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United States Department of Agriculture

Agricultural Research Service

Research Project: RESPONSE OF DIVERSE RICE GERMPLASM TO BIOTIC AND ABIOTIC STRESSES
2009 Annual Report


1a.Objectives (from AD-416)
The long-term objective of this project is to better understand rice responses to pests, pathogens, and weather stress in the environment, and to use that information to enhance pest protection and production efficiency for a more sustainable U.S. rice production. Over the next 5 years we will focus on the following objectives: Objective 1: Map rice genes associated with resistance to sheath blight and blast diseases and identify sources of resistance to kernel and false smut diseases. Sub-objective 1.A. Map candidate genes for sheath blight resistance in rice. Sub-objective 1.B. Develop high-resolution genetic maps of Rhizoctonia solani phytotoxin. Sub-objective 1.C. Elucidate recognition mechanisms of the rice blast resistance gene, Pi-ta, to the pathogen avirulence gene AVR-Pita. Sub-objective 1.D. Identify sources of resistance to grain quality reducing diseases: false smut and kernel smut. Objective 2: Identify the physiological, environmental, and genetic factors associated with tillering and seedling vigor under cold temperatures in rice. Sub-objective 2.A. Identify environmental and cultural factors that induce early tillering in indica germplasm and identify early tillering QTL in mapping populations. Sub-objective 2.B. Identify genomic regions associated with cold temperature stress at the seedling stage. Objective 3: Develop chromosome segment substitution lines (CSSLs) and advanced backcross mapping populations using selected Oryza wild species to study the chromosomal location of grain shape, pest resistance, and domestication traits. Sub-objective 3.A. Introgress novel sheath blight resistance genes into U.S. rice cultivars using Oryza wild species accessions. Sub-objective 3.B. Exploring transgressive variation in rice. Objective 4: Identify growth factors associated with changes in weed species populations in response to the use of herbicide–resistant rice cultivar technology and other non-conventional cultural management practices. Sub-objective 4.A. Determine the efficacy of weed-suppressive rice in reduced input rice production systems. Sub-objective 4.B. Quantify gene flow of herbicide resistance genes in commercial rice production.


1b.Approach (from AD-416)
Genetic markers associated with QTLs linked to sheath blight resistance, sheath blight toxin sensitivity, and tillering will be identified in various mapping populations. Sequence diversity for the Pi-ta blast resistance gene in several species of rice and of the avirulence gene in the pathogen will be determined. Interactions of predicted host and pathogen proteins will be used to identify critical amino acid residues important for disease resistance. Cultivars and parents of existing mapping populations will be evaluated to identify sources of resistance to kernel and false smuts. The impact of different tillage methods, fertilizer rates, and crop rotation systems on incidence of smut will be determined to give cultural management recommendations to farmers. To identify QTL underlying tillering in rice, we will identify growth conditions including include soil temperature, planting depth, and fertilizer rates, sources, and timing that maximize the phenotypic differences between high- and low-tillering genotypes. Germplasm lines will be evaluated for coleoptile emergence under cold temperatures (11oC) to identify sources that can be used to develop mapping populations. Novel sources of sheath blight resistance identified in wild species of rice (i.e. O. meridionalis, O. nivara, O. rufipogon) will introgressed into a susceptible Southern U.S. cultivar through backcrossing. Putative sheath blight resistance QTL will be verified using inoculated field tests and greenhouse toxin assays. The O. rufipogon wild species of rice and several O. sativa sub-populations will be used to identify adapted gene complexes responsible for positive transgressive variation. Chromosome segment substitution lines and near isogenic lines (NILs) will be used to systematically explore the relationship between diversity and transgressive variation. In addition, lines containing O. rufipogon introgressions that alter flowering time, grain size and weight, and number of grains per plant will be analyzed to determine the impact of the introgressions on agronomic traits. High-tillering indica rice lines and commercial hybrids will be evaluated to determine if they have sufficient weed-suppression capabilities when coupled with low rates of herbicide and/or alternative production systems that result in effective weed control. Competitive interactions between rice and barnyardgrass (C4 weed species) will be assessed using 13C isotope depletion analyses of roots extracted from soil core samples. Alternative cultural practices including early planting, reduced irrigation, and decreased seeding rates will be evaluated for their savings in water use and impact on weed control. Reciprocal outcrossing rates between commercial hybrid rice cultivars and common U.S. red rice biotypes will be investigated to determine the likelihood of herbicide resistance gene flow. Putative outcrosses will be verified using herbicide screening when herbicide resistant cultivars serve as the male or assessment of unique plant characteristic and genetic markers when non-herbicide resistant rice serves as the male.


