鸽圆环病毒浙江株全基因组的克隆及序列分析

Cloning and Analysis of the Complete Genomes of Pigeon Circovirus from Zhejiang Province

鸽圆环病毒（Pigeon circovrius,PiCV）是圆环病毒科的一个成员,是一种免疫抑制性病原,鸽子感染圆环病毒后可以出现昏睡、嗜眠、厌食、体重减轻、呼吸窘迫等症状。本研究应用建立的PCR方法,检测了从浙江省5个鸽场采集的样品,结果在杭州某鸽场检测到阳性。进一步设计引物分段PCR扩增、克隆并测定了2株鸽圆环病毒的全基因组序列,分别命名为PiCV-zj1和PiCV-zj2,序列的GenBank登录号为DQ090945和DQ090944。基因组序列分析表明,PiCV-zj1及PiCV-zj2全长均为2039bp,具有圆环病毒共同的与病毒复制相关的茎环结构和Rep蛋白保守基序等特征,与已发表的国外11株鸽圆环病毒序列在全基因组核苷酸水平有86%~89.1%的同源性,在Rep和外壳蛋白的氨基酸水平分别有92.1%~94.7%和76.6%~81.4%的同源性。应用PHYLIP方法作进化树分析并应用bootstrap重复1000次进行检验,结果显示：10个欧美株组成一个大的分支,而浙江株和澳洲株各自独立分支。这是我国首次检测确认鸽圆环病毒的存在,并测定了2株PiCV病毒的全基因组序列。

Pigeon circovrius （PiCV） is a member of circovirus, which is usually regarded as an immunosuppression agent. There were reports that pigeons infected by PiCV showed symptoms of lethargy, weight loss, vomiting, diarrhea, respiratory distress, etc. In this study, we established a PCR method for the detection of PiCV DNA. Samples from 5 different farms in Zhejiang Province were examined and samples from a farm in Hangzhou were positive. Furthermore, the genomic segments of 2 strains of PiCV were amplified, cloned and sequenced using designed primers and the complete genomes of the strains were then assembled and named as PiCV-zj1 and PiCV-zj2, respectively. The sequences were deposited in GenBank under the GenBank Accession number of DQ090945 and DQ090944, respectively. Sequence Analysis had shown that the complete genomes of 2 strains of PiCV from Zhejiang Province had 2 039 nucleotides totally in length and common characters of eircovirus such as a stem-loop structure and conserved motifs for Rep protein, which were supposed to be related to the replication of the virus. Pairwise comparisons showed that the nueleotide sequence of the genome of PiCV strains from Zhejiang Province had 86%-89.1% identities to that of 11 published PiCV strains, and that the amino acid identities of the replication-associated protein （Rep） and capsid protein （Cap） displayed 92. 1%-94.7% and 76.6%-81.4%, respectively. A phylogenic tree was built using PHYLIP with bootstrap support for 1 000 replicates. The result showed that 10 strains from Europe and America formed one big branch and the others from Zhejiang Province and Australia formed the other two, respectively. This was the first report on the detection and full genome sequencing of PiCV in China.