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Background and Objectives: Background: Cypress and olive pollen are the most prevalent sensitizers in the Mediterranean area. Some patients exhibit dual sensitization, which has not been well documented to date. Objective: To identify the allergens involved in dual cypress and olive allergy (C+O) and to study the relationship between phenotype and allergen sensitization., Methods: C+O patients were selected. Those monosensitized to olive or cypress were used as a reference. Specific IgE to whole extracts and purified allergens from olive and cypress was determined. Immunoblotting was performed to analyze IgG and IgE binding using olive polyclonal antibodies and patients' sera, respectively. Mutual immunoblotting inhibition of olive and cypress extracts and inhibition of cypress extract immunoblotting with olive allergens were performed. Multiple correspondence analysis and hierarchical cluster classifications were conducted to analyze the relationships between the clinical presentation of C+O (symptoms, seasonality) and allergen profile., Results: C+O patients were clustered in 4 phenotypes. The most frequent one (58.4%) was rhinoconjunctivitis in winter (February) and spring (May), with asthma in 38% of patients. Ole e 1 and Cup s 1 were the major allergens. Proteins homologous to Ole e 1, Ole e 9, and Ole e 11 in cypress pollen were identified and shown to inhibit IgE binding to cypress extract., Conclusions: The exclusive C+O results from cosensitization to Cup s 1 and Ole e 1 and cross-reactivity due to previously unreported Ole e 1-like, Ole e 9-like, and Ole e 11-like allergens in cypress pollen. Our findings point to 4 clinical phenotypes of winter and/or spring or perennial rhinoconjunctivitis with and without asthma.

Food allergy is a global public health problem that until recent years lacked any aetiological treatment supported by academy, industry and regulators. Food immunotherapy (AIT) is an evolving treatment option, supported by clinical practice and industry trial data. Recent AIT meta-analyses have highlighted the difficulty in pooling safety and efficacy data from AIT trials, due to secondary heterogeneity in the study. An EAACI task force (CO-FAITH) initiated by the Paediatric Section was created to focus on AIT efficacy outcomes for milk, egg and peanut allergy rather than in trial results. A systematic search and a narrative review of AIT controlled clinical trials and large case series was conducted. A total of 63 manuscripts met inclusion criteria, corresponding to 23, 21 and 22 studies of milk, egg and peanut AIT, respectively. The most common AIT efficacy outcome was desensitization, mostly defined as tolerating a maintenance phase dose, or reaching a particular dose upon successful exit oral food challenge (OFC). However, a large degree of heterogeneity was identified regarding the dose quantity defining this outcome. Sustained unresponsiveness and patient-reported outcomes (e.g. quality of life) were explored less frequently, and to date have been most rigorously described for peanut AIT versus other allergens. Change in allergen threshold assessed by OFC remains the most common efficacy measure, but OFC methods suffer from heterogeneity and methodological disparity. This review has identified multiple heterogeneous outcomes related to measuring the efficacy of AIT. Efforts to better standardize and harmonize which outcomes, and how to measure them must be carried out to help in the clinical development of safe and efficacious food allergy treatments., (© 2024 European Academy of Allergy and Clinical Immunology and John Wiley & Sons Ltd.)

Background: Primary hazelnut allergy is a common cause of anaphylaxis in children, as compared to birch-pollen associated hazelnut allergy. Population-based data on hazelnut and concomitant birch-pollen allergy in children are lacking. We aimed to investigate the prevalence of primary and pollen-associated hazelnut allergy and sensitization profiles in school-aged children in Berlin, Germany., Methods: 1570 newborn children were recruited in Berlin in 2005-2009. The school-age follow-up (2014-2017) was based on a standardized web-based parental questionnaire and clinical evaluation by a physician including skin prick tests, allergen specific immunoglobulin E serum tests and placebo-controlled double-blind oral food challenges, if indicated., Results: 1004 children (63.9% response) participated in the school-age follow-up assessment (52.1% male). For 1.9% (n = 19, 95%-confidence interval 1.1%-2.9%) of children their parents reported hazelnut-allergic symptoms, for half of these to roasted hazelnut indicating primary hazelnut allergy. Symptoms of birch-pollen allergy were reported for 11.6% (n = 116 95%-CI 9.7%-13.7%) of the children. Both birch-pollen allergy and hazelnut allergy associated symptoms affected 0.6% (n = 6, 95%-CI 0.2%-1.3%) of children. Assessment of allergic sensitization was performed in 261 participants and showed that almost 20% of these children were sensitized to hazelnut, being the most frequent of all assessed food allergens, or birch-pollen, the majority to both., Conclusions: Based on parental reports hazelnut-allergic symptoms were far less common than sensitization to hazelnut. This needs to be considered by physicians to avoid unnecessary changes in diet due to sensitization profiles only, especially when there is a co-sensitization to hazelnut and birch-pollen., (Copyright © 2021 Japanese Society of Allergology. Production and hosting by Elsevier B.V. All rights reserved.)

Background: In allergology, the intradermal approach is generally used to establish an aetiological diagnosis, with limited experience in specific allergen immunotherapy., Objective: To evaluate the efficacy and safety of immunotherapy with an allergen extract of glutaraldehyde-polymerized Phleum pratense, administered intradermally, in patients with rhinoconjunctivitis sensitized to grass pollen., Methods: Multicentre, randomized, double-blind, placebo-controlled clinical trial in patients from 12 to 65 years of age with rhinitis or rhinoconjunctivitis, with or without asthma, due to grass pollen allergy. Patients were divided into three groups and received a total of six doses in a weekly interval, of either placebo; 0.03 or 0.06 μg of protein per dose of P pratense allergoid. The primary objective was to evaluate the combined symptoms and medication consumption score (CSMS). The secondary objectives were symptoms and medication, tolerance to the conjunctival provocation test, specific IgE and IgG4 antibodies and the safety profile according to the WAO scale., Results: The dose of 0.06 μg of protein proved to be effective versus the placebo by significantly reducing CSMS and increasing tolerance to the allergenic extract in the conjunctival provocation test, after the first pollen season. This group showed a significant reduction in specific IgE after the second pollen season relative to the baseline. There were no variations in IgG4 levels. Only one grade 2 systemic reaction was recorded., Conclusion & Clinical Relevance: Intradermal immunotherapy with P pratense allergoid has been shown to be effective and safe, reducing CSMS, increasing tolerance to the conjunctival provocation test and reducing IgE levels., (© 2020 The Authors. Clinical & Experimental Allergy published by John Wiley & Sons Ltd.)

