The non-specific lipid transfer protein, Ara h 9, is an important allergen in peanut.

BACKGROUND: Plant food allergy in the Mediterranean area is mainly caused by non-specific lipid transfer proteins (nsLTP). The aim of this study was to characterize peanut nsLTP in comparison with peach nsLTP, Pru p 3, and assess its importance in peanut allergy. METHODS: Peanut-allergic patients from Spain (n=32) were included on the basis of a positive case history and either a positive skin prick test or specific IgE to peanut. For comparison, sera of 41 peanut-allergic subjects from outside the Mediterranean area were used. Natural Ara h 9 and two isoforms of recombinant Ara h 9, expressed in Pichia pastoris, were purified using a two-step chromatographic procedure. Allergen characterization was carried out by N-terminal sequencing, circular dichroism (CD) spectroscopy, immunoblotting, IgE inhibition tests and basophil histamine release assays. RESULTS: Compared with natural peanut nsLTP, the recombinant proteins could be purified in high amounts from yeast supernatant (> or =45 mg/L). The identity of the proteins was verified by N-terminal amino acid sequencing and with rabbit nsLTP-specific antibodies. CD spectroscopy revealed similar secondary structures for all preparations and Pru p 3. The Ara h 9 isoforms showed 62-68% amino acid sequence identity with Pru p 3. IgE antibody reactivity to rAra h 9 was present in 29/32 Spanish and 6/41 non-Mediterranean subjects. Recombinant Ara h 9 showed strong cross-reactivity to nPru p 3 and similar IgE-binding capacity as nAra h 9. The two Ara h 9 isoforms displayed similar IgE reactivity. In peanut-allergic patients with concomitant peach allergy, Ara h 9 showed a weaker allergenic potency than Pru p 3 in histamine release assays. CONCLUSIONS: Ara h 9 is a major allergen in peanut-allergic patients from the Mediterranean area. Ara h 9 is capable of inducing histamine release from basophils, but to a lesser extent than Pru p 3.

The CREATE project: development of certified reference materials for allergenic products and validation of methods for their quantification.

Allergen extracts have been used for diagnosis and treatment of allergy for around 100 years. During the second half of 20th century, the notion increasingly gained foothold that accurate standardization of such extracts is of great importance for improvement of their quality. As a consequence, manufacturers have implemented extensive protocols for standardization and quality control. These protocols have overall IgE-binding potencies as their focus. Unfortunately, each company is using their own in-house reference materials and their own unique units to express potencies. This does not facilitate comparison of different products. During the last decades, most major allergens of relevant allergen sources have been identified and it has been established that effective immunotherapy requires certain minimum quantities of these allergens to be present in the administered maintenance dose. Therefore, the idea developed to introduce major allergens measurements into standardization protocols. Such protocols based on mass units of major allergen, quantify the active ingredients of the treatment and will at the same time allow comparison of competitor products. In 2001, an EU funded project, the CREATE project, was started to support introduction of major allergen based standardization. The aim of the project was to evaluate the use of recombinant allergens as reference materials and of ELISA assays for major allergen measurements. This paper gives an overview of the achievements of the CREATE project.

Human seminal plasma allergy and successful pregnancy.

Human seminal plasma allergy in women is an uncommon phenomenon. A great variety of reactions ranging from local swelling to generalized systemic reactions have been described, and local symptoms have often been misdiagnosed as chronic vulvovaginitis. Sperm barriers, such as condoms, are the most widely advocated method for avoiding these reactions; however this is not acceptable to couples who wish to have children. We present a case of a woman with human seminal plasma allergy who became pregnant after a fourth cycle of artificial insemination. Sodium dodecyl sulfate polyacrylamide gel electrophoresis immunoblotting showing an IgE binding band at 28kDa in the husband's seminal fluid identified the culprit allergen. Artificial insemination is an effective way to achieve a pregnancy in patients with seminal plasma allergy.

Exotic pets are new allergenic sources: allergy to iguana.

Although furry animals are known sources of respiratory allergy, scaly animals are assumed not to be allergenic. Exotic animals such as iguanas are becoming increasingly common pets. Nevertheless, these animals are not suspected to be allergenic. We present the case of a 42-year-old woman suffering from allergic rhinoconjunctivitis and asthma caused by a pet iguana. Clear IgE-sensitization and respiratory allergy to iguana scales is demonstrated, suggesting that scaly pets should be taken into account as possible allergenic sources.

