SARS-CoV-2 RNA Test Assessed Among Recovered COVID-19 Patients

Some patients who have recovered from coronavirus disease 2019 (COVID-19) with documented negative real-time polymerase chain reaction (RT-PCR) results at the time of recovery have had subsequent positive RT-PCR test results for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2).

In the absence of any symptoms suggestive of new SARS-CoV-2 infection and it is unknown whether such patients are infectious and whether they should be quarantined. Real-time PCR is not a viral culture and does not allow determination of whether the virus is viable and transmissible.

A team of medical scientists associated with the Università Cattolica del Sacro Cuore (Rome, Italy) studied 176 recovered patients with COVID-19 who were admitted to the post-acute outpatient service of their institution from April 21 to June 18, 2020, for COVID-19 follow-up. Before that, patients had discontinued isolation according to current criteria which require no fever for three consecutive days, improvement in other symptoms, and two negative RT-PCR results for SARS-CoV-2 RNA 24 hours apart.

Nasal/oropharyngeal swab (NOS) samples from patients at follow-up were analyzed for total (genomic) and replicative (subgenomic) SARS-CoV-2 RNA using RT-PCR assays. Total SARS-CoV-2 RNA Detection and Quantification Total viral RNA was extracted from NOS samples using the Nimbus automated system (Seegene, Seoul, South Korea) which uses STARMag Universal Cartridge kit for both RNA extraction and PCR assay setup.

The RT-PCR testing was performed using the Seegene Allplex 