In vivo diagnosis with purified tropomyosin in mite and shellfish allergic patients.

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.

Different sensitization to storage mites depending on the co-exposure to house dust mites.

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.

Prevalence of sensitization to Cannabis sativa. Lipid-transfer and thaumatin-like proteins are relevant allergens.

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.

Recently introduced foods as new allergenic sources: sensitisation to Goji berries (Lycium barbarum).

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.

Identification and quantification of tomato allergens: in vitro characterization of six different varieties.

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.

In vitro cross-reactivity between tomato and other plant 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.

Sensitization and allergy to Cannabis sativa leaves in a population of tomato (Lycopersicon esculentum)-sensitized patients.

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).

The use of raw or boiled crustacean extracts for the diagnosis of seafood allergic individuals.

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. Cross-reactivity 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.