摘要:
根据已发表的CSFV序列合成1对引物,以猪瘟病毒(CSFV)兔化弱毒株为模板,建立并优化了检测CSFV的RT-PCR方法,并与直接荧光抗体试验(DFA)进行比较,检测了15份临床疑似病料。结果如下:应用RT-PCR对CSFV兔化弱毒株RNA进行扩增,获得与预期大小相符长为509bp的特异性目的片段,敏感性达到10pg的CSFV-RNA,对猪繁殖与呼吸综合征病毒(PRRSV)、猪细小病毒(PPV)、猪伪狂犬病毒(PRV)和猪2型圆环病毒(PCV2)进行同条件扩增均为阴性,RT-PCR产物测序结果与文献报道的CSFV不同毒株的核苷酸序列同源性达到95%～99%;15份临床疑似病料应用RT-PCR的检出率为66.7%,而应用免疫荧光试验的检出率为60.0%,二者的总符合率为80%。表明RT-PCR方法比DFA更敏感,两种方法均可用于猪瘟的快速诊断。
Abstract:
A pair of primers for RT-PCR to detect the classical swine fever virus (CSFV) were synthesized according to the published genome sequences of CSFV strains.After the RT-PCR method was established and optimized,15 clinic specimens were detected for CSFV by RT-PCR in comparison with the direct immunofluoresence assay (DFA).The result indicated that a 509 bp specifical fragment was obtained from the CSFV vaccine strains by RT-PCR with the sensitivity reaching 10 pg of CSFVRNA.Negative results were obtained from porcine reproductive and respiratory syndrome virus (PRRSV),porcine parvovirus (PPV),pseudorabies virus (PRV) and porcine circovirus Ⅱ (PCV2).The nucleotide sequence homology of RT-PCR products was 95%-99% of the other CSFV strains.The positive detection rate of the 15 clinic specimens reached 66.7% by RT-PCR,as compared to 60.0% by DFA.The two method’s coincidence rate was 80%.These results indicated that the RTPCR method was more sensitive than DFA,and that both methods were suitable for rapid detection of CSFV.
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