树鼩粪便中病毒三种浓缩方法的效果比较  

Methods for Concentration of Enteroviruses from Fecal of Tupaia belangeri: a Comparative Study

作　　者：陈玲霞[1] 尹博文[1] 苗雨润[1] 宋庆凯 李晓飞[1] 代解杰[1] CHEN Lingxia;YIN Bowen;MIAO Yurun;SONG Qingkai;LI Xiaofei;DAI Jiejie(The Institute of Medical Biology, The Chinese Academy of Medical Science and Peking Union Medical College. Yunnan Key Laboratory of Vaccine Research and Development on Severe Infectious Diseases, Center of Tree Shrew Germplasm Resources, Kunming 650118, China)

机构地区：[1]中国医学科学院/北京协和医学院医学生物学研究所云南省重大传染病疫苗研发重点实验室树鼩种质资源中心,昆明650118

出　　处：《实验动物科学》2018年第2期38-41,46,共5页Laboratory Animal Science

基　　金：国家科技支撑计划项目(No.2014BAI01B00);云南省联合支持国家计划项目(No.2015GA009)

摘　　要：目的比较树鼩粪便中病毒的三种浓缩方法,评价各方法的浓缩效果,为病毒浓缩方法的最佳选择提供依据。方法将指示病毒(树鼩呼肠孤病毒,tree shrew’s reovirus,TRV)加入经离心过滤处理后的野生树鼩粪便上清中,采用PEG(聚乙二醇,polyehhylene glycol)沉淀结合超声分离法、铁离子絮凝的化学方法和GBviroFast病毒浓缩试剂盒对树鼩粪便上清分别进行浓缩,经实时荧光定量PCR技术对指示病毒进行绝对定量,比较各方法的浓缩效果。结果 TRV病毒悬液的载量为1.34E+05;铁离子絮凝的化学方法浓缩的TRV载量为1.86E+04,浓缩效率为49.57%;GBviroFast病毒浓缩试剂盒浓缩的TRV载量为0.98E+04,浓缩效率为26.12%;PEG沉淀结合超声分离法浓缩的TRV载量为2.30E+03,浓缩效率为6.13%。结论三种方法均能浓缩粪便上清中低浓度的肠道病毒,铁离子絮凝方法最佳,GBviroFast病毒浓缩试剂盒其次,PEG结合超声分离效果最差。Objective To provide a simple and efficient method for concentration of the virus in feces of Tupaia belangeri,the sensitivity,different concentrations and recovery of three method for enterovirus concentration were observed. Method The centrifuged and filtered wild tree shrews stool supernatant was added indicative virus（tree shrew＇s reovirus TRV）,and the fecal supernatants were concentrated by PEG precipitation combined with ultrasonic separation,Fe^3＋-based virus occulation and GBviro Fast concentrated virus kit. To determine the effect of concentrating three method,real-time PCR technique was used. Result Load of TRV virus suspension is1. 34 E ＋ 05; TRV load concentrated by Fe^3＋-based chemical flocculation is 1. 86 E ＋ 04,and the concentration efficiency is 49. 57%. TRV load concentrated by GBviro Fast concentrated virus kit is 0. 98 E ＋ 04,with concentrated efficiency of 26. 12%. TRV load concentrated by PEG precipitation combined with ultrasonic separation is 2. 30 E ＋ 03,and the concentration efficiency only is 6. 13%. Conclusion Three method can concentrate stool supernatant which contain low concentrations of enteroviruses,Fe-based chemical flocculation is the best,GBviro Fast concentrated virus kit secondly,PEG combined with ultrasonic separation method is the worst.

关 键 词：实时荧光定量PCR 病毒浓缩 粪便 树鼩 

分 类 号：Q95-33[生物学—动物学]