Development of a Sequence Searchable Database of Celiac Disease-Associated Peptides and Proteins for Risk Assessment of Novel Food Proteins.

Celiac disease (CeD) is an autoimmune enteropathy induced by prolamin and glutelin proteins in wheat, barley, rye, and triticale recognized by genetically restricted major histocompatibility (MHC) receptors. Patients with CeD must avoid consuming these proteins. Regulators in Europe and the United States expect an evaluation of CeD risks from proteins in genetically modified (GM) crops or novel foods for wheat-related proteins. Our database includes evidence-based causative peptides and proteins and two amino acid sequence comparison tools for CeD risk assessment. Sequence entries are based on the review of published studies of specific gluten-reactive T cell activation or intestinal epithelial toxicity. The initial database in 2012 was updated in 2018 and 2022. The current database holds 1,041 causative peptides and 76 representative proteins. The FASTA sequence comparison of 76 representative CeD proteins provides an insurance for possible unreported epitopes. Validation was conducted using protein homologs from Pooideae and non-Pooideae monocots, dicots, and non-plant proteins. Criteria for minimum percent identity and maximum E-scores are guidelines. Exact matches to any of the 1,041 peptides suggest risks, while FASTA alignment to the 76 CeD proteins suggests possible risks. Matched proteins should be tested further by CeD-specific CD4/8+ T cell assays or in vivo challenges before their use in foods.

Updated population minimal eliciting dose distributions for use in risk assessment of 14 priority food allergens.

Food allergy and allergen management are important global public health issues. In 2011, the first iteration of our allergen threshold database (ATDB) was established based on individual NOAELs and LOAELs from oral food challenge in roughly 1750 allergic individuals. Population minimal eliciting dose (EDp) distributions based on this dataset were published for 11 allergenic foods in 2014. Systematic data collection has continued (2011-2018) and the dataset now contains over 3400 data points. The current study provides new and updated EDp values for 14 allergenic foods and incorporates a newly developed Stacked Model Averaging statistical method for interval-censored data. ED01 and ED05 values, the doses at which 1%, and respectively 5%, of the respective allergic population would be predicted to experience any objective allergic reaction were determined. The 14 allergenic foods were cashew, celery, egg, fish, hazelnut, lupine, milk, mustard, peanut, sesame, shrimp (for crustacean shellfish), soy, walnut, and wheat. Updated ED01 estimates ranged between 0.03 mg for walnut protein and 26.2 mg for shrimp protein. ED05 estimates ranged between 0.4 mg for mustard protein and 280 mg for shrimp protein. The ED01 and ED05 values presented here are valuable in the risk assessment and subsequent risk management of allergenic foods.

Deriving individual threshold doses from clinical food challenge data for population risk assessment of food allergens.

BACKGROUND: Food allergies are a significant public health issue, and the only effective management option currently available is strict avoidance of all foods containing the allergen. In view of the practical impossibility of limiting risks to zero, quantitative allergen risk assessment and management strategies are needed. OBJECTIVE: We sought to develop appropriate methods for informing population-based risk assessments and risk management programs to benefit all stakeholders but particularly patients with food allergy. METHODS: Individual thresholds for food allergens (maximum tolerable doses and minimum eliciting doses) can ideally be established through double-blind, placebo-controlled food challenges. If double-blind, placebo-controlled food challenge data are not available, data from widely used open food challenges using predefined objective criteria can also provide useful data regarding minimum eliciting doses. For more than 20 years, the Netherlands Organisation for Applied Scientific Research and the Food Allergy Research and Resource Program at the University of Nebraska-Lincoln have been collecting individual maximum tolerable doses and minimum eliciting doses that produce objective symptoms from published and unpublished clinical data to better refine knowledge regarding the sensitivity of the population to food allergens. RESULTS: In this article we provide in-depth insights into the methodology applied by the Netherlands Organisation for Applied Scientific Research and Food Allergy Research and Resource Program to derive individual maximum tolerable doses and minimum eliciting doses for objective symptoms from clinical food challenge data. More than 90 examples for determining individual allergic thresholds are presented. CONCLUSION: With the methodology presented in this article, we aim to stimulate harmonization and transparency in quantitative food allergen risk assessment and risk management programs, encouraging their wider adoption.

