Location: Food Processing and Sensory Quality Research
2025 Annual Report
Objectives
Objective 1: Decipher the molecular, structural and immunological properties of purified native and recombinant allergens that contribute to allergenic potency towards development of therapeutic products. [NP306, C1, PS1C]
Objective 2: Use serum from verified nut allergic and non-allergic individuals to identify and compare IgE and IgG binding sites (or epitopes) of known nut allergens with peptide microarrays to understand cross-reactivity between multiple allergens and improve diagnosis of nut allergy. [NP306, C1, PS1A]
Objective 3: Characterize, quantify and monitor allergen characteristics and levels pre- and post-harvest, and pre- and post-processing of commercial nuts and nut-containing foods, and during nut seed development to produce hypoallergenic, prophylactic or therapeutic food products. [NP306, C1, PS1B]
The immunoglobulin E (IgE) binding sites (epitopes) that are responsible for the symptoms of allergic disease and cross-reactivity among peanut, tree nut and pollen allergens will be identified with peptide microarray technology. The IgE and immunoglobulin G4 (IgG4, thought to act as an IgE-blocking antibody) epitopes will be identified for the most potent nut allergens. These will be modeled on the surface of allergen structures to identify location and common or cross-reactive (or potentially blocked) sequences and structures of allergens among nuts and pollens. Simultaneously, the changes in peanut and tree nut extracts or purified allergens thereof (recombinant or native) will be assessed before and after processing treatments for changes in allergenic properties. Proteins found to be immunologically altered by processing will be assessed within a total nut extract or they will be purified and analyzed for alterations in size, structure, digestibility, binding to various serum IgE and allergen-specific antibodies. The specific amino acid residues, or peptides thought to be modified during different processing events, and thought to contribute to altered allergenic properties, will be identified by mass spectrometry. The studies above will be combined to identify the influence of the processing-induced alteration in relationship to the immunoglobulin binding sites of nut allergens. This will guide the development of better diagnostics and therapeutics as well as processing technologies to reduce allergenicity of nuts and products thereof. Early intervention methods to reduce the allergenic potential of nuts, the natural variation in allergen gene sequence, expression, post-translational modification, stability and accumulation patterns in a model tree nut (pecan) will be studied. Less allergenic variants and factors that can be used to interfere with allergen accumulation in plants will also be characterized in detail. Collectively our studies will also contribute to the development of better detection tools and labeling practices for industry and regulatory agencies resulting in better protection of consumers.
Approach
The immunoglobulin E (IgE) binding sites (epitopes) that are responsible for the symptoms of allergic disease and cross-reactivity among peanut, tree nut and pollen allergens will be identified with peptide microarray technology. The IgE and immunoglobulin G4 (IgG4, thought to act as an IgE-blocking antibody) epitopes will be identified for the most potent nut allergens. These will be modeled on the surface of allergen structures to identify location and common or cross-reactive (or potentially blocked) sequences and structures of allergens among nuts and pollens. Simultaneously, the changes in peanut and tree nut extracts or purified allergens thereof (recombinant or native) will be assessed before and after processing treatments for changes in allergenic properties. Proteins found to be immunologically altered by processing will be assessed within a total nut extract or they will be purified and analyzed for alterations in size, structure, digestibility, binding to various serum IgE and allergen-specific antibodies. The specific amino acid residues, or peptides thought to be modified during different processing events, and thought to contribute to altered allergenic properties, will be identified by mass spectrometry. The studies above will be combined to identify the influence of the processing-induced alteration in relationship to the immunoglobulin binding sites of nut allergens. This will guide the development of better diagnostics and therapeutics as well as processing technologies to reduce allergenicity of nuts and products thereof. Early intervention methods to reduce the allergenic potential of nuts, the natural variation in allergen gene sequence, expression, post-translational modification, stability and accumulation patterns in a model tree nut (pecan) will be studied. Less allergenic variants and factors that can be used to interfere with allergen accumulation in plants will also be characterized in detail. Collectively our studies will also contribute to the development of better detection tools and labeling practices for industry and regulatory agencies resulting in better protection of consumers.
Progress Report
This final report summarizes five years of research under project 6054-43440-052-00D. The project addressed four Objectives related to food allergen characterization, mitigation, and improved detection, focusing on peanut and tree nut allergens. A new project, 6054-43440-0XX-00D, will build on this work.
