Bridging the Diagnostic Gap: A Rapid, Cost-Effective, and Equitable Hepatitis C Virus RNA Detection Method for Resource-Limited Settings.

Hepatitis C virus (HCV) remains a major global health challenge, particularly in resource-limited settings where access to molecular diagnostics is restricted. The loop-mediated isothermal amplification (LAMP) assay offers a rapid, cost-effective alternative to polymerase chain reaction (PCR) for HCV RNA detection. Unlike PCR, LAMP uses isothermal amplification (60-65°C), eliminating the need for expensive thermal cyclers and yielding results in under 60 minutes. We evaluated two LAMP detection methods - hydroxynaphthol blue (HNB, colorimetric) and calcein (fluorescent) - against an in-house TaqMan qPCR assay. Both LAMP methods demonstrated high sensitivity (99.6%) and specificity (95.6% for HNB, 99.2% for calcein), with a broad dynamic range (10-10⁶ copies/mL). Clinical validation against real-time PCR showed strong agreement, with a positive predictive value of 98.6% and a negative predictive value of 94.4%. The HNB-LAMP assay, in particular, provides a simple visual readout (blue to sky blue), requiring no specialized equipment, while calcein-LAMP offers fluorescence-based detection under UV light. Both methods showed no significant correlation (p > 0.5), confirming their reliability. LAMP's advantages - minimal infrastructure, ambient temperature stability, and low cost (<$5 per test) - make it ideal for decentralized testing in low-resource settings. This approach could revolutionize HCV diagnosis by enabling same-day test-and-treat strategies, improving linkage to care, and supporting global HCV elimination efforts. Future steps include field validation in remote clinics, manufacturing scale-up, and integration into point-of-care platforms to maximize accessibility. By bridging the diagnostic gap, LAMP can potentially transform HCV management in underserved regions worldwide.