Phage Displayed Domain Antibodies (dAb) for Detection of Allergenic Pistachio Proteins in Foods.
Pistachio nuts (Pistacia vera) have been consumed by past and present-day civilizations because of their organoleptic characteristics and potential health benefits. However, they can also produce moderate to severe IgE-mediated reactions in allergic individuals. In this work, we report the isolation of the first recombinant antibodies against pistachio nut, produced without animal immunization, to be used in immunoassays for detection of allergenic pistachio in food products. Several phage display biopanning strategies were evaluated to screen the human-based domain antibody library (dAb) in search for pistachio-specific probes. The clone producing the PVF4 phage-dAb was finally selected, and it does not cross-react with cashew despite the phylogenetic proximity with pistachio. Western blot and matrix-assisted laser desorption/ionization tandem mass spectrometry (MALDI-TOF/TOF) analysis demonstrated that this clone recognised a unique band of ∼22 kDa related to the basic subunit of pistachio 11S globulin (allergen Pis v 2). The PVF4 phage-dAb allowed detection of pistachio in a food matrix with a limit of detection (LOD) of 3983 mg kg-1 in an indirect phage-enzyme-linked immunosorbent assay (ELISA). The ELISA method developed was used to assess applicability of the PVF4 phage-dAb for analysis of 77 commercial food products.
Sesame, Pistachio, and Macadamia Nut: Development and Validation of New Allergenic Systems for Fast Real-Time PCR Application.
Food allergy is a worldwide health problem that concerns infants to adults. The main health risk for sensitised individuals is due to the presence of traces of allergens as the result of an accidental contamination during food processing. The labelling of allergens such as sesame, pistachio, and macadamia nut on food products is mandatory according to Regulation (EU) N. 1169/2011; therefore, the development of suitable and specific analytical methodologies is advisable. The aim of this study was to perform a multi-allergen real-time PCR system that works well in fast mode at the same annealing temperature and with the same thermal profile. The real-time PCR was developed designing new, specific, and efficient primer and probe systems for the 2S albumingene for sesame and pistachio and for the vicilin precursorgene for macadamia nut. These systems were subjected to a robust intra-laboratory qualitative validation process prior to their application, by DNA extraction and fast real-time PCR, on some real market samples to reproduce a potential allergen contamination along the food chain. The developed system results were specific and robust, with a sensible limit of detection (0.005% for sesame; 0.004% for pistachio; 0.006% for macadamia nut). The performance and the reliability of the target systems were confirmed on commercial food samples. This molecular approach could be used as a screening or as a support tool, in association with the other widespread monitoring techniques (such as ELISA).
Are current analytical methods suitable to verify VITAL® 2.0/3.0 Allergen Reference doses for EU Allergens in Foods?
Food allergy affects up to 6% of Europeans. Allergen identification is important for the risk assessment and management of the inadvertent presence of allergens in foods. The VITAL® initiative for voluntary incidental trace allergen labeling suggests protein reference doses, based on clinical reactivity in food challenge studies, at or below which voluntary labelling is unnecessary. Here, we investigated if current analytical methodology could verify the published VITAL® 2.0 doses, that were available during this analysis, in serving sizes between 5 and 500 g. Available data on published and commercial ELISA, PCR and mass spectrometry methods, especially for the detection of peanuts, soy, hazelnut, wheat, cow's milk and hen's egg were reviewed in detail. Limit of detection, quantitative capability, matrix compatibility, and specificity were assessed. Implications by the recently published VITAL® 3.0 doses were also considered. We conclude that available analytical methods are capable of reasonably robust detection of peanut, soy, hazelnut and wheat allergens for levels at or below the VITAL® 2.0 and also 3.0 doses, with some methods even capable of achieving this in a large 500 gram serving size. Cow's milk and hen's egg are more problematic, largely due to matrix/processing incompatibility. An unmet need remains for harmonized reporting units, available reference materials, and method ring-trials to enable validation and the provision of comparable measurement results.
Risk of shared equipment in restaurants for peanut-allergic consumers: a simulation for preparing Asian foods.
Background: Allergic reactions to meals consumed outside the home are common, can be severe and sometimes fatal. Objective: To quantify the risk reduction potentially achieved by increasing an individual's threshold sensitivity to peanut (such as via immunotherapy) in scenarios of peanut exposure through shared kitchen materials in a restaurant setting. Methods: Three versions of popular peanut-containing sauces were selected to represent common ingredients used in Asian cooking. Different combinations of utensils, equipment, sauces and test conditions were prepared by a professional chef, with or without common cleaning procedures to represent normal daily practice. Residue amounts of peanut-containing material on kitchen equipment and utensils were measured and used for quantitative risk assessment to model the risk reduction associated with increasing an individual's threshold. Results: Shared utensils had mean residue amounts of 23-1519 mg peanut protein (no cleaning), and 3-82 mg peanut protein (after water rinse). Shared woks and pans had up to 20 mg peanut protein after rinsing. Individuals who reach a threshold of 300 mg peanut protein have a predicted relative risk reduction of 94.9 to >99.99% with brief cleaning. With no cleaning, relative risk reductions were 63.5 to 91.1% for individuals with a baseline threshold ≤100 mg peanut protein who reach a threshold of 300 mg peanut protein, increasing to 91 to 99.7% when reaching a threshold value of 1000 mg peanut protein. Conclusion: In all shared kitchen material scenarios that we studied, achieving an eliciting dose of 300 or 1000 mg peanut protein appears clinically relevant for the peanut-allergic population.