3.Progress Report
The 250 LJ F5 recombinant inbred lines (RIL) were used to identify 10 sheath blight resistant QTLs including the major effect QTL-qSB9-2 in FY09. A total of 1000 individuals of BC2F2 populations will be produced for genotyping using polymorphic SSRs and 100 SNPs spanning the qSB9-2 region to identify molecular recombinants to delimit the chromosomal region for ShB resistance and for selecting identified candidate genes, including three ABC transporters in FY10. Several candidate genes from rice, including an ethylene responsive transcription factor, were identified in FY09, and are being verified for their interactions and should be completed in FY10;.
2)Recombinant AVR-Pita in several US virulent isolates was shown to trigger resistance on rice cultivars carrying Pi-ta resistance genes in FY09. The effects of recombinant AVR-Pita for infection on rice cultivars without Pi-ta will be examined in FY10;.
3)Genomic region harboring Ptr(t) was delimited within a 50-kb region on chromosome 12 in FY09, and these regions are being sequenced for candidate gene identification. These candidate genes will be verified in FY10;.
4)DNA markers for P42(t)and Pi43(t) were identified in FY09 and these markers with previously identified markers for Pi-z, Pi-ta, Pi-k will be used to characterize rice germplasm in FY10; and.
5)a mapping population of Nipponbare and 93-11 (N9) was developed in FY09 and is being analyzed for yield components, and results will be repeated in FY10. Tissue was harvested from 2 warm-sprouting and 3 cold-sprouting lines of rice near-isogenic to each other. The tissue has been extracted for mRNA and is awaiting hybridization for analysis by microarray. A BC2F2 mapping population developed from a cross between Bengal/O. nivara (IRGC 104705) was evaluated for reaction to sheath blight disease in the greenhouse using the micro-chamber method and in the field. The set of 400 rice accessions was genotyped with a novel 1,536 SNP panel using the Illumina GoldenGate SNP assay to reveal the population substructure and identify introgressions for plant stature, blast resistance and amylose content. This set of rice accessions is now in the process of being genotyped with a newly designed 44K SNP assay. With cooperators at Texas A&M Univ., showed that the weed-suppressive indica cultivar 'TN1' suppressed water weevil infestation in roots and yielded well even without insecticide. In a field study with Univ. of Arkansas cooperators, outcrossing from a hybrid cultivar to red rice was greater than from an inbred cultivar, and was favored by elevated late-morning relative humidity.


4.Accomplishments
1. Finding the Gene Associated with Sheath Blight Disease Resistance in Rice: Sheath blight disease causes yield losses in rice production areas around the world. It has been difficult to develop sheath blight-resistant cultivars because there are no known sources of complete resistance and resistance is conditioned by many genes that are modulated by the growing environment. ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified DNA markers associated with major sheath blight disease resistance QTLs in a project partially funded by the USDA-CSREES-NRI RiceCAP grant. A newly developed greenhouse screening method for evaluating sheath blight resistance was used to evaluate a mapping population segregating for reaction to the sheath blight pathogen. A total of 10 QTLs on several rice chromosomes were identified. A major QTL for sheath blight resistance was found on chromosome 9. It is anticipated that use of these markers linked to the major sheath blight-resistant QTL will allow breeders to accelerate the introduction of genetic resistance into new cultivars.

