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Background Methods Results Conclusion In contrast to sublingual immunotherapy (SLIT) with recombinant Mal d 1 (rMal d 1‐SLIT), SLIT with rBet v 1 (rBet v 1‐SLIT) induced Mal d 1‐cross‐reactive antibodies without IgE‐blocking activity. To elucidate whether the development of cross‐protective IgG responses depends on the degree of molecular identity of allergens we compared the cross‐reactivity, cross‐blocking activity, and affinity of SLIT‐induced antibodies with allergens of varying amino acid sequence identities to Bet v 1 and Mal d 1, namely Cor a 1.04 (hazelnut), Pru av 1 (cherry), and Dau c 1 (carrot).Allergen‐specific antibodies were quantified by ELISA. IgE blocking was analyzed by inhibition of allergen‐induced basophil activation and IgE‐facilitated allergen‐presentation to T cells. The affinity of SLIT‐induced antibodies was studied by acidic dissociation ELISA and competition ELISA. Identical surface areas on allergens were predicted using an in‐house designed script based on structural alignments.rBet v 1‐SLIT‐induced IgG antibodies cross‐reacted with all allergens except Dau c 1. rMal d 1‐SLIT‐induced antibodies predominantly cross‐reacted with Pru av 1 and displayed significantly higher IgE blocking to Pru av 1 than rBet v 1‐SLIT‐induced antibodies. rMal d 1‐SLIT‐induced IgG1 showed higher affinity to Mal d 1 and Pru av 1. Surface analysis revealed 84% identical area on Mal d 1 and Pru av 1. Furthermore, we identified two surface areas potentially containing epitopes present on these allergens and absent on Bet v 1.In summary, our findings suggest that a relatively high threshold of similarity is required to establish effective cross‐blocking antibodies to related allergens. Apparently, the structural identity between Bet v 1 and Mal d 1 is below this threshold. Therefore, this study may explain why immunotherapy with birch pollen allergen often fails to reduce birch pollen‐related apple allergy. [ABSTRACT FROM AUTHOR]

ABSTRACT Penicillins are the most frequently prescribed class of medications worldwide and first‐line antibiotic of choice for most bacterial infections. They are also commonly labelled as the culprit of drug ‘allergy’; leading to obligatory use of second‐line antibiotics, suboptimal antibiotic therapy and increased antimicrobial resistance. However, the majority of reported penicillin ‘allergy’ labels are found to be incorrect after allergy testing, emphasising the importance of proper drug allergy testing and evaluation. Penicillin skin testing (PST) remains an important component of drug allergy diagnosis; however, its practice and policies significantly differ across the world. Inappropriate and non‐evidence‐based PST practices can lead to consequences associated with allergy mislabelling. Even within different regions of China, with a population exceeding 1.4 billion, there are marked differences in the implementation, execution and interpretation of PST. This review aims to examine the differences in PST between Mainland China, Hong Kong and the rest of the world. We critically analyse the current practice of ‘pre‐emptive’ PST in Mainland China, which has a significant false‐positive rate leading to high levels of penicillin allergy mislabelling. Non‐evidence‐based practices further compound the high false‐positive rates of indiscriminatory PST. We postulate that inappropriate PST policies and practices may exacerbate the mislabelling of penicillin allergy, leading to unnecessary overuse of inappropriate second‐line antibiotics, increasing antimicrobial resistance and healthcare costs. We advocate for the importance of more collaborative research to improve the contemporary workflow of penicillin allergy diagnosis, reduce mislabelling and promote the dissemination of evidence‐based methods for allergy diagnosis. [ABSTRACT FROM AUTHOR]

A systematic review and meta-analysis compared the efficacy and safety of subcutaneous immunotherapy (SCIT) and sublingual immunotherapy (SLIT) in treating allergic rhinitis. The study found that SCIT had a significantly lower symptom score compared to SLIT, but there was no significant difference in medication scores between the two groups. Patients treated with SCIT had a higher incidence of systemic reaction grades 3 and 4 compared to SLIT. While SCIT is more effective in controlling symptoms, clinicians should consider patient preference, adherence, convenience, and cost-effectiveness when choosing a treatment. Further research is needed in this area. [Extracted from the article]

Background: IgE antibodies to cross‐reactive carbohydrate determinants (CCD) are usually clinically irrelevant but they can be a cause of false positive outcomes of allergen‐specific IgE tests in vitro. Their prevalence and levels have been so far cross‐sectionally examined among adult allergic patients and much less is known about their origins and relevance in childhood. Methods: We examined CCD with a cross‐sectional approach in 1263 Italian pollen allergic children (Panallergen in Paediatrics, PAN‐PED), as well as with a longitudinal approach in 612 German children (Multicenter Allergy Study, MAS), whose cutaneous and IgE sensitization profile to a broad panel of allergen extracts and molecules was already known. The presence and levels of IgE to CCD were examined in the sera of both cohorts using bromelain (MUXF3) as reagent and a novel chemiluminescence detection system, operating in a solid phase of fluorescently labelled and streptavidin‐coated paramagnetic microparticles (NOVEOS, HYCOR, USA). Results: IgE to CCD was found in 22% of the Italian pollen allergic children, mainly in association with an IgE response to grass pollen. Children with IgE to CCD had higher total IgE levels and were sensitized to more allergenic molecules of Phleum pratense than those with no IgE to CCD. Among participants of the German MAS birth cohort study, IgE to CCD emerged early in life (even at pre‐school age), with IgE sensitization to group 1 and 4 allergen molecules of grasses, and almost invariably persisted over the full observation period. Conclusions: Our results contribute to dissect the immunological origins, onset, evolution and risk factors of CCD‐sIgE response in childhood, and raise the hypothesis that group 1 and/or 4 allergen molecules of grass pollen are major inducers of these antibodies through an antigen‐specific, T‐B cell cognate interaction. [ABSTRACT FROM AUTHOR]

This article discusses the diagnosis of paediatric mild acute Food Protein-Induced Enterocolitis Syndrome (FPIES). The current diagnostic criteria for FPIES may not accurately diagnose milder cases of the disease. The article proposes new criteria for the diagnosis of paediatric mild acute FPIES. The study retrospectively reviewed cases of paediatric mild acute FPIES and found that the new diagnostic and suspicion criteria were able to identify all 15 cases. The article provides detailed information on the patients' demographics and symptoms. These new diagnostic and suspicion criteria aim to include all mild forms of pediatric acute FPIES and may be useful for community pediatricians, although they have not been prospectively validated and are based on expert opinion. [Extracted from the article]

Background: In birch‐dominated areas, allergies to pollen from trees of the order Fagales are considered to be initiated by the major birch pollen allergen Bet v 1. However, the sensitizing activity of Bet v 1‐homologs in Fagales pollen might be underestimated. Allergen‐specific T‐cells are crucial in the sensitization process. The T‐cell response to major allergens from alder, hazel, oak, hornbeam, chestnut, beech, and chestnut pollen has not yet been analyzed. Here, we characterized the cellular cross‐reactivity of major allergens in Fagales pollen with Bet v 1. Methods: T‐cell‐lines (TCL) were established from allergic individuals with Aln g 1, Car b 1, Ost c 1, Cor a 1, Fag s 1, Cas s 1, and Que a 1, and tested for reactivity with Bet v 1 and synthetic overlapping 12‐mer peptides representing its primary sequence. Aln g 1‐specific TCL was additionally tested with Aln g 1‐derived peptides and all allergens. IgE‐competition experiments with Aln g 1 and Bet v 1 were performed. Results: T‐cell‐lines initiated with Fagales pollen allergens varied strongly in their reactivity with Bet v 1 and by the majority responded stronger to the original stimulus. Cross‐reactivity was mostly restricted to the epitope Bet v 1142–153. No distinct cross‐reactivity of Aln g 1‐specific T‐cells with Bet v 1 was detected. Among 22 T‐cell epitopes, Aln g 1 contained two immunodominant epitopes. Bet v 1 inhibited IgE‐binding to Aln g 1 less potently than Aln g 1 itself. Conclusion: The cellular cross‐reactivity of major Fagales pollen allergens with Bet v 1 was unincisive, particularly for Aln g 1, most akin to Bet v 1. Our results indicate that humoral and cellular responses to these allergens are not predominantly based on cross‐reactivity with the major birch pollen allergen but suggest a Bet v 1‐independent sensitization in individuals from birch tree‐dominated areas. [ABSTRACT FROM AUTHOR]

