Location: Mycotoxin Prevention and Applied Microbiology Research
2021 Annual Report
Objectives
Objective 1: Define diversity of mycotoxin-producing Fusarium species. [C1, PS1, PS2]
Sub-objective 1.A: Elucidate phylogenetic diversity, mycotoxin potential, and pathogenicity to cereals of fungi in the F. tricinctum species complex.
Sub-objective 1.B: Identify genomic and phenotypic differences in collections of F. proliferatum and F. verticillioides isolates to aid discovery of targets for control of fumonisins in corn.
Objective 2: Identify targets to reduce fumonisin contamination in corn. [C1, PS1, PS2, PS5]
Sub-objective 2.A: Determine whether the corn zmCRR1 protein contributes to resistance to fumonisin contamination.
Sub-objective 2.B: Identify corn genes encoding papain-like cysteine proteases involved in fumonisin contamination to aid genomics-assisted breeding.
Sub-objective 2.C: Reduce fumonisin contamination in corn by engineering kernel-specific expression of RNAi targeting the fumonisin biosynthetic gene FUM1.
Sub-objective 2.D: Determine how corn oxylipins control fumonisin production in F. verticillioides.
Sub-objective 2.E: Determine whether the killer meiotic drive element SkK can be used to drive biased transmission of a gene that blocks fumonisin production in F. verticillioides.
Approach
Fusarium species are fungi with potentially the greatest negative impact on agriculture. This is because of their collective abilities to produce mycotoxins and cause destructive diseases in crops, including the important cereals: corn, wheat, and rice. The Fusarium mycotoxins fumonisins and trichothecenes are among the mycotoxins of most concern to food and feed safety due to their toxicity and frequent occurrence in crops. However, Fusarium species produce other mycotoxins whose effects on food and feed safety are poorly understood. In the U.S., harmful impacts of mycotoxins on health are mitigated by removing contaminated grain from food/feed supply chains. Despite these efforts, however, the toxins continue to cause billions of dollars in agricultural losses. This project plan addresses knowledge gaps that hinder control of mycotoxins caused by two groups of Fusarium: the Fusarium tricinctum species complex (FTSC), which includes multiple species that cause head blight of small-grain cereals and produce multiple mycotoxins; and the F. fujikuroi species complex, specifically Fusarium proliferatum and Fusarium verticillioides, which are the primary causes of fumonisin contamination in corn. The proposed research has two objectives: i) define diversity of mycotoxin-producing Fusarium species, specifically members of the FTSC, F. proliferatum, and F. verticillioides; and ii) identify targets to reduce fumonisins in corn. To address the first objective, we propose to elucidate variation in genome sequences, mycotoxin production, and pathogenicity within and among Fusarium species. This will aid development of broadly effective control practices for Fusarium mycotoxins. To address the second objective, we propose to identify corn and Fusarium proteins/genes that can be used to enhance breeding or engineering strategies aimed at reducing fumonisin contamination. To address the second objective, we also propose to develop fumonisin reduction methods based on two biological phenomena: RNA interference and meiotic drive elements. The research accomplishments will aid efforts to reduce mycotoxin contamination in corn and other cereal crops, and will benefit growers, processors, regulatory agencies, and ultimately American consumers.
Progress Report
The fungus Fusarium poses a dual threat to agriculture because it can cause destructive crop diseases and contaminates crops with toxins (mycotoxins) that pose health risks to humans, pets, and livestock. For example, fumonisins are a group of mycotoxins produced by multiple Fusarium species and are of concern because they are frequent contaminants in corn kernels and they can induce multiple animal diseases, including cancer. There are also studies that link fumonisins to several human diseases, including esophageal cancer and neural tube defects. Therefore, the two objectives of this project focus on developing information that contributes to agricultural practices that reduce crop diseases and mycotoxin contamination caused by Fusarium. Objective 1 of the project is focused on understanding the diversity of two groups of Fusarium. The first group is the Fusarium tricinctum species complex (FTSC), which occurs on diverse crop species but is of particular concern as a cause of head blight, a disease of wheat and barley that occurs in many regions of the world where these crops are grown. We examined a global collection of isolates of the FTSC recovered from cereals and determined 1) they are comprised of multiple species; 2) produce diverse mycotoxins; and 3) the type of mycotoxins they produce play a role in their ability to cause head blight of wheat. This research provides plant pathologists, plant breeders, and regulatory organizations with information on which species within the FTSC cause head blight and mycotoxin contamination of wheat. The information will enhance efforts to develop methods to reduce both the disease and contamination problems.