Background: Peanut allergy is the leading cause of food-related anaphylaxis. Current management options can negatively affect food allergy-related quality of life. We aimed to investigate the efficacy of an investigational oral biologic drug (AR101)., Methods: The AR101 Trial in Europe Measuring Oral Immunotherapy Success in peanut-allergic children (ARTEMIS) trial was a multicentre, double-blind, randomised, placebo-controlled phase 3 trial done at 18 hospitals in Ireland, France, Germany, Italy, Spain, Sweden, and the UK. Children and adolescents with peanut allergy, aged 4-17 years, who developed dose-limiting symptoms to 300 mg or less peanut protein (equivalent to approximately one peanut kernel) during a double-blind placebo-controlled food challenge test at study entry were enrolled. Participants were randomly assigned (3:1) to receive daily doses of either AR101 oral immunotherapy (AR101 group) or a taste-masked placebo (placebo group). All participants, investigators, and care providers were masked to treatment allocation until the study was completed. Doses were increased every 2 weeks over 6 months until a dose of 300 mg was reached and maintained for 3 months. The primary endpoint was the proportion of participants in the intention-to-treat or safety population (defined as those participants who had been randomly assigned and had received at least one dose of the assigned drug) who could consume a single dose of 1000 mg (cumulative dose 2043 mg) peanut protein without developing dose-limiting allergic symptoms at an exit double-blind placebo-controlled food challenge after 9 months of treatment. Additional endpoints included safety (ie, the frequency and severity of adverse events) and changes in food allergy-related quality of life, assessed by use of age-appropriate Food Allergy Quality of Life Questionnaires (FAQLQs) and the Food Allergy Independent Measure (FAIM). The study is registered with ClinicalTrials.gov, NCT03201003, and is completed., Findings: Between June 12, 2017, and Feb 15, 2018, 227 patients were screened, of whom 175 were randomly assigned to the AR101 group (n=132) and the placebo group (n=43). All primary and secondary endpoints were met. 77 (58%) of 132 participants in the AR101 group tolerated 1000 mg peanut protein at the exit food challenge versus one (2%) of 43 participants in the placebo group (AR101-placebo treatment difference 56·0% [95% CI 44·1-65·2], p<0·0001). Adverse events were reported by almost all participants. The maximum severity of adverse events reported was mild or moderate for most participants who received AR101 (mild, 66 [50%] of 132 participants; moderate, 63 [48%]; and severe, one [1%]) or placebo (mild, 24 [56%] of 43 participants; moderate, 18 [42%]; severe, none). Participants aged 8-12 years in the AR101 group reported improvements that exceeded the minimum clinically important difference between the two groups across all FAQLQ domains. Additionally, participants in the AR101 group and their caregivers reported improvements that exceeded the minimum clinically important difference in FAIM domains related to the perceived likelihood and outcomes of a severe allergic reaction., Interpretation: AR101 oral immunotherapy treatment led to rapid desensitisation to peanut protein, with a predictable safety profile that improved with treatment, and an associated improvement in self-reported and caregiver-reported food allergy-related quality of life. These patient-oriented outcomes provide invaluable data to help physicians, patients, and caregivers make informed, shared decisions on the management of peanut allergy., Funding: Aimmune Therapeutics., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Background: The geographical variation and temporal increase in the prevalence of food sensitization (FS) suggest environmental influences., Objective: To investigate how environment, infant diet, and demographic characteristics, are associated with FS in children and adults, focusing on early-life exposures., Methods: Data on childhood and adult environmental exposures (including, among others, sibship size, day care, pets, farm environment, and smoking), infant diet (including breast-feeding and timing of introduction to infant formula and solids), and demographic characteristics were collected from 2196 school-age children and 2185 adults completing an extensive questionnaire and blood sampling in the cross-sectional pan-European EuroPrevall project. Multivariable logistic regression was applied to determine associations between the predictor variables and sensitization to foods commonly implicated in food allergy (specific IgE ≥0.35 kU A /L). Secondary outcomes were inhalant sensitization and primary (non-cross-reactive) FS., Results: Dog ownership in early childhood was inversely associated with childhood FS (odds ratio, 0.65; 95% CI, 0.48-0.90), as was higher gestational age at delivery (odds ratio, 0.93 [95% CI, 0.87-0.99] per week increase in age). Lower age and male sex were associated with a higher prevalence of adult FS (odds ratio, 0.97 [95% CI, 0.96-0.98] per year increase in age, and 1.39 [95% CI, 1.12-1.71] for male sex). No statistically significant associations were found between other evaluated environmental determinants and childhood or adult FS, nor between infant diet and childhood FS, although early introduction of solids did show a trend toward prevention of FS., Conclusions: Dog ownership seems to protect against childhood FS, but independent effects of other currently conceived environmental and infant dietary determinants on FS in childhood or adulthood could not be confirmed., (Copyright © 2020 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Understanding consumer perceptions is crucial if effective food safety policy and risk communication are to be developed and implemented. We sought to understand how those living with food allergy assess risk with precautionary allergen labelling (PAL) and their preference in how risks are communicated within a quantitative risk assessment (QRA) framework., Methods: The Integrated Approaches to Food Allergen and Allergy Risk Management (iFAAM) labelling online survey was developed for adults and parents of children with food allergy and distributed across Germany, Ireland, Netherlands, Spain and UK via patient support groups., Results: There were 1560 complete responses. 'This product is not suitable for' was selected as first choice for PAL by 46% overall and 'May contain' was selected as the first choice by 44%. Seventy-three percent reported that it would improve their trust in a product if a QRA process had been used to make a decision about whether to include 'may contain'. Overall, 66% reported that a 'statement + symbol' on the label indicating a QRA, would help them to understand the risk assessment process that had been used by the food manufacturer., Conclusions: Consumers want to know what process has actually taken place for the placing of a PAL and/or risk assessment statement on a particular food product. Our findings provide a basis for the development of more informative communication around food allergen risk and safety and support evidence-based policy-making in the context of the legislative requirements of the European Union's Food Information for Consumers Regulation., (© 2019 John Wiley & Sons Ltd.)

Allergen immunotherapy (AIT) has been in use for the treatment of allergic disease for more than 100 years. Asthma treatment relies mainly on corticosteroids and other controllers recommended to achieve and maintain asthma control, prevent exacerbations, and improve quality of life. AIT is underused in asthma, both in children and in adults. Notably, patients with allergic asthma not adequately controlled on pharmacotherapy (including biologics) represent an unmet health need. The European Academy of Allergy and Clinical Immunology has developed a clinical practice guideline providing evidence-based recommendations for the use of house dust mites (HDM) AIT as add-on treatment for HDM-driven allergic asthma. This guideline was developed by a multi-disciplinary working group using the Grading of Recommendations Assessment, Development and Evaluation (GRADE) approach. HDM AIT was separately evaluated by route of administration and children and adults: subcutaneous (SCIT) and sublingual AIT (SLIT), drops, and tablets. Recommendations were formulated for each. The important prerequisites for successful treatment with HDM AIT are (a) selection of patients most likely to respond to AIT and (b) use of allergen extracts and desensitization protocols of proven efficacy. To date, only AIT with HDM SLIT-tablet has demonstrated a robust effect in adults for critical end points (exacerbations, asthma control, and safety). Thus, it is recommended as an add-on to regular asthma therapy for adults with controlled or partially controlled HDM-driven allergic asthma (conditional recommendation, moderate-quality evidence). HDM SCIT is recommended for adults and children, and SLIT drops are recommended for children with controlled HDM-driven allergic asthma as the add-on to regular asthma therapy to decrease symptoms and medication needs (conditional recommendation, low-quality evidence)., (© 2019 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

Background: Peanut allergy, for which there are no approved treatment options, affects patients who are at risk for unpredictable and occasionally life-threatening allergic reactions., Methods: In a phase 3 trial, we screened participants 4 to 55 years of age with peanut allergy for allergic dose-limiting symptoms at a challenge dose of 100 mg or less of peanut protein (approximately one third of a peanut kernel) in a double-blind, placebo-controlled food challenge. Participants with an allergic response were randomly assigned, in a 3:1 ratio, to receive AR101 (a peanut-derived investigational biologic oral immunotherapy drug) or placebo in an escalating-dose program. Participants who completed the regimen (i.e., received 300 mg per day of the maintenance regimen for approximately 24 weeks) underwent a double-blind, placebo-controlled food challenge at trial exit. The primary efficacy end point was the proportion of participants 4 to 17 years of age who could ingest a challenge dose of 600 mg or more, without dose-limiting symptoms., Results: Of the 551 participants who received AR101 or placebo, 496 were 4 to 17 years of age; of these, 250 of 372 participants (67.2%) who received active treatment, as compared with 5 of 124 participants (4.0%) who received placebo, were able to ingest a dose of 600 mg or more of peanut protein, without dose-limiting symptoms, at the exit food challenge (difference, 63.2 percentage points; 95% confidence interval, 53.0 to 73.3; P<0.001). During the exit food challenge, the maximum severity of symptoms was moderate in 25% of the participants in the active-drug group and 59% of those in the placebo group and severe in 5% and 11%, respectively. Adverse events during the intervention period affected more than 95% of the participants 4 to 17 years of age. A total of 34.7% of the participants in the active-drug group had mild events, as compared with 50.0% of those in the placebo group; 59.7% and 44.4% of the participants, respectively, had events that were graded as moderate, and 4.3% and 0.8%, respectively, had events that were graded as severe. Efficacy was not shown in the participants 18 years of age or older., Conclusions: In this phase 3 trial of oral immunotherapy in children and adolescents who were highly allergic to peanut, treatment with AR101 resulted in higher doses of peanut protein that could be ingested without dose-limiting symptoms and in lower symptom severity during peanut exposure at the exit food challenge than placebo. (Funded by Aimmune Therapeutics; PALISADE ClinicalTrials.gov number, NCT02635776 .).

