Boletus edulis: a digestion-resistant allergen may be relevant for food allergy.

BACKGROUND: Fungal components can cause allergic symptoms either through inhalation, ingestion or contact. Whereas respiratory allergy is thought to be induced by spores, allergic reactions following ingestion are attributed to other parts of the mushroom. Reports of food-related allergic reactions due to the edible mushroom Boletus edulis have occasionally been reported. OBJECTIVE: The aim of the study was to investigate whether separate allergens may be detected in alimentary allergy to Boletus edulis. METHODS: Sera of two subjects, one with recurrent anaphylaxis and the other with a predominantly oral allergy syndrome following ingestion of Boletus edulis, have been analysed by a time-course digestion assay using simulated gastric fluid and by SDS-PAGE immunoblotting. Sera of four Boletus edulis skin prick test-negative subjects and all without clinical symptoms to ingested Boletus edulis served as controls. RESULTS: In lyophilized Boletus edulis extract, at least four water-soluble proteins were detected, the most reactive at 55 kDa and at 80 kDa. Following the time-course digestion assay, IgE binding was found to a 75-kDa protein, but only if the sera of the subject with recurrent anaphylaxis was used. CONCLUSION: The data indicate that Boletus edulis can cause an IgE-mediated food allergy due to a digestion-stabile protein at 75 kDa. No IgE immune response to this protein was detected in the serum of a subject with respiratory allergy and oral allergy syndrome to Boletus edulis nor in control sera.

Coprinus comatus (shaggy cap) is a potential source of aeroallergen that may provoke atopic dermatitis.

BACKGROUND: Basidiospores are universal components in the air and established as important causes of respiratory allergies. Recent reports indicate that aeroallergens may aggravate eczematous skin lesions in subjects with atopic dermatitis (AD). OBJECTIVE: The aim of the study was to investigate whether spores of Coprinus comatus, a species of basidiomycetes, may elicit delayed-type skin reactions in subjects with an atopic predilection, especially dermatitis. METHODS: Sixty-six study subjects were categorized in groups having AD or respiratory allergy with regard to the skin prick test (SPT) reactivity to C comatus extract. Twenty nonatopic individuals served as control subjects. Atopy patch tests (APTs) were performed with extract of C comatus spore containing tissue at a concentration of 1. 35 mg of protein per gram of petrolatum (Vaseline) and C comatus cap at a concentration of approximately 5 mg of protein per gram of petroleum jelly. APT reactions were evaluated after 48 and 72 hours. RESULTS: Of the subjects with AD completing the study, 12 (32%) of 38 showed a positive APT reaction, with 8 (57%) also having a positive SPT response to C comatus. Only 1 (9%) of 11 subjects with asthma had a positive SPT and APT response to C comatus. No positive test reaction was observed in the nonatopic control subjects or in subjects with respiratory allergy and negative SPT responses to C comatus. CONCLUSION: Our results demonstrate that C comatus can induce delayed-type reactions in atopic individuals, particularly in those with AD. Because spores of Coprinus species are ubiquitous, basidiomycetes have to be considered as possible aeroallergens when investigating causes of eczematous skin lesions in AD.

IgE-binding proliferative responses and skin test reactivity to Cop c 1, the first recombinant allergen from the basidiomycete Coprinus comatus.