Double-blind, placebo-controlled study of immunotherapy with Parietaria judaica: clinical efficacy and tolerance.

Allergy to Parietaria causes significant morbidity in most Mediterranean areas. The aim of this study is to investigate the efficacy and tolerance of Parietaria depot extract at 25 BU/mL (1.5 microg/mL Par j 1). We performed a multicenter double-blind, placebo-controlled study in rhinitic patients with/without asthma, sensitized to Parietaria. 42 patients followed 20-month immunotherapy. Clinical efficacy was based on symptom and medication scores and the percentage of healthy days (days without symptoms or medication). Severity of asthma/rhinitis scales, visual analogue scale, evaluation of the treatment by doctors and patients, immediate and delayed cutaneous response and quality of life questionnaires were also studied. The active group showed a sustained decrease in symptoms (p = 0.008), medication (p = 0.009) and both (p = 0.001), and an increase in healthy days (p = 0.001) throughout the study, with a threefold increase of healthy days and almost a three time reduction in medication only after one year of treatment. Asthma and rhinitis severity scales also decreased after immunotherapy, and blinded clinical evaluation by physicians confirmed efficacy in 85% and 77% of the active patients. Patient's self-evaluation returned similar results. None of these changes were observed with placebo. Immediate cutaneous response was significantly reduced at the maintenance phase in the active group and remained reduced throughout the study. Late-phase response after intradermal testing also showed a statistical decrease in actively treated patients. Immunotherapy was well tolerated and every systemic reaction reported was mild. In conclusion, immunotherapy with Parietaria 25 BU/mL is an effective and safe treatment for patients with respiratory allergies.

Molecular cloning and immunological characterisation of potential allergens from the mould Fusarium culmorum.

BACKGROUND: High quality and stability are essential requirements of commercial allergen preparations. Recently we have demonstrated the very low stability of protein allergens in an extract of the ubiquitous mould Fusarium culmorum. OBJECTIVE: The present study was performed to identify, isolate and characterise allergens of F. culmorum as a basis for a stable allergenic reference material. In addition, the significance of IgE binding to carbohydrate structures in the natural allergen source was investigated. METHODS: Sera of 52 subjects with suspected mould allergy were used to determine the IgE binding capacity of a commercial F. culmorum extract and an in-house extract by immunoblotting and enzyme allergo sorbent test (EAST). Binding of IgE-antibodies to putative carbohydrate structures located on glycoproteins was verified by periodate treatment of blot strips prior to immunodetection. A complementary (c)DNA expression library of F. culmorum was prepared and screened for IgE-binding clones using sera from F. culmorum-sensitive individuals. Positive clones were isolated, and the open reading frames were subcloned into expression vectors to produce recombinant proteins in E. coli. The recombinant proteins were tested for their IgE reactivity by immunoblotting and EAST. RESULTS: Using the in-house extract for EAST and immunoblot experiments 44% (23/52) of the sera were found to contain F. culmorum-specific IgE antibodies. Compared to the in-house extract, nearly all IgE-reactivties in the range of 15-30kD were lacking in the commercial preparation as examined by immunoblot analysis and only 10% (5/52) of the sera were found to contain F. culmorum-specific IgE by EAST. IgE binding to putative carbohydrate structures was observed in the high molecular weight range in approximately 50% (12/23) of the IgE-positive sera by both extracts. Three IgE binding clones were isolated from the cDNA-library. One clone (Fus c 1) is homologous to the highly conserved 60S acidic ribosomal protein P2 described as minor allergen in other moulds. The second (Fus c 2) shows high similarity (64%) to a respiratory allergen from the basidiomycete Coprinus comatus (Cop c 2). The third clone (Fus c 3) was not related to known proteins. With sera from 26 individuals sensitised to F. culmorum the IgE prevalence of recombinant proteins rFus c 1, rFus c 2 and rFus c 3 was found to be 35, 50, and 15%, respectively. CONCLUSIONS: F. culmorum may represent an underestimated source of aeroallergens. In contrast to highly labile and poorly standardised F. culmorum extracts, the new recombinant allergens may serve as stable allergenic reference material. A combination of rFus c 1 and rFus c 2 is suitable to diagnose 81% of F. culmorum-sensitised subjects. IgE reactivity to putative carbohydrate structures is relatively frequent, and can not be detected by these allergens.

Chronic urticaria associated with chronic myelomonocytic leukemia.