2. Characterizing Blast Resistance Genes in US Rice Germplasm Collection: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified blast resistance genes in the US rice germplasm collection. Rice blast disease is a serious threat to rice production in the US and worldwide due to constant development of more virulent races of the pathogen. The Pi-ta gene in rice has been effectively deployed in the US for preventing infection caused by most races. However, race IE-1k overcomes the Pi-ta-mediated resistance and has been frequently recovered from commercial rice fields. A total of 1800 rice accessions were analyzed for the presence of various blast resistance genes using DNA markers and disease evaluation. Among them, diverse accessions were found to possess the Pi-ta resistance gene and others appeared to possess new genes for resistance. These rice accessions will be important to breeding programs for blast resistance in US and worldwide.

3. Genetic Variability Identified at the Pi-ta Locus: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified 12 variants in a disease resistance gene. The Pi-ta gene in rice confers resistance to races of the blast pathogen that contain the corresponding AVR-Pita avirulence gene. Understanding genetic regulation at the Pi-ta locus will benefit the development of durable resistance to this plant disease. Genetic variation in the Pi-ta gene was identified that affected expression of the resistance gene. This study demonstrates that the Pi-ta gene in rice may produce complex isoforms that have profound significance in the evolution of plant resistance genes.

4. Evolutionary Mechanisms of Pathogenicty in the Rice Blast Pathogen: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified molecular mechanisms of disease susceptibility by studying the pathogen that causes rice blast. The AVR-Pita1 gene in the blast pathogen Magnaporthe oryzae is necessary to elicit a resistant response in rice plants that possess the Pi-ta resistance gene. Over 150 blast isolates from the field were characterized for DNA sequence variation in the AVR-Pita1 gene and were tested for their ability to cause disease. By analyzing DNA sequence variation of the AVR-Pita1 gene it was found that many changes occurred in expressed regions of the gene that allow the pathogen to cause disease on a diversity of rice cultivars. This suggests that this gene has evolved under intensified selection pressure, allowing it to continue to cause disease in rice. This information will help researchers better understand the fundamentals of plant-pathogen interactions that will lead to better control of crop disease.

5. One of the Largest Genetic Linkage Blocks Identified in Rice: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, have identified a large linkage block in rice associated with a disease resistance gene. Studying the size of genomic introgressions will lead to a better understanding of genetic linkage and how it impacts crop breeding. Using a rice mapping population, genomic fragments ranging from half to the entire chromosome were found around Pi-ta blast resistance gene. Large segments of the Pi-ta genomic region originating from the rice variety Tetep from Vietnam were in the Pi-ta containing US rice cultivars Katy, Madison, Kaybonnet, and Drew. The entire chromosome 12, which contains Pi-ta, was found to be identical in Tetep and the cultivar Tadukan from the Philippines. In addition, IR64, the most widely grown rice cultivar in the world, was found to be genetically the same as these for a very large region around Pi-ta. These findings suggest that a large linked set of genes is present as a super locus on chromosome 12 and is necessary for blast resistance. This information will help breeders as they develop new rice cultivars that are resistant to this global pathogen.

6. Tracking Evolutionary Changes in Disease Resistance in Rice: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified molecular mechanisms of evolution of disease resistance genes. The Pi-ta blast resistance gene in rice can detect a signaling molecule of the pathogen, thus eliciting a resistance response by the plant that prevents disease development. To understand the evolutionary changes around this disease resistance gene, DNA sequences of the Pi-ta locus in 159 rice accessions composed of seven species (O. sativa, O. rufipogon, O. nivara, O. meridionalis, O. glaberrima, O. barthii, and O. glumaepatula) were analyzed in a study partially supported by the National Science Foundation. The genetic fingerprint of the Pi-ta gene in cultivated rice, O. sativa, was demonstrated to have originated from the weedy species O. rufipogon. These findings demonstrated that Pi-ta evolved under extensive selection pressure and genetic variation within specific sites of this gene impact disease resistance.

