IgE response to Aed al 13 and Aed al 14 recombinant allergens from Aedes albopictus saliva in humans.

Background: Mosquito bite is normally associated with mild allergic responses, but severe localized or systemic reactions are also possible. Reliable tools for the diagnosis of mosquito allergy are still unavailable. Here, we investigated the IgE response to 3 potential salivary allergens identified in the saliva of the tiger mosquito Aedes albopictus. Methods: Serum from 55 adult individuals (28 controls and 27 allergic people), were analysed using an in-house Enzyme Linked ImmunoSorbent Assay (ELISA) against the Salivary Gland Extract (SGE) and the recombinant proteins albD7l2 (Aed al 2), albAntigen5-3 (Aed al 13) and albLIPS-2 (Aed al 14). Results: Fifteen of the 27 (56%) individuals having hypersensitive reactions to mosquito bites had IgE serum levels recognizing SGE. Negative sera did not show detectable levels of IgE targeting the SGE from the most common sympatric mosquito Culex pipiens. Among the positive individuals, 2 subjects displayed IgE targeting Aed al 2 (13%), while IgE recognizing Aed al 13 and Aed al 14 were detected in ten (67%) and seven (47%) individuals, respectively. Two sera from non-hypersensitive subjects had detectable levels of IgE targeting Aed al 13, suggesting possible cross-reaction with the homologue salivary proteins of multiple mosquito species or, more generally, of hematophagous insects. Conclusions: Our results indicate that Aed al 13 and Aed al 14 hold the potential to be developed as tools for the diagnosis of allergy to Ae. albopictus bites. Such tools would facilitate epidemiological studies on tiger mosquito allergy in humans and might foster the development of further protein-based assays to investigate cross-species allergies.

Identification of risk factors of severe hypersensitivity reactions in general anaesthesia.

BACKGROUND: Hypersensitivity reactions to anaesthetic agents are rare but often severe, with a mortality ranging from 4 to 9% in IgE-mediated events. Identification of the risk factors may contribute to limit the incidence of these reactions. The aim of our study was to search for possible risk factors of severe perioperative hypersensitivity reactions in our study population. METHODS: For this study we retrospectively reviewed data from 193 patients who experienced drug hypersensitivity reactions during general anaesthesia. The diagnostic protocol consisted of 1) history of the reaction, 2) measurement of serum baseline tryptase and specific IgE-assays for latex, beta-lactams and succinylcholine, 3) skin tests for the agents listed in the anaesthesia chart and for others likely to be safe for future use, latex, and others medications administered during the perioperative period (i.e. antibiotics), 4) subdivision of our patients on the basis of two criteria: a) grade of severity of clinical reactions according to the Ring and Messmer classification; b) results of skin tests and/or serum specific IgE-assays. RESULTS: One hundred of 193 patients had reactions of grade I, 32/193 patients had reactions of grade II, 55/193 patients had reactions of grade III and 6/193 patients had reactions of grade IV. A diagnosis of IgE-mediated reaction was established in 55 cases (28.50%); the most common causes were neuromuscular blocking agents, followed by latex and beta-lactams. Severe reactions were associated with older age (p = 0.025), asthma (p = 0.042), history of hypertension (p = 0.001), intake of serum angiotensin converting enzyme inhibitor medication (p = 0.012) or serum angiotensin II antagonist (p = 0.033), higher levels of basal tryptase (p = 0.0211). Cardiovascular symptoms (p = 0.006) and history of hypersensitivity to antibiotics (p = 0.029) were more frequently reported in IgE-mediated reactions. CONCLUSIONS: We confirmed the relevance of several clinical features as risk factors for anaphylactic reactions induced by anaesthetic agents: older age, asthma, hypertension and antihypertensive drugs. We observed increased levels of serum basal tryptase in severe reactions: this finding may signify that this biomarker is useful for the identification of patients at risk.

Anti-Amoxicillin Immunoglobulin E, Histamine-2 Receptor Antagonist Therapy and Mast Cell Activation Syndrome Are Risk Factors for Amoxicillin Anaphylaxis.

