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ARS Home » Southeast Area » Raleigh, North Carolina » Plant Science Research » Research » Research Project #434206

Research Project: Genetic Improvement of Small Grains and Characterization of Pathogen Populations

Location: Plant Science Research

2019 Annual Report

1a. Objectives (from AD-416):
Objective 1. Identify and develop improved small grain germplasm with resistance to rusts, powdery mildew, Fusarium head blight, necrotrophic pathogens, and tolerance to freezing conditions during winter and spring. Sub-objective 1a. Develop wheat germplasm with resistance to stripe rust, leaf rust, stem rust, and powdery mildew. Sub-objective 1b. Develop wheat germplasm with resistance to Fusarium head blight (FHB). Sub-objective 1c. Develop wheat germplasm with resistance to Stagonospora nodorum blight (SNB). Sub-objective 1d. Identify oat, wheat and barley germplasm with tolerance to freezing. Objective 2. Develop improved methods of marker-assisted selection and genomic prediction, and apply markers in development of small grains cultivars. Sub-objective 2a. Identify and characterize new QTL for important traits in eastern winter wheat germplasm. Sub-objective 2b. Evaluate important traits in eastern winter wheat using molecular markers. Sub-objective 2c. Develop new eastern winter wheat germplasm using marker-assisted breeding and genomic selection. Objective 3. Develop new wheat and barley germplasm and cultivars having enhanced end-use characteristics for the eastern United States. Objective 4. Target resistance breeding efforts accurately by determining the relevant geographic variation in pathogen virulence profiles and the range of mycotoxin potential in pathogen populations. Sub-objective 4a. Determine the virulence frequencies and population structure in the wheat powdery mildew pathogen, Blumeria graminis f. sp. tritici, from different regions in the U.S. Sub-objective 4b. Identify and determine toxicological importance of minority Fusarium species causing FHB of wheat in North Carolina.

1b. Approach (from AD-416):
1a. Cross elite, adapted lines with sources of seedling and adult plant resistance to stripe rust, leaf rust, stem rust, and powdery mildew. Coordinate efforts to identify resistant lines in field breeding nurseries evaluated throughout the southeastern United States and in Njoro, Kenya (for Ug99). Evaluation with reliable molecular markers for known resistance genes. 1b. Continue use of inoculated, misted screening nurseries to evaluate regional and in-house breeding materials. Develop, evaluate and refine genomic selection models for scab resistance traits. 1c. Conduct appropriate phenotyping of regional and in-house breeding materials, including mapping populations, in inoculated Stagonospora blight nurseries to assist in locating the genes and associated markers to allow for marker-assisted selection. 1d. Select wheat and oat germplasm with superior resistance to freezing first by identifying genotypic differences in freezing patterns using IR technology. Crosses will be made with adapted cultivars to develop germplasm with improved resistance to freezing conditions. We will continue coordinating an oat and barley uniform nursery. 2a. Use sequencing based genotyping techniques to develop high-density genetic linkage maps of bi-parental mapping populations and association mapping populations as they are developed. Populations are phenotyped in conjunction with other unit scientists to identify regions of the genome involved in resistance to LR, YR, SR, PM, and SNB. 2b. Evaluate diverse germplasm with molecular markers linked to genes for pest resistance, agronomic and end-use quality, determine the level of marker polymorphism and the presence of favorable alleles in breeding lines. 2c. Apply MAS to introgress and pyramid new fungal resistance genes into eastern winter wheat germplasm. Genotype three-way cross and backcross F1s for populations entering into a doubled-haploid (DH) production pipeline. 2c. Use different parameters based on genomic position and linkage disequilibrium to select SNP sets that can be tested via cross-validation to identify the optimum number and most informative markers for GS. 3. Each year, approximately 600 crosses will be made to combine superior quality, yield, agronomic, and disease and insect resistance using recurrent parents from the program, as well as new sources of diversity. Utilize combinations of molecular markers with phenotypic selection and screening to accumulate favorable agronomic traits. Phenotyping and selection for improved hard wheats lines; grow and select populations under organic and conventional conditions. 4a. Samples will be gathered in each of two years from two to five states per region and derive single-pustule isolates; The phenotyping will be done in growth chambers using standard detached-leaf methodology. Population structure will be evaluated using molecular markers. 4b. Scabby wheat spikes will be collected from fields across a broad geographic range of North Carolina. Sequencing of the transcription elongation factor will be used to determine species. Population genetic analyses will determine if the Fusarium species are geographically clustered, or evenly distributed.

