Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay

Sayamon Hongjaisee, Natteewan Doungjinda, Woottichai Khamduang, Tanawan Samleerat Carraway, Jiraprapa Wipasa, Jose D. Debes, Khuanchai Supparatpinyo

Research output: Contribution to journalArticlepeer-review

13 Scopus citations

Abstract

Objectives: The aim was to develop a reverse transcription loop-mediated isothermal amplification (RT-LAMP) assay for the detection of hepatitis C virus (HCV) in a single closed tube. Methods: Plasma samples were collected from 200 HCV-infected patients. HCV-RNA was detected by one-step RT-LAMP processed at 65 °C for 60 min. The amplified products were detected by hydroxynaphthol blue (HNB)-dependent visual method and gel electrophoresis. Specificity was tested against other viruses. Sensitivity was determined using serial dilutions of extracted RNA. Results: The RT-LAMP assay detected 97.5% of HCV-RNA genotype 1, 91.1% of genotype 3, and 100% of genotype 6. The color change was evidenced with the naked eye. The assay demonstrated a clinical sensitivity of 95.5% and specificity of 100%, as well as no cross-reactivity with other viruses (i.e., hepatitis B virus, HIV). The limit of detection was as low as 10 ng per reaction for HCV genotypes 1a and 6, while it was 100 ng for genotype 3a. The assay showed a 100% detection threshold at a viral load of 5.00 log10 IU/mL in the clinical samples tested. Conclusions: This study demonstrated the use of an RT-LAMP assay for the detection of HCV in a simple, rapid, and cost-effective manner, which will be useful in resource-limited settings to allow the identification of individuals in need of HCV treatment.

Original languageEnglish (US)
Pages (from-to)440-445
Number of pages6
JournalInternational Journal of Infectious Diseases
Volume102
DOIs
StatePublished - Jan 2021

Bibliographical note

Funding Information:
The research reported in this publication was supported by the F ogarty International Center of the National Institutes of Health under grant number #D43TW009345 awarded to the Northern Pacific Global Health Fellows Program. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This work was also supported in part by Chiang Mai University, Thailand. The authors would like to express their gratitude to Nipapan Leetrakul, Blood Bank Section, Maharaj Nakorn Chiangmai Hospital, Chiangmai, Thailand.

Funding Information:
The research reported in this publication was supported by the Fogarty International Center of the National Institutes of Health under grant number #D43TW009345awarded to the Northern Pacific Global Health Fellows Program. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health. This work was also supported in part by Chiang Mai University, Thailand. The authors would like to express their gratitude to Nipapan Leetrakul, Blood Bank Section, Maharaj Nakorn Chiangmai Hospital, Chiangmai, Thailand.

Publisher Copyright:
© 2020 The Author(s)

Keywords

  • Hepatitis C virus
  • Hydroxynaphthol blue
  • Loop-mediated isothermal amplification
  • Reverse transcription

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