一株雏鹅痛风型鹅星状病毒的分离鉴定及其生物学特性研究  

Identification and characterization of a goose astrovirus causing gout in goslings

作　　者：徐蓉[1] 陆明青 张海涛 黄灿平 张冠卿 鲍恩东[1] 孙卫东[1] 吕英军[1] XU Rong;LU Mingqing;ZHANG Haitao;HUANG Canping;ZHANG Guanqing;BAO Endong;SUN Weidong;L Yingjun(College of Veterinary Medicine,Nanjing Agricultural University,Nanjing 210095,China;Lihua Muye Company Limited,Changzhou 213168,China;Asset Management Company,Nanjing Agricultural University,Nanjing 210095,China)

机构地区：[1]南京农业大学动物医学院,江苏南京210095 [2]立华牧业股份有限公司,江苏常州213168 [3]南京农业大学资产经营公司,江苏南京210095

出　　处：《南京农业大学学报》2019年第6期1124-1132,共9页Journal of Nanjing Agricultural University

基　　金：江苏高校优势学科建设工程资助项目(PAPD);中央高校基本科研业务费专项资金(KYZ201629)

摘　　要：[目的]2017年以来,在中国东部地区的许多鹅场中,4-16日龄雏鹅暴发了严重的痛风,导致2%~20%的死亡率,给养鹅场带来巨大经济损失,本试验旨在分离、鉴定导致此次雏鹅痛风的病原并分析其生物学特性。[方法]将发病雏鹅病料接种10日龄鹅胚,分离病毒,通过PCR、基因测序和系统进化分析对分离的病毒进行鉴定;病毒尿囊液接种鹅肾脏上皮细胞测定病毒滴度;攻毒扬州白鹅雏鹅进行动物回归实验,观察剖检和组织病理变化,PCR方法检测肛拭子和各脏器病毒含量,ELISA方法检测血清中尿酸含量。[结果]从第3代鹅胚尿囊液中分离到一株鹅星状病毒(GAstV),命名为JSHA株。对星状病毒ORF2基因氨基酸进化树的分析结果表明,JSHA毒株与2018年从患痛风的鹅上分离得到的星状病毒毒株(SD01、SDPY、GD)具有99%以上的同源性,而与其他禽星状病毒同源性仅为27.3%~57.0%。阳性尿囊液感染鹅胚肾脏上皮细胞后出现变圆、聚团和脱落等细胞病变,病毒滴度为104.25 TCID50·mL^-1。动物回归实验结果显示,该病毒可致雏鹅25%的死亡率,发病鹅体质量减轻、肾脏发白肿胀、脏器表面和关节腔内尿酸盐沉积以及血清尿酸升高;组织病变表现为肾小管上皮细胞变性坏死,尿酸盐结晶形成,大量炎性细胞浸润,肝脏和脾脏中出现坏死区和炎性细胞浸润;组织病毒载量结果显示肾脏、肝脏和脾脏的病毒载量高于其他组织;雏鹅从感染后3 d开始排毒,排毒高峰出现在6~9 d。[结论]鹅星状病毒JSHA毒株是此次雏鹅发生痛风的主要原因。[Objectives]The paper aimed to identify and characterize the etiological agent of the gout which many goose flocks suffered from in the severe outbreaks of gout in 4 to 16-day-old goslings which had caused about 2% to 20% mortality in the Eastern China since 2017,bringing huge economic losses to the goose flocks. [Methods]Tissue samples from diseased goslings were inoculated into goose embryos to isolate pathogens,and the isolates were identified by PCR detection,genome sequencing and phylogenetic analysis. Positive allantoic fluid was inoculated into goose embryonic kidney epithelial cells to determinate virus titer. Goslings(Yangzhou white goose)were inoculated with the virus as an animal reproduction experiment,and observing anatomical and histopathological changes,detecting virus shedding and distribution in clocal sample and organ samples were detected by PCR,and serum uric acid was measured by ELISA. [Results]A virus designated goose astrovirus(GAstV)JSHA was isolated from allantoic fluid of goose embryos following three serial passages. Phylogenetic analysis indicated that the JSHA strain had more than 99% identity with GAstV strains(SD01,SDPY,GD)isolated from geese with gout while it was highly divergent from all other avastroviruses(only 27.3% to 57.0%)according to the amino acid sequence of the full-length ORF2. Cytopathic changes including rounding,clumping and detachment of cells from the surface were found in goose embryonic kidney epithelial cells with positive allantoic fluid,and the virus titer was 104.25 TCID50·mL^-1. Animal reproduction experiment showed that the virus caused the mortality of 25%(5/20),the reduction of body weight,pale and swollen kidney,visceral gout and the increase of serum uric acid. Histopathological changes were observed in the kidneys,livers and spleens,including necrosis of renal tubular,liver and spleen,the deposition of urates and inflammatory infiltration. Accordingly,the virus load was higher in kidney,liver and spleen than that in other tissues. Virus shedding of g

关 键 词：星状病毒 分离 系统进化分析 痛风 雏鹅 

分 类 号：S835[农业科学—畜牧学]