3. Progress Report:
New sources of resistance to wheat stripe rust have been developed in hard and soft wheat germplasm lines under Objective 1. ARS researchers used exotic sources and relatives of common wheat to cross into wheats adapted to the eastern United States. These new germplasm lines were confirmed to have stripe rust resistance in both Kenya and North Carolina. Combined disease resistance to leaf rust, stripe rust, stem rust, powdery mildew, and Fusarium head blight has been identified in wheat germplasm lines adapted to the eastern U.S., as well to the southern Great Plains. Under Objective 1, we tested 40 advanced lines at field locations in Texas, Louisiana, North Carolina, and Virginia to confirm the combination of multiple disease resistances in the lines. In addition, we identified the specific genes responsible for the resistances through use of genetic markers associated with each resistance trait. In October and November 2018 under Objective 1, ARS researchers evaluated 8,600 lines of wheat, barley, and triticale lines from breeding programs across all geographic and market classes of the crops in the United States and in Kenya to stem rust disease. Under Objective 1, an international team of researchers organized in 2018 began a detailed analysis of freezing in wheat with the aim of understanding age-dependant freezing in small grains that was discovered using infrared thermography. Protein and carbohydrate concentrations as well as the presence of bacterial and fungal populations and anatomical measurements were all investigated to better understand why older wheat leaves freeze before younger ones. This research could help explain why certain tissues in wheat never freeze and how they contribute to the hardiness of wheat as compared to less freezing tolerant species such as barley and oat. In addition, 30 barley and 21 oat experimental lines were evaluated at 10 and 15 locations respectively, worldwide. Data from cooperators is still being collected. Under Objective 2, DNA sequence based genotyping was done on approximately 10,000 samples for trait mapping and genomic selection. Exome capture and sequencing performed on 55 soft red winter wheat lines resulted in identification of approximately one million polymorphisms. High density genetic linkage maps were developed for four bi-parental soft red winter wheat mapping populations. Analysis identified loci affecting plant height, kernel size, leaf, stripe and stem rust resistance, as well as resistance to Septoria nordorum leaf and glume blotch. Germplasm in collaborative nurseries and breeding populations were genotyped with markers associated with 59 different genes of interest to breeders. Eight new marker assays were developed for genes conferring resistance to leaf, stem, and stripe rust, powdery mildew, and Septoria nordorum glume blotch. Significant progress was made under Objective 3 to develop high-quality bread wheat and malting barley for the eastern United States. ARS researchers did this in wheat by making 645 new crosses; evaluating 1,430 early-generation populations; selecting superior lines from 28,000 individual, unique rows; and yield-testing 1,600 lines. In barley, we made 110 new crosses; evaluated 95 early-generation populations; selected superior lines from 1,200 individual, unique rows; and yield-tested 120 lines. Grain from three of our experimental malting barley lines were requested by industry to be tested on a pilot-scale for malting quality. As part of Objective 4, a large study has revealed that so far, the U.S. wheat powdery mildew population does not appear to have key mutations that render it resistant to either of the two main classes of fungicides (demethylase inhibitors and quinone outside inhibitors, or strobilurins) that are used to manage wheat diseases here and worldwide. However, the study has shown that there are regional differences in sensitivity to DMIs, suggesting some erosion of efficacy. The basis for reduced DMI sensitivity has been investigated, and a mutation (Y136F) in the CYP51 gene has been detected in some isolates. Data on CYP51 expression and copy number have been obtained. Together, these data are allowing us to understand the current status of evolution to fungicide resistance in the U.S. wheat powdery mildew population. Also, a collaboration led to publication of characterization of Pm 65, a new powdery mildew resistance gene from the cultivar Xinmai 208 that is effective in several U.S. regions. In a collaboration between two USDA labs, ARS researchers are studying the Fusarium head blight-causing species from 59 wheat fields sampled across North Carolina’s wheat-growing area under Objective 4. Our research is demonstrating that while F. graminearum is the predominant FHB-causing species, other Fusarium species were found in the wheat heads of some fields, in some cases in high frequencies. These species produce mycotoxins different from those produced by F. graminearum.

4. Accomplishments
1. Identification of gene underlying awn suppression in wheat. Awns are stiff, hair-like structures that grow from the lemmas of wheat. Awns are thought to contribute to yield in warmer, drier environments, supplying carbohydrates to developing grain. However, awnless wheat varieties are the dominant type in many areas of the world, suggesting that absence of awns may be favored in some environments. Genes underlying the awnless trait in wheat had not previously been characterized. ARS scientists in Raleigh, North Carolina, identified a transcription repressor as the major gene responsible for awn inhibition in global wheat germplasm. This gene was also found to be associated with increased number of spikelets per spike, increased kernel number and decreased kernel size. This new study suggests that differences in grain size between awned and awnless wheats may be primarily due to changes in development, rather than differences in availability of photosynthate as previously assumed. Improved understanding of the relationship between awn development and control of spikelets per spike, grain number and grain size in wheat is providing new insight in to pathways that can be manipulated to increase grain yield.