Objective 1: Research focused on identifying and characterizing small protein fragments called vicilin leader sequences (VLS), which were discovered by ARS scientists and registered with the WHO-Allergen Nomenclature Committee as a new class of allergens. The VLS were once thought to degrade during plant seed development. ARS researchers at New Orleans, Louisiana discovered that these fragments remain intact and can trigger allergic reactions when ingested. The researchers produced these leader sequences from peanut, walnut, pistachio, cashew, and tomato in recombinant form in bacteria or purified them from the plant seed. Structural analysis using nuclear magnetic resonance confirmed the stability of these fragments. Comparative studies with blood serum from allergic individuals showed that some leader sequences contribute to allergic cross-reactivity between different nuts.
Researchers developed specialized immunoassays using antibodies created to detect peanut and tree nut allergens. These assays helped characterize peanut allergens in different peanut flours and to identify how processing, such as roasting, alters allergenic properties. Results from these studies led to the classification of peanut flour as a drug product by Aimmune Therapeutics Inc. and approved for human use by the FDA, opening the door to the usage of peanut flour in therapeutic applications.
ARS scientists also evaluated allergen stability and potency in peanut flour and extracts provided by industry collaborators. They determined optimal storage conditions and extraction methods, which helped improve the quality and consistency of these allergenic materials. In additional work, researchers tested how bacterial and fungal fermentation affects peanut and tree nut flours. Although fermentation reduced some allergenic content, the products were still not safe for individuals with allergies.
Objective 2: The team worked to identify which parts of allergens trigger immune responses in people with food allergies. They focused on how two types of antibodies, immunoglobulin E (IgE) and immunoglobulin G4 (IgG4), interact with specific allergen segments. IgE is associated with allergic symptoms, while IgG4 may help reduce those symptoms.
To investigate this, researchers placed allergen fragments onto glass slides and exposed them to blood serum from over 400 individuals with peanut and tree nut allergies. This method allowed them to map IgE and IgG4 binding sites on more than 40 allergens. They observed how binding patterns changed before and after oral immunotherapy, which can help doctors monitor how well treatments work. Researchers also analyzed data using computational tools to improve predictions of allergenicity and inform diagnostic tests.
In additional studies, scientists synthesized altered versions of allergen peptides to determine how changes affect antibody recognition. This work identified key regions responsible for triggering immune responses and showed how genetic changes may reduce or alter allergenic potential.
Objective 3: Researchers evaluated genetic diversity and allergen profiles in multiple pecan cultivars and related hickory species. They used proteomic and immunological techniques to track how allergenic pecan proteins, such as Car i 1, Car i 2, and Car i 4, accumulated during nut development. The team identified hundreds of proteins from different developmental stages and found that allergen accumulation typically began in late September, when the nut tissue began to solidify.
They also developed antibodies that detect heat-stable markers of pecan allergens. These markers can help verify the presence of pecans in processed foods and may support food safety labeling.
Although the genetic variation among common pecan cultivars was limited, researchers found some differences in related hickory species. These findings provide targets for breeding new cultivars with reduced allergen content or better adaptation to climate stress. Scientists also discovered that one allergen, Car i 4, is prone to clumping during heating, which could affect its detectability in food processing.
In partnership with New Mexico State University, researchers analyzed new samples from diverse pecan cultivars and tissues using novel antibodies for allergen detection. They exchanged protocols and materials with collaborators to support the development of geographic and climate-adapted pecan trees.
Objective 4: Throughout the project, researchers worked to share their findings and improve methods for allergen detection and food safety. They contributed data to a large database to support broader allergen prediction models and shared tools and insights with other scientists, clinicians, and industry partners. They also assessed how processing steps such as roasting and fermentation affect allergen stability and immunoreactivity. Although some treatments reduced detectable allergen levels, none were sufficient to make products safe for allergic consumers.
In the final year, researchers evaluated proteins from glandless cottonseed and found that some cross-reacted with peanut and tree nut IgE antibodies. This result suggests a potential allergenic risk for new ingredients derived from cottonseed protein. Researchers also screened bacterial and fungal enzymes to identify candidates that could better degrade allergenic proteins and improve food safety.
Across all five years, this project made significant progress in understanding food allergens, developing new therapeutics and detection methods, and identifying strategies to reduce nut seed allergenicity. This work supports future efforts to improve food safety, develop hypoallergenic products, and provide better tools for diagnosis and therapy.
