African swine fever (ASF) is an infectious disease of domestic and wild pigs, caused by ASF virus (ASFV). In this study, a triplex real-time PCR assay was developed to detect and differentiate the gene-deleted and wild-type ASFV strains. Three pairs of primers and probes were designed to target the conserved region of B646L gene (p72), MGF_360-14L gene (located in the middle of MGF360-505R gene) and CD2v gene, respectively. Gene-deleted (with MGF360-505R and / or CD2v genes deletion) and wild-type ASFV strains were detected specifically and simultaneously by the assay developed without cross-reactions with other nucleic acids of PCV-2, CSFV, PRRSV, FMDV or SVA. The detection limits of the triplex rPCR were 7.9 copies, 9.7 copies, and 9.6 copies of standard plasmid DNA containing B646L gene, MGF_360-14L gene and CD2v gene, respectively. A total of 1215 field samples were tested in parallel by the triplex rPCR and real-time PCR recommended by OIE, and the B646L gene detection results were completely consistent between these two assays. The triplex rPCR assay was successfully developed to identify pigs infected with wild-type ASFV strains or immunized with the ASFV gene-deleted vaccine.

中文翻译：

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开发用于检测和区分基因缺失和野生型非洲猪瘟病毒的三重实时PCR分析方法。

非洲猪瘟（ASF）是由ASF病毒（ASFV）引起的家猪和野猪的传染病。在这项研究中，开发了三重实时荧光定量PCR检测和区分基因缺失和野生型ASFV菌株。设计三对引物和探针分别靶向B646L基因（p72），MGF_360-14L基因（位于MGF360-505R基因中间）和CD2v基因的保守区。通过开发的检测方法，可以特异性地同时检测基因缺失的基因（具有MGF360-505R和/或CD2v基因缺失）和野生型ASFV菌株，而不会与PCV-2，CSFV，PRRSV，FMDV或SVA的其他核酸发生交叉反应。三重rPCR的检出限分别为含有B646L基因，MGF_360-14L基因和CD2v基因的标准质粒DNA的7.9个拷贝，9.7个拷贝和9.6个拷贝。通过OIE推荐的三重rPCR和实时PCR平行测试了总共1215个田间样品，这两种测定之间的B646L基因检测结果完全一致。成功开发了三重rPCR分析法，以鉴定感染了野生型ASFV株或用ASFV基因缺失疫苗免疫的猪。