猪流行性腹泻病毒和猪传染性胃肠炎病毒一步法双重RT-PCR检测方法的建立与应用

doi:10.16431/j.cnki.1671-7236.2016.09.009

Abstract

Abstract:

The study was aimed to establish a rapid one-step duplex RT-PCR detection method,which could be used to identify and diagnose PEDV and TGEV in clinical diarrhea cases.According to the gene sequences of PEDV and TGEV from GenBank,two pairs of specific primes were designed.Through optimizing and selecting of the best reaction conditions,we finally pinpointed the duplex one-step RT-PCR detection method with strong specificity,which could detect 1×10^-5 diluent degree of vaccine. Suspected samples,which were collected from different pig farms in 2015,were detected of PEDV with 100% positive rate.The method was rapid,high sensitivity and specificity,which could be used for clinical detection of PEDV and TGEV,and also for epidemiological investigation.

Key words: porcine epidemic diarrhea virus (PEDV); porcine transmissible gastroenteritis virus (TGEV); duplex RT-PCR

CLC Number:

S858.28

HU Hong-hui, NAN Wen-jin, HUANG Jian-qiang, WU Jing-bo, PENG Guo-liang. Establishment and Application of a Double RT-PCR Method for Detection of PEDV and TGEV[J]. , 2016, 43(9): 2279-2284.

References

[1] Song D,Moon H,Kang B.Porcine epidemic diarrhea:A review of current epidemiology and available vaccines[J].Clin Exp Vaccine Res,2015,4(2):166-176.
[2] Lee C.Porcine epidemic diarrhea virus:An emerging and re-emerging epizootic swine virus[J].Virol J,2015,12:193
[3] 林初文,庄金秋,谢印乾.我国主要猪病毒性腹泻疾病的特点及其防治对策[J].中国畜牧兽医,2007,34(5):121-122.
[4] Lin C M,Gao X,Oka T,et al.Antigenic relationships among porcine epidemic diarrhea virus and transmissible gastroenteritis virus strains[J].J Virol,2015,89(6):3332-3342.
[5] Chung H C,Nguyen V G,Moon H J,et al.Isolation of porcine epidemic diarrhea virus during outbreaks in South Korea,2013-2014[J].Emerg Infect Dis,2015,21(12):2238-2240.
[6] Suzuki T,Murakami S,Takahashi O,et al.Molecular characterization of pig epidemic diarrhea viruses isolated in Japan from 2013 to 2014[J].Infect Genet Evol,2015,36:363-368.
[7] Kim Y K,Lim S I,Lim J A,et al.A novel strain of porcine epidemic diarrhea virus in Vietnamese pigs[J].Arch Virol,2015,160:1573-1577.
[8] Wang L,Byrum B,Zhang Y.New variant of porcine epidemic diarrhea virus,United States,2014[J].Emerg Infect Dis,2014,20:917-919.
[9] Zhang J,Tsai Y L,Lee P A,et al.Evaluation of two singleplex reverse transcription-insulated isothermal PCR tests and a duplex real-time RT-PCR test for the detection of porcine epidemic diarrhea virus and porcine deltacoronavirus[J].J Virol Methods,2016,234:34-42.
[10] Yamamoto T,Suzuki T,Ohashi S,et al.Genomic motifs as a novel indicator of the relationship between strains isolated from the epidemic of porcine epidemic diarrhea in 2013-2014[J].PLoS One,2016,11(1):e0147994
[11] Choe S E,Park K H,Lim S I,et al.Complete genome sequence of a porcine epidemic diarrhea virus strain from Vietnam,HUA-14PED96,with a large genomic deletion[J].Genome Announc,2016,4(1):e00002-16.
[12] Cai Y,Yin W,Zhou Y,et al.Molecular detection of porcineastrovirus in Sichuan province,China[J].Virol J,2016,13(1):6.
[13] 郭容利,何孔旺,倪艳秀,等.PEDV、TGEV、PARV多重RT-PCR检测方法的建立及其应用[J].江苏农业学报,2013,29(5):1065-1069.
[14] 赵雯雯,李敬双,赵晓岚,等.猪传染性胃肠炎病毒和猪流行性腹泻病毒二联RT-PCR检测方法的建立[J].黑龙江畜牧兽医,2010,2:109-110.
[15] 李一经,刘博,姜艳平,等.双抗体夹心ELISA检测排泄物PEDV[J].东北农业大学学报,2015,46(2):1-10.
[16] 杜晓莉,王一成,吴润,等.2010-2013年浙江省猪流行性腹泻病毒临床检测及PEDV-S基因型分析[J].浙江农业学报,2014,26(3):581-587.
[17] 赵梦姣,陈书民,成岩,等.山东省部分地区猪流行性腹泻流行病学调查及其M基因遗传变异分析[J].中国兽医学报,2013,33(10):1504-1508.
[18] 卢冰霞,秦毅斌,何颖,等.2011年-2014年广西猪流行性腹泻病毒检测及其M基因的序列分析[J].中国畜牧兽医,2015,42(3):549-557.
[19] 田云,焦颖,王福广,等.2012-2013年广东猪流行性腹泻流行情况调查报告[J].广东畜牧兽医科技,2015,40(5):18-21.
[20] 田云,孙彦伟,王福广,等.2012年广东猪群腹泻流行病学调查[J].广东畜牧兽医科技,2015,40(3):19-22.
[21] 蔡汝健,张乐宜,宋长绪.2010-2013年华南地区猪流行性腹泻病流行情况调查及防控效果[J].广东农业科学,2013,11:104-114.
[22] 孙振钊,赵伟男,王靖.猪腹泻病例中PEDV、TGEV及PRV的感染调查[J].动物医学进展,2014,35(5):132-135.

Establishment and Application of a Double RT-PCR Method for Detection of PEDV and TGEV

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