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Heterogeneity of immune responses to various Aspergillus species in patients with allergic respiratory diseases
Abstract
In the present study, allergenic significance of thirteen species of Aspergillus and their allergenic and antigenic relationship was studied. Of the 3025 ID tests performed with the 13 species of Aspergillus on 289 patients suffering with allergic respiratory diseases, 627 (20.7%) were positive (1+ to 4+), 386 (12.8%) being significantly positive (2+ to 4+) . Of the 64 patients eliciting a positive cutaneous response to at least one species, 42(65.6%) were positive to 5 or less number of species while others showed a broad spectrum of positive skin reactivity to different Aspergillus extracts. In RAST inhibition assays using pooled sera ofpatients sensitive to A. tamarii dose related inhibition was produced by homologous as well as 5 of the 12 heterologous species. Similarly, in A. terreus RAST inhibition was observed with homologous and A. tamarii extracts only. Our results suggested the presence of both species specific as well as shared allergenic components among different Aspergillus species. In TDIEP experiments using rabbit antisera to A. tamarii and A. terreus extracts multiple precipitin bands were observed with the homologous extracts. However, only 1-2 bands were produced by 6 heterologous Aspergillus species in each system. Collectively, these results gave evidence that there is heterogeneity of immune response in the patients with allergic respiratory diseases to different species of Aspergillus and also in rabbits immunized with Aspergillus extracts.
... [12][13][14][15] Crude fungal extracts used for diagnostic purposes only provide information of the allergen source to which the patient has been sensitized. [16][17][18][19] However, the clinical sensitivity of patients depends on the number and type of IgE-binding proteins present in these extracts. 18,19 Hence, it is crucial to determine the profile of allergenic proteins in crude fungal extracts and identify the IgE reactivity profile of each patient. ...
... [16][17][18][19] However, the clinical sensitivity of patients depends on the number and type of IgE-binding proteins present in these extracts. 18,19 Hence, it is crucial to determine the profile of allergenic proteins in crude fungal extracts and identify the IgE reactivity profile of each patient. ...
... Of the various fungi, Aspergillus species have been reported to be one of the important inhalant allergens in different geographical regions of the world. [6][7][8][9][10][13][14][15][16]18,[20][21][22] The reported frequency of sensitization with various species of Aspergillus in patients of allergic respiratory diseases varies from 15.3-38% worldwide. 18,[23][24][25][26] A number of A fumigatus allergens have been cloned from cDNA/phage display libraries, characterized and purified as recombinant proteins. ...
Aspergillus species (A flavus and A niger) are important sources of inhalant allergens. Current diagnostic modalities employ crude Aspergillus extracts which only indicate the source to which the patient has been sensitized, without identifying the number and type of allergens in crude extracts. We report a study on the identification of major and minor allergens of the two common airborne Aspergillus species and heterogeneity of patients' IgE response to them. Skin prick tests were performed on 300 patients of bronchial asthma and/or allergic rhinitis and 20 healthy volunteers. Allergen specific IgE in patients' sera was estimated by enzyme allergosorbent test (EAST). Immunoblots were performed to identify major/minor allergens of Aspergillus extracts and to study heterogeneity of patients' IgE response to them. Positive cutaneous responses were observed in 17% and 14.7% of patients with A flavus and A niger extracts, respectively. Corresponding EAST positivity was 69.2% and 68.7%. In immunoblots, 5 allergenic proteins were identified in A niger extract, major allergens being 49, 55.4 and 81.5 kDa. Twelve proteins bound patients' IgE in A flavus extract, three being major allergens (13.3, 34 and 37 kDa). The position and slopes of EAST binding and inhibition curves obtained with individual sera varied from patient to patient. The number and molecular weight of IgE-binding proteins in both the Aspergillus extracts varied among patients. These results gave evidence of heterogeneity of patients' IgE response to major/minor Aspergillus allergens. This approach will be helpful to identify disease eliciting molecules in the individual patients (component resolved diagnosis) and may improve allergen-specific immunotherapy. Copyright© Summer 2015, Iran J Allergy Asthma Immunol. All rights reserved.
... Aspergillus species have been reported to be one of the most predominant constituents of aeromycoflora worldwide (4)(5)(6)(7)(8)(13)(14)(15)(16). Besides conidiospores (2-5 µm), its hyphal fragments, mostly conidiophores, are also present in the air throughout the year (17). ...
