Location: Peanut and Small Grains Research Unit
2021 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
In order to discover new sources of sorghum germplasm with resistance to sugarcane aphid (SCA), a group of highly diverse germplasm (consisting of 242 accessions) collected by the Indian sorghum community were obtained for this research purpose. Seedlings developed from those germplasm were screened for resistance to sugarcane aphids in a greenhouse, and survived lines have been identified as the new sources of sorghum with resistance to this pest aphid. Thus, these newly identified sugarcane aphid resistant lines are being utilized in breeding resistant sorghum. (Objective 1A)
Sixteen hundred barley accessions from the National Small Grains Collection (NSGC) were screened for bird cherry-oat aphid (BCOA) resistance and 150 rescued resistant plants grown in the greenhouse. (Objective 1A)
Developed six populations for greenbug resistance, three of which can be used to map greenbug resistance genes next year. (Objective 1A)
Three recombinant inbred line (RIL) populations were phenotyped for greenbug resistance and 6 allelism tests were conducted to determine genetic diversity for greenbug resistance in barley. (Objective 2B)
We have screened a large set of wheat accessions available at NSGC for resistance to greenbug and BCOA, leading to the identification of 6 resistance sources each for greenbug and BCOA. Given that resistance genes are urgently needed, we decided to focus on identification of resistance genes in these accessions. Therefore, we develop mapping populations for these resistance sources. Currently, we have one F2, four F3:4, and one F5:6 recombinant inbred line (RIL) populations for BCOA resistance, and the F5:6 RIL population can be used for genetic analysis next year. (Objective 2C)
We have developed a genetic map for the SCA-resistant line with a high-density of DNA markers, particularly with two flanking markers at the resistant locus; thus, it allowed us to identify a major quantitative trait locus (QTL) on chromosome 6. This locus accounted for a large portion of phenotypic variation observed for sugarcane aphid resistance. This result confirms the precise location of a major aphid resistant QTL and will be useful for the experiment of gene cloning. (Objective 2C)
One of the important research objectives is to identify genes and gene products that are related to the host plant defense against aphids. During the past year, the cutting-edge technologies including RNA-sequencing and proteomics were used to develop gene expression profiles of sorghum seedlings in response to infestation by sugarcane aphids. Those aphid-induced data are being analyzed to compare differential expression of defense-related genes between resistant and susceptible lines, and the resulted gene expression profiling will provide us with new insights and tremendous promise for dissecting the host defense and molecular resistance mechanisms. (Objective 2D)
Eighty-six crosses were made to transfer BCOA resistance into adapted malting barley backgrounds. (Objective 3A)
Genotyped a RIL population derived from TA3516 x Bainong 418 using genotyping by sequencing and evaluated responses of 246 RILs to BCOA infestation in 2020 and 2021. To get more accurate results, the plan is to increase sample size in BCOA infestation experiments. Therefore, we increased seeds of the RIL population this year, and will collect additional phenotypic data next year (Objective 2C). In addition, a breeding line carrying greenbug and Russian wheat aphid (RWA) resistance genes was evaluated for resistance to U.S. RWA and greenbug biotypes. This line will be released after further evaluation in field. (Objective 3A)
Newly identified sugarcane aphid resistance sources have been crossed into several elite sorghum lines for development of both mapping populations and breeding populations. Several resulted populations are being tested and the pedigree has been advancing simultaneously, and a couple of such populations are already at F5 or F6 generation (i.e., recombinant inbred lines). The promising SCA inbred lines may be released within 1-2 years. (Objective 3B)
There were 2,957 single rows planted for evaluation in Woodward, Oklahoma and 310 rows were selected and harvested. Advanced yield trials were planted and harvested in Lahoma and Chickasha, Oklahoma. Grain yield, test weight, plant height, lodging, and percent plump and thin kernels were measured. 115 second filial generation (F2) populations were planted in Woodward, Oklahoma and 30 heads were selected from each population. All heads were threshed and seed counted. 3,420 lines were screened to both RWA and greenbug and 7,058 resistant selections from screening were grown and harvested in the greenhouse for field evaluation in Fiscal Year 2022. 30 second filial generation (F2) populations were increased and harvested in the greenhouse for head selection in Fiscal Year 2022. 121 crosses were made towards transferring malting quality into adapted winter barley backgrounds. (Objective 3B)