Regulatory approaches for allergen immunotherapy (AIT) products and the availability of high-quality AIT products are inherently linked to each other. While allergen products are available in many countries across the globe, their regulation is very heterogeneous. First, we describe the regulatory systems applicable for AIT products in the European Union (EU) and in the United States (US). For Europe, a depiction of the different types of relevant procedures, as well as the committees involved, is provided and the fundamental role of national agencies of the EU member states in this complex and unique network is highlighted. Furthermore, the regulatory agencies from Australia, Canada, Japan, Russia, and Switzerland provided information on the system implemented in their countries for the regulation of allergen products. While AIT products are commonly classified as biological medicinal products, they are made available by varying types of procedures, most commonly either by obtaining a marketing authorization or by being distributed as named patient products. Exemptions from marketing authorizations in exceptional cases, as well as import of allergen products from other countries, are additional tools applied by countries to ensure availability of needed AIT products. Several challenges for AIT products are apparent from this analysis and will require further consideration., (© 2017 EAACI and John Wiley and Sons A/S. Published by John Wiley and Sons Ltd.)

Background: The European Academy of Allergy and Clinical Immunology (EAACI) is developing Guidelines for Allergen Immunotherapy (AIT) for IgE-mediated Food Allergy. To inform the development of clinical recommendations, we sought to critically assess evidence on the effectiveness, safety and cost-effectiveness of AIT in the management of food allergy., Methods: We undertook a systematic review and meta-analysis that involved searching nine international electronic databases for randomized controlled trials (RCTs) and nonrandomized studies (NRS). Eligible studies were independently assessed by two reviewers against predefined eligibility criteria. The quality of studies was assessed using the Cochrane Risk of Bias tool for RCTs and the Cochrane ACROBAT-NRS tool for quasi-RCTs. Random-effects meta-analyses were undertaken, with planned subgroup and sensitivity analyses., Results: We identified 1814 potentially relevant papers from which we selected 31 eligible studies, comprising of 25 RCTs and six NRS, studying a total of 1259 patients. Twenty-five trials evaluated oral immunotherapy (OIT), five studies investigated sublingual immunotherapy, and one study evaluated epicutaneous immunotherapy. The majority of these studies were in children. Twenty-seven studies assessed desensitization, and eight studies investigated sustained unresponsiveness postdiscontinuation of AIT. Meta-analyses demonstrated a substantial benefit in terms of desensitization (risk ratio (RR) = 0.16, 95% CI 0.10, 0.26) and suggested, but did not confirm sustained unresponsiveness (RR = 0.29, 95% CI 0.08, 1.13). Only one study reported on disease-specific quality of life (QoL), which reported no comparative results between OIT and control group. Meta-analyses revealed that the risk of experiencing a systemic adverse reaction was higher in those receiving AIT, with a more marked increase in the risk of local adverse reactions. Sensitivity analysis excluding those studies judged to be at high risk of bias demonstrated the robustness of summary estimates of effectiveness and safety of AIT for food allergy. None of the studies reported data on health economic analyses., Conclusions: AIT may be effective in raising the threshold of reactivity to a range of foods in children with IgE-mediated food allergy whilst receiving (i.e. desensitization) and post-discontinuation of AIT. It is, however, associated with a modest increased risk in serious systemic adverse reactions and a substantial increase in minor local adverse reactions. More data are needed in relation to adults, long term effects, the impact on QoL and the cost-effectiveness of AIT., (© 2017 The Authors. Allergy Published by John Wiley & Sons Ltd.)

Background: The conduct of oral food challenges as the preferred diagnostic standard for food allergy (FA) was harmonized over the last years. However, documentation and interpretation of challenge results, particularly in research settings, are not sufficiently standardized to allow valid comparisons between studies. Our aim was to develop a diagnostic toolbox to capture and report clinical observations in double-blind placebo-controlled food challenges (DBPCFC)., Methods: A group of experienced allergists, paediatricians, dieticians, epidemiologists and data managers developed generic case report forms and standard operating procedures for DBPCFCs and piloted them in three clinical centres. The follow-up of the EuroPrevall/iFAAM birth cohort and other iFAAM work packages applied these methods., Recommendations: A set of newly developed questionnaire or interview items capture the history of FA. Together with sensitization status, this forms the basis for the decision to perform a DBPCFC, following a standardized decision algorithm. A generic form including details about severity and timing captures signs and symptoms observed during or after the procedures. In contrast to the commonly used dichotomous outcome FA vs no FA, the allergy status is interpreted in multiple categories to reflect the complexity of clinical decision-making., Conclusion: The proposed toolbox sets a standard for improved documentation and harmonized interpretation of DBPCFCs. By a detailed documentation and common terminology for communicating outcomes, these tools hope to reduce the influence of subjective judgment of supervising physicians. All forms are publicly available for further evolution and free use in clinical and research settings., (© 2016 The Authors. Allergy Published by John Wiley & Sons Ltd.)

Anaphylaxis has been defined as a 'severe, life-threatening generalized or systemic hypersensitivity reaction'. However, data indicate that the vast majority of food-triggered anaphylactic reactions are not life-threatening. Nonetheless, severe life-threatening reactions do occur and are unpredictable. We discuss the concepts surrounding perceptions of severe, life-threatening allergic reactions to food by different stakeholders, with particular reference to the inclusion of clinical severity as a factor in allergy and allergen risk management. We review the evidence regarding factors that might be used to identify those at most risk of severe allergic reactions to food, and the consequences of misinformation in this regard. For example, a significant proportion of food-allergic children also have asthma, yet almost none will experience a fatal food-allergic reaction; asthma is not, in itself, a strong predictor for fatal anaphylaxis. The relationship between dose of allergen exposure and symptom severity is unclear. While dose appears to be a risk factor in at least a subgroup of patients, studies report that individuals with prior anaphylaxis do not have a lower eliciting dose than those reporting previous mild reactions. It is therefore important to consider severity and sensitivity as separate factors, as a highly sensitive individual will not necessarily experience severe symptoms during an allergic reaction. We identify the knowledge gaps that need to be addressed to improve our ability to better identify those most at risk of severe food-induced allergic reactions., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Data are lacking regarding the prevalence of food sensitization and probable food allergy among general population in India. We report the prevalence of sensitization and probable food allergy to 24 common foods among adults from general population in Karnataka, South India., Methodology: The study was conducted in two stages: a screening study and a case-control study. A total of 11 791 adults in age group 20-54 were randomly sampled from general population in South India and answered a screening questionnaire. A total of 588 subjects (236 cases and 352 controls) participated in the case-control study involving a detailed questionnaire and specific IgE estimation for 24 common foods., Results: A high level of sensitization (26.5%) was observed for most of the foods in the general population, higher than that observed among adults in Europe, except for those foods that cross-react with birch pollen. Most of the sensitization was observed in subjects who had total IgE above the median IgE level. A high level of cross-reactivity was observed among different pollens and foods and among foods. The prevalence of probable food allergy (self-reports of adverse symptoms after the consumption of food and specific IgE to the same food) was 1.2%, which was mainly accounted for cow's milk (0.5%) and apple (0.5%)., Conclusion: Very high levels of sensitization were observed for most foods, including those not commonly consumed in the general population. For the levels of sensitization, the prevalence of probable food allergy was low. This disassociation needs to be further explored in future studies., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The availability of allergen molecules ('components') from several protein families has advanced our understanding of immunoglobulin E (IgE)-mediated responses and enabled 'component-resolved diagnosis' (CRD). The European Academy of Allergy and Clinical Immunology (EAACI) Molecular Allergology User's Guide (MAUG) provides comprehensive information on important allergens and describes the diagnostic options using CRD. Part A of the EAACI MAUG introduces allergen molecules, families, composition of extracts, databases, and diagnostic IgE, skin, and basophil tests. Singleplex and multiplex IgE assays with components improve both sensitivity for low-abundance allergens and analytical specificity; IgE to individual allergens can yield information on clinical risks and distinguish cross-reactivity from true primary sensitization. Part B discusses the clinical and molecular aspects of IgE-mediated allergies to foods (including nuts, seeds, legumes, fruits, vegetables, cereal grains, milk, egg, meat, fish, and shellfish), inhalants (pollen, mold spores, mites, and animal dander), and Hymenoptera venom. Diagnostic algorithms and short case histories provide useful information for the clinical workup of allergic individuals targeted for CRD. Part C covers protein families containing ubiquitous, highly cross-reactive panallergens from plant (lipid transfer proteins, polcalcins, PR-10, profilins) and animal sources (lipocalins, parvalbumins, serum albumins, tropomyosins) and explains their diagnostic and clinical utility. Part D lists 100 important allergen molecules. In conclusion, IgE-mediated reactions and allergic diseases, including allergic rhinoconjunctivitis, asthma, food reactions, and insect sting reactions, are discussed from a novel molecular perspective. The EAACI MAUG documents the rapid progression of molecular allergology from basic research to its integration into clinical practice, a quantum leap in the management of allergic patients., (© 2016 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

