Portable microfluidic-LAMP assay for rapid on-site detection of eight highly pathogenic viruses

Background

The recent global spread of highly pathogenic viruses, such as Mpox and Ebola viruses, highlights the urgent need for multiplex detection technologies. Current methods lack the capability for simultaneous identification, which hampers outbreak control in resource-limited settings and global surveillance efforts. Traditional techniques, like real-time quantitative polymerase chain reaction (qPCR), are often impractical in resource-limited areas due to laboratory dependency, and specialized personnel requirements. A portable, rapid, and cost-effective method for multiplex field detection of highly pathogenic viruses is critically needed to address the growing threat of outbreaks in resource-limited settings.

Results

In this study, a simple and rapid nucleic acid extraction method was performed using a paper-based dipstick, which was equipment-free and could be completed within 2 min. Subsequently, a microfluidic chip was integrated with loop-mediated isothermal amplification (LAMP) technology to develop a field-deployable, portable detection method termed the microfluidic-LAMP assay. This assay allows for the simultaneous detection of eight highly pathogenic viruses: Ebola virus (EBOV), Marburg virus (MARV), Rift Valley fever virus (RVFV), Lassa virus (LASV), yellow fever virus (YFV), Congo Basin Mpox virus (C-MPXV), West African Mpox virus (W-MPXV), and variola virus (VARV), all within a span of 70 min. The assay demonstrated a sensitivity of 7.02 × 10²-8 × 10⁴ copies/mL across different targets using simulated clinical samples. The limits of detection of this field available assay were comparable to or lower than those of the real-time quantitative PCR. The microfluidic-LAMP assay demonstrated good specificity in cross-tests with 21 pathogens, including the above 8 viruses and 13 other respiratory pathogens.

Significance

This study presents the first portable microfluidic-LAMP assay capable of simultaneously detecting eight highly pathogenic viruses in resource-limited settings. The assay's equipment-free nucleic acid extraction (2 min) and rapid detection (70 min) make it a practical solution for on-site detection in resource-limited regions, facilitating outbreak control and global surveillance efforts.