Sensitivity analysis to derive a food consumption point estimate for deterministic food allergy risk assessment.

One of the input parameters in food allergy risk assessment is the amount of a given food consumed at an eating occasion. There is no consensus on how to use food consumption data when assessing the risk from unintended allergen presence in food products. A sensitivity analysis was performed to establish the optimal food consumption estimate for a deterministic food allergy risk assessment. Exposure was calculated for consumption percentiles (50th percentile, P50 to maximum) using the iFAAM consumption database in conjunction with an allergen concentration range from 1 to 1000 ppm. The resulting allergen intakes were compared to the allergic population reference doses proposed by Taylor et al. (2014) for 10 major allergenic foods. Optimal consumption percentiles were defined as those which predicted an intake below the relevant reference dose and met the defined acceptable risk level confirmed by probabilistic risk assessments. Analysis showed that, for 99% of the food groups, the P50 consumption met our criteria, while the P75 did so for 100% of the food groups. We suggest that the P75 is the optimal point estimate for use in deterministic food allergy risk assessment. It meets the safety objective and is adequately conservative for a public health context.

Quantitative Assessment of the Safety Benefits Associated with Increasing Clinical Peanut Thresholds Through Immunotherapy.

BACKGROUND: Peanut immunotherapy studies are conducted with the aim to decrease the sensitivity of patients to peanut exposure with the outcome evaluated by testing the threshold for allergic response in a double-blind placebo-controlled food challenge. The clinical relevance of increasing this threshold is not well characterized. OBJECTIVE: We aimed to quantify the clinical benefit of an increased threshold for peanut-allergic patients. METHODS: Quantitative risk assessment was performed by matching modeled exposure to peanut protein with individual threshold levels. Exposure was modeled by pairing US consumption data for various food product categories with potential contamination levels of peanut that have been demonstrated to be present on occasion in such food products. Cookies, ice cream, doughnuts/snack cakes, and snack chip mixes were considered in the risk assessment. RESULTS: Increasing the baseline threshold before immunotherapy from 100 mg or less peanut protein to 300 mg peanut protein postimmunotherapy reduces the risk of experiencing an allergic reaction by more than 95% for all 4 food product categories that may contain trace levels of peanut residue. Further increase in the threshold to 1000 mg of peanut protein had an additional quantitative benefit in risk reduction for all patients reacting to 300 mg or less at baseline. CONCLUSIONS: We conclude that achieving thresholds of 300 mg and 1000 mg of peanut protein by peanut immunotherapy is clinically relevant, and that the risk for peanut-allergic patients who have achieved this increased threshold to experience an allergic reaction is reduced in a clinically meaningful way.

Economic Factors Impacting Food Allergen Management: Perspectives from the Food Industry.

Food allergies affect up to 8% of children in the United States and may occasionally lead to severe life-threatening reactions. Because there is currently no cure for food allergies, strict avoidance of the allergen-containing foods is the only means of preventing an allergic reaction. Consumers rely on food manufacturers to reliably track and declare the presence of food allergens in products. Over the past 10 to 20 years, the food industry has increasingly adopted allergen control approaches in its processing facilities. However, the major industry costs related to food allergen management have not been fully described. The objective of this study was to characterize the factors that contribute to the economic impact of food allergen control practices on the food industry. A focus group (n = 100) was conducted with food industry professionals to identify key areas of cost for food allergen management. A survey based on the domains identified was then developed and disseminated to a convenience sample (n = 50) of quality control food industry specialists with knowledge of their company's food allergen management practices. Nearly all companies (92%) produced food products containing one or more of the top eight allergenic foods recognized by the U.S. Food and Drug Administration or sesame seeds. Cleaning procedures, employee training, and the potential for a recall due to allergen cross-contact were most frequently rated as the important factors in food allergen management. Recalls due to food allergen cross-contact, cleaning procedures, equipment and premises design, and employee training were ranked as the greatest allergen management expenses. Although 96% of companies had a food allergen control plan in place, nearly half (42%) had at least one food allergen-related recall within the past 5 years. The industry appears to endorse a willingness to unify precautionary allergen labeling to communicate a clear message more effectively to consumers.

