title: Early protein ORF086 is an effective vaccine candidate for infectious spleen and kidney necrosis virus in mandarin fish Siniperca chuatsi.
abstract: Infectious spleen and kidney necrosis virus (ISKNV) has caused significant loss in the Mandarin fish (Siniperca chuatsi) aquaculture industry. Vaccination is an important measure to prevent fatal ISKNV infection. In this study, the ORF086 gene encoding an early protein helicase of ISKNV was cloned into the prokaryotic pET32a (+) and eukaryotic pcDNA3.1 (+) expression vectors and designated as pET086 and pcDNA086, respectively. A recombinant 36 kDa protein was detected in Escherichia coli BL21 (harboring pET086) after isopropyl β-d-1-thiogalactopyranoside (IPTG) induction. Polyclonal antibodies against the purified ORF086 protein were raised in rabbits. The antibody reaction and the pcDNA086 expression in muscle tissues of vaccinated fish were confirmed using Western blot analysis. The protective efficacy of ORF086 was also investigated. The cumulative mortality rates of Mandarin fish were significantly different between immune and control groups (P < 0.05) after ISKNV challenge. The relative percentage survival (RPS) values of the recombinant ORF086 protein emulsified with ISA763A adjuvant and pcDNA086 added with QCDC adjuvant were 73% and 63%, respectively. Transcriptional analysis of non-specific and specific immune related genes revealed that the expression levels of IRF-7, IRAK1, Mx, Viperin, and IgM were strongly up-regulated in the vaccinated groups post-immunization. In particular, the expression levels in the QCDC + pcDNA086 group was higher than those in the control groups (P < 0.05). These results indicated that the early protein ORF086 could be an effective antigen candidate for controlling ISKNV disease in Mandarin fish.
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Xiaozhe
authors.initials: X
authors.lastname: Fu
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China. Electronic address: liningq@126.com.
authors.forename: Ningqiu
authors.initials: N
authors.lastname: Li
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Qiang
authors.initials: Q
authors.lastname: Lin
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Huizhi
authors.initials: H
authors.lastname: Guo
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Lihui
authors.initials: L
authors.lastname: Liu
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China.
authors.forename: Zhibin
authors.initials: Z
authors.lastname: Huang
authors.affiliation: Pearl River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Key Laboratory of Fishery Drug Development, Ministry of Agriculture, Key Laboratory of Aquatic Animal Immune Technology, Guangdong Province, Guangzhou 510380, China; Freshwater Aquaculture Collaborative Innovation Center of Hubei Province, Huazhong Agricultural University, Wuhan, Hubei 430070, China.
authors.forename: Shuqin
authors.initials: S
authors.lastname: Wu
mesh_terms: D000818:Animals; D004266:DNA Virus Infections / Q000276:immunology / Q000662:veterinary* / Q000821:virology; D004926:Escherichia coli / Q000235:genetics; D005393:Fish Diseases / Q000276:immunology* / Q000821:virology; D007113:Immunity, Innate; D007497:Iridoviridae / Q000276:immunology*; D030781:Organisms, Genetically Modified / Q000235:genetics; D010473:Perciformes; D010957:Plasmids / Q000235:genetics; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014764:Viral Proteins / Q000235:genetics* / Q000378:metabolism; D014765:Viral Vaccines / Q000276:immunology*
chemical_list: D011994:Recombinant Proteins; D014764:Viral Proteins; D014765:Viral Vaccines
keywords: Infectious spleen and kidney necrosis virus; ORF086; Siniperca chuatsi; Vaccine