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Field-applicable simultaneous multiplex LAMP assay for screening HBV and HCV co-infection in a single tube.
- Source :
-
BMC infectious diseases [BMC Infect Dis] 2024 Aug 09; Vol. 24 (1), pp. 805. Date of Electronic Publication: 2024 Aug 09. - Publication Year :
- 2024
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Abstract
- Background: Globally, around 7 to 20 million people are believed to be suffering from coinfection with both hepatitis B virus (HBV) and hepatitis C virus (HCV). The loop-mediated isothermal amplification (LAMP) approach, introduced by Notomi and colleagues, has undergone substantial advancements as an effective molecular tool that enables the simultaneous analysis of multiple samples in a single tube.<br />Methods: The present study examined the simultaneous detection of HBV and HCV in a single tube using melt curve analysis multiplex LAMP (mLAMP), which is based on the identification of unique melting peak temperatures. Selected regions for primer design including the S gene of HBV and the UTR gene of HCV. Primer optimization is initially performed through individual HBV and HCV LAMP analysis. Following the optimization process, the mLAMP assay was evaluated by optimizing the multiplex reaction mixture, determining the reaction time, and analyzing the limit of detection (LOD). The results are also analyzed using lateral flow dipsticks (LFD), which enable the visual detection of HBV and HCV by adding 20 pmol FITC-labeled LF primers into the reaction mixture prior the mLAMP.<br />Results: The LOD for the mLAMP assay was determined as 10 copies/µl, and no cross-reactivity with other microorganisms was detected. The detection results obtained from patient plasma were also visually demonstrated using LFD, and displayed significant concordance with those obtained from Real-Time Polymerase Chain Assay. The mLAMP assay revealed a diagnostic sensitivity of 95% for detecting the HBV, and LOD is 90% for HCV. The overall diagnostic sensitivity of the mLAMP assay for both viruses was 85%. The assay confirmed a specificity of 100%.<br />Conclusion: The mLAMP assay displays significant promise for analyzing coinfected samples by simultaneously detecting the dual targets HBV and HCV within a set temperature of 62 °C, all within a time frame of 1 h. Additionally, when paired with disposable LFD, the mLAMP assay enables rapid visual detection of assay results in a matter of minutes. The result contributes to the mLAMP assay being highly suitable for coinfection screening, particularly in field conditions.<br /> (© 2024. The Author(s).)
- Subjects :
- Humans
Limit of Detection
DNA Primers genetics
Nucleic Acid Amplification Techniques methods
Hepatitis C diagnosis
Hepatitis C virology
Hepatitis C complications
Hepatitis B diagnosis
Hepatitis B virology
Hepatitis B virus genetics
Hepatitis B virus isolation & purification
Hepacivirus genetics
Hepacivirus isolation & purification
Coinfection diagnosis
Coinfection virology
Molecular Diagnostic Techniques methods
Sensitivity and Specificity
Subjects
Details
- Language :
- English
- ISSN :
- 1471-2334
- Volume :
- 24
- Issue :
- 1
- Database :
- MEDLINE
- Journal :
- BMC infectious diseases
- Publication Type :
- Academic Journal
- Accession number :
- 39123117
- Full Text :
- https://doi.org/10.1186/s12879-024-09567-8