TY - JOUR
T1 - Rapid visual detection of hepatitis C virus using a reverse transcription loop-mediated isothermal amplification assay
AU - Hongjaisee, Sayamon
AU - Doungjinda, Natteewan
AU - Khamduang, Woottichai
AU - Carraway, Tanawan Samleerat
AU - Wipasa, Jiraprapa
AU - Debes, Jose D.
AU - Supparatpinyo, Khuanchai
N1 - Publisher Copyright:
© 2020 The Author(s)
PY - 2021/1
Y1 - 2021/1
N2 - 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.
AB - 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.
KW - Hepatitis C virus
KW - Hydroxynaphthol blue
KW - Loop-mediated isothermal amplification
KW - Reverse transcription
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U2 - 10.1016/j.ijid.2020.10.082
DO - 10.1016/j.ijid.2020.10.082
M3 - Article
C2 - 33130211
AN - SCOPUS:85097065349
SN - 1201-9712
VL - 102
SP - 440
EP - 445
JO - International Journal of Infectious Diseases
JF - International Journal of Infectious Diseases
ER -
