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Open AccessArticle

Development and Validation of a S1 Protein-Based ELISA for the Specific Detection of Antibodies against Equine Coronavirus

Virology Division, Department of Infectious Diseases & Immunology, Faculty of Veterinary Medicine, Utrecht University, Yalelaan 1, 3584CL Utrecht, The Netherlands
GD Animal Health, Department of Small Ruminants, Horses and Companion Animals, Arnsbergstraat 7, 7418EZ Deventer, The Netherlands
Department of Medicine and Epidemiology, School of Veterinary Medicine, University of California, Davis, One Shields Ave., Davis, CA 95616, USA
Authors to whom correspondence should be addressed.
These authors contributed equally to the work.
Viruses 2019, 11(12), 1109;
Received: 29 October 2019 / Revised: 26 November 2019 / Accepted: 29 November 2019 / Published: 30 November 2019
(This article belongs to the Special Issue Equine Viruses)
Equine coronavirus (ECoV) is considered to be involved in enteric diseases in foals. Recently, several outbreaks of ECoV infection have also been reported in adult horses from the USA, France and Japan. Epidemiological studies of ECoV infection are still limited, and the seroprevalence of ECoV infection in Europe is unknown. In this study, an indirect enzyme-linked immunosorbent assay (ELISA) method utilizing ECoV spike S1 protein was developed in two formats, and further validated by analyzing 27 paired serum samples (acute and convalescent sera) from horses involved in an ECoV outbreak and 1084 sera of horses with unknown ECoV exposure. Both formats showed high diagnostic accuracy compared to virus neutralization (VN) assay. Receiver-operating characteristic (ROC) analyses were performed to determine the best cut-off values for both ELISA formats, assuming a test specificity of 99%. Employing the developed ELISA method, we detected seroconversion in 70.4% of horses from an ECoV outbreak. Among the 1084 horse sera, seropositivity varied from 25.9% (young horses) to 82.8% (adult horses) in Dutch horse populations. Further, sera of Icelandic horses were included in this study and a significant number of sera (62%) were found to be positive. Overall, the results demonstrated that the ECoV S1-based ELISA has reliable diagnostic performance compared to the VN assay and is a useful assay to support seroconversion in horses involved with ECoV outbreaks and to estimate ECoV seroprevalence in populations of horses. View Full-Text
Keywords: equine coronavirus; spike S1 protein; ELISA; virus neutralization; seroprevalence equine coronavirus; spike S1 protein; ELISA; virus neutralization; seroprevalence
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Zhao, S.; Smits, C.; Schuurman, N.; Barnum, S.; Pusterla, N.; van Kuppeveld, F.; Bosch, B.-J.; van Maanen, K.; Egberink, H. Development and Validation of a S1 Protein-Based ELISA for the Specific Detection of Antibodies against Equine Coronavirus. Viruses 2019, 11, 1109.

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