牛乳头瘤病毒基因1型广西GX01流行株全基因组克隆及序列分析

为了解牛乳头瘤病毒1型（bovine papillomavirus genotype 1，BPV-1）广西GX01株全基因组序列、结构特征及遗传变异情况，同时了解该毒株引起宿主产生的病理组织学变化情况，本研究选取广西贺州市患病牛皮肤肿瘤样物制作石蜡切片后镜检观察，提取病料DNA，以乳头瘤病毒L1基因的简并引物FAP59/FAP64进行PCR扩增以确定此病毒的基因型，根据GenBank中BPV参考株设计嵌套引物，对GX01株进行全基因组扩增、克隆测序及序列分析。病理组织学检查结果显示，可在病变部位发现表皮细胞增生、肿胀，角质过度及挖空细胞等乳头瘤病毒感染的特征性病变。序列分析结果表明，GX01株为BPV-1，其全基因组长为7 945 bp，包含E1、E2、E4、E5、E6、E7、L1、L2 8个开放阅读框，符合BPV-1型基因组的结构特征；GX01与BPV-1参考株全基因组核苷酸序列同源性为98.6%~99.6%，与BPV-2型参考株（M20219.1）、BPV-13型参考株（JQ798171.1）同源性分别为86.9%和87.2%。GX01株为广西地区首次经检测确认并测定全基因组序列的牛乳头瘤病毒。本研究为广西地区乃至全国的牛乳头状瘤的病原鉴定、流行规律、遗传变异、疫源追溯及科学防控提供了基础数据。

Cloning and Sequence Analysis of the Complete Genome of Bovine Papillomavirus Genotype 1 GX01 Strain from Guangxi

The aim of this study was to investigate the complete genomic sequence,genomic characteristics and genetic variation of bovine papilloma virus genotype 1 (BPV-1) GX01 strain from Guangxi,and the histopathological changes of host was caused by it as well.Firstly,histopathological examination was conducted using the cutaneous papilloma samples of cattle collected from Hezhou city of Guangxi.DNA were extracted and employed as the template of PCR for BPV detection and genotyping using FAP59/FAP64 primers.Based on the complete genomes of the BPV-1 reference strains,specific primer pairs were designed,and the complete genome of the GX01 strain was further amplified and sequenced.Histopathological analysis of cutaneous papilloma showed hyperkeratosis,acanthosis and koilocytosis,which were the typical histopathological characteristics of BPV infection.Genotyping showed GX01 belonged to BPV-1.The full-length genome of GX01 was 7 945 bp,and contained 8 open reading frames (E1,E2,E4,E5,E6,E7,L1 and L2),which matched the structural characteristics of BPV-1.GX01 shared 98.6% to 99.6% homology with the complete genome sequences of BPV-1 reference strains,while 86.9% and 87.2% homology with those of the BPV-2 (M20219.1) and BPV-13 (JQ798171.1) reference strains,respectively.The GX01 strain was the first BPV-1 strain from Guangxi being confirmed and fully sequenced.This study provided theoretical basis for pathogen identification,epidemic regularity,genetic variation,epidemic source tracking and scientific prevention and control of bovine papilloma in Guangxi and China.