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The importance of apple allergens, in particular Mal d 1, a Bet v 1 homologue for the pollen-fruit syndrome in Northern Europe, and Mal d 3, responsible for true fruit allergy in Southern Europe, has been repeatedly emphasized. However, little is known about the distribution pattern of major allergens in fruits and whether differences exist among different cultivars. Transcript expression of Mal d 1 isoforms and Mal d 3 was examined by RealTime-PCR and Northern analysis, respectively. An immuno-tissue-print (ITP) assay was developed to localise major allergens in apple fruit tissue and a Mal d 1 specific, patient independent ELISA was established. ITP analyses show that Mal d 1 and Mal d 2 are distributed throughout the apple pulp and peel, while Mal d 3 is restricted to the peel. Data obtained by ELISA reveal a variation of Mal d 1 content ranging from 0.84 to 33.17 μg/g fresh weight in 39 selected cultivars. Different apple cultivars show a markedly different expression of major allergens; this finding will influence the development of diagnostic tools as well as the dietary management of allergic individuals.

Background Lipid transfer proteins (LTPs) are clinically relevant plant food panallergens and have been proposed as ideal tools to study true food allergy. Pru p 3, the major peach allergen in the Mediterranean area, is among the best-characterized allergenic members of the LTP family. Its diagnostic value for Rosaceae fruit allergy has been demonstrated both in vivo and in vitro. Objective We sought to locate major IgE-binding epitopes of Pru p 3. Methods A serum pool and individual sera from patients with peach allergy and positive skin prick test results to Pru p 3 were used. Three-dimensional modeling was achieved by using experimentally available structures of Pru p 3 homologues as templates. Theoretical prediction of potential IgE-binding regions was performed by selecting specific residues on the molecular surface displaying prominent electrostatic potential features. Point mutants of Pru p 3 were constructed by standard polymerase chain reaction procedures with the appropriate primers. Mutants were expressed in P pastoris by means of the pPIC 9 vector and purified from the corresponding supernatants by gel-filtration chromatography followed by RP-HPLC. IgE binding by Pru p 3 mutants was tested by immu-nodetection and quantified by ELISA and ELISA inhibition assays. Synthetic peptides (10 mer; 5 amino acids overlapping) covering the full Pru p 3 sequence were used to detect IgE epitopes by 125 I–anti-IgE immunodetection. Results Pru p 3 showed a 3-dimensional structure comprising 4 α-helixes and a nonstructured C-terminal coil (residues 73 to 91). Regions around amino acids in positions 23 to 36, 39 to 44, and 80 to 91, particularly residues R39, T40, and R44, K80 and K91, were predicted as potential antibody recognition sites according to their relevant surface and electrostatic properties. Point mutants K80A and K91A were found to have an IgE-binding capacity similar to that of recombinant Pru p 3, but the triple mutant R39A/T40A/R44A showed a substantial decrease (approximately 5 times) of IgE binding. IgE immu-nodetection of synthetic peptides led to the identification of Pru p 3 sequence regions 11 to 25, 31 to 45, and 71 to 80 as major IgE epitopes. Conclusions Main IgE-binding regions of the Pru p 3 amino acid sequence were identified. The three major ones comprised the end of an α-helix and some residues of the following interhelix loop. These data can help to search for Pru p 3 hypoallergenic forms.

Two cDNA clones (named Pp-LTP1 and Pp-LTP2 ) corresponding to different lipid transfer protein (LTP) genes have been isolated from peach ( Prunus persica L. Batsch) epicarp and ovary, respectively. Sequence analysis revealed that the two fragments share 54% identity at nucleotide level and show common features of plant LTP genes, such as conserved cysteine residues and lipid-binding motifs. Phylogenetic analysis grouped Pp-LTP1 and Pp-LTP2 in two distinct clusters, the former with most of LTP genes sequenced in the Rosaceae family, the latter only with one almond LTP. Genomic Southern data indicated that a small LTP gene family is present in peach. Pp-LTP1 and Pp-LTP2 have been used as gene-specific probes to describe expression in flowers and fruits throughout development. In petals, sepals and stamen only Pp-LTP1 was expressed whereas transcripts of Pp-LTP2 strongly accumulated in non-pollinated and pollinated ovary with a decreasing trend in the period of four weeks after pollination. In fruits, a dramatic accumulation of Pp-LTP1 mRNA was detected in epicarp at all stages of fruit development and, with the exception of the early growth stage, no Pp-LTP1 transcripts have been detected in mesocarp. When Pp-LTP2 was used as a probe in the same fruit tissues, a faint hybridisation signal was observed only in epicarp of fruitlets collected at an early growth stage. Infection with Monilia induced only a slight increase of Pp-LTP1 transcript in epicarp of pre-climacteric and climacteric fruits. These results support the hypothesis of multiple roles played by LTPs and, considering that LTPs have been recognised as the major allergen of peach, indicate that Pp-LTP1 could be related to the allergenicity of peach.