Prevalence of physician-reported food allergy in Canadian children.
Background: Food allergy prevalence data has largely been derived from self-report, and estimates vary.
Objective: Determine the prevalence of physician-reported food allergy in children using Electronic Medical Record (EMR) data from the Canadian Primary Care Sentinel Surveillance Network (CPSSN). Methods: Retrospective cohort study using the CPCSSN repository, Canada's only primary care practice-based surveillance system. Machine learning algorithms were applied to assess for food allergy documentation. Demographic information, chronic diseases of interest, prescribed medications, and health behaviours from the CPCSSN repository were identified. Results: The prevalence of physician-reported food allergy in Canadian children was 2.53% (95% CI:2.48%-2.59%). The most common food allergies documented were peanut (0.8% of children), tree nut (0.6%), cow's milk (0.4%), egg (0.3%), fruit (0.2%), finned fish (0.2%) and shellfish (0.2%). Among children with food allergy only 33.7% had an epinephrine autoinjector prescription. In logistic regression analysis, children with food allergy were more likely to have an atopic comorbidity (OR 2.20, 95%CI,2.06-2.35) and less likely to be obese than non-food allergic children (OR 0.84, 95%CI,0.78-0.90). In the age and sex-adjusted models, patients with food allergy were significantly more likely to have a psychiatric morbidity, specifically: ADHD (OR 1.81, 95%CI,1.66-1.96), autism (OR 1.89, 95%CI,1.63-2.19), and depression (OR 1.17, 95%CI,1.02-1.35). Conclusion: Our study is the first to estimate national physician-reported prevalence of food allergy, and demonstrates a lower rate than that based on self-report. Further studies into the association of food allergy and psychiatric comorbidities (ADHD, autism, depression) and the association of food allergy and obesity are needed.
Heat-Stable Hazelnut Profilin: Molecular Dynamics Simulations and Immunoinformatics Analysis.
Heat treatment can modify the allergenic potential, reducing allergenicity in specific proteins. Profilins are one of the important hazelnut allergens; these proteins are considered panallergens due to their high capacity for cross-reactivity with other allergens. In the present work, we evaluated the thermostability of hazelnut profilin, combining molecular dynamics simulation and immunoinformatic techniques. This approach helped us to have reliable results in immunogenicity studies. We modeled Cor a 2 profilin and applied annealing simulation, equilibrium, and production simulation at constant temperatures ranging from 300 to 500 K using Gromacs software. Despite the hazelnut profilins being able to withstand temperatures of up to 400 K, this does not seem to reduce its allergenicity. We have found that profilin subjected to temperatures of 450 and 500 K could generate cross-reactivity with other food allergens. In conclusion, we note a remarkable thermostability of Cor a 2 at 400 K which avoids its structural unfolding.
Implementation of the Addendum Guidelines for Peanut Allergy Prevention by US Allergists, a Survey conducted by the NIAID, in Collaboration with the AAAAI.
Background: In 2017, the Addendum Guidelines for the Prevention of Peanut Allergy were published with recommendations on early introduction of peanut-containing foods based on infants' clinical history. Objective: To conduct a nationwide U.S. survey to assess Guidelines implementation among allergists and immunologists who manage infants for food allergy. Methods: Survey invitations were delivered to 3281 non-retired, U.S. members of the American Academy of Asthma, Allergy, and Immunology, board certified in allergy and immunology. The survey assessed awareness and implementation of the Guidelines and barriers to implementation. Descriptive statistics were generated. Results: Twenty-nine percent (946 of 3281) of surveyed allergists/immunologists responded, and 87.1% (825 of 946) of responders met eligibility criteria. Among eligible responders, 97.1% were aware of the Guidelines. Of those, 64.5% reported full implementation of the Guidelines as published, 34.4% reported partial implementation, and 1.1% reported using none of the Guidelines. Barriers to Guidelines use included parental (47.6%) and self (21.8%) concerns about allergic reactions, lack of referrals (33.6%), parents uninterested in early feeding (28.2%), and lack of clinic time (20.9%). The two most common deviations from the Guidelines were considering additional factors not specified in the Guidelines such as family history (50.2%) and conducting skin prick testing in non-high-risk children (43.9%). Of respondents using the Guidelines, 45.7% indicated they needed more education or training. Conclusion: Essentially all allergists/immunologists who responded to the survey reported full or partial Guidelines implementation. Parental concerns and lack of referrals are major identifiable barriers. Improved Guidelines messaging to parents and referring physicians is warranted.
