Equine Research Institute, Japan Racing Association, 1400-4 Shiba, Shimotsuke, Tochigi 329-0412, Japan.
J Virol Methods. 2019 Sep;271:113681. doi: 10.1016/j.jviromet.2019.113681. Epub 2019 Jun 15.
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.
马感染克沙奇病毒后会出现发热、皮疹和四肢水肿等症状。为了寻找一种高通量、省时的血清学诊断方法，我们探索了克沙奇病毒的免疫原性抗原，并使用重组蛋白建立了酶联免疫吸附试验（ELISA）。Western blot 分析显示，感染马血清与约 46 kDa 的病毒抗原发生强烈反应，该抗原对应 E1 或 E2 糖蛋白。Western blot 和 ELISA 结果均显示，重组 E2（rE2）蛋白比重组 E1（rE1）蛋白与感染马血清的反应更强。在使用 rE2 蛋白的 ELISA（rE2-ELISA）中，所有经实验感染克沙奇病毒的马（n=7）在感染后 14 天的 OD 值均超过了截断值。9 只接种疫苗马中有 5 只的 OD 值在接种后也略高于截断值。在自然感染的马（n=28）中，24 匹急性血清呈阴性，恢复期血清转为阳性。在急性血清呈阳性的 4 匹马中，采用配对血清系列稀释的终点法检测到滴度增加了 4 倍以上。总之，我们建立了 rE2-ELISA，可以检测实验和自然感染后马针对克沙奇病毒的抗体；这对于克沙奇病毒感染的诊断和监测应该是有用的。
