In wheat-dependent exercise-induced anaphylaxis (WDEIA), cofactors such as exercise, acetylsalicylic acid (ASA), alcohol or unfavorable climatic conditions are required to elicit a reaction to wheat products. The mechanism of action of these cofactors is unknown, but an increase of gliadin absorption has been speculated. Our objectives were to study gliadin absorption with and without cofactors and to correlate plasma gliadin levels with factors influencing protein absorption in healthy volunteers. Twelve healthy probands (six males, six females; aged 20–56 years) ingested 32 g of gluten without any cofactor or in combination with cofactors aerobic and anaerobic exercise, ASA, alcohol and pantoprazole. Gliadin serum levels were measured up to 120 min afterwards and the intestinal barrier function protein zonulin in stool was collected before and after the procedure; both were measured by ELISA. Stool microbiota profile was obtained by 16S gene sequencing. Within 15 min after gluten intake, gliadin concentrations in blood serum increased from baseline in all subjects reaching highly variable peak levels after 15–90 min. Addition of cofactors did not lead to substantially higher gliadin levels, although variability of levels was higher with differences between individuals (p < 0.001) and increased levels at later time points. Zonulin levels in stool were associated neither with addition of cofactors nor with peak gliadin concentrations. There were no differences in gut microbiota between the different interventions, although the composition of microbiota (p < 0.001) and the redundancy discriminant analysis (p < 0.007) differed in probands with low versus high stool zonulin levels. The adsorption of gliadin in the gut in healthy volunteers is less dependent on cofactors than has been hypothesized. Patients with WDEIA may have a predisposition needed for the additional effect of cofactors, e.g., hyperresponsive or damaged intestinal epithelium. Alternatively, other mechanisms, such as cofactor-induced blood flow redistribution, increased activity of tissue transglutaminase, or increases in plasma osmolality and acidosis inducing basophil and mast cell histamine release may play the major role in WDEIA.

Idiopathic anaphylaxis (IA) is an unresolved concern. Hidden allergens may be relevant in IA and in non-steroidal anti-inflammatory drug hypersensitivity (NSAID-HS). To identify hidden elicitors for IA and NSAID-HS by a skin prick test (SPT; 13 allergens) and allergen-specific IgE (sIgE; 12 allergens) panel and to determine the value of each tested allergen. We retrospectively included all patients from 2018 to 2021 referred with a suspicion of IA or NSAID-HS by history in whom SPT and/or sIgE to allergens of the IA panel were performed. Patient characteristics from patients' records included comorbidities, history and symptoms of anaphylaxis, serum baseline tryptase level (sBT), total IgE level, SPT, sIgE, challenge results and final diagnoses. 134 patients (77 female; mean age 39.7±14.6 years) were included. Median sBT and total IgE levels were 4.23 μg/l and 133.5 kU/L, respectively. Allergological workup with the IA panel resulted in positive SPT and sIgE in 61 (47%) and 66 (60%) patients, respectively. In those, confirmation or exclusion of allergy, mostly by challenge, led to a definitive diagnosis in 61/134 patients (46%). SPT was most frequently positive to gluten (22.4%) and sIgE to ω5-gliadin (21.6%), which correlated with the history (r=.310 p<0.001 and r=.407 p<0.001, respectively). In 28/134 (21%) of patients with initially suspected IA or NSAID-HS, challenges confirmed occult food allergy with wheat allergy dependent on augmentation factors (WANDA) being the most frequent cause of anaphylaxis (19%). WANDA should be considered in all patients with anaphylaxis of unknown cause or after NSAID intake.

