Matsuda Sayaka, Nerome Reiko, Maegawa Kenichi, Kotaki Akira, Sugita Shigeo, Kawasaki Kazunori, Kuroda Kazumichi, Yamaguchi Ryoji, Takasaki Tomohiko, Nerome Kuniaki
The Institute of Biological Resources, 893-2, Nakayama, Nago-shi, Okinawa 905-0004, Japan.
Equine Research Institute, Japan Racing Association, 1400-4, Shiba, Shimotsuke-shi, Tochigi 329-0412, Japan.
Heliyon. 2017 Apr 11;3(4):e00286. doi: 10.1016/j.heliyon.2017.e00286. eCollection 2017 Apr.
We have successfully prepared a Japanese encephalitis virus (JEV) - Nakayama virus like particle (NVLP) vaccine using synthetic codon-optimized prM and E genes. The expression of the recombinant JEV Nakayama-BmNPV (JEV-NNPV) virus was determined in infected silkworm Bm-N cells by fluorescence and Western blot analysis. The recombinant was inoculated into silkworm pupae and the yield of Nakayama VLP (NVLP) reached a peak in the homogenates after 3 days. Additionally, in the peptide analysis of infected pupae homogenate, it appeared approximately 300-500 μg E protein/pupa were produced. When purified the above eluates on the discontinuous sucrose density gradient centrifugation, NVLP showed a strong hemagglutination (HA) activity by using chicken red blood cell in phosphate-buffered saline (PBS) free from Mg and Ca ions. The immune antisera against NVLP strain could efficiently neutralize the plaque formation of Nakayama, Beijing-1 and Muar strains, showing tendency of much higher reaction with heterologous Muar strain than homologous Nakayama strain. Our findings suggest that the JEV-NVLP may be useful for JEV epidemic control in many endemic areas of Asian countries as a widely effective and less expensive JE vaccine.
我们已成功使用合成密码子优化的prM和E基因制备了一种日本脑炎病毒（JEV）- 中山病毒样颗粒（NVLP）疫苗。通过荧光和蛋白质印迹分析在感染的家蚕Bm-N细胞中测定重组JEV中山 - 家蚕核型多角体病毒（JEV-NNPV）病毒的表达。将重组体接种到家蚕蛹中，3天后中山VLP（NVLP）产量在匀浆中达到峰值。此外，在感染蛹匀浆的肽分析中，每只蛹产生约300 - 500μg E蛋白。当在不连续蔗糖密度梯度离心中纯化上述洗脱液时，NVLP在不含镁和钙离子的磷酸盐缓冲盐水（PBS）中使用鸡红细胞显示出很强的血凝（HA）活性。针对NVLP株的免疫抗血清可以有效中和中山株、北京-1株和麻坡株的蚀斑形成，显示出与异源麻坡株的反应比同源中山株高得多的趋势。我们的研究结果表明，JEV-NVLP作为一种广泛有效且成本较低的日本脑炎疫苗，可能对亚洲国家许多流行地区的日本脑炎疫情控制有用。
