Immunochemical Characterization of Prosopis Juliflora Pollen Allergens and Evaluation of Cross-Reactivity Pattern with the Most Allergenic Pollens in Tropical Areas

Allergy to Prosopis juliflora (mesquite) pollen is one of the common causes of respiratory allergy in tropical countries. Mesquite is widely used as street trees in towns and ornamental shade trees in parks and gardens throughout arid and semiarid regions of Iran. The inhalation of mesquite pollen and several species of Amaranthus/Chenopodiaceae family is the most important cause of allergic respiratory symptoms in Khuzestan province. This study was designed to evaluate IgE banding proteins of mesquite pollen extract and its IgE cross-reactivity with other allergenic plants. Twenty patients with allergic symptoms and positive skin prick tests (SPT) for mesquite pollen extract participated in the study. Crude pollen extract was prepared from local mesquite trees and used for the evaluation of allergenic profiles of P. juliflora pollen extract by Sodium dodecylsulfate polyacrylamide gel electrophoresis (SDS-PAGE) and IgE-immunoblotting. There were several protein bands in mesquite pollen extract using SDS-PAGE with the approximate range of molecular weight of 10-85 kDa. The most frequent IgE reactive bands among the patients' sera were approximately 20 and 66 kDa. However, there were other IgE reactive protein bands among the patients' sera with molecular weights of 10, 15, 35, 45, 55 and 85 kDa. Inhibition experiments revealed high IgE cross-reactivity between mesquite and acacia. There are several IgE-binding proteins in P. juliflora pollen extract. Results of this study indicate that proteins with a molecular weight of 10 to 85 kDa are the major allergens in P. juliflora pollen extract

Allergy to Salsola Kali in a Salsola Incanescens-rich Area: Role of Extensive Cross Allergenicity

Background: Pollens from the Salsola spp. are an important source of respiratory allergy in tropical countries. Our aim was to characterize the IgE binding proteins of S. incanescens pollen extract and study its cross-reactivity with S. kali pollen allergens. Methods: Prick tests with S. kali and S. incanescens pollen extracts were performed on eight respiratory allergy patients from Mashhad, Northeast Iran. The antigenic profiles and IgE-binding patterns of S. kali and S. incanescens pollen extracts were compared by SDS-PAGE and Western blotting, using individual sera from the salsola pollen-sensitive patients. Cross-reactivity of proteins in the two weeds was assessed by IgE- immunoblotting inhibition. Results: S. kali and S. incanescens pollen extracts showed similar IgE-binding profiles in Western blotting. The IgE binding components of 39, 45, 66 and 85 kDa were detected in both pollen extracts. Furthermore, inhibition of the immunoblots revealed extensive inhibition of IgE binding to proteins and a close relationship between these two weeds allergens. Conclusions: S. incanescens pollen is a potent allergen source with several IgE binding components that shows a close allergenic relationship with S. kali. Our results suggest that in S. incanescens-rich areas, S. kali pollen extracts could be used as a diagnostic reagent for allergic patients to S. incanescens pollen

Designing a T-cell epitope-based vaccine using in silico approaches against the Sal k 1 allergen of Salsola kali plant

Abstract Allergens originated from Salsola kali (Russian thistle) pollen grains are one of the most important sources of aeroallergens causing pollinosis in desert and semi-desert regions. T-cell epitope-based vaccines (TEV) are more effective among different therapeutic approaches developed to alleviate allergic diseases. The physicochemical properties, and B as well as T cell epitopes of Sal k 1 (a major allergen of S. kali) were predicted using immunoinformatic tools. A TEV was constructed using the linkers EAAAK, GPGPG and the most suitable CD4+ T cell epitopes. RS04 adjuvant was added as a TLR4 agonist to the amino (N) and carboxyl (C) terminus of the TEV protein. The secondary and tertiary structures, solubility, allergenicity, toxicity, stability, physicochemical properties, docking with immune receptors, BLASTp against the human and microbiota proteomes, and in silico cloning of the designed TEV were assessed using immunoinformatic analyses. Two CD4+ T cell epitopes of Sal k1 that had high affinity with different alleles of MHC-II were selected and used in the TEV. The molecular docking of the TEV with HLADRB1, and TLR4 showed TEV strong interactions and stable binding pose to these receptors. Moreover, the codon optimized TEV sequence was cloned between NcoI and XhoI restriction sites of pET-28a(+) expression plasmid. The designed TEV can be used as a promising candidate in allergen-specific immunotherapy against S. kali. Nonetheless, effectiveness of this vaccine should be validated through immunological bioassays

Identification of a New Allergen from Amaranthus retroflexus Pollen, Ama r 2

Background: Pollinosis from Amaranthus retroflexus pollen is a common cause of respiratory allergy in Iran with a high positive rate (68.8%) among Iranian allergic patients. The aim of the present study was to evaluate the allergenicity of the A. retroflexus pollen profilin. Methods: Using sera from twelve patients allergic to A. retroflexus pollen, IgE-binding proteins from the A. retroflexus pollen extract was identified by immunoblotting. The cDNA of A. retroflexus pollen profilin was amplified, then cloned into the pET-21b (+) vector, expressed in Escherichia coli, and finally purified by metal affinity chromatography. The IgE-binding capacity of the recombinant protein was then analyzed by the ELISA, immunoblotting, and inhibition assays, as well as by the skin prick test (SPT). Results: Immunoblotting results indicated a 14.6 kDa protein with IgE-reactivity to 33% (4/12) among A. retroflexus pollen-allergic patients. Nucleotide sequencing of the cDNA revealed an open reading frame of 399 bp encoding for 133 amino acid residues which was belonged to the profilin family and designated as Ama r 2. A recombinant Ama r 2 (rAma r 2) was then produced in E. coli as a soluble protein which showed a strong IgEreactivity via ELISA confirmed by the SPT. Inhibition experiments revealed high IgE cross-reactivities with the profilins from other plants. Conclusions: The profilin from the A. retroflexus pollen, Ama r 2, was firstly identified as an allergen. Moreover, rAma r 2 was produced in E. coli as a soluble immunoreactive protein with an IgE-reactivity similar to that of its natural counterpart