Background: Food allergy (FA) is an important health problem for which epidemiologic studies are needed. Objective: We performed an epidemiologic survey in France to determine the prevalence, clinical pictures, allergens, and risk factors of FA. Methods: This study was conducted on 33,110 persons who answered a questionnaire addressed to a representative sample of the French population on a scale of 1:1000 (44,000 subjects aged ≤60 years). One thousand one hundred twenty-nine persons with FA selected during phase 1 received a second questionnaire. Results: The reported prevalence of FA is 3.52%: 3.24% evolutionary FA; 0.12% asymptomatic cases thanks to eviction diets; and 0.17% cured FA. The subjects were characterized by overrepresentation of city dwellers (80% vs 76%), women (63% vs 50%), and health care personnel (11% vs 4%). Fifty-seven percent (vs 17%) presented with atopic diseases (P < .01). FA was often persistent, lasting more than 7 years in 91% of the adults. The most frequent allergens were 14% Rosaceae, 9% vegetables, 8% milk, 8% crustaceans, 5% fruit cross-reacting with latex, 4% egg, 3% tree nuts, and 1% peanut. Sensitization to pollen was significantly correlated with angioedema, asthma, rhinitis, and fruit allergy (P < .01). FA was 4 times more frequent in patients with latex allergy. The main manifestations of FA were atopic dermatitis for subjects under 6 years of age, asthma for subjects between 4 and 6 years of age, and anaphylactic shock in adults over 30 years of age (P < .007). Shocks were correlated with alcohol or nonsteroidal anti-inflammatory drug intake (P < .01 and P < .04, respectively). Conclusion: The prevalence of FA is estimated at 3.24% (range, 3.04% to 3.44%) in France. This study emphasizes the increasing risk of FA in well-developed countries and draws attention to certain FA risk factors, such as the intake of drugs (nonsteroidal anti-inflammatory drugs, β-blockers, and angiotensin-converting enzyme inhibitors) or alcohol, intolerance of latex gloves, and socioprofessional status. (J Allergy Clin Immunol 2001;108:133-40.)

Background: Few studies have evaluated IgE-mediated hypersensitivity to melon with details of clinical reactions confirmed by double-blind, placebo-controlled, food challenges (DBPCFCs). Objective: We sought to investigate clinical features (type and severity of reactions, age at onset, results of skin prick and in vitro tests, and incidence of other allergic diseases and associated food allergies) of acute allergic reactions to melon confirmed by DBPCFCs. Methods: Fifty-three consecutive adult patients complaining of adverse reactions to melon were included in the study. Skin prick tests and detection of specific IgE were performed in all patients with melon, avocado, kiwi, banana, chestnut, latex, pollen, and other offending foods. Patients first underwent an open food challenge, unless they had a convincing history of severe anaphylaxis. Positive open food challenge reactions were subsequently evaluated by DBPCFCs. Results: Actual clinical reactivity was confirmed in 19 (36%) of 53 patients. The most frequent symptom was oral allergy syndrome (n = 14), but two patients experienced life-threatening reactions, including respiratory symptoms and hypotension. The positive predictive value for a skin prick test was 42%, and that for specific IgE measurement was 44%. Forty-five reactions to 15 other foods were confirmed in 18 patients. The most common foods associated with melon allergy were avocado (n = 7), banana (n = 7), kiwi (n = 6), watermelon (n = 6), and peach (n = 5). Onset of melon-induced allergic symptoms occurred from 6 to 45 years (median, 20 years), preceded by seasonal rhinitis, asthma, or both in 88% (15/17). Conclusion: About one third of reported reactions to melon are confirmed by means of DBPCFC, which has been proven to be the most reliable procedure in the diagnosis of clinical fruit allergy. Isolated melon allergy is rare, with most patients either having allergic rhinitis, asthma, or both and associated food allergies. (J Allergy Clin Immunol 2000;106:968-72.)

Background: Fruit allergy has been attributed to cross-reactive IgE to pollens and has been associated with a particular pollen sensitization. Objective: The aim of the study was to evaluate sensitization to several taxonomically unrelated pollens in peach- and pollen-allergic patients and to study cross-reactivity between them. Methods: One hundred sixty-five patients were evaluated: 70 peach- allergic patients together with 95 pollen-allergic patients (control group). Pollen skin tests in duplicate were performed to 5 grasses, 8 trees, and 7 weeds. Cross-reactivity between peach and taxonomically diverse pollens was determined by radioallergosorbent inhibition and Western blot inhibition tests. Experiments were also carried out after preadsorption of the sera with purified natural profilin. Results: The skin test results revealed that peach-allergic patients frequently reacted to most pollens—grasses, weeds, and trees—even when some of these are not found in our geographic area. There was a statistically significant increase in sensitization frequency to most trees and weeds, with a statistically higher occurrence of asthma (odds ratio 2.98, 95% confidence interval 1.46-6.09). Inhibition test results provided evidence that taxonomically unrelated grasses, weeds, and trees produced various and substantial degrees of inhibition in specific IgE to peach and that the peach extract elicited strong inhibitions to those pollens. Profilin was found to be a relevant cross-reactive antigen in these patients. Conclusion: The results of this study provide evidence that peach allergy is linked to sensitization to several taxonomically unrelated pollens. This is attributable to the ubiquitous nature of the IgE binding determinants—such as profilins—between peach and taxonomically unrelated pollens. (J Allergy Clin Immunol 1999;104:688-94.)