Background: House dust mites (HDM) are among the most important sources for airborne allergens with high relevance for atopic diseases. Routine tests contain only 4 of 32 registered allergens of Dermatophagoides pteronyssinus. Clinical relevance and pathomechanistic properties of many allergens are not well understood. Objective: The association of several HDM allergens with allergic rhinitis, allergic asthma, and atopic dermatitis was investigated to identify allergens with biomarker potential and to transfer them into diagnostics. Methods: Eight out of nine D. pteronyssinus allergens (nDer p 1, rDer p 2, rDer p 5, rDer p 7, rDer p 10, rDer p 13, rDer p 20, rDer p 21, rDer p 23) were recombinantly expressed and purified. Sensitization patterns of 384 HDM‐allergic individuals exhibiting different clinical phenotypes were analyzed with a serum‐saving multiplex array. Results: Sensitization to more than three mite allergens (sensitization count) was associated with allergic asthma and/or atopic dermatitis. Reactions to Der p 5 and Der p 21 were more frequent in allergic asthma compared to allergic rhinitis. Atopic dermatitis patients were more often sensitized to Der p 5, Der p 20, and Der p 21 among others. Der p 20‐IgE > 80 kU/L was associated with severe atopic dermatitis in 75% of patients. Conclusion: This study demonstrates the clinical importance of the sensitization count and of certain allergens (Der p 5, Der p 20, and Der p 21) not available for routine diagnostics yet. Implementing them as well as the sensitization count in diagnostic measures will improve diagnosis and risk assessment of HDM‐allergic patients. [ABSTRACT FROM AUTHOR]

Background: Immunoglobulin E (IgE) sensitization is associated with asthma and allergic diseases. Gestational age influences early immune system development, thereby potentially affecting the process of tolerance induction to allergens. Objective: To study IgE sensitization to common allergens by gestational age from childhood up to early adulthood. Methods: Population‐based birth cohort, data from the Swedish BAMSE study were used. Allergen‐specific IgE antibodies to a mix of common food (fx5) and inhalant (Phadiatop) allergens were analysed at 4, 8, 16 and 24 years. Sensitization was defined as allergen‐specific IgE ≥0.35 kUA/L to fx5 and/or Phadiatop at each time point. Using logistic regression and generalized estimated equations, adjusted odds ratios (aORs) for sensitization in relation to gestational age were calculated. Replication was sought within the Swedish twin study STOPPA. Results: In BAMSE, 3522 participants were screened for IgE antibodies during follow‐up; of these, 197 (5.6%) were born preterm (<37 gestational weeks) and 330 (9.4%) post‐term (≥42 weeks). Preterm birth reduced the risk of sensitization to common food and/or inhalant allergens up to early adulthood by 29% (overall aOR = 0.71; 95% CI: 0.52–0.98), and to food allergens specifically by 40% (overall aOR = 0.60; 95% CI: 0.38–0.93). No relation was found between post‐term birth and IgE sensitization at any time point. Replication analyses in STOPPA (N = 675) showed similar risk estimates for sensitization to food and/or inhalant allergens (aOR = 0.72; 95% CI: 0.42–1.21), which resulted in a combined meta‐analysis aOR = 0.71 (95% CI: 0.54–0.94). Conclusions: Our study suggests an inverse association between preterm birth and long‐term IgE sensitization. [ABSTRACT FROM AUTHOR]

Charlene M Prazma,1 Marco Idzko,2,3 Jo Anne Douglass,4 Arnaud Bourdin,5,6 Stephen Mallett,7 Frank C Albers,8 Steven W Yancey1 1Respiratory Therapeutic Area, GSK, Research Triangle Park, NC, USA; 2Division of Pneumology, Medical University of Vienna, Vienna, Austria; 3Department of Pneumology, University Hospital Freiburg, Freiburg, Germany; 4The Royal Melbourne Hospital and University of Melbourne, Melbourne, VIC, Australia; 5Departement de Pneumologie et Addictologie, Hôpital Arnaud de Villeneuve, CHU Montpellier, Montpellier, France; 6PhyMedExp, University of Montpellier, Montpellier, France; 7Clinical Statistics, GSK, Uxbridge, Middlesex, UK; 8Respiratory Medical Franchise, GSK, Research Triangle Park, NC, USACorrespondence: Charlene M PrazmaRespiratory Therapeutic Area, GSK, 5 Moore Drive, PO Box 13398, Research Triangle Park, NC, 27709, USATel +1 919 483 9013Email charlene.m.prazma@gsk.comPurpose: Improved understanding of characteristics that may influence treatment response across phenotypes may help guide treatment decisions.Patients and Methods: This was a post hoc analysis of MENSA, a multicenter, randomized, double-blind, placebo-controlled trial (NCT01691521). Patients aged ≥ 12 years with severe eosinophilic asthma received mepolizumab (75 mg intravenously or 100 mg subcutaneously) or placebo, plus standard of care, every 4 weeks for 32 weeks. Outcomes assessed were the annualized rate of clinically significant exacerbations and change from baseline in Asthma Control Questionnaire (ACQ)-5 score. Subgroup analyses were performed by baseline blood eosinophil count (< 150, ≥ 150– 300, ≥ 300 cells/μL) within atopic subgroups (non-atopic [specific immunoglobulin E < 0.35 kU/L], atopic [≥ 0.35– 17.5 kU/L], strongly atopic [> 17.5 kU/L]), and by house dust mite (HDM) sensitivity.Results: Of 576 patients analyzed, 272 were non-atopic, 181 were atopic and 94 were strongly atopic; 29 had missing atopy data. In patients with blood eosinophil counts ≥ 300 cells/μL, mepolizumab versus placebo reduced clinically significant exacerbations by 74%, 43% and 25% in the non-atopic, atopic and strongly atopic subgroups. Similar reductions were observed in all atopic subgroups in other blood eosinophil count categories where there were sufficient patient numbers for analysis, except for non-atopic patients with baseline blood eosinophil counts of < 150 cells/μL. Improvements in ACQ-5 scores of – 0.75, – 0.73 and – 0.78 with mepolizumab versus placebo were observed in non-atopic, atopic and strongly atopic patients with blood eosinophil counts ≥ 300 cells/μL; consistent improvements in ACQ-5 were not observed in patients with blood eosinophil counts < 150 or ≥ 150– 300 cells/μL. Reductions in clinically significant exacerbations with mepolizumab versus placebo were also observed irrespective of sensitivity to HDMs.Conclusion: Mepolizumab was associated with a trend for reductions in clinically significant exacerbations and improved asthma control versus placebo in patients with severe eosinophilic asthma, irrespective of atopic status or HDM sensitivity.Keywords: allergens and epitopes, eosinophils, asthma, asthma treatments, biologics

The 5-year mean total pollen concentrations of Cupressaceae, I Platanus i , I Quercus i and I Olea i , but not Poaceae, increased more dramatically, and were significantly correlated with the increase in temperature (Figure 1). Quercus ilex pollen allergen, Que i 1, responsible for pollen food allergy syndrome caused by fruits in Spanish allergic patients. Severity of pollen-induced symptoms depends on the number of pollen grains and their allergenicity, variables related to pollution and local climate.2 Therefore, climate change could potentially change pollen exposure, sensitization and symptoms.3-5 However, only sparse data exist regarding the direct correlation between climate changes, pollen seasons and allergic sensitization. Keywords: allergens and epitopes; basic mechanisms; climate change; clinical immunology; environment and hygiene hypothesis; IgE; Madrid; pollinosis EN allergens and epitopes basic mechanisms climate change clinical immunology environment and hygiene hypothesis IgE Madrid pollinosis 574 577 4 04/04/22 20220401 NES 220401 Key Messages We observed a 1.3ºC increase in temperature in Madrid over 40 years. [Extracted from the article]

Keywords: allergens and epitopes; anaphylaxis; COVID-19 vaccines; drug allergy; education; IgE; polyethylene glycol (PEG); polysorbate 80; skin prick test EN allergens and epitopes anaphylaxis COVID-19 vaccines drug allergy education IgE polyethylene glycol (PEG) polysorbate 80 skin prick test 465 469 5 02/24/22 20220301 NES 220301 Key Messages We evaluated 60 adult patients with adverse reactions to a COVID-19 vaccine. Patients with a convincing history of vaccine anaphylaxis with objective symptoms or with suspected PEG and/or PS80 allergy were invited for allergy skin testing. Specialist allergy advice allows vaccination in patients with reactions to COVID-19 vaccines Allergens and epitopes, anaphylaxis, drug allergy, education, IgE, skin prick test, COVID-19 vaccines, polyethylene glycol (PEG), polysorbate 80. [Extracted from the article]

