ABSTRACT 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.

This prospective cohort study aimed to monitor the humoral immune response to SARS-CoV-2 proteins over 3 years in relation to repeated vaccination or infection. Quantitative immunoassays traceable to international IgG units were used to measure and compare IgG levels against Wuhan type and Omicron spike-S1 and nucleocapsid proteins over 3 years. In addition, the Euroimmun-N-test was used to determine positive IgG values against the nucleocapsid. A total of 1223 serum samples from 126 participants without evidence of COVID-19 at enrollment were analyzed and used to calculate antibody half-lives following vaccination or infection, as reported by the questionnaire. Antibody levels and half-life against both variants of the spike S1 protein increased significantly with each additional vaccination or after infection. IgG against the Wuhan type was, on average, three times higher than against the Omicron variant. The half-lives of antibodies against the Wuhan type after the second and third vaccinations and after infection were significantly longer than those of IgG against the Omicron variant. Two-thirds of the cohort reported COVID-19 infection, detected with a sensitivity of 70% by the quantitative nucleocapsid IgG assay and 83% by the Euroimmun-N-test. The level of anti-nucleocapsid-IgG after infection in the vaccinated cohort was significantly lower than anti-S1-IgG and also lower than in unvaccinated infected persons. Despite repeated vaccinations and progressively increasing IgG antibody levels targeting the spike protein, most of the cohort reported breakthrough infections, possibly due to the lower concentration and reduced half-life of antibodies against the Omicron variant.

This occupational case report describes a 28-year-old woman, employed since November 2020 as a research assistant working with mealworms (Tenebrio molitor), who developed symptoms of mild shortness of breath and severe redness and swelling of the eyes in early 2022, leading to the diagnosis of occupational allergic asthma due to respiratory sensitization to mealworm. Subsequent tests confirmed mono-sensitization to T. molitor and additional cross-sensitization to flour beetle (Tribolium confusum). Further studies aimed to determine the antigenic and allergenic potency of different protein fractions and rearing material isolated from T. molitor regarding protein pattern and immunological activity by T. molitor-specific polyclonal rabbit IgG and human IgE. The highest antigen content of T. molitor was measured in the rearing material, followed by mealworm flour and aqueous extraction. Allergenic proteins are particularly detected in mealworm flour and aqueous fraction at molecular weights of 24 and 11 kDa, which appear to be specific for primary airway sensitization. In addition, the airborne T. molitor antigen levels in the workplace were monitored using electrostatic dust collectors to identify hotspots of exposure. The disposal of dry insect material was shown to be responsible for the unintentional release of potential allergens. Therefore, awareness of the potential risk of a type I allergy due to the release of airborne insect allergens in the workplace should be raised.

Mite sensitization is notable in several occupational settings. Elevated house dust mite concentrations are primarily detected in workplaces where people congregate and are active. Allergy to storage mites and spider mites has commonly been reported in agricultural and various food processing occupations. Rapid expansion of biological pest control has resulted in increased exposure to predatory mites causing sensitization of greenhouse workers. Globally, mite populations in workplaces are likely to change due to climate change. Occupational relevant mites produce a variety of allergens and adjuvants that trigger both innate and adaptive immune responses. Cross-reactivity between allergens occurs due to shared IgE-binding epitopes to different allergens. Occupational allergy to mites typically causes rhinitis and asthma. Challenges of distinguishing the role of occupational exposure to allergens, also present in non-occupational environments, complicate the diagnosis of occupational mite allergy and asthma. Nevertheless, preventive measures to reduce exposure to mite allergens in workplaces are essential in mitigating occupational hazards. Further research is needed to better understand the incidence of occupational mite allergy and asthma. It is essential to identify the risk factors in different occupational settings, assess the impact of climate change on exposure, and determine the relevant allergens and their potential cross-reactivity.

New protein-rich flour blends from legumes are increasingly being processed for gluten-free bakery products, which may increase the risk of IgE-mediated sensitization through inhalation exposure. In the described case of a 29-year-old baker, work incapacity occurred due to respiratory complaints following exposure to various gluten-free baking ingredients. Occupation-related sensitization to "chestnut pea flour" (flour from lightly roasted and ground yellow peas) was identified, while the patient showed no IgE sensitization to five other ingredients. Significant allergic respiratory reactions occurred during bronchial challenge test with "chestnut pea" extract. Based on the test results and typical workplace-related symptoms, recognition of occupational disease was recommended. Further identification of potential causal allergens using IgE blot and allergen component-resolved diagnostics revealed IgE bindings to typical storage proteins, 2S albumin and vicilin, as well as to Bet v 1 homologous of "chestnut pea" and non-specific lipid transfer proteins. This case illustrates that the processing of protein-rich legume flours can pose new sensitization risks for workers, which should be considered in the future.