... Once inhaled, they either reach terminal airways or are deposited in large clusters in the upper respiratory tract and cause sensitization, resulting in allergic symptoms. The reported frequency of sensitization with Aspergillus species in patients with allergic diseases has varied from 15.3% to 38% worldwide (13,(18)(19)(20)(21)(22). As such, the knowledge of seasonal patterns of occurrence and concentrations of various Aspergillus species in a given geographical area becomes very important for proper diagnosis and management of Aspergillus hypersensitive patients. ...
... For studying allergenic significance of Aspergillus species, a majority of the air surveys worldwide have usually enumerated them up to genus level only (4,5,33,(40)(41)(42). However, some Aspergillus species have been consistently encountered in the air surveys conducted in Delhi metropolitan area, such as A. flavus, A. fumigatus, A. niger, and A. tamarii (4,13). In the present air survey, these four common species of Aspergillus were identified and quantified. ...
Airborne Aspergillus species are significant environmental components involved in the pathogenesis and persistence of allergic respiratory diseases. The detection and quantification of airborne allergens is important to elucidate the clinical implications of environmental exposure of patients suffering with allergic asthma and/or allergic rhinitis.
The authors report a simple volumetric approach to measure atmospheric concentration of four common airborne species of Aspergillus-A. flavus, A. fumigatus, A. niger, and A. tamarii.
As particulate aeroallergens may also exist in amorphous form in addition to morphologically identifiable fungal spores/hyphae, a volumetric technique using membrane filters was developed for simultaneous quantification of (a) viable Aspergillus counts, i.e., colony-forming units (cfu)/m(3), and (b) actual Aspergillus allergen content (ng/m(3)) in the air. Further, immunochemically quantified airborne Aspergillus allergens were compared with their corresponding colony counts.
The average monthly aerial counts of the four Aspergillus species recorded during the sampling year were A. flavus: 0.25-15.2 cfu/m(3); A. fumigatus: 1.25-15.6 cfu/m(3); A. niger: 0.75-16.0 cfu/m(3); and A. tamarii: 0.5-11.8 cfu/m(3) of air. Aerial Aspergillus allergen(s) concentration varied from species to species: A. flavus: 26.8-680.8 ng; A. fumigatus: 18.0-380.4 ng; A. niger: 28.2-1879.0 ng; and A. tamarii: 9.2-238.3 ng/m(3) of air. Seasonal distribution of airborne colony counts of the four species didn't correlate with their respective allergen content.
Aspergillus allergens were present in the air of Delhi area throughout the year with seasonal variations. The authors feel that by using the immunochemical technique it will be possible to measure actual exposure of patients to various airborne Aspergillus allergens.
... The most important fungi causing Abbreviations: A tamarii, Aspergillus tamarii; BSA, bovine serum albumin; GPBS, 50% glycerinated phosphate buffered saline; BCIP/NBT, 5-bromo-4-chloro-3-indolyl phosphate/Nitro blue tetrazolium chloride; NHV, non-allergic healthy volunteers; NHS, normal human serum; OD, optical density; PPS (Ata), pool of sera from A tamarii hypersensitive patients; EAST, enzyme allergosorbent test; SDS-PAGE, sodium dodecyl sulphate-polyacrylamide gel electrophoresis; SPT, skin prick test; TBS, Tris buffered saline. respiratory allergies include Aspergillus, Cladosporium, Penicillium, Alternaria, Epicoccum, Curvularia, Bipolaris, Candida, etc. Aggarwal et al., 2000;Agarwal and Shivpuri 1974;Crameri et al. 2008;Horner et al. 1995;Kurup and Banerjee 2000;Shivpuri and Agarwal 1969;Simon-Nobbe et al. 2008). ...
... Among the fungi, airborne Aspergillus species are sizable environmental components involved in the pathogenesis and persistence of allergic respiratory diseases (Aggarwal et al. 2000;G Model IMBIO-50612; No. of Pages 9 2 M. Vermani et al. / Immunobiology xxx (2010) xxx-xxx Crameri et al. 2008;Kurup and Banerjee 2000;Simon-Nobbe et al. 2008). The fungi grow outdoors as well as indoors, release large quantities of conidiospores and hyphal fragments, which act as sources of inhalant allergens (Crameri et al. 2008; Kurup and Banerjee 2000;Simon-Nobbe et al. 2008). ...
