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Background: Cross-reactivity reactions between allergenic polygalacturonases (PGs) from different biological sources, especially foods and pollens from the Oleaceae family, have been described using Salsola kali PG (Sal k 6). No PG from olive pollen has been characterized to date, hampering further knowledge about cross-reactions through PGs., Objectives: The aim of this work was to determine the potential allergenicity of the PG from olive pollen and clarify its role in cross-reactivity., Methods: A cDNA-encoding olive pollen PG sequence was subcloned into the pET41b vector and used to transform BL21(DE3) Escherichia coli cells to produce a His-tag fusion recombinant protein. The allergenic properties of olive pollen PG were determined by immunoblotting and ELISA in comparison to Sal k 6. The cross-reactivity potential of the protein with other pollen sources was analyzed by inhibition immunoassays., Results: The existence of other isoforms of Ole e 14 with different allergenicity was confirmed by proteomics and a meta-analysis of the recently reported olive genome. Sal k 6 showed a higher IgE recognition than Ole e 14 regardless of patient sensitization, suggesting the existence of more allergenic Ole e 14 isoforms in olive pollen. IgG and IgE inhibition assays supported the existence of cross-reactions between them and with other PGs from Oleaceae and Poaceae plant families., Conclusions: A new allergen from olive pollen, Ole e 14, has been identified, produced as a recombinant isoform, and structurally and immunologically characterized. Its role in cross-reactivity has been confirmed and, due to its smaller IgE binding capacity, it could have an important role for therapeutic purposes., (© 2018 S. Karger AG, Basel.)

Background: The incidence of Amaranthaceae pollen allergy has increased due to the desertification occurring in many countries. In some regions of Spain, Salsola kali is the main cause of pollinosis, at almost the same level as olive and grass pollen. Sal k 1 - the sensitization marker of S. kali pollinosis - is used in clinical diagnosis, but is purified at a low yield from pollen. We aimed to produce a recombinant (r)Sal k 1 able to span the structural and immunological properties of the natural isoforms from pollen, and validate its potential use for diagnosis., Methods: Specific cDNA was amplified by PCR, cloned into the pET41b vector and used to transform BL21 (DE3) Escherichia coli cells. Immunoblotting, ELISA, basophil activation and skin-prick tests were used to validate the recombinant protein against Sal k 1 isolated from pollen. Sera and blood cells from S. kali pollen-sensitized patients and specific monoclonal and polyclonal antisera were used., Results: rSal k 1 was produced in bacteria with a yield of 7.5 mg/l of cell culture. The protein was purified to homogeneity and structural and immunologically validated against the natural form. rSal k 1 exhibited a higher IgE cross-reactivity with plant-derived food extracts such as peanut, almond or tomato than with pollen sources such as Platanus acerifolia and Oleaceae members., Conclusions: rSal k 1 expressed in bacteria retains intact structural and immunological properties in comparison to the pollen-derived allergen. It spans the immunological properties of most of the isoforms found in pollen, and it might substitute natural Sal k 1 in clinical diagnosis., (© 2015 S. Karger AG, Basel.)

Background: Salsola kali is an Amaranthaceae weed with important repercussions for pollinosis in temperate areas. Ole e 1-like members are relevant allergens in pollen from different species. We aimed to characterize and produce as recombinant allergen S. kali Ole e 1-like protein., Methods: The natural allergen was purified at homogeneity after three chromatographic steps. Specific cDNA was sequenced and expressed in Pichia pastoris yeast. Structural relationships of natural and recombinant forms were carried out by 2D electrophoresis and spectroscopic analyses. Its immunological relevance was analyzed by ELISA and immunoblotting using an IgG antiserum and monoclonal antibodies specific to Ole e 1, as well as sera from 57 allergic patients recruited from two Spanish regions where this pollinosis is frequent., Results: The purified allergen, Sal k 5, is an acidic glycoprotein of 151 amino acid residues and 17,628 Da of molecular mass. Its amino acid sequence exhibits 68 and 32% identity with the allergens of Che a 1 and Ole e 1, respectively. The recombinant protein was correctly processed and its structural and immunologic equivalence to the natural form was proven. A sensitization frequency between 30 and 40% was observed in pollinic patients from the center and east coast of Spain., Conclusions: Sal k 5 is a member of the Ole e 1-like protein family which can be considered an important allergen from S. kali. Its inclusion in diagnosis protocols would allow the accurate defining of patients allergic to this pollen.

Background: Sensitivity to Chenopodiaceae is a frequent cause of allergic respiratory diseases in geographic areas where sensitization to Salsola kali and Chenopodium album has been reported. The objective of this study was to evaluate the pattern of sensitization to 3 Salsola species in patients residing on the Mediterranean coast of south-eastern Spain., Methods: S. kali, S. vermiculata and S. oppositifolia pollen extracts were prepared. Patients reporting respiratory and/or cutaneous symptoms were skin prick tested with the 3 Salsola extracts. Individuals with positive skin prick tests to at least 1 of the 3 Salsola species were included. Specific IgE was determined by direct ELISA. SDS-PAGE and 2-D analysis were conducted to elucidate the protein profile. The allergenic profile was investigated by immunoblot. Inhibition experiments were conducted to establish cross-reactivity between different species., Results: 246 patients were included. 237 patients (96.3%) tested positive to S. oppositifolia, 189 (76.8%) to S. kali and 185 (75.2%) to S. vermiculata. Protein profile and immunoblot demonstrated similar patterns in all extracts, except in low-molecular-weight allergens of S. oppositifolia. Immunoblot inhibition experiments demonstrated that most high-molecular-weight allergens of S. oppositifolia were inhibited by S. kali whereas low-molecular-weight allergens were totally inhibited only by C. album., Conclusions: This study confirms the allergenic importance of other Salsola species, especially S. oppositifolia. We have demonstrated that the 3 species show a high degree of cross-reactivity, but S. oppositifolia shares more allergenic similarities with C. album than S. kali.