheat leaf rust is a fungal disease of wheat that can significantly impact crop yield, as has been documented in several outbreaks over recent years. Popular wheat cultivars in the southern Great Plains region of the United States are susceptible to new leaf rust races. Leaf rust races with virulence to major plant resistance genes, Lr11, Lr18, and Lr26, have increased in the eastern soft red winter wheat region and have caused higher levels of infection on cultivars such as Shirley that have Lr26 and Lr18. Leaf rust races with virulence to a major plant resistance gene Lr39 have increased in the hard red winter wheat area of the southern-mid Great Plains. These genes are present in many hard red winter wheat cultivars. Other races with virulence to genes Lr17, Lr24, and Lr26 are also common in the southern Great Plains region. Cultivars with these genes are susceptible to the common leaf rust races in this area. Monitoring of pathogen populations and identifying more plant resistance genes by ARS scientists is crucial to production of healthy wheat in the United States.

4. Wheat leaf rust is a fungal disease of wheat that can significantly impact crop yield, as has been documented in several outbreaks over recent years. A unique leaf rust race in Ethiopia has been identified. Collections of leaf rust isolates from Ethiopia in 2011, 2012, and 2013 were tested for virulence and genotyped with DNA markers. Three distinct forms or races of P. triticina were found in Ethiopia. One race type is virulent to common bread wheat and has virulence to major plant leaf rust resistance genes Lr3bg and Lr17, and has also been found in the US and other countries in Europe, South America, and the Middle East. This race type may have migrated from North America to other worldwide regions in the early 2000s. The second race type was collected from durum wheat and is avirulent to most resistance genes found in hexaploid common wheat. This race type has also been found on durum wheat in North America, South America, Europe and the Middle East, indicating the possible role of recent migration between the durum producing regions. The third race type is found only on tetraploid wheat in Ethiopia and is avirulent to common hexploid wheat. This race type has not been found in other wheat producing regions of the world. All three race types had highly distinct molecular genotypes. Monitoring the diversity, distribution and movement of leaf rust worldwide by ARS scientists is critical to continued development of resistant varieties for the growers in the United States.

5. Possible routes of introduction and migration of wheat leaf rust has been established. A total of 759 isolates of leaf rust from North America, South America, Central Asia, Europe, Russia, China, New Zealand and South Africa were tested for virulence to 20 plant resistance or Lr genes in Thatcher near-isogenic lines and for molecular genotype with 23 molecular markers. The worldwide populations were highly distinct for their marker genotype. Groups of isolates from the United States were related to isolates from South America, Europe, New Zealand, and the Middle East, which suggested possible routes of initial introduction of wheat leaf rust to North America and migration between populations in North America and other regions. Future monitoring of this disease from these regions will help guide researchers in developing resistant varieties before new virulent isolates impact the United States.

6. Alternate hosts of wheat stem rust are plants harboring the fungus and allowing sexual reproduction leading to greater variation in the pathogen population. A functional alternate host of wheat stem rust in Africa was established. Sustained efforts in field surveys of aecial infections on Berberis holstii and analyses of aecial samples led to the identification of P. graminis in aecial samples collected from Ethiopia in 2014, and established for the first time that B. holstii is a functional alternate host of P. graminis in Africa. This finding provides evidence as to the possible source of new variations, and possibly the virulent races, in the pathogen population. This finding allows ARS scientists to further analyze this source for new and more virulent forms of the pathogen in effort to develop resistant wheat cultivars.

7. Two Ug99 resistance genes were mapped in diploid wheat. The diploid relative of wheat, Triticum monococcum, has previously been used as a source of disease resistance. We identified the genomic location of a gene that provides resistance to Ug99 from T. monococcum and identified molecular markers linked to this resistance gene called SrTm4. In addition, resistance gene Sr21 from T. monococcum was previously characterized as ineffective to Ug99, but we demonstrated that this gene is effective to Ug99 at relatively high temperatures. We also identified the genomic location and linked molecular markers for Sr21. The two Ug99 resistance genes are both on chromosome arm 2A, approximately 38 cM apart. Mapping these genes and the identification of linked molecular markers will facilitate the breeding of Ug99-resistant wheat varieties to protect United States wheat production from yield losses caused by stem rust.

8. Single race stem rust screening nurseries established in Ethiopia. ARS researchers in St. Paul led an international effort to select wheat lines with field resistance to multiple virulent races of the stem rust pathogen. Four different virulent races of the stem rust pathogen were isolated in Ethiopia by Ethiopian Institute of Agricultural Research (EIAR) and ARS researchers. These races were used to inoculate four stem rust screening nurseries to evaluate 201 breeding lines and cultivars for field reaction to each race. Data were collected and distributed to collaborators at EIAR and CIMMYT. The development of these single race nurseries is significant because lines selected as resistant to Ug99 in Kenya were susceptible to a different race of the stem rust pathogen in Ethiopia in 2013 and 2014. Assessment of breeding lines with multiple virulent races is necessary for selecting wheat lines that are resistant to stem rust.


