Performance of the TaqMan COVID-19 Pooling Kit for detection of SARS-CoV-2 in asymptomatic and symptomatic populations

Abstract

Clinical evidence for asymptomatic cases of coronavirus disease (COVID-19) has reinforced the significance of effective surveillance testing programs. Quantitative reverse transcriptase polymerase chain reaction (RT-qPCR) assays are considered the 'gold standard' for detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA. However, the labor and resource requirements can be prohibitive with respect to large testing volumes associated with the pandemic. Pooled testing algorithms may serve to increase testing capacity with more efficient resource utilization. Due to the lack of carefully curated cohorts, there is limited evidence for the applicability of RT-PCR pooling in asymptomatic COVID-19 cases. In this study, we compared the analytical sensitivity of the TaqMan™ SARS-CoV-2 Pooling Assay to detect one positive sample in a pool of five anterior nares swabs in symptomatic and asymptomatic cohorts at an institute of higher education. Positive pools were deconvoluted and each individual sample was retested using the TaqPath™ COVID-19 Combo Kit. Both assays target the open reading frame (ORF) 1ab, nucleocapsid (N), and spike (S) gene of the strain that originated in Wuhan, Hubei, China. Qualitative results demonstrated absolute agreement between pooled and deconvoluted samples in both cohorts. Independent t-test performed on Ct shifts supported an insignificant difference between cohorts with p-values of 0.306 (Orf1ab), 0.147 (N), and 0.052 (S). All negative pools were correctly reported as negative. Pooled PCR testing up to five samples is a valid method for surveillance testing of students and staff in a university setting, especially when the prevalence is expected to be low.

Fig 1. Trend of C_t shift from pooled and deconvoluted result.

Pairwise comparisons of pooled and deconvoluted samples for each SARS-CoV-2 gene target from Asymptomatic (a,b,c) and Symptomatic cohorts (d,e,f); ORF1ab, circles; N gene, squares; S gene, triangles; mean C_t values, dashed intersects.

Fig 2. Asymptomatic and symptomatic C_t shift.

Box-and-whisker plot describing C_t shift (ΔC_t) for each gene from each cohort; whiskers, 5–95 percentiles; min-max range, dash mark; p-values annotated; A, Asymptomatic; S, Symptomatic; O, ORF1ab gene; N, N gene; S, S gene.