Structure and IgE Cross-Reactivity among Cashew, Pistachio, Walnut, and Peanut Vicilin-Buried Peptides.

Peanut and tree-nut allergies are frequently comorbid for reasons not completely understood. Vicilin-buried peptides (VBPs) are an emerging family of food allergens whose conserved structural fold could mediate peanut/tree-nut co-allergy. Peptide microarrays were used to identify immunoglobulin E (IgE) epitopes from the N-terminus of the vicilin allergens Ara h 1, Ana o 1, Jug r 2, and Pis v 3 using serum from three patient diagnosis groups: monoallergic to either peanuts or cashew/pistachio, or dual allergic. IgE binding peptides were highly prevalent in the VBP domains AH1.1, AO1.1, JR2.1, and PV3.1, but not in AO1.2, JR2.2, JR2.3, and PV3.2 nor the unstructured regions. The IgE profiles did not correlate with diagnosis group. The structure of the VBPs from cashew and pistachio was solved using solution-NMR. Comparisons of structural features suggest that the VBP scaffold from peanuts and tree-nuts can support cross-reactivity. This may help understand comorbidity and cross-reactivity despite a distant evolutionary origin.

Oleosin Cor a 15 is a novel allergen for Italian hazelnut allergic children.

BACKGROUND: Hazelnut allergy, which is characterized by symptoms that range from mild to severe, is one of the most common allergies in children throughout Europe, and an accurate diagnosis of this allergy is therefore essential. However, lipophilic allergens, such as oleosins, are generally underrepresented in diagnostic tests. We therefore sought to characterize the IgE reactivity of raw and roasted hazelnut oleosins, using the sera of hazelnut-allergic pediatric patients. METHODS: Raw and roasted hazelnut oil body-associated proteins were analyzed by means of 1D and 2D electrophoresis and MS. Oleosin IgE reactivity was assessed by immunoblotting with the sera of 27 children who have confirmed hazelnut allergies and from 10 tolerant subjects. A molecular characterization of the oleosins was performed by interrogating the C. avellana cv. Jefferson and cv. TGL genomes, and through expression and purification of the recombinant new allergen. RESULTS: A proteomic and genomic investigation allowed two new oleosins to be identified, in addition to Cor a 12 and Cor a 13, in hazelnut oil bodies. One of the new oleosins was registered as a new allergen, according to the WHO/IUIS Allergen Nomenclature Subcommittee criteria, and termed Cor a 15. Cor a 15 was the most frequently immunorecognized oleosin in our cohort. Oleosins resulted to be the only immunorecognized allergens in a subgroup of allergic patients who showed low ImmunoCAP assay IgE values and positive OFC and PbP. Hazelnut roasting resulted in an increase in oleosin immunoreactivity. CONCLUSION: A novel hazelnut oleosin, named Cor a 15, has been discovered. Cor a 15 could play a role in eliciting an allergic reaction in a subgroup of pediatric patients that exclusively immunorecognize oleosins. The high prevalence of hazelnut oleosin sensitization here reported further confirms the need to include oleosins in routine diagnostic procedures.

MRM3-based LC-MS multi-method for the detection and quantification of nut allergens.

Food allergies have become a global challenge to food safety in industrialized countries in recent years. With governmental monitoring and legislation moving towards the establishment of threshold allergen doses, there is a need for sensitive and quantitative analytical methods for the determination of allergenic food contaminants. Targeted proteomics employing liquid chromatography-mass spectrometry (LC-MS) has emerged as a promising technique that offers increased specificity and reproducibility compared to antibody and DNA-based technologies. As the detection of trace levels of allergenic food contaminants also demands excellent sensitivity, we aimed to significantly increase the analytical performance of LC-MS by utilizing multiple reaction monitoring cubed (MRM3) technology. Following a bottom-up proteomics approach, including a straightforward sample preparation process, 38 MRM3 experiments specific to 18 proteotypic peptides were developed and optimized. This permitted the highly specific identification of peanut, almond, cashew, hazelnut, pistachio, and walnut. The analytical performance of the method was assessed for three relevant food matrices with different chemical compositions. Limits of detection were around 1 µg/g or below in fortified matrix samples, not accounting for the effects of food processing. Compared to an MRM-based approach, the MRM3-based method showed an increase in sensitivity of up to 30-fold. Regression analysis demonstrated high linearity of the MRM3 signal in spiked matrix samples together with robust intersample reproducibility, confirming that the method is highly applicable for quantitative purposes. To the best of our knowledge, we describe here the most sensitive LC-MS multi-method for food allergen detection thus far. In addition, this is the first study that systematically compares MRM3 with MRM for the analysis of complex foods. Graphical abstract Allergen detection by MRM3.

Consumo de nozes e amendoim durante a gestação pode diminuir risco de doençasalérgicas em crianças / Consumption of nuts and peanuts during pregnancy may reduce risk of doençasalérgicas in children

Esse estudo não encontrou associação significativa entre oconsumo de amendoim e nozes na gravidez e aumento no riscode asma nas crianças. Na verdade, a ingestão materna de amendoime de nozes, pelo menos uma vez por semana durante agravidez, foi associada a risco significativamente reduzido deasma em seus filhos.

CD63 expression on basophils as a tool for the diagnosis of pollen-associated food allergy: sensitivity and specificity.

BACKGROUND: Basophil activation is associated with the expression of CD63. Because allergens can induce basophil activation by cross-linking specific IgE, increased CD63 expression has been proposed as a novel in vitro test for immediate type allergy. OBJECTIVE: We compared the CD63-based basophil activation test (BAT) in the diagnosis of allergy to carrot, celery and hazelnut with skin prick tests (SPT) and measurement of allergen-specific IgE. METHODS: Twenty-nine patients with a history of an oral allergy syndrome induced by carrot, celery or hazelnut (n = 20 for each allergen) and 20 controls were studied. SPT were performed with standardized and native carrot, celery and hazelnut extracts. Allergen-specific IgE was determined by the CAP FEIA method and basophil activation was determined by flow cytometry upon double staining with anti-IgE/anti-CD63 mAb. RESULTS: SPT with native carrot, celery and hazelnut showed sensitivities of 100%, 100% and 90%, and specificities of 80%, 80% and 90%. SPT with commercial extracts of the same allergens gave sensitivities of 85%, 80% and 85%, and specificities of 80%, 80% and 90%. Sensitivity of allergen-specific IgE and the BAT for carrot, celery and hazelnut was 80% vs. 85%, 70% vs. 85%, and 80% vs. 90%, with corresponding specificities of 80% vs. 85%, 80% vs. 80%, and 95% vs. 90%. The cut-off for a positive BAT was 10% CD63+ basophils. Moreover, there was a positive correlation between IgE reactivity and the number of CD63+ basophils for all food allergens (carrot: r = 0.69, celery: r = 0.67, hazelnut: r = 0.66). CONCLUSIONS: Quantification of basophil activation by CD63 expression is a valuable new in vitro method for diagnosis of immediate type food sensitization. Although double-blind placebo-controlled food challenges remain the gold standard, the CD63-based BAT may supplement routine diagnostic tests such as SPT or allergen-specific IgE in the future.