Background: Mouse allergy is an important cause of indoor asthma and allergic rhinoconjunctivitis. The major mouse allergen, Mus m 1, is a complex of homologous pheromone‐binding lipocalins called major urinary proteins (MUPs). Methods: We analyzed the proteome of MUPs in mouse urine, commercial mouse epithelial extracts, and environmental samples using several approaches. These include as follows: two‐dimensional electrophoresis and immunoblotting; liquid chromatography‐high‐resolution mass spectrometry (LC/HRMS); multiple reaction monitoring (MRM) mass spectrometry; and LC/HRMS analysis of glycans at the N‐66 residue of MUP3. Results: Albumin is predominant in the extracts, while MUPs are predominant in urine. LC/HRMS of 4 mouse allergen extracts revealed surprising heterogeneity. Of 22 known mouse MUPs, only 6 (MUP3, MUP4, MUP5, MUP13, MUP20, and MUP21) could be identified with MRM using unique peptides. Assessment of MUP content in urine, extracts, and dust samples showed good correlation between MRM and other methods working with different detection principles. All 6 identifiable MUPs were found in electrophoretically separated urine bands, but only MUP3 and MUP20 were above LOQ in unseparated mouse urine, and only MUP3, MUP4, and MUP20 were found in mouse epithelial extracts. Glycan heterogeneity was noted among 4 individual inbred mice: of 13 glycan structures detected, 8 were unique to one mouse, and only 2 glycan modifications were present in all 4 mice. Conclusions: Using mass spectrometry and MRM, mouse allergen extracts and urine samples are shown to be complex and heterogeneous. The efficacy and safety of commercial mouse allergen extracts will be improved with better controls of allergen content. [ABSTRACT FROM AUTHOR]

While 2020 will be remembered for the global coronavirus pandemic, there were also important advances in the field of allergy. In this review article, we summarize key findings reported in Clinical and Experimental Allergy during 2020. We hope this provides readers with an accessible snapshot of the work published in our journal during this time. [ABSTRACT FROM AUTHOR]

Keywords: allergens and epitopes; anaphylaxis; clinical immunology; drug allergy; epidemiology EN allergens and epitopes anaphylaxis clinical immunology drug allergy epidemiology 1514 1518 5 10/29/21 20211101 NES 211101 Key Messages The incidence of chemotherapy-induced anaphylaxis in the United States is 1 in 1,611. Patients with anaphylaxis had higher total hospital costs (median $38,226) compared with hospitalized patients receiving chemotherapy without anaphylaxis (median $25,344) ( I p i < .0001). Allergens and epitopes, anaphylaxis, clinical immunology, drug allergy, epidemiology. [Extracted from the article]

Solely in allergics, an interplay of mucosal antibody subclass interaction led to outcompeting of allergen-IgG binding by the high abundance of low-affinity SIgA. Nasal allergen-neutralizing IgG4 antibodies block IgE-mediated responses: novel biomarker of subcutaneous grass pollen immunotherapy. High-affinity Bet v 1-specific secretory IgA antibodies in nasal fluids protect against birch pollen allergy Keywords: allergens and epitopes; allergy diagnosis; immunologic tests; mucosal immunity; pollen EN allergens and epitopes allergy diagnosis immunologic tests mucosal immunity pollen 2267 2270 4 07/02/21 20210701 NES 210701 Nasal fluid antibodies act as specific barrier molecules against inhaled antigens through neutralization and shielding mechanisms. [Extracted from the article]

We showed that only 29% of positive patch test reactions observed before dupilumab treatment, could be re-elicited in AD patients using dupilumab treatment. Keywords: allergens and epitopes; allergic contact dermatitis; atopic dermatitis; dermatology EN allergens and epitopes allergic contact dermatitis atopic dermatitis dermatology 959 967 9 06/30/21 20210701 NES 210701 DATA AVAILABILITY STATEMENT The data that support the findings of this study are available in our manuscript (Table 2, Appendix S2). Future studies investigating patch test reactions in patients who discontinued dupilumab treatment and evaluation of the effect of dupilumab in patients with recalcitrant ACD would be of added value. Adult AD patients who had at least 1 positive reaction in a PT conducted before the start of dupilumab treatment, elicited by an allergen that was re-tested during dupilumab treatment (>=12 weeks) were eligible. [Extracted from the article]

Background: Sensitization to thermotolerant fungi, including filamentous fungi and Candida albicans, is associated with poor lung function in adults with severe asthma. Data in children are lacking. Environmental exposure to fungi is linked with acute severe asthma attacks, but there are few studies reporting the presence of fungi in the airways during asthma attacks. Methods: We investigated the association between fungal sensitization and/or positive fungal sputum culture and markers of asthma severity in children with chronic and acute asthma. Sensitization was determined using serum‐specific IgE and skin prick testing against a panel of five fungi. Fungal culture was focused towards detection of filamentous fungi from sputum samples. Results: We obtained sensitization data and/or sputum from 175 children: 99 with chronic asthma, 39 with acute asthma and 37 controls. 34.1% of children with chronic asthma were sensitized to thermotolerant fungi compared with no children without asthma (p =< 0.001). These children had worse pre‐bronchodilator lung function compared with asthmatics without sensitization including a lower FEV1/FVC ratio (p <.05). The isolation rate of filamentous fungi from sputum was higher in children with acute compared with chronic asthma. Conclusions: Fungal sensitization is a feature of children with chronic asthma. Children sensitized to thermotolerant fungi have worse lung function, require more courses of systemic corticosteroids and have greater limitation of activities due to asthma. Asthma attacks in children were associated with the presence of filamentous fungi positive sputum culture. Mechanistic studies are required to establish whether fungi contribute directly to the development of acute asthma. [ABSTRACT FROM AUTHOR]

Background: Children with atopic dermatitis (AD) are often sensitized to food and aeroallergens, but sensitization patterns have not been analysed with biologic measures of disease pathogenicity. Objective: We sought to define allergen sensitization grouping(s) using unbiased machine learning and determine their associations with skin filaggrin (FLG) and transepidermal water loss (TEWL) (assesses skin barrier integrity), S100A8 and S100A9 expression (assesses skin inflammation) and AD severity. Methods: We studied 400 children with AD in the Mechanisms of Progression from Atopic Dermatitis to Asthma in Children (MPAACH) cohort to identify groupings of food and aeroallergen sensitizations. MPAACH is a paediatric AD cohort, aged 1–2, recruited through hospital/community settings between 2016 and 2018. We analysed these groupings' associations with AD biomarkers: skin FLG, S100A8 and S100A9 expression, total IgE, TEWL and AD severity. Results: An unbiased machine learning approach revealed five allergen clusters. The most common cluster (N = 131), SPTPEP, had sensitization to peanut, egg and/or pets. Three low prevalence clusters, which included children with allergen sensitization other than peanut, egg or pets, were combined into SPTOther. SPTNEG included children with no sensitization(s). SPTPEP children had higher median non‐lesional TEWL (16.9 g/m2/h) and IgE (90 kU/L) compared with SPTOTHER (8.8 g/m2/h and 24 kU/L; p =.01 and p <.001) and SPTNEG (9 g/m2/h and 26 kU/L; p =.003 and p <.001). SPTPEP children had lower median lesional (0.70) and non‐lesional (1.09) FLG expression compared with SPTOTHER (lesional: 0.9; p =.047, non‐lesional: 1.78; p =.01) and SPTNEG (lesional: 1.47; p <.001, non‐lesional: 2.21; p <.001). There were no differences among groupings in S100A8 or S100A9 expression. Conclusions and clinical relevance: In this largely clinic‐based cohort of young children with AD, allergic sensitization to peanut, egg, cat or dog was associated with more severe disease and skin barrier function but not markers of cutaneous inflammation. These data need replicating in a population‐based cohort but may have important implications for understanding the interaction between AD and allergic sensitization. [ABSTRACT FROM AUTHOR]

Background: Inducible laryngeal obstruction (ILO) is often misdiagnosed as, or may coexist with, asthma. Identifying differences in triggering factors may assist clinicians to differentiate between the two conditions and could give mechanistic insights. Objective: To identify and compare patient‐reported triggers in ILO and asthma. Methods: This was a two‐part study. Initially, we conducted a retrospective case note review of the triggers of ILO from endoscopically confirmed ILO patients to generate a Breathlessness Triggers Survey (BrTS). Triggers were categorized as scents, environmental factors, temperature, emotions, mechanical factors and daily activities. Secondly, ILO and/or asthma patients completed the BrTS prospectively, rating the likelihood of each item triggering their symptoms using a five‐point Likert scale (strongly disagree to strongly agree). Chi‐square testing was performed to compare responses by cohort. Results: Data from 202 patients with ILO [73% female, mean (SD) age 53(16) years] were included in the case note review. For the prospective study, 38 patients with ILO only [63% females, age 57(16) years], 39 patients with asthma only [(56% female, age 53(13) years] and 12 patients with both ILO and asthma [83% female, mean age, 57 (14) years)] completed the BrTS. The triggers identified in the case note review were confirmed in the independent sample of patients with ILO and/or asthma and identified several difference in prevalence of the triggers between disease types. Mechanical factors (talking [P <.001], shouting [P =.007] and swallowing [P =.002]) were more common in the ILO cohort compared to patients with asthma. Environmental factors (pollen/flowers [P =.005] and damp air [P =.012]) were more common in asthma. There were no differences between groups in frequency of reporting scents as triggers (except for vinegar, more common in ILO, P =.019), temperature, emotions or daily activities. Conclusion: There were notable differences between patient‐reported triggers of ILO and asthma, which may support clinician differential diagnosis. [ABSTRACT FROM AUTHOR]

