title: Rapid detection of all known ebolavirus species by reverse transcription-loop-mediated isothermal amplification (RT-LAMP).
abstract: Ebola virus disease (EVD), a highly virulent infectious disease caused by ebolaviruses, has a fatality rate of 25-90%. Without a licensed chemotherapeutic agent or vaccine for the treatment and prevention of EVD, control of outbreaks requires accurate and rapid diagnosis of cases. In this study, five sets of six oligonucleotide primers targeting the nucleoprotein gene were designed for specific identification of each of the five ebolavirus species using reverse transcription-loop mediated isothermal amplification (RT-LAMP) assay. The detection limits of the ebolavirus species-specific primer sets were evaluated using in vitro transcribed RNAs. The detection limit of species-specific RT-LAMP assays for Zaire ebolavirus, Sudan ebolavirus, Taï Forest ebolavirus, and Bundibugyo ebolavirus was 256 copies/reaction, while the detection limit for Reston ebolavirus was 64 copies/reaction, and the detection time for each of the RT-LAMP assays was 13.3±3.0, 19.8±4.6, 14.3±0.6, 16.1±4.7, and 19.8±2.4min (mean±SD), respectively. The sensitivity of the species-specific RT-LAMP assays were similar to that of the established RT-PCR and quantitative RT-PCR assays for diagnosis of EVD and are suitable for field or point-of-care diagnosis. The RT-LAMP assays were specific for the detection of the respective species of ebolavirus with no cross reaction with other species of ebolavirus and other viral hemorrhagic fever viruses such as Marburg virus, Lassa fever virus, and Dengue virus. The species-specific RT-LAMP assays developed in this study are rapid, sensitive, and specific and could be useful in case of an EVD outbreak.
authors.affiliation: Department of Emerging Infectious Diseases, Institute of Tropical Medicine (NEKKEN), Nagasaki University, Nagasaki, Japan; Graduate School of Biomedical Sciences and Program for Nurturing Global Leaders in Tropical and Emerging Communicable Diseases, Nagasaki University, Nagasaki, Japan. Electronic address: o.oloniniyi@gmail.com.
authors.forename: Olamide K
authors.initials: OK
authors.lastname: Oloniniyi
authors.affiliation: Department of Emerging Infectious Diseases, Institute of Tropical Medicine (NEKKEN), Nagasaki University, Nagasaki, Japan. Electronic address: ykuro@nagasaki-u.ac.jp.
authors.forename: Yohei
authors.initials: Y
authors.lastname: Kurosaki
authors.affiliation: Division of Global Epidemiology, Research Center for Zoonosis Control, Hokkaido University, Sapporo, Japan. Electronic address: hirom@czc.hokudai.ac.jp.
authors.forename: Hiroko
authors.initials: H
authors.lastname: Miyamoto
authors.affiliation: Division of Global Epidemiology, Research Center for Zoonosis Control, Hokkaido University, Sapporo, Japan; Global station for Zoonosis Control, Global Institution for Collaborative Research and Education, Hokkaido University, Sapporo, Japan. Electronic address: atakada@czc.hokudai.ac.jp.
authors.forename: Ayato
authors.initials: A
authors.lastname: Takada
authors.affiliation: Department of Emerging Infectious Diseases, Institute of Tropical Medicine (NEKKEN), Nagasaki University, Nagasaki, Japan; Graduate School of Biomedical Sciences and Program for Nurturing Global Leaders in Tropical and Emerging Communicable Diseases, Nagasaki University, Nagasaki, Japan. Electronic address: j-yasuda@nagasaki-u.ac.jp.
authors.forename: Jiro
authors.initials: J
authors.lastname: Yasuda
mesh_terms: D017931:DNA Primers; D003716:Dengue Virus / Q000235:genetics; D029043:Ebolavirus / Q000145:classification / Q000235:genetics / Q000302:isolation & purification*; D019142:Hemorrhagic Fever, Ebola / Q000175:diagnosis*; D006801:Humans; D007836:Lassa virus / Q000235:genetics; D057230:Limit of Detection; D029024:Marburgvirus / Q000235:genetics; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation / Q000379:methods*; D019095:Point-of-Care Systems; D012367:RNA, Viral / Q000235:genetics*; D048348:Reverse Transcription; D012680:Sensitivity and Specificity; D013696:Temperature
chemical_list: D017931:DNA Primers; D012367:RNA, Viral
keywords: Diagnosis; Ebola virus disease; Ebolavirus; RT-LAMP