Review Publications
Kolmer, J.A. 2015. First Report of a wheat leaf rust (Puccinia triticina) phenotype with high virulence to durum wheat in the Great Plains region of the United States. Plant Disease. 99:56.
Kumssa, T.T., Baenziger, P.S., Rouse, M.N., Guttieri, M., Dweikat, I., Brown Guedira, G.L., Williamson, S.M., Graybosch, R.A., Wegulo, S.N., Lorenz, A.J., Poland, J. 2015. Characterization of stem rust resistance in wheat cultivar 'Gage'. Crop Science. 55:229-239.
Echeverry-Solarte, M., Kumar, A., Kianian, S., Simsek, S., Alamri, M.S., Mantovani, E.E., Mcclean, P.E., Deckard, E.L., Elias, E., Schatz, B., Xu, S.S., Mergoum, M. 2015. New QTL alleles for quality-related traits in spring wheat revealed by RIL population derived from supernumerary x non-supernumerary spikelet genotypes. Theoretical and Applied Genetics. 128:893-912.
Talajoor, M., Jin, Y., Wan, A.M., Chen, X., Bhavani, S., Tabe, L., Lagudah, E., Huang, L. 2015. Specificity of a rust resistance suppressor on 7DL in the spring wheat cultivar Canthatch. Phytopathology. 105:477-481.
Glover, K.D., Hall, R.G., Jin, Y., Osborne, L.E., Ingemansen, J.A., Turnipseed, E.B., Hareland, G.A. 2015. Registration of 'Advance' Hard Red Spring Wheat. Journal of Plant Registrations. 9:83-88.
Chen, S., Rouse, M.N., Zhang, W., Jin, Y., Akhunov, E., Wei, Y., Dubcovsky, J. 2015. Fine mapping and characterization of Sr21, a temperature-sensitive diploid wheat resistance gene effective against the Puccinia graminis f.sp. tritici Ug99 race group. Theoretical and Applied Genetics. 128:645–656.
Berg, J.E., Wichman, D.M., Kephart, K.D., Eckhoff, J.L., Stougaard, R.N., Lamb, P.F., Miller, J.H., Nash, D.L., Grey, W.E., Johnston, M., Gettel, D., Larson, R., Jin, Y., Chen, X., Bruckner, P.L. 2014. Registration of ‘WB3768’ wheat. Journal of Plant Registrations. 8:288–290.
Singh, R., Hodson, D., Jin, Y., Lagudah, E., Ayliffe, M.A., Bhavani, S., Rouse, M.N., Szabo, L.J., Pretorius, Z.A., Huerta-Espino, J., Basnet, B.R., Lan, C., Hovmøller, M.S. 2015. Emergence and spread of new races of wheat stem rust fungus: Continued threat to food security and prospects of genetic control. Phytopathology. 105(7):872-884.
Kolmer, J.A. 2015. A QTL on chromosome 5BL in wheat enhances leaf rust resistance of Lr46. Molecular Breeding. 35:74-81.
Kolmer, J.A. 2015. Collections of Puccinia triticina in different provinces of China are highly related for virulence and molecular genotype. Phytopathology. 105:700-706.
Olivera, P., Newcomb, M., Szabo, L.J., Rouse, M.N., Johnson, J.L., Hodson, D., Luster, D.G., Cox, J., Burgin, L., Gilligan, C., Patpour, M., Hovmoller, M., Woldeab, G., Hailu, E., Hundie, B., Tadesse, K., Pumphrey, M., Singh, R., Jin, Y. 2015. Phenotypic and genotypic characterization of race TKTTF of Puccinia graminis f. sp. tritici that caused a wheat stem rust epidemic in southern Ethiopia in 2013-2014. Phytopathology. 105(7):917-928.
Babiker, E.M., Bonman, J.M., Gordon, T.C., Chao, S., Newcomb, M.S., Rouse, M.N., Jin, Y., Wanyera, R., Acevedo, M., Brown Guedira, G.L., Williamson, S. 2015. Mapping resistance to the Ug99 race group of the stem rust pathogen in a spring wheat landrace. Journal of Theoretical and Applied Genetics. 128:605-612. doi:10.1007/s00122-015-2456-6.
Anderson, J.A., Wiersma, J.J., Linkert, G.L., Reynolds, S., Kolmer, J.A., Jin, Y., Dill-Macky, R., Hareland, G.A. 2015. Registration of 'Rollag' spring wheat. Journal of Plant Registrations. 9:201-207.
Maazaheri, M., Kianian, P., Mergoum, M., Valentini, G., Seetan, R., Pirseyedi, S., Kumar, A., Gu, Y.Q., Stein, N., Kubalakova, M., Dolezel, J., Denton, A., Kianian, S. 2014. Transposable element junctions in marker development and genomic characterization of barley. The Plant Genome. 7:1-8.