The purpose of this brief communication is to highlight emerging evidence regarding potential benefits of supporting early rather than delayed peanut introduction during the period of complementary food introduction in infants. This document should be considered as interim guidance based on consensus among the following organizations: American Academy of Allergy, Asthma, and Immunology, American Academy of Pediatrics, American College of Allergy, Asthma, and Immunology, Australasian Society of Clinical Immunology and Allergy, Canadian Society of Allergy and Clinical Immunology, European Academy of Allergy and Clinical Immunology, Israel Association of Allergy and Clinical Immunology, Japanese Society for Allergology, Society for Pediatric Dermatology, and World Allergy Organization. More formal guidelines regarding early-life, complementary feeding practices and the risk of allergy development will follow in the next year from the National Institute of Allergy and Infectious Diseases-sponsored Working Group and the European Academy of Allergy and Clinical Immunology., (© 2015 the Authors. Published by Wiley Periodicals, Inc.)

Precautionary allergen labelling (PAL) was introduced by the food industry to help manage and communicate the possibility of reaction from the unintended presence of allergens in foods. However, in its current form, PAL is counterproductive for consumers with food allergies. This review aims to summarize the perspectives of all the key stakeholders (including clinicians, patients, food industry and regulators), with the aim of defining common health protection and risk minimization goals. The lack of agreed reference doses has resulted in inconsistent application of PAL by the food industry and in levels of contamination that prompt withdrawal action by enforcement officers. So there is a poor relationship between the presence or absence of PAL and actual reaction risk. This has led to a loss of trust in PAL, reducing the ability of consumers with food allergies to make informed choices. The result has been reduced avoidance, reduced quality of life and increased risk-taking by consumers who often ignore PAL. All contributing stakeholders agree that PAL must reflect actual risk. PAL should be transparent and consistent with rules underpinning decision-making process being communicated clearly to all stakeholders. The use of PAL should indicate the possible, unintended presence of an allergen in a consumed portion of a food product at or above any proposed action level. This will require combined work by all stakeholders to ensure everyone understands the approach and its limitations. Consumers with food allergy then need to be educated to undertake individualized risk assessments in relation to any PAL present., (© 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.)

Background: Traffic-related particulate matter (PM) has been linked to a heightened incidence of asthma and allergic diseases. However, the molecular mechanisms by which PM exposure promotes allergic diseases remain elusive., Objective: We sought to determine the expression, function, and regulation of pathways involved in promotion of allergic airway inflammation by PM., Methods: We used gene expression transcriptional profiling, in vitro culture assays, and in vivo murine models of allergic airway inflammation., Results: We identified components of the Notch pathway, most notably Jagged 1 (Jag1), as targets of PM induction in human monocytes and murine dendritic cells. PM, especially ultrafine particles, upregulated TH cytokine levels, IgE production, and allergic airway inflammation in mice in a Jag1- and Notch-dependent manner, especially in the context of the proasthmatic IL-4 receptor allele Il4raR576. PM-induced Jag1 expression was mediated by the aryl hydrocarbon receptor (AhR), which bound to and activated AhR response elements in the Jag1 promoter. Pharmacologic antagonism of AhR or its lineage-specific deletion in CD11c(+) cells abrogated the augmentation of airway inflammation by PM., Conclusion: PM activates an AhR-Jag1-Notch cascade to promote allergic airway inflammation in concert with proasthmatic alleles., (Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: Hazelnut allergy is birch pollen-driven in Northern/Western Europe and lipid transfer protein-driven in Spain and Italy. Little is known about other regions and other allergens., Objective: Establishing a molecular map of hazelnut allergy across Europe., Methods: In 12 European cities, subjects reporting reactions to hazelnut (n = 731) were evaluated and sensitization to 24 foods, 12 respiratory allergen sources, and latex was tested by using skin prick test and ImmunoCAP. A subset (124 of 731) underwent a double-blind placebo-controlled food challenge to hazelnut. Sera of 423 of 731 subjects were analyzed for IgE against 7 hazelnut allergens and cross-reactive carbohydrate determinants by ImmunoCAP., Results: Hazelnut allergy was confirmed in 70% of those undergoing double-blind placebo-controlled food challenges. Birch pollen-driven hazelnut sensitization (Cor a 1) dominated in most cities, except in Reykjavik, Sofia, Athens, and Madrid, where reporting of hazelnut allergy was less frequent anyhow. In Athens, IgE against Cor a 8 dominated and strongly correlated with IgE against walnut, peach, and apple and against Chenopodium, plane tree, and mugwort pollen. Sensitization to seed storage proteins was observed in less than 10%, mainly in children, and correlated with IgE to nuts, seeds, and legumes. IgE to Cor a 12, observed in all cities (10% to 25%), correlated with IgE to nuts, seeds, and pollen., Conclusions: In adulthood, the importance of hazelnut sensitization to storage proteins, oleosin (Cor a 12), and Cor a 8 is diluted by the increased role of birch pollen cross-reactivity with Cor a 1. Cor a 8 sensitization in the Mediterranean is probably driven by diet in combination with pollen exposure. Hazelnut oleosin sensitization is prevalent across Europe; however, the clinical relevance remains to be established., (Copyright © 2015 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

The purpose of this brief communication is to highlight emerging evidence to existing guidelines regarding potential benefits of supporting early, rather than delayed, peanut introduction during the period of complementary food introduction in infants. This document should be considered as interim guidance based on consensus among the following organizations: American Academy of Allergy, Asthma & Immunology, American Academy of Pediatrics, American College of Allergy, Asthma & Immunology, Australasian Society of Clinical Immunology and Allergy, Canadian Society of Allergy and Clinical Immunology, European Academy of Allergy and Clinical Immunology, Israel Association of Allergy and Clinical Immunology, Japanese Society for Allergology, Society for Pediatric Dermatology, and World Allergy Organization. More formal guidelines regarding early-life, complementary feeding practices and the risk of allergy development will follow in the next year from the National Institute of Allergy and Infectious Diseases-sponsored Working Group and the European Academy of Allergy and Clinical Immunology., (Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Precautionary labeling is used to warn consumers of the presence of unintended allergens, but the lack of agreed allergen thresholds can result in confusion and risk taking by patients with food allergy. The lack of data on threshold doses below which subjects are unlikely to react is preventing the development of evidence-based allergen management strategies that are understood by clinician and patient alike., Objective: We sought to define threshold dose distributions for 5 major allergenic foods in the European population., Methods: Patients with food allergy were drawn from the EuroPrevall birth cohort, community surveys, and outpatient clinic studies and invited to undergo a food challenge. Low-dose, double-blind, placebo-controlled food challenges were undertaken with commercially available food ingredients (peanut, hazelnut, celery, fish, and shrimp) blinded into common matrices. Dose distributions were modeled by using interval-censoring survival analysis with 3 parametric approaches., Results: Of the 5 foods used for challenge, 4 produced similar dose distributions, with estimated doses eliciting reactions in 10% of the allergic population (ED10), ranging from 1.6 to 10.1 mg of protein for hazelnut, peanut, and celery with overlapping 95% CIs. ED10 values for fish were somewhat higher (27.3 mg of protein), although the CIs were wide and overlapping between fish and plant foods. Shrimp provided radically different dose distributions, with an ED10 value of 2.5 g of protein., Conclusion: This evidence base will contribute to the development of reference doses and action levels for allergens in foods below which only the most sensitive subjects might react., (Copyright © 2014 American Academy of Allergy, Asthma & Immunology. Published by Elsevier Inc. All rights reserved.)

