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Background Tropomyosin is the most studied shellfish allergen and has been involved in cross-reactivity among different invertebrates (crustacean, mollusks, mites, insects, and nematodes). Objective To determine the relevance of tropomyosin in mite- and shellfish-sensitized patients using tropomyosin skin testing. Methods Patients were divided into 3 groups: group M included mite allergic patients (ie, individuals with respiratory symptoms and a positive result on skin prick testing [SPT] to house dust mites), group S included shellfish allergic patients (ie, individuals who reported symptoms with shellfish), and group MS included mite- and shellfish allergic patients (ie, individuals who simultaneously fulfilled the inclusion criteria for groups M and S). Tropomyosin was purified from shrimp, characterized, and used in SPT for diagnosis in the patient population. Results Eight hundred fifty patients were included in the study: 790 (92.9%) in group M, 21 (2.5%) in group S, and 39 (4.6%) in group MS. Tropomyosin was purified from shrimp with a purity higher than 95%. Forty-two individuals tested positive to tropomyosin: the prevalence was 2.7% in group M, 28.6% in group S, and 38.5% in patients of group MS. Twenty-one (50%) of the tropomyosin-positive individuals had symptoms with shellfish, and 3 (14.3%) reported anaphylaxis. Conclusion The prevalence of tropomyosin was low in mite-sensitized patients (2.7 %) and high in shellfish allergic patients (28.6%). The higher prevalence of tropomyosin was found in patients sensitized to both mite and shellfish (38.5%). The selection of tropomyosin-sensitized patients by SPT might help in the choice of appropriate treatments or management for these patients.

Goji berries (GB) have been introduced in Western diet. Preliminary reports have demonstrated its allergenic capacity. The objectives of the study were to investigate the frequency of sensitisation and the allergens involved. 566 individuals, with respiratory or cutaneous symptoms were skin-prick tested with GB extract. Thirty three were positive (5.8%). 94% were sensitised to other allergens. Specific IgE to GB, peach, tomato and nut-mix was measured. Thirteen individuals from 24 available sera (54.2%) had positive specific IgE. 92.3% of GB positive patients were positive to peach. Seven individuals recognised 8 bands and six recognised a 7kDa band. This band was identified as a LTP by MS/MS. Cross-reactivity was demonstrated with tomato, tobacco, nutmix, Artemisia pollen and purified Lyc e 3 and Pru p 3. GB are a new allergenic source with high prevalence of sensitisation. LTP seems to be the major allergen involved in sensitisation and cross-reactivity.

Background: Although allergy to Cannabis sativa was first reported over 40 years ago, the allergenicity has scarcely been studied. The objectives of this study were to investigate the frequency of sensitization to this plant, to analyze the clinical characteristics and allergenic profile of sensitized individuals and to identify the allergens involved. Methods: Five hundred and forty-five individuals in Spain attending allergy clinics with respiratory or cutaneous symptoms underwent a skin-prick test (SPT) with C. sativa leaf extract. The extract was characterized by SDS-PAGE and 2-dimensional electrophoresis. Specific IgE to C. sativa was measured in positive SPT individuals. The clinical and allergenic profiles of sensitized individuals were investigated and the most-recognized allergens sequenced and characterized by liquid chromatography-mass spectrometry/mass spectrometry. Results: Of this preselected population, 44 individuals had positive SPT to C. sativa (prevalence 8.1%). Prevalence was higher in individuals who were C. sativa smokers (14.6%). Two individuals reported mild symptoms with C. sativa. Twenty-one individuals from 32 available sera (65.6%) had positive specific IgE to C. sativa. Twelve sera recognized at least 6 different bands in a molecular-weight range of between 10 and 60 kDa. Six of them recognized a 10-kDa band, identified as a lipid transfer protein (LTP) and 8 recognized a 38-kDa band, identified as a thaumatin-like protein. Conclusions: There is a high prevalence of sensitization to C. sativa leaves. The clinical symptoms directly attributed to C. sativa were uncommon and mild. The sensitization profile observed suggests that C. sativa sensitization may be mediated by two mechanisms, i.e. cross-reactivity, mainly with LTP and thaumatin-like protein, and exposure-related ‘de novo' sensitization.

