猪繁殖障碍病毒性疫病六重PCR检测方法的建立及应用

Abstract

Abstract: According to the gene sequences in GenBank, six pairs of specific primers were designed to amplify part fragments of the specific genes of PRV gE, PPV NS1, PCV ORF2, PRRSV ORF7, CSFV E2 and JEV E, respectively. The results of sensitivity and specificity tests showed that the minimum amounts of nucleic acid detection by the mPCR were as follows: PRV 6.6 pg, PPV 96 pg, PCV2 12.9 pg, PRRSV 10.5 pg, CSFV 51 pg, and JEV 46 pg respectively, and the amplification results of Swine Influenza Virus (SIV) and Escherichia coli (E. coli) were both negative. The results of 103 clinical samples from Guizhou province revealed the six viral diseases exist universally in pig farms of Guizhou province, and the mixed infection was much serious. The Six-plex PCR method could detect the infection of PRV, PPV, PCV, PRRSV, CSFV and JEV simultaneously, could detect DNA and RNA virus synchronously, and could differentiate and diagnose the clinical samples of single infection or mixed infection infected by the six viruses quickly.

CLC Number:

S854.43

LIU Zhi-Jie, ZENG Zhi-Yong, TANG De-Yuan, ZHOU Li, LIANG Hai-Ying, XIAO Chao-Neng. Establishment and Application of a Multiplex PCR Method for Single-step Simultaneous Detection of Six Viral Pathogens of Porcine Reproductive Disorder[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2012, 43(9): 1429-1436.

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References

［1］CHEN H Y, WEI Z Y, ZHANG H Y, et al. Use of a multiplex RTPCR assay for simultaneous detection of the North American genotype porcine reproductive and respiratory syndrome virus, swine influenza virus and Japanese encephalitis virus［J］. Agr Sci China, 2010, 9(7):1050-1057.

［2］OGAWA H, TAIRAA O, HIRAI T, et al. Multiplex PCR and multiplex RT-PCR for inclusive detection of major swine DNA and RNA viruses in pigs with multiple infections ［J］. J Virol Methods, 2009, 160:210-214.

［3］曹洪志. 多重PCR诊断猪5种繁殖障碍性疾病的方法研究［D］.雅安：四川农业大学，2007.

［4］YUE F X, CUI S J, ZHANG C F, et al. A multiplex PCR for rapid and simultaneous detection of porcine circovirus type 2, porcine parvovirus, porcine pseudorabies virus, and porcine reproductive and respiratory syndrome virusin clinical specimens［J］.Virus Genes, 2009, 38(3):392-397.

［5］CHABERLAIN J S,GIBBS R A,RANIER J E, et al. Deletion screening of the Duchenne muscular dystrophy locus via multiplex DNA amplification［J］. Nucleic Acids Res, 1988, 16(23): 11141-11156.

［6］BELLAU-PUJOL S, VABRET A, LEGRAND L, et al. Development of three multiplex RT-PCR assays for the detection of 12 respiratory RNA viruses［J］. J Virol Methods, 2005, 126: 53-63.

［7］曾智勇，周莉，汤德元，等.猪伪狂犬病病毒GZ-Z1株的分离鉴定［J］.中国兽医科学, 2011,41(05):459-463.

［8］刘霞，汤德元，黄涛，等.猪繁殖与呼吸综合征病毒贵州分离株ORF5基因的克隆及其表达载体的构建［J］.黑龙江畜牧兽医，2010，(2):102-104.

［9］黄涛，汤德元，嵇辛勤，等.猪瘟病毒E2基因的克隆及原核和真核表达载体的构建［J］.黑龙江畜牧兽医，2009，（10）：70-72.

［10］徐健，汤德元，黄涛，等.日本乙型脑炎病毒的分离与鉴定［J］.畜牧与兽医，2009,41（12）:10-12.

［11］殷华平，郭万柱，徐志文，等.猪细小病毒(PPV)SC1株的分离鉴定［J］.黑龙江畜牧兽医，2006，（7）：63-65.

［12］王稳，屈武斌，申志勇，等. 利用MPprimer设计引物并优化扩增条件以提高多重PCR效率的试验研究［J］.生物化学与生物物理进展，2010，37（3）：342-346.

［13］殷震，刘景华.动物病毒学［M］.北京:科学出版社，1997.

［14］ELNIFRO E M, ASHSHI A M, COOPER R J, et al. Multiplex PCR: Optimization and application in diagnostic virology ［J］. Clin Microbiol Rev, 2000, 4(13):559-570.

［15］LIU S S, ZHAO Y R, HUA Q B, et al. A multiplex RTPCR for rapid and simultaneous detection of porcine teschovirus, classical swine fever virus, and porcine reproductive and respiratory syndrome virus in clinical specimens ［J］. J Virol Methods, 2011, 172(1-2):88-92.

［16］方立，徐辉，陈伟杰，等. 猪圆环病毒2型猪伪狂犬病病毒和猪细小病毒三重PCR检测方法的建立［J］.中国兽医科技，2005，35(12):954-958.

［17］MURDOCH D R．Molecular genetic methods in the diagnosis of lower respiratory tract infections［J］．APMIS，2004，112(11-12):713-727．

［18］刘志杰，李如举，周莉，等.猪繁殖障碍性疾病的病原鉴定［J］.贵州农业科学，2011，39（8）：136-137.

［19］李昌文，仇华吉，童光志，等．一株内源性猪细小病毒的分离与鉴定［J］．中国预防兽医学报，2000，22(9):90-93.

［20］JOO H S，DONALDSON-WOOD C R，JOHNSON R H．A standardised haemagglutination inhibition test for porcine parvovirus antibody ［J］.Aust Vet J，1976，52(9):422-424.

［21］刘志杰，李如举，周莉，等.猪伪狂犬病毒病与圆环病毒病2型混合感染的诊断［J］. 贵州农业科学，2011，39(11):153-154.

［22］OPRIESSNIG T，MADSON D M，PRICKETT J R，et al. Effect of porcine circovirus type 2 (PCV2) vaccination on porcine reproductive and respiratory syndrome virus (PRRSV) and PCV2 coinfection ［J］.Vet Microbiol, 2008, 18:103-114.

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Establishment and Application of a Multiplex PCR Method for Single-step Simultaneous Detection of Six Viral Pathogens of Porcine Reproductive Disorder

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