Background: The FAST (food allergy-specific immunotherapy) project aims at developing safe and effective subcutaneous immunotherapy for fish allergy, using recombinant hypoallergenic carp parvalbumin, Cyp c 1., Objectives: Preclinical characterization and good manufacturing practice (GMP) production of mutant Cyp (mCyp) c 1., Methods: Escherichia coli-produced mCyp c 1 was purified using standard chromatographic techniques. Physicochemical properties were investigated by gel electrophoresis, size exclusion chromatography, circular dichroism spectroscopy, reverse-phase high-performance liquid chromatography and mass spectrometry. Allergenicity was assessed by ImmunoCAP inhibition and basophil histamine release assay, immunogenicity by immunization of laboratory animals and stimulation of patients' peripheral blood mononuclear cells (PBMCs). Reference molecules were purified wild-type Cyp c 1 (natural and/or recombinant). GMP-compliant alum-adsorbed mCyp c 1 was tested for acute toxicity in mice and rabbits and for repeated-dose toxicity in mice. Accelerated and real-time protocols were used to evaluate stability of mCyp c 1 as drug substance and drug product., Results: Purified mCyp c 1 behaves as a folded and stable molecule. Using sera of 26 double-blind placebo-controlled food-challenge-proven fish-allergic patients, reduction in allergenic activity ranged from 10- to 5,000-fold (1,000-fold on average), but with retained immunogenicity (immunization in mice/rabbits) and potency to stimulate human PBMCs. Toxicity studies revealed no toxic effects and real-time stability studies on the Al(OH)3-adsorbed drug product demonstrated at least 20 months of stability., Conclusion: The GMP drug product developed for treatment of fish allergy has the characteristics targeted for in FAST: i.e. hypoallergenicity with retained immunogenicity. These results have warranted first-in-man immunotherapy studies to evaluate the safety of this innovative vaccine., (© 2015 S. Karger AG, Basel.)

Although English walnut is a commonly allergenic tree nut, walnut allergens have been poorly characterized to date. The objective of this work was to characterize the natural, low molecular weight (LMW) allergens from walnut. A protocol was developed to purify LMW allergens (specifically 2S albumins) from English walnuts. In addition to 2S albumins, a series of peptides from the N-terminal region of the 7S seed storage globulin proprotein were also identified and characterized. These peptides comprised a four-cysteine motif (C-X-X-X-C-X10-12-C-X-X-X-C) repeated throughout the 7S N-terminal region. Upon IgE immunoblotting, 3/11 and 5/11 sera from walnut-allergic subjects showed IgE reactivity to the 7S N-terminal fragments and 2S albumin, respectively. The mature 7S protein and the newly described 7S N-terminal peptides represent two distinct types of allergens. Because the proteolytic processing of 7S globulins has not been elucidated in many edible plant species, similar protein fragments may be present in other nuts and seeds.

Background: Kiwifruit is a common cause of food allergy. Symptoms range from mild to anaphylactic reactions., Objective: We sought to elucidate geographic differences across Europe regarding clinical patterns and sensitization to kiwifruit allergens. Factors associated with the severity of kiwifruit allergy were identified, and the diagnostic performance of specific kiwifruit allergens was investigated., Methods: This study was part of EuroPrevall, a multicenter European study investigating several aspects of food allergy. Three hundred eleven patients with kiwifruit allergy from 12 countries representing 4 climatic regions were included. Specific IgE to 6 allergens (Act d 1, Act d 2, Act d 5, Act d 8, Act d 9, and Act d 10) and kiwifruit extract were tested by using ImmunoCAP., Results: Patients from Iceland were mainly sensitized to Act d 1 (32%), those from western/central and eastern Europe were mainly sensitized to Act d 8 (pathogenesis-related class 10 protein, 58% and 44%, respectively), and those from southern Europe were mainly sensitized to Act d 9 (profilin, 31%) and Act d 10 (nonspecific lipid transfer protein, 22%). Sensitization to Act d 1 and living in Iceland were independently and significantly associated with severe kiwifruit allergy (odds ratio, 3.98 [P = .003] and 5.60 [P < .001], respectively). Using a panel of 6 kiwifruit allergens in ImmunoCAP increased the diagnostic sensitivity to 65% compared with 20% for skin prick tests and 46% ImmunoCAP using kiwi extract., Conclusion: Kiwifruit allergen sensitization patterns differ across Europe. The use of specific kiwifruit allergens improved the diagnostic performance compared with kiwifruit extract. Sensitization to Act d 1 and living in Iceland are strong risk factors for severe kiwifruit allergy., (Copyright © 2012 American Academy of Allergy, Asthma & Immunology. Published by Mosby, Inc. All rights reserved.)

Background: Lentils are increasingly consumed in many parts of the world.Two allergens, Len c 1 and 2, have been reported previously. Recently, peanut and green bean lipid transfer proteins (LTPs) have been identified as the first two members of an important group of allergens that might be associated with severe food allergies., Objective: To investigate lentil LTP as a potential new allergen., Methods: Efficacy of LTP extraction was monitored at different acidic pH values, using immunoblotting with cross-reactive anti-peach LTP antiserum. Natural LTP was purified from lentil extract and expressed as recombinant allergen in Escherichia coli. Sera from 10 lentil-allergic and/or -sensitized patients (Spain: 6, Italy: 1 and the Netherlands: 3) were used to further characterize lentil LTP., Results: Natural lentil LTP, purified from the homogenized germinated seeds and optimally extracted at pH 3, was identified and designated as allergen Len c 3. By CAP, 9/10 sera showed specific IgE to Len c 3. Recombinant (r) Len c 3 was successfully purified. The natural (n) Len c 3 CAP was completely inhibited by rLen c 3/rPru p 3. IgE binding to lentil pH 3 extract blot was completely inhibited by rLen c 3., Conclusion: The availability of immunochemically active nLen/rLen c 3 as a novel legume allergen facilitates further development and implementation of a third (next to peanut and green bean) legume LTP in component-resolved diagnosis strategies and contributes to evaluate the clinical importance of legume LTPs. Preferential extraction of Len c 3 (pH 3) will affect the production of sensitive extract-based diagnostic tests., (Copyright © 2011 S. Karger AG, Basel.)