BACKGROUND: ß-Lactam antibiotics (mainly amoxicillin, AX) are the drugs that most frequently induce systemic drug allergy reactions. OBJECTIVE: We attempted to identify the risk factors associated with systemic reactions to AX. METHODS: All patients who were referred to our department for suspected hypersensitivity reactions to AX over a 6-month period were evaluated for anti-AX immunoglobulin E (IgE) levels and skin-test positivity for ß-lactams. Age, sex, concomitant diseases, therapies, total IgE, serum tryptase levels and signs and symptoms suggesting mast cell activation syndrome (MCAS) were analyzed in relation to the severity of the reaction in accordance with the Mueller classification. RESULTS: Sixty-seven patients were selected: 39 with mild reactions such as cutaneous or gastrointestinal symptoms (grades I and II) and 28 with severe systemic reactions (grades III and IV). Anti-AX IgE levels and total IgE were significantly higher in severe reactions than in mild ones (p < 0.00005, p = 0.0037). Treatment with histamine-2 receptor antagonists (anti-H2) was significantly related to severe reactions (p = 0.007). No significant correlations were found between the severity of the reactions and dyslipidemia or levels of angiotensin-converting enzyme and tryptase. CONCLUSION: Anti-AX IgE levels were the most significant immunological parameter distinguishing patients who presented with severe reactions to AX and those with mild reactions. Higher values of total IgE, the use of gastroprotective drugs and signs and symptoms suggesting an MCAS significantly increased the odds ratio of having a severe reaction. The risk of serious adverse reactions to AX increased in older patients and in males, but this trend was not significant.

Accuracy of a questionnaire for identifying respiratory allergies in epidemiological studies.

BACKGROUND: The assessment of allergic asthma (AA) and allergic rhinitis (AR) in epidemiological studies is often challenging. We performed a cross-sectional study to test the accuracy of a Questionnaire aimed at Identifying subjects with Respiratory Allergy (QIRA) in a simple and fast way. METHODS: One hundred-thirty subjects, 18-76 years of age, admitted consecutively at the Allergy Center of the Niguarda Ca` Granda Hospital of Milan were included. The questionnaire (index test) investigated the presence of AA and AR with seven questions enquiring history of symptoms, diagnosis made by a doctor, allergy tests performed, and treatments. After completing the questionnaire, all subjects were subsequently diagnosed by an allergist (reference standard). RESULTS: The accuracy of the questionnaire for the diagnosis of AA and AR was high (sensitivity 94.7% [95% confidence interval CI: 74.0-99.9] and specificity 99.1% [95% CI 95.1-100.0] for AA; sensitivity 82.8% [95% CI 71.3-91.1] and specificity 98.5% [95% CI 91.8-100.0] for AR). CONCLUSION: The questionnaire significantly distinguished subjects with respiratory allergy from those without. The QIRA represents a valid and accurate tool for classifying subjects as having or not AA and/or AR in epidemiological studies.

Anti-rPru p 3 IgE levels are inversely related to the age at onset of peach-induced severe symptoms reported by peach-allergic adults.

Sensitisation to peach lipid transfer protein (LTP; Pru p 3) is significantly associated with severe allergic symptoms in adults, but little is known about the age at onset of peach allergy. We investigated a possible correlation between specific IgE levels to Pru p 3 and the age at onset of peach allergy. One hundred and forty-eight patients allergic to peach were divided into 6 classes according to the age at onset. Sera were analyzed for IgE antibodies to peach, rPru p 3, rPru p 1, rPru p 4, rBet v 1, rBet v 2, total IgE titre, and tryptase; all collected data were statistically analysed. A significant inverse correlation was found between the age at onset of peach allergy and anti-rPru p 3 IgE levels at diagnosis (p < 0.0005; Spearman's ρ = -0.3833). In contrast, the age at onset was directly correlated with both anti-rPru p 1 IgE levels (p = 0.0001; Spearman's ρ = 0.3197) and anti-rBet v 1 IgE levels (p = 0.0006; Spearman's ρ = 0.2914) at diagnosis. No correlations were detected between the reported age at onset and anti-peach, anti-rPru p 4, anti-rBet v 2 IgE and total IgE values and serum tryptase levels. At diagnosis, when peach allergy starts at a younger age, it is likely associated with Pru p 3 sensitisation, and the younger the onset, the higher the IgE titres. When peach allergy starts at an older age, it is more likely the result of cross-reactivity to Bet v1.

Rice allergy demonstrated by double-blind placebo-controlled food challenge in peach-allergic patients is related to lipid transfer protein reactivity.