Background: Allergy to fresh fruits and vegetables is mostly observed in subjects with pollinosis, especially from birch, because of cross-reacting allergens in vegetable foods and pollens. However, allergic reactions to fruits, specifically Rosaceae fruits, have been reported in subjects without pollinosis. Objective: This study evaluated the pattern of IgE reactivity, identifying the allergen responsible in 2 groups of patients with oral allergy syndrome to peach with or without birch pollinosis. Methods: The allergenic components of peach were detected by SDS-PAGE and immunoblotting. The major peach allergen was purified by HPLC with a cation-exchange column followed by gel filtration chromatography. Its IgE-binding capacity and its homology with the protein of the crude extract were demonstrated by immunoblotting inhibition techniques. To better characterize this allergen, periodic acid-Schiff stain and isoelectrofocusing were used. The amino acid sequencing was done with a gas-phase sequencer. Results: SDS-PAGE and immunoblotting of the 15 patients allergic to peach, 8 without and 7 with birch pollinosis, showed that they all recognized a protein with a molecular weight of 9 kd. This was the only allergen recognized by patients not sensitized to pollen, whereas the birch pollen–sensitive patients had IgE binding to other allergenic proteins at higher molecular weights. The purified 9-kd protein retained its IgE-binding capacity, was negative to periodic acid-Schiff stain, and had an isoelectric point value of greater than 9. A search in the Swiss Prot Bank showed this was a lipid transfer protein, belonging to a group of molecules involved in the defensive system of plants. Conclusions: The major allergen of peach is a 9-kd protein belonging to the group of lipid transfer proteins. This is the only allergen recognized by patients allergic to peach but not sensitized to birch pollen. (J Allergy Clin Immunol 1999;103:520-6.)

Background: Allergy to apple and Prunus fruits is frequently associated with birch pollinosis, with the principal cross-reacting allergens involved being members of the Bet v 1 family. However, a major 13-kd component, nonimmunologically related to Bet v 1, has been implicated as allergen in patients allergic to Prunoideae fruit but not to birch pollen. Objective: We sought to isolate and characterize the 13-kd allergen present in apple and peach. Methods: Sera from patients allergic to both fruits were selected on the basis of clinical symptoms, skin prick tests responses, and specific IgE levels. Allergens were purified by reverse-phase HPLC and characterized by N-terminal amino acid sequencing, MALDI analysis, specific IgE immunodetection, and immunoblot inhibition assays. Results: A 13-kd protein band was recognized in crude apple and peach extracts by 9 of 10 and 10 of 10 sera from patients allergic to fruit, respectively. The isolation and characterization of the corresponding allergens allowed their identification as lipid-transfer proteins, with a molecular mass of 9058 d for the apple protein and 9138 d for the peach protein. Both purified allergens were recognized by sera from patients allergic to fruit and fully inhibited the IgE binding by the 13-kd component present in the 2 crude fruit extracts. Conclusion: Lipid-transfer proteins are relevant apple and peach allergens and, considering their ubiquitous distribution in tissues of many plant species, could be a novel type of panallergen of fruits and vegetables. (J Allergy Clin Immunol 1999;103:514-9.)

Rationale Ornamental Ficus benjamina (FB) is a new indoor allergen potentially associated with hypersensitivity to fig fruit (Ficus carica). We evaluated the prevalence of sensitization and studied the cross-reactivity with other fruits and with Hevea latex. Methods A prick test solution from FB latex was tested in 4930 consecutive patients with suspect inhalant allergy. Patients positive to FB were further skin tested with exotic fruits by the prick-to-prick method. RAST inhibition using fig fruit as solid phase was performed in 11 patients with clinical fruit allergy. Results Among 2662 patients reacting to any aeroallergen, 66 (2.5%) reacted to FB. Ten patients showed isolated sensitization. Only 10.6% of FB-positive subjects were also positive to Hevea latex. Sensitization to FB was specifically associated with positive skin tests to fresh fig (83%), dried fig (37%), kiwi (28%), papaya (22%), avocado (19%), banana (15%), and pineapple (10%) (n=54). Clinical reactions were reported mainly from fresh and dried fig and kiwi (47, 60, and 64% of skin test-positive patients). Fig fruit RAST was inhibited 87% by FB, 89% by fresh fig, 80% by dried fig, 38% by kiwi, and 59 and 44% by the thiolproteases ficin and papain, respectively. No inhibition was obtained with Hevea latex and rHev b 6.02 (hevein). Conclusions Sensitization to FB latex is found in 2.5% of atopics and is largely independent of Hevea latex allergy. Sensitization is commonly associated with allergy to figs and other tropical fruits (“Ficus-fruit syndrome”). This cross-reactivity is mediated, at least in part, by thiolproteases but not by hevein-like allergens.