Background Knowledge of the domestic mite fauna and allergen levels is important for a correct diagnosis and treatment of mite allergy. Our objectives were to describe the domestic mite fauna in the region of Murcia, Spain, to quantify mite allergens in dust samples obtained from mattresses of this area and to assess the influence of geographical, climatic and dwelling factors. Methods Dust samples were collected in a transversal descriptive study from mattresses of 51 patients who went to the Allergology Service, and from mattress of 81 neighbours or family members of these patients. A questionnaire about home environment was filled in and obtained by all participants. Mite identification was done by light microscopy and allergen determinations (Der p 1 and Der f 1) by monoclonal antibodies. Results Sixteen mite species were identified in the 132 dust samples collected. The most frequent species were Dermatophagoides farinae (36% of the samples), Dermatophagoides pteronyssinus (32%) and Tyrophagus putrescentiae (5.3%). There were significant differences among climatic regions. The coastal sector had greater mite abundance, being D. pteronyssinus more frequent and abundant than D. farinae . In inland areas D. farinae was the predominant mite species. Allergen levels correlated with the concentration of Dermatophagoides , with higher levels detected in coastal regions. Average annual temperature was the main outdoor factor that correlated with higher mite concentrations. Indoor main predictor of higher levels of mites was the presence of obvious signs of humidity in the home. Conclusion This study demonstrates the existence of a mite fauna dominated by D. pteronyssinus and D. farinae with a strong influence of climatic factors and residential characteristics.

Date palm pollen allergy is frequently associated with polysensitisation. Observational studies have suggested that date-palm-sensitised individuals could be included in a distinct group of polysensitised patients. The objectives of the study were to analyse the clinical characteristics of a group of patients diagnosed of date-palm pollen allergy and to compare them with pollen allergic patients without date-palm sensitisation.Forty-eight palm-pollen sensitised individuals were classified as Group A. A control group of 48 patients sensitised to pollens but without palm-pollen allergy were included as Group B. All individuals were skin prick tested with a common battery of aeroallergens. Information about age, sex, family history of atopy, respiratory symptoms, food allergy and sensitisation to other pollens were considered variables of the study. Specific IgE and the allergogram to date-palm pollen were determined in a subgroup of Group A.Significant differences in the family history of atopy and number of sensitisations were observed. Both parameters were significantly higher in Group A. Group A showed high prevalence of asthma and higher level of sensitisation to foods (p0.05). Significant differences were obtained for sensitisation to epithelia and pollens. Pho d 2 was the most commonly recognised allergen (83.3%) in the palm-pollen allergic group.Date-palm pollen allergic patients constitute a homogeneous group characterised for showing bronchial asthma, sensitisation to food allergens and polysensitisation. These results suggest that the reasons for sensitisation to date-palm pollen remain to be elucidated, but could relate to the existence of as yet non-identified pan-allergens.

Background Cross-reactivity among fruits and different pollen and fruit species has been extensively reported. Objectives To investigate the in vitro cross-reactivity between tomato and pollen, fruit, and latex extracts and to identify the proteins involved. Methods A serum pool was prepared from 18 individuals residing on the Spanish Mediterranean coast (9 men and 9 women; mean [SD] age, 27.4 [10.1] years) who had positive skin prick test reactions to tomato peel. Extracts from 10 pollens, 12 fruits, and latex were tested. Levels of specific IgE to each extract were measured. The allergenic profile was evaluated by means of immunoblot. The percentage of inhibition between extracts and tomato peel extract was analyzed by means of CAP inhibition, and the allergens implicated were elucidated by immunoblot inhibition. Results For pollens, the highest specific IgE values were obtained for grasses. Most pollen extracts showed a capacity of inhibition similar to that of tomato peel extract; high percentages were obtained with Artemisia vulgaris and Poa pratensis . The most strongly inhibited allergens in tomato corresponded to bands of 32 and 45 kDa. For fruits, the highest value of specific IgE was detected for peach. High percentages of inhibition were obtained with peach and hazelnut. No inhibition was detected with latex. Peach, chestnut, and melon inhibited high molecular weight bands (32 and 45 kDa) and a band of approximately 10 kDa. Conclusions Cross-reactivity between tomato and pollen and fruit extracts has been demonstrated. Allergens with a high molecular weight range seem to be responsible in pollen extracts. A 10-kDa band seems to be responsible in Platanus acerifolia, Salsola kali , peach, chestnut, and melon.

