Rapid, low-cost, and portable molecular detection of HIV-1 subtype B and HCV 1a directly from blood

Accessible molecular diagnostics for human immunodeficiency virus-1 (HIV-1) and hepatitis C virus (HCV) remain constrained by the cost and infrastructure requirements of existing molecular point-of-care platforms. In this study, we describe a portable, proof-of-concept molecular workflow with a total runtime of 32 min for the simultaneous qualitative detection of HIV-1 subtype B and HCV genotype 1a directly from contrived whole blood. The workflow employs a battery-powered heating device, ambient-stable reagents, and lateral-flow dipstick readout. Contrived whole blood samples were diluted twofold with nuclease-free water and added directly to rehydrated reverse-transcription loop-mediated amplification (RT-LAMP) reactions without nucleic acid extraction, and reaction parameters were systematically varied to determine lower limits of detection. Both viral targets were consistently detected at clinically relevant input levels following a 30 min isothermal amplification step, with visual lateral flow results available within 2 min. In addition, CRISPR-Cas12a detection was used as an orthogonal method to verify RT-LAMP target amplicons during assay development. Together, this work establishes the analytical feasibility of a rapid, low-cost, portable molecular workflow and provides a foundation for future development of decentralized HIV-1 and HCV molecular diagnostics.IMPORTANCEOngoing changes in human immunodeficiency virus (HIV) and hepatitis C virus (HCV) epidemiology highlight the need for more adaptable and accessible diagnostic tools. In this study, we developed a fast, low-cost test that detects HIV-1 and HCV from a small blood sample. The test is designed to work outside traditional laboratories, using minimal equipment. It does not require complex sample preparation, refrigeration, or expensive machines, and produces results in about 30 min. The workflow combines a rapid genetic amplification method with an easy-to-read test strip and a small, battery-powered heater, making it portable and suitable for use in low-resource or remote settings. An additional molecular method was used during development to confirm that the test was accurately detecting the correct viral targets. By focusing on simplicity, speed, and affordability, this work aims to expand access to reliable HIV and HCV testing in settings where these infections often occur together and laboratory access is limited.