7. Identification of genes controlling plant defense to the sheath blight toxin: Sheath blight disease in rice is caused by Rhizoctonia solani, a pathogen that attacks many crops worldwide. The pathogen produces a toxin that kills the plant, reducing grain yield. Researchers at the Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, have isolated a phytotoxin from the sheath blight pathogen and have demonstrated that plant sensitivity to it is correlated with sheath blight susceptibility. Using a rice mapping population that is segregating for sheath blight susceptibility, a region on chromosome 7 was identified as associated with sensitivity to the toxin. This research has demonstrated a new means of evaluating rice germplasm for a genetic component that is important in the plant's defense response to the sheath blight pathogen and will aid breeders in developing disease resistant cultivars.

8. New methods for controlling grain smuts in rice: False smut and kernel smut are two diseases found in rice that dramatically reduce grain quality. Because of the sporadic occurence of these diseases, little is known about control methods. Researchers at the Dale Bumpers National Rice Research Center, Suttgart, Arkansas, in cooperation with the University of Arkansas, have identified rice cultivars resistant to kernel smut and false smut. These newly identified sources of smut resistance can be used to identify the corresponding disease resistance genes that can be used by breeders to develop improved cultivars.

9. Identification of rice cropping systems that eliminate false smut disease in rice: False smut is a rice disease that develops on the seed head and can reduce yield and quality. Little is known about methods to control this disease. At the Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, in cooperation with the University of Arkansas, the effects of reduced (upland) irrigation on smut severity were evaluated. Upland irrigation was found to eliminate false smut from susceptible cultivars, but had no effect on kernel smut. These findings demonstrate a cultivation practice that can be used by growers to successfully manage this disease without using fungicides.

10. Discovery of novel sheath blight resistance QTL from weedy rice species: Rice sheath blight disease causes significant economic damage to rice production worldwide, but no source of complete resistance to this disease has been identified in cultivated rice. Researchers at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, identified moderate resistance to sheath blight disease in an accession of Oryza nivara, a wild ancestral species of cultivated rice. Subsequently, the chromosomal location of three possible sheath blight resistance genes (QTL) on chromosomes 2, 6, and 11 contributed by O. nivara were identified, and efforts are underway to transfer this resistance into the popular Southern U.S. medium grain variety, Bengal. Ultimately, these adapted progeny lines with improved sheath blight resistance will be made available to rice breeders to develop superior sheath blight resistant varieties for the U.S. rice industry.

11. Exploring the agronomic and seed traits among five genetic sub-groups in rice: Substantial increases in rice yield have been obtained in hybrid rice developed from crosses between the two major rice sub-species identified as indica and japonica. These two groups have been divided into five different sub-groups (indica, aus, aromatic, temperate, and tropical japonica) using genetic markers. Researchers at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, evaluated the agronomic and seed characteristics of 400 rice accessions from around the world based on the five sub-groups and found that each group was defined by a particular set of agronomic and seed traits. This study will be used to further exploit the variation available in the five rice sub-groups to attempt to achieve even greater increases in yield and improved agronomic performance in new rice varieties under development. This study is part of a National Science Foundation funded collaborative research project with Cornell University.

12. Genetic diversity within weedy red rice and its impact on geneflow: ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, demonstrated that a small number of diverse weedy red rice accessions in the southern U.S. exhibit a history of low-level gene exchange with other red rice biotypes and cultivated rice, and that genetic differences distinguish weedy red rice accessions collected from various geographic regions. Thirty-one SSR markers were used to analyze approximately 50 U.S. red rice accessions in comparison to more than 100 international and national rice cultivars. Knowledge of the genetic diversity within U.S. red rice populations and the gene exchange potential between red rice and cultivated rice will be useful in the understanding and prevention of gene movement between herbicide-resistant cultivars and red rice biotypes prevalent in the southern U.S. and will extend life of herbicide-resistant rice technology.