Accomplishments
1. Identification and classification of the Lipoxygenase gene family and their roles in sorghum-aphid interaction. Sorghum is an important crop grown worldwide and ranks fifth among the major cereal crops in terms of both production and area planted. But it is currently threatened by sugarcane aphid (SCA). SCA has spread to all sorghum growing areas in the U.S. and it has become a major pest of sorghum. Thus, the current research at USDA-ARS laboratory in Stillwater, Oklahoma, has focused on studies of genetic mechanisms underlying host plant resistance in sorghum using a cutting-edge genomic approach. Lipoxygenase (LOX) pathway is crucial for plant defense responses to infection by pathogens and insects. ARS researchers have conducted a genome-wide analysis of the sorghum LOXs genome, which led to the identification of all members of the LOX gene family, showing at least nine individual genes in this gene family. Furthermore, expression of those genes was investigated, which provided the evidence that some of the LOX genes are closely associated with plant responses to aphid attack, implying that those genes and their expression products play important roles in plant resistance to SCA. This new information generated from this study will facilitate future research to characterize the roles of each individual LOXs gene in a stress-related function. This research will be of great interest to plant scientists and crop breeders for genetic manipulation and breeding for better resistant varieties for our sorghum producers. It is believed that host plant resistance is the most economical and effective method to manage this pest.
2. Elevated production of reactive oxygen species is related to host plant resistance to sugarcane aphid in sorghum. Sugarcane aphid is a devastating insect pest that causes severe damage to sorghum plants and huge reduction in crop production in the U.S. and elsewhere. ARS scientists in Stillwater, Oklahoma, conducted research exploring molecular mechanisms of host plant resistance to sugarcane aphids. One of the research efforts focuses on examining production of reactive oxygen species (ROS) in plants that were being attacked by the aphids and analyzing altered expression of related plant enzymes in response to aphid attack. According to their recent results, those aphid-induced reactive chemical species such as hydrogen peroxide (H2O2), were rapidly produced in the aphid-attacking plants. Significant increase in H2O2 accumulation was observed in resistant genotype at all time points surveyed as compared to susceptible plants. In addition, some related ROS enzymes were induced in aphid-challenged plants, leading to much higher expression in resistant plants compared to susceptible plants. On other hand, research data showed that aphid survival and fecundity were significantly inhibited in resistant plants when compared to susceptible plants. Taken together, those findings suggest that the elevated accumulation of H2O2 and the strong up-regulation of the antioxidant genes in sorghum may have contributed to host plant resistance to sugarcane aphid in resistant cultivars, but the susceptible cultivar resulted in the failure of attempting defense against sugarcane aphid. The resulted information will contribute to a better understanding of the molecular mechanisms of host plant defense against sugarcane aphids, contributing a good strategy to manage aphid pest for the U.S. sorghum farmers.
3. Identified and characterized a new leaf rust resistance gene Lr470121 in wheat. Leaf rust is a major disease that causes significant yield losses worldwide and is particularly important in the Great Plains where new P. triticina races frequently occur. Currently, only a few resistance genes originating from wheat relatives remain effective against P. triticina races in this region. Therefore, identification of novel resistance genes is imperative for sustainable wheat production. A USDA-ARS scientist at Stillwater, Oklahoma, identified a leaf rust resistance gene Lr470121 conferring high resistance to P. triticina races collected in southern Great Plains, and allelism test indicated that Lr470121 is a new gene. Lr470121, which is being transferred to Oklahoma wheat breeding lines, is a valuable gene that can be widely used to enhance leaf rust resistance in the U.S., and four breeding lines combining Lr470121, Lr34, Lr37, and Lr39 have been developed.