Background: Sensitivity to Chenopodiaceae is a frequent cause of allergic respiratory diseases in geographic areas where sensitization to Salsola kali and Chenopodium album has been reported. The objective of this study was to evaluate the pattern of sensitization to 3 Salsola species in patients residing on the Mediterranean coast of south-eastern Spain. Methods:S. kali, S. vermiculata and S. oppositifolia pollen extracts were prepared. Patients reporting respiratory and/or cutaneous symptoms were skin prick tested with the 3 Salsola extracts. Individuals with positive skin prick tests to at least 1 of the 3 Salsola species were included. Specific IgE was determined by direct ELISA. SDS-PAGE and 2-D analysis were conducted to elucidate the protein profile. The allergenic profile was investigated by immunoblot. Inhibition experiments were conducted to establish cross-reactivity between different species. Results: 246 patients were included. 237 patients (96.3%) tested positive to S. oppositifolia, 189 (76.8%) to S. kali and 185 (75.2%) to S. vermiculata. Protein profile and immunoblot demonstrated similar patterns in all extracts, except in low-molecular-weight allergens of S. oppositifolia. Immunoblot inhibition experiments demonstrated that most high-molecular-weight allergens of S. oppositifolia were inhibited by S. kali whereas low-molecular-weight allergens were totally inhibited only by C. album. Conclusions: This study confirms the allergenic importance of other Salsola species, especially S. oppositifolia. We have demonstrated that the 3 species show a high degree of cross-reactivity, but S. oppositifolia shares more allergenic similarities with C. album than S. kali.

Background: Cases of allergy to Cannabis sativa have occasionally been reported, but both the allergenic profile and eventual cross-reactivity pattern remain unknown. Objective: To analyze the allergenic profile of a population of patients from Spain sensitized to C. sativa and to characterize the C. sativa leaf extract. Methods: A total of 32 subjects were enrolled in the study: group A, 10 individuals sensitized to tomato, reporting reactions by contact or inhalation to Cannabis; group B, 14 individuals sensitized to tomato, without reactions to Cannabis; group C, 8 individuals not sensitized to tomato and without reactions to Cannabis. Sensitivity to Cannabis, tomato and peach peel, Platanus hybrida and Artemisia vulgaris pollen extracts was measured by skin tests and specific IgE. Individual immunoblots and inhibition experiments with a pool of sera were conducted. Results: All tomato-sensitized subjects (and 1 negative) had positive skin tests to C. sativa leaves and hashish. Specific IgE to C. sativa and peach peel was more common than to tomato. Immunoblot experiments showed 2 prominent bands of 10 and 14 kDa and 2 weakly recognized bands of 30 and 45 kDa. Tomato, peach and A. vulgaris extracts inhibited most of the bands present in C. sativa. P. hybrida inhibited only the high-molecular-weight bands. Conclusion: Sensitization to C. sativa with or without symptoms is frequent among patients in Spain sensitized to tomato. C. sativa leaves are a potential allergenic source and their allergens may cross-react with other allergenic sources from plants (fruit peels and pollen).