Quantitative risk assessment of foods containing peanut advisory labeling.

Foods with advisory labeling (i.e. "may contain") continue to be prevalent and the warning may be increasingly ignored by allergic consumers. We sought to determine the residual levels of peanut in various packaged foods bearing advisory labeling, compare similar data from 2005 and 2009, and determine any potential risk for peanut-allergic consumers. Of food products bearing advisory statements regarding peanut or products that had peanut listed as a minor ingredient, 8.6% and 37.5% contained detectable levels of peanut (>2.5 ppm whole peanut), respectively. Peanut-allergic individuals should be advised to avoid such products regardless of the wording of the advisory statement. Peanut was detected at similar rates and levels in products tested in both 2005 and 2009. Advisory labeled nutrition bars contained the highest levels of peanut and an additional market survey of 399 products was conducted. Probabilistic risk assessment showed the risk of a reaction to peanut-allergic consumers from advisory labeled nutrition bars was significant but brand-dependent. Peanut advisory labeling may be overused on some nutrition bars but prudently used on others. The probabilistic approach could provide the food industry with a quantitative method to assist with determining when advisory labeling is most appropriate.

Soy in wheat--contamination levels and food allergy risk assessment.

In the United States, packaged food ingredients derived from allergenic sources must be clearly labeled. However, no requirement exists to declare the presence of residues of raw agricultural commodities due to agricultural commodity comingling. Clinical reports of allergic reactions to undeclared soy in wheat-based products do not exist suggesting that a rather low degree of risk is posed by wheat-based products that are comingled with soy. Detectable soybean residues (>2.5 ppm soy flour) were found in 62.8% of commercially available wheat flours at concentrations of 3-443 ppm soy flour (1.6-236 ppm soy protein). Conservative probabilistic risk assessments predict a risk of allergic reaction among the most sensitive soy-allergic individuals of 2.8±2.0 per 1000 soy-allergic user eating occasions of foods containing wheat flour. However, the predicted reactions occur at exposure levels below the lowest eliciting dose observed to provoke objective reactions in clinical oral soy challenges. Given this low level of predicted risk and the lack of evidence for allergic reactions among soy-allergic consumers to wheat-based products, the avoidance of wheat-based products by soy-allergic consumers does not appear to be necessary.

Survey of peanut levels in selected Irish food products bearing peanut allergen advisory labels.

Peanut allergy affects up to 2% of consumers and is responsible for the majority of fatalities caused by food-induced anaphylaxis. Peanut-containing products must be clearly labelled. Manufacturers are not legally required to label peanut if its inclusion resulted from unintentional cross contact with foods manufactured in the same facility. However, the use of allergen advisory statements alerting consumers of the potential presence of peanut allergen has increased in recent years. In previous studies, the vast majority of foods with precautionary allergen statements did not contain detectable levels of peanut, but no data are available on Irish food products. Thirty-eight food products bearing peanut/nut allergen-related statements were purchased from multiple locations in the Republic of Ireland and analysed for the presence of peanut. Peanut was detected in at least one lot in 5.3% (2 of 38) of the products tested. The doses of peanut detected ranged from 0.14 mg to 0.52 mg per suggested serving size (0.035-0.13 mg peanut protein). No detectable levels of peanut were found in the products that indicated peanut/nuts as a minor ingredient. Quantitative risk assessment, based on the known distribution of individual threshold doses for peanut, indicates that only a very small percentage of the peanut-allergic population would be likely to experience an allergic reaction to those products while the majority of products with advisory labels appear safe for the peanut-allergic population. Food manufacturers should be encouraged to analyse products manufactured in shared facilities and even on shared equipment with peanuts for peanut residues to determine whether sufficient risk exists to warrant the use of advisory labelling. Although it appears that the majority of food products bearing advisory nut statements are in fact free of peanut contamination, advice to peanut allergy sufferers to avoid said foods should continue in Ireland and therefore in the wider European Union.