Wheat allergy dependent on augmentation factors (WALDA) is the most common gluten allergy in adults. IgE-mediated sensitizations are directed towards ω5-gliadin but also to other wheat allergens. The value of the different in vitro cellular tests, namely the basophil activation test (BAT) and the active (aBHRA) and passive basophil histamine-release assays (pBHRA), in the detection of sensitization profiles beyond ω5-gliadin has not been compared. Therefore, 13 patients with challenge-confirmed, ω5-gliadin-positive WALDA and 11 healthy controls were enrolled. Specific IgE (sIgE), skin prick tests, BATs, aBHRA, and pBHRA were performed with allergen test solutions derived from wheat and other cereals, and results were analyzed and compared. This study reveals a distinct and highly individual reactivity of ω5-gliadin-positive WALDA patients to a range of wheat allergens beyond ω5-gliadin in cellular in vitro tests and SPT. In the BAT, for all tested allergens (gluten, high-molecular-weight glutenin subunits, α-amylase/trypsin inhibitors (ATIs), alcohol-free wheat beer, hydrolyzed wheat proteins (HWPs), rye gluten and secalins), basophil activation in patients was significantly higher than in controls (p = 0.004–p < 0.001). Similarly, significant histamine release was detected in the aBHRA for all test substances, exceeding the cut-off of 10 ng/mL in all tested allergens in 50% of patients. The dependency of tests on sIgE levels against ω5-gliadin differed; in the pBHRA, histamine release to any test substances could only be detected in patients with sIgE against ω5-gliadin ≥ 7.7 kU/L, whereas aBHRA also showed high reactivity in less sensitized patients. In most patients, reactivity to HWPs, ATIs, and rye allergens was observed. Additionally, alcohol-free wheat beer was first described as a promising test substance in ω5-gliadin-positive WALDA. Thus, BAT and aBHRA are valuable tools for the identification of sensitization profiles in WALDA.

Background: Wheat allergy dependent on augmentation factors (WALDA) represents the most common form of IgE‐mediated wheat allergy in adults. Although ω5‐gliadin is the major allergen in WALDA, sIgE reactivity to non‐gluten wheat proteins remains poorly characterized. Methods: Serum samples from 54 adults with challenge‐confirmed WALDA and 21 tolerant controls (including 5 with positive sIgE to ω5‐gliadin) from two German centres were analysed for sIgE reactivity to 19 recombinant wheat proteins and the cross‐reactive carbohydrate determinant (CCD) MUXF3. Results: sIgE to α−/β‐gliadin (Tri a 21.0101) was highly specific for WALDA, present in 29/54 patients (53.7%) but absent in controls (100% specificity, p < 0.001). Receiver operating characteristic analysis revealed excellent discriminatory ability (area under the curve = 0.85) with an optimal cut‐off of 0.22 kU/l. Reactivity to non‐gluten wheat proteins was limited, with 24.1% of patients showing sIgE to thioredoxin, 11.3% to dehydrin, 9.3% to any α‐amylase/trypsin inhibitor subunit, and minimal sIgE to non‐specific lipid transfer proteins (Tri a 14), thiol reductase, triosephosphate isomerase, peroxiredoxin, serpin, glyceraldehyde‐3‐phosphate dehydrogenase, serine protease inhibitor‐like allergen, and peroxidase. Non‐gluten reactivity correlated strongly with MUXF3 CCD (ρ = 0.776), total IgE (ρ = 0.646), and wheat extract sIgE (ρ = 0.534; all p < 0.001). Grass pollen‐sensitized individuals showed increased non‐gluten protein reactivity ( p < 0.001). Controls demonstrated higher MUXF3 CCD reactivity (38.1% vs. 11.6%, p < 0.05). Conclusion: The data suggest that WALDA is characterized by highly specific reactivity to gliadin proteins, with limited sensitization to non‐gluten wheat components. sIgE to α−/β‐gliadin represents a promising diagnostic biomarker with superior specificity compared with ω5‐gliadin alone.

Sensitisation leading to food allergy may occur through the skin. Quantitative data on epidermal allergen penetration as a prerequisite for this remain limited. This study quantifies epidermal penetration of gluten and hydrolysed wheat protein (HWP) in 18 patients with challenge-confirmed WALDA (wheat allergy dependent on augmentation factors) and 12 healthy controls (HC). After 1 h of epicutaneous wheat application, 20 consecutive tape strips (TS) were collected, and wheat protein concentration was quantified by gliadin-specific ELISA. Skin barrier status was assessed by electrical impedance spectroscopy (EIS). Serum gliadin levels were measured before and 90 min after wheat application. Gliadin was detected across all TS layers for gluten and HWP. Penetration levels did not differ between patients and controls. Skin barrier status assessed by EIS did not differ significantly between individuals with and without a history of atopic dermatitis (p = 0.27). No correlation was observed between EIS-assessed skin barrier status and gliadin penetration. Serum gliadin was not increased after epicutaneous wheat application. This study using TS demonstrates for the first time that wheat allergens penetrated into the epidermis but could not be detected in the serum. Neither wheat allergy nor skin barrier status was associated with increased stratum corneum penetration. These findings suggest that epidermal uptake alone may not be sufficient to explain sensitisation.