2. Improved understanding of fungicide sensitivity in the wheat powdery mildew pathogen. Powdery mildew is one of the most common and damaging diseases in global wheat production. Blumeria graminis f. sp. tritici (Bgt), the fungus that causes the disease, can evolve rapidly and has developed high levels of resistance to fungicides used to manage it in Europe. It is important to understand current levels of Bgt sensitivity to commonly used fungicides such as demethylase inhibitors (DMIs, also known as triazoles) in the U.S. ARS researchers at Raleigh, North Carolina, conducted a large study of the U.S. wheat powdery mildew population and learned that eastern US Bgt populations are less sensitive to two common DMI fungicides, tebuconazole and propiconazole, than central US Bgt populations. This suggests Bgt has evolved to reduced sensitivity in the eastern states, which have more frequent powdery mildew epidemics and fungicide applications to wheat. The study highlights the need to conserve the remaining efficacy of DMIs, which are one of two main chemistry classes used to control all wheat diseases in the U.S.

3. Identification of stem rust resistance in soft red winter wheat. Stem rust of wheat, caused by Puccinia graminis F. sp. tritici, has historically impacted bread wheat yields worldwide and is generally managed through genetic resistance in deployed cultivars. The stem rust race-group Ug99 represents a re-emergence of this threat that is virulent to most wheat varieties grown in the United States. Having strong resistance to this new race of stem rust in U.S. cultivars is important to reduce the potential of Ug99 stem rust spreading and establishing in new wheat growing regions. ARS scientist at Raleigh, North Carolina, identified a soft red winter wheat breeding line having reduced stem rust symptoms at the adult stage in field nurseries in Kenya. Genetic analysis identified a gene located on the short arm of chromosome 6D, contributing to significantly reduced stem rust symptoms. A DNA marker predictive for the presence of the resistance gene was developed. This resistance has not been previously reported in Eastern U.S. breeding programs. The germplasm and associated DNA markers are being used to develop wheat cultivars with polygenic resistance to the Ug99 race-group.

4. Resistance to ozone damage identified in wheat. Ground-level ozone is an air pollutant and a greenhouse gas that has increased in ambient air since the industrial era. Wheat is sensitive to ozone and loses yield and grain quality when exposed to elevated ozone amounts. In order to breed for resistance to ozone, we must identify the location of ozone resistance in wheat. We found several genetic locations for the trait, including one specific chromosome that had a large effect on resistance, and several other locations that had smaller effects in reducing ozone damage. The locations having smaller effects appeared to be associated with some resistance genes to an unrelated disease, named leaf rust. This research allows ARS scientists to identify the correct parents to use in crosses for breeding for ozone resistance and provides information to pinpoint genes for resistance. This research by ARS scientists in Raleigh, North Carolina, can lead to reducing some of the estimated annual 7-12% global yield loss in wheat caused by ozone damage.

5. Improved understanding to freezing during reproductive phase of growth in wheat. Freezing weather during spring can devastate winter cereal crops when they are in the reproductive phase of growth. An ARS Scientist in Raleigh, North Carolina, developed a procedure for evaluating large numbers of wheat varieties for their ability to withstand an unexpected freeze in the spring. The procedure confirmed that contrary to existing literature there are genetic differences in spring freeze tolerance between wheat genotypes. Use of this new procedure will allow researchers to evaluate to populations to determine the genetic basis of spring-freeze tolerance.