Although chronic urticaria is usually idiopathic, in rare cases it may be a sign of underlying malignancy. We describe the first case of chronic urticaria associated with chronic myelomonocytic leukemia. The urticarial lesions healed successfully with etoposide, an antineoplastic agent. This case demonstrates that cases of chronic urticaria should not to be labeled as idiopathic until diligent evaluation has failed to reveal a cause.

Disseminated and recurrent sarcoid-like granulomatous panniculitis due to bovine collagen injection.

Three to five percent of patients treated with injectable collagen implants have adverse local reactions. Systemic signs or symptoms are infrequent. We describe the case of a 53-year-old woman who developed local and general complaints 2 months after receiving two collagen injections in both nasolabial folds. She presented asthenia, malaise, polyarthralgia and inflammatory nodular panniculitis in the places of injection as well as on the forearms and lower extremities. Skin testing was compatible with hypersensitivity to collagen. This case raises the question as to whether the injection of animal-derived collagen induces manifestations similar to those observed in connective autoimmune diseases or human adjuvant disease.

Recurrent urticaria as a rare manifestation of familial Mediterranean fever.

Familial Mediterranean fever (FMF) is a genetic disorder characterized by acute episodes of fever with some combination of severe abdominal pain, pleurisy, arthritis, and skin rash. The case of a patient with recurrent urticaria referred for study of drug allergy is presented. After allergy had been ruled out, the urticaria was attributed to previously undiagnosed symptoms of an underlying systemic disease: FME. Urticaria is the least frequent cutaneous manifestation of this disease, and genetic analysis was required to confirm the diagnosis.

Tolerance and effects on skin reactivity to latex of sublingual rush immunotherapy with a latex extract.

BACKGROUND: Specific immunotherapy could be a therapeutic tool for the increasing problem of sensitisation to Natural Rubber Latex (NRL). OBJECTIVE: To investigate the tolerability of SLIT for Latex and its effects on skin reactivity. METHODS: Twenty-six patients (mean age 35.5 years) with an average history of 7.5 years of cutaneous symptoms plus respiratory symptoms (23/26) due to NRL were studied. All underwent rush sublingual therapy (4 days) with a standardized NRL extract followed by a 9-week maintenance treatment. Local and systemic adverse reactions were monitored throughout the treatment. Skin reactivity to NRL extract was evaluated before, during and at the end of the treatment by latex glove-use test, rubbing test and skin prick test. RESULTS: All patients reached the maintenance dose. Out of 1044 administered doses, 257 (24.6%) produced adverse reactions from which 21.4% were local. Only 10.1% of cases required treatment, mainly with antihistamines alone (5.8%), with 2-agonists alone (0.8%) or associated to antihistamines and/or corticosteroids (2.7%). One patient was precautionary treated twice with adrenaline but completed the treatment without further problems. The glove-use test improved significantly after 5 days and 10 weeks of treatment (p = 0.003, p = 0.0004 respectively), whereas the rubbing test improved significantly only after 10 weeks of treatment. Doctor's assessments confirmed the results obtained with the glove-use test (p = 0.003 after 5 days, and p = 0.004 after 10 weeks) but not those obtained with the rubbing test. No change was detected for SPTs. CONCLUSION: SLIT for NRL allergy is able to modify skin reactivity to NRL in days as assessed with methods reproducing HCWs normal exposure to the allergen. Tolerance of SLIT is better than tolerance reported for injective therapy with NRL, but the build up phase should be administered under medical surveillance until sufficient experience has been accumulated. The long-term effect of the treatment deserves further investigation.

IgE reactivity to profilin in Platanus acerifolia pollen-sensitized subjects with plant-derived food allergy.

