Development and evaluation of a multiplex reverse-transcription real-time PCR assay for detection of equine respiratory disease viruses. (November 2018)

• Record Type: Journal Article
• Authors: Ghoniem, Shimaa M.; El Deeb, Ayman H.; Aggour, Mohammed G.; Hussein, Hussein A.
• Is Part Of: Journal of veterinary diagnostic investigation. Volume 30:Number 6(2018:Nov.)
• Page Start: 924
• Page End: 928
• Publication Date: 2018-11
• Subjects: Equine -- multiplex real-time PCR -- respiratory viruses; Veterinary medicine -- Diagnosis -- Periodicals; 636.0896075
• DOI: 10.1177/1040638718799388
• Languages: English
• ISSNs: 1040-6387

Abstract

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 2 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.