A Novel Saliva RT-LAMP Workflow for Rapid Identification of COVID-19 Cases and Restraining Viral Spread

Kobayashi, Gerson Shigeru; Brito, Luciano Abreu; Moreira, Danielle de Paula; Suzuki, Angela May; Hsia, Gabriella Shih Ping; Pimentel, Lylyan Fragoso; de Paiva, Ana Paula Barreto; Dias, Carolina Regoli; Lourenço, Naila Cristina Vilaça; Oliveira, Beatriz Araujo; Manuli, Erika Regina; Corral, Marcelo Andreetta; Cavaçana, Natale; Mitne-Neto, Miguel; Sales, Maria Mirtes; Dell’ Aquila, Luiz Phellipe; Filho, Alvaro Razuk; Parrillo, Eduardo Fagundes; Mendes-Corrêa, Maria Cássia; Sabino, Ester Cerdeira; Costa, Silvia Figueiredo; Leal, Fabio Eudes; Sgro, Germán Gustavo; Farah, Chuck Shaker; Zatz, Mayana; Passos-Bueno, Maria Rita

Rapid diagnostics is pivotal to curb SARS-CoV-2 transmission, and saliva has emerged as a practical alternative to naso/oropharyngeal (NOP) specimens. We aimed to develop a direct RT-LAMP (reverse transcription loop-mediated isothermal amplification) workflow for viral detection in saliva, and to provide more information regarding its potential in curbing COVID-19 transmission. Clinical and contrived specimens were used to optimize formulations and sample processing protocols. Salivary viral load was determined in symptomatic patients to evaluate the clinical performance of the test and to characterize saliva based on age, gender and time from onset of symptoms. Our workflow achieved an overall sensitivity of 77.2% (n = 90), with 93.2% sensitivity, 97% specificity, and 0.895 Kappa for specimens containing >10(2) copies/μL (n = 77). Further analyses in saliva showed that viral load peaks in the first days of symptoms and decreases afterwards, and that viral load is ~10 times lower in females compared to males, and declines following symptom onset. NOP RT-PCR data did not yield relevant associations. This work suggests that saliva reflects the transmission dynamics better than NOP specimens, and reveals gender differences that may reflect higher transmission by males. This saliva RT-LAMP workflow can be applied to track viral spread and, to maximize detection, testing should be performed immediately after symptoms are presented, especially in females.