... The fungi grow outdoors as well as indoors, release large quantities of conidiospores and hyphal fragments, which act as sources of inhalant allergens (Crameri et al. 2008; Kurup and Banerjee 2000;Simon-Nobbe et al. 2008). The allergenic significance of various airborne Aspergillus species like A fumigatus, A flavus, A niger, A nidulans, A versicolor and A oryzae has been extensively studied (Aggarwal et al. 2000;Kurup and Banerjee 2000;Simon-Nobbe et al. 2008;Shen et al. 1999). A number of A fumigatus allergens have been cloned from cDNA/phage display libraries, characterized and purified as recombinant proteins (Crameri and Blaser 1996;Crameri et al. 2006;Vailes et al. 2001). ...
Aspergillus-derived inhalant allergens play an important role in the etiology of allergic respiratory diseases. In the present study, we investigated the allergenic potential of Aspergillus tamarii, quantified its airborne content, identified its major/minor allergens, evaluated heterogeneity of patients' IgE response to its allergens and cross-reactivity of its allergens with other Aspergillus allergens. Skin prick tests with A tamarii extract were performed on 300 patients of bronchial asthma/allergic rhinitis and 20 healthy volunteers. Sixty-six patients (22%) elicited positive cutaneous reactions to A tamarii extract. Only one of the 20 non-allergic healthy volunteer showed a mild positive cutaneous reaction. Allergen-specific IgE levels increased with increase in patients' cutaneous response (0% in negative to 100% in 3+/4+). The skin positivity and allergen-specific IgE levels were significantly higher in patients compared to healthy volunteers (P>0.05). However, no differences were found for these two parameters among patients of bronchial asthma, allergic rhinitis and bronchial asthma with allergic rhinitis. The airborne A tamarii allergen content was highest in February and October. A tamarii extract revealed at least 22 proteins (13.3-120 kDa). Seventeen of these proteins bound patients' IgE with six being major allergens (13.3, 23, 25, 34, 39.5, 43 kDa). Three major allergens (13.3, 34, 43 kDa) were found to cross-react with A flavus and one (34 kDa) with A niger. Our results revealed that A tamarii allergen(s) are present in the air, which might serve as important inhalant allergens in IgE-mediated allergic respiratory diseases.
... It required 9e12 ng self-proteins for 50% ELISA-inhibition. Individual patients' sera identified 23,28, 35, 38, 40, 49, 72, 78 and 97 kDa as major IgE binding components in PA extract. Nymph extract exhibited similar potency and protein profile to PA extract with 72 and 78 kDa proteins present in high intensities. ...
... PA extract and one of the commercial extract) was injected into the forearm of the patient raising a bleb of 2e3 mm. The reactions were graded after 20 min on the basis of wheal size of positive control i.e. histamine diphosphate (100 mg/ml) [23]. Blood was collected from patients showing marked positive skin reactions (wheal size equal to positive control and or more), sera separated and stored at À70 C. Informed consent of each patient was obtained for skin tests and blood collection. ...
... Proteins of 26, 31, 45, 51 kDa identified here can be designated as Per a 1, which shows variations in molecular weights [14]. Thus, PA extract contains allergens as identified in previous studies and also establishes the importance of few more allergenic components (23,35,40, 49 and 97 kDa). The disparity in IgE reactivity of present extract and the extracts used in previous investigations may be due to variability of allergenic components among extracts and also the serum sources [12]. ...
Commercial cockroach extracts for diagnosis and therapy show batch-to-batch variation. This study aimed to standardize Periplaneta americana extract based on major IgE binding components using hypersensitive patients' sera. Extracts were prepared in phosphate buffered saline (PBS) or NH(4)HCO(3), from freeze-dried or 37 degrees C dried material and compared with commercial extracts by immunobiochemical methods. Cockroach positive patients' sera were collected after intradermal tests and specific IgE enzyme linked immunosorbent assay (ELISA). Allergenic proteins were identified by western-blotting and potency of extracts determined by ELISA-inhibition. Adult P. americana extract from freeze dried source material in PBS (PA extract) resolved into 45 protein bands and showed 22 IgE binding components with pooled patients' sera. It required 9-12 ng self-proteins for 50% ELISA-inhibition. Individual patients' sera identified 23, 28, 35, 38, 40, 49, 72, 78 and 97 kDa as major IgE binding components in PA extract. Nymph extract exhibited similar potency and protein profile to PA extract with 72 and 78 kDa proteins present in high intensities. Commercial extracts exhibited only 6-11 IgE reactive bands compared to PA extract and required 40 folds or more protein for 50% ELISA-inhibition. PA extract from freeze-dried source material seems a potent allergen preparation with 9-major IgE binding components. It can be referred to upgrade the quality of commercial extracts exhibiting low potencies due to poor quality source material, inadequate extraction procedures and improper storage.
