猪圆环病毒2型2010 AHCY 株的分离鉴定及遗传变异分析

Isolation,Identification and Genetic Variation of Porcine Circovirus Type 2 Isolate 2010AHCY

旨为分离得到1株猪圆环病毒2型毒株,对其生物学特性进行鉴定,测定其TCID50,并进行全基因组比对分析。利用细胞传代的方法对PCR检测为PCV1阴性、PCV2阳性的病料进行病毒分离,分离毒株接种PK15细胞,通过PCR检测和间接免疫荧光进行鉴定。并对分离株的TCID50和全基因组序列进行了测定与分析。成功分离得到猪圆环病毒2型,命名为2010AHCY株,其基因组全长为1 767 bp,在PK15细胞上的TCID50为104.23。全基因组序列分析表明,该分离株与其他参考毒株核苷酸同源性为93.2%～99.9%,与HM038030 PCV2d的同源性最高,为99.9%。系统进化树分析表明,分离株2010AHCY属于PCV2d基因型,与PCV2b基因型亲缘关系较近。为分析华东地区PCV2分子流行病学、遗传变异和PMWS的防治奠定了基础。

The aim of this study was to obtain the porcine circovirus type 2(PCV2) isolate 2010AHCY and to identify its biological characteristics ,and to analyze the 50%tissue culture infective dose ( TCID50 ) and the whole genome of this isolate .The samples which were detected PCV 1 negative and PCV2 positive were inoculated and pas-saged on PK15 cells.The isolate was identified by PCR and confirmed by immunofluorescent assay ( IFA).The TCID50 of 2010 AHCY isolate was determined and the whole genome was sequenced and its molecular variations was analyzed by DNAStar software .A PCV2 strain 2010 AHCY was successfully isolated from samples .The results showed that the genome length of 2010AHCY was 1 767 bp and the TCID50 was 104.23.Comparison analysis of the complete genomic sequences of 2010 AHCY and other strains indicated that the homologies among these 11 strains were 93.2%-99.9%and the highest homology of 99.9%to the HM038030 PCV2d strain.The phylogenetic trees revealed that the isolate 2010 AHCY belonged to genotype PCV 2 d and had a close relationship with PCV 2 b geno-type.This study provide foundations for studying the molecular epidemiology ,variation and prevention of PMWS in Eastern China .