A Pilot Study on the Effects of Nut Consumption on Cardiovascular Biomarkers.
Background: Cardiovascular disease (CVD) is a leading cause of morbidity and mortality in the United States and changes in lifestyle can minimize the likelihood of succumbing to heart disease. Anti-inflammatory agents are commonly used to reduce the chronic inflammatory state behind the pathogenesis of CVD. Multiple studies have been published correlating nut consumption with a reduction in both heart attacks and strokes. The goal of this study is to determine to what extent the consumption of almonds, hazelnuts, and walnuts have on the blood markers associated with cardiac disease and inflammation. Methods: This was a six-week study in which subject's baseline values act as controls. Fasting blood draws occurred at week 0, week 2, and after four weeks of intervention (week 6). All participants had undesirable lipid profiles and no medications related to heart disease. Results: Total cholesterol (TC): high-density lipoprotein (HDL-C) ratio was lowered a statistically significant amount at the six-week time point (3.89 ± 0.74) compared to both the zero-week (4.93 ± 1.16, p < 0.01) and two-week (4.63 ± 1.20, p < 0.5) timepoints. Low-density lipoprotein (LDL) measurements were lowered a statistically significant amount at the six-week time point (135.6 ± 15.0 mg/dL) compared to the zero-week (159.7 ± 12.3 mg/dL, p < 0.01). Erythrocyte sedimentation rate (ESR) was lowered a statistically significant amount at six-week time point (10.44 ± 5.05 mm/h) compared to the zero-week (14.44 ± 5.12 mm/h, p < 0.01). Conclusions: Blood markers associated with CVD specifically and the general marker for inflammation associated with many chronic diseases can be favorably modified with the consumption of specific nuts as demonstrated by this study.
10.7759/cureus.8798
Long-Term, Open-Label Extension Study of the Efficacy and Safety of Epicutaneous Immunotherapy for Peanut Allergy in Children: PEOPLE 3-Year Results.
Background: We previously reported the safety and efficacy of epicutaneous immunotherapy (EPIT™) for peanut allergy (250 μg, daily epicutaneous peanut protein; DBV712 250μg) in a 12-month randomized controlled study (PEPITES) of peanut-allergic children aged 4-11 years. Objective: To assess interim safety and efficacy of an additional 2 years of EPIT from the ongoing (5-year treatment) open-label extension PEOPLE study. Methods: Subjects who completed PEPITES were offered enrollment in PEOPLE. Following an additional 2 years of daily DBV712 250μg, subjects who had received DBV712 250μg in PEPITES underwent Month-36 double-blind, placebo-controlled, food challenge (DBPCFC) with an optional Month-38 sustained unresponsiveness (SU) assessment. Results: 198 (93%) of 213 eligible subjects who had received DBV712 250μg in PEPITES entered PEOPLE, of whom 141 (71%) had assessable DBPCFC at Month 36. At Month 36, 51.8% (73/141) of subjects reached an eliciting dose (ED) of ≥1000 mg, compared with 40.4% (57/141) at Month 12. 75.9% (107/141) demonstrated increased ED compared to baseline. 13.5% (19/141) tolerated the full DBPCFC of 5444 mg. Median cumulative reactive dose increased from 144 to 944 mg. 18 subjects underwent an optional SU assessment; 14/18 (77.8%) maintained an ED of ≥1000 mg at Month 38. Local patch-site skin reactions were common but decreased over time. There was no treatment-related epinephrine use in years 2 or 3. Compliance was high (96.9%), and withdrawals due to treatment-related adverse events low (1%). Conclusion: These results demonstrate that daily EPIT treatment for peanut allergy beyond 1 year leads to continued response from a well-tolerated, simple-to-use regimen.
Natural Variation of Hazelnut Allergenicity: Is There Any Potential for Selecting Hypoallergenic Varieties?.
Hazelnuts (Corylus avellana L.) have an important role in human nutrition and health. However, they are a common cause of food allergy. Due to hazelnut varietal diversity, variety-dependent differences in the IgE-binding properties may be suspected, which could allow therapeutic strategies based on the use of hypoallergenic varieties to induce desensitization. In a proteogenomic approach, we aimed to evaluate the allergenic potential of a genetically diverse set of hazelnuts (n = 13 varieties). Minor differences were found at the level of genes encoding important allergens, namely Cor a 8, Cor a 9, and Cor a 14. Nevertheless, IgE-reactivity was similar for all varieties using sera from seven allergic individuals. The predominant IgE-reactive proteins were Cor a 9 (100%) and Cor a 1.04 (60%), with the former being the most frequently identified by a two-dimensional gel electrophoresis (2-DE)-based proteomic approach. Therefore, it seems that the conventional exclusion diet will hold its ground for the time being.