Review Publications
Huang, M., Ward, B., Van Sanford, D., McKendry, A., Brown Guedira, G.L., Tyagi, P., Sneller, C. 2018. The accuracy for genomic prediction between environments and populations for soft wheat traits. Crop Science. 58:1-15.
Huang, M., Mheni, N., Brown Guedira, G.L., McKendry, A., Griffey, C., Vansanford, D., Costa, J. 2018. Genetic analysis of heading date in winter and spring wheat. Euphytica. 214:128.
Case, A.J., Bhavani, S., Macharia, G., Pretorius, Z., Coetzee, V., Kloppers, F., Tyagi, P., Brown Guedira, G.L., Steffenson, B.J. 2018. Mapping adult plant stem rust resistance in barley accessions Hietpas-5 and GAW-79. Theoretical and Applied Genetics. 131:2245-2266.
Daba, S., Tyagi, P., Brown Guedira, G.L., Mohammadi, M. 2018. Genome-wide association studies to identify loci and candidate genes controlling kernel weight and length in a historical United States wheat population. Frontiers in Plant Science. 9:1045.
Beyer, S., Daba, S., Tyagi, P., Bockelman, H.E., Brown Guedira, G.L., Mohammadi, M. 2019. Loci and candidate genes controlling root traits in wheat seedlings—a wheat root GWAS. Functional and Integrative Genomics. 19:91-107.
Ward, B.P., Brown Guedira, G.L., Kolb, F.L., Van Sanford, D.A., Tyagi, P., Sneller, C.H., Griffey, C.A. 2019. Multi-environment and multi-trait genomic selection models in unbalanced early generation wheat yield trials. Crop Science. 59:491-507.
Sarinelli, J.M., Murphy, J.P., Tyagi, P., Holland, J.B., Johnson, J.W., Mergoum, M., Mason, R.E., Babar, A., Harrison, S., Sutton, R., Griffey, C.A., Brown Guedira, G.L. 2019. Training population selection and use of fixed effects to optimize genomic predictions in a historical USA winter wheat panel. Theoretical and Applied Genetics. 132:1247.
Ward, B.P., Brown Guedira, G.L., Kolb, F.L., Van Sanford, D.A., Tyagi, P., Sneller, C.H., Griffey, C.A. 2019. Genome-wide association studies for yield-related traits in soft red winter wheat grown in Virginia. PLoS One.
Li, G., Carver, B., Cowger, C., Bai, G., Xu, X. 2018. Pm223899, a new recessive powdery mildew resistance gene identified in Afghanistan landrace PI 223899. Theoretical and Applied Genetics. 131(12):2775-2783.
Niu, Z., Chao, S., Cai, X., Whetten, R.B., Breiland, M., Cowger, C., Chen, X., Friebe, B., Gill, B.S., Rasmussen, J.B., Klindworth, D.L., Xu, S.S. 2018. Molecular and cytogenetic characterization of six wheat-Aegilops markgrafii disomic addition lines and their resistance to rusts and powdery mildew. Frontiers in Plant Science.
Tan, C., Li, G., Cowger, C., Carver, B.F, Xu, X. 2019. Characterization of Pm63, a new powdery mildew resistance gene identified in Iranian landrace PI 628024. Journal of Theoretical and Applied Genetics. 132(4):1137-1144.
Li, G., Xu, X., Tan, C., Carver, B.F., Bai, G., Wang, X., Bonman, J.M., Wu, Y., Hunger, R., Cowger, C. 2019. Identification of powdery mildew resistance loci in wheat by integrating genome-wide association study (GWAS) and linkage mapping. The Crop Journal. 7(3):294-306.
Beccari, G., Arellano, C., Covarelli, L., Tini, F., Sulyok, M., Cowger, C. 2019. Effect of wheat infection timing on fusarium head blight causal agents and secondary metabolites in grain. International Journal of Food Microbiology. 290:214-225.
Cowger, C., Arellano, C., Marshall, D.S., Fitzgerald, J. 2019. Managing fusarium head blight in winter barley with cultivar resistance and fungicide. Plant Disease.
Kimball, J., Tuong, T.D., Arellano, C., Livingston, D.P., Milla-Lewis, S.R. 2017. Freeze-Testing in St. Augustinegrass II: Evaluation of acclimation effects. European Journal of Agronomy. 213:282-286.
Dunne, J., Tuong, T.D., Livingston, D.P., Reynolds, C., Milla-Lewis, S. 2018. Field and laboratory evaluation of Bermudagrass (Cynodon spp.) germplasm for cold hardiness and freezing tolerance. Crop Science. 59:392-399.
Livingston, D.P. 2018. Investigating freezing patterns in plants using infrared thermography. In: Iwaya-Inoue M., Sakurai M., Uemura M. (eds) Survival Strategies in Extreme Cold and Desiccation. Advances in Experimental Medicine and Biology, vol 1081. Springer, Singapore. Book Chapter. p. 117-127.
Kimbal, J., Tuong, T.D., Arellano, C., Livingston, D.P., Milla-Lewis, S. 2018. QTL linkage analysis and identification of quantitative trait loci associated with freeze tolerance and turf quality traits in St. Augustinegrass. Molecular Breeding. 38:67.
Livingston, D.P., Tuong, T.D., Hoffman, M., Fernandez, G. 2018. Protocol for producing three-dimensional infrared video of freezing in plants. Journal of Visualized Experiments. 139:e58025.
Livingston, D.P., Tuong, T.D., Noguiera, M., Sinclair, T.R. 2019. 3-D reconstruction of soybean nodules provides an update on vascular structure. American Journal of Botany. 106:507-513.