4. New barley greenbug resistance gene Rsg1.a3 in wild barley. Greenbug not only causes barley yield losses, but also facilitates the spread of several viruses destructive to barley. Planting greenbug-resistant cultivars is the most economical and environment-friendly way to reduce losses. Currently, more than 30 different types of greenbug (known as biotype) have been identified, and two greenbug resistance genes (Rsg1 and Rsg2) are available for barley breeding, with each one conferring resistance to only a few greenbug biotypes. However, greenbug biotype H is virulent to both Rsg1 and Rsg2, making it urgent to identify novel greenbug resistance genes. USDA-ARS scientists at Stillwater, Oklahoma, and Manhattan, Kansas, discovered a new barley greenbug resistance gene, officially designated Rsg1.a3, in WBDC336 and developed Kompetitive allele specific PCR (KASP) markers for its introgression into elite barley cultivars. WBDC336 is a wild barley collected from Turkmenistan, and Rsg1.a3 confers resistance to economically important greenbug biotypes and are valuable for barley breeding. Especially, Rsg1.a3, the only gene conferring resistance to greenbug biotype H, makes it feasible for the first time to develop barley cultivars resistant to all economically important greenbug biotypes.
5. Identified a new greenbug resistance gene Rsg2.a3 in wild barley. Greenbug is a worldwide pest that poses a serious threat to cereal production, and new greenbug resistance genes are urgently needed to meet the challenges of the diverse greenbug biotypes. USDA-ARS scientists at Stillwater, Oklahoma, and Manhattan, Kansas, identified a new greenbug resistance gene Rsg2.a3 in wild barley accession WBDC053, and mapped Rsg2.a3 to the terminal region of chromosome 3HS. Two user-friendly, high throughput Kompetitive allele specific PCR (KASP) markers co-segregating with Rsg2.a3 were developed. Rsg2.a3 is valuable, and the KASP markers developed in this study can expedite its use in barley breeding.
Review Publications
Shrestha, K., Pant, S.R., Huang, Y. 2021. Genome-wide identification and classification of the Lipoxygenase gene family and their roles in sorghum-aphid interaction. Plant Molecular Biology. https://doi.org/10.1007/s11103-020-01107-7.
Pant, S.R., Huang, Y. 2020. Elevated production of reactive oxygen species is related to host plant resistance to sugarcane aphid in sorghum. Plant Signaling and Behavior. http://doi.org/10.1080/15592324.2020.1849523.
Xu, X., Li, G., Bai, G., Bernardo, A.E., Carver, B.F., St Amand, P.C., Bian, R. 2021. Characterization of an incomplete leaf rust resistance gene and development of KASP markers for the leaf rust resistance gene Lr47 in wheat. Phytopathology. https://doi.org/10.1094/PHYTO-07-20-0308-R.
Xu, X., Li, G., Bai, G., Bernardo, A., Carver, B.F., St Amand, P., Armstrong, S. 2020. Development of KASP markers for wheat greenbug resistance gene Gb5. Crop Science. 61(1):490–499. https://doi.org/10.1002/csc2.20339.
Puterka, G.J., Xu, X., Li, G., Carver, B.F., Guo, P. 2020. Mechanisms of resistance of new wheat gene Dn10 in comparison with other Dn genes resistant to Russian wheat aphid. Crop Science. 60:1782-1788. https://doi.org/10.1002/csc2.20051.
Niu, F., Xu, Y., Liu, X., Zhao, L., Bernardo, A.E., Yaoguang, L., Guixia, L., Chen, M., Cao, L., Hu, Z., Xu, X., Bai, G. 2020. The Hessian fly recessive resistance gene H4 mapped to chromosome 1A of the wheat cultivar ‘Java’ using genotyping-by-sequencing. Journal of Theoretical and Applied Genetics. https://doi.org/10.1007/s00122-020-03642-9.
Xu, X., Mornhinweg, D.W., Bai, G., Steffenson, B., Bian, R., Li, G., Bernardo, A. 2021. Rsg1.a3: A new allele conferring unique resistance to greenbug biotype H at the Rsg1 locus in Hordeum vulgar ssp spontaneum. Crop Science. 61:3578-3585. https://doi.org/10.1002/csc2.20581.
