Profiling RT-LAMP tolerance of sequence variation for SARS-CoV-2 RNA detection

The ongoing SARS-CoV-2 pandemic has necessitated a dramatic increase in our ability to conduct molecular diagnostic tests, as accurate detection of the virus is critical in preventing its spread. However, SARS-CoV-2 variants continue to emerge, with each new variant potentially affecting widely-used nucleic acid amplification diagnostic tests. RT-LAMP has emerged as a quick, inexpensive diagnostic alternative to RT-qPCR, but has not been studied as thoroughly. Here we interrogate the effect of SARS-CoV-2 sequence mutations on RT-LAMP amplification, creating 572 single point mutation “variants” covering every position of the LAMP primers in 3 SARS-CoV-2 assays and analyzing their effects with over 4,500 RT-LAMP reactions. Remarkably, we observed only minimal effects on amplification speed and no effect on detection sensitivity, highlighting RT-LAMP as an extremely robust technique for viral RNA detection. Additionally, we describe the use of molecular beacons to sensitively identify variant RNA sequences. Together these data add to the growing body of knowledge on the utility of RT-LAMP and increase its potential to further our ability to conduct molecular diagnostic tests outside of the traditional clinical laboratory environment.