Location: Wheat, Sorghum and Forage Research
2019 Annual Report
Objectives
Objective 1: Identify and manipulate the genetic and biochemical mechanisms controlling lignin deposition to develop improved sorghum germplasm for bioenergy and forage uses.
Subobjective 1A: Characterize the effects six recently identified brown midrib mutants (bmr) on phenylpropanoid metabolism and lignin deposition.
Subobjective 1B: Evaluate ways to increase lignin deposition and alter phenolic composition of biomass through overexpression of monolignol genes.
Objective 2: Identify and manipulate the genetic and biochemical mechanisms controlling starch and phosphorus composition of grain to develop novel sorghum traits for food, biofuel and livestock production.
Subobjective 2A: Identify and characterize mutants that alter phosphorus composition and reduce phytate in grain.
Subobjective 2B: Develop germplasm with altered starch composition and content in grain.
Objective 3: Identify resistance to fungal pathogens in lignin modified sorghum germplasm for development of stalk rot-resistant grain, bioenergy, and forage sorghums.
Subobjective 3A: Determine the responses of sorghum lines with six recently identified bmr mutants to stalk pathogens.
Subobjective 3B: Assess impact of the stalk pathogen Fusarium thapsinum on sorghum with altered the monolignol synthesis.
Subobjective 3C: Determine the response of sorghum stalk moisture phenotypes on stalk rot pathogens.
Subobjective 3D: Determine whether beneficial microorganisms increase protection of bmr mutants against root and stalk pathogens.
Objective 4: Identify resistance interactions between sorghum grain with novel composition and fungal pathogens for food, fuel, and feed uses.
Subobjective 4A: Determine whether pericarp pigments provide protection against grain pathogens.
Subobjective 4B: Determine whether grain tannins prevent fungal infection.
Approach
Sorghum (Sorghum bicolor) is a climate resilient crop, which is capable of providing grain and forage (biomass) to both the existing agricultural markets and the emerging bioenergy markets in the United States. To compete in these markets, compositional improvements to both sorghum grain and forage are needed and an understanding how these changes affect the fungal pathogens of sorghum. The objectives of this project will focus on the genetic, biochemical, and physiological mechanisms affecting the composition of sorghum biomass and grain. Efforts will result in sorghum with altered lignin content and/or composition of biomass, and increased starch content and reduced phytate content of grain for improved bioenergy conversion, livestock utilization and human nutrition. The impacts of fungal pathogens on sorghum with compositionally modified biomass and grain will be determined. Sorghum germplasm with desirable traits enhancing sorghum biomass and grain utilization will be developed, fully characterized, released and deposited into USDA–ARS National Plant Germplasm System (NPGS) for use by public and private sector plant breeders for developing improved hybrids and cultivars. The project consists of three integrated components: germplasm development, molecular biology, and plant pathology. Molecular and conventional methodologies will be utilized, and the project scale will range from gene-level to field-level. The project also has extensive formal and informal collaborations enhancing our ability to conduct this research. Anticipated products include improved sorghum germplasm for the sorghum seed industry with value-added traits and biotic stress tolerance, and tools to assess these biological pathways and fungal pathogen responses of sorghum.
