Development of rapid immunochromatographic strip test for the detection of porcine epidemic diarrhoea virus

Lyoo, Kwang-Soo; Yeom, Minjoo; Kim, Jungho; Kim, Donghyuk; Ha, Gunwoo; Na, Woonsung; Le, Van Phan; Song, Daesub

Porcine epidemic diarrhoea virus (PEDV) is a highly contagious pathogen that causes acute and severe watery diarrhoea, vomiting and dehydration, and shows 50-100 per cent mortality in up to one-week-old piglets and less severe disease in older pigs. 1 PEDV belongs to genus Alphacoronavirus, family Coronaviridae, and is an enveloped single-stranded positive sense RNA virus. 2 3 Outbreaks of porcine epidemic diarrhoea (PED) have been reported in several countries in Europe, as well as in Asian countries including China, Thailand and South Korea, since the disease was first identified in England in the early 1970s. 1 3 Recently, PEDV was diagnosed in the midwestern region of the USA in 2013. 4 5 Since then, the virus has spread rapidly and was confirmed throughout 30 states in the USA. PEDV has significant economic impacts on the pig industry, causing the loss of approximately seven million piglets within the one-year epidemic period in the USA. 3 Moreover, the emergence of PEDV has been reported in western, central and eastern Europe, and the genome sequences were found to be closely related to PEDV strains from the USA, showing a sequence identity of more than 99 per cent. [6] [7] [8] The genome of PEDV is approximately 28 kb in size and encodes four structural proteins, including spike (S), membrane (M), envelope (E) and nucleocapsid (N), and four non-structural proteins, including 1a, 1b, 3a and 3b. 2 The S protein is critical to regulating interactions between receptor glycoproteins and virus ligands and for mediating viral entry into host cells, while the N protein is a basic phosphor-protein important for maintaining the nucleocapsid structure. Epitopes of the N protein may play a role in inducing cell-mediated immunity. 2 Additionally, the N protein may be an appropriate target because it is the predominant antigen expressed in coronavirus-infected cells. 9 To diagnose PEDV, laboratory-based techniques are necessary, because PED is clinically indistinguishable from other diarrhoea diseases such as transmissible gastritis-enteritis virus infection. 10 To detect the PEDV antigen, virus isolation, immunofluorescence testing, immunohistochemical techniques, ELISA, and molecular techniques including RT-PCR and real-time RT-PCR are currently used in most laboratories. 1 However, to obtain accurate results, these methods require welltrained technicians and specific equipment, and are time-consuming. Moreover, issues such as transportation delays or temperature changes during shipping from outbreak locations to diagnostic laboratories may affect the diagnosis results. 1 10 Therefore, a rapid test kit that is specific and sensitive to PEDV is critical to monitoring this disease on pig farms. Here, we aimed to develop and validate an immunochromatographic assay (ICA) for the rapid and qualitative detection of PEDV antigen from diseased animals. The assay detects N protein of PEDV using highly selective monoclonal antibodies.