Keywords: allergens and epitopes; IgE; immunotherapy vaccines and mechanisms EN allergens and epitopes IgE immunotherapy vaccines and mechanisms 1503 1507 5 06/01/20 20200601 NES 200601 Studies demonstrated that among the identified I B tropicalis i allergens, Blo t 5 is a major allergen,[[1], [3]] which makes the protein a target for diagnosis and allergen-specific immunotherapy (AIT). Cross-inhibition results showed that rBlo t 5 SB short sb cannot always inhibit the rDer p 5-specific IgE, but rDer p 5 possesses a moderate inhibitory capacity on rBlo t 5 SB short sb -specific IgE binding (Figure S4F). Blomia tropicalis Blo t 5 and Blo t 21 recombinant allergens might confer higher specificity to serodiagnostic assays than whole mite extract. [Extracted from the article]

Background: Food processing, including heat‐treatment, can affect protein structure and stability, and consequently affect protein immunogenicity and allergenicity. A few studies have shown that structural changes induced by heat‐treatment impact the intestinal protein uptake and suggest this as a contributing factor for altered allergenicity. Objective: To investigate the impact of heat‐treatment of a whey‐based protein product on allergenicity and tolerogenicity as well as on intestinal uptake in various animal models. Methods: Immunogenicity and sensitizing capacity of the heat‐treated whey product were compared to that of the unmodified product by intraperitoneal and oral exposure studies, while tolerogenic properties were assessed by oral primary prevention and desensitization studies in high‐IgE responder Brown Norway rats. Results: Heat‐treatment of whey induced partial protein denaturation and aggregation, which reduced the intraperitoneal sensitizing capacity but not immunogenicity. In contrast, heat‐treatment did not influence the oral sensitizing capacity, but the heat‐treated whey showed a significantly reduced eliciting capacity compared to unmodified whey upon oral challenge. Heat‐treatment did not reduce the tolerogenic properties of whey, as both products were equally good at preventing sensitization in naïve rats as well as desensitizing already sensitized rats. Results from inhibitory ELISA and immunoblots with sera from sensitized rats demonstrated that heat‐treatment caused an altered protein and epitope reactivity. Protein uptake studies showed that heat‐treatment changed the route of uptake with less whey being absorbed through the epithelium but more into the Peyer's patches. Conclusion and Clinical Relevance: These results support the notion that the physicochemical features of proteins affect their route of uptake and that the route of uptake may affect the protein allergenicity. Furthermore, the study highlights the potential for heat‐treatment in the production of efficient and safe cow's milk protein‐based products for prevention and treatment of cow's milk allergy. [ABSTRACT FROM AUTHOR]

Background: Allergen extracts are the primary tool for diagnosis and treatment of allergic diseases. In the United States, most allergen extracts are non‐standardized. More sophisticated analytical approaches are needed to characterize these products and enable manufacturers and regulators to better determine potency. Objective: To expand the multiple reaction monitoring (MRM) assay for an in‐depth characterization of German cockroach (GCr; Blattella germanica) allergen extracts. Methods: We applied advanced liquid chromatography (LC) and mass spectrometry (MS) techniques including MRM. The expanded LC/MRM‐MS method was optimized to measure known GCr allergens and their isoforms/variants in commercial extracts and environmental samples. We performed isoform‐specific allergen measurements in multiple extracts from four commercial sources and extracts prepared using environmental samples from urban homes. To investigate causes of heterogeneity, we examined over 30 extraction process variables. Results: Evaluation of the commercial extracts confirmed the variability of production lots and commercial sources. Commonly used defatting and extraction protocols yielded extracts with comparable allergen profiles and content. However, the identity and quality of source materials was a major contributor to variability. In comparing commercial GCr extracts to environmental samples, relative quantities of Bla g 1, Bla g 2, Bla g 3, Bla g 4 and Bla g 11 were similar, while Bla g 5, Bla g 6, Bla g 7 and Bla g 8 were present in the environmental samples but largely absent for the commercial extracts. Conclusions and Clinical Relevance: LC/MRM‐MS can be used to measure all known GCr allergens in commercial allergen extracts and environmental samples. Significant differences exist between allergen profiles of commercial extracts and the profiles of environmental samples from dwellings. This analytical platform can serve as a template to achieve better product characterization of similarly complex products. [ABSTRACT FROM AUTHOR]

Background: Skin patch testing is still seen as the gold standard for the diagnosis of allergic hypersensitivity. For several metals and for patients with a suspected adverse reaction to their medical device implant material, patch testing can be unreliable. The current alternative to metal allergy patch testing is the in vitro lymphocyte proliferation test (LPT) using tritiated thymidine. This method is well‐established but requires handling of radioactive material, often uses heat‐inactivated allogenic human pooled serum and cannot determine T cell subsets. Objective: To develop a radioactive free LPT by using carboxyfluorescein succinimidyl ester (CFSE) and to evaluate the influence of serum source (heat‐inactivated human pooled serum [HI HPS] vs autologous serum) on the sensitivity and specificity of the nickel‐specific LPT. Methods: Peripheral blood mononuclear cells derived from nickel‐allergic patients and healthy controls were collected, labelled with CFSE and cultured in medium containing 10% HI HPS or 10% autologous serum with or without additional T cell skewing cytokine cocktails (Th1: IL‐7/IL‐12, Th2: IL‐7/IL‐4 or Th17: IL‐7/IL‐23/IL‐1β) in the absence or presence of NiSO4. The stimulation index (SI) was calculated as the ratio of divided cells, that is the percentage of CFSElow/neg CD3+CD4+ T‐lymphocytes upon nickel stimulation compared to the percentage of CFSElow/neg CD3+CD4+ T‐lymphocytes without antigen. These results were compared with the history of Ni allergy, patch test results and the MELISA test. Results: Autologous serum positively influenced Ni‐specific proliferation while HI HPS negatively influenced Ni‐specific proliferation. The test protocol analysing CD4+ cells and autologous serum without skewing cytokines scored the best diagnostic values (sensitivity 95%; specificity 93%; and overall accuracy 94%) compared to the parallel test using HI HPS (accuracy 60%). Cytokine supplements did not further improve the test protocol which used autologous serum. The protocol using HI HPS could be further improved by addition of the cytokine skewing cocktails. Conclusions: Here, we describe an optimized and highly accurate flow cytometric LPT which comprises of CFSE‐labelled cells cultured in autologous serum (not heat inactivated) and without the presence of T cell skewing cytokines. Clinical relevance: The sensitivity and specificity of LPT is enhanced, compared to HI HPS, when autologous serum without skewing cytokines is used. [ABSTRACT FROM AUTHOR]

Enhanced frequency and function of follicular T cells in the tonsils of house dust mite-sensitized children Keywords: allergens and epitopes; B cells; mucosal immunology; pediatrics; T cells EN allergens and epitopes B cells mucosal immunology pediatrics T cells 1240 1243 4 05/20/20 20200501 NES 200501 Abbreviations GC germinal center HDM house dust mite OSA obstructive sleep apnea TFH T follicular helper cells TFR T follicular regulatory cells T follicular helper cells (TFH) are a specialized subset of CD4 SP + sp T cells that regulate B cell responses within the germinal center (GC). CD4 SP + sp T cells associated with B cell follicles were separated into those found within the GC, GC TFH (CXCR5 SP hi sp PD1 SP hi sp Foxp3 SP - sp CD25 SP - sp ), and those that reside outside of the GC, non-GC TFH (CXCR5 SP int sp PD1 SP int sp Foxp3 SP - sp CD25 SP - sp ). [Extracted from the article]

Background: Small, basic peanut proteins are often poorly extracted in pH‐neutral buffers that are optimal for the extraction of peanut storage proteins such as Ara h 1. As a result, such proteins are easily missed as potential allergens. Objective: To analyse the allergenic composition of the basic peanut protein (BPP) fraction. Methods: A peanut extract prepared at pH 4 was fractionated by physicochemical procedures. Chemical analysis was performed by SDS‐PAGE and mass spectrometry. Because immunoblotting was found to be inefficient for most of these small basic proteins, IgE‐binding activity was measured by coupling the fractions to CNBr‐activated Sepharose, followed by incubation with sera from 55 Dutch peanut‐allergic children and 125I‐labelled anti‐IgE. Results: Most IgE reactivity of the BPP fraction was due to the 5‐7 kDa amino‐terminal fragment of Ara h 1. This finding was confirmed by the use of the fragment in recombinant form, to which 25/55 of the sera was IgE‐positive. Conclusion: The amino‐terminal fragment of Ara h 1, a member of a family of small anti‐microbial proteins, is an allergen independent of the carboxy‐terminal fragment of Ara h 1. [ABSTRACT FROM AUTHOR]

