Rapid Detection of Porcine Circovirus Type 2 Using a SYBR Green Ⅰ Real-time PCR

doi:10.7668/hbnxb.2014.02.012

Abstract

Abstract: Porcine circovirus type 2 (PCV2) is the causative factor of Post-weaning multi-systemic wasting syndrome (PMWS) in pigs. Rapid detection of PCV2 is very important for the effective prophylaxis and treatment of PMWS. To establish a sensitive, specific assay for the detection and quantitation of PCV2, we designed and synthe- sized specific primers in the open reading frame 1 (ORF1 ), and to develop a SYBR Green Ⅰ Real-time PCR. The results indicated that the Real-time PCR assay could detect 10-100 copies of the genomic DNA per reaction, and its sensitivity was 10 times of the conventional PCR. The assay did not cross-react with classical swine fever virus (CSFV), porcine parvovirus(PPV), porcine reproductive and respiratory syndrome virus (PRRSV) and pseudora- bies virus (PRV) . The limits of detection and quantitation were 10 and 100 copies, respectively. Using the estab- lished Real-time PCR system,43 of the 50 samples we tested were detected as positive. In conclusion, the Real-time PCR assay is sensitive, specific, accurate and can be used for the monitoring of PCV2 infection.

Key words: Porcine circovirus type 2, SYBR Green Ⅰ, Real-time PCR

CLC Number:

LI Peng, GUO Jun-qing, JIN Qian-yue, LI Qing-mei, LI Qing-zhou, WAN Bo, WANG Xuan-nian, WANG Chuan-qing, ZHANG Gai-ping. Rapid Detection of Porcine Circovirus Type 2 Using a SYBR Green Ⅰ Real-time PCR[J]. ACTA AGRICULTURAE BOREALI-SINICA, 2014, 29(2): 66-70. doi: 10.7668/hbnxb.2014.02.012.

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References

[1] Fu F, Tian H, Li X, et al. C-terminal heat shock protein70 of Mycobacterium tuberculosis as a molecular adjuvantfor DNA vaccination with the porcine circovirus type 2ORF2 (capsid) gene in mice[J]. Vet J, 2013, 195(2):244-247.
[2] 苏金存, 常孝勇, 王隆绥, 等. 猪圆环病毒 2 型感染后猪血液学指标研究[J]. 河南农业科学, 2012, 41 (8):163-166.
[3] Borghetti P, Morganti M, Saleri R, et al. Innate pro-inflam-matory and adaptive immune cytokines in PBMC of vacci-nated and unvaccinated pigs naturally exposed to porcinecircovirus type 2 (PCV2) infection vary with the occur-rence of the disease and the viral burden[J]. Vet Micro-biol,2013,163(1-2):42-53.
[4] Yang K, Li W, Niu H, et al. Efficacy of single dose of aninactivated porcine circovirus type 2 (PCV2) whole-virusvaccine with oil adjuvant in piglets[J]. Acta Vet Scand,2012,54:67.
[5] Grasland B, Blanchard P, Kéranflec'h A, et al. Evaluationof the transmission of porcine circovirus type 2 (PCV-2)genogroups a and b with semen from infectedspecific-pathogen-free boars[J]. Vet Microbiol, 2013, 162(2 /4):381-387.
[6] Wen L, He K, Li B, et al. In vitro and in vivo isolation of anovel rearranged porcine circovirus type 2 [J]. J Virol,2012,86(23):13120.
[7] Priscilla F G, Flávia M G, ?ngela M Q L, et al. Fetal in-fections and antibody profiles in pigs naturally infectedwith porcine circovirus type 2 (PCV2) [J]. Can J VetRes,2012,76(1):38-44.
[8] Meng Ge, Wei Luo, Daliang Jiang, et al. Development andapplication of a double-antigen sandwich enzyme-linkedimmunosorbent assay for detection of antibodies to porcinecircovirus 2 [J]. Clin Vaccine Immunol, 2012, 19 (9 ):1480-1486.
[9] Siegfried K, Titus S, Dieter Z, et al. Coreplication of themajor genotype group members of porcine circovirus type2 as a prerequisite to coevolution may explain the variabledisease manifestations [J ]. J Virol, 2011, 85 (21 ):11111-11120.
[10] Lakshman N A G, Kathleen A M, Sarah P, et al. Efficacyof parenteral vaccination against porcine circovirus type2 (PCV2) in seropositive piglets[J]. Can J Vet Res,2012,76(4):301-307.
[11] 王宏魁, 彭志峰, 孙彦婷, 等. 猪圆环病毒复合 PCR 检测方法的建立及应用[J]. 河南农业科学, 2009(1 ):109-111.
[12] Zhao K, Han F, Zou Y, et al. Rapid detection of porcinecircovirus type 2 using a TaqMan-based Real-time PCR[J]. Virol J,2010(7):374.
[13] 祁艳华, 王爱萍, 李学武, 等. 猪圆环病毒 2 型 ORF2基因片段的克隆与原核表达 [J]. 河南农业科学,2010 (3):112-115.
[14] Grau-Roma L, Fraile L, Segalés J. Recent advances inthe epidemiology, diagnosis and control of diseasescaused by porcine circovirus type 2 [J]. Vet J, 2011(1):23-32.
[15] Esther S, Carole B, Heidi G, et al. Cellular adaptive im-mune response against porcine circovirus type 2 in sub-clinically infected pigs[J]. BMC Vet Res,2009(5):45.
[16] Chung W B, Chan W H, Chaung H C, et al . Real-timePCR for quantitation of porcine reproductive and respira-tory syndrome virus and porcine circovirus type 2 in nat-urally-infected and challenged pigs[J]. J Virol Methods,2005,124:11-19.
[17] Yang Z Z, Habib M, Shuai J B, et al . Detection of PCV2DNA by SYBR Green Ⅰ -based quantitative PCR[J]. JZhejiang Univ Sci B,2007,8(3):162-169.

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