Location: Cereal Disease Lab2013 Annual Report
1a. Objectives (from AD-416):
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.
1b. Approach (from AD-416):
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.
3. Progress Report:
Progress was made on all three objectives, which are under National Program 303: Component 1, Disease Diagnosis and Etiology; Component 2, Biology and Epidemiology of Plant Disease; Component 3, Plant Disease Management. Under objective 1, 2013 annual survey of wheat stem and leaf rust, oat stem and crown rust, and barley leaf rust in the United States were completed. Samples from the 2012 annual survey were characterized for virulence phenotypes. In the central wheat growing areas of the U.S. QFCSC remained the dominant race of the wheat stem rust pathogen. MCCDC was the only other race of the wheat stem rust pathogen identified in this region. In the Pacific Northwest more than 60 races of the stem rust pathogen were identified from wheat and barley collections where a moderate stem rust epidemic occurred. More than 50 races of the wheat leaf rust pathogen was found in the 2012 sample set and the most common races have virulence to resistance genes present in the leading winter wheat cultivars. Virulence phenotypes of the 2012 survey samples of crown rust pathogen were similar to what was observed in 2011. Differential responses were observed on host lines possessing resistance genes Rph3, Rph7, Rph5, and Rph10 in the 2012 barley leaf rust samples. Genotyping using SSR markers of global populations of wheat leaf rust pathogen has continued. Validation of a SNP chip for wheat stem rust pathogen was completed and representative isolates from the United States, Middle East and Africa were analyzed. Samples of the Ug99 race group represent a distinct genetic group. Sixty isolates of stem rust pathogen from aecial collections on Mahonia repens and M. aquifolium were characterized for virulence on a standard set of wheat and barley lines. Under objective 2, a partial high-density genetic map of the wheat stem rust pathogen was developed. Using this genetic map, a candidate effector gene was identified for Sr28. Under objective 3, wheat and oat lines from United States national and regional nurseries, and individual breeding programs were evaluated for rust resistance in seedling greenhouse tests and field plots. Adult plant wheat leaf rust resistance was mapped and indicated that Lr23, Lr46 and Lr68 were likely present in variety Ulen. The adult plant leaf rust resistance in the Tc*3/Caldwell population was mapped to chromosome 3B and is likely a new wheat leaf rust resistance gene. Markers linked to wheat stem rust resistance gene Sr44 were identified and this alien gene was moved into a wheat line, facilitating the use of Sr44 in plant breeding programs. New stem rust resistance was described from Aegilops tauschii and introgressed into bread wheat. Progress was made on describing the genetics of adult plant resistance in adapted United States hard red spring wheat through phenotyping for Ug99 resistance in Kenya and Ethiopia nurseries. Candidate genes were identified for the Ug99 resistance conferred by wheat line Gabo 56.
1. Host specialization in the wheat leaf rust fungus is correlated with wheat evolution. Distinct forms of the leaf rust fungus Puccinia triticina are found on common hexaploid wheat with the ABD genomes; durum wheat with the AB genomes, and Aegilops speltoides with the B genome. The objectives of this research were to determine if P. triticina evolved to different forms of wheat in a manner that was consistent with the development of modern wheat. DNA sequence data of isolates from different hosts indicated that P. triticina from a wild grass – Aegilops speltoides that is a wild wheat relative was the first form of the rust, followed by isolates from durum wheat in Ethiopia. A form of the rust that occurs on durum wheat in Europe, Mexico, California, South America, and the Middle East evolved very recently from the group of isolates that occurs on common wheat that is found worldwide, the most recently derived form is of isolates from common wheat. The evolutionary relationship between the different forms of P. triticina were congruent with the history of wheat evolution. These results show that P. triticina can quickly adapt to different types of wheat very quickly. These results provide further insight into the evolutionary processes of pathogen adaptation to important crops.
2. Development of a molecular diagnostic assay for the Ug99 race group of the wheat stem rust pathogen. Ug99 race group of the wheat stem rust pathogen is a current threat to wheat crop in the United States and around the world. Standard methods for identification of this race group requires characterizing live samples on a standard set of wheat plants under containment conditions. A two-stage assay based on polymorphisms in target regions of the pathogen's DNA was developed. The first stage assay determines if the sample belongs to the Ug99 race group. The second stage assay confirms the results of the first stage and predicts the specific race phenotype based on genotype at 15 independent loci. This assay is currently being used to track the movement of Ug99 race group in Africa. The use of DNA based diagnostic assay for Ug99 race group allows for rapid identification and does not require working with living cultures. The deployment of this assay in regional plant pathogen diagnostic laboratories in the United States will greatly enhance the ability to identify introductions into the United States of the Ug99 race group of the wheat stem rust pathogen. Adoption of this assay in regional diagnostic labs is dependent upon coordination between ARS and APHIS, which is currently being discussed. Deployment of this assay in regional diagnostic labs will allow for the rapid detection of presence and spread of Ug99 if it arrives in the United States. This will allow for timely mitigation procedures to be made in order to prevent Ug99 from establishing in the United States. Preventing the establishment of Ug99 in the United States would prevent losses in wheat production to this dangerous disease
3. Determined the first gene sequence of a Ug99 resistance gene. Resistance gene Sr35 provides near-immunity to avirulent stem rust races such as the Ug99 race group. The gene was initially mapped in both hexaploid and diploid wheat. Screening of resistant and susceptible T. monococcum accessions with and without Sr35 combined with sequencing of these accessions allowed for the identification of the Sr35 gene sequence. Once the sequence was identified and cloned, susceptible wheat lines were made to be resistant by inserting the Sr35 gene, confirming the resistance function. ARS also collaborated with Australian scientists to verify the gene sequence of the Sr33 resistance gene. Obtaining the Sr35 and Sr33 gene sequences is significant for two reasons: 1. The gene sequences open up the possibility for further experiments to identify the molecular mechanisms of Ug99 stem rust resistance in wheat. Understanding the molecular mechanisms will guide gene deployment in agriculture leading to enhanced resistance durability. 2. The gene sequences allow for the use of transgenic wheat to provide resistance to Ug99 stem rust. Since backcrossing stem rust resistance genes into modern cultivars takes years and slows down breeding progress, the transgenic option could speed up the resistance breeding and also allow for multiple genes (such as Sr33 and Sr35) to be stacked together in single wheat lines to enhance resistance durability.
Carlson, G.R., Berg, J.E., Kephart, K.D., Wichman, D.M., Lamb, P.F., Miller, J.H., Stougaard, R.N., Eckhoff, J.L., Riveland, N.R., Nash, D.L., Grey, W.E., Jin, Y., Kolmer, J.A., Chen, X., Bai, G., Bruckner, P.L. 2013. Registration of ‘Judee’ wheat. Journal of Plant Registrations. 7:191-194.