Simple, Rapid and Sensitive Portable Molecular Diagnosis of SFTS Virus Using Reverse Transcriptional Loop-Mediated Isothermal Amplification (RT-LAMP)
2
조회
0
다운로드
4
인용
0
참고문헌
키워드
초록
Recently, human infections caused by severe fever with thrombocytopenia syndrome virus (SFTSV), which can lead to fatality, have dramatically increased in East Asia. With the unavailability of vaccines or antiviral drugs to prevent and/or treat SFTSV infection, early rapid diagnosis is critical for prevention and control of the disease. Here, we report the development of a simple, rapid and sensitive portable detection method for SFTSV infection applying reverse transcription–loop mediated isothermal amplification (RT-LAMP) combined with one-pot colorimetric visualization and electro-free reaction platform. This method utilizes a pocket warmer to facilitate diagnosis in a resource-limited setting. Specific primers were designed to target the highly-conserved region of L gene of SFTSV. The detection limit of the RT-LAMP assay was approximately 100 viral genome copies from three different SFTSV strains. This assay exhibited comparable sensitivity to qRT-PCR and 10-fold more sensitivity than conventional RT-PCR, with a rapid detection time of 30 to 60 minutes. The RT-LAMP assay using SFTSV clinical specimens has demonstrated a similar detection rate to qRT-PCR and a higher detection rate compared to conventional RT-PCR. Moreover, there was no observed cross-reactive amplification of other human infectious viruses including Japanese Encephalitis Virus (JEV), Dengue, Enterovirus, Zika, Influenza and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). This highly sensitive, electro- and equipment-free rapid colorimetric visualization method is feasible for resource-limited SFTSV field diagnosis.
Abstract
Recently, human infections caused by severe fever with thrombocytopenia syndrome virus (SFTSV), which can lead to fatality, have dramatically increased in East Asia. With the unavailability of vaccines or antiviral drugs to prevent and/or treat SFTSV infection, early rapid diagnosis is critical for prevention and control of the disease. Here, we report the development of a simple, rapid and sensitive portable detection method for SFTSV infection applying reverse transcription–loop mediated isothermal amplification (RT-LAMP) combined with one-pot colorimetric visualization and electro-free reaction platform. This method utilizes a pocket warmer to facilitate diagnosis in a resource-limited setting. Specific primers were designed to target the highly-conserved region of L gene of SFTSV. The detection limit of the RT-LAMP assay was approximately 100 viral genome copies from three different SFTSV strains. This assay exhibited comparable sensitivity to qRT-PCR and 10-fold more sensitivity than conventional RT-PCR, with a rapid detection time of 30 to 60 minutes. The RT-LAMP assay using SFTSV clinical specimens has demonstrated a similar detection rate to qRT-PCR and a higher detection rate compared to conventional RT-PCR. Moreover, there was no observed cross-reactive amplification of other human infectious viruses including Japanese Encephalitis Virus (JEV), Dengue, Enterovirus, Zika, Influenza and Middle East Respiratory Syndrome Coronavirus (MERS-CoV). This highly sensitive, electro- and equipment-free rapid colorimetric visualization method is feasible for resource-limited SFTSV field diagnosis.