... [40][41][42] Among the different types of fungi, the Aspergillus species have been reported as important inhalant allergens in different geographical regions of the world. [14][15][16][43][44][45][46][47][48][49][50][51][52] Aspergillus species are the most common causes of invasive mold infections worldwide, especially in immunocompromised patients. A. fumigatus is the main etiologic agent of allergic, chronic, and invasive bronchopulmonary fungoides among the infections caused by Aspergillus species. ...
Allergies are pathological manifestations originating from a trigger-sensitized immune system. Aspergillus species have been reported to be one of the important inhalant allergens in different geographical regions of the world. House dust mite (HDM) allergens play a major role in causing allergic diseases. The emerging literature indicates the allergenicity and contribution of Aspergillus species and HDMs. Allergies erupt when innocuous foreign components are confused as foes by the immune surveillance. The incidence of fungal sensitization in patients with allergic respiratory diseases has been reported from 2.3% to even 80% in various studies worldwide. Human skin scales provide food for both mites and fungi. Fungi may either constitute a food supplement for mites or may have an indirect effect by decomposing human dander, thus making it more accessible for HDMs. There is a mutual relationship between fungi and HDMs. In addition to avoid exposure to an allergen as a secondary or tertiary preventive strategy, which is often not sufficiently effective against domestic mites, the treatment of mite allergy is mainly based on allergen-specific immunotherapy (AIT). Treatment with azole antifungal drugs in patients with severe asthma is effective and improves patient quality of life.
... The presence of allergen-specific IgE on mast cells is evident by weal and flare response after application of allergen extract on skin [68][69][70][71]. Weal diameter is compared with positive control and equal to control or more were considered as marked positive skin reactions [72]. If smaller weal as compared to positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. ...
Legumes belonging to Fabaceae family of the order Fabales are a rich and important source of proteins and many essential elements. Due to its nutritious elements, these are preferably included in human diet in most part of the world. But, unfortunately, IgE binding proteins have been identified in majority of legumes, and allergenic response to these legumes may range from mild skin reactions to life-threatening anaphylactic reaction. Overall, allergenicity due to consumption of legumes in decreasing order may be peanut, soybean, lentil, chickpea, pea, mung bean, and red gram. So far, several allergens from different legumes have been identified and characterized. Most of identified allergens belong to storage proteins family, profilins, or the pathogenesis-related proteins. Legumes also have property of immunological cross-reactivity among themselves and from other sources that also increases the severity of allergenic response to a particular legume. This review summarizes the currently available knowledge on legume allergy and describes the allergenic problems associated with different legumes. It also tries to explore about the legume allergens identified so far by different scientific groups. The culmination of knowledge about identification and characterization of allergens from different legumes will be helpful in diagnosis and treatment of allergy, for development of novel therapeutic strategies, for strict avoidance of particular legume in diet by susceptible individual and also to produce hypoallergenic cultivars of leguminous crop through conventional breeding or genetic modification.
... The presence of allergen-specific IgE on mast cells is evident by weal and flare response after application of allergen extract on skin68697071. Weal diameter is compared with positive control and equal to control or more were considered as marked positive skin reactions [72]. If smaller weal as compared to positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. ...
... Skin prick tests (SPT) were performed with allergen kit (Alicit India Private Limited) on allergic patients (n = 200) and normal healthy control (n = 10) at the Department of Pulmonary Medicine, Chhatrapati Shahuji Maharaj Medical University (CSMMU), Lucknow. Weal diameter equal to positive control or more were considered as marked positive skin reactions (Aggarwal et al., 2000). If smaller weal as compared to positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. ...
Allergy to chickpea or Garbanzo bean (Cicer arietinum) has been reported in the Indian population. Little information is found regarding allergenic events involved in the chickpea allergy; therefore, chickpea allergenicity assessment was undertaken. In vivo and ex vivo studies were carried out using BALB/c mice. Chickpea skin prick test positive patients have been used to extend this study in humans. Identification of allergens was carried out by simulated gastric fluids assay for pepsin resistant polypeptides and validated by IgE western blotting using chickpea sensitive humans and sensitized mice sera. Our data have shown the occurrence of a systemic anaphylactic reaction resulting in reduced body temperature after challenge along with significantly increased levels of IgE, IgG1, MMCP-1, CCL-2 as well as histamine. Further, increased Th1/Th2 (mixed) cytokine response was observed in spleen cell culture supernatants. Jejunum, lungs and spleen showed prominent histopathological changes specific for allergic inflammation. Immunoblotting with pooled sera of either sensitized mice or human sera recognized seven similar IgE binding polypeptides that may be responsible for chickpea induced hypersensitivity reactions. This study has addressed the allergenic manifestations associated with chickpea consumption and identifies the proteins responsible for allergenicity which may prove useful in diagnosis and management of allergenicity of legumes especially chickpea.
