Location: Peanut and Small Grains Research Unit
2023 Annual Report
Objectives
Objective 1: Identify new germplasm sources of resistance to cereal aphids, including greenbugs, Russian wheat aphid, bird cherry-oat aphid, and other important insect pests, in wheat, barley, and sorghum.
Subobjective 1A. Evaluate available germplasm resources (U.S. germplasm collections and accessible exotic resources) to identify new sources resistant to insect pests [Russian wheat aphid (RWA), greenbug (GB), bird cherry-oat aphid (BCOA)], and other important insect pests in wheat, barley, sorghum, and related species.
Objective 2: Characterize the genes controlling pest resistance and other related traits that are important for sustainable cereal crop production.
Subobjective 2.A. Develop and evaluate genetic populations to determine the genetic control of host resistance to GB, RWA, and BCOA in barley.
Subobjective 2.B. Develop and evaluate genetic populations to determine levels of genetic diversity of host resistance and genes controlling the resistance to GB, RWA, SCA, and BCOA in wheat, barley, and sorghum.
Subobjective 2.C. Map genes conferring resistance to cereal aphids, and develop genomic tools for cloning and marker-assisted selection of aphid resistance genes.
Subobjective 2.D. Conduct functional genomics studies on host response to attack by GB and sugarcane aphids (SCA), leading to advanced understanding of the defense mechanisms in the hosts and discovery of genes and factors that affect host defense against insect pests (i.e. GB and SCA) in grain sorghum, forage sorghum, and related species.
Subobjective 2.E. Reveal the genetic architecture of BCOA resistance using genomewide association study (GWAS), and develop genomic tools to facilitate rapid introgression of aphid resistance genes into adapted germplasm.
Objective 3: Develop enhanced germplasm and new varieties of sorghum, wheat, and barley with resistance to insect pests.
Subobjective 3.A. Develop high performance wheat, barley, and sorghum germplasm with enhanced resistance to GB, RWA, SCA, or BCOA, and release to the public.
Subobjective 3.B. Develop winter malting barley germplasm and cultivars for the Great Plains with enhanced insect resistance.
Approach
Wheat, barley and sorghum are the major cereal grains in the United States but their production is often threatened by destructive aphid pests, specifically the Russian wheat aphid, greenbug, bird cherry-oat aphid, and sugarcane aphid. Identification of natural resistance and use of genetically pest-resistant cultivars and hybrids in an integrated pest management program are the most economical and environmentally sound methods to reduce the negative economic impact of these damaging aphids. The overall goal of this project is to develop high performance wheat, barley, and sorghum with resistance to aphid pests. To accomplish this goal, the project will search available germplasm collections to find new, effective sources of resistance to aphid pests that are notorious for overcoming resistance through biotype evolution. The genetic diversity and resistance mechanisms will be analyzed, and resistance genes will be characterized and transferred into adapted genetic backgrounds. Plant genotyping will be conducted to map aphid resistance genes to the crop chromosomes and to develop molecular markers to facilitate marker-assisted selection and map-based gene cloning. The research team of the project will work closely with collaborating plant breeding programs to obtain elite breeding lines to use as parents in backcrossing procedures to transfer aphid resistance and other value-added traits. The genetically improved germplasm and varieties will be field-tested for agronomic and quality performance prior to release. The project will provide testing and selecting support to assure that these desirable genes move through the various breeding programs on their way to producers via improved cultivar and hybrid releases.
Progress Report
This is the final report for Project Number 3070-21000-008-000D, entitled “Genetic mechanisms and Improvement of Insect Resistance in Wheat, Barley, and Sorghum”, which terminated in 02/28/2023 and continues as Project Number 3070-21000-010-000D, entitled “Genetic Improvement for Resistance to Aphids and Major Diseases in Wheat, Barley, and Sorghum”. Progress was made toward the completion of all three objectives.
For Sub-objective 1A, we have evaluated over 6000 wheat germplasm for resistance to the Russian wheat aphid (RWA), greenbug (GB), and bird cherry oat aphid (BCOA). A set of 14 wheat landraces that are resistant or highly resistant to all five predominant U.S. biotypes—RWA1, RWA2, RWA3, RWA6, and RWA8—were identified. Five new greenbug resistance sources that may carry novel resistance gene(s) were discovered, and mapping and breeding populations have been developed to identify and transfer the new gene(s) into locally adapted breeding lines. In addition, a novel BCOA assay protocol has been developed, leading to reproducible BCOA assay results.
