Health hazards associated with bio-aerosol is gaining immense importance in recent time. However, studies linking the probable source characterization of bio-aerosol, and their potential effect as respiratory ailments on the local people are extremely rare over the Indian subcontinent as well as in global perspective. To fill our knowledge gap, we have conducted a 3-year study on fungal bio-aerosol characterization, possible source segregation, identification of health hazardous fungal species and their role in causing allergy from seven different micro-environments over a semi-rural site of lower Indo-Gangetic Plain, West Bengal, India. The result showed the highest fungal spore concentration as well as spore diversity in the crop field (4477 ± 1343 spores m −3) and the lowest in the classroom (1994 ± 651 spores m −3). Ascospore, basidiospore, Cladosporium, and Aspergilli group were the primary indoor spores. The marker species for the crop field were Curvularia and Drechslera, whereas Ganoderma was for the factory environment. Source profile of the fungal spore of different micro-environments indicates the domination of outdoor species, whereas enclosed microenvironments have been dominated with spores of indoor origin. Aspergillus, Curvularia, Penicillium, and Rhizopus oryzae showed maximum correlation with the local allergic rhinitis and asthmatic patients. In skin prick test, Aspergillus sp., contributing a maximum percentage of the total culturable fungi, was found to be an expected potent allergic candidate in both non-asthmatic and asthmatic patients, as well. This study will directly help the inhabitants to avoid the hazardous fungal sensitization in different micro-environments.

Aspergillus terreus is an allergenic fungus in addition to causing infections in both humans and plants. However, the allergens in this fungus are still unknown, limiting the development of diagnostic and therapeutic strategies. We used a proteomic approach to search for allergens, identifying sixteen allergens based on two-dimensional immunoblotting with A. terreus susceptible patient sera. We further characterized triosephosphate isomerase (Asp t 36), one of the dominant immunoglobulin E (IgE)-reactive proteins. The gene was cloned and expressed in E. coli. Phylogenetic analysis showed Asp t 36 to be highly conserved with close similarity to the triosephosphate isomerase protein sequence from Dermatophagoides farinae, an allergenic dust mite. We identified four immuno-dominant epitopes using synthetic peptides, and mapped them on a homology-based model of the tertiary structure of Asp t 36. Among these, two were found to create a continuous surface patch on the 3D structure, rendering it an IgE binding hotspot. Biophysical analysis indicated that Asp t 36 shows similar secondary structure content and temperature sensitivity with other reported triosephosphate isomerase allergens. In vivo studies using a murine model displayed that the recombinant Asp t 36 was able to stimulate airway inflammation, as demonstrated by an influx of eosinophils, goblet cell hyperplasia, elevated serum immunoglobulins and induction of Th2 cytokines. Collectively, our results reveal the immunogenic property of Asp t 36, a major allergen from A. terreus, and define a new fungal allergen more broadly. This allergen could serve as a potent candidate for investigating component resolved diagnosis and immunotherapy.

As this particular abbreviation is a fungal allergen name (Asp t 36) given by the WHO/IUIS, it should not be included in the abbreviation list of the manuscript.

This study is aimed to identify the allergens and determine their allergenic potency through immuno-biochemical methods from A. terreus, the most prevalent airborne allergenic fungus in West Bengal, India. METHODS: Proteins from a two-week old culture of A. terreus were extracted for determining its allergenicity by SPT, ELISA and histamine release assay. Protein profile was analyzed by 1D and 2D gel electrophoresis, followed by immunoblotting with 15 individual sensitized patients' sera and identification by mass spectrometry using MALDI-TOF/TOF. One of the major allergens, Triose phosphate isomerase (TPI) has been purified for bio-physical and biochemical studies, followed by in silico allergenicity assessment. The 3D structure of TPI was homology modelled by using crystal structure of TPI from Tenebrio molitor as template. Putative IgE binding peptides were synthesized and subsequently validated by in vitro experiments. RESULTS: Histamine release, specific IgE, and skin sensitivity were found in higher amount in susceptible patients. 2D immunoblot revealed sixteen IgE reactive spots which were identified by MALDI-TOF/TOF. Amongst these, TPI was purified by two stage chromatography. It demonstrated a stable structure upon thermal denaturation using circular dichroism. Predicted IgE binding epitopes mapped on homology modelled structure showed that they were surface accessible. Synthetic epitope containing peptides showed significantly high IgE binding and histamine release with hypersensitive patients' sera. CONCLUSIONS: This finding unearthed novel allergens from A terreus and suggests that TPI is a major allergen with respect to immunoreactivity. Further, B cell peptides, identified by in silico methods can be potentially used in drug designing, allergy diagnosis and therapy.

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