Journal
VACCINES
Volume 10, Issue 5, Pages -Publisher
MDPI
DOI: 10.3390/vaccines10050730
Keywords
SARS-CoV-2; COVID-19 testing; direct RT-qPCR; saliva
Categories
Funding
- Secretaria de Educacion, Ciencia, Tecnologia e Innovacion (SECTEI, CDMX) [SECTEI/277/219]
- Consejo Nacional de Ciencia y Tecnologia (CONACyT) [CVU: 1005898]
- SECTEI fellowships
- CONACyT
- Fondo FORDECyT-PRONACES
- DGAPA/PAPIIT-UNAM project [IN203820]
- CONACYT [CB-2016-01]
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This study optimized a buffer composition that allows for direct detection of SARS-CoV-2 from saliva without RNA extraction. The buffer showed good sensitivity and correlations in comparison to the standard RNA-extraction-based protocol. The study also proposed an auto-collection protocol for saliva samples and a multiplex reaction to minimize costs and processing time for massive testing.
Massive testing is a cornerstone in efforts to effectively track infections and stop COVID-19 transmission, including places with good vaccination coverage. However, SARS-CoV-2 testing by RT-qPCR requires specialized personnel, protection equipment, commercial kits, and dedicated facilities, which represent significant challenges for massive testing in resource-limited settings. It is therefore important to develop testing protocols that are inexpensive, fast, and sufficiently sensitive. Here, we optimized the composition of a buffer (PKTP), containing a protease, a detergent, and an RNase inhibitor, which is compatible with the RT-qPCR chemistry, allowing for direct SARS-CoV-2 detection from saliva without extracting RNA. PKTP is compatible with heat inactivation, reducing the biohazard risk of handling samples. We assessed the PKTP buffer performance in comparison to the RNA-extraction-based protocol of the US Centers for Disease Control and Prevention in saliva samples from 70 COVID-19 patients finding a good sensitivity (85.7% for the N1 and 87.1% for the N2 target) and correlations (R = 0.77, p < 0.001 for N1, and R = 0.78, p < 0.001 for N2). We also propose an auto-collection protocol for saliva samples and a multiplex reaction to minimize the PCR reaction number per patient and further reduce costs and processing time of several samples, while maintaining diagnostic standards in favor of massive testing.
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