A Rapid Assay for Hepatitis C Virus RNA Detection Using Reverse-Transcription Loop-Mediated Isothermal...

Area of Science:

• Molecular diagnostics
• Viral RNA detection
• Point-of-care testing

Background:

• Current Hepatitis C virus (HCV) diagnosis relies on antibody and RNA tests, causing care drop-offs.
• Existing HCV RNA tests are slow, expensive, and have limited availability.
• Need for rapid, cost-effective HCV RNA detection, especially for remote and marginalized populations.

Purpose of the Study:

• To develop a cost-effective, rapid, and sensitive assay for Hepatitis C virus (HCV) RNA detection.
• To improve HCV screening and diagnosis, particularly in underserved populations.

Main Methods:

• RNA extraction from plasma followed by reverse transcription and loop-mediated isothermal amplification (RT-LAMP).
• HCV RNA amplification using specific primers and detection via CRISPR-Cas12b with fluorescence readout.

Main Results:

• The assay detected HCV RNA across genotypes 1a, 1b, 2, 3a, and 4 with high sensitivity and specificity.
• Lower limit of detection (LLOD) for genotype 2 was 250 copies/ml; for clinical samples (genotype 1), it was ≥100 IU/ml within 45 minutes.
• Clinical sensitivity was 100% in 72 patient samples (including acute HCV and co-infections), with 100% specificity in 33 negative samples.

Conclusions:

• The developed assay demonstrates high specificity and sensitivity for direct HCV RNA detection from plasma.
• The test provides results within 45 minutes, enabling efficient global screening and diagnosis of HCV infection.