13. A new method to quantify rice-weed root interactions in the field: Identifying weed-suppressive rice cultivars that can out-compete troublesome weeds such as barnyardgrass has been difficult due to the intertwining growth of roots in the field. ARS scientists at Dale Bumpers National Rice Research Center in Stuttgart, Arkansas, demonstrated that a new method that accurately determines the amount and distribution of barnyardgrass and rice roots in soil using a naturally occurring stable isotope of carbon (C-13). C3 plants (e.g., rice) and C4 plants (e.g., barnyardgrass) have different photosynthetic pathways for carbon fixation. Thus, 13C isotope depletion analysis can be used to estimate the proportion of rice and barnyardgrass roots present in soil samples. The method proved to be reliable over several years and environments. A weed-suppressive cultivar, PI 312777, produced consistently greater root growth and allowed less barnyardgrass root growth in comparison to a typical U.S. rice cultivar. Having a method that can evaluate interactions between rice and weed roots under the soil surface will help in the development of weed-suppressive rice cultivars that will provide farmers with an additional tool that can help control this damaging pest while minimizing their input costs.


6.Technology Transfer

Number of the New/Active MTAs (providing only)1
Number of Invention Disclosures Submitted1

Review Publications
Gealy, D.R., Bryant, R.J. 2009. Seed physicochemical characteristics of field-grown U.S. weedy red rice biotypes: Contrasts with commercial cultivars. Journal of Cereal Science. 49:239-245.

Prasad, B., Eizenga, G.C. 2008. Sheath blight disease screening methods to identify resistant Oryza spp. accessions. Plant Disease. 92:1503-1509.

Jia, Y., Lee, F.N., McClung, A.M. 2009. Determination of resistance spectra of the Pi-ta and Pi-k genes to U.S. races of Magnaporthe oryzae causing rice blast in a recombinant inbred line population. Plant Disease. 93(6):639-644.

Jia, Y., Liu, G., Costanzo, S., Lee, S., Dai, Y. 2009. Current progress on genetic interactions of rice with rice blast and sheath blight fungi. Frontiers of Agriculture in China. 3(3):231-239.

Huang, L., Brooks, S.A., Li, W., Fellers, J.P., Nelson, J., Gill, B.S. 2009. Evolution of New Disease Specificity at a Single Resistance Locus in a Crop-Weed Complex: Reconstitution of the Lr21 Gene in Wheat. Genetics 182:595-602.

Shivrain, V.K., Burgos, N.R., Gealy, D.R., Baquireza, C.J. 2008. Maximum outcrossing rate and genetic compatibility between red rice (Oryza sativa) biotypes and Clearfield rice. Weed Science. 56(6):807-813.

Liu, G., Jia, Y., Correa-Victoria, F.J., Prado, G.A., Yeater, K.M., McClung, A.M., Correll, J.C. 2009. Mapping quantitative trait loci responsible for resistance to rice sheath blight disease using greenhouse assays. Phytopathology. 99(9):1078-1084.

Wang, X., Jia, Y., Shu, Q., Wu, D. 2008. Haplotype diversity at the Pi-ta locus in cultivated rice and its wild relataives. Phytopathology. 98:1305-1311.

Lee, S., Wasmishe, Y., Jia, Y., Liu, G., Jia, M.H. 2009. Identification of two major resistance genes against race IE-1k of Magnaporthe oryzae in the indica rice cultivar ZHE733. Molecular Breeding. 24:127-134.

Eizenga, G.C., Agrama, H.A., Lee, F.N., Jia, Y. 2009. Exploring genetic diversity and potential novel disease resistance genes in a collection of rice wild relatives. Genetic Resources and Crop Evolution. 56:65-76.

Costanzo, S., Jia, Y. 2009. Alternatively spliced transcripts of Pi-ta blast resistance gene in Oryza sativa. Plant Science. 177:468-478.

Shivrain, V.K., Burgos, N.R., Gealy, D.R., Smith, K.L., Scott, R.C., Mauromoustakos, A., Black, H.L. 2009. Red rice (Oryza sativa L.) emergence characteristics and influence on rice (O. sativa) yield at different planting dates. Weed Science. 57(1):94-102.

Shivrain, V.K., Burgos, N.R., Salesa, M.A., Mauromoustakos, A., Gealy, D.R., Smith, K.L., Black, H.L., Jia, M.H. 2009. Factors affecting the outcrossing rate between Clearfield rice and red rice (Oryza sativa). Weed Science. 57:394-403.

Last Modified: 11/24/2014
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