The second group of Fusarium consists of two closely related species, Fusarium proliferatum and F. verticillioides, that are the primary cause of contamination of crops with fumonisin mycotoxins. F. verticillioides is of particular concern for fumonisin contamination in corn, whereas F. proliferatum is of concern for contamination in corn and other crops. Despite the importance of these fungi to agriculture, there is only limited knowledge of differences among strains of each species recovered from different geographic regions and/or hosts. Therefore, we have initiated an investigation into a worldwide collection of isolates of F. proliferatum and F. verticillioides to determine if and how strains recovered from different geographic regions and crops vary in genetic make-up, ability to produce fumonisins, and ability to cause disease. Initial assessments of ~20 isolates of each species indicate that F. proliferatum exhibits greater genetic diversity than F. verticillioides, a finding that is consistent with the former fungus causing disease and mycotoxin contamination on a greater diversity of crop species than the latter. Knowledge of diversity that exists within the F. proliferatum and F. verticillioides will aid in developing robust agricultural practices aimed at reducing fumonisin contamination in corn.
Existing agricultural practices to control contamination of corn and other cereal crops with Fusarium mycotoxins are not effective under all circumstances. Therefore, additional novel practices are needed. Because there is generally a positive correlation between severity of Fusarium-incited crop diseases and mycotoxin contamination, one approach to reduce contamination is to reduce severity of the crop diseases. To this end, we are investigating how plant disease defense proteins interact with proteins produced by fungal pathogens. The scientists found that the defense protein CRR1, which occurs in corn and cotton, can be cleaved into two pieces by proteins produced by the Fusarium species F. verticillioides and F. graminearum, which are pathogens of corn, and the related fungus Verticillium dahliae, which is a pathogen of cotton. While the role of CRR1-like proteins in plant defense is well established, identification of fungal proteins that cleave CRR1 is completely novel. The scientists are currently determining whether the activity of the cleaved CRR1 protein differs from the intact protein. Understanding how CRR1 and fungal proteins interact will provide insight into how plants defend themselves against plant diseases. Such insight should contribute to development of crops with increased resistance to fungal diseases, which should reduce mycotoxin contamination.
Meiotic drive elements (MDEs) are a class of genes that exhibit unusually high rates of transmission from parents to offspring. This phenomenon can result in rapid spread of MDEs in populations of organisms that undergo sexual reproduction. The fumonisin-producing fungus F. verticillioides has an MDE called Spore Killer. When a strain of the fungus with Spore Killer mates with a strain that lacks the gene, all the resulting offspring have the gene. We are investigating whether Spore Killer can be manipulated to spread a gene that blocks fumonisin production in populations of F. verticillioides as a novel strategy to reduce fumonisin contamination in corn. This approach requires that the Spore Killer gene be fused to the gene that blocks fumonisin production. Therefore, we used a fine-tuned mutation approach to determine that Spore Killer corresponds to 213-nucleotide region on chromosome 5 of F. verticillioides. This finding will allow the researchers to conduct experiments to test how effective Spore Killer is at spreading the gene that blocks fumonisin production in a laboratory population of F. verticillioides.
For additional progress during FY21 see the Annual Report for the recently expired project 5010-42000-050-00D.
Accomplishments
1. Determining the causes of fungal disease and toxin contamination in barley and wheat. Head blight is an economically important disease of cereal crops worldwide and is caused by multiple species of the fungus Fusarium, particularly a subset of species known as the Fusarium sambucinum species complex (FSAMSC). These fungi are also food and feed safety concerns because they can contaminate grain with toxins that are health hazards to humans, pets, and livestock. ARS researchers at Peoria, Illinois, in collaboration with Italian universities in Bologna and Perugia determined the species composition in a collection of 117 Fusarium isolates recovered from diseased barley and wheat grown in diverse regions of Italy. DNA sequence analysis indicated that the isolates consisted of nine genetically distinct species that belonged to the Fusarium tricinctum species complex (FTSC), which is distinct from the FSAMSC. Although most of the isolates could produce two toxins of moderate concern to food safety, none produced trichothecenes, a group of toxins that are of major concern and produced by most members of FSAMSC. These findings indicate that FTSC is a significant cause of head blight of barley and wheat, and control strategies targeted at reducing trichothecenes will not be effective against all fungi that cause the disease.
Review Publications
Susca, A., Anelli, P., Haidukowski, M., Probyn, C.E., Epifani, F., Logrieco, A.F., Moretti, A., Proctor, R.H. 2021. A PCR method to identify ochratoxin A-producing Aspergillus westerdijkiae strains on dried and aged food. International Journal of Food Microbiology. 344. Article 109113. https://doi.org/10.1016/j.ijfoodmicro.2021.109113.
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.