Background:   Double-blind placebo-controlled food challenge (DBPCFC) is the gold standard for diagnosing food allergy. Standardized materials and protocols are essential for comparing DBPCFC results for multicentre studies such as EuroPrevall. This required the development and piloting of a standardized vehicle and low-dose protocol for confirming food allergy and determination of minimum eliciting doses (MEDs)., Methods:   A low-dose DBPCFC protocol was developed, with eight titrated protein doses from 3 μg to 1 g. This was delivered using a simple, microbiologically stable food base incorporating allergenic food ingredients manufactured at three sites and centrally distributed to clinical centres. Allergen blinding was assessed by a professional sensory testing panel using a triangle test. Homogeneity and allergen content were confirmed by ELISA and clinical efficacy was assessed in a pilot study, using celeriac and hazelnut as exemplars., Results:   Celeriac and hazelnut ingredients were sufficiently blinded in the dessert. The dessert meals were successfully piloted with hazelnut in allergy clinics in Spain, the Netherlands and Italy and with celeriac and hazelnut in Zurich. The challenges elicited a range of subjective and objective reactions ranging in severity from mild itching of the oral mucosa to bronchospasm., Conclusions:   A standardized challenge vehicle proven to sufficiently blind processed, powdered hazelnut and celeriac ingredients and that can be reproducibly manufactured has been developed. This pilot study shows that the vehicle is promising for the confirmation of food allergy and determination of MEDs in adults and children with body weight >28.8 kg (approximately 7-11 years old)., (© 2011 John Wiley & Sons A/S.)

Background: Fish allergens represent one of the most common causes of adverse reactions to food worldwide. Double-blind placebo-controlled food challenges (DBPCFC) are the gold standard for food allergy diagnosis. However, no standardised recipes are available for common food allergens such as fish, and a well trained dietitian is essential for creating and standardising them. The present study aimed to create and standardise recipes for use in DBPCFCs to fish., Methods: Three recipes were prepared. Employing a standardised procedure, a total of 35 panelists evaluated the different matrices using an evaluation form. A paired comparison test was used to estimate total evaluation's outcome. Fish allergic patients were challenged with different fish species blinded with the selected matrix and evaluated the recipe using the same form., Results: From a base recipe and step-by-step modifications, a low fat recipe was selected among other recipes tested, which proved to be appropriate for fish blinding, in terms of taste, odour, appearance and blinding. Patients challenged with the final matrix found it acceptable, no matter which fish type was used., Conclusions: In this pilot study, a recipe with satisfactory organoleptic characteristics was developed and validated for DBPCFC to fish.

Whiff (Lepidorhombus whiffiagonis) is a fish frequently consumed in Spain. Lep w 1, its major allergen, is a calcium-binding beta-parvalbumin. The resistance of Lep w 1 to heat denaturation and to digestion were studied by circular dichroism spectroscopy and by in vitro gastric digestion systems. Purified Lep w 1 was thermally stable up to 65 degrees C at neutral pH. Calcium depletion resulted in a change of its structure as determined by circular dichroism spectroscopy. A partial loss of structure was also observed at acidic pH; however, the allergen retained its full IgE-binding ability. The partially denatured Lep w 1 was easily digested by pepsin within 2 min. Further, the IgE reactivity of proteins extracted from cooked fish and their stability to proteolysis were analyzed. The extract revealed a higher number of IgE reactive bands than an extract from uncooked fish. IgE binding to these proteins could not be inhibited by an extract from uncooked fish. In contrast to a raw fish extract, the cooked extract showed higher resistance to pepsinolysis. The stability of Lep w 1 to thermal denaturation and digestion explains the high allergenicity of whiff.

Background: Parvalbumins are the most important fish allergens. Polysensitization to various fish species is frequently reported and linked to the cross-reactivity of their parvalbumins. Studies on cross-reactivity and its association to the allergenicity of purified natural parvalbumins from different fish species are still lacking. In addition, some studies indicate that dark muscled fish such as tuna are less allergenic., Methods: Total protein extracts and purified parvalbumins from cod, whiff, and swordfish, all eaten frequently in Spain, were tested for their IgE-binding properties with 16 fish allergic patients' sera from Madrid. The extent of cross-reactivity of these parvalbumins was investigated by IgE ELISA inhibition assays. Additionally, the cDNA sequences of whiff and swordfish parvalbumins were determined., Results: Extractable amounts of parvalbumins from cod were 20 times and from whiff 30 times higher than from swordfish. Parvalbumins were recognized by 94% of the patients in extracts of cod and whiff, but only by 60% in swordfish extracts. Nevertheless, a high cross-reactivity was determined for all purified parvalbumins by IgE inhibition. The amino acid sequence identities of the three parvalbumins were in a range of 62-74%., Conclusions: The parvalbumins of cod, whiff and swordfish are highly cross-reactive. The high amino acid sequence identity among cod, whiff and swordfish parvalbumins results in the observed IgE cross-reactivity. The low allergenicity of swordfish is due to the low expression levels of its parvalbumin.

Background: Grass pollen allergy is one of the most common allergies worldwide, and patients often show sensitization to an array of phylogenetically related species., Objective: To determine the effect of specific immunotherapy (SIT) with Phleum pratense extract on induction of the immune response to a mixture composed of 5 grass pollen extracts., Methods: Forty-six adult patients suffering from rhinitis and sensitized to a mix of grass pollen allergen extracts were randomized 3:1 to receive a short course of SIT with P. pratense or to an open control group without SIT. At baseline and after 3-4 months, we evaluated levels of specific immunoglobulin (Ig) E and IgG4, as well as the immediate and delayed cutaneous responses to the grass mix and P. pratense. IgG4 to Lolium perenne was also determined., Results: Levels of IgE and IgG4 to grass mix and P. pratense increased significantly during treatment (P<.001). However, this increase was only significantly higher in the SIT group than in the control group for IgG4 (P<.001). The levels of IgG4 to Phl p 5 and Lol p 5 were highly correlated (r=0.99, P<.001). The immediate and delayed cutaneous responses were significantly diminished to both extracts after SIT (P<.001)., Conclusions: Patients with rhinoconjunctivitis diagnosed using skin prick testing with a grass mix allergen extract and treated with a short course of SIT based on a single species P. pratense allergen extract are able to develop an immune response that targets not only the immunizing species, but also the grass mix allergen extract.

Background: Pru p 3, the nonspecific lipid transfer protein from peach, is an important plant food allergen that frequently induces systemic reactions., Objective: We sought to analyze the allergic T-cell response to Pru p 3., Methods: PBMCs from Italian and Spanish patients with peach allergy were stimulated with purified natural Pru p 3. Allergen-specific T-cell lines were used to identify T-cell epitopes of Pru p 3. Pru p 3-specific T-cell clones (TCCs) were analyzed for allergen-induced secretion of IL-4, IFN-gamma, and IL-10 and expression of the integrin beta7, a receptor critical for gut homing., Results: No difference in T-cell responses of Italian and Spanish patients was found. Among several T cell-activating regions, Pru p 3(13-27), Pru p 3(34-48), Pru p 3(43-57), and Pru p 3(61-75) were most frequently recognized in 18 Pru p 3-specific T-cell lines. The majority of 32 Pru p 3-specific TCCs belonged to the T(H)2 subset. In contrast to TCCs specific for other plant food and pollen allergens, only a limited number of Pru p 3-specific TCCs produced significant amounts of IL-10. The expression of integrin beta7 on Pru p 3-specific TCCs was comparable with that observed on peanut-specific TCCs and higher compared with that seen in different pollen-specific TCCs., Conclusion: The T-cell response to Pru p 3 is dominated by T(H)2 cells presumably primed in the gut. The identification of relevant T cell-activating regions provides a basis for engineering hypoallergenic variants of Pru p 3 with less IgE binding and retained T-cell stimulatory capacity for safe immunotherapy of peach allergy.

Background: Peach allergy is highly prevalent in the Mediterranean area; it is persistent and potentially severe, and therefore a prime target for immunotherapy. We aimed to study the efficacy and safety of sublingual immunotherapy (SLIT) with a peach extract quantified in mass units for Pru p 3, the peach lipid transfer protein., Methods: Randomized, double-blind, placebo-controlled (DBPC) clinical trial. The main efficacy outcome was the change in the response to a DBPC food challenge (DBPCFC) with peach. Secondary efficacy outcomes were the changes in skin prick test (SPT), and in specific immunoglobulin E (IgE) and IgG(4) to Pru p 3. Tolerance was assessed with a careful recording of adverse events., Results: After 6 months of SLIT, the active group tolerated a significantly higher amount of peach (three- to ninefold), presented a significant decrease (5.3 times) in SPT, and a significant increase in IgE and IgG(4) to Pru p 3. No significant changes were observed within the placebo group. Statistically significant inter-group differences were only observed in the SPT and IgG(4) responses. No serious adverse events were reported. Systemic reactions were mild, and observed with a similar frequency in both groups. Local reactions were significantly more frequent in the active group (three times) and 95% of them restricted to the oral cavity., Conclusion: In this first exploratory clinical trial, SLIT for peach allergy seems to be a promising therapeutic option that could modify the clinical reactivity of the patients to peach intake and the underlying immunological response with a good tolerance.

