山羊痘病例的病理学诊断及病毒鉴定

Isolation,identification and pathological diagnosis of goat pox virus

菏泽市某养羊场从散户购买青山羊50余只,购进后1周左右陆续发病,发病羊体温41～42℃,2周内死亡率超过60%,部分羊体表有痘疹样病变,疑似羊痘。对该病例进行了现场调查、尸体剖检、病理组织学检查及病毒分离鉴定。结果显示：病羊眼睑、口、鼻部有痘疹结节,皮肤有大量红色丘疹。剖检病死羊可见肺、胃肠黏膜、肾及肝表面有数量不等的灰白色丘疹。皮肤及黏膜痘疹部的上皮细胞增生、水泡变性,并见胞浆红染包涵体;肺、肝、肾丘疹完全符合痘疹的病变特征,通过Macehiavello包涵体染色在皮肤、黏膜增生的上皮细胞及肝、肾痘细胞的胞浆内有紫红染包涵体。将获得p32基因序列与GenBank上登录的相应序列进行同源性分析表明该分离株与山羊痘病毒分离株的同源性为99.4%～99.8%,但与绵羊痘病毒的同源性偏低（97.2%～98.1%）。系统进化树分析表明该分离株与山羊痘病毒处于同一分支。将纯化后的p32基因扩增产物用Hinf I酶切位点分析表明,该病毒仅在688bp处存在1个酶切位点,酶切后产生2条目的片段（688和311bp）。对p32基因序列进一步分析表明,在169～171位点上存在3个碱基（GAT）的插入,而山羊痘病毒和其他痘病毒在该位点上有3个碱基缺失,证明此次疫情是由山羊痘病毒引起的。据病理学及病原学诊断结果判定该病例为山羊痘重症感染。按《中华人民共和国动物防疫法》规定,扑杀并无害化处理发病羊群,羊舍、场地、用具严格消毒,粪便堆积发酵处理,受威胁区的羊群实施疫苗紧急接种羊痘疫苗,疫情未发生进一步扩散。

Two weeks after purchased 50 domestic grey goats in one goat farm in Heze,Shandong,China,the goats began to exhibit signs of disease,including papules on the skin and pyrexia（41to42℃）,and morbidity exceeded 60%.Clinical,necropsy,histopathological examinations and molecular biological diagnosis were performed.Greyish-white papules were observed in the lung,gastrointestinal mucosa,kidney,and liver.Most of the papular lesions were characterized by epithelial cell hyperplasia,vacuolar degeneration and eosinophilic inclusion bodies.The viral P32 gene sequence was compared with the genomic sequences in the GenBank nucleotide database,and was found to share 99.4%-99.5%similarity with the GP virus.The presence of only one Hinf I site in the P32 gene sequence distinguished the isolate from the sheep pox virus.Thus,a comprehensive diagnosis of GP was made.According to the Animal Epidemic Prevention Laws of the Peoples Republic of China,emergency measures were implemented to control the outbreak,which included the slaughter and proper disposal of the diseased flock,disinfecting contaminated facilities,and the emergency vaccination of threatened flocks.These measures were eventually successful in eliminating GP from the affected area.