Agriculture Ministry Key Laboratory of Healthy Freshwater Aquaculture, Key Laboratory of Fish, Health and Nutrition of Zhejiang Province, Zhejiang Institute of Freshwater Fisheries, Huzhou, 313001, China.
Agriculture Ministry Key Laboratory of Healthy Freshwater Aquaculture, Key Laboratory of Fish, Health and Nutrition of Zhejiang Province, Zhejiang Institute of Freshwater Fisheries, Huzhou, 313001, China.
J Virol Methods. 2020 Jun;280:113877. doi: 10.1016/j.jviromet.2020.113877. Epub 2020 Apr 23.
In recent years, crucian carp hematopoietic necrosis caused by Cyprinid herpesvirus 2 (CyHV-2) infection has caused an enormous economic loss to the aquaculture industry.
In this study antigenic epitope analysis was performed on the membrane proteins of CyHV-2, and 8 antigen-rich peptide fragments were selected for prokaryotic expression. Then, the immunogenicity of the recombinant proteins was analyzed. On this basis, DNA vaccines were constructed for immunization of hybridized Prussian carps. The protective effect of DNA vaccines against challenge in hybridized Prussian carps was evaluated.
The results showed that all 8 recombinant proteins were successfully expressed. Among the recombinant proteins, ORF16, tORF25, tORF64, and ORF146, gave a positive serum reaction with CyHV-2. Of the four proteins used for the immunization of silver crucian carps, the antibody titer induced by tORF25 was the highest. The DNA vaccine, pEGFP-N1-ORF25, was constructed based on ORF25 and able to induce production of specific antibodies in carps, while up-regulating the expression of MHCⅠ, IL-1β, C3, and TF-A in the kidneys of carps. Moreover, the immunoprotective rate was increased to 70% in hybridized Prussian carps.
The results showed that the DNA vaccine constructed based on the ORF25 gene had a greater immune protective effect and can be used as a candidate vaccine for immunoprotection against CyHV-2.
近年来，由鲤疱疹病毒 2 （CyHV-2）感染引起的鲫鱼造血坏死病给水产养殖业造成了巨大的经济损失。
本研究对 CyHV-2 的膜蛋白进行抗原表位分析，选择 8 个富含抗原的肽段进行原核表达，并分析重组蛋白的免疫原性。在此基础上，构建杂交鲫鱼的 DNA 疫苗进行免疫接种，评价 DNA 疫苗对杂交鲫鱼的攻毒保护效果。
结果表明，8 种重组蛋白均成功表达。在重组蛋白中，ORF16、tORF25、tORF64 和 ORF146 与 CyHV-2 血清呈阳性反应。在对银鲫进行免疫的 4 种重组蛋白中，tORF25 诱导的抗体滴度最高。基于 ORF25 构建了 DNA 疫苗 pEGFP-N1-ORF25，能够诱导鲤鱼产生特异性抗体，同时上调鲤鱼肾脏中 MHCⅠ、IL-1β、C3 和 TF-A 的表达。此外，在杂交鲫鱼中的免疫保护率提高到 70%。
结果表明，基于 ORF25 基因构建的 DNA 疫苗具有更大的免疫保护效果，可作为防治 CyHV-2 的候选疫苗。