Background: Allergy to German cockroach (CR) is common in urban environments and is an important allergen in children with asthma. Objective: We hypothesize that the evolution of allergic sensitization and clinical disease is associated with distinct patterns of allergen‐specific T cell reactivity. To test this hypothesis, a subset of high‐risk inner‐city children participating in the URECA (Urban Environment and Childhood Asthma) birth cohort were selected to evaluate CR‐specific T cell reactivity from three distinct groups based on acquisition of aeroallergen sensitivity from ages 2 to 10: low atopy with minimal to no sensitivity (n = 26), early‐onset allergic sensitization (n = 25) and late‐onset allergic sensitization (n = 25). Methods: Using pools of previously identified CR‐derived T cell epitopes, we characterized the allergen‐specific T cell response in these 76 subjects from blood samples obtained at age 10. CR‐specific production of IL‐5, IFNγ and IL‐10 was measured by ELISPOT following two‐week in vitro culture with CR extract. Results: T cell responses were significantly higher in the early‐onset atopy group compared to low atopy (P = 0.01), and a trend for higher cytokine production in the late onset compared to the low atopy cohort was also observed (P = 0.06). T cell responses were similar between early‐ and late‐onset cohorts. Furthermore, a comparison of T cell reactivity between asthmatic and non‐asthmatic individuals revealed significantly higher cytokine production in asthmatics compared to non‐asthmatics (P = 0.02) within both the CR‐allergic and non‐allergic cohorts. Conclusions and clinical relevance: In conclusion, the present study reports that higher T cell reactivity is associated with allergen sensitization and asthma. Interestingly, no significant difference in T cell reactivity was observed in allergic children with early‐onset versus late‐onset atopy. [ABSTRACT FROM AUTHOR]

Background: Peach is a common elicitor of food allergic reactions. Peach‐induced immediate reactions may occur as benign pollen‐food syndromes, usually due to birch pollen‐related PR‐10 cross‐reactivity in temperate climates, and as potentially severe primary food allergies, predominantly related to nsLTP Pru p 3 in Mediterranean regions. The newly described peach allergen Pru p 7 has gained recent attention as a potential peach allergy severity marker. Sensitization to Pru p 7 and its allergenic homologues of the gibberellin‐regulated protein family occurs in areas with high Cupressaceae tree pollen exposure. Objective: We sought to investigate the distribution, clinical characteristics and molecular associations of Pru p 7 sensitization among subjects with suspected peach allergy in different regions of France. Methods: Subjects with suspected peach allergy (n = 316) were included. Diagnostic work‐up was performed according to current guidelines, including open food challenge when required. IgE antibody measurements and competition experiments were performed using the ImmunoCAP assay platform. Results: Sensitization to Pru p 7 was present in 171 (54%) of all subjects in the study and in 123 of 198 (62%) diagnosed as peach allergic, more than half of whom were sensitized to no other peach allergen. Frequency and magnitude of Pru p 7 sensitization were associated with the presence of peach allergy, the clinical severity of peach‐induced allergic reactions and the level of cypress pollen exposure. Cypress pollen extract completely outcompeted IgE binding to Pru p 7. Pru p 7 was extremely potent in basophil activation tests. Conclusion and Clinical Relevance: A subtype of Cupressaceae pollinosis, characterized by Pru p 7 sensitization, can be an underlying cause of severe peach allergy. [ABSTRACT FROM AUTHOR]

Summary: Background: Previous clinical trials with birch pollen subcutaneous immunotherapy have been conducted over a 1‐ to 2‐year treatment period and involved mostly a single geographic location. Objective: This study (EudraCT‐Number: 2005‐000025‐35) intended to evaluate the effect of subcutaneous immunotherapy with high‐dose hypoallergenic birch pollen allergoid in patients with confirmed moderate to severe seasonal allergic rhinitis/rhinoconjunctivitis over a 3‐year course in 19 European centres. Methods: Adults with confirmed birch pollen allergy (n = 253) were randomized to preseasonal placebo (n = 129) or active treatment (n = 124). Primary endpoint was change in Symptom Medication Score after 2 years treatment (2007). Results: The change in Symptom Medication Score of active‐ vs placebo‐treated patients for the Full Analysis Set (n = 227, 15.2% reduction, P = 0.0710) and Per‐Protocol Set (n = 216, 16.7% reduction, P = 0.0523) showed a positive trend, although significance was not achieved. The primary endpoint, assessed in 2007, coincided with the lowest pollination during the study period. In a subgroup analysis of patients in the north‐eastern region (n = 102), where birch is the major tree and consequently patients' exposure is higher, changes in Symptom Medication Score (32.7% reduction, P = 0.0034) and median number of well days (P = 0.0232) were highly significant in favour of the active group. During the open‐label third year of treatment, the mean Symptom Medication Score of active‐treated patients was further reduced despite an increased pollen count. Subcutaneous immunotherapy was well tolerated and consistent with the known safety profile. Conclusions and clinical relevance: Although the primary endpoint was not reached for the Full Analysis Set, a significant and clinically relevant effect on Symptom Medication Score was clearly demonstrated for the subgroup of patients in the north‐eastern region of Europe, where birch is the predominant tree species. Proving efficacy of birch allergen subcutaneous immunotherapy is challenging due to the numerous factors influencing birch pollen allergen exposure in field studies. [ABSTRACT FROM AUTHOR]

Summary: Background: The prevalence and predisposing factors of asymptomatic sensitization to Hymenoptera venom marker allergens are largely unknown. Objective: To evaluate sensitization profiles in a group of 490 dermatologic patients without a history of sting‐induced anaphylaxis. Methods: Clinical data were collected using a structured questionnaire; sera were tested for total IgE and specific IgE to venom preparations, recombinant venom marker allergens, inhalative allergens, and cross‐reactive carbohydrate determinants. Results: The lifetime prevalence of Hymenoptera stings was 85.3%. IgE rates exceeding cut‐off values of 0.35 kUA/L were 17.3% for rVes v 1, 18.0% for rVes v 5, and 3.5% for rApi m 1. Median specific/total IgE ratios for the above mentioned marker allergens were 0.05%, 0.02%, and 0.00%, respectively. Marker allergen‐directed sensitization was detectable in 85.5% of 138 Vespula venom‐reactive sera. Of 68 bee venom‐reactive participants, 23.5% were sensitized to rApi m 1 and 64.7% to any one or several of five commercially available bee venom allergens. Although double reactivity to bee and Vespula venom was clearly associated with sensitization to cross‐reactive carbohydrate determinants (P < 0.001), sensitization to marker allergens of both species was detectable in most double‐reactive sera (56.5%). Vespula venom marker allergen‐directed sensitization was associated with recent stings (P = 0.010), large local reactions (P = 0.009), total IgE elevation (P < 0.001), and sensitization to cross‐reactive carbohydrate determinants (P = 0.008). Conclusions and Clinical Relevance: The high sensitization rates observed in individuals without a history of sting‐induced anaphylaxis as well as total IgE levels and cross‐reactive carbohydrate determinant‐directed reactivity as potential confounders need to be considered in any interpretation of positive test results for Hymenoptera venom marker allergens. [ABSTRACT FROM AUTHOR]

Summary: Background: Screening for specific IgE against 2S albumin proteins Ara h 2 and 6 has good positive predictive value in diagnosing peanut allergy. From the third 2S member Ara h 7, 3 isoforms have been identified. Their allergenicity has not been elucidated. Objective: This study investigated the allergenicity of Ara h 7 isoforms compared to Ara h 2 and 6. Methods: Sensitization of 15 DBPCFC‐confirmed peanut‐allergic patients to recombinant Ara h 2.0201, Ara h 6.01 and isoforms of recombinant Ara h 7 was determined by IgE immunoblotting strips. A basophil activation test (BAT) was performed in 9 patients to determine IgE‐cross‐linking capacities of the allergens. Sensitivity to the allergens was tested in 5 patients who were sensitized to at least 1 Ara h 7 isoform, by a concentration range in the BAT. 3D prediction models and sequence alignments were used to visualize differences between isoforms and to predict allergenic epitope regions. Results: Sensitization to Ara h 7.0201 was most frequent (80%) and showed to be equally potent as Ara h 2.0201 and 6.01 in inducing basophil degranulation. Sensitization to Ara h 7.0201 together with Ara h 2.0201 and/or 6.01 was observed, indicating the presence of unique epitopes compared to the other 2 isoforms. Differences between the 3 Ara h 7 isoforms were observed in C‐terminal cysteine residues, pepsin and trypsin cleavage sites and 3 single amino acid substitutions. Conclusion & clinical relevance: The majority of peanut‐allergic patients are sensitized to isoform Ara h 7.0201, which is functionally as active as Ara h 2.0201 and 6.01. Unique epitopes are most likely located in the C‐terminus or an allergenic loop region which is a known allergenic epitope region for Ara h 2.0201 and 6.01. Due to its unique epitopes and allergenicity, it is an interesting candidate to improve the diagnostic accuracy for peanut allergy. [ABSTRACT FROM AUTHOR]

