Allergic sensitization to Mal d 1 without detectable specific serum IgE

To the Editor, In clinical practice it is sometimes observed that allergic patients exhibit allergic symptoms or positive skin prick test results in the absence of detectable allergenspecific IgE in serum. One example for such a condition is local allergic rhinitis (LAR) where it is hypothesized that IgE is produced only locally but does not reach the systemic circulation.1 In this study, 25 subjects, age from 13 to 59 years [Mean 32.6 ± 13.6], 16 males and nine females, with birch pollenrelated allergic rhinoconjunctivitis (ARC) with or without allergic asthma (AA), with (group 1: n = 13) or without crossreactive food allergy to apple (group 2: n = 12) (oral allergy syndrome, OAS) were enrolled (Table S1) and investigated at three time points (i.e., before the birch pollen seasontime point 1, shortly after the birch pollen seasontime point 2 and thereafter in autumntime point 3) (Figure 1). The diagnosis of birch pollen allergy and OAS, demographic, serological and clinical features of the patients are described in the Supplemental materials (Table S1). Two patients from group 1 and group 2, (patients 1 and 2) stood out. They contained low but clearly positive Bet v 1specific IgE levels before the birch pollen season (time point 1, patient 1: 0.16 kUA/L; patient 2: 0.11 kUA/L), showed increases of Bet v 1specific IgE shortly after the birch pollen season (time point 2, patient 1: 0.75 kUA/L, patient 2: 0.53 kUA/L), which then declined again until September at time point 3 (patient 1: 0.53 kUA/L; patient 2: 0.43 kUA/L) (Tables S2 and S3, Figures 2 and 3). Interestingly, Mal d 1specific IgE could not be detected in the sera obtained from patients 1 and 2 at any of the three time points even by sensitive and quantitative ImmunoCAP technology (Table S2; Figures 2 and 3). The two patients were young females who suffered from birch pollenrelated allergic rhinoconjunctivitis and one (patient 1) suffered also from birch polleninduced asthma (Table S4). Importantly, one patient suffered from OAS upon consumption of apples, especially after the birch pollen season and was positive in open food challenge with apple (Table S4). The two patients were sensitized only to few other allergens besides Bet v 1 and Mal d 1 and sensitization to other crossreactive food allergens (e.g., profilin, lipid transfer proteins) could be excluded (Table S5). An increase of Bet v 1specific IgE levels was observed in both patients 1 and 2 at time point 2 shortly after the birch pollen season (Figures 2 and 3, Table S2), which was associated with a strong increase in Bet v 1specific basophil sensitivity in patient 2 (Figure 3, Table S6). The Bet v 1specific basophil sensitivity in patient 1 at time points 1 and 2 was comparable whereas at time point 3 patient 1 showed basophil activation with a tenfold lower Bet v 1 concentration indicating increased Bet v 1specific basophil sensitivity at time point 3 (Figure 2, Table S6). Exposure of basophils to antiIgE antibodies indicated strongly reduced IgEmediated basophil sensitivity in patient 1 at time point 3 as compared to time points 1 and 2 (Table S6). However, antiIgEinduced basophil activation cannot be compared with allergeninduced basophil activation because of the different molarity of the crosslinkers and the different modes of crosslinking (i.e., crosslinking of IgE constant regions versus different IgE epitopes on allergens). Importantly, antiIgE will induce not only crosslinking of Bet v 1 and/or Mal d 1specific IgE also but of IgE antibodies with different other specificities. We could not detect at any of the three time points Mal d 1specific IgE in the sera of patients 1 and 2 (Table S2, Figures 2 and 3) but we found a strongly increased Mal d 1specific basophil sensitivity in both patients at time points 2 and 3 (Figures 2 and 3, Table S6) after the birch pollen season indicating increases of Mal d 1specific IgE on the patients basophils after birch pollen exposure. At time point 2 it appeared that Mal d 1 activated more basophils than Bet v 1 although the latter is the genuinely sensitizing allergen. This may be explained by the interplay of IgE antibodies with different affinities, different numbers of IgE epitopes and epitope proximity.2– 4 The increase of Mal d 1specific basophil sensitivity was associated with marked increased appleinduced skin responses at time points 2 and 3 after the birch pollen season as compared to time point 1 before the birch pollen season (Figures 2 and 3, Table S7). By contrast, skin responses to histamine were comparable at all three time points in patients 1 and 2 (Table S7). It is of note that the cumulative allergenspecific IgE levels calculated according to ImmunoCAP ISAC results in patient 1 and patients 2 were the lowest (i.e., patient 1: 11.08 ISU; patient 2: 12.49 ISU) among all patients analyzed (Table S8) because it may explain the high activity of the low levels of cellbound Mal d 1specific IgE. Using purified allergen molecules, our study has demonstrated that basophils and mast cells of allergic patients contain already allergenspecific IgE before it can be detected in serum in the form