Clinical, immunological and virological characterization of COVID-19 patients that test re-positive for SARS-CoV-2 by RT-PCR

Abstract

Background: Some COVID-19 cases test positive again for SARS-CoV-2 RNA following negative test results and discharge, raising questions about the meaning of virus detection. Better characterization of re-positive cases is urgently needed.

Methods: Clinical data were obtained through Guangdong's COVID-19 surveillance network. Neutralization antibody titre was determined using microneutralization assays. Potential infectivity of clinical samples was evaluated by cell inoculation. SARS-CoV-2 RNA was detected using three different RT-PCR kits and multiplex PCR with nanopore sequencing.

Findings: Among 619 discharged COVID-19 cases, 87 re-tested as SARS-CoV-2 positive in circumstances of social isolation. All re-positive cases had mild or moderate symptoms at initial diagnosis and were younger on average (median, 28). Re-positive cases (n = 59) exhibited similar neutralization antibodies (NAbs) titre distributions to other COVID-19 cases (n = 218) tested here. No infectious strain could be obtained by culture and no full-length viral genomes could be sequenced from re-positive cases.

Interpretation: Re-positive SARS-CoV-2 cases do not appear to be caused by active reinfection and were identified in ~14% of discharged cases. A robust NAb response and potential virus genome degradation were detected in almost all re-positive cases, suggesting a substantially lower transmission risk, especially through respiratory routes.

Keywords: COVID-19; Neutralizing antibody; Re-positive; SARS-CoV-2; Virology.

Fig. 1

(a) Summary of sampling scheme in this study; (b) Comparison of Nab titres among infections that were re-positive, general discharged COVID-19 cases, and cases in hospitalization. General discharged cases refer to COVID-19 recovered cases detected as SARS-CoV-2 negative in 14 days after discharge. To plot these results into one figure, we recorded 2 for antibody titre less than 4 (highlighted in red), and 2048 for titre larger than 1024. The start point of serum dilution (titre of 4) was highlighted with red dash line. (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)

Fig. 2

Timeline of 32 COVID-19 re-positive cases sampled and tested between 28 February–1 March.

Fig. 3

(a) Virus genome sequences (for those samples with genome coverage >10%) obtained from re-positive cases. Single nucleotide polymorphisms (with respect to the reference genome MN908947.3) are coloured in red. Genome sequence fragments are coloured blue, orange to indicate whether they were obtained from anal, nasopharyngeal and throat swabs, respectively. The colored bars at the top indicate the targeting regions for three PCR kits. The corresponding Ct values from three different RT-PCR kits were shown on the right side. (b) Coverage of the consensus sequence among nasopharyngeal swabs, throat swabs and anal swabs. A solid circle refers a RT-PCR positive sample, and a hollow circle refers a RT-PCR negative sample. The red dash line refers to the sequencing coverage of 10%; (c) Coverage of consensus sequence measured from respiratory tract (nasopharyngeal swabs/throat swabs) and matched digestive tract (anal swabs). (For interpretation of the references to color in this figure legend, the reader is referred to the web version of this article.)