Abstract: Background: Impaired antiviral interferon expression may be involved in asthma exacerbations commonly caused by rhinovirus infections. Allergy is a known risk factor for viral‐induced asthma exacerbation, but little is known whether allergens may affect interferon responses. Objective: Our hypothesis is that house dust mite (HDM) impairs viral stimulus‐induced antiviral signalling. Methods: Experimental asthma exacerbations were produced in vitro in human bronchial epithelial cells (HBECs) and in mice using sequential challenges with HDM and a viral infection mimic, Poly(I:C). We examined rhinovirus pattern recognition receptors (PRRs) signalling pathways and potential mechanisms of impaired interferon response. Results: HBECs and mice exposed to HDM prior to Poly(I:C) exhibited a reduced antiviral response compared to Poly(I:C) alone, including reduced IFN‐β, IFN‐λ, TLR3, RIG‐I, MDA5, IRF‐3 and IRF‐7. Heat inactivation of HDM partially restored the TLR3‐induced interferon response in vitro and in vivo. Our HBEC‐data further showed that HDM directly affects TLR3 signalling by targeting the receptor glycosylation level. Conclusions: Direct effects of allergens such as HDM on PRRs can present as potential mechanism for defective antiviral airway responses. Accordingly, therapeutic measures targeting inhibitory effects of allergens on antiviral PRRs may find use as a strategy to boost antiviral response and ameliorate exacerbations in asthmatic patients. [ABSTRACT FROM AUTHOR]

Summary: Background: Fish is a well‐recognized cause of food allergy and anaphylaxis. The evolutionary and taxonomic diversity of the various consumed fish species pose a challenge in the identification and characterization of the major fish allergens critical for reliable diagnostics. Globally, fish is a rising cause of food allergy complicated by a large under‐investigated variety of species as well as increasing global tourism and trade. This is the first comprehensive study on allergen profiles of heat‐processed fish from Vietnam. Objective: The aim of this study was to identify the major heat‐stable allergens from frequently exported Asia‐Pacific freshwater and marine fish and to characterize the major allergen parvalbumin (PV) from one of the most consumed and exported fish species from Asia, the Indian mackerel (Rastrelliger kanagurta). Methods: Heated protein extracts from 33 fish species were separated by gel electrophoresis. PV isoforms were identified by immunoblotting utilizing 3 different PV‐specific monoclonal and polyclonal antibodies and further characterized by mass spectrometry. IgE reactivity was investigated using sera from 21 patients with confirmed fish allergy. Results: Heat‐stable IgE‐reactive PVs, with up to 5 isoforms per species, were identified in all 33 analysed fish species. In the Indian mackerel, 7 PV isoforms were identified by 2D‐gel electrophoresis combined with mass spectrometric analyses. The amino acid sequence deduced from cDNA of the most expressed isoform showed a high identity (>90%) to PVs from 2 other mackerel species. Conclusions and Clinical Relevance: Different PVs were identified as the major heat‐stable allergens in all 33 analysed freshwater and marine fish species from Vietnam, many of which are exported world‐wide and 21 species that have never been investigated before. The Indian mackerel PV represents a novel fish allergen, now officially registered as Ras k 1. Improved diagnostics for fish allergy against Asia‐Pacific species should be developed with focus on PV. [ABSTRACT FROM AUTHOR]

Abstract: Background: Art v 1, Amb a 4, and Par h 1 are allergenic defensin‐polyproline–linked proteins present in mugwort, ragweed, and feverfew pollen, respectively. We aimed to investigate the physicochemical and immunological features underlying the different allergenic capacities of those allergens. Methods: Recombinant defensin‐polyproline–linked proteins were expressed in E. coli and physicochemically characterized in detail regarding identity, secondary structure, and aggregation status. Allergenic activity was assessed by mediator releases assay, serum IgE reactivity, and IgE inhibition ELISA using sera of patients from Austria, Canada, and Korea. Endolysosomal protein degradation and T‐cell cross‐reactivity were studied in vitro. Results: Despite variations in the proline‐rich region, similar secondary structure elements were observed in the defensin‐like domains. Seventy‐four percent and 52% of the Austrian and Canadian patients reacted to all three allergens, while Korean patients were almost exclusively sensitized to Art v 1. This was reflected by IgE inhibition assays demonstrating high cross‐reactivity for Austrian, medium for Canadian, and low for Korean sera. In a subgroup of patients, IgE reactivity toward structurally altered Amb a 4 and Par h 1 was not changed suggesting involvement of linear epitopes. Immunologically relevant endolysosomal stability of the defensin‐like domain was limited to Art v 1 and no T‐cell cross‐reactivity with Art v 125‐36 was observed. Conclusions: Despite structural similarity, different IgE‐binding profiles and proteolytic processing impacted the allergenic capacity of defensin‐polyproline–linked molecules. Based on the fact that Amb a 4 demonstrated distinct IgE‐binding epitopes, we suggest inclusion in molecule‐based allergy diagnosis. [ABSTRACT FROM AUTHOR]

Background IgE reactivity to antigens from Gram-positive and Gram-negative bacteria is common in patients suffering from respiratory and skin manifestations of allergy, but the routes and mechanisms of sensitization are not fully understood. The analysis of the genome, transcriptome and microbiome of house dust mites ( HDM) has shown that Staphylococcus aureus ( S. aureus) and Escherichia coli ( E. coli) species are abundant bacteria within the HDM microbiome. Therefore, our aim was to investigate whether HDM are carriers of bacterial antigens leading to IgE sensitization in patients suffering from atopic dermatitis. Methods Plasma samples from patients with AD (n = 179) were analysed for IgE reactivity to a comprehensive panel of microarrayed HDM allergen molecules and to S. aureus and E. coli by IgE immunoblotting. Antibodies specific for S. aureus and E. coli antigens were tested for reactivity to nitrocellulose-blotted extract from purified HDM bodies, and the IgE-reactive antigens were detected by IgE immunoblot inhibition experiments. IgE antibodies directed to bacterial antigens in HDM were quantified by IgE Immuno CAP™ inhibition experiments. Results IgE reactivity to bacterial antigens was significantly more frequent in patients with AD sensitized to HDM than in AD patients without HDM sensitization. S. aureus and E. coli antigens were detected in immune-blotted HDM extract, and the presence of IgE-reactive antigens in HDM was demonstrated by qualitative and quantitative IgE inhibition experiments. Conclusion House dust mites ( HDM) may serve as carriers of bacteria responsible for the induction of IgE sensitization to microbial antigens. [ABSTRACT FROM AUTHOR]

Background: The clinical benefit of allergen‐specific immunotherapy (AIT) involves induction of blocking antibodies. It is not clear if these antibodies function via steric hindrance alone or a combination of levels, avidities, and epitope specificities, and clinical outcome cannot be predicted. We aim to in‐depth characterize serum antibody profiles during birch pollen AIT, investigate therapy‐induced antibodies for their capacity to block IgE binding to Bet v 1 and correlate data with clinical outcomes. Methods: Immune responses of five birch pollen allergic patients were monitored during the first year of AIT by nasal provocation tests (NPTs), ImmunoCAP, immunoblots, direct and avidity enzyme‐linked immunosorbent assays, mediator release assays, facilitated antigen binding (FAB) assays, and inhibition mediator release assays. Results: There was no correlation between NPT results and therapy‐induced changes in levels (IgE, IgG, IgA, IgM), avidities, or mediator release potency of Bet v 1‐specific antibodies. In FAB assays, blocking antibodies initiated upon AIT were shown to prevent formation of Bet v 1‐IgE complexes of an indicator serum pool and significantly correlated with clinical readout. Inhibition mediator release assays using patient‐specific IgE for passive sensitization revealed therapy‐induced blocking capacities with very good correlation to NPT results. Notably, this assay was the only one to detect a non‐responder during treatment in this pilot study. Conclusions: Clinical outcome of AIT depends on induction of blocking antibodies able to prevent the patient's own IgE from allergen binding. Monitoring of clinical efficacy seems to be best achieved using the inhibition mediator release assay, as development of relevant blocking antibodies can be verified in a patient‐tailored manner. [ABSTRACT FROM AUTHOR]

