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. 2020 Dec;26(12):1687.e1-1687.e5.
doi: 10.1016/j.cmi.2020.09.008. Epub 2020 Sep 10.

Sample pooling for SARS-CoV-2 RT-PCR screening

Adolfo de Salazar  1 , Antonio Aguilera  2 , Rocio Trastoy  2 , Ana Fuentes  1 , Juan Carlos Alados  3 , Manuel Causse  4 , Juan Carlos Galán  5 , Antonio Moreno  6 , Matilde Trigo  7 , Mercedes Pérez-Ruiz  8 , Carolina Roldán  9 , Maria José Pena  10 , Samuel Bernal  11 , Esther Serrano-Conde  1 , Gema Barbeito  2 , Eva Torres  3 , Cristina Riazzo  4 , Jose Luis Cortes-Cuevas  12 , Natalia Chueca  1 , Amparo Coira  2 , Juan Manuel Sanchez-Calvo  3 , Eduardo Marfil  4 , Federico Becerra  12 , María José Gude  6 , Ángeles Pallarés  7 , María Luisa Pérez Del Molino  2 , Federico García  13
Affiliations

Sample pooling for SARS-CoV-2 RT-PCR screening

Adolfo de Salazar et al. Clin Microbiol Infect. 2020 Dec.

Abstract

Objective: To evaluate the efficacy of sample pooling compared to the individual analysis for the diagnosis of coronavirus disease 2019 (COVID-19) by using different commercial platforms for nucleic acid extraction and amplification.

Methods: A total of 3519 nasopharyngeal samples received at nine Spanish clinical microbiology laboratories were processed individually and in pools (342 pools of ten samples and 11 pools of nine samples) according to the existing methodology in place at each centre.

Results: We found that 253 pools (2519 samples) were negative and 99 pools (990 samples) were positive; with 241 positive samples (6.85%), our pooling strategy would have saved 2167 PCR tests. For 29 pools (made out of 290 samples), we found discordant results when compared to their correspondent individual samples, as follows: in 22 of 29 pools (28 samples), minor discordances were found; for seven pools (7 samples), we found major discordances. Sensitivity, specificity and positive and negative predictive values for pooling were 97.10% (95% confidence interval (CI), 94.11-98.82), 100%, 100% and 99.79% (95% CI, 99.56-99.90) respectively; accuracy was 99.80% (95% CI, 99.59-99.92), and the kappa concordant coefficient was 0.984. The dilution of samples in our pooling strategy resulted in a median loss of 2.87 (95% CI, 2.46-3.28) cycle threshold (Ct) for E gene, 3.36 (95% CI, 2.89-3.85) Ct for the RdRP gene and 2.99 (95% CI, 2.56-3.43) Ct for the N gene.

Conclusions: We found a high efficiency of pooling strategies for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) RNA testing across different RNA extraction and amplification platforms, with excellent performance in terms of sensitivity, specificity and positive and negative predictive values.

Keywords: Pooled analysis; RT-PCR; SARS-CoV-2; Sample pooling; Sensitivity.

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Figures

Fig. 1
Fig. 1
Pooling effect on cycle threshold (Ct) results of the specific E, RdRP and N genes from Allplex 2019-nCoV Assay (Seegene, Seoul, South Korea).
See this image and copyright information in PMC

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References

    1. Zou L., Ruan F., Huang M., Liang L., Huang H., Hong Z. SARS-CoV-2 viral load in upper respiratory specimens of infected patients. N Engl J Med. 2020;382:1177–1179. - PMC - PubMed
    1. Pan Y., Zhang D., Yang P., Poon L.L.M., Wang Q. Viral load of SARS-CoV-2 in clinical samples. Lancet Infect Dis. 2020;20:411–412. - PMC - PubMed
    1. Dorfman R. The detection of defective members of large populations. Ann Math Stat. 1943;14:436–440.
    1. Xiong W., Ding J., He Y., Li Q. Improved matrix pooling. Stat Methods Med Res. 2019;28:211–222. - PubMed
    1. Nguyen N.T., Aprahamian H., Bish E.K., Bish D.R. A methodology for deriving the sensitivity of pooled testing, based on viral load progression and pooling dilution. J Transl Med. 2019;17:252. - PMC - PubMed