Evaluating a Modified Sputum Preparation Method for Improved qRT-PCR Detection of COVID-19

Rinjong Kendal; Pantitra  Sudadet; Jindarphorn  Suttayamuk; Panchanita Tuenkplee

doi:10.33165/rmj.48.03.e270736

Authors

Rinjong Kendal Department of Medical Technology, Khon Kaen Hospital, Khon Kaen, Thailand https://orcid.org/0009-0002-8471-6198
Pantitra Sudadet Department of Medical Technology, Khon Kaen Hospital, Khon Kaen, Thailand
Jindarphorn Suttayamuk Department of Medical Technology, Khon Kaen Hospital, Khon Kaen, Thailand
Panchanita Tuenkplee Department of Medical Technology, Khon Kaen Hospital, Khon Kaen, Thailand

DOI:

https://doi.org/10.33165/rmj.48.03.e270736

Keywords:

qRT-PCR, Sputum sample, Modified sample preparation method, COVID-19

Abstract

Background: An appropriate specimen is the crucial for quantitative reverse transcriptase–polymerase chain reaction (qRT-PCR) detection of COVID-19. Sputum is suitable specimen due to its noninvasive and simple collection process. For reference method, sputum samples were prepared by adding normal saline in a 1:2 ratio, followed by mixing and vortexing until fully dissolved. However, its sticky and vicious nature can hinder the preparation process and delay results. Therefore, a modified sample preparation was required during the COVID-19 pandemic.

Objective: To compare a modified sample preparation method with reference method for sputum specimens in the detection of COVID-19 by qRT-PCR.

Methods: Leftover sputum specimens were included and prepared in parallel using both the modified sample preparation and reference methods. qRT-PCR for COVID-19 was performed using the Seegene Allplex SAR-CoV2 master assay. Sensitivity, specificity, and other performance parameters were statistically analyzed. Cycle threshold (Ct) values of target genes (E, RdRp/S, N, and S variants) from both methods were compared.

Result: A total of 202 samples were examined. The modified sample preparation and reference methods yield positive results of 32 (15.8%) and 31 (15.3%), inconclusive results of 1 (0.5%) and 5 (2.5%), invalid results of 4 (2.0%) and 27 (13.4%), and negative results of 165 (81.7%) and 139 (68.8%), respectively. The sensitivity and specificity were 86.7% and 100%, respectively, with positive predictive value of 100% and negative predictive value of 97.1%. There were no statistically significant differences in Ct values for the E, RdRp/S, N, and S variants between the 2 methods with P values of .169, .421, .513, and .172, respectively.

Conclusions: The modified sample preparation method can be used during a COVID-19 pandemic while maintaining the reliable accuracy and reducing turnaround time.

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