Background Arachidonic acid metabolites regulate several aspects of airway function including inflammation, muscle contraction and mucous secretion. Objective The aim of this study was to evaluate concentration of selected 5-lipoxygenase- and cyclooxygenase-derived eicosanoids in exhaled breath condensate (EBC) during allergen-induced bronchoconstriction. Methods The study was performed on 24 allergic rhinitis/asthma patients sensitized to a house dust mite (HDM) Dermatophagoides pteronyssinus (Dp) and 13 healthy controls (HCs). Bronchial challenge with Dp extract was performed only in the allergic patients. EBC samples were collected before ( T0) and during Dp-induced bronchoconstriction ( TEAR). Eicosanoid concentration was measured using HPLC-tandem mass spectrometry. Results Significant bronchoconstriction after Dp challenge was demonstrated in 15 patients (Rs), while in 9 patients (NRs) no asthmatic response could be detected. At T0 the most abundant eicosanoids in EBC of HDM-allergic patients were LTB4 and 5-oxo-ETE, while in HCs EBC concentration of LTB4 was significantly greater than that of 5-oxo-ETE. Allergen challenge resulted in significant increase in EBC concentration of 5-oxo-ETE, LTD4 and 8-iso-PGE2 only in Rs. At TEAR, the relative change of 5-oxo-ETE concentration in EBC correlated with decrease of peripheral blood eosinophilia ( R = −0.774; P = .0012). Moreover, the relative increase of 5-oxo-ETE in EBC at TEAR significantly correlated with the severity of the subsequent late asthmatic response ( R = 0.683, P = .007). Conclusion Our study demonstrates significant up-regulation of 5-oxo-ETE synthesis in HDM-allergic patients and indicates possible involvement of that mediator in the pathogenesis of allergic asthma. [ABSTRACT FROM AUTHOR]

Background Microarray-based component-resolved diagnostics (CRD) has become an accepted tool to detect allergen-specific IgE sensitization towards hundreds of allergens in parallel from one drop of serum. Nevertheless, specificity and sensitivity as well as a simultaneous detection of allergen-specific IgG4, as a potential parameter for tolerance development, remain to be optimized. Objective We applied the recently introduced silicon chip coated with a functional polymer named copoly(DMA-NAS-MAPS) to the simultaneous detection of food allergen-specific IgE and IgG4, and compared it with ImmunoCAP and ImmunoCAP ISAC. Inter- and intraslide variation, linearity of signal and working range, sensitivity and application of internal calibrations for IgE and IgG4 were assessed. Methods Native and recombinant allergenic proteins from hen's egg and cow's milk were spotted on silicon chips coated with copoly(DMA-NAS-MAPS) along with known concentrations for human IgE and IgG4. A serum pool and 105 patient samples were assessed quantitatively and semi-quantitatively with the ImmunoCAP and ImmunoCAP ISAC and correlated with IgE- and IgG4-specific fluorescence on silicon microarrays. Results Allergen-specific IgE and IgG4 were detected in parallel using two fluorescent dyes with no crosstalk. Results from the ImmunoCAP correlated better with microarray fluorescence than with ImmunoCAP ISAC except for the allergen ovomucoid. The working range of the silicon microarray for total hen's egg-specific IgE was comparable to the range of 0.1 to >100 kUA/L of the ImmunoCAP system, whereas for total cow's milk, the silicon microarray was less sensitive. Detectable allergen-specific IgG4 could be determined only for low concentrations, but still correlated positively with ImmunoCAP results. Conclusions We confirmed the ability of the polymer coated silicon microarray to be comparably sensitive to the ImmunoCAP ISAC for various food allergens. This suggests that the copoly(DMA-NAS-MAPS) microarray is a low-cost, self-producible alternative to the commercial ImmunoCAP ISAC in allergy research. [ABSTRACT FROM AUTHOR]

Background Microarray-based component-resolved diagnostics (CRD) has become an accepted tool to detect allergen-specific IgE sensitization towards hundreds of allergens in parallel from one drop of serum. Nevertheless, specificity and sensitivity as well as a simultaneous detection of allergen-specific IgG4, as a potential parameter for tolerance development, remain to be optimized. Objective We applied the recently introduced silicon chip coated with a functional polymer named copoly(DMA-NAS-MAPS) to the simultaneous detection of food allergen-specific IgE and IgG4, and compared it with ImmunoCAP and ImmunoCAP ISAC. Inter- and intraslide variation, linearity of signal and working range, sensitivity and application of internal calibrations for IgE and IgG4 were assessed. Methods Native and recombinant allergenic proteins from hen's egg and cow's milk were spotted on silicon chips coated with copoly(DMA-NAS-MAPS) along with known concentrations for human IgE and IgG4. A serum pool and 105 patient samples were assessed quantitatively and semi-quantitatively with the ImmunoCAP and ImmunoCAP ISAC and correlated with IgE- and IgG4-specific fluorescence on silicon microarrays. Results Allergen-specific IgE and IgG4 were detected in parallel using two fluorescent dyes with no crosstalk. Results from the ImmunoCAP correlated better with microarray fluorescence than with ImmunoCAP ISAC except for the allergen ovomucoid. The working range of the silicon microarray for total hen's egg-specific IgE was comparable to the range of 0.1 to >100 kUA/L of the ImmunoCAP system, whereas for total cow's milk, the silicon microarray was less sensitive. Detectable allergen-specific IgG4 could be determined only for low concentrations, but still correlated positively with ImmunoCAP results. Conclusions We confirmed the ability of the polymer coated silicon microarray to be comparably sensitive to the ImmunoCAP ISAC for various food allergens. This suggests that the copoly(DMA-NAS-MAPS) microarray is a low-cost, self-producible alternative to the commercial ImmunoCAP ISAC in allergy research. [ABSTRACT FROM AUTHOR]

Background IgG to galactose-α-1,3-galactose (α-gal) are highly abundant natural antibodies (Ab) in humans. α-Gal-specific IgE Ab cause a special form of meat allergy characterized by severe systemic reactions 3-7 h after consumption of red meat. We investigated 20 patients who experienced such reactions and characterized their α-gal-specific IgE and IgG responses in more detail. Methods α-Gal-specific IgE was determined by Immuno CAP. IgE reactivity to meat extract and bovine gamma globulin ( BGG) was assessed by immunoblotting and ELISA, respectively. In some experiments, sera were pre-incubated with α-gal or protein G to deplete IgG Ab. α-Gal-specific IgG1-4 Ab in individuals with and without meat allergy were assessed by ELISA. Results In immunoblots, BGG was the most frequently recognized meat protein. Binding of IgE and IgG to BGG was confirmed by ELISA and completely abolished after pre-incubation with α-gal. Neither the depletion of autologous α-gal-specific IgG Ab nor the addition of α-gal-specific IgG Ab from nonallergic individuals changed the IgE recognition of BGG of meat-allergic patients. Meat-allergic patients showed significantly higher α-gal-specific IgG1 and IgG3 Ab than nonallergic individuals, whereas the latter showed significantly higher levels of α-gal-specific IgG4 Ab. Conclusion Patients with delayed meat allergy display IgE and IgG Ab that selectively recognize the α-gal epitope on BGG. Their enhanced α-gal-specific IgE levels are accompanied by high levels of α-gal-specific IgG1 devoid of IgE-blocking activity. This subclass distribution is atypical for food allergies and distinct from natural α-gal IgG responses in nonallergic individuals. [ABSTRACT FROM AUTHOR]

Background There is limited understanding of how maternal diet affects breastmilk food allergen concentrations, and whether exposure to allergens through this route influences the development of infant oral tolerance or sensitization. Objective To investigate how maternal dietary egg ingestion during early lactation influences egg protein (ovalbumin) levels detected in human breastmilk. Methods In a randomized controlled trial, women were allocated to a dietary group for the first six weeks of lactation: high-egg diet (> 4 eggs per week), low-egg diet (one-three eggs per week) or an egg-free diet. Breastmilk samples were collected at 2, 4 and 6 weeks of lactation for the measurement of ovalbumin. The permeability of the mammary epithelium was assessed by measuring the breastmilk sodium : potassium ratio. Egg-specific IgE and IgG4 were measured in infant plasma at 6 weeks, and prior to the introduction of egg in solids at 16 weeks. Results Average maternal egg ingestion was associated with breastmilk ovalbumin concentration. Specifically, for each additional egg ingested per week, there was an average 25% increase in ovalbumin concentration (95% CI: 5-48%, P = 0.01). Breastmilk ovalbumin concentrations were significantly higher in the 'high-egg' group (> 4 eggs per week) compared with the 'egg-free' group ( P = 0.04). However, one-third of women had no breastmilk ovalbumin detected. No detectable associations were found between mammary epithelium permeability and breastmilk ovalbumin concentrations. Infant plasma egg-specific IgG4 levels were also positively associated with maternal egg ingestion, with an average 22% (95% CI: 3-45%) increase in infant egg-specific IgG4 levels per additional egg consumed per week ( P = 0.02). Conclusions and Clinical Relevance Increased maternal egg ingestion is associated with increased breastmilk ovalbumin, and markers of immune tolerance in infants. These results highlight the potential for maternal diet to benefit infant oral tolerance development during lactation. [ABSTRACT FROM AUTHOR]

