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Detection of Middle East respiratory syndrome coronavirus using reverse transcription loop-mediated isothermal amplification (RT-LAMP).
Virol J. 2014 Aug 08; 11:139.VJ

Abstract

BACKGROUND

The first documented case of Middle East Respiratory Syndrome coronavirus (MERS-CoV) occurred in 2012, and outbreaks have continued ever since, mainly in Saudi Arabia. MERS-CoV is primarily diagnosed using a real-time RT-PCR assay, with at least two different genomic targets required for a positive diagnosis according to the case definition of The World Health Organization (WHO) as of 3 July 2013. Therefore, it is urgently necessary to develop as many specific genetic diagnostic methods as possible to allow stable diagnosis of MERS-CoV infections.

METHODS

Reverse transcription-loop-mediated isothermal amplification (RT-LAMP) is a genetic diagnostic method used widely for the detection of viral pathogens, which requires only a single temperature for amplification, and can be completed in less than 1 h. This study developed a novel RT-LAMP assay for detecting MERS-CoV using primer sets targeting a conserved nucleocapsid protein region.

RESULTS

The RT-LAMP assay was capable of detecting as few as 3.4 copies of MERS-CoV RNA, and was highly specific, with no cross-reaction to other respiratory viruses. Pilot experiments to detect MERS-CoV from medium containing pharyngeal swabs inoculated with pre-titrated viruses were also performed. The RT-LAMP assay exhibited sensitivity similar to that of MERS-CoV real-time RT-PCR.

CONCLUSIONS

These results suggest that the RT-LAMP assay described here is a useful tool for the diagnosis and epidemiologic surveillance of human MERS-CoV infections.

Authors+Show Affiliations

Laboratory of Acute Respiratory Viral Diseases and Cytokines, Department of Virology III, National Institute of Infectious Disease, Laboratory of Acute Respiratory Viral Diseases and Cytokines, 4-7-1 Gakuen, Musashimurayama, Tokyo 208-0011, Japan. shirato@nih.go.jp.No affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

25103205

Citation

Shirato, Kazuya, et al. "Detection of Middle East Respiratory Syndrome Coronavirus Using Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP)." Virology Journal, vol. 11, 2014, p. 139.
Shirato K, Yano T, Senba S, et al. Detection of Middle East respiratory syndrome coronavirus using reverse transcription loop-mediated isothermal amplification (RT-LAMP). Virol J. 2014;11:139.
Shirato, K., Yano, T., Senba, S., Akachi, S., Kobayashi, T., Nishinaka, T., Notomi, T., & Matsuyama, S. (2014). Detection of Middle East respiratory syndrome coronavirus using reverse transcription loop-mediated isothermal amplification (RT-LAMP). Virology Journal, 11, 139. https://doi.org/10.1186/1743-422X-11-139
Shirato K, et al. Detection of Middle East Respiratory Syndrome Coronavirus Using Reverse Transcription Loop-mediated Isothermal Amplification (RT-LAMP). Virol J. 2014 Aug 8;11:139. PubMed PMID: 25103205.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Detection of Middle East respiratory syndrome coronavirus using reverse transcription loop-mediated isothermal amplification (RT-LAMP). AU - Shirato,Kazuya, AU - Yano,Takuya, AU - Senba,Syouhei, AU - Akachi,Shigehiro, AU - Kobayashi,Takashi, AU - Nishinaka,Takamichi, AU - Notomi,Tsugunori, AU - Matsuyama,Shutoku, Y1 - 2014/08/08/ PY - 2014/04/25/received PY - 2014/08/04/accepted PY - 2014/8/9/entrez PY - 2014/8/12/pubmed PY - 2015/3/31/medline SP - 139 EP - 139 JF - Virology journal JO - Virol J VL - 11 N2 - BACKGROUND: The first documented case of Middle East Respiratory Syndrome coronavirus (MERS-CoV) occurred in 2012, and outbreaks have continued ever since, mainly in Saudi Arabia. MERS-CoV is primarily diagnosed using a real-time RT-PCR assay, with at least two different genomic targets required for a positive diagnosis according to the case definition of The World Health Organization (WHO) as of 3 July 2013. Therefore, it is urgently necessary to develop as many specific genetic diagnostic methods as possible to allow stable diagnosis of MERS-CoV infections. METHODS: Reverse transcription-loop-mediated isothermal amplification (RT-LAMP) is a genetic diagnostic method used widely for the detection of viral pathogens, which requires only a single temperature for amplification, and can be completed in less than 1 h. This study developed a novel RT-LAMP assay for detecting MERS-CoV using primer sets targeting a conserved nucleocapsid protein region. RESULTS: The RT-LAMP assay was capable of detecting as few as 3.4 copies of MERS-CoV RNA, and was highly specific, with no cross-reaction to other respiratory viruses. Pilot experiments to detect MERS-CoV from medium containing pharyngeal swabs inoculated with pre-titrated viruses were also performed. The RT-LAMP assay exhibited sensitivity similar to that of MERS-CoV real-time RT-PCR. CONCLUSIONS: These results suggest that the RT-LAMP assay described here is a useful tool for the diagnosis and epidemiologic surveillance of human MERS-CoV infections. SN - 1743-422X UR - https://www.unboundmedicine.com/medline/citation/25103205/Detection_of_Middle_East_respiratory_syndrome_coronavirus_using_reverse_transcription_loop_mediated_isothermal_amplification__RT_LAMP__ L2 - https://virologyj.biomedcentral.com/articles/10.1186/1743-422X-11-139 DB - PRIME DP - Unbound Medicine ER -