The AllergoVet study longitudinally examines the influence of animal exposure on the development of sensitization and allergic diseases among veterinary medicine students. In this group, contact to animals usually existed long before the study began. Therefore, the aim of this analysis was to investigate lifelong animal species-specific exposure and the prevalence of sensitizations and allergic symptoms already existing before the start of the study. Questionnaire data, including exposure history, were summarized to determine the duration and intensity of animal-related exposure as well as the prevalence of allergic symptoms to animals. Serologically, specific IgE was determined against ubiquitous inhalant allergens (atopy screen sx1) and against animal allergens using ImmunoCAP. The association between animal-specific sensitization, allergic symptoms, and exposure was analyzed using Fisher's exact test or CochranArmitage trend test. All study participants (n = 313) had previous contact with animals, with dogs mentioned most frequently (91.1%) followed by cats (89.5%) and horses (72.2%). Sensitization to ubiquitous allergens (positive sx1 value) was detected in 38.4% of subjects. Approximately 11%, 7%, and 5% were sensitized to cats, dogs, and horses, respectively. Only a small proportion of these sensitizations were associated with self-reported symptoms (41% for cat, 9% for dog, and 13% for horse). While no significant association between animal-specific exposure and sensitization was found for cats and horses, a clear trend emerged for dogs. With increasing duration of exposure to dogs, the number of dog-specific sensitizations decreased significantly (p = 0.0069). Furthermore, a decreasing trend in sx1 sensitization was noted with increasing cat (p = 0.0288) and dog (p = 0.0107) exposure. None of the subjects who grew up on a farm (n = 40) had any sensitization to animals. The sensitization prevalence determined among first-year students in veterinary medicine roughly corresponds to that in the general population. Most animal sensitizations were not clinically relevant. In this collective, a protective effect of increasing exposure to animals in childhood and adolescence was found on sensitization, which was particularly pronounced during contact with dogs.

The detection of occupational allergies is a complex undertaking: A case study According to the recommendations for the diagnosis of occupational asthma, suspected cases require a confirmed asthma diagnosis, work-associated symptoms, evidence of sensitization to an occupational allergen and a close relationship between the occupation and the symptoms. In addition to a detailed medical history, several examinations and tests and a differentiated consideration of them are required in order to arrive at a final expert assessment. We report the case of a 58-year-old butcher who had been complaining of shortness of breath and coughing at his workplace in a slaughterhouse for 5 years. It was only after extensive investigations and a detailed medical history that a powdered kebab spice used in an adjacent room was suspected as the potential allergen. Diagnostic tests showed that the patient was sensitized to the spice. While the specific inhalation challenge (SIC) with kebab spice did not elicit an asthmatic reaction, a non-specific bronchial hyperreactivity in the insured person was observed. On the day after the SIC, the bronchial hyperreactivity was increased by a factor of three and there were further indications of eosinophilic or allergic reaction. In summary, the individual diagnostic components led to the assessment of this case as allergic occupational asthma. This case illustrates that an assessment of the individual diagnostic components in the diagnosis of occupational asthma is quite complex and not without pitfalls. Keywords: occupational asthma – diagnosis – sensitization – bronchial hyperreactivity

In order to ensure valid diagnostics for occupational test allergen solutions despite the ongoing reduction in the availability of commercial test extracts, a plan B was initiated for the possible production of skin prick test (SPT) solutions in public pharmacies. For important occupational allergen sources (wheat and rye, storage mites, animal epithelia, mold material) laboratory extraction methods were analyzed in comparison to pharmacy compatible extraction methods regarding protein quantity and quality in SDS-PAGE combined with silver staining. Subsequently, using the example of bovine epithelia, adapted extraction procedures as well as in-process and final product controls were transferred to a public pharmacy. Allergen sources with a high protein content, such as wheat and rye grains as well as storage mites, showed good comparability of the extractable protein quantity and protein pattern, regardless of the applied extraction method. In contrast, allergen source materials with a low total protein content, such as animal epithelia and molds, can benefit from laboratory extraction conditions such as mechanical disruption and specific buffer additives. In the qualitative protein silver staining, characteristic protein patterns were identified for each allergen source. Depending on the extraction method, only minor differences in total protein patterns were observed in animal epithelia and molds. Using source materials from two suppliers, the resulting allergen extracts displayed clear differences in protein content in storage mites and quantitative and qualitative differences in molds. A practical preparation attempt of SPT solutions in a public pharmacy was successful. SPT solutions prepared with adapted pharmacy extraction methods showed a comparable protein and Bos d 2 allergen content and equivalent qualities in the protein pattern compared to a previously available commercial SPT solution. Accordingly, it can be assumed that standardized SPT solutions with sufficient allergen quality for occupational allergen sources can be prepared in public pharmacies if certified allergen sources with appropriate protein content are available.