BACKGROUND: The risk factors for sensitisation to rice and the involved allergens are still partially unknown. In this study we evaluated the clinically relevant aspects of rice allergy in DBPCF-positive patients, the major rice allergens, the severity of peach- and rice-induced symptoms in respect to Pru p 3 sensitisation and the role of anti-rPru p 3 IgE levels as a risk factor for rice allergy. METHODS: In 148 peach-allergic subjects, patients with allergic reactions to rice and rice-positive serum IgE were selected. Symptoms were verified by double-blind placebo-controlled food challenges (DBPCFCs), performed at a maximum dosage of 25 g. Rice allergens, identified by IgE immunoblotting, were characterised by N-terminal amino acid sequencing. The relationship between anti-rPru p 3, 1 and 4 IgE levels and rice symptoms were statistically analysed. RESULTS: Eight out of 10 recruited rice-allergic patients had positive DBPCFCs, while 2 patients were not challenged due to their previously documented severe reactions. All patients with rice-induced symptoms were Pru p 3 positive and presented with higher anti-rPru p 3 levels than the rice-sensitised but tolerant patients. A 9-kDa lipid transfer protein, which was highly homologous to Pru p 3, was identified as the major rice allergen and elicited a positive response in all of the patients. Five patients reacted to a putative 15- to 17-kDa rice allergenic protein, and 3 patients reacted to an [alpha]-amylase/subtilisin inhibitor that was approximately 20 kDa. CONCLUSION: Rarely, allergic reactions to rice can arise in patients with peach allergies who are sensitised to Pru p 3, particularly in patients with high anti-rPru p 3 IgE levels.

Fennel allergy is a lipid-transfer protein (LTP)-related food hypersensitivity associated with peach allergy.

Fennel allergy has been rarely reported, and the association with peach allergy has never been described. Our aim was to (i) study the correlation between symptom severity of peach and fennel and (ii) identify fennel allergens and the role of rPru p 3 antibodies in severe reactions to fennel. In 148 patients with peach allergy, we investigated 58 patients with symptoms and IgE antibodies positive to fennel. IgE to rPru p 1, 3, and 4 and rBet v 1, 2, and 4 were measured by immunoblotting, and the N-terminal amino acid sequences and relevant allergens were determined. We found significant association between severe reactions to fennel and peach (p = 0.0009). A major allergen was ~9 kDa lipid-transfer protein (LTP), cross-reactive with Pru p 3, a 15 kDa protein identified as a pathogenesis-related protein 1 of the Bet v 1 family. In conclusion, peach and fennel severe allergic symptoms are significantly related, and LTP is a major fennel allergen. Fennel should be included in the LTP syndrome.

Pru p 3-sensitised Italian peach-allergic patients are less likely to develop severe symptoms when also presenting IgE antibodies to Pru p 1 and Pru p 4.

BACKGROUND: The roles played by different peach allergens with respect to symptom severity have not been completely ascertained. We have evaluated the diagnostic efficacy of peach recombinant allergens ImmunoCAP compared to peach in the identification of subjects at an increased risk for severe reactions to peaches. METHODS: 148 peach-allergic patients were divided based on their symptom severity into 2 groups: mild oral allergy syndrome (OAS) and severe OAS. Anti-rPru p 1, 3 and 4 IgE levels were measured. Statistical analyses were carried out using parametric and non-parametric tests. RESULTS: anti-rPru p 1 and anti-rPru p 4 IgE levels were significantly higher in patients with mild OAS than in patients with severe OAS (p = 0.0001); in contrast, anti-rPru p 3 IgE levels were significantly higher in patients with severe OAS than in patients with mild OAS (p < 0.00005). Moreover, we found that any unitary increase in anti-rPru p 1 IgE values corresponded to a 2.48% reduction in the odds of having severe OAS (p = 0.048), whereas any unitary increase in anti-rPru p 3 IgE values corresponded to a 9.02% increase in the probability of having severe OAS (p = 0.001). Unexpectedly, we found that patients positive to rPru p 3 as well as rPru p 1 and 4 demonstrated a significant reduction of the odds of developing severe symptoms than those positive to rPru p 3 alone. Anti-rPru p 3 IgE levels were a significantly better indicator than anti-peach IgE values (p = 0.016) of patients with the highest risk for severe OAS. A cutoff of 2.69 kUA/l for anti-rPru p 3 IgE values better discriminated peach-allergic patients at a higher risk for symptoms. CONCLUSIONS: Italian patients with positive anti-rPru p 1, 4 and 3 IgE levels seemed less likely to experience the clinical effects of high anti-rPru p 3 IgE values.

Green bean (Phaseolus vulgaris): a new source of IgE-binding lipid transfer protein.