참고문헌 (27)
- Yu X-J, "Fever with thrombocytopenia associated with a novel bunyavirus in China", N Engl. J. Me, 364, pp.1523~1532 (2011)
- Zhan J, "Current status of severe fever with thrombocytopenia syndrome in China", Virol. Sin, 32, pp.51~62 (2017)
- Park S-W, "Epidemiological and clinical features of severe fever with thrombocytopenia syndrome during an outbreak in South Korea, 2013-2015", Am. J. Trop. Med. Hyg, 95, pp.1358~1361 (2015)
- Niu G, "Severe fever with thrombocytopenia syndrome virus among domesticated animals, China", Emerg. Infect. Dis, 19, pp.756~ (2013)
- Zhang Y-Z, "The emergence and cross species transmission of newly discovered tick-borne Bunyavirus in China", Curr. Opin. Virol, 16, pp.126~131 (2016)
- Kim WY, "Nosocomial transmission of severe fever with thrombocytopenia syndrome in Korea", Clin. Infect. Dis, 60, pp.1681~1683 (2015)
- Liu Y, "Personto-person transmission of severe fever with thrombocytopenia syndrome virus", Vector Borne Zoonotic Dis, 12, pp.156~160 (2012)
- Bao C-j, "A family cluster of infections by a newly recognized bunyavirus in eastern China, 2007: further evidence of personto-person transmission", Clin. Infect. Dis, 53, pp.1208~1214 (2011)
- Li S, "Sporadic case infected by severe fever with thrombocytopenia syndrome bunyavirus in a non-epidemic region of China", Biosci. Trends, 5, pp.273~276 (2011)
- Xu B, "Metagenomic analysis of fever, thrombocytopenia and leukopenia syndrome (FTLS) in Henan Province, China:discovery of a new bunyavirus", PLoS Pathog, 7, pp.e1002369~ (2011)
- Zhang Y-Z, "Hantavirus infections in humans and animals, China", Emerg. Infect. Dis, 16, pp.1195~ (2010)
- Sun Y, "Early diagnosis of novel SFTS bunyavirus infection by quantitative real-time RT-PCR assay", J. Clin. Virol, 53, pp.48~53 (2012)
- Jiao Y, "Preparation and evaluation of recombinant severe fever with thrombocytopenia syndrome virus nucleocapsid protein for detection of total antibodies in human and animal sera by double-antigen sandwich enzyme-linked immunosorbent assay", J. Clin. Microbiol, 50, pp.372~377 (2012)
- Notomi T, "Loop-mediated isothermal amplification of DNA", Nucleic Acids Res, 28, pp.e63~e63 (2000)
- Fukuta S, "Detection of Japanese yam mosaic virus by RTLAMP", Arch. Virol, 148, pp.1713~1720 (2003)
- Hatano B, "Mobile and accurate detection system for infection by the 2009 pandemic influenza A (H1N1) virus with a pocket-warmer reverse-transcriptase loop-mediated isothermal amplification", J. Med. Virol, 83, pp.568~573 (2011)
- Hatano B, "LAMP using a disposable pocket warmer for anthrax detection, a highly mobile and reliable method for anti-bioterrorism", Jpn. J. Infect. Dis, 63, pp.36~40 (2010)
- Xu H, "Establishment of a novel one-step reverse transcription loop-mediated isothermal amplification assay for rapid identification of RNA from the severe fever with thrombocytopenia syndrome virus", J. Virol. Methods, 194, pp.21~25 (2013)
- Kim K-H, "Severe f ever with thrombocytopenia syndrome, South Korea, 2012", Emerg. Infect. Dis, 19, pp.1892~ (2013)
- Li Z, "Seroprevalence of antibodies against SFTS virus infection in farmers and animals, Jiangsu, China", J. Clin. Virol, 60, pp.185~189 (2014)
- Park J-S, "Immunostained plaque assay for detection and titration of rabies virus infectivity", J. Virol. Methods, 228, pp.21~25 (2016)
- Wang S, "SFTS virus in ticks in an endemic area of China", Am. J. Trop. Med. Hyg, 92, pp.684~689 (2015)
- Huang X-Y, "Detection of new bunyavirus rna by reverse transcription–loop-mediated isothermal amplification", J. Clin. Microbiol, 2, pp.531~535 (2014)
- Yang G, "Development and evaluation of a reverse transcription loopmediated isothermal amplification assay for rapid detection of a new SFTS bunyavirus", Arch. Virol, 157, pp.1779~1783 (2012)
- Ablordey A, "Detection of Mycobacterium ulcerans by the loop mediated isothermal amplification method", PLoS Negl. Trop. Dis, 6, pp.e1590~ (2012)
- Priye A, "A smartphone-based diagnostic platform for rapid detection of Zika, chikungunya, and dengue viruses", Sci. Rep, 7, pp.44778~ (2017)
- Lanciotti RS, "Rapid detection and typing of dengue viruses from clinical samples by using reverse transcriptase-polymerase chain reaction", J. Clin. Microbiol, 30, pp.545~551 (1992)