BACKGROUND: The presence of profilin-specific IgE antibodies is a cause of cross-reactivity between botanically-unrelated allergen sources. Recently, the association between Platanus acerifolia pollinosis and plant-derived food allergy has been described. The aim of this study was to ascertain whether the P. acerifolia profilin is involved in such cross-reactivity. METHODS: Twenty-three patients suffering from Platanus acerifolia pollinosis and plant-derived food allergy were evaluated in an allergy department. Specific IgE levels to P. acerifolia pollen, P. acerifolia profilin and food extracts were measured. Molecular masses of IgE-binding proteins were calculated by Western blotting and cross-reactivity studies among P. acerifolia profilin and different food extracts were evaluated by Enzyme AllergoSorbent Test (EAST)-inhibition assays. Also, EAST-inhibition assays with the two known P. acerifolia allergens, Pla a 1 and Pla a 2, were performed. RESULTS: Surprisingly, a high IgE-binding prevalence (90%) of P. acerifolia profilin was found. EAST-inhibition showed high inhibition values when Platanus acerifolia pollen extract was used as free phase and plant-derived food extracts as solid phase, whereas the other way round showed low inhibition values. IgE reactivity to profilin was studied using a pool of patient sera, by EAST-inhibition assays with hazelnut, apple peel, peanut, chickpea and peanut extracts as solid phase and no inhibition was obtained when P. acerifolia profilin was used as inhibitor phase. The same results were obtained when purified Pla a 1 and Pla a 2 were also used as inhibitor phase. CONCLUSIONS: The clinical association observed between Platanus acerifolia pollen and plant-derived food could be explained by the in vitro IgE cross-reactivity detected by EAST-inhibition. However, it appears that neither P. acerifolia profilin nor the two major allergens described (Pla a 1 and Pla a 2) can explain such a strong cross-reactivity.

Olerance of a cluster schedule with a house dust mite extract quantified in mass units: multicentre study.

The standardisation of allergenic extracts in micrograms of the major allergen has encouraged the search for new treatment schedules, with the purpose of shortening the number of visits and doses required to reach the maintenance dose without eliciting a greater risk of adverse reactions for the patients. With this objective, a prospective multicentre pharmacovigilance study was designed that included 200 patient with allergic rhinoconjunctivitis and/or allergic asthma sensitised to mites (Dermatophagoides pteronyssinu and/or farinae). The dose increment period was carried out using a cluster schedule, where the optimal dose wa reached after 4 visits, administering two doses in each visit. The duration of the study was 5 months and a total o 1902 doses were administered. At the end of the trial, 31 adverse reactions in 23 patients were recorded. Six of these were systemic (0.3% of t administered doses) recorded in 6 patients (3% of the sample). One was an immediate reaction (grade 1) and delayed (4 mild and 1 moderate). Two were asthmatic exacerbations, 2 cutaneous reactions, 1 rhinitis and 1 an unspecific symptom (not IgE-mediated). Two appeared upon administration of the first vial and the remaining 4 after administration of the third cluster. Therefore, the schedule tested presents an adequate tolerance profile, suggesting savings (compared to th conventional schedule of 13 doses per patient) of 1800 visits and 1000 treatment doses in the whole study.

Influence in the quality of life of the respiratory patients by environmental control and the acaricide Frontac®.

The present prospective, open, observational study assessed the effect of the use of standard environmental control measures together with the application of an acaricide-containing aerosol of esbiol/benzyl benzoate/piperonyl butoxide/2-phenylphenol (Frontac) on several clinical and environmental parameters in patients suffering from perennial allergic rhinoconjunctivitis related to house dust mites (HDM). Fifty-five patients were enrolled and studied for one year. Both symptoms and the patients' quality of life (evaluated by the Rhinoconjunctivitis Quality of Life Questionnaire (RQLQ)) were assessed at several clinic visits. The amount of HDM antigen was semi-quantitatively estimated using the guanine test. We found a significant decrease (p < 0.001) in most nasal symptoms scores: sneezing, runny nose, stuffy nose and itching. The overall quality of life (QoL) score also showed a significant improvement during the study period; the mean score decrease found was 0.86 (CI 95 %, 0.54-1.17) (p < 0.001). Five of the seven domains included in the questionnaire decreased significantly: non-hayfever symptoms; practical problems; nasal symptoms; eye symptoms, and emotions. These QoL score decreases were found during the first 3 months and later remained stable at lower levels than at baseline. The content of guanine in dust samples decreased significantly from 2.17 0.75 to 1.43 0.68 (p < 0.001). No toxic adverse events were recorded. In conclusion, these results show an improvement in the quality of life and a clinical benefit of the long-term use of the new chemical acaricide tested, together with environmental measures, in the home of patients suffering from allergic respiratory pathology.

Amoxicillin-induced aseptic meningitis: 2 case reports and appraisal of the literature

No disponible

Enhancement of hazelnut extract for IgE testing by recombinant allergen spiking.

