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SalivaSTAT: Direct-PCR and Pooling of Saliva Samples Collected in Healthcare and Community Setting for SARS-CoV-2 Mass Surveillance

Authors :
Nikhil S. Sahajpal
Ashis K. Mondal
Sudha Ananth
Allan Njau
Pankaj Ahluwalia
Gary Newnam
Adriana Lozoya-Colinas
Nicholas V. Hud
Vamsi Kota
Ted M. Ross
Michelle D. Reid
Sadanand Fulzele
Alka Chaubey
Madhuri Hegde
Amyn M. Rojiani
Ravindra Kolhe
Source :
Diagnostics, Vol 11, Iss 5, p 904 (2021)
Publication Year :
2021
Publisher :
MDPI AG, 2021.

Abstract

Objectives: Limitations of widespread current COVID-19 diagnostic testing exist in both the pre-analytical and analytical stages. To alleviate these limitations, we developed a universal saliva processing protocol (SalivaSTAT) that would enable an extraction-free RT-PCR test using commercially available RT-PCR kits. Methods: We optimized saliva collection devices, heat-shock treatment, and homogenization. Saliva samples (879) previously tested using the FDA-EUA method were reevaluated with the optimized SalivaSTAT protocol using two widely available commercial RT-PCR kits. A five-sample pooling strategy was evaluated as per FDA guidelines. Results: Saliva collection (done without any media) showed performance comparable to that of the FDA-EUA method. The SalivaSTAT protocol was optimized by incubating saliva samples at 95 °C for 30-min and homogenization, followed by RT-PCR assay. The clinical sample evaluation of 630 saliva samples using the SalivaSTAT protocol with PerkinElmer (600-samples) and CDC (30-samples) RT-PCR assay achieved positive (PPA) and negative percent agreements (NPAs) of 95.0% and 100%, respectively. The LoD was established as ~60–180 copies/mL by absolute quantification. Furthermore, a five-sample-pooling evaluation using 250 saliva samples achieved a PPA and NPA of 92% and 100%, respectively. Conclusion: We have optimized an extraction-free RT-PCR assay for saliva samples that demonstrates comparable performance to FDA-EUA assay (Extraction and RT-PCR).

Details

Language :
English
ISSN :
20754418
Volume :
11
Issue :
5
Database :
Directory of Open Access Journals
Journal :
Diagnostics
Publication Type :
Academic Journal
Accession number :
edsdoj.35618974a0ff410f917919e1e553da3b
Document Type :
article
Full Text :
https://doi.org/10.3390/diagnostics11050904