[Development of Novel Multiplex Real-time RT-PCR Assays for Detection of MERS-CoV Infection]

We aimed to develop a novel laboratory assay for the detection of Middle East respiratory syndrome coronavirus (MERS-CoV) infection .Several novel multiplex real-time RT-PCR assays were developed using the upE,ORF1 band N2genes of MERS-CoV as targets; the novel assays were compared with previous monoplex real-time RT-PCR assays. For validation, we tested a MERS-CoV strain (hCoVEMC), clinical specimens from patients with fever in Shanghai, and specimens from the first imported MERS case in China. The detection limit of the novel multiplex real-time RT-PCR assays was 10 PFU of MERS-CoV per ml, the same as that in monoplex real-time RT-PCR assays based on upE or N2. The detection was specific for MERS-CoV. In validation using clinical samples, pharyngeal swabs from Shanghai patients were detected as negative, while swabs from the first imported MERS case in China were detected as positive. Using whole blood samples from a MERS case, better detection results were obtained with N2 as the target than upE. We conclude that all the novel assays established in this study could be used for the detection of MERS-CoV; they show potential for improvement compared with monoplex real-time RT-PCR assay based on upE.