Rapid Detection of Knowlesi Malaria Using Room-Temperature Stable and Ready-to-Use Colorimetric Loop-Mediated Isothermal Amplification Reagents.

A room temperature-stable and ready-to-use dried-format loop-mediated isothermal amplification (LAMP) diagnostic tool is proposed for detecting knowlesi malaria in the present study. Traditional LAMP assays require a cold chain to maintain the stability of their reagents during storage and transport. This requirement limits the use of LAMP-based methods in environments with limited infrastructure. Results can be obtained by directly observing color changes without the need for specialized instruments; positive samples appear yellow, whereas negative reactions remain pink. The developed dehydrated colorimetric LAMP was tested using 75 clinical blood samples collected from district and state hospitals in Malaysia. By using nested polymerase chain reaction-based assays as a reference method, dehydrated LAMP exhibited 100% sensitivity (95% CI: 92.1-100.0%) and 100% specificity (95% CI: 88.4-100.0%). The dehydrated LAMP reagents can be stored at room temperature for up to 15 days. This dehydrated LAMP assay offers an ideal diagnostic tool for resource-limited settings where knowlesi malaria is endemic.