Cycle threshold values in RT-PCR to determine dynamics of SARS-CoV-2 viral load: An approach to reduce the isolation period for COVID-19 patients

Abstract

Severe acute respiratory syndrome coronavirus (SARS-CoV-2) has affected all inhabited continents, and India is currently experiencing a devastating second wave of coronavirus disease-2019 (COVID-19). Here, we examined the duration of clearance of SARS-CoV-2 in respiratory samples from 207 infected cases by real-time reverse-transcription polymerase chain reaction (RT-PCR). A substantial proportion of COVID-19 positive cases with cycle threshold (Ct) values more than or equal to 31 (45.7%) were subsequently tested negative for SARS-CoV-2 RNA within 7 days of initial detection of the viral load. A total of 60% of all the patients with COVID-19, irrespective of their Ct values, cleared SARS-CoV-2 RNA within 14 days of the initial detection. Longitudinal assessment of RT-PCR test results in individuals requiring 15-30 days to clear SARS-CoV-2 RNA showed a significant reduction of the viral load in samples with high or intermediate viral loads (Ct values ≤ 25 and between 26 and 30, respectively) but the follow-up group with low viral RNA (Ct values ≥ 31) exhibited a stable viral load. Together, these results suggest that COVID-19 positive cases with Ct values more than or equal to 31 require reduced duration to clear SARS-CoV-2, and thus, a shorter isolation period for this group might be considered to facilitate adequate space in the COVID Care Centres and reduce the burden on healthcare infrastructure.

Keywords: COVID 19; RT-PCR; SARS-CoV-2; cycle threshold value; isolation.

Figure 1

Distribution of COVID‐19 cases that tested negative when followed up over a period of 30 days at 3 days, 7 days, between 8 and 14 days, and between 15 and 30 days. Stacked graph bar chart depicts the COVID‐19 cases that tested negative for SARS‐CoV‐2 RNA in individuals with Ct values of ≤25, 26–30, and ≥31, respectively at the mentioned follow‐up days. COVID‐19, coronavirus disease 2019; SARS‐CoV‐2, severe acute respiratory syndrome coronavirus 2

Figure 2

Cycle threshold (Ct) values for the O and E genes at two‐time intervals of Day 0 (initial day of testing) and Day 8–14 in follow‐up patients that required 15–30 days to clear SARS‐CoV‐2 viral load. O‐gene stands for ORF1, a nonstructural region that is specific for SARS‐CoV‐2 (target 1) and E‐gene stands for a structural envelope gene that is common to all Sarbecovirus subgenus (target 2). Scatter plot showing Ct values for O and E genes with (A) Ct values of ≤25 (n = 19), (B) Ct values between 26 and 30 (n = 13), and (C) Ct values of ≥31 (n = 29). RT‐PCR tests between 8 and 14 days revealed a significantly lowered SARS‐CoV‐2 viral load in individuals with initial Ct values of ≤25 and Ct values between 26 and 30 (p < 0.0001), but there was no significant change in the viral load of individuals with initial Ct values of ≥31 (nonsignificant). Each dot represents an individual and the horizontal lines indicate mean values. The Mann–Whitney test was used as the test of significance. RT‐PCR, reverse‐transcription polymerase chain reaction; SARS‐CoV‐2, severe acute respiratory syndrome coronavirus 2