Since severe acute respiratory syndrome (SARS) and Middle East respiratory 
syndrome (MERS) coronaviruses (CoVs) share similar characteristics with respect 
to clinical signs, etiology, and transmission, methods for a rapid and accurate 
differential diagnosis are important. Therefore, the aim of this study was to 
develop a duplex real-time reverse transcription (RT)-PCR method for the 
simultaneous detection of these viruses. Primers and probes that target the 
conserved spike S2 region of human SARS-CoV, MERS-CoV, and their related bat 
CoVs were designed. The results of real-time RT-PCR showed specific reactions 
for each virus with adequate detection limits of 50-100 copies/mL and 
5-100 copies/mL using pUC57-SARS-pS2 (a template for SARS-CoV) and pGEM-MERS-S2 
(a template for MERS-CoV), respectively. In addition, this real-time RT-PCR 
system was able to detect the target viruses SARS-like bat CoV and MERS-CoV in 
bat fecal samples and sputum of MERS patients, respectively. Therefore, this 
newly developed real-time RT-PCR method is expected to detect not only SARS-CoV 
and MERS-CoV in humans but also several bat CoVs that are closely related to 
these viruses in bats.