Allergenicity assessment of genetically modified crops--what makes sense?

GM crops have great potential to improve food quality, increase harvest yields and decrease dependency on certain chemical pesticides. Before entering the market their safety needs to be scrutinized. This includes a detailed analysis of allergenic risks, as the safety of allergic consumers has high priority. However, not all tests currently being applied to assessing allergenicity have a sound scientific basis. Recent events with transgenic crops reveal the fallacy of applying such tests to GM crops.

Assessment of the potential allergenicity of a Milk Basic Protein fraction.

BACKGROUND: A specific basic fraction of bovine milk, termed Milk Basic Protein (MBP), has the potential to provide nutritionally important benefits if used as a food ingredient. Although derived from milk, MBP is intended for use as an ingredient in other foods. Cows' milk is a well studied, commonly allergenic food. Although the proteins in MBP are not identified as milk allergens, food products containing MBP will be labelled as containing milk as a caution to milk allergic consumers under food labelling guidelines in the US and the European Union as MBP has not been demonstrated to be free of milk allergens. However, as part of an overall safety evaluation of MBP, the developers sought to evaluate the potential allergenicity of the primary protein components for characteristics of allergenic food proteins and to assess whether intake of these proteins at intended use levels could present a significant new allergenic risk for consumers. OBJECTIVE: To evaluate the potential allergenicity of the five identified proteins in MBP. While extensive studies have not demonstrated allergenicity of lactoferrin, the four other proteins are less studied. The four were tested here by sequence identity comparison to known allergens, and for stability of these proteins in acidic pepsin as a characteristic common to many food allergens. METHODS: Sequences of the proteins were compared to those listed in AllergenOnline.com, by methods recommended for the evaluation of proteins introduced in crops through genetic engineering. Pepsin stability was assessed by incubating the various proteins in simulated gastric fluid at pH 1.2 with porcine pepsin for up to 60 min at 37 degrees C, with samples withdrawn and analyzed at specific times. RESULTS: No significant sequence similarities were identified for the MBP proteins compared to known allergens. All but one of the protein components of MBP were digested relatively quickly by pepsin. The more stable protein will be of low abundance as consumed in contrast to most pepsin-stable food allergens. CONCLUSIONS: Based on molecular characteristics and expected exposure, the protein components in MBP are unlikely to present any increased risk of allergy for milk allergic subjects or of cross-reactivity for other allergic subjects. However, since the proteins are derived from milk, products containing MBP will need to be labelled as containing milk proteins to warn milk allergic subjects of the potential risk of allergic reactions.

Safety assessment of foods produced through agricultural biotechnology.

Often the main criticism of foods derived from biotechnology is concerns about food safety. Whereas most present-day biotechnology-derived foods are approximately 99% similar to their non-biotechnology counterparts, the scientific community must ensure the safety of the novel aspects of these foods. The three phases of safety assessment are discussed and the concept of substantial equivalence is explained.

Protein allergenicity assessment of foods produced through agricultural biotechnology.

Foods produced through agricultural biotechnology are reaching the consumer marketplace. These novel foods should be assessed for their safety, including their potential allergenicity. Agricultural biotechnology involves the introduction of novel proteins into the modified foods, and proteins can be allergenic. The potential allergenicity of the introduced proteins can be evaluated by focusing on the source of the gene, the homology of the newly introduced protein to known allergens, the reactivity of the novel protein with IgE antibodies from the serum of individuals with known allergies to the source of the transferred DNA or to materials that are broadly related to the source of the transferred DNA, the resistance of the novel protein to pepsin, and the immunoreactivity of the novel protein in appropriate animal models. Additional factors, such as the level of expression of the novel protein in the modified food and expression in the edible portion of the food, may also yield valuable insights. Applying such criteria provides a reasonable approach to determining whether or not the novel protein is likely to become an allergen.