Different Respiratory Samples for COVID-19 Detection by Standard and Direct Quantitative RT-PCR: A Literature Review

Abstract

The most common diagnostic method for detecting severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection is real-time quantitative reverse transcriptase-polymerase chain reaction (RT-qPCR). Upper respiratory tract samples, including nasopharyngeal swab (NPS), oropharyngeal swab (OPS), saliva and lower respiratory tract samples such as sputum, are the most widely used specimens for diagnosis of SARS-CoV-2 using RT-qPCR. This study aimed to compare the diagnostic performance of different samples for Coronavirus disease 2019 (COVID-19) detection. It was found that NPS, the reference respiratory specimen for COVID-19 detection, is more sensitive than OPS. However, the application of NPS has many drawbacks, including challenging sampling process and increased risk of transmission to healthcare workers (HCWs). Saliva samples can be collected less invasively and quickly by HCWs with less contact or by own patients, and they can be considered as an alternative to NPS for COVID-19 detection by RT-qPCR. Additionally, sputum, which demonstrates higher viral load can be applied in patients with productive coughs and negative results from NPS. Commonly, after viral RNA purification from patient samples, which is time-consuming and costly, RT-qPCR is performed to diagnose SARS-CoV-2. Herein, different approaches including physical (heat inactivation) and chemical (proteinase K treatment) methods, used in RNA extraction free- direct RT-qPCR, were reviewed. The results of direct RT-qPCR assays were comparable to the results of standard RT-qPCR, while cost and time were saved. However, optimal protocol to decrease cost and processing time, proper transport medium and detection kit should be determined.

Keywords: COVID-19; Nasopharyngeal swab; Oropharyngeal swab; RT-qPCR; SARS-CoV-2; Sputum; saliva.

Figure 1

Different respiratory samples for COVID-19 detection. The respiratory tract samples for COVID-19 detection are divided into the upper (nasopharyngeal swab (NPS)/oropharyngeal swab (OPS), NP wash or saliva) and the lower (sputum) parts

Figure 2

Schematic representation of standard and direct RT-qPCR. In standard RT-qPCR, respiratory samples were collected, and in the case of swabs, they were transferred into a transport medium. Then, viral RNA was extracted, and an RT-qPCR reaction was performed. In direct RT-qPCR, the virus was inactivated by different methods, including heating, proteinase K treatment, pH changing, and then these inactivated samples were used for COVID-19 detection by RT-qPCR. In this process, time and cost were saved