Isothermal nucleic acid amplification for monitoring hand-foot-and-mouth disease: current status and future implications

With the global prevalence of the hand-foot-and-mouth disease (HFMD) epidemic, the development of reliable point-of-care testing (POCT) is crucial for the timely identification and prevention of outbreaks. Isothermal nucleic acid amplification techniques (INAATs) have attracted much attention because of their high efficiency for rapid diagnosis. In this work, we systematically summarize the current status of INAATs for HFMD and discuss advantages and drawbacks of various INAATs for HFMD. The INAATs for HFMD detection mainly include loop-mediated isothermal amplification (LAMP), simultaneous amplification and testing (SAT), and recombinase polymerase amplification (RPA). Among them, LAMP has excelled in several diagnostic metrics and has made significant progress in the field of POCT. SAT has been effective in overcoming the problem of RNA degradation. RPA is suited for on-site testing due to its rapid amplification rate and low reaction temperature. In addition, this study explores the potential of INAATs in lateral flow strips (LFS) test and microfluidic devices for HFMD. LFS is typically used for qualitative analysis and supports multiple detection. Microfluidics can integrate necessary processes of sample pre-processing, amplification, and signal output, enabling high-throughput qualitative or quantitative detection and demonstrating the potential of monitoring HFMD. We hope the current work will provide insights into INAATs for monitoring HFMD and serve as a reference for the implementation of on-site EV detection for public health.