Background Seafood plays an important role in human nutrition and is responsible for severe hypersensitivity reactions. Objective To evaluate how the cooking process may alter the in vivo and in vitro allergenicity of these extracts. Methods Raw and boiled extracts of shrimps and 2 types of lobsters were manufactured. Boiled extracts were prepared after the raw material was boiled for 15 minutes in phosphate-buffered saline. Raw and boiled extracts were homogenized and extracted for 4 hours. Afterward, the extracts were centrifuged, dialyzed, filtered, and freeze-dried. Seventy-eight patients were skin prick tested with these raw and boiled extracts. Specific IgE against the 6 extracts was measured by enzyme-linked immunosorbent assay (ELISA). Immunoblots and ELISA inhibition studies were performed with a pool of sera. Results In vivo results showed that boiled extracts induced statistically significant larger wheals than raw extracts. More patients with positive results were also detected with boiled extracts. In vitro experiments by direct ELISA confirmed the in vivo results. The protein content in the boiled extracts decreased, and important differences were detected by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Crossreactivity experiments showed that both types of extracts retained similar allergenic characteristics, even if the immunoblots revealed some differences in IgE binding. Conclusions More patients were identified using boiled extracts of shrimp and American and spiny lobsters than with raw extracts. The wheal sizes of the skin test reactions and specific IgE levels were also significantly greater using boiled extracts. The use of boiled extracts seems to be more effective in diagnosing seafood allergy.

Three tomato allergens (Lyc e 1, Lyc e 2 and Lyc e 3) have been characterised until now. Previous studies have demonstrated the existence of other proteins with capacity to bind IgE. The objective of this study was to identify new tomato allergens by molecular biology techniques. A tomato peel cDNA library was screened using a specific pool of sera from sensitised individuals. A clone was obtained and its sequence compared to other described allergens. This clone contained 34 aminoacids from the C-terminal region of an acidic ribosomal protein 60S. This protein showed more than 85% of homology with fungal allergens and 93.8% of homology with the almond allergen Pru du 5. The clone also contains 169 base pairs without homology with other known genes and could correspond to non-coding sequences. The IgE detection with a pool of sera was positive which confirms that is a new tomato allergen.

Background Sensitisation to pan-allergens has become an interesting tool for the study of the allergenic profile of different populations. Profilins are one of the most common pan-allergens to be studied because they are responsible for a large number of sensitisations and are clearly related to cross-reactivity and co-sensitisation. Objectives The objective of this study was to investigate the profile of sensitisation to profilins and to correlate it with sensitisation to foods and pollens. Methods Six hundred and fifty-four consecutive patients were skin-prick tested with a battery of common allergens including pollens, epithelia, mites and moulds and profilin and divided into three groups depending on their sensitisation profile (non-atopic, atopic with pollinosis and atopic without pollinosis). Patients with symptoms were challenged and diagnosed with the offending food extracts. Profilin sensitisation was identified and analysed in detail. Results According to the classification of the population, the prevalence of profilin sensitisation was estimated at 2.9% in patients suffering respiratory allergy, 4.2% in atopic patients, and 5.9% in pollen-sensitised individuals. Positive association was observed between pollen (except Cupressus and olive) and profilin but not with moulds, mites or epithelia. With respect to foods, positive association was only observed between profilin and melon sensitisation. Lastly, in terms of symptoms, positive association was only observed between profilin sensitisation and OAS. Conclusion Profilin sensitisation seems to be a marker of pollen-related poly-sensitisation in our area. Pan-allergen diagnosis seems to be an essential tool for developing and improving selection of the correct treatment for allergic patients.

Background The prevalence of sensitization to tomato has been estimated in 6.5% in a Mediterranean population. Objectives To characterize the allergenic profile of 6 commonly ingested varieties of tomato and to identify and quantify the most representative allergens. Methods Rama, Rambo, Canario, Kumato, Pera and Raf tomato varieties were carefully peeled and extracts from peel and pulp prepared. The extracts were characterized by SDS-PAGE and immunoblot. Lyc e 3 and osmotin-like protein (OLP) were identified by LC MS/MS and PG2A by immunoblot with a specific polyclonal antibody. Protein content and allergen concentration (Lyc e 3 and PG2A) were measured by scanning densitometry after identification. One hundred and ninety-one patients were enrolled in the study. Specific IgE to all tomato extracts was evaluated by direct ELISA. Results Eighty (41.7%) individuals reported symptoms after contact/ingestion with tomato, most of them oral symptoms. Protein profile and allergen quantification of the 6 varieties showed differences in the allergen composition. Lyc e 3 was visualized in all extracts, being the most abundant in all cases. PG2A and OLP were identified in all peel extracts and in some pulp extracts. Higher specific IgE values were obtained for the Kumato variety. Immunoblot showed the existence of several bands with IgE-binding capacity, especially at 12 and 45-50 kDa. Conclusions The study demonstrates differences in the antigenic and allergenic profiles of the analyzed varieties and variations in their allergenic composition, affecting the diagnostic capacity. LTP and PG2A are important tomato allergens in our population but other IgE-binding capacity proteins are also involved.

