SLAMP: A Rapid Fluorometric RT-LAMP Assay for Sensitive and Specific Detection of SARS-CoV-2 from Human Saliva

BackgroundDiagnostic testing can identify outbreaks and inform preventive strategies for slowing the spread of SARS-CoV-2, the virus that causes Covid-19. The "gold standard" method for detection of SARS-CoV-2 is reverse transcription quantitative polymerase chain reaction (RT-qPCR) performed on samples collected using nasopharyngeal (NP) swabs. While NP RT-qPCR achieves high sensitivity, it requires trained personnel to administer and suffers from lengthy time-to-result. Instead, rapid saliva-based reverse transcription loop-mediated amplification (RT-LAMP) screening methods may offer advantages in sample collection and speed. MethodsRegardless of symptomatic presentation, a total of 233 individuals were tested for SARS-CoV-2 using NP RT-qPCR, alongside saliva-based RT-qPCR (SalivirDetect) and RT-LAMP (SLAMP), a simple and rapid fluorometric RT-LAMP assay performed directly on heat-inactivated saliva without any additional treatments or RNA extraction. SLAMP is conducted in triplicate and takes 45 min. Samples found negative using both saliva-based methods but positive under CDC NP RT-qPCR above the saliva method LoD were excluded from evaluation, suggesting significant differences in viral titer between sampling sites. Individuals who consumed potential inhibitors in the form of food, drink, and oral health products within 30 min of sampling were identified using a self-reported questionnaire. ResultsOf the 233 NP RT-qPCR tests, 58 were positive and 175 were negative. Comparatively, SLAMP resulted in 95% sensitivity and 98% specificity and SalivirDetect 97% sensitivity and 98% specificity. Prior consumption had no measurable effect on test outcomes, except for drinking, which lowered Ct values in saliva. ConclusionsSLAMP requires less technician and instrument time than CDC-approved NP RT-qPCR and demonstrates that saliva-based RT-LAMP can enable frequent and rapid identification of pre-symptomatic and asymptomatic SARS-CoV-2 infections with high sensitivity and specificity.