Wu Minglin, Li Haiyang, Jiang He, Hou Guanjun, He Jixiang, Jiang Yangyang, Chen Honglian
Fisheries Research Institute, Anhui Academy of Agricultural Sciences, No. 40 South Nongke Road, Luyang District, Hefei, 230000 Anhui China.
Virusdisease. 2017 Mar;28(1):26-32. doi: 10.1007/s13337-016-0357-1. Epub 2017 Jan 16.
A highly virulent grass carp reovirus (GCRV) strain, named GCRV-AH528, was recently purified from a diseased grass carp with hemorrhage disease in Anhui, China. GCRV-AH528 S9 segment was 1320 nucleotides in length and encoded a 418 amino acid VP6 protein. BLAST search showed that the VP6 protein owned a conserved domain belonging to the reoviral σ2 family. Phylogenetic analysis of VP6 presented that GCRV-AH528 belonged to GCRV genotype II, which was more closely related to than GCRV genotype I and genotype III. Further analysis revealed that GCRV-AH528 S9 and mammalian orthoreovirus S8 might have evolved from a common ancestral precursor and have identical mechanism in virus assembly. The expression level of gene was detected by quantitative real-time PCR (qRT-PCR). Over time, the expression level of gradually increased in kidney cells. However, the level of expression in blood sharply increased at 4-6 days, and then decreased to a low level after GCRV-AH528 challenge ( < 0.05). The gene was detected in all tissues examined, whereas at relatively higher levels in blood, kidney, and liver ( < 0.05). The yeast two-hybrid (Y2H) system was used to identify VP6 self-interaction, while no interaction was detected in VP6-VP6. This study not only revealed the S9 segment structure and expression pattern but also analyzed the VP6 mechanism by yeast hybridization method. The present study provides valuable informations for further experimental design and investigation of VP6 functions.
一种高致病性草鱼呼肠孤病毒（GCRV）毒株，命名为GCRV - AH528，最近从中国安徽一条患出血病的草鱼中纯化得到。GCRV - AH528的S9片段长度为1320个核苷酸，编码一个418个氨基酸的VP6蛋白。BLAST搜索显示VP6蛋白拥有一个属于呼肠孤病毒σ2家族的保守结构域。VP6的系统发育分析表明GCRV - AH528属于GCRV基因型II，与GCRV基因型I和基因型III相比关系更密切。进一步分析表明，GCRV - AH528的S9和哺乳动物正呼肠孤病毒的S8可能从一个共同的祖先前体进化而来，并且在病毒组装中具有相同的机制。通过定量实时PCR（qRT - PCR）检测基因的表达水平。随着时间的推移，基因在肾细胞中的表达水平逐渐升高。然而，在GCRV - AH528攻击后，血液中的表达水平在4 - 6天急剧上升，然后降至低水平（<0.05）。在所检测的所有组织中都检测到了基因，而在血液、肾脏和肝脏中的水平相对较高（<0.05）。利用酵母双杂交（Y2H）系统鉴定VP6的自我相互作用，而在VP6 - VP6中未检测到相互作用。本研究不仅揭示了S9片段的结构和表达模式，还通过酵母杂交方法分析了VP6的机制。本研究为进一步的实验设计和VP6功能研究提供了有价值的信息。