Allergic reactions to fruits and vegetables are frequently observed in older children and adolescents. They can result from a primary sensitization to food allergens or from a primary sensitization to inhalant allergens such as pollens or latex. In the case of fruit allergies, the stability of the allergens involved is crucial to the sensitization pathway and in the clinical presentation of the food allergy. Two patients allergic to fruits are presented and discussed in the light of the allergens involved. Patient 1 was a 14 yr-old girl with a grass and olive pollen allergy who developed oropharyngeal symptoms typical of the oral allergy syndrome (OAS) with multiple fruits from taxonomically unrelated families, and who was sensitized to profilin. Patient 2 was an 8 yr-old girl, with no pollen allergies, who developed systemic reactions to peach and apple, and who was sensitized to non-specific lipid transfer proteins (LTP). Profilins are labile allergens present in pollens and foods, and sensitization occurs through the respiratory route to pollen profilin. The cross-reactive IgE antibodies generated can elicit local reactions in the oropharyngeal mucosa (OAS) when exposed to fruit profilins. In contrast, LTPs are a family of stable allergens that resist thermal treatment and enzymatic digestion, and can thus behave as true food allergens inducing primary (non-pollen related) sensitizations and triggering systemic reactions. These two cases represent two distinct patterns of sensitization and clinical expression of fruit allergies that are determined by the panallergens involved (LTPs and profilins) and their intrinsic physicochemical properties. Additionally, these two cases also show the improved diagnostic value of Component Resolved Diagnosis, and strengthen its utility in the routine diagnosis and management of patients.

Allergic reaction following fish consumption can trigger life-threatening reactions in predisposed individuals. Parvalbumins from different species have been identified as the major fish allergens. There are two distinct phylogenetic lineages of parvalbumins, alpha and beta. Most allergic reactions are caused by beta-parvalbumins. We cloned and expressed cDNAs encoding cod (Gadus morhua) and carp (Cyprinus carpio) beta-parvalbumins and purified natural cod beta-parvalbumin. CD spectra of the purified proteins showed that their overall secondary structure contents were very similar. No differences in thermal stability were monitored in the calcium-bound or calcium-depleted form of natural cod parvalbumin. IgE reactivity was assessed using 26 sera of fish allergic patients from Spain, The Netherlands, and Greece in immunoblot and ELISA experiments. Twenty-five of the 26 patients with IgE reactivity to native and recombinant cod parvalbumin also reacted to the recombinant carp parvalbumin. IgE inhibition assays were performed using cod and carp extracts and purified recombinant parvalbumin of cod and carp. High crossreactivity among cod and carp parvalbumins was observed in immunoblots as well as in fluid phase assays. Natural and recombinant parvalbumins gave comparable results when performing various in vitro diagnostic assays.

A lipid transfer protein (LTP, Cor a 8) together with the 11S (Cor a 9) and 7S seed storage globulins (Cor a 11) are major food allergens present in hazelnut. Methods are described for their purification and characterisation using in-gel tryptic digestion mass spectrometry to confirm their identities and circular dichroism and Fourier-transform infrared spectroscopies to demonstrate that they are authentically folded. Preliminary immunochemical studies have also confirmed that the purified preparations retain their immunological properties in terms of immunoglobulin E binding, determined by immunoblotting using serum from hazelnut allergic patients. These preparations form a basis for development of improved methods of diagnosis of food allergy based on the concept of component-resolved diagnosis.

Egg proteins represent one of the most important sources evoking food allergic reactions. In order to improve allergy diagnosis, purified and well-characterized proteins are needed. Although the egg white allergens Gal d 1, 2, 3 and 4 (ovomucoid, ovalbumin, ovotransferrin, and lysozyme) are commercially available, these preparations contain impurities, which affect exact in vitro diagnosis. The aim of the present study was to set up further purification protocols and to extend the characterization of the physicochemical and immunological properties of the final batches. The egg white allergens Gal d 1-4 were purified from commercial preparations, whereas Gal d 5 (alpha-livetin) was purified from egg yolk. The final batches of Gal d 1-5 consisted of a range of isoforms with defined tertiary structure. In addition, the IgE binding capacity of the purified egg allergens was tested using allergic patients' sera. The allergen batches will be further used to set up allergen specific diagnostic assays and to screen a larger collection of patients' sera.

Safety assessment for genetically-engineered crop plants includes assessment for allergic responses. To facilitate this assessment, serum banks should contain well-characterised sera from patients with confirmed food allergies. A serum is defined as well-characterised if it is taken from a patient who has a convincing history of allergic responses to a known allergen or an allergen-containing food, a positive skin prick test (or elevated IgE response), and a positive response in a clinical food challenge.

EuroPrevall is an EU-funded multidisciplinary project including 62 institutions from 22 countries. EuroPrevall studies the prevalence and distribution of food allergies in infants, children, adolescents, and adults in Europe, threshold doses for allergenic foods, the role of the environment in food allergy, the socioeconomic impact of food allergy, and novel diagnostic tools for food allergies. The EuroPrevall serum bank (EPASB), containing samples from approximately 70,000 subjects, is a major tool to achieve these goals. EPASB is coordinated by the Paul-Ehrlich-Institut, Langen, Germany. Local sera collections are administered at the University of Amsterdam (NL), the University Hospital of Manchester (UK), Charité Hospital (DE) and the Paul-Ehrlich-Institut. The EPASB coordinator and managing partners distribute samples for experimental work and regulate access. The overall aim is to provide sera to fulfil EuroPrevall research goals. The EPASB coordinator and managing partners suggest appropriate sera for addressing specific scientific and diagnostic questions. The serum bank will be maintained after termination of the project, but subsequent investigations must be in accordance with the original research goals of EuroPrevall. Thus, the contributors of the sera retain control over their future use. This rule prevents investigation of questions outside the scope of EuroPrevall, e.g. the allergenicity of genetically-modified foods.

Allergen extracts have been used for diagnosis and treatment of allergy for around 100 years. During the second half of 20th century, the notion increasingly gained foothold that accurate standardization of such extracts is of great importance for improvement of their quality. As a consequence, manufacturers have implemented extensive protocols for standardization and quality control. These protocols have overall IgE-binding potencies as their focus. Unfortunately, each company is using their own in-house reference materials and their own unique units to express potencies. This does not facilitate comparison of different products. During the last decades, most major allergens of relevant allergen sources have been identified and it has been established that effective immunotherapy requires certain minimum quantities of these allergens to be present in the administered maintenance dose. Therefore, the idea developed to introduce major allergens measurements into standardization protocols. Such protocols based on mass units of major allergen, quantify the active ingredients of the treatment and will at the same time allow comparison of competitor products. In 2001, an EU funded project, the CREATE project, was started to support introduction of major allergen based standardization. The aim of the project was to evaluate the use of recombinant allergens as reference materials and of ELISA assays for major allergen measurements. This paper gives an overview of the achievements of the CREATE project.