BACKGROUND: Basidiomycetes spores are ubiquitously distributed, found throughout the year in outdoor and indoor air, and represent relevant sources of aeroallergens associated with allergy and asthma. OBJECTIVE: Cloning and characterization of Coprinus comatus (shaggy cap mushroom) allergens is essential to elucidate their molecular characteristics and to improve the diagnosis of allergy. METHODS: A complementary DNA (cDNA) library of C comatus displayed on phage surface was screened with sera of basidiomycete-sensitized individuals. Subcloning and high-level expression of one of the enriched cDNAs allowed the isolation of a [His](6)-tagged recombinant protein formally termed rCop c 1. The allergenic properties of rCop c 1 were investigated in vitro by ELISA, inhibition experiments, immunoblots, and proliferation assays and in vivo by skin tests. RESULTS: The rCop c 1-encoding cDNA spans 435 bp and contains an open reading frame of 246 bp, predicting a protein of 8.96 kd without significant sequence homology to known proteins. Immunoblots with [His](6)-rCop c 1 fusion protein show a background free IgE-binding band of the expected size that can be completely inhibited by crude C comatus extracts in ELISA. rCop c 1 induced specific proliferative responses in PBMCs of C comatus-sensitized individuals. The incidence of sensitization to rCop c 1 among 92 sera of basidiomycete-sensitized individuals tested in ELISA was 25%, indicating that Cop c 1 is an intermediate allergen. However, prick tests showed that less than 2 pmol of the rCop c 1 protein was able to induce strong specific skin reactions in sensitized individuals. CONCLUSIONS: rCop c 1, the first cloned allergen from the genus Coprinus, fulfills all the criteria required to be classified as a clinically relevant allergen. The data demonstrate at the molecular level the presence of sensitizing molecules among Basidiomycetes, the most important source contributing to the total spore load in the outdoor air.

Respiratory allergy to mushroom spores: not well recognized, but relevant.

BACKGROUND: Although basidiospores are a major component of the air spora in many parts of the world, their clinical significance as triggers of respiratory allergy has rarely been demonstrated. Therefore, the class of basidiomycetes as an aeroallergen is not well known. OBJECTIVE: To demonstrate a cause and effect relationship between respiratory allergy and basidiospores, we illustrate this case report of a 38-year-old housewife. METHODS: Skin prick test, immunoblot, and active anterior rhinomanometry were used as diagnostic tools to verify specific reactivity of a Pleurotus pulmonalis spore extract. Two atopic subjects served as controls. RESULTS: The skin prick test positive study subject reacted with subjective and objective signs including a significant drop of the FEV1 by nasal challenge at a concentration of 0.1 mg/mL of the Pleurotus spore extract while both controls were negative even at a higher test concentration. IgE-immunoblot revealed several distinct bands in the serum of the Pleurotus-sensitized subject. CONCLUSION: Spores of Pleurotus pulmonalis, a common mushroom of the fungal class of basidiomycetes, can cause specific, IgE-mediated acute rhinoconjuncivitis and asthma in sensitized individuals.

Mushroom (Basidiomycete) allergy: diagnosis established by skin test and nasal challenge.

BACKGROUND: Fungal spores are universal components in the air and established as important causes of respiratory allergies. Whereas fungi imperfecti are accepted sources of allergic asthma and rhinitis, the significance of basidiomycetes as respiratory allergens is not established. OBJECTIVES: The aims of the present study were to investigate the rate of sensitization in subjects referred to an allergy clinic to 3 basidiomycetes commonly found in Europe. We demonstrate the clinical relevance of basidiomycete sensitization by active anterior rhinomanometry and identified basidiomycete allergens by immunoblotting. METHODS AND RESULTS: Consecutive outpatient clinic attendees (n = 1207, >12 years of age) were given the skin prick test with a panel of common inhalant allergens and extracts of Boletus, Coprinus, and Pleurotus. To evaluate a cause-and-effect relation between basidiomycete allergens and respiratory allergies, 12 Pleurotus pulmonalis spore-sensitized subjects with respiratory symptoms and 6 control subjects were challenged by anterior rhinomanometry. SDS-PAGE immunoblots were used to identify basidiomycete-specific IgE antibodies in sera from subjects with positive skin prick test results. Of 1207 subjects tested, 48 (4%) reacted to at least 1 of the basidiomycete extracts. Whereas all of the 12 subjects with Pleurotus pulmonalis-positive skin test results showed a significant decrease in nasal air flow (mean 73%), none of the control subjects reacted. Immunoblots revealed several different IgE-binding proteins in Pleurotus and Coprinus extracts. CONCLUSIONS: Our results demonstrate that sensitization to basidiomycetes in subjects with respiratory allergies is frequent. Furthermore, Pleurotus pulmonalis spore extracts can definitely induce respiratory allergy in sensitized subjects. Immunoblot assays disclosed different IgE-reactive bands indicating the existence of basidiomycete allergens.

HLA-restricted, processing- and metabolism-independent pathway of drug recognition by human alpha beta T lymphocytes.