Progress Report
Lignin is a component of the plant cell wall and its presence affects the use of sorghum as livestock forage or bioenergy feedstock. In Subobjective 1A, next-generation sequencing technology was used to determine a chromosome map position for brown midrib (bmr) loci 29, 31 and 32. Candidate genes at each locus were identified for bmr29 and bmr32. The gene identified for bmr30 in the previous FY was shown to no longer be a valid candidate, so an alternative gene was identified within this chromosomal interval. Experiments are being performed to verify that the mutations identified are responsible for the changes to lignin observed in bmr29, 30 and 31 mutant plants. The identification of these bmr genes with this approach will validate the use of this technology for identifying candidate genes of other mutants in sorghum. The newly developed bmr29 through 32 mutant lines in three sorghum varieties were grown in the field for the first year of a two-year trial at two Nebraska locations. Field traits were measured and biomass from these lines was collected for future analyses. The discovery of the genes underlying these bmr mutants through this approach will pave the way for the development of rational strategies to combine bmr mutants to alter lignin content and composition for bioenergy and forage uses in sorghum and other grasses. Understanding the impact of each brown midrib mutant in different varieties will lay the foundation for the development of the next-generation of brown midrib hybrids that will benefit the livestock and bioenergy industries. In Sub-objective 1B, lines overexpressing different genes in lignin synthesis were combined to further elevate phenolic compounds in sorghum biomass. The analyses of biomass from this strategy are currently underway. This approach may lead to new resources for renewable chemical applications, because the elevated phenolic compounds in the biomass do not affect plant growth.
Phytate is a major phosphorus storage compound in seeds, but it is also an antinutrient for both animals and humans, and it causes phosphorus management problems in their waste. In Subobjective 2A, several sorghum mutants with reduced levels of phytate were identified. Further testing narrowed the focus to two mutants, which are in two separate genes. Reduced phytate levels in grain were not observed in the subsequent generation for these two mutants. Previous generation of seeds were planted to reassess the genetic transmission of this low phytate trait. The ability to develop reduced phytate sorghum would increase the use of sorghum in animal feed.
Stalk rot pathogens are destructive to sorghum, particularly under environmental stresses such as drought. Sorghum with impaired lignin synthesis appears to be a source of stalk rot resistance. In Subobjective 3A, the lignin synthesis-impaired lines were shown to have increased resistance under reduced water conditions. Another set of sorghum lines, which had changes to a biosynthesis pathway to increase lignin, either had no increased susceptibility or some lines even had increased stalk rot in Subobjective 3B. There are also beneficial microorganisms in the soil that may prevent stalk rot pathogens from infecting sorghum. In Subobjective 3D, a set of microorganisms were identified that were antagonistic to stalk rot pathogens. Preliminary studies using two strains of these microorganisms increased the growth of sorghum seedlings under adequate water compared to a well-known biocontrol agent or no microorganism treatment; one strain also increased seedling growth under drought conditions. The genomes of these strains will be evaluated through DNA sequencing funded through the USDA Alternatives to Antibiotics Program with the goal of identifying the genes responsible for promoting plant growth and inhibiting fungal diseases. Biological controls are emerging tools to control fungal diseases in sorghum grain, forage and bioenergy production, which reduce use of commercial fungicides and the development of resistance to these compounds.
The phenolic pigments of sorghum grain affect its end-uses, but they are also sources of antioxidants and may help defend against grain molds. In Sub-objective 4A, sorghum with red, yellow or unpigmented grain has been planted in both Texas (high disease pressure) and Nebraska (moderate disease pressure) to assess the role of grain pigments in grain mold resistance. Understanding the role of grain pigments is critical for controlling grain mold, because people consume both unpigmented and pigmented sorghum grain.
Accomplishments
1. Demonstrated a lignin-related enzyme increases energy content of sorghum biomass. Sorghum biomass serves as an important forage crop for livestock, and it is being developed as a bioenergy crop. The caffeoyl-CoA 3-O-methyltransferase (CCoAOMT) gene encodes an enzyme involved in the synthesis of the biomass component lignin. To understand the role of this enzyme in lignin synthesis and its effect on cell wall composition, ARS scientists in Lincoln, Nebraska, and their collaborators used biotechnology to greatly elevate levels of the CCoAOMT enzyme in sorghum plants. The increased levels of this enzyme led to increased levels of phenolic compounds in the biomass, which increased the energy content without affecting lignin levels or plant growth. Engineering of CCoAOMT in plants provides a new way to change biomass composition and increase the energy content of biomass, which may allow the plant to store more energy for renewable chemical applications.