OBJECTIVES:The study aimed to determine the allergen, endotoxin and β-(1,3)-glucan concentrations at various areas on a university campus of veterinary medicine. METHODS:Dust samples were collected four times a year for three years using electrostatic dust collectors (EDC) at 25 different locations on a campus of veterinary medicine and in laboratories of inorganic chemistry as a control area representing animal-free environment. Major animal allergens from dog, cat, horse, cattle and mouse, domestic mite (DM) allergens, and β-(1,3)-glucan were measured using enzyme immunoassays and endotoxin using the limulus amoebocyte lysate (LAL) assay. Seasonal, annual and local influences on exposure levels were analyzed using Bayesian mixed models. RESULTS:With the exception of mouse allergens, all other determinants were found in almost all locations on the campus and in the control area, but in up to 10.000-fold variable concentrations. By far the highest levels of feline, canine, equine and bovine allergens were detected in buildings where the respective species were examined. The highest levels of mouse and DM allergens, β-(1,3)-glucan and endotoxin occurred together and were associated with locations where large animals were present. In buildings without animals, allergen levels were considerably lower but still elevated at several locations compared to the control area, especially for dog and horse allergens, and β-(1,3)-glucan. Significant seasonal effects were observed for dog, cat, horse and DM allergens, and β-(1,3)-glucan. Variations between years were less apparent than between seasons (except for β-(1,3)-glucan). CONCLUSIONS:The strongest influencing factor on the concentration of mammalian allergens was the presence of the corresponding animal at the collection site. Seasonal influence on allergen concentrations was observed, while the overall exposure remained constant over the years. At locations with horses, elevated levels of mite allergens, endotoxin, and β-(1,3)-glucan can be expected, probably due to passive transfer from stable environment.

The availability of high-quality skin test allergens is a prerequisite for the reliable diagnosis of occupational type I allergies. Due to the withdrawal of existing marketing authorizations (MAs) by pharmaceutical companies and the lack of new MAs for commercial test allergens, there is an increasing diagnostic gap in Germany and other EU member states, which makes it necessary to investigate alternative ways of providing in vivo diagnostics. The German Medicinal Products Act (Arzneimittelgesetz = AMG) allows for the possibility of preparing medicinal products in pharmacies without the need for an MA or a manufacturing authorization pursuant to Section 13 (2) No. 1 in conjunction with Section 13 (2a) Sentence 2 No. 3 AMG. This also includes test allergens. In addition to the AMG, the requirements of the German Ordinance on the Operation of Pharmacies (Apothekenbetriebsordnung - ApBetrO) and the European Pharmacopoeia apply in particular. Medicolegal and practical challenges, as well as potentials of manufacturing skin prick test solutions in public pharmacies are presented based on examples of different allergen source materials.

AbstractObjectiveStandardised quantitative analysis of the humoral immune response to SARS‐CoV‐2 antigens may be useful for estimating the extent and duration of immunity. The aim was to develop enzyme‐linked immunosorbent assays (ELISAs) for the quantification of human IgG antibodies against SARS‐CoV‐2 antigens.MethodsEnzyme‐linked immunosorbent assays were developed based on monoclonal antibodies against human IgG and recombinant SARS‐CoV‐2 antigens (Spike‐S1 and Nucleocapsid). The WHO 67/086 immunoglobulin and WHO 20/136 SARS‐CoV‐2 references were used for standardisation. Sera of a study group of COVID‐19‐positive subjects (n = 144), pre‐pandemic controls (n = 135) and individuals vaccinated with BioNTech–Pfizer BNT162b2 vaccine (n = 48) were analysed. The study group sera were also tested using EuroImmun SARS‐CoV‐2‐ELISAs and a quantitative S1‐specific fluorescence enzyme immunoassay (FEIA) from Thermo Fisher.ResultsThe ELISA results were repeatable and traceable to international units because of their parallelism to both WHO references. In the study group, median anti‐S1‐IgG concentrations were 102 BAU mL−1, compared to 100 and 1457 BAU mL−1 in the vaccination group after first and second vaccination, respectively. The ELISAs achieved an area under the curve (AUC) of 0.965 (S1) and 0.955 (Nucleocapsid) in receiver operating characteristic (ROC) analysis, and a specificity of 1 (S1) and 0.963 (Nucleocapsid) and sensitivity of 0.903 (S1) and 0.833 (Nucleocapsid) at the maximum Youden index. In comparison, the commercial assays (S1‐FEIA, S1 and Nucleocapsid ELISA EuroImmun) achieved sensitivities of 0.764, 0.875 and 0.882 in the study group, respectively.ConclusionsThe quantitative ELISAs to measure IgG binding to SARS‐CoV‐2 antigens have good analytical and clinical performance characteristics and units traceable to international standards.