Shedding of infectious virus in hospitalized patients with COVID-19

COVID-19: Shedding of infectious virus in hospitalized patients with COVID-19 Jefferson T, Spencer EA.  Heneghan C.

https://www.cebm.net/study/shedding-of-infectious-virus-in-hospitalized-patients-with-covid-19/

Published on August 11, 2020

Reference van Kampen JJA, van de Vijver DAMC, Fraaij PLA, Haagmans BL, Lamers MM, Okba N, et al. Shedding of infectious virus in hospitalized patients with coronavirus disease-2019 (COVID-19): duration and key determinants. medRxiv. 2020:2020.06.08.20125310. 2020
Study type
Country Netherlands
Setting Hospital
Funding Details EU COVID-19 grant RECOVER
Transmission mode Viral load
Exposures

Bottom Line

Patients with severe or critical COVID-19 may shed infectious virus for longer periods of time compared to what has been reported for in patients with mild COVID-19. Quantitative viral RNA load assays and serological assays should be used for test-based strategies to discontinue or de-escalate infection prevention and control precautions.

Evidence Summary

The presence of infectious virus was tested in 690 respiratory samples from 129 patients using cell culture and determined the viral RNA load with RT-qPCR. 

Of these 62 samples yielded Infectious SARS-CoV-2 from respiratory tract samples (9.0%) of 23 patients (17,8%). 

Median time of infectious virus shedding was 8 days post onset symptoms (IQR 5 – 11, range 0 – 20) and probability of ≤ 5% for isolating infectious SARS-CoV-2 when the duration of symptoms was 15.2 days (95% CI 13.4 to 17.2) or more. 

The median viral load was significantly higher in culture positive samples than in culture negative samples (8.1 versus 5.9  Log10 RNA copies/mL, p<0,0001).  

The probability of isolating infectious SARS-CoV-2 was less than 5% when the viral load was below 6.6 Log10 RNA copies/mL (95% CI 6.2 – 6.9).

For conversion of logs see:

Log10 copies/mL
8 100,000,000
7 10,000,000
100,0000
5 10,000
4 1,000
3 100
2 10

In 112 serum samples from 27 patients, neutralizing antibody titers were obtained on the same day as a respiratory tract sample was available. The probability of isolating infectious virus was less than 5% when the neutralizing antibody titer was 1:80 or higher. RT-PCRs detecting SARS-CoV-2 subgenomic messenger RNA in the 112 corresponding respiratory tract samples outlasted the detection of infectious virus and was a poor predictor of positive virus cultures (PPV 37,5%).

What did they do?

The study looked at the shedding of infectious virus in 129 severely ill COVID-19 patients aged around 65. 

The authors used viral cultures on sputum and upper respiratory tract samples and addressed the relationship and predictive value of viral RNA load in respiratory tract samples, detection of subgenomic viral RNA in respiratory tract samples, neutralizing antibody titer in serum, the duration of symptoms, and/or immunocompromised status on virus shedding.  

Between March 8th 2020 and April 8th 2020, diagnostic respiratory samples of COVID-19 patients from the Erasmus Medical Centre were sent to the laboratory for PCR on diagnostic respiratory samples and for SARS-CoV-2 neutralizing antibodies. These samples were also submitted for virus culture. 

The following information recorded from the patient files: date of onset of symptoms, disease severity, immunocompetence and whether the patients were still alive by April 17th 2020.

Cycle threshold (ct) values were converted to Log10 RNA copies/mL using calibration curves based on quantified E-gene in vitro RNA transcripts, subgenomic RNAs were detected with RT-PCR. 

Isolation of infectious SARS-CoV-2 was done from Vero cells from respiratory tract samples.18 Samples were cultured for seven days and when cytopathic effect was visible, the presence of SARS-CoV-2 was confirmed with immunofluorescent detection of nucleocapsid proteins. A viral load exceeding 7 Log10 RNA copies/mL (10,000,000 copies per ml)  less than 7 days of symptoms, absence of serum neutralizing antibodies and being immunocompromised were all associated with a positive virus culture in univariate analysis. 

In a multivariate analysis only a viral load above 7 Log10 RNA copies/mL and absence of serum neutralizing antibodies were independently associated with isolation of infectious SARS-CoV-2 from the respiratory tract.

Study reliability

The patients are not completely described as well as their selection which appears to have been made on the basis of severity. Sampling was not carried out on the basis of a protocol.
Vero cells appear to be a good culture medium but may not be the best according to the authors.

Clearly defined setting Demographic characteristics described Follow-up length was sufficient Transmission outcomes assessed Main biases are taken into consideration
Yes No Unclear Yes No

What else should I consider?

The design and results are similar to those of Bullard and more studies of this type are required. A Ct threshold in Bullard of 25 relates roughly to the 7 Log10 RNA copies/mL 

A varied number of contextual and patient variables need to be assessed to achieve prediction of infectivity

About the authors

Carl Heneghan

Carl Heneghan

Carl is Professor of EBM & Director of CEBM at the University of Oxford. He is also a GP and tweets @carlheneghan. He has an active interest in discovering the truth behind health research findings

Elizabeth Spencer

Elizabeth Spencer

Dr Elizabeth Spencer; MMedSci, PhD. Epidemiologist, Nuffield Department for Primary Care Health Sciences, University of Oxford.

Tom Jefferson

Tom Jefferson

Tom Jefferson is a senior associate tutor and honorary research fellow, Centre for Evidence-Based Medicine, University of Oxford.