T cell recognition of drugs is explained by the hapten-carrier model, implying covalent binding of chemically reactive drugs to carrier proteins. However, most drugs are nonreactive and their recognition by T cells is unclear. We generated T cell clones from allergic individuals specific to sulfamethoxazole, lidocaine (nonreactive drugs), and cef-triaxone (per se reactive beta-lactam antibiotic) and compared the increase of intracellular free calcium concentration ([Ca2+]i) and the kinetics of T cell receptor (TCR) downregulation of these clones by drug-specific stimulations. All drugs tested induced an MHC-restricted, dose- and antigen-presenting cell (APC)-dependent TCR downregulation on specific CD4(+) and CD8(+) T cell clones. Chemically nonreactive drugs elicited an immediate and sustained [Ca2+]i increase and a rapid TCR downregulation, but only when these drugs were added in solution to APC and clone. In contrast, the chemically reactive hapten ceftriaxone added in solution needed > 6 h to induce TCR downregulation. When APC were preincubated with ceftriaxone, a rapid downregulation of the TCR and cytokine secretion was observed, suggesting a stable presentation of a covalently modified peptide. Our data demonstrate two distinct pathways of drug presentation to activated specific T cells. The per se reactive ceftriaxone is presented after covalent binding to carrier peptides. Nonreactive drugs can be recognized by specific alphabeta+ T cells via a nonconventional presentation pathway based on a labile binding of the drug to MHC-peptide complexes.

Aerobic fermentation in tobacco pollen.

We characterized the genes coding for the two dedicated enzymes of ethanolic fermentation, alcohol dehydrogenase (ADH) and pyruvate decarboxylase (PDC), and show that they are functional in pollen. Two PDC-encoding genes were isolated, which displayed reciprocal regulation: PDC1 was anaerobically induced in leaves, whereas PDC2 mRNA was absent in leaves, but constitutively present in pollen. A flux through the ethanolic fermentation pathway could be measured in pollen under all tested environmental and developmental conditions. Surprisingly, the major factor influencing the rate of ethanol production was not oxygen availability, but the composition of the incubation medium. Under optimal conditions for pollen tube growth, approximately two-thirds of the carbon consumed was fermented, and ethanol accumulated into the surrounding medium to a concentration exceeding 100 mM.

Highly conserved genes coding for eukaryotic translation initiation factor eIF-4A of tobacco have specific alterations in functional motifs.

Eukaryotic translation initiation factor eIF-4A is an ATP-dependent RNA helicase that is required for the binding of mRNA to ribosomes. Plant eIF-4A-like proteins are highly homologous to eIF-4As from yeast, mouse and Drosophila melanogaster. The pattern of intron-exon boundaries in eIF-4A-like genes are conserved within tobacco, but are not conserved with other organisms. Fixed spacings between the functionally important sequence motifs, GKT-PTRELA (72 bp), DEAD-SAT (81 bp) and SAT-HRIGR (426 bp), are conserved between plants, mouse, Drosophila and yeast.

A pollen-specific DEAD-box protein related to translation initiation factor eIF-4A from tobacco.

A pollen-specific sequence, NeIF-4A8, has been isolated from a cDNA library from mature pollen of Nicotiana tabacum cv. Samsun. NeIF-4A8 is a full-length cDNA whose deduced amino acid sequence exhibits high homology to the eucaryotic translation initiation factor eIF-4A from mouse, Drosophila and tobacco. eIF-4A is an RNA helicase which belongs to the supergene family of DEAD-box proteins. Northern blot analysis with a gene-specific probe showed strict anther-specific expression of NeIF-4A8 starting at microspore mitosis. With antibodies raised against tobacco eIF-4A the presence of abundant eIF-4A-related proteins in developing anthers and pollen grains was demonstrated. The genomic analysis shows that the coding region is split by three introns whereas a large, fourth intron is situated in the 5'-untranslated region. A promoter construct with 2137 bp of upstream sequence fused to the GUS reporter gene was used to confirm that the expression is confined to the haploid cells within the anther. NeIF-4A8 is a prime candidate formediating translational control in the developing male gametophyte.