Green beans belong to the Fabaceae family, which includes widely consumed species, such as beans, peanuts, and soybeans. In the literature, few cases have described allergic reactions upon the exposure to green bean boiling steam or ingestion. Here, we describe five patients reporting documented adverse reactions upon the ingestion of cooked green beans, and we characterize the responsible allergen. Fresh and cooked green beans were tested by a prick + prick technique. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and IgE immunoblotting were performed with boiled vegetable extract, and the N-terminal sequence of the immunoreactive protein was obtained by analyzing the excised band in a protein sequencer. Immunoblotting inhibition of cooked green bean with in-house-purified peach lipid transfer protein (LTP) Pru p 3 was performed. An interesting green bean protein was chromatographically purified, tested with a pool serum, and inhibited with Pru p 3. Moreover, its molecular mass was determined by mass spectrometry. Prick + prick tests with raw and cooked green beans were positive for all of the patients. IgE immunoblotting showed that all of the patients reacted toward a unique IgE-binding protein at about 9 kDa. The obtained N-terminal sequence revealed the following amino acids: Ala-Ile-Ser-X-Gly-Qln-Val-Thr-Ser-Ser-Leu-Ala, corresponding to an LTP. A complete inhibition of the IgE binding to this protein, in both raw and purified extract, was obtained by purified peach Pru p 3, confirming previous IgE immunoblotting results.

Tomato allergy: detection of IgE-binding lipid transfer proteins in tomato derivatives and in fresh tomato peel, pulp, and seeds.

There is an increasing consumption of tomatoes worldwide: fresh in salads, cooked in household sauces, or industrially processed. Although many tomato allergens have been identified, there is no information in the literature on the allergenic components found in commercial tomato products. The primary aim of the study was to evaluate the allergenic profile of commercial tomato products by skin prick tests (SPTs) and IgE/immunoblotting in tomato-allergic subjects. The secondary end point was the study of the IgE-binding profile of tomato peel, pulp, and seeds. Forty tomato-allergic patients, reporting oral allergy syndrome (OAS) at different grades of severity for fresh and, in some cases, also for cooked tomato, were selected on the basis of positive tomato allergy history or open food challenge (OFC). They were evaluated by SPTs with different experimental tomato extracts. SDS-PAGE/immunoblotting was performed to detect tomato allergens, which were then identified by Edman degradation. Twenty-three patients (57.5%) presented first-grade OAS at the OFC, whereas 17 (42.5%) reported severe symptoms. Ten of these 17 patients (25%) reported allergic reactions to cooked tomatoes; in immunoblotting tests, their sera reacted only to lipid transfer protein (LTP). In commercial products, LTP was the only detectable allergen. In contrast to other LTP-containing fruits, in tomato, an IgE-binding LTP was identified not only in the peel but also in the pulp and seeds. This study demonstrates that, in fresh tomato, different LTP isoforms are present and allergenic. Industrial tomato derivatives still contain LTP, thus presenting a problem for LTP-allergic patients.

Effects of specific immunotherapy on the B7 family of costimulatory molecules in allergic inflammation.

The effect of allergen-specific immunotherapy (IT) on Ag presentation and T lymphocyte stimulation was evaluated by verifying the expression of costimulatory molecules in allergic patients. Thus, CD28 and CTLA-4, B7, and B7-H molecules on immune cells, as well as cytokine production, were analyzed in and out of the pollination period in 30 patients allergic to Betulaceae that had or had nor undergone specific IT. Results showed that IT attenuated the increase in the percentage of CD28(+)CD4 T cells and the decrease in the percentage of CTLA-4(+)CD4(+) T cells seen in untreated individuals. CD19(+)/CD80, CD19(+)/CD86(+), and CD14(+)/CD80(+) APCs were significantly augmented during pollination in unvaccinated individuals. B7-H1-expressing monocytes (CD14(+)) and B lymphocytes (CD19) as well as CD14 and CD19 B7-H1(+)/IL-10(+) APC were augmented in Betulaceae Ag-stimulated cell cultures of vaccinated patients independently of pollination, and were further increased in these individuals during pollination. As a result, the IL-10-IFN-gamma ratio in CD4(+), CD14(+), and CD19(+) cells increased in vaccinated patients, but decreased in unvaccinated individuals during pollination. These data clarify the cellular and molecular basis underlying the recent observation that peripheral expansion of IL-10-producing cells is associated with successful IT. B7-H1 could be an optimal target for IT of allergic diseases using mAbs.