Cloning, production and characterization of antigen 5 like proteins from Simulium vittatum and Culicoides nubeculosus, the first cross-reactive allergen associated with equine insect bite hypersensitivity.
Abstract: Insect bite hypersensitivity (IBH) is an IgE-mediated seasonal dermatitis of the horses associated with bites of Simulium (black fly) and Culicoides (midge) species. Although cross-reactivity between Simulium and Culicoides salivary gland extracts has been demonstrated, the molecular nature of the allergens responsible for the observed cross-reactivity remains to be elucidated. In this report we demonstrate for the first time in veterinary medicine that a homologous allergen, present in the salivary glands of both insects, shows extended IgE cross-reactivity in vitro and in vivo. The cDNA sequences coding for both antigen 5 like allergens termed Sim v 1 and Cul n 1 were amplified by PCR, subcloned in high level expression vectors, and produced as [His](6)-tagged proteins in Escherichia coli. The highly pure recombinant proteins were used to investigate the prevalence of sensitization in IBH-affected horses by ELISA and their cross-reactive nature by Western blot analyses, inhibition ELISA and intradermal skin tests (IDT). The prevalence of sensitization to Sim v 1 and Cul n 1 among 48 IBH-affected horses was 37% and 35%, respectively. In contrast, serum IgE levels to both allergens in 24 unaffected horses did not show any value above background. Both proteins strongly bound serum IgE from IBH-affected horses in Western blot analyses, demonstrating the allergenic nature of the recombinant proteins. Extended inhibition ELISA experiments clearly showed that Sim v 1 in fluid phase is able to strongly inhibit binding of serum IgE to solid phase coated Cul n 1 in a concentration dependent manner and vice versa. This crucial experiment shows that the allergens share common IgE-binding epitopes. IDT with Sim v 1 and Cul n 1 showed clear immediate and late phase reactions to the allergen challenges IBH-affected horses, whereas unaffected control horses do not develop relevant immediate hypersensitivity reactions. In some horses, however, mild late phase reactions were observed 4h post-challenge, a phenomenon reported to occur also in challenge experiments with Simulium and Culicoides crude extracts probably related to lipopolysaccaride contaminations which are also present in E. coli-expressed recombinant proteins. In conclusion our data demonstrate that IgE-mediated cross-reactivity to homologous allergens, a well-known clinically relevant phenomenon in human allergy, also occurs in veterinary allergy.
Copyright 2010 Elsevier B.V. All rights reserved.
Publication Date: 2010-04-28 PubMed ID: 20537727DOI: 10.1016/j.vetimm.2010.04.012Google Scholar: Lookup
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- Journal Article
- Research Support
- Non-U.S. Gov't
- Allergies
- Amino Acid Sequence
- Animal Science
- Antigen
- Biotechnology
- Cloning
- Enzyme-Linked Immunosorbent Assay (ELISA)
- Equine Diseases
- Equine Health
- Escherichia coli
- Immune Response
- Immunoglobulin E
- Immunology
- In Vitro Research
- In Vivo
- Insect Bite Hypersensitivity
- Protein
- Veterinary Medicine
- Veterinary Research
- Western Blot
Summary
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This research article explores how the antigens found in the saliva of certain insect species can cause a type of allergic reaction in horses known as insect bite hypersensitivity (IBH). The researchers have isolated and studied these allergens, revealing their molecular nature and detailing a high rate of sensitization among horses with IBH.
Introduction to the Research
- The study investigates Insect Bite Hypersensitivity (IBH), which is a type of allergic reaction in horses caused by bites from the insects Simulium (black fly) and Culicoides (midge).
- Though it was known that horses display a cross-reactive immune response to the saliva of these two insects, this paper presents the first detailed analysis of the molecular nature of these allergens.
- Specifically, the researchers identify an allergen common to both insects, which accounts for the cross-reactive nature of the allergic response.
Methodology
- The research involves cloning and producing the allergenic proteins (Sim v 1 and Cul n 1) found in the saliva of Simulium and Culicoides.
- These were done using high level expression vectors and the bacterium Escherichia coli as host for the production.
- The allergenic proteins produced were then used to assess sensitization among 48 horses with IBH by tests such as ELISA, Western Blot analyses, inhibition ELISA and intradermal skin tests (IDT).
Results
- Findings revealed that 37% and 35% of the horses with IBH were sensitized to Sim v 1 and Cul n 1, respectively.
- The serum IgE levels (an indicator of allergic reaction) to both allergens in 24 unaffected horses did not show any value above background, indicating no sensitization.
- Both the allergenic proteins strongly bound serum IgE from IBH-affected horses demonstrating their allergenic nature.
- The allergens were shown to share common IgE-binding sites, explaining the cross-reactive nature of the immune response in IBH-affected horses.
- IDT with Sim v 1 and Cul n 1 showed clear immediate and late phase reactions to the allergen challenges in IBH-affected horses.
Conclusion
- The study concludes that, like in human allergies, cross-reactivity can occur in veterinary allergies too.
- The findings also underline the need for further investigations on the nature of the common allergenic proteins from different insect species, and how these can be used to better understand and manage allergies.
Cite This Article
APA
Schaffartzik A, Marti E, Crameri R, Rhyner C.
(2010).
