Division of Fish Disease, Yangtze River Fisheries Research Institute, Chinese Academy of Fishery Sciences, Wuhan, Hubei, 430223, PR China.
Fish Shellfish Immunol. 2018 May;76:206-215. doi: 10.1016/j.fsi.2018.02.039. Epub 2018 Feb 23.
The hemorrhagic disease of grass carp (Ctenopharyngodon idellus) induced by grass carp reovirus (GCRV) leads to huge economic losses in China and currently, there are no effective methods available for prevention and treatment. The various GCRV genotypes may be one of the major obstacles in the pursuit of an effective antiviral treatment. In this study, we exploited CRISPR/Cas9 gene editing to specifically knockout the DNA sequence of the grass carp Junctional Adhesion Molecule-A (gcJAM-A) and evaluated in vitro resistance against various GCRV genotypes. Our results show that CRISPR/Cas9 effectively knocked out gcJAM-A and reduced GCRV infection for two different genotypes in permissive grass carp kidney cells (CIK), as evidenced by suppressed cytopathic effect (CPE) and GCRV progeny production in infected cells. In addition, with ectopic expression of gcJAM-A in cells, non-permissive cells derived from Chinese giant salamander (Andrias davidianus) muscle (GSM) could be highly infected by both GCRV-JX0901 and Hubei grass carp disease reovirus (HGDRV) strains that have different genotypes. Taken together, the results demonstrate that gcJAM-A is necessary for GCRV infection, implying a potential approach for viral control in aquaculture.
草鱼出血病（Ctenopharyngodon idellus）由草鱼呼肠孤病毒（GCRV）引起，给中国造成了巨大的经济损失，目前尚无有效的预防和治疗方法。不同的 GCRV 基因型可能是寻求有效抗病毒治疗的主要障碍之一。在本研究中，我们利用 CRISPR/Cas9 基因编辑技术特异性敲除草鱼连接黏附分子-A（gcJAM-A）的 DNA 序列，并评估其对各种 GCRV 基因型的体外抗性。我们的结果表明，CRISPR/Cas9 有效地敲除了 gcJAM-A，并降低了两种不同基因型在许可性草鱼肾细胞（CIK）中的 GCRV 感染，这表现在受感染细胞中细胞病变效应（CPE）和 GCRV 后代产量的抑制。此外，通过在细胞中外源表达 gcJAM-A，可使来自中国大鲵（Andrias davidianus）肌肉的非许可性细胞（GSM）高度感染具有不同基因型的 GCRV-JX0901 和湖北草鱼疾病呼肠孤病毒（HGDRV）株。总之，这些结果表明 gcJAM-A 是 GCRV 感染所必需的，这为水产养殖中的病毒控制提供了一种潜在的方法。
