Related Experiment Video
Updated: Sep 30, 2026
Measuring Dengue Virus RNA in the Culture Supernatant of Infected Cells by Real-time Quantitative Polymerase Chain Reaction
Published on: November 1, 2018
Development and evaluation of reverse transcription loop-mediated isothermal amplification for rapid and real-time
Jyoti S Kumar1, Pragya D Yadav2, Anita M Shete2
1Division of Virology, Defence Research and Development Establishment, Gwalior 474002, India.
Significance:
Kyasanur forest disease (KFD), a re-emerging tick-borne viral disease, causes severe hemorrhagic fever in humans and nonhuman primates. KFD virus (KFDV) is a member of the genus Flavivirus. KFD is now increasingly reported outside its endemic zone in India. Rapid and specific detection of the KFDV plays a critical role in containment of the outbreak. The diagnosis of KFD currently relies on real-time RT-PCR, nested RT-PCR, end point RT-PCR, and serodiagnostic assay. These assays are tedious, time-consuming, and cannot be used as a routine screening platform.
Objective:
The present study was aimed to develop a one-step reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for molecular diagnosis of KFD.
Design:
The gene amplification reaction was accomplished by incubation at a constant temperature of 63°C for 60min.
Results:
The limit of detection of RT-LAMP assay was 10 copies. KFD RT-LAMP assay was successfully evaluated with diverse host samples including humans, monkeys, and tick. The assay correctly picked up different KFD isolates indicating its applicability for divergent strains. Comparative evaluation of RT-LAMP assay with quantitative TaqMan real-time RT-PCR revealed 100% concordance. No cross-reaction with related flavi and other hemorrhagic fever viruses was observed, indicating its high specificity.
Conclusion And Relevance:
The RT-LAMP test developed in this study will serve as a rapid, sensitive alternate detection method for KFDV infection and would be useful for high throughput screening of clinical samples in resource limited areas during outbreaks.
