中文摘要：

利用RT-PCR从广东某猪场疑似猪瘟（Classical swine fever,CSF）病例中检测出CSFV,并对扩增的E2全长基因进行了序列测定和系统进化分析。结果表明：引发该起猪瘟疫情的毒株属于CSFV 2.1b基因亚亚型,命名为GD176。为了进一步了解该毒株的复制特点,利用PK-15细胞对GD176进行了体外细胞适应性传代,并对获得的细胞适应毒GD176-F46进行了全基因组测序。结果表明：GD176通过在PK-15上不断传代,最终获得了高滴度的适应毒株,其滴度从第6代的10^2.61TCID50/m L提高至第46代的10^8.06TCID50/m L。病毒生长动力学结果显示,GD176-F46在感染后72 h病毒滴度达到最高值（10^8.17TCID50/m L）。为进一步分析GD176在细胞传代适应过程中的稳定性,对不同代次细胞毒的E2全长基因进行扩增和序列测定,结果发现：GD176-F0与GD176-F6、F10、F15、F20、F25、F30、F35、F40和F46的E2基因序列完全一致,这表明囊膜蛋白E2在病毒适应PK-15细胞过程中非常稳定。通过比对GD176和其他2.1亚型毒株各蛋白的氨基酸序列发现,不同毒株之间同源性最小的病毒蛋白是NS5A,低于病毒囊膜糖蛋白E2。获得了高度适应PK-15细胞的GD176细胞毒,为进行病毒毒力评价、复制和致病特性研究奠定了基础。

英文摘要：

Classical swine fever virus（ CSFV） is the causative agent of highly contagious swine disease CSF that has occurred in many countries over the world including China. Recently a CSF- suspected clinical sample from Guangdong province was collected and detected as CSFV positive by RT- PCR. Phylogenetic analysis based on the nucleotide sequence of full-length E2 gene showedthat the CSFV isolate GD176 belongs to sub-subgenotype 2. 1b of CSFV. To further understand the replication characteristics of this sub-subgenotype 2. 1 isolate,GD176 was isolated and adapted in PK- 15 cells,whole-genome sequence of GD176cell-adapted virus was amplified and sequenced.Adaptation of GD176 in PK- 15 cells started at F6 with 10^2. 61TCID50/ mL and was increased to10^8. 06TCID50/ mL at F46. Growth dynamics of GD176-F46 with a MOI at 0. 1 indicated that the peak titer reached at 72 h postinfection（ 10^8. 17TCID50/ mL）. To further define the stability of GD176 during cellular adaptation,full-length E2 genes of different cell-adapted viruses of GD176 were sequenced. As a result,E2 gene of GD176-F0 was found to be identical with that of GD176-F6,F10,F15,F20,F25,F30,F35,F40 and F46,indicating the genetic stability of E2 gene during in vitro adaptation of GD176 in PK- 15 cells. In addition,comparison of individual viral protein showed that the most variable protein between GD176 and other 2. 1 sub-subgenotype isolates is NS5 A rather than viral envelope protein E2. Overall,highly cell-adapted CSFV sub-subgenotype 2. 1b strain GD176 was obtained in this study and it will be useful for future study on viral virulence,replication and pathogenesis.