Summary Background Tomatoes (Lycopersicon esculentum) are consumed world-wide. The prevalence of sensitization to tomatoes remains unknown. Objective To determine the prevalence of skin test reactivity to tomato and to describe the characteristics of tomato-sensitized subjects. Methods Individuals attending for the first time during the period of the study to six Allergy centres, located along the Mediterranean coast of Spain, reporting respiratory and/or cutaneous symptoms, were included. All patients were skin prick tested with a battery of inhalant allergens and with peel and pulp of Canary tomato extracts. Results The study included 1734 individuals (757 males, 977 females; 31.9±17.8 years old). The prevalence of sensitization to tomato was 6.52% (113 patients; 65 males, 48 females; 29.5±13 years old). The peel extract was positive in 110 patients and the pulp extract in 47 patients; three patients were positive exclusively to pulp. Only 1.8% of individuals reported symptoms with tomato; 44% of them had skin test negative to both extracts. Among tomato-sensitized subjects, 16% reported symptoms with tomato, 97% were sensitized to inhalant aeroallergens, including 84% to pollens (mainly Artemisia vulgaris and Platanus hybrida), with differences between Northern and Southern centres. Conclusions The prevalence found of skin test sensitivity to tomato is high. Peel extracts detected most of the sensitized subjects. Most of the sensitized subjects were asymptomatic and some patients reported symptoms without skin test sensitivity. Positive subjects were very frequently sensitized to pollens, suggesting allergen cross-reactivity. Regional differences may exist, possibly related to the pattern of sensitization to cross-reacting pollens.

Background Co-sensitization to house dust mites (HDMs) and storage mites (SMs) is very frequent, although the clinical relevance is not well established. Objective To describe the pattern of sensitization and immunologic characterization of patients with positive skin prick test reactions to HDMs and SMs in 4 areas of Spain, selected according to high exposure to HDMs and variable exposure to SMs. Methods One hundred sixty-nine individuals with positive skin prick test reactions to HDMs and SMs were included. Specific IgE levels to different mite species and to Der p 1, Der p 2, and Der p 10 were determined. Immunoblots to Dermatophagoides pteronyssinus, Lepidoglyphus destructor, and Tyrophagus putrescentiae extracts were carried out, and allergograms were obtained. Results of different areas were compared. Results A high rate of polysensitization to SMs was observed, although 12% of participants did not have specific IgE to any SM species. Sensitization to Dermatophagoides species, Der p 2, and L destructor were predominant, although significant differences were observed among areas depending on the grade of exposure to SMs. In areas with high exposure, the SM allergogram showed greater recognition of group 2 allergen. Conclusion Sensitization patterns to SMs in patients sensitized to HDMs and SMs differ depending on the exposure to SMs. Sensitization, mainly to L destructor, seems to exist in areas with high exposure, possibly with group 2 allergens mainly involved. However, in areas with low SM populations, sensitizations observed by skin prick testing appear to be related to HDM exposure.

