Location: Mycotoxin Prevention and Applied Microbiology Research
2021 Annual Report
Objectives
Objective 1: Identify Fusarium graminearum (Fg) virulence factors and/or fitness traits that can be targeted to reduce grain mycotoxin contamination. [C1, PS2]
Sub-objective 1.A: Identify and characterize core effectors of Fg that can be targeted to reduce initial infection of wheat and barley.
Sub-objective 1.B: Identify and characterize Fg population-specific factors that contribute to differences in virulence and mycotoxin contamination of wheat and barley.
Objective 2: Identify germplasm that can be used by breeders to simultaneously target climate resilient mycotoxin resistance and high grain quality traits. [C1, PS2]
Sub-objective 2.A: Evaluate the resilience of FHB resistance to e[CO2] in MR and S wheat cultivars.
Sub-objective 2.B: Determine the impact of e[CO2] on the production of Fg mycotoxins and other secondary metabolites during growth on wheat and barley grains.
Sub-objective 2.C: Determine the impact of e[CO2] on the nutritional quality of FHB MR parent wheat lines and identify potential breeding strategies to maintain grain quality.
Objective 3: Manipulate microbial populations or metabolites to control trichothecene contamination of grain and malting barley. [C1, PS2, PS5]
Sub-objective 3.A: Evaluate the efficacy of Sarocladium and Paenibacillus as biocontrol agents to control FHB and mycotoxin contamination of grains.
Sub-objective 3.B: Determine how perithecial pigmentation affects production, discharge, and germination of Fusarium ascospores so that these metabolites can be manipulated to reduce Fg inoculum in the field.
Sub-objective 3.C: Develop a biofumigant from plant derived metabolites to inhibit fungal growth and mycotoxin production during barley malting.
Approach
Mycotoxins are poisonous fungal metabolites that contaminate cereals, making them unsafe for human or livestock consumption. Contamination originates in the field during grain development when crops become infected by mycotoxigenic fungal pathogens that can result in diseases with significant economic losses. Fusarium head blight (FHB), is a devastating disease of wheat and barley that is caused primarily by the fungal pathogen Fusarium graminearum (Fg) which produces mycotoxins, including trichothecenes and zearalenone. FHB is a complex ecological problem that has been difficult to eradicate because infection is dependent on multiple interacting factors. The severity of FHB is contingent on the prevalence, virulence, and aggressiveness of the pathogen, the genetic potential of the host plant’s resistance, as well as other abiotic and biotic environmental factors that influence the outcome of the plant-pathogen interactions. Previous efforts by scientists in the FHB community have laid a foundation of information on pathogen virulence and host resistance, but we need to further understand how environmental factors shape the outcome and impact of interactions. Using a holistic approach that tackles the problem from multiple angles, we propose to target factors that can be manipulated to impose mycotoxin control: 1) Fg pathogenic fitness, 2) resilience of crop resistance, and 3) beneficial microorganisms and microbial or plant metabolites. Knowledge obtained from this approach will aid in the development of integrated climate-resilient control strategies for FHB and mycotoxin contamination of grains, thereby reducing the impact of mycotoxins on our food supply. These studies will ultimately benefit growers; small grain breeders; stakeholders in the food and feed industry; other research scientists; regulatory agencies (United States Food and Drug Administration, USDA Federal Grain Inspection Service, and Animal Plant Health Inspection Service); and most importantly the consumer.
Progress Report
Mycotoxins are toxic chemicals produced by fungal pathogens that infect crops and contaminate food and feed making them unsafe to eat. The fungus Fusarium graminearum produces a class of mycotoxins called trichothecenes and is the primary cause of a devastating disease of small cereal crops called Fusarium head blight (FHB). FHB and trichothecene contamination are estimated to cost the U.S. wheat and barley industry approximately $1.5 billion annually. The goal of this project is to develop ways to mitigate losses and food safety concerns caused by FHB and grain contamination.
Plant pathogens secrete small molecules, known as effector proteins or virulence factors, that enable them to overcome plant defenses and cause disease. These small molecules are essential for the pathogen to be successful and therefore are ideal targets to control disease. The aim of Objective 1 is to identify effector proteins and/or virulence factors that make Fusarium graminearum a successful pathogen. While some of these small molecules are produced by many Fusarium species that cause agricultural problems, others are unique to F. graminearum, or specific F. graminearum populations. To accurately predict effectors that have the potential to be important for FHB or other diseases, we tested several effector prediction software tools, and then combined individual prediction tools into a computational pipeline based on sophisticated machine learning approaches. We are now using this pipeline to evaluate genomes of 38 Fusarium species and identify effector genes that are shared by these pathogens and likely contribute to their success.