In response to the invasive sorghum aphid, Melanaphis sacchari (Zehntner), a new devastating pest moving onto sorghum crops recently, a sorghum geneticist at Stillwater, Oklahoma, collected and evaluated more than 10,000 sorghum germplasm lines, including elite lines, landraces and wild relatives harboring useful genetic variation for the desirable traits such as aphid resistance, wide adaptation, and high yield potential. All germplasm materials were screened for their reaction to artificial infestation in greenhouse, which led to the identification of 45 germplasm lines with varied levels of resistance to the aphid. These survived resistant lines were rescreened to confirm the stable aphid resistance. Then, the confirmed resistant germplasm have been purified and selected through pre-breeding approach, which can be more easily utilized in sorghum breeding. Now some of the aphid resistant trait/genes have been already transferred into the elite breeding lines and are ready for releasing, and others will be ready for releasing soon.
For Sub-objective 1A, 8,000 accessions of barley from the National Small Grains Collection (NSGC) were screened with BCOA. Two accessions were identified with some level of resistance.
For Sub-objective 2A and 2B, genetic populations were developed, phenotyped, and genotyped to determine the inheritance of resistance, genetic diversity for resistance, and to map resistant genes from 6 GB resistant lines. Two new alleles for resistance and 2 new genes for resistance were identified. For Sub-objective 2C, these new alleles and gene were mapped, and markers developed for marker assisted selection by interested breeding programs.
In support of Objective 2B, the sorghum geneticists have developed at least three recombinant inbred line (RIL) populations for studies of aphid resistance in sorghum. At the same time, several breeding populations have been also developed, which are at various generations, from F2 to F6 generations. In addition, they have identified about 60 pairs of new primers based on the available genome sequences of sorghum in the DNA databases and developed more simple sequence repeats (SSR) markers, which are useful for fine-mapping quantitative trait loci (QTLs) in sorghum.
For Sub-objective 2C, genotyping these mapping populations using several hundred of DNA markers, they have constructed genetic maps from the above-mentioned sorghum mapping populations that were developed in the lab recently. In continuous efforts, they have been successful in identifying the QTLs linked to aphid resistance in sorghum, which represents as a major QTL located on chromosome 6 of sorghum. This locus (gene) accounted for a large portion of phenotypic variation observed for aphid resistance. This result confirms the precise location of a major aphid resistant QTL and will be useful for both map-based gene cloning and marker-assisted breeding.
A new greenbug resistance gene, officially designated as Gb8, was identified in the wheat reselection line PI 697274, and this gene is currently used in wheat breeding programs. Wheat greenbug resistance genes Gb1, Gb5 and Gb7 were characterized, and user-friendly, high throughput markers were developed for their use in wheat molecular breeding. A novel gene conferring resistance to all U.S. predominant RWA biotypes was revealed, and this gene was officially named as Dn10. Dn10 has the potential to play an important role in reducing economic losses caused by RWA. In addition, a set of six wheat powdery mildew resistance genes have been discovered through genetic analysis, and three of them, Pm59, Pm63, and Pm65, won international approval of permanent designations. A novel leaf rust resistance gene, Lr81, and a QTL for leaf rust resistance, QLr.stars-1RS, were also identified. Both Lr81 and QLr.stars-1RS are valuable for sustainable wheat production.
For Sub-objective 2D, recent studies by ARS scientists at Stillwater, Oklahoma, focused on the identification of plant resistance genes in sorghum plants using cutting-edge genomic tools. In the genome-wide analysis of plant-aphid interactions, 308 resistance (R) genes were discovered in the sorghum genome; subsequently, the gene structures and their phylogenetic relationship among the members of the R gene family were characterized through bioinformatic analysis. Furthermore, differential expression of those R genes responding to aphids were analyzed using both RNA Sequencing (RNA-Seq) and quantitative real-time polymerase chain reaction (PCR) (qRT-PCR) technologies, leading to the identification of several important R genes that confer aphid resistance in sorghum. The information generated in this research advanced the understanding of the defense mechanisms in the hosts and the genes and regulatory factors that affect host defense against aphids in sorghum and possibly other related species.