... Weal diameter equal to positive control or more were considered as marked positive skin reactions. 28 If smaller weal as compared with positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. Blood was collected from mustard SPT positive patients and normal healthy volunteers. ...
... Skin prick tests (SPT) were performed with allergen kit (Alicit India Private Limited) on allergic patients (n = 200) and normal healthy control (n = 10) at the Department of Pulmonary Medicine, Chhatrapati Shahuji Maharaj Medical University (CSMMU), Lucknow. Weal diameter equal to positive control or more were considered as marked positive skin reactions (Aggarwal et al., 2000). If smaller weal as compared to positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. ...
... Skin prick tests (SPT) were performed with allergen kit (Alicit India Private Limited) on allergic patients (n = 200) and normal healthy control (n = 10) at the Department of Pulmonary Medicine, Chhatrapati Shahuji Maharaj Medical University (CSMMU), Lucknow. Weal diameter equal to positive control or more were considered as marked positive skin reactions ( Aggarwal et al., 2000). If smaller weal as compared to positive control then +1, equal +2, larger 3 and for bigger with pseudopodia then +4. ...
Genetically modified (GM) mustard line (V4) with increased carotenoid content was compared with native mustard to find the difference in allergenic potential, if any. Simulated gastric fluid (SGF) digestibility of crude protein extract from GM as well as its native counterpart mustard crop was envisaged to understand the intended or unintended changes in GM crop along with IgE immunoblotting. BALB/c mice were used as model for allergenicity studies for monitoring total and specific IgE, specific IgG1, histamine level, histopathology, and systemic anaphylaxis score. Allergenicity of mustard was checked in humans by clinical history, skin prick test and IgE levels. Similar results were evident by significant increase in total IgE, specific IgE, IgG1, histamine levels, in GM and native mustard in comparison to control group. Prominent anaphylactic symptoms (score 2: 60%; score 3: 20%; score 4: 20% in native mustard and score 2: 40%; score 3: 40%; score 4: 20% in GM mustard) and eruptive histopathological changes were observed in both GM and native mustard when compared with controls. One protein of approximately 16 kDa was found stable up to 1 h in both GM as well as non GM mustard. IgE immunoblotting detected three protein components of approximately 29, 24 and 16 kDa in both GM and non GM varieties. Collectively, our data demonstrate substantially equivalent allergic responses against GM as well as its native counterpart. Therefore, the GM mustard may be as safe as its native counterpart with reference to allergenic responses.
Assessing the allergenicity and toxicity of genetically modified (GM) crops is essential before they become a regular part of our food supply. The present study aimed to assess the allergenicity of Brassica juncea (mustard) expressing choline oxidase (codA) gene from Arthrobacter globiformis that provides resistance against abiotic stresses.
SDAP, Farrp, and Swiss-Prot databases were used to study allergenicity of choline oxidase. Digestibility of choline oxidase was assessed in simulated gastric fluid (SGF). Specific immunoglobulin E (IgE) reactivity of native and GM mustard was compared by using enzyme-linked immunosorbent assay (ELISA) and skin tests in respiratory-allergic patients. Allergenicity of GM and native mustard proteins was compared in Balb/c mice.
Choline oxidase showed no significant homology with allergenic proteins in SDAP and Farrp databases. Cross-reactive epitope search showed a stretch similar to Hev b 6 having some antigenic properties. Purified choline oxidase showed complete degradation with SGF. Skin prick test of native and GM mustard extract on respiratory allergic patients showed significant correlation (P < 0.05). ELISA with 96 patients' sera showed comparable IgE reactivity. Balb/c mice immunized with native and GM mustard proteins showed low IgE response. Presensitized mice on intravenous challenge with Brassica extract showed no anaphylactic symptoms unlike ovalbumin (OVA) sensitization that showed anaphylactic reaction in mice. Lung histology of OVA-sensitized mice showed narrowing of airway and large eosinophilic infiltration, whereas native and GM Brassica extract showed normal airway.
Genetically modified mustard with the codA gene possessed allergenicity similar to that of native mustard and no enhancement of IgE binding was observed due to genetic manipulation.
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