BACKGROUND: Hazelnuts are a common cause of food allergic reactions. Most hazelnut allergic individuals in central and northern Europe are sensitized to Cor a 1, a member of the PR-10 protein family, while the lipid transfer protein Cor a 8 acts as a major allergen in the south of Europe. Other allergens, including profilin and seed storage proteins, may be important in subgroups of patients. Reliable detection of specific IgE in the clinical diagnosis of food allergy requires allergen reagents with a sufficient representation of all relevant allergen components. Some reported observations suggest that natural hazelnut extract may not be fully adequate in this respect. METHODS: The capacity of immobilized natural hazelnut extract to bind Cor a 1-, Cor a 2- and Cor a 8-specific IgE and IgG antibodies was investigated by serum adsorption and extract dilution experiments and by the use of allergen specific rabbit antisera. All measurements were performed with the ImmunoCAP assay platform. RESULTS: The experimental results revealed an incomplete capacity of immobilized hazelnut extract to capture IgE antibodies directed to the major allergen Cor a 1. Spiking of hazelnut extract with recombinant Cor a 1.04 prior to solid phase coupling gave rise to significantly enhanced IgE antibody binding from Cor a 1 reactive sera. The spiking did not negatively affect the measurement of IgE to extract components other than Cor a 1. CONCLUSION: A hazelnut allergen reagent with enhanced IgE detection capacity can be generated by supplementing the natural food extract with recombinant Cor a 1.04.

An observational study on outgrowing food allergy during non-birch pollen-specific, subcutaneous immunotherapy.

BACKGROUND: Birch pollen-specific immunotherapy (SIT) decreases allergy to foods containing birch pollen-homologous allergens. Cross-reactivity was also observed between plane tree pollen and some vegetable foods. OBJECTIVE: The aim of this study was to evaluate the outgrowing of food allergy by patients suffering from vegetable food allergy associated with plane tree pollinosis (rhinoconjunctivitis and/or asthma) during plane tree pollen SIT. METHODS: An observational and prospective study was conducted in 16 adult patients suffering from vegetable food allergy (hazelnut, walnut, lettuce, peach and cherry) and from plane tree pollinosis receiving plane tree pollen SIT for 1 year. Open oral challenges with the implicated food were performed before and after SIT. Blood samples were drawn for measurement of pollen- and food-specific IgE and IgG4 before and after treatment. RESULTS: Plane tree SIT resulted in a significant decrease in food allergy, since the mean food quantity provoking objective symptoms increased from 2.19 to 13.74 g (p < 0.05), and 6 of the 11 patients tolerated the highest level (25 g) of the challenged food after plane tree SIT. Laboratory data also showed a decrease in IgE levels and an increase in IgG4 levels after immunotherapy. CONCLUSION: SIT with plane tree pollen has a positive impact on food allergy in plane tree pollen-allergic subjects.

Identification of a plane pollen lipid transfer protein (Pla a 3) and its immunological relation to the peach lipid-transfer protein, Pru p 3.

BACKGROUND: An association between plane tree pollen allergy and plant food allergy has been described, but the cross-reacting allergens have not yet been identified. The aim of this study was the identification of homologous non-specific lipid-transfer proteins (nsLTPs) in plane pollen, and to investigate its immunological relationship with the peach LTP, Pru p 3. METHODS: Three different patient groups were recruited in Spain: 22 plane pollen-allergic patients without food allergy (A), 36 plane pollen-allergic patients with peach allergy (B) and 10 peach-allergic patients without plane pollen allergy (C). Proteins from plane pollen extract were fractionated by ion-exchange and reversed-phase chromatography. Further methods applied were N-terminal amino acid sequence analysis, immunoblotting, enzyme allergosorbent test, CAP and basophil histamine release assays. RESULTS: A 10 kDa IgE-reactive protein was purified from plane pollen and identified as nsLTP. Pla a 3 was characterized as a minor allergen (27.3%) in plane pollen-allergic patients without food allergy (A) and as a major allergen in plane pollen-allergic patients with peach allergy (B) showing a prevalence of IgE-reactivity of 63.8%. Group B contained patients sensitized to Pru p 3 without IgE-reactivity to plane-LTP (16.6%). By contrast, Pla a 3 IgE-reactive patients without sensitization to Pru p 3 could be found (16.6%). The sera of patients sensitized to both LTPs (50%), Pla a 3 and Pru p 3, showed different biological activity in histamine release assay: depending on individual patient's sera tested, Pla a 3 showed a similar, a stronger or a weaker allergenic potency in comparison with Pru p 3. CONCLUSIONS: Plane LTP is a major allergen in plane pollen-allergic patients with peach allergy recruited in the Mediterranean area. The results of histamine release tests and different IgE-binding profiles pointed towards the existence of species-specific IgE epitopes. Likewise, no general conclusion on the sensitizer could be made.