一株猪Ⅱ型圆环病毒的分离鉴定及基因组序列分析

Isolation,Identification and Genome Sequence Analysis of Porcine Circovirus Type Ⅱ Isolate

2011年11月,北京某猪场暴发以全身斑块状出血、腹泻及腹股沟淋巴结明显肿大为主要特征的传染性疾病。采集发病猪淋巴结、肾脏和脾脏组织,研磨成组织混悬液,经0.2μm滤膜过滤后接种PK15细胞,然后用D-氨基葡萄糖处理,连续盲传3代后收取细胞病毒液并提取DNA,经PCR检测、免疫荧光鉴定、全基因组测定及用DNAStar软件对序列进行拼接和分析,结果表明,该分离株为猪Ⅱ型圆环病毒(porcine circovirus type 2,PCV2),命名为BJ20111113株,其全基因组长度为1767bp,与国内外毒株核苷酸序列同源性为94.5%～99.8%,其中与HM038017(BDH株)亲缘关系最近,同源性高达99.8%;与EU148504亲缘关系较远,同源性为94.5%。进化分析结果表明,本研究2011年分离的BJ20111113株属于基因型PCV2b。

In November 2011, an infectious disease characterized by systemic macular hemorrhage, diarrhea and obviously inguinal lymph node enlargement, broke out in a swine farm located in Beijing. The lymph node, kidney and spleen tissues were taken from infected pigs and grinded to obtain tissue suspension which was then filtered through 0.2 μm filter and inocula- ted into the PK15 cells. IYglucosamine was added to the infected cell which was then gone through 3 blind passages,and DNA was extracted from the harvested virus suspension. Identification were done by PCR method and immunofluorescent assay,and the complete genome was also amplified and sequenced,which was then analyzed by using DNAStar software. The results dem- onstrated that the isolated strain belonged to PCV2 （named BJ20111113 strain） with whole genome size of 1767 bp. At nucleo- tide sequence level, its homology could reach 94. 5% to 99. 8% compared with the domestic and foreign reference strains. Among them, the most closely related strain was HM038017 （BDH strain） with homology of 99.8%, and the far distantly re- lated strain was EU148504 with homology of only 94.5%. The phylogenetic analysis result indicated that BJ20111113 strain belonged to genotype PCV2b.