title: Recombinase Polymerase Amplification Assay for Rapid Diagnostics of Dengue Infection.
abstract: BACKGROUND
  Over 2.5 billion people are exposed to the risk of contracting dengue fever (DF). Early diagnosis of DF helps to diminish its burden on public health. Real-time reverse transcription polymerase amplification assays (RT-PCR) are the standard method for molecular detection of the dengue virus (DENV). Real-time RT-PCR analysis is not suitable for on-site screening since mobile devices are large, expensive, and complex. In this study, two RT-recombinase polymerase amplification (RT-RPA) assays were developed to detect DENV1-4.
  
  
  RESULTS
  Using two quantitative RNA molecular standards, the analytical sensitivity of a RT-RPA targeting the 3´non-translated region of DENV1-4 was found to range from 14 (DENV4) to 241 (DENV1-3) RNA molecules detected. The assay was specific and did not cross detect other Flaviviruses. The RT-RPA assay was tested in a mobile laboratory combining magnetic-bead based total nucleic acid extraction and a portable detection device in Kedougou (Senegal) and in Bangkok (Thailand). In Kedougou, the RT-RPA was operated at an ambient temperature of 38 °C with auxiliary electricity tapped from a motor vehicle and yielded a clinical sensitivity and specificity of 98% (n=31) and 100% (n=23), respectively. While in the field trial in Bangkok, the clinical sensitivity and specificity were 72% (n=90) and 100%(n=41), respectively.
  
  
  CONCLUSIONS
  During the first 5 days of infection, the developed DENV1-4 RT-RPA assays constitute a suitable accurate and rapid assay for DENV diagnosis. Moreover, the use of a portable fluorescence-reading device broadens its application potential to the point-of-care for outbreak investigations.
authors.affiliation: Unit of Infection Models, German Primate Center, Goettingen, Germany; Department of Virology, Mansoura University, Dakahlia, Egypt.
authors.forename: Ahmed
authors.initials: A
authors.lastname: Abd El Wahed
authors.affiliation: CBS1-Highly Pathogenic Viruses, Center for Biological Threats and Special Pathogens, Robert Koch Institute, Berlin, Germany.
authors.forename: Pranav
authors.initials: P
authors.lastname: Patel
authors.affiliation: Arbovirus Unit, Pasteur Institute, Dakar, Senegal.
authors.forename: Oumar
authors.initials: O
authors.lastname: Faye
authors.affiliation: Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Sasikanya
authors.initials: S
authors.lastname: Thaloengsok
authors.affiliation: Department of Virology, University Medical Center, Goettingen, Germany.
authors.forename: Doris
authors.initials: D
authors.lastname: Heidenreich
authors.affiliation: Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand; Systems Biology of Diseases Research Unit at Faculty of Science and Center of Emerging and Neglected Infectious Diseases, Mahidol University, Bangkok, Thailand.
authors.forename: Ponpan
authors.initials: P
authors.lastname: Matangkasombut
authors.affiliation: Department of Microbiology, Faculty of Science, Mahidol University, Bangkok, Thailand.
authors.forename: Khajohnpong
authors.initials: K
authors.lastname: Manopwisedjaroen
authors.affiliation: Functional Genetics of Infectious Diseases Unit, Institute Pasteur, Paris, France.
authors.forename: Anavaj
authors.initials: A
authors.lastname: Sakuntabhai
authors.affiliation: Arbovirus Unit, Pasteur Institute, Dakar, Senegal.
authors.forename: Amadou A
authors.initials: AA
authors.lastname: Sall
authors.affiliation: Institute of Microbiology and Virology, Brandenburg Medical School Theodor-Fontane, Senftenberg, Brandenburg, Germany.
authors.forename: Frank T
authors.initials: FT
authors.lastname: Hufert
authors.affiliation: Institute of Aquaculture, University of Stirling, Stirling, Scotland, United Kingdom.
authors.forename: Manfred
authors.initials: M
authors.lastname: Weidmann
mesh_terms: D003715:Dengue / Q000175:diagnosis* / Q000821:virology*; D003716:Dengue Virus / Q000145:classification / Q000235:genetics*; D006801:Humans; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D012367:RNA, Viral / Q000235:genetics; D015203:Reproducibility of Results; D012675:Senegal; D012680:Sensitivity and Specificity; D013785:Thailand
chemical_list: D012367:RNA, Viral