1Guangxi Key Laboratory of Marine Natural Products and Combinatorial Biosynethesis Chemistry, Guangxi Academy of Sciences, Nanning, PR China.
2College of Marine Sciences, South China Agricultural University, Guangzhou, PR China.
J Gen Virol. 2019 Mar;100(3):380-391. doi: 10.1099/jgv.0.001226. Epub 2019 Feb 4.
Nervous necrosis virus (NNV), is one of the most fatal viruses in marine fish aquaculture, and is capable of infecting over 50 different fish species. Trachinotus ovatus NNV (GTONNV) was isolated from diseased golden pompano. This T. ovatus strain was isolated from Guangxi, China. Single-stranded DNA (ssDNA) aptamers with high specificity for GTONNV-infected T. ovatus cerebellum cells (TOCC) were produced by Systematic Evolution of Ligands by Exponential Enrichment (SELEX). The characterization of these aptamers was performed using flow cytometry and laser scanning confocal microscopy. The selected aptamers showed significant specificity for GTONNV-infected cells. Based on MFOLD prediction, aptamers formed distinct stem-loop structures that could form the basis for the aptamers' specific binding to their cellular targets. Protease treatment results revealed that the target molecules for aptamers TNA1, TNA4 and TNA19 within GTONNV-infected cells may be membrane proteins that were trypsin-sensitive. Specific endocytosis of aptamer TNA1, TNA4 and TNA19 into GTONNV-infected cells was also shown. The selected aptamers demonstrated antiviral effects against GTONNV both in vitro and in vivo. This is the first time that aptamers targeting GTONNV-infected T. ovatus cells have been selected and characterized. These aptamers hold promise as rapid diagnostic reagents or targeted therapeutic drugs against GTONNV.
神经坏死病毒（NNV）是海水养殖鱼类中最致命的病毒之一，能够感染 50 多种不同的鱼类。金黄鰤神经坏死病毒（GTONNV）是从患病的金鲳鱼中分离出来的。该金黄鰤毒株分离自中国广西。通过指数富集的配体系统进化（SELEX）技术，从感染 GTONNV 的金黄鰤小脑细胞（TOCC）中产生了针对 GTONNV 的高特异性单链 DNA（ssDNA）适体。通过流式细胞术和激光共聚焦显微镜对这些适体进行了表征。所选适体对 GTONNV 感染的细胞表现出显著的特异性。基于 MFOLD 预测，适体形成了独特的茎环结构，这可能是适体与其细胞靶标特异性结合的基础。蛋白酶处理结果表明，GTONNV 感染细胞中适体 TNA1、TNA4 和 TNA19 的靶分子可能是对胰蛋白酶敏感的膜蛋白。还显示了适体 TNA1、TNA4 和 TNA19 特异性地内吞进入 GTONNV 感染的细胞。所选适体在体外和体内均显示出对 GTONNV 的抗病毒作用。这是首次筛选和表征针对 GTONNV 感染的金黄鰤细胞的适体。这些适体有望成为针对 GTONNV 的快速诊断试剂或靶向治疗药物。
