Identification, expression and characterisation of a major salivary allergen (Cul s 1) of the biting midge Culicoides sonorensis relevant for summer eczema in horses

Abstract

Salivary proteins of Culicoides biting midges are thought to play a key role in summer eczema (SE), a seasonal recurrent allergic dermatitis in horses. The present study describes the identification, expression and clinical relevance of a candidate allergen of the North American midge Culicoides sonorensis. Immunoblot analysis of midge saliva revealed a 66 kDa protein (Cul s 1) that was bound by IgE from several SE-affected (SE+) horses. Further characterization by fragmentation, mass spectrometry and bioinformatics identified Cul s 1 as maltase, an enzyme involved in sugar meal digestion. A cDNA encoding Cul s 1 was isolated and expressed as a polyhistidine-tagged fusion protein in a baculovirus/insect cell expression system. The clinical relevance of the affinity-purified recombinant Cul s 1 (rCul s 1) was investigated by immunoblotting, histamine release testing (HRT) and intradermal testing (IDT) in eight SE+ and eight control horses. Seven SE+ horses had rCul s 1-specific IgE, whereas only one control animal had IgE directed against this allergen. Furthermore, the HRT showed rCul s 1 induced basophil degranulation in samples from seven of eight SE+ horses but in none of the control animals. rCul s 1 also induced immediate (7/8), late-phase (8/8) and delayed (1/8) skin reactivity in IDT on all SE+ horses that had a positive test with the whole body extract (WBE) of C. sonorensis. None of the control horses showed immediate or delayed skin reactivity with rCul s 1, and only one control horse had a positive late-phase response, while several nonspecific late-phase reactions were observed with the insect WBE. Thus, we believe rCul s 1 is the first specific salivary allergen of C. sonorensis to be described that promises to advance both in vitro and in vivo diagnosis and may contribute to the development of immunotherapy for SE in horses.

Developing a preventive immunization approach against insect bite hypersensitivity using recombinant allergens: A pilot study

“…Today many of the causative allergens of IBH have been identified and produced as recombinant proteins. This provides the opportunity to develop an allergen specific immunotherapy against IBH (Langner et al, 2009;Schaffartzik et al, 2011;van der Meide et al, 2013).…”

“…The first salivary gland proteins that bind IgE from IBH affected horses to be identified and produced as recombinant proteins were derived from laboratory-produced species. These comprised one protein from Culicoides sonorensis (Langner et al, 2009) and eleven from Culicoides nubeculosus (Schaffartzik et al, 2010(Schaffartzik et al, , 2011. These two species are not very common in Europe, but the salivary proteins from them bind IgE from IBH horses with variable frequency.…”

Skin Immune System and Allergic Skin Diseases

“…7,77,87 Culicoides numbers increase with warm and humid weather, when wind is minimal (not strong fliers), and at dawn and dusk (prime feeding times). * The data on Simulium, Stomoxys, and Haematobia are mostly anecdotal and unconvincing.…”

“…68 In another study, IgE-binding proteins of 32 and 70 kDa (range of seven recognized proteins 13-70 kDa) were commonly recognized by horses with insect-bite hypersensitivity. 77 Peripheral blood leukocytes (primarily basophils) from horses with insect-bite hypersensitivity release leukotrienes when incubated with C. nubeculosis antigen. 7,68,69 A 66-kDa protein (Cul s1; a maltase) was reported to be a major allergen in salivary gland extracts from C. sonorensis.…”

Longitudinal analysis of allergen‐specific IgE and IgG subclasses as potential predictors of insect bite hypersensitivity following first exposure to Culicoides in Icelandic horses

“…Absence of the insects in winter leads to temporary remission. The causative allergens for IBH, salivary gland proteins from Culicoides , have been characterized at the molecular level and produced as recombinant proteins . These recombinant allergens (r‐allergens) bind serum IgE from IBH‐affected animals, but only rarely from healthy control horses .…”