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J Virol Methods. 2019 Sep;271:113681. doi: 10.1016/j.jviromet.2019.113681. Epub 2019 Jun 15.

Development of an enzyme-linked immunosorbent assay for Getah virus infection in horses using recombinant E2 protein as an antigen.

Author information

1
Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. Electronic address: hiroshi_bannai@jra.go.jp.
2
Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. Electronic address: manabu_nemoto@jra.go.jp.
3
Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. Electronic address: koji_tsujimura@jra.go.jp.
4
Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. Electronic address: takashi_yamanaka@jra.go.jp.
5
Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan. Electronic address: hiroshi_kokado@jra.go.jp.

Abstract

Getah virus causes fever, skin eruptions, and limb edema in horses. For a high-throughput and time-saving method for serodiagnosis, we explored immunogenic antigens of Getah virus, and established an enzyme-linked immunosorbent assay (ELISA) using a recombinant protein. Western blot analysis using sera from infected horses showed strong reaction with viral antigens around 46 kDa corresponding to E1 or E2 glycoproteins. Recombinant E2 (rE2) protein reacted more strongly with infected horse sera than did rE1 protein in both Western blotting and ELISA. In ELISA using rE2 protein (rE2-ELISA), for all horses experimentally infected with Getah virus (n = 7), optical density (OD) exceeded the cutoff value at 14 days post-infection. ODs in five of nine vaccinated horses also slightly exceeded the cutoff value after vaccination. Among naturally infected horses (n = 28), 24 were seronegative in the acute sera, which turned seropositive in the convalescent sera. For the four horses seropositive in the acute sera, an endpoint method with serial dilutions of paired sera detected a ≥4-fold increase in titer. In conclusion, we established rE2-ELISA that could detect horse antibodies against Getah virus after experimental and natural infections; this should be useful in the diagnosis and surveillance of Getah virus infection.

KEYWORDS:

E2 glycoprotein; ELISA; Getah virus

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