2. Identification of sorghum resistance factors to sugarcane aphid. The sugarcane aphid has emerged as a significant pest for sorghum in the U.S. and around the world. Sugarcane aphid resistant sorghum lines are one way to combat this pest. Through a series of experiments, ARS scientists in Lincoln, Nebraska, and their collaborators characterized the differences between a sugarcane aphid resistant sorghum line and a susceptible one to determine the factors that make sorghum susceptible or resistant to this highly destructive pest. Gene expression analysis of the entire sorghum genome identified several disease resistance/defense genes whose expression supports a role in aphid resistance. Analyses of aphid feeding showed that the aphids spent about a quarter of the time actually feeding on resistant plants compared to the susceptible ones, which shows the resistant plants inhibit aphid feeding. A single dominant gene was responsible for aphid resistance in sorghum, which allows resistant plants to grow normally and impede aphid feeding. Together these discoveries are enabling ARS scientists to develop DNA sequence-based methods to identify the sugarcane aphid resistance gene in sorghum, which will enable sorghum breeders to rapidly create sugarcane aphid-resistant hybrids to combat this devastating pest.
3. Identification of stalk rot resistance in sorghum lines overexpressing lignin biosynthesis genes. Sorghum lines were developed with increased levels of enzymes required for lignin synthesis through biotechnology to provide chemical subunits for renewable fuel and chemical uses from biomass. ARS researchers in Lincoln, Nebraska, tested whether increased enzyme levels affected plant resistance to stalk rots, which can impair biomass production. Three lines had increased resistance to Fusarium stalk rot compared to normal plants, and only one line showed increased susceptibility to charcoal rot. The research demonstrates that changes to lignin synthesis may also increase stalk rot resistance in sorghum. Sources of stalk rot resistant sorghums are critical for the successful development of sorghum as a bioenergy crop.
4. Identification of Fusarium head blight (FHB) resistance in “waxy” wheat. The grain of waxy wheat lacks amylose starch, which increases shelf-life of baked goods containing waxy flour and makes starch more digestible. Mattern, the first Great Plains waxy variety is vulnerable to FHB, which produces a dangerous toxin that can make the grain unconsumable for both humans and animals. ARS scientists in Lincoln, Nebraska, developed several new waxy wheats, and screened their grain for disease symptoms and the presence of this toxin. Two new lines had noticeably less disease symptoms than Mattern. These newly-developed waxy lines are valuable materials for breeding the next-generation of Northern Great Plains waxy wheats with modified starch for specialty products. FHB resistant wheat varieties are vitally important way to tackle this destructive disease, because severe FHB outbreaks are intermittent.
Review Publications
Jun, S., Vermerris, W., Sattler, S.E., Kang, C. 2018. Biochemical and structural analysis of substrate specificity of a phenylalanine ammonia-lyase. Plant Physiology. 174:1452-1468. https://dol.org/10.1104/pp.17.01608.
Grover, S., Wojahn, B., Varsani, S., Sattler, S.E., Louis, J. 2019. Resistance to greenbugs in the sorghum nested association mapping population. Arthropod-Plant Interactions. 13(2):261-269. https://doi.org/10.1007/s11829-019-09679-y.
Tetreault, H.M., Sajjan, G., Scully, E.D., Gries, T.L., Sattler, S.E., Palmer, N.A., Sarath, G., Louis, J. 2019. Global responses of resistant and susceptible sorghum (sorghum bicolor) to sugarcane aphid (melanaphis sacchari). Frontiers in Plant Science. 10:145.Available: https://doi.org/10.3389/fpls.2019.00145.
Tetreault, H.M., Scully, E.D., Gries, T.L., Palmer, N.A., Sattler, S.E., Funnell-Harris, D.L., Baird, L., Seravalli, J., Sarath, G., Clemente, T.E. 2018. Over-expression of the sorghum bicolor SbCCoAOMT alters cell wall associated hydroxycinnamoyl moieties. Plant Physiology. 13(10): e0204153. Available: https://doi.org/10.1371/journal.pone.0204153.
