Sensitivity and Efficiency of RNA Sample Pooling for RT-qPCR Testing for SARS-CoV-2

Abstract

Background: Use of RT-qPCR pool testing in the diagnosis of SARS-CoV-2 infection has been limited due to a lack of data on its sensitivity and efficiency.Methods: We mixed single specimens of extracted RNA positive for the SARS-CoV-2 E gene by RT-qPCR with negative specimens, in pools of 4 (n=89), 8 (n=92), 16 (n=102), and 32 (n=105) specimens each. We estimated the average change in Ct for each pool size and added it to the Ct values of the first 1,350 tests in our lab, to obtain dilution-corrected Ct values. We estimated pool sensitivity as the proportion of samples with dilution-corrected Ct>40, and used it in simulations of the efficiency (tests used/true case detected) of binary split pool testing.Findings: We tested 388 pools. Average Ct changes were 2.21, 2.51, 3.27, and 3.94 cycles, for pools of 4, 8, 16, and 32 specimens, respectively. Corresponding pool tests sensitivities were 91.1%, 89.6%, 85.8% and 82.5%. Pool testing was substantially more efficient than individual testing. For prevalence of 0.5% to 2.0%, the efficiency of pools of ≥8 specimens was 30% to 280% higher, and the number of people tested was 4.4 to 13.9 times higher than those of individual testing were.Interpretation: Binary split pool testing substantially increases the number of people tested and the number of true cases detected per test. This strategy is key to curtail the transmission of SAR-CoV-2, by increasing efficiency in the identification and isolation of symptomatic and asymptomatic infected individuals.Funding: Major Office of Bucaramanga, Colombia.Declaration of Interests: None.Ethics Approval Statement: The study was approved by both the UIS’ and the University of Wisconsin-Madison’s Institutional Review Boards.