M O N D A Y 651 Allergenic Profile Differences In Individuals Sensitized To House Dust And Storage Mites In Different Climatic And Geographical Areas V. Iraola, M. Morales, A. Huertas, M. Boquete, F. Rodriguez, J. Bartra, M. Paniagua, S. D iaz Angulo, F. Carballada, J. Carnes; Laboratorios LETI S.L., Tres Cantos, SPAIN, Hospital Santa Mar ia del Rosell, Cartagena, SPAIN, Hospital Xeral de Calde, Lugo, SPAIN, Hospital de Valdecilla, Santander, SPAIN, Hospital Cl inico de Barcelona, Barcelona, SPAIN, Hospital San Pere Clavert, Barcelona, SPAIN. RATIONALE: Previous studies demonstrated different exposure and sensitisation patterns to house dust (HDM) and storage mite (SM) depending on the geographic area in Spain. TheAtlantic area (more humid) has high exposure to HDM-SM, whereas the Mediterranean area (warmer) has high HDM abundance. METHODS: Serum samples were taken from patients with positive skin prick test to HDM and at least to one SM from two climatic areas with high exposure to mites (Atlantic-Mediterranean). Specific IgE levels to HDM, SM, Der p1 and Der p2 were determined. Immunoblot to D.pteronyssinus, L.destructor and T.putrescentiae were done using sera with sIgE to HDM and SM. 152 patients (71 Mediterranean; 81 Atlantic) were included. RESULTS: In the Mediterranean area 14% patients were monosensitised to only one SM,whereas 55%were sensitised to all SM. In theAtlantic area percentages were 4% and 75%. SM with highest sIgE were L.destructor and T.putrescentiae, although they were inferior to those obtained with HDM. sIgE against all SM (except A. siro) and Der p2 were significant greater in the Atlantic area, whereas sIgE to D. farinae and E. maynei were higher in Mediterranean. D.pteronyssinus allergen recognition patternwas similar in both areas. Regarding SM,most ofAtlantic area patients recognised allergens of 13-15 kDa, whereas in theMediterranean areamost individuals recognised high MW allergens. CONCLUSIONS: HDM are the main sensitising species in Spain, although higher exposure to SM is related with greater sIgE levels. In patients with sIgE to HDM and SM there are different allergens involved depending on the geographic area.

Background Commercial available skin prick test with fruits can be negative in sensitized or allergic patients due to a reduction in biological activity during the manufacturing process. Prick-prick tests with fresh foods are often preferred, but they are a non-standardized procedure. The usefulness of freeze-dried extracts of Canary Islands tomatoes, comparing the wheal sizes induced by prick test with the prick-prick method in the diagnosis of tomato sensitization has been analyzed. The objective of the study was to assess the potential diagnostic of freeze-dried extracts of Canary Islands tomatoes, comparing the wheal sizes induced by prick test with the prick-prick method. Methods Two groups of patients were analyzed: Group I: 26 individuals reporting clinical symptoms induced by tomato contact or ingestion. Group II: 71 control individuals with no symptoms induced by tomato: 12 of them were previously skin prick test positive to a tomato extract, 39 were atopic and 20 were non-atopic. All individuals underwent prick-prick with fresh ripe peel Canary tomatoes and skin prick tested with freeze-dried peel and pulp extracts obtained from peel and pulp of Canary tomatoes at 10 mg/ml. Wheal sizes and prick test positivity (≥ 7 mm2) were compared between groups. Results In group I, 21 (81%) out of 26 patients were prick-prick positive. Twenty patients (77%) had positive skin prick test to peel extracts and 12 (46%) to pulp extracts. Prick-prick induced a mean wheal size of 43.81 ± 40.19 mm2 compared with 44.25 ± 36.68 mm2 induced by the peel extract (Not significant), and 17.79 ± 9.39 mm2 induced by the pulp extract (p < 0.01). In group II, 13 (18%) out of 71 control patients were prick-prick positive. Twelve patients (all of them previously positive to peel extract) had positive skin prick test to peel and 3 to pulp. Prick-prick induced a mean wheal size of 28.88 ± 13.12 mm2 compared with 33.17 ± 17.55 mm2 induced by peel extract (Not significant), and 13.33 ± 4.80 mm2 induced by pulp extract (p < 0.05 with peel extract and prick-prick). Conclusion Canary peel tomato extract seems to be as efficient as prick-prick tests with ripe tomatoes to diagnose patients sensitized to tomato. The wheal sizes induced by prick-prick and peel extracts were very similar and showed a high correlation coefficient.