Background Birch pollen-related soya allergy is mediated by Gly m 4. Conformational IgE epitopes of Gly m 4 are unknown. Objective To identify the IgE epitope profile of Gly m 4 in subjects with birch pollen-related soya allergy utilizing an epitope library presented by Gly m 4-type model proteins. Methods Sera from patients with ( n = 26) and without ( n = 19) allergy to soya as determined by oral provocation tests were studied. Specific IgE (Bet v 1/Gly m 4) was determined by Immuno CAP. A library of 59 non-allergenic Gly m 4-type model proteins harbouring individual and multiple putative epitopes for IgE was tested in IgE binding assays. Primary, secondary and tertiary protein structures were assessed by mass spectrometry, circular dichroism and nuclear magnetic resonance spectroscopy. Results All subjects were sensitized to Gly m 4 and Bet v 1. Allergen-specific serum IgE levels ranged from 0.94 to > 100 kUA/L. The avidities of serum IgE were 5.06 ng (allergic) and 1.8 ng (tolerant) as determined by EC50 for IgE binding to Gly m 4. 96% (46/48) of the protein variants bound IgE. Model proteins had Gly m 4-type conformation and individual IgE binding clustered in six major surface areas. Gly m 4-specific IgE binding could be inhibited to up to 80% by model proteins harbouring individual IgE binding sites in an epitope-wise equimolar fashion. Receiver operating curve analysis revealed an area under fitted curve of up to 0.88 for model proteins and 0.66 for Gly m 4. Conclusion and Clinical Relevance Serum levels and avidity of Gly m 4-specific IgE do not correlate with clinical reactivity to soya. Six IgE-binding areas, represented by 23 amino acids, account for more than 80% of total IgE binding capacity of Gly m 4. Model proteins may be used for epitope-resolved diagnosis to differentiate birch-soya allergy from clinical tolerance. [ABSTRACT FROM AUTHOR]

The author comments on the mapping of conformational immunoglobulin E (IgE) epitopes of food allergens. He discusses the side effects of food allergies, the aim of grafting of surface areas of allergens, X-ray crystallography of an allergen-antibody complex, and technologies for the mapping of conformational IgE epitopes.

Background Personal exposure to fungal bioaerosols derived from contaminated building materials or agricultural commodities may induce or exacerbate a variety of adverse health effects. The genomic mechanisms that underlie pulmonary immune responses to fungal bioaerosols have remained unclear. Objective The impact of fungal viability on the pulmonary micro RNA and messenger RNA profiles that regulate murine immune responses was evaluated following subchronic inhalation exposure to Aspergillus fumigatus conidia. Methods Three groups of naïve B6C3F1/N mice were exposed via nose-only inhalation to A. fumigatus viable conidia, heat-inactivated conidia (HIC), or HEPA-filtered air twice a week for 13 weeks. Total RNA was isolated from whole lung 24 and 48 h postfinal exposure and was further processed for gene expression and micro RNA array analysis. The molecular network pathways between viable and HIC groups were evaluated. Results Comparison of data sets revealed increased Il4, Il13 and Il33 expression in mice exposed to viable vs. HIC. Of 415 micro RNAs detected, approximately 50% were altered in mice exposed to viable vs. HIC 48 h postexposure. Significantly down-regulated ( P ≤ 0.05) miR-29a-3p was predicted to regulate TGF-β 3 and Clec7a, genes involved in innate responses to viable A. fumigatus. Also significantly down-regulated ( P ≤ 0.05), miR-23b-3p regulates genes involved in pulmonary IL-13 and IL-33 responses and SMAD2, downstream of TGF-β signalling. Using Ingenuity Pathway Analysis, a novel interaction was identified between viable conidia and SMAD2/3. Conclusions and Clinical Relevance Examination of the pulmonary genetic profiles revealed differentially expressed genes and micro RNAs following subchronic inhalation exposure to A. fumigatus. Micro RNAs regulating genes involved in the pulmonary immune responses were those with the greatest fold change. Specifically, germinating A. fumigatus conidia were associated with Clec7a and were predicted to interact with Il13 and Il33. Furthermore, altered micro RNAs may serve as potential biomarkers to evaluate fungal exposure. [ABSTRACT FROM AUTHOR]

Background: House dust mite (HDM) acts on the airway epithelium to induce airway inflammation in asthma. We previously showed that the ability of HDM to induce allergic sensitization in mice is related to airway epithelial CCL20 secretion. Objective: As a disintegrin and metalloprotease (ADAM)s have been implicated in chemokine shedding, we sought to determine their involvement in HDMinduced release of chemokines, including CCL20, by airway epithelial cells. Methods: We studied the effects of pharmacological ADAM inhibitors as well as ADAM10 and ADAM17 siRNA downregulation on chemokine release using (multiplex) ELISA in supernatants from HDM-exposed human bronchial epithelial 16HBE cells and primary normal human bronchial epithelial cells (NHBE) at 4-24 h. Results: House dust) mite markedly increased CCL20 levels in both 16HBE and NHBE cells (16-24 h). In 16HBE cells, the HDM-induced increase was observed as early as 4 h upon exposure and the use of specific inhibitors indicated the involvement of ADAM10/17-mediated shedding. siRNA knockdown of ADAM10, but not of ADAM17, significantly reduced the HDM-induced release of CCL20 in both 16HBE and NHBE cells. A similar effect was observed for HDM-induced CCL2, CCL5, and CXCL8 release in NHBE cells. The HDMinduced increase in CCL20 levels was not affected by protein synthesis inhibitor cycloheximide nor protein transport inhibitor monensin, indicating that HDM induces surface shedding of chemokines. Conclusion: Our data show for the first time that ADAM10 activity contributes to HDM-induced shedding of chemokines, including CCL20. The ADAM10/ CCL20 axis may be a target for novel therapeutic strategies in asthma. [ABSTRACT FROM AUTHOR]

Allergic bronchopulmonary aspergillosis ( ABPA) is characterized by an allergic immunological response to Aspergillus fumigatus. In this study, we investigated whether certain Aspergillus antigens are more allergenic than others, as was postulated previously. We stimulated peripheral blood mononuclear cells from patients with ABPA with the classically described A. fumigatus allergens Aspf1, Aspf2, Aspf3, and Aspf4, as well as two other Aspergillus antigens, Crf1 and Catalase1. Activated CD4+ T cells displayed a T helper 2 phenotype with the production of IL-4 in response to stimulation with several of these different antigens. Immune responses were not limited to the classically described A. fumigatus allergens. In healthy individuals, we demonstrated a similar recognition profile to the different antigens, but in contrast the activated CD4+ T cells exerted a T helper 1 phenotype and mainly produced IFN-γ after stimulation with A. fumigatus antigens. In conclusion, irrespective of the A. fumigatus antigen, the T-cell immune response in patients with ABPA is skewed to a T helper 2 cytokine secretion profile. [ABSTRACT FROM AUTHOR]

Background Exposure to indoor allergens during early life may play a role in the development of the immune system and inception of asthma. Objective To describe the house dust mite ( HDM) allergen concentrations in bedroom dust during early life and to evaluate its associations with HDM sensitization, wheezing, and asthma, from birth to school age, in 5 geographically spread European birth cohorts. Methods We included 4334 children from INMA- Menorca ( Spain), BAMSE ( Sweden), LISAplus and MAS ( Germany), and PIAMA- NHS (the Netherlands). Dust samples were collected from bedrooms during early life and analyzed for Dermatophagoides pteronyssinus ( Der p1) and Dermatophagoides farinae ( Der f1). HDM concentrations were divided into four categories. Sensitization was determined by specific Ig E. Wheezing and asthma information up to 8/10 years was collected through questionnaires. We performed mixed-effects logistic regression models and expressed associations as odds ratios with 95% confidence intervals. Results House dust mite concentrations varied across cohorts. Mean allergen concentrations were highest in INMA-Menorca (geometric mean ( GM) Der p1 = 3.3 μg/g) and LISAplus ( GM Der f1 = 2.1 μg/g) and lowest in BAMSE ( GM Der p1 = 0.1 μg/g, Der f1 = 0.3 μg/g). Moderate and high HDM concentrations were significantly ( P-values < 0.05) associated with 50-90% higher prevalence of HDM sensitization. No significant associations were observed with respiratory outcomes. Conclusion Our study based on geographically spread regions, a large sample size, and a wide range of allergen concentration shows that HDM allergen concentrations vary across regions and that exposure during early life plays a role in the development of allergic sensitization but not in the development of respiratory outcomes. [ABSTRACT FROM AUTHOR]