Ann Lab Med.  2017 Nov;37(6):494-498. 10.3343/alm.2017.37.6.494.

Performance Evaluation of the PowerChek MERS (upE & ORF1a) Real-Time PCR Kit for the Detection of Middle East Respiratory Syndrome Coronavirus RNA

• Huh HJ
• Kim JY
• Kwon HJ
• Yun SA
• Lee MK
• Ki CS
• Lee NY
• Kim JW

• KMID: 2425989
• DOI: http://doi.org/10.3343/alm.2017.37.6.494

Abstract

BACKGROUND
Molecular detection of Middle East respiratory syndrome coronavirus (MERS-CoV) using real-time reverse transcription (rRT)-PCR assays is the method of choice for diagnosis of MERS. We evaluated the performance of the PowerChek MERS (upE & ORF1a) real-time PCR Kit (PowerChek MERS assay; Kogene Biotech, Korea) a one-step rRT-PCR assay for the qualitative detection of MERS-CoV.
METHODS
We evaluated PowerChek MERS assay performance in comparison with nested RT-PCR and sequencing of the RNA-dependent RNA polymerase (RdRp) and N genes. To evaluate diagnostic sensitivity and specificity, 100 clinical specimens (50 positive and 50 negative for MERS-CoV) were simultaneously tested by using the PowerChek MERS and sequencing assays. Assay performance, including limit of detection and precision, was evaluated in vitro by using MERS-CoV RNA transcripts. Analytical specificity was evaluated with a diverse collection of 16 respiratory virus-positive clinical specimens and 14 respiratory bacterial isolates.
RESULTS
The 95% limits of detection of the PowerChek MERS assay for the upE and the open rading frame (ORF)1a were 16.2 copies/µL and 8.2 copies/µL, respectively. No cross-reactivity was observed. The diagnostic sensitivity and specificity of the PowerChek MERS assay were both 100% (95% confidence interval, 91.1-100%).
CONCLUSIONS
The PowerChek MERS assay is a straightforward and accurate assay for detecting MERS-CoV RNA. The assay will be a useful tool for the rapid diagnosis of MERS and could prove especially important for MERS outbreak control.

Keyword

MERS-CoV; Real-time reverse-transcription PCR; Performance

MeSH Terms

Coronavirus Infections*
Diagnosis
In Vitro Techniques
Limit of Detection
Methods
Middle East Respiratory Syndrome Coronavirus*
Middle East*
Real-Time Polymerase Chain Reaction*
Reverse Transcription
RNA
RNA Replicase
Sensitivity and Specificity
RNA
RNA Replicase

Reference

1. Chan JF, Lau SK, To KK, Cheng VC, Woo PC, Yuen KY. Middle East respiratory syndrome coronavirus: another zoonotic betacoronavirus causing SARS-like disease. Clin Microbiol Rev. 2015; 28:465–522. PMID: 25810418.

2. World Health Organization. Laboratory testing for Middle East respiratory syndrome coronavirus (MERT-CoV). Interim guidance. Updated on June 2015. http://www.who.int/csr/disease/coronavirus_infections/mers-laboratory-testing/en/.

3. Corman VM, Eckerle I, Bleicker T, Zaki A, Landt O, Eschbach-Bludau M, et al. Detection of a novel human coronavirus by real-time reverse-transcription polymerase chain reaction. Euro Surveill. 2012; 17:pii:20285.

4. Corman VM, Müller MA, Costabel U, Timm J, Binger T, Meyer B, et al. Assays for laboratory confirmation of novel human coronavirus (hCoV-EMC) infections. Euro Surveill. 2012; 17:pii:20334.

5. Corman VM, Ölschläger S, Wendtner CM, Drexler JF, Hess M, Drosten C. Performance and clinical validation of the RealStar MERS-CoV Kit for detection of Middle East respiratory syndrome coronavirus RNA. J Clin Virol. 2014; 60:168–171. PMID: 24726679.

6. Kim MN, Ko YJ, Seong MW, Kim JS, Shin BM, Sung H. Analytical and clinical validation of six commercial Middle East Respiratory Syndrome coronavirus RNA detection kits based on real-time reverse-transcription PCR. Ann Lab Med. 2016; 36:450–456. PMID: 27374710.

7. Lu X, Whitaker B, Sakthivel SK, Kamili S, Rose LE, Lowe L, et al. Real-time reverse transcription-PCR assay panel for Middle East respiratory syndrome coronavirus. J Clin Microbiol. 2014; 52:67–75. PMID: 24153118.

8. Chan JF, Choi GK, Tsang AK, Tee KM, Lam HY, Yip CC, et al. Development and evaluation of novel real-time reverse transcription-PCR assays with locked nucleic acid probes targeting leader sequences of human-pathogenic coronaviruses. J Clin Microbiol. 2015; 53:2722–2726. PMID: 26019210.

9. Centers for Disease Control and Prevention. Novel coronavirus 2012 real-time RT-PCR assay. Updated on June 10, 2014. http://www.fda.gov/downloads/MedicalDevices/Safety/EmergencySituations/UCM355572.pdf.

10. Shirato K, Yano T, Senba S, Akachi S, Kobayashi T, Nishinaka T, et al. Detection of Middle East respiratory syndrome coronavirus using reverse transcription loop-mediated isothermal amplification (RT-LAMP). Virol J. 2014; 11:139. PMID: 25103205.

11. Seong MW, Lee SJ, Cho SI, Ko K, Kim MN, Sung H, et al. External quality assessment of MERS-CoV molecular diagnostics during the 2015 korean outbreak. Ann Lab Med. 2016; 36:230–234. PMID: 26915611.

12. Jung YJ, Kwon HJ, Huh HJ, Ki CS, Lee NY, Kim JW. Comparison of the AdvanSure real-time RT-PCR and Seeplex(®) RV12 ACE assay for the detection of respiratory viruses. J Virol Methods. 2015; 224:42–46. PMID: 26277911.

13. Bibby DF, McElarney I, Breuer J, Clark DA. Comparative evaluation of the Seegene Seeplex RV15 and real-time PCR for respiratory virus detection. J Med Virol. 2011; 83:1469–1475. PMID: 21678451.

14. Modjarrad K, Moorthy VS, Ben Embarek P, Van Kerkhove M, Kim J, Kieny MP. A roadmap for MERS-CoV research and product development: report from a World Health Organization consultation. Nat Med. 2016; 22:701–705. PMID: 27387881.

15. Drosten C, Doerr HW, Lim W, Stöhr K, Niedrig M. SARS molecular detection external quality assurance. Emerg Infect Dis. 2004; 10:2200–2203. PMID: 15663861.

16. Centers for Disease Control and Prevention. Interim guidelines for collecting, handling, and testing clinical specimens from patients under investigation (PUIs) for Middle East Respiratory Syndrome Coronavirus (MERS-CoV). V2.1. Updated on June 16, 2015. http://www.cdc.gov/coronavirus/mers/guidelines-clinical-specimens.html.

17. Korea Ministry of Health and Welfare and Center for Disease Control and Prevention. Guidelins for MERS-CoV control. 4-1ed. http://cdc.go.kr/CDC/info/CdcKrHealth0289.jsp?menuIds=HOME001-MNU2374-MNU2375-MNU1509-MNU1913&fid=5742&q_type=&q_value=&cid=73196&pageNum=.