Cloning, production and characterization of antigen 5 like proteins from Simulium vittatum and Culicoides nubeculosus, the first cross-reactive allergen associated with equine insect bite hypersensitivity.
Vet Immunol Immunopathol, 137(1-2), 76-83.
https://doi.org/10.1016/j.vetimm.2010.04.012 Publication
Researcher Affiliations
- Swiss Institute of Allergy and Asthma Research (SIAF), University of Zürich, Obere Strasse 22, CH-7270 Davos, Switzerland.
MeSH Terms
- Allergens / genetics
- Allergens / immunology
- Animals
- Ceratopogonidae / immunology
- Cloning, Molecular
- Cross Reactions
- Enzyme-Linked Immunosorbent Assay
- Horse Diseases / etiology
- Horses
- Hypersensitivity / etiology
- Hypersensitivity / veterinary
- Immunoglobulin E / immunology
- Insect Bites and Stings / veterinary
- Recombinant Proteins / immunology
- Simuliidae / immunology
- Skin Tests
- Wasp Venoms / immunology
Citations
This article has been cited 14 times.- Simonin EM, Wagner B. IgE-binding monocytes upregulate the coagulation cascade in allergic horses.. Genes Immun 2023 Jun;24(3):130-138.
- Cvitas I, Oberhaensli S, Leeb T, Marti E. Equine keratinocytes in the pathogenesis of insect bite hypersensitivity: Just another brick in the wall?. PLoS One 2022;17(8):e0266263.
- Birras J, White SJ, Jonsdottir S, Novotny EN, Ziegler A, Wilson AD, Frey R, Torsteinsdottir S, Alcocer M, Marti E. First clinical expression of equine insect bite hypersensitivity is associated with co-sensitization to multiple Culicoides allergens.. PLoS One 2021;16(11):e0257819.
- Willen L, Basáñez MG, Dvorak V, Veriegh FBD, Aboagye FT, Idun B, Osman ME, Osei-Atweneboana MY, Courtenay O, Volf P. Human immune response against salivary antigens of Simulium damnosum s.l.: A new epidemiological marker for exposure to blackfly bites in onchocerciasis endemic areas.. PLoS Negl Trop Dis 2021 Jun;15(6):e0009512.
- Larson EM, Babasyan S, Wagner B. IgE-Binding Monocytes Have an Enhanced Ability to Produce IL-8 (CXCL8) in Animals with Naturally Occurring Allergy.. J Immunol 2021 May 15;206(10):2312-2321.
- Hemmer W, Wantke F. Insect hypersensitivity beyond bee and wasp venom allergy.. Allergol Select 2020;4:97-104.
- Novotny EN, White SJ, Wilson AD, Stefánsdóttir SB, Tijhaar E, Jonsdóttir S, Frey R, Reiche D, Rose H, Rhyner C, Schüpbach-Regula G, Torsteinsdóttir S, Alcocer M, Marti E. Component-resolved microarray analysis of IgE sensitization profiles to Culicoides recombinant allergens in horses with insect bite hypersensitivity.. Allergy 2021 Apr;76(4):1147-1157.
- Larson EM, Babasyan S, Wagner B. Phenotype and function of IgE-binding monocytes in equine Culicoides hypersensitivity.. PLoS One 2020;15(5):e0233537.
- François L, Hoskens H, Velie BD, Stinckens A, Tinel S, Lamberigts C, Peeters L, Savelkoul HFJ, Tijhaar E, Lindgren G, Janssens S, Ducro BJ, Buys N, Schurink AA. Genomic Regions Associated with IgE Levels against Culicoides spp. Antigens in Three Horse Breeds.. Genes (Basel) 2019 Aug 8;10(8).
- Hempolchom C, Sookrung N, Srisuka W, Reamtong O, Sakolvaree Y, Chaicumpa W, Dedkhad W, Jariyapan N, Takaoka H, Saeung A. Characterization of IgE-binding proteins in the salivary glands of Simulium nigrogilvum (Diptera: Simuliidae).. Parasitol Res 2019 Aug;118(8):2353-2359.
- Torsteinsdottir S, Scheidegger S, Baselgia S, Jonsdottir S, Svansson V, Björnsdottir S, Marti E. A prospective study on insect bite hypersensitivity in horses exported from Iceland into Switzerland.. Acta Vet Scand 2018 Nov 3;60(1):69.
- Einhorn L, Hofstetter G, Brandt S, Hainisch EK, Fukuda I, Kusano K, Scheynius A, Mittermann I, Resch-Marat Y, Vrtala S, Valenta R, Marti E, Rhyner C, Crameri R, Satoh R, Teshima R, Tanaka A, Sato H, Matsuda H, Pali-Schöll I, Jensen-Jarolim E. Molecular allergen profiling in horses by microarray reveals Fag e 2 from buckwheat as a frequent sensitizer.. Allergy 2018 Jul;73(7):1436-1446.
- Mueller RS, Janda J, Jensen-Jarolim E, Rhyner C, Marti E. Allergens in veterinary medicine.. Allergy 2016 Jan;71(1):27-35.
- Rohoušová I, Subrahmanyam S, Volfová V, Mu J, Volf P, Valenzuela JG, Jochim RC. Salivary gland transcriptomes and proteomes of Phlebotomus tobbi and Phlebotomus sergenti, vectors of leishmaniasis.. PLoS Negl Trop Dis 2012;6(5):e1660.
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