For Sub-objective 2E, a set of 260 lines were genotyped and evaluated for responses to BCOA infestation. Genome-wide association analysis was performed to identify genetic variance associated with BCOA resistance or tolerance. A set of over 400 synthetic hexaploid wheat (SHW) lines were also evaluated for BCOA resistance, leading to the identification of a couple of SHW lines that provide a high level of BCOA resistance. These SHW lines can be directly used in wheat breeding and are precious for the wheat industry. Development of mapping and breeding population is underway to pinpoint the BCOA resistance genes in the SHW lines and transfer those genes into locally adapted cultivars.
For Subobjective 3A, three wheat germplasm, including PI 697274, PI 682675, and PI 700925, have been developed and released to the wheat community. PI 697274, PI 682675, and PI 700925 carry greenbug resistance gene Gb8, RWA resistance gene Dn10, and leaf rust resistance gene Lr81, respectively.
The sorghum geneticist at Stillwater, Oklahoma, recently focused searching for new sources of aphid resistance and subsequently incorporating the genetic resistance to elite sorghum breeding lines through traditional crossing breeding approach. This work has led to development and releasing of two improved sorghum lines, STARS 1801S with genetic resistance to both sugarcane aphid and greenbug and STARS 1802S with resistance to sugarcane aphid and head smut disease.
For Sub-objective 3A and 3B, over the 5-year project, more than 750 crosses were made to transfer aphid resistance (RWA, GB, and BCOA) into elite feed and malting barleys.
For Sub-objective 3B, crosses were also made to transfer malting quality from elite winter malting barleys into winter feed barleys adapted to the southern plains. Fifty to 100 heads were selected from a total of 547 resulting F2 populations and seeds from over 5,000 heads screened to both RWA and GB as seedlings in the greenhouse. Each year approximately 4,000 F3 resistant seedlings were selected and increased in the greenhouse to the F4. These F4 were evaluated in the field the following season as F4 plant rows and 10% selected for post-harvest evaluation (grain yield, test weight, potential malting quality, row type). Superior lines were entered into yield trials at 2 locations in Oklahoma. One winter feed barley, USDA Fortress, with resistance to RWA, GB, and a new invasive aphid Hedgehog grain aphid, (HGA), was released in 2020. USDA Fortress is a plant variety protected and licensed to Oklahoma Genetics, Inc.
Accomplishments
1. Identification of a novel Pm65 allele conferring a wide spectrum of resistance to wheat powdery mildew. Powdery mildew is caused by the most adaptive biotrophic fungus Blumeria graminis f. sp. tritici (Bgt) inflicting wheat worldwide. Novel powdery mildew resistance genes are urgently needed that can be rapidly used in wheat cultivar development with minimal disruption of trait advances elsewhere. ARS scientists in Stillwater, Oklahoma, and Manhattan, Kansas, identified a new powdery mildew resistance gene conferring resistance to all U.S. representative Bgt isolates in German cultivar PI 351817. This new gene, designated Pm351817, was delimited to a 634-kb interval in the terminal region of the long arm of chromosome 2A, and tests of allelism indicated Pm351817 is a new allele at the Pm65 locus. User-friendly, high throughput markers were developed for the introgression of Pm351817 into locally adapted lines. Pm351817 can be widely used to enhance wheat powdery mildew resistance in the U.S.
2. Evaluation and improvement of sorghum germplasm for resistance to sugarcane aphids and high yield potential. The invasive sugarcane aphid (SCA), Melanaphis sacchari (Zehntner), is a devastating new pest of sorghum crops. Currently, this pest affects all sorghum production areas of North America and causes severe plant damage and significant yield loss in sorghum. Thus, sorghum producers and farmers face the great challenge and urgently need an effective tactic to protect their crops. In response to this important issue, ARS researchers in Stillwater, Oklahoma, efforts have focused on (1) collecting a wide range of sorghum germplasm with diverse genetic background, (2) searching for new resistant sources through experimental screening, and (3) transferring the newly identified SCA resistance into the well-adapted breeding materials. Initially, more than 10,000 sorghum germplasm lines were collected from the important sorghum production areas, which include elite lines, landraces and wild relatives harboring useful genetic variation for traits such as aphid resistance, wide adaptation, and high yield potential. All germplasm materials were screened for their reaction to artificial infestation in greenhouse, which led to the identification of 45 germplasm lines with varied levels of resistance to SCA. The putative resistant lines were rescreened to confirm the SCA resistance. Most of the confirmed lines have been purified and selected through pre-breeding approach, which can be more easily utilized in sorghum breeding. In fact, several SCA resistant sources/alleles were already transferred into the elite breeding lines and are ready for releasing, and others will be ready for releasing soon. In summary, the new sources of SCA resistance will be useful for sorghum breeding and more importantly, use of the new resistant varieties offers the most economical and environmentally sound method to avoid or reduce the negative economic impact of the damaging aphids on sorghum production.
