Submitted to: International Journal for Parasitology
Publication Type: Peer reviewed journal
Publication Acceptance Date: 6/13/2008
Publication Date: 1/15/2009
Citation: Langner, K.F., Jarvis, D.L., Schuberth, H., Nimtz, M., Heselhaus, J.E., Mcholland, L.E., Leibold, W., Drolet, B.S. 2009. Identification, expression, and characterization of a major salivary allergen (Cul s 1) of the biting midge Culicoides sonorensis relevant for summer eczema in horses. International Journal for Parasitology. Vol 39 (2009) 243–250. Interpretive Summary: Summer eczema (SE) is a seasonal recurrent allergic dermatitis in horses thought to be caused by the repeated biting of Culicoides midges. Salivary proteins of these midges are thought to play a key role in the induction of this hypersensitivity. A candidate allergen in the saliva of the North American species Culicoides sonorensis was identified, expressed, purified, and its clinical relevance was examined in horses. Horses affected with SE reacted to the protein, named rCul s 1 whereas control horses did not. Thus, rCul s 1 is the first described recombinant allergen of Culicoides spp. that may greatly improve in vitro and in vivo allergy diagnosis and may contribute to future immunotherapy of SE in horses.
Technical Abstract: Salivary proteins of Culicoides biting midges are thought to play a key role in the induction of summer eczema (SE), a seasonal recurrent allergic dermatitis in horses. The present study describes the identification of a candidate allergen in artificially collected saliva of the North American species Culicoides sonorensis by immunoblot analysis and mass spectrometry. This candidate was expressed as a polyhistidine-tagged fusion protein in a baculovirus/insect cell expression system, purified using cobalt affinity chromatography, and its clinical relevance was investigated by immunoblotting, histamine release testing (HRT) and intradermal testing (IDT) in eight SE-affected (SE+) and eight control horses. Analysis of native saliva revealed seven proteins ranging from 90 to 15 kDa that were bound by immunoglobulin E (IgE) of horse sera. One immunoreactive protein of 66 kDa (Cul s 1) was further characterized by internal peptide sequencing and identified as maltase, an enzyme involved in sugarmeal digestion. Expressed and purified recombinant Cul s 1 (rCul s 1) was detected by specific IgE in sera of seven SE+ horses whereas only one control animal had rCul s 1-specific IgE. The HRT showed rCul s 1 induced basophil degranulation for all SE+ horses and revealed a considerable higher sensitivity for the recombinant allergen as compared to whole body extracts (WBE) of C. sonorensis. In IDT, rCul s 1 induced immediate, late-phase and delayed skin reactivity in all SE+ horses that had a positive test with Culicoides WBE. None of the control horses showed reactivity with rCul s 1 in HRT and IDT although unspecific reactions were observed for WBE in both test systems. Thus, rCul s 1 is the first described recombinant allergen of Culicoides spp. that may greatly improve in vitro and in vivo allergy diagnosis and may contribute to future immunotherapy of SE in horses.