Three populations of F. graminearum (NA1, NA2, NA3) have been identified in North America. NA1 and NA2 population isolates primarily produce the trichothecene deoxynivalenol (also known as vomitoxin), while NA3 population isolates produce a chemical variant known as NX. Vomitoxin has been shown to function as a virulence factor and is required for F. graminearum to spread throughout the wheat head. However, the role of NX in NA3 isolate success has not been similarly demonstrated. This year, we investigated the role of NX in F. graminearum infection of wheat. We generated mutant strains unable to produce NX. Experiments showed that NX production in NA3 isolates contributes not only to disease spread, but also initial infection.
Complete FHB resistant wheat genotypes have not been identified, but cultivars that are moderately resistant to FHB have been developed through extensive breeding efforts. During our previous project, we discovered that some FHB moderately resistant wheat cultivars were more vulnerable to disease and had less nutritional value when grown at elevated atmospheric carbon dioxide. Research under Objective 2 is designed to identify wheat germplasm that will maintain high grain quality and resistance to FHB at elevated carbon dioxide. This year, we obtained 12 near isogenic lines with or without the primary marker that breeders currently use to develop FHB moderately resistant cultivars and started plant growth experiments to measure physiological responses to elevated carbon dioxide. We also evaluated the mycotoxin accumulation and fungal biomass of F. graminearum isolates selected from the three North American populations grown on grain from 15 wheat cultivars grown at ambient or elevated carbon dioxide.
Objective 3 is designed to develop methods to manipulate microbial populations or metabolites to control trichothecene contamination of grain and malting barley. Use of beneficial microbes to reduce FHB is an effective ecofriendly approach. We previously showed that under controlled growth chamber conditions wheat plants that harbor beneficial bacteria within their tissues were less susceptible to FHB and mycotoxin contamination. This year we evaluated two ways to introduce beneficial bacteria, on wheat heads or on wheat seed prior to planting. Treatment of mature wheat heads with bacteria did not control FHB, and in some cases enhanced disease symptoms. However, bacteria applied to the seeds successfully controlled FHB without reducing seed germination.
Grain infected with F. graminearum can continue to accumulate mycotoxins during storage and processing. Mycotoxin accumulation during malting of barley results in more than $406 million annual losses to the U.S. malting and brewing industry. This year we continued experiments to determine if volatile antifungal chemicals released from wetted mustard seed meal kill F. graminearum in barley being stored prior to malting. We tested volatiles from three mustards and found that Penny Cress and Ethiopian Mustard had the most antifungal activity. Initial experiments found that the volatiles killed Fusarium living on the surface of the barley seed but Fusarium within the seed survived. We also found that mustard seed meal fumigation effectively eliminated three common grain storage insect pests.
Accomplishments
1. Gene from fungus helps protect wheat and barley from fungal toxins. The cereal crop fungal pathogen Fusarium graminearum causes a cereal disease called Fusarium Head Blight (FHB) and produces vomitoxin and related mycotoxins. These toxins contaminate grain and if consumed can cause serious health problems to humans and animals. Control of F. graminearum infection and mycotoxin contamination remains a challenge due to a lack of completely resistant plant varieties and the emergence of fungicide resistant strains. ARS researchers at Peoria, Illinois, tested a strategy to enhance crop resistance to vomitoxin by using a fungal gene (Tri101) that protects the fungus from its own toxin. Transgenic model plants, Arabidopsis, expressing the fungal gene effectively detoxified vomitoxin. Additionally, the transgenic plants were protected by pumping the toxin out of the plant cells. If successfully duplicated in cereal crops, this research could provide an alternative strategy to enhance cereal crop resistance to mycotoxins and reduce FHB and toxin contamination of grain.
Review Publications
Hao, G., McCormick, S., Tiley, H., Usgaard, T. 2021. Detoxification and excretion of trichothecenes in transgenic Arabidopsis thaliana expressing Fusarium graminearum trichothecene 3-O-acetyltransferase. Toxins. 13(5). https://doi.org/10.3390/toxins13050320.
Senatore, M.T., Ward, T.J., Cappelletti, E., Beccari, G., McCormick, S.P., Busman, M., Laraba, I., O'Donnell, K., Prodi, A. 2021. Species diversity and mycotoxin production by members of the Fusarium tricinctum species complex associated with Fusarium head blight of wheat and barley in Italy. International Journal of Food Microbiology. https://doi.org/10.1016/j.ijfoodmicro.2021.109298.
