09/01/2021

Correlation Between 3790 Quantitative Polymerase Chain Reaction–Positives Samples and Positive Cell Cultures, Including 1941 Seve…

CORRECTED PROOF

Correlation Between 3790 Quantitative
Polymerase Chain Reaction–Positives
Samples and Positive Cell Cultures,
Including 1941 Severe Acute Respiratory
Syndrome Coronavirus 2 Isolates 
Rita Jaafar, Sarah Aherfi, Nathalie Wurtz, Clio Grimaldier,Thuan Van Hoang, Philippe Colson, Didier Raoult, Bernard La Scola Clinical Infectious Diseases, ciaa1491,
https://doi.org/10.1093/cid/ciaa1491
Published: 28 September 2020 Article history Issue Section: Correspondence

TO THE EDITOR—The outbreak of the coronavirus disease 2019
(COVID-19) pandemic due to severe acute respiratory syndromecoronavirus 2 (SARS-CoV-2) was declared a pandemic on 12 March
2020 by the World Health Organization [1]. A major issue related tothe outbreak has been to correlate viral RNA load obtained after
reverse-transcription polymerase chain reaction (RT-PCR) and
expressed as the cycle threshold (Ct) with contagiousness andtherefore duration of eviction from contacts and discharge fromspecialized infectious disease wards. Several recent publications,based on more than 100 studies, have attempted to propose a cutoPDF
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Ct value and duration of eviction, with a consensus at approximately
Ct >30 and at least 10 days, respectively [2–5]. However, in an article
published in Clinical Infectious Diseases, Bullard et al reported thatpatients could not be contagious with Ct >25 as the virus is notdetected in culture above this value [6]. This limit was then evoked in
the French media during an interview with a member of the French
Scienti c Council Covid-19 as a possible value above which patientsare no longer contagious [7].
At the beginning of the outbreak, we correlated Ct values obtainedusing our PCR technique based on ampli cation of the E gene and thehttps://academic.oup.com/cid/advance-article/doi/10.1093/cid/ciaa1491/5912603

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Correlation Between 3790 Quantitative Polymerase Chain Reaction–Positives Samples and Positive Cell Cultures, Including 1941 Seve…

results of the culture [8]. Since the beginning of the pandemic, wehave performed 250 566 SARS-CoV-2 RT-PCR for 179 151 patients, ofwhom 13 161 (7.3%) tested positive. Up to the end of May, 3790 ofthese samples, reported as positive on nasopharyngeal samples, wereinoculated and managed for culture as previously described [8]. Of
these 3790 inoculated samples, 1941 SARS-CoV-2 isolates could beobtained after the rst inoculation or up to 2 blind subcultures. The
correlation between the scanner values and the positivity of theculture allows us to observe that the image obtained with 10 times
more isolates than in our preliminary work (1941 vs 129) does notchange signi cantly (Figure 1). It can be observed that at Ct = 25, up
to 70% of patients remain positive in culture and that at Ct = 30 thisvalue drops to 20%. At Ct = 35, the value we used to report a positive
result for PCR, <3% of cultures are positive. Our Ct value of 35,initially based on the results obtained by RT-PCR on control negative
samples in our laboratory and initial results of cultures [8], isvalidated by the results herein presented and is in correlation withwhat was proposed in Korea [9] and Taiwan [10]. We could observethat subcultures, especially the rst one, allow an increasing
percentage of viral isolation in samples with Ct values, con rmingthat these high Ct values are mostly correlated with low viral loads.
From our cohort, we now need to try to understand and de ne theduration and frequency of live virus shedding in patients on a caseby-case basis in the rare cases when the PCR is positive beyond 10days, often at a Ct >30. In any cases, these rare cases should not
impact public health decisions.
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Correlation Between 3790 Quantitative Polymerase Chain Reaction–Positives Samples and Positive Cell Cultures, Including 1941 Seve…

Figure 1.

Percentage of positive viral cultures of severe acute respiratory syndrome
coronavirus 2 polymerase chain reaction–positive nasopharyngeal samples fromcoronavirus disease 2019 patients, according to Ct value (plain line). The dashedcurve indicates the polynomial regression curve. Abbreviations: Ct, cyclethreshold; Poly., polynomial.

Notes
Ethical approval. The protocol was approved by the University
Hospital Institute Méditerranée Infection Ethical Committee. All
patients provided informed consent in accordance with theDeclaration of Helsinki.
Financial support. This research was funded by the French

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Government under the Investissements d’avenir (Investments for theFuture) program managed by the Agence Nationale de la Recherche(French National Agency for Research; reference: Méditerranée
Infection 10-IAHU-03) and by Région Provence-Alpes-Côte d’Azurand European funding Fond Europen de Recheche et dedeveloppement (FEDER) PRIMI.
Potential con icts of interest. D. R. reports grants from the Hitachi
High-Tech Corporation outside the submitted work. All other authors
report no potential con icts. All authors have submitted the ICMJE
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Form for Disclosure of Potential Con icts of Interest. Con icts thatthe editors consider relevant to the content of the manuscript havebeen disclosed.

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