Spring viremia of carp virus (SVCV) is a highly lethal virus in common carp (Cyprinus carpio) and other cyprinid fish species. The aim of the present study was to develop an in vivo therapeutic measure against SVCV using artificial microRNA (AmiRNA) targeting the SVCV P gene transcript. Three candidates of AmiRNAs (AmiR-P1, -P2, and -P3) were selected, and their ability to downregulate SVCV P gene transcript was analyzed by both synthesized AmiRNA mimics and AmiRNA-expressing vector system, in which AmiR-P3 showed the strongest inhibitory activity among the three candidates. To overcome in vivo limitation of miRNA mimics or plasmid-based miRNA expression systems, we rescued recombinant snakehead rhabdoviruses (SHRVs) expressing SVCV P gene–targeting AmiRNA (rSHRV-AmiR-P3) or control AmiRNA (rSHRV-AmiR-C) using reverse genetic technology. The successful expression of AmiR-P3 and AmiR-C in cells infected with the rescued viruses was verified by quantitative PCR. To evaluate the availability of rSHRV-AmiR-P3 for in vivo control of SVCV, zebrafish (Danio rerio) were (i) infected with either rSHRV-AmiR-C or rSHRV-AmiR-P3 followed by SVCV infection or (ii) infected with SVCV followed by either rSHRV-AmiR-C or rSHRV-AmiR-P3 infection. Fish infected with rSHRVs before and after SVCV infection showed significantly higher survival rates than fish infected with SVCV alone. There was no significant difference in survival rates between groups of fish infected with rSHRV-AmiR-C and rSHRV-AmiR-P3 before SVCV infection; however, fish infected with SVCV followed by infection with rSHRV-AmiR-P3 showed significantly higher survival rates than fish infected with rSHRV-AmiR-C. These results suggest that rSHRV-AmiR-P3 has therapeutic potential against SVCV in fish when administered after SVCV infection, and rSHRVs expressing artificial microRNAs targeting SVCV transcripts could be used as a tool to control SVCV infection in fish for a therapeutic purpose.

鲤鱼春季病毒血症病毒（SVCV）是鲤鱼（Cyprinus carpio）和其他鲤科鱼类的一种高度致死性病毒。本研究的目的是使用针对 SVCV P 基因转录本的人工 microRNA (AmiRNA) 开发针对 SVCV 的体内治疗措施。选择了三个候选AmiRNA（AmiR-P1、-P2和-P3），并通过合成的AmiRNA模拟物和AmiRNA表达载体系统分析了它们下调SVCV P基因转录本的能力，其中AmiR-P3表现出最强三个候选者之间的抑制活性。为了克服 miRNA 模拟物或基于质粒的 miRNA 表达系统的体内限制，我们使用反向拯救表达 SVCV P 基因靶向 AmiRNA (rSHRV-AmiR-P3) 或对照 AmiRNA (rSHRV-AmiR-C) 的重组蛇头弹状病毒 (SHRV)基因技术。通过定量PCR验证了AmiR-P3和AmiR-C在被拯救的病毒感染的细胞中的成功表达。为了评估 rSHRV-AmiR-P3 对 SVCV 体内控制的可用性，斑马鱼 ( Danio rerio ) (i) 感染 rSHRV-AmiR-C 或 rSHRV-AmiR-P3，然后感染 SVCV，或 (ii) 感染SVCV 随后感染 rSHRV-AmiR-C 或 rSHRV-AmiR-P3。在SVCV感染前后感染rSHRV的鱼比单独感染SVCV的鱼表现出显着更高的存活率。SVCV感染前感染rSHRV-AmiR-C和rSHRV-AmiR-P3的鱼组间存活率无显着差异；然而，感染SVCV后再感染rSHRV-AmiR-P3的鱼的存活率明显高于感染rSHRV-AmiR-C的鱼。这些结果表明，在 SVCV 感染后施用时，rSHRV-AmiR-P3 具有针对鱼类 SVCV 的治疗潜力，并且表达靶向 SVCV 转录物的人工 microRNA 的 rSHRV 可用作控制鱼类 SVCV 感染的工具，以达到治疗目的。