Detection of Monkeypox virus Using Helicase Dependent Amplification and Recombinase Polymerase Amplification Combined with Lateral Flow Test

Abstract

The monkeypox virus (MPXV) is a zoonotic DNA virus that belongs to the poxvirus family. Conventional laboratory methods for detecting MPXV are complex and expensive, making them unsuitable for detecting the virus in regions with limited resources. In this study, we using the Helicase dependent amplification (HDA) method and the Recombinase polymerase amplification (RPA) technique in combination with the lateral flow test (LFT), together with a self-designed qPCR technique for the detection of the monkeypox virus-specific conserved fragment F3L, to compare the sensitivity and specificity of the three assays. The assay results showed that the detection limits for HDA-LFT and RPA-LFT were ༜ 10 copies/µL and for qPCR ༜ 500 copies/µL. The three methods described above showed good specificity in the detection of monkeypox viruses among their congeners. The simple, highly sensitive, and specific MPXV assay developed in this study is anticipated to provide a solid foundation for future applications in the early screening, diagnosis, and evaluation of the efficacy of MPXV. This is the first time the HDA-LFT assay has been utilized to detect MPXV infection.