3. Release of USDA Fortress winter feed barley. Feed barley was produced in acreage equal to wheat in Oklahoma until the 1970s. Interest in barley as an alternative to wheat has increased in the past 5 years. The last barley released in Oklahoma in 1990 was resistant to greenbug, GB, but not the then new invasive pest Russian wheat aphid, RWA. USDA Fortress, released in 2020, the first multiple aphid resistant barley in the U.S. is resistant to GB, RWA, and new invasive aphid, Hedgehog grain aphid, HGA. It is plant variety protected and licensed to Oklahoma Genetics, Inc.
4. Discovery of a novel barley greenbug resistance gene Rsg3. Greenbug is a pest that poses a serious threat to cereal production worldwide. Yield losses caused by greenbug are predicted to increase because of global warming, and new greenbug resistance genes are urgently needed to meet the challenge caused by the continuous occurrence of novel greenbug biotypes. ARS scientists in Stillwater, Oklahoma, and Manhattan, Kansas, identified a novel greenbug resistance gene, designated Rsg3, in the Chinese landrace PI 565676, and delimited Rsg3 to an interval of 93,140 bp between 667,558,306 and 667,651,446 bp on the long arm of chromosome 3H. Two Kompetitive Allele Specific PCR (KASP) markers flanking Rsg3 were developed to facilitate rapid introgression of Rsg3 into locally adapted barley cultivars. Rsg3 and the two KASP markers developed in this study are valuable for sustainable barley production.
Review Publications
Huang, J., Shrestha, K., Huang, Y. 2022. Revealing differential expression of phytohormones in sorghum in response to aphid attack using the metabolomics approach. International Journal of Molecular Sciences. 23. Article 13782. https://doi.org/10.3390/ijms232213782.
Huang, Y., Huang, J. 2023. Analysis of transcriptome profiles revealed that aphid attack triggered dynamic defense responses in sorghum plant. Frontiers in Genetics. https://doi.org/10.3389/fgene.2023.1194273.
Mornhinweg, D.W., Armstrong, J.S. 2023. Resistance to bird cherry-oat aphid, Rhopalosiphum padi (L.), in barley, Hordeum vulgare (L.). Southwestern Entomologist. 48(1):75-82. https://doi.org/10.3958/059.048.0107.
Xu, X., Mornhinweg, D.W., Bai, G., Li, G., Bian, R., Bernardo, A.E., Armstrong, J.S. 2022. Characterization of Rsg3, a novel greenbug resistance gene from the Chinese barley landrace PI 565657. The Plant Genome. 16(1). Article e20287. https://doi.org/10.1002/tpg2.20287.
Xu, X., Li, G., Bai, G., Carver, B.F., Bian, R., Bernardo, A.E., Armstrong, J.S. 2023. Genomic location of Gb1, a unique gene conferring wheat resistance to greenbug biotype F. The Crop Journal. https://doi.org/10.1016/j.cj.2023.02.002.
Xu, X., Li, G., Cower, C., Bai, G., Carver, B.F., Bian, R., Bernardo, A. 2023. Identification of a novel Pm65 allele conferring a wide spectrum of resistance to powdery mildew in wheat accession PI 351817. Phytopathology. https://doi.org/10.1094/PHYTO-01-23-0032-R.
Xu, X., Li, G., Lhamo, D., Carver, B.F., Wulff, B., Gu, Y.Q., Xu, S.S., Armstrong, J.S. 2023. Identification of bird-cherry oat aphid and greenbug resistance sources from Ae. tauschii for wheat improvement. Crop Science. https://doi.org/10.1002/csc2.21042.
