Ghoniem Shimaa M, El Deeb Ayman H, Aggour Mohammed G, Hussein Hussein A
Departments of Virology (Ghoniem), Animal Health Research Institute, Dokki, Egypt.
Biotechnology (Aggour), Animal Health Research Institute, Dokki, Egypt.
J Vet Diagn Invest. 2018 Nov;30(6):924-928. doi: 10.1177/1040638718799388. Epub 2018 Sep 21.
We developed a multiplex reverse-transcription real-time PCR (RT-rtPCR) assay for the simultaneous detection of the main equine respiratory viruses: equid alphaherpesviruses 1 and 4 (EHV-1, -4) and equine influenza virus (EIV; species Influenza A virus). The primers and probes amplified only the targeted viruses, and there were no inter-assay cross-amplifications or nonspecific interactions. The multiplex assay efficiencies were 92.5%, 97%, and 90% for EHV-1, EHV-4, and EIV, respectively. The R values of the monoplex and multiplex assays were ⩾0.990, and the slopes were -3.37 to -3.59. The performance of the assay was evaluated by analyzing 152 samples from clinically infected horses. EHV-1 DNA was detected in 12 samples, EHV-4 DNA in 9 samples, and both EHV-1 and EHV-4 in 4 samples. The accuracy of the assay was confirmed by comparing these results using commercial rtPCR and RT-rtPCR kits. Our multiplex RT-rtPCR was a sensitive, specific, accurate, and cost-effective method for the detection of the target viruses whether they occur alone or as part of coinfections.
我们开发了一种多重逆转录实时荧光定量PCR（RT-rtPCR）检测方法，用于同时检测主要的马呼吸道病毒：马α疱疹病毒1型和4型（EHV-1、-4）以及马流感病毒（EIV；甲型流感病毒种）。引物和探针仅扩增目标病毒，不存在检测间的交叉扩增或非特异性相互作用。EHV-1、EHV-4和EIV的多重检测效率分别为92.5%、97%和90%。单重和多重检测的R值均≥0.990，斜率为-3.37至-3.59。通过分析152份来自临床感染马匹的样本对该检测方法的性能进行了评估。在12份样本中检测到EHV-1 DNA，9份样本中检测到EHV-4 DNA，4份样本中同时检测到EHV-1和EHV-4。通过使用商业rtPCR和RT-rtPCR试剂盒比较这些结果，证实了该检测方法的准确性。我们的多重RT-rtPCR是一种灵敏、特异、准确且经济高效的方法，可用于检测目标病毒，无论它们单独出现还是作为混合感染的一部分。