Background: Coconut (Cocos nucifera) is a monocotyledonous plant of the Arecaceae family. Allergy to coconut is infrequent, with only 5 cases reported so far in the medical literature., Objective: To identify coconut allergens in 2 patients allergic to this food., Methods: We describe 2 patients allergic to coconut: an adult pollen-allergic patient monosensitized to coconut who presented with severe oropharyngeal symptoms and a child with a previous allergy to walnut, not allergic to pollen, who developed anaphylaxis on coconut ingestion. Both patients had positive skin prick test results and serum specific IgE (CAP) to coconut. IgE sodium dodecyl sulfate-polyacrylamide gel electrophoresis immunoblotting was performed to identify the allergens involved, and a strong IgE binding band detected in both patients was further analyzed by matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF MS). Stability to pepsin digestion of the coconut extract and its cross-reactivity with tree nuts were studied., Results: An immunoblot showed an almost identical profile of IgE binding proteins in the coconut extract in both patients who reacted strongly to a band of approximately 29 kDa. The peptide analysis by MALDI-TOF MS of this band obtained the sequence GHGKREDPEKR. The protein with the highest correlation with this peptide was found to be a 7S globulin from Elaeis guineensis, another oil palm species also belonging to the Arecaceae family. The 29-kDa band was digested by pepsin in less than 1 minute. Cross-reactivity among coconut, walnut, and hazelnut was demonstrated by CAP inhibition in patient 2., Conclusion: We have identified a 7S storage protein as a novel coconut allergen.

Background: Allergy to a plant food can either result from direct sensitization to that food or from primary sensitization to pollen, latex, or another food., Objective: We sought to investigate the primary sensitizers in apple allergy across Europe, the individual allergens involved, and whether these differences determine the clinical presentation., Methods: Patients (n = 389) with positive case histories and skin prick test responses to fresh apple were selected in the Netherlands, Austria, Italy, and Spain. Skin prick tests and RASTs to a panel of pollens and plant foods were performed, as well as RASTs to Bet v 1 and the apple allergens Mal d 1, 2, 3, and 4., Results: In the Netherlands, Austria, and Italy apple allergy is mild (>90% isolated oral symptoms) and related to birch pollinosis and sensitization to Bet v 1 and its apple homologue, Mal d 1, which has an odds ratio of local reactions of 2.85 (95% CI, 1.47-5.55). In Spain apple allergy is severe (>35% systemic reactions) and related to peach allergy and sensitization to Mal d 3 (nonspecific lipid transfer protein), which has an odds ratio of systemic reactions of 7.76 (95% CI, 3.87-15.56)., Conclusion: The analysis of individual apple allergens in a clinical context has provided insight into the sensitization pathway and into the intrinsic risk an allergen bears to induce mild or severe food allergy., Clinical Implications: Information on the sensitization pathway is essential to develop preventive strategies in food allergy. The application of individual food allergens with a known intrinsic risk will improve the prognostic value of diagnostic tests.

Immune reactivity is a key issue in the evaluation of the quality of recombinant allergens as potential reference materials. Within the frame of the CREATE project, the immune reactivity of the natural and recombinant versions of the major allergens of birch pollen (Bet v 1), grass pollen (Phl p 1 and 5), olive pollen (Ole e 1), and house dust mite (Der p 1 and 2, and Der f 1 and 2) was analysed. The IgE binding capacity of the allergens was studied by direct RAST and RAST inhibition, and their biological activity by basophil histamine release, using sera of allergic patients selected across Europe. For birch pollen, rBet v 1 is an excellent mimic of the natural allergen. For grass pollen, rPhl p 1 showed a significant lower IgE reactivity and was not considered a suitable candidate, whereas rPhl p 5a exhibited an immune reactivity closer to that of its natural counterpart. For olive, rOle e 1 had a lower IgE binding capacity in RAST but a higher biological activity in histamine release. For house dust mite, recombinant group 1 allergens were significantly less potent than their natural counterparts, but recombinant group 2 allergens were close mimics of their natural homologues.

Background: Pepsin resistance of allergens like lipid transfer protein and 2S albumin has been suggested as explanation for the severity of symptoms often induced by these allergens. Component-resolved diagnosis with purified labile and stable allergens has therefore been proposed to better characterize the risk involved in a positive in vitro IgE test. However, for many foods, purified allergens are not (yet) available., Objective: It was the aim of this study to evaluate the potential of pepsin-digested whole-food extracts to distinguish between IgE responses to stable (potentially severe) and labile (mild) allergens., Methods: Sera (n = 143) from Italian, Spanish and Dutch patients with hazelnut and/or apple ingestion-related symptoms were analyzed for residual IgE binding to pepsin-resistant hazelnut and/or apple allergens. Control and pepsin-digested hazelnut and apple extracts were used for radioallergosorbent test analysis and immunoblot analysis., Results: Pepsin digestion of food extracts, like from hazelnut and apple used for in vitro diagnostic tests, provides a way to distinguish sensitization to pepsin-resistant allergens from that to pepsin-susceptible allergens. In this selected group of patients, IgE reactivity to pepsin-digested extracts correlated with sensitization to the stable allergen lipid transfer protein. The analysis further revealed that the use of soluble pepsin can result in false-positive in vitro tests (2/143)., Conclusion: Pepsin-digested food extracts are a convenient tool to identify patients with IgE antibodies against potentially dangerous stable allergens, in particular for those foods where the relevant stable allergens have not yet been identified. This can increase the clinical prognostic value of food allergy serology., (Copyright (c) 2005 S. Karger AG, Basel.)

Background: Apple cultivars have been reported to differ in allergenicity on the basis of in vitro and skin prick tests with apple extracts., Objectives: We sought to evaluate the efficacy of the prick-to-prick method in assessing differences in allergenicity of apple cultivars and to confirm differences by means of double-blind, placebo-controlled food challenge (DBPCFC)., Methods: Intra-assay and intracultivar variation of prick-to-prick test results were determined in 6 Dutch and 8 Spanish patients with apple allergy by using 5 apples of the cultivars Golden Delicious, Fuji, and Ecolette in duplicate. In addition, 21 cultivars were screened for allergenicity in 15 Dutch patients with birch pollen and apple allergy. Two selected cultivars (Golden Delicious and Santana) were tested with DBPCFCs. The influence of storage conditions on allergenicity was assessed in 5 cultivars., Results: Intra-assay variation of skin prick testing was 3.9%, and intracultivar variation was 4.1%. A ranking of 21 cultivars was made on the basis of prick-to-prick tests in 9 patients. Apple cultivars were classified as of low, intermediate, and high allergenicity, with a significant difference between low and high allergenicity (P < .001). A significant difference in allergenicity determined between Golden Delicious and Santana cultivars (P < .05) was confirmed by means of DBPCFC. With 5 cultivars, controlled atmosphere (2.5% oxygen/1% carbon dioxide) was shown to reduce allergenicity (P < .001) by 15% compared with storage at 2 degrees C., Conclusions: Prick-to-prick testing with fresh apples is a reproducible method of assessing allergenicity. Apples can be classified as of low or high allergenicity for the majority of patients. This was confirmed by using DBPCFCs. Selection of cultivars and control of storage conditions are both viable strategies for reduction of symptoms in patients with apple allergy.

Background: Non-specific lipid transfer proteins (LTPs) are involved in allergy to fresh and processed fruits. We have investigated the effect of thermal treatment and glycation on the physico-chemical and IgE-binding properties of the LTP from apple (Mal d 3)., Methods: Mal d 3 was purified from apple peel and the effect of heating in the absence and presence of glucose investigated by CD spectroscopy, electrospray and MALDI-TOF mass spectrometry. IgE reactivity was determined by RAST and immunoblot inhibition, SPT and basophil histamine release test., Results: The identity and IgE reactivity of purified Mal d 3 was confirmed. Mild heat treatment (90 degrees C, 20 min) in the absence or presence of glucose did not alter its IgE reactivity. More severe heat treatment (100 degrees C, 2 h) induced minor changes in protein structure, but a significant decrease in IgE-binding (30-fold) and biological activity (100- to 1000-fold). Addition of glucose resulted in up to four glucose residues attached to Mal d 3 and only a 2- and 10-fold decrease of IgE-binding and biological activity, respectively., Conclusions: Only severe heat treatment caused a significant decrease in the allergenicity of Mal d 3 but glycation had a protective effect. The presence of sugars in fruits may contribute to the thermostability of the allergenic activity of LTP in heat-processed foods.