Accomplishments
1. National and global allergy data improve diagnostics and treatment potential. Immunoglobulin E (IgE) antibodies from the blood of allergic individuals cause allergic symptoms, while immunoglobulin G4 (IgG4) may help block those symptoms. Yet it remains unclear how these antibody responses differ across people in different regions. Researchers in New Orleans, Louisiana analyzed blood serum from about 1,000 individuals across the United States and other countries to study how IgE and IgG4 recognize peanut and tree nut allergens. Using microarray technology, they identified which parts of the allergens triggered immune responses and how those responses varied by geographic location. This work resulted in a large, searchable database that links allergen structure to regional antibody responses. This valuable resource will help scientists, healthcare providers, and allergy researchers better understand food allergies and develop more personalized and accurate allergy tests and treatments.
2. Identification of glandless cottonseed vicilin proteins as an allergen risk. Glandless cottonseed is a promising new source of plant-based protein because it lacks toxic compounds found in regular cottonseed. ARS researchers in New Orleans, Louisiana found that this protein breaks down easily during digestion and contains peptides that may offer health benefits. However, they also identified two proteins in glandless cottonseed that resemble a known peanut allergen. Lab tests using blood samples from people with peanut and tree nut allergies showed that about half reacted to these cottonseed proteins. This suggests that glandless cottonseed could pose a hidden risk to individuals with these allergies. The findings are important for food developers, allergy specialists, and the agricultural industry, as it highlights both the nutritional potential and the allergen concerns of this new ingredient. The research also provides clues about what makes peanut and tree nut allergens so reactive in the human body.
3. New insight into why peanut and tree nut allergies often overlap. Many people with peanut allergies also react to tree nuts, a problem known as cross-reactivity. This overlap can make allergy diagnoses difficult, especially since common tests rely on measuring how immune system antibodies called immunoglobulin E (IgE) respond to food proteins. ARS researchers in New Orleans, Louisiana studied a group of protein fragments called vicilin leader sequences, which are found in peanuts and tree nuts. They discovered that these fragments have physical similarities across different nuts, even though their sequences differ slightly. This small difference affects how IgE antibodies recognize them. In lab tests, these protein fragments helped distinguish between people with peanut allergies, walnut allergies, or both. These results could lead to more accurate testing for nut allergies and improve how doctors diagnose and treat allergic individuals. People with food allergies, healthcare providers, and diagnostic developers all benefit from a better understanding of what causes cross-reactions among nuts.
Review Publications
Vuong, T.T., Dupre, R.A., He, Z., Minkiewicz, P., Darewicz, M., Mattison, C.P. 2025. Glandless cottonseed C72 and GC72 vicilins cross-react with peanut and tree nut allergic IgE. ACS Food Science and Technology. https://doi.org/10.1021/acsfoodscitech.4c00577.
Brown, C.N., Dupre, R.A., Ebmeier, C.C., Patil, S.L., Smith, B., Mattison, C.P. 2025. Heating differentiates pecan allergen stability: Car i 4 is more heat labile than Car i 1 and Car i 2. Food Science and Nutrition. https://doi.org/10.1002/fsn3.4747.
Mattison, C.P., Dupre, R.A., Clermont, K., Gibbons, J.D., Yu, J. 2024. Proteomic characterization of peanut flour fermented by rhizopus oryzae. Heliyon. https://doi.org/10.1016/j.heliyon.2024.e34793.
Sabaghi, M., Maleki, S.J. 2024. Mitigating food protein allergy with biopolymers, bioactive compounds, and enzymes. Allergies. https://doi.org/10.3390/allergies4040016.
Musa, I., Ardalani, F., Yang, N., Maleki, S.J., Li, X. 2025. Murine model of cross-IgE sensitization and cross-anaphylactic reactions among multiple group food allergens. Frontiers in Immunology. https://doi.org/10.3389/fimmu.2024.1497368.
Gipson, S.A., Swientoniewski, L.T., Rogers, S.I., Mustafa, S.S., Dreskin, S.C., Teuber, S.S., Cheng, H., Maleki, S.J. 2025. Purification and epitope mapping of Jug r 4, a major walnut allergen. Allergies. https://doi.org/10.3390/allergies5010008.
Kabasser, S., Kamath, .T., Eber, E., Podzhilkova, A., Lupinek, C., Hemmer, W., Bublin, ., Croote, D., Maleki, S.J., Breiteneder, H., Hoffman-Sommergruber, K., Radauer, C., Bublin, M. 2025. Convergent monoclonal IgE antibodies from peanut allergic patients are multispecific to immunodominant epitopes of unrelated major peanut and tree nuts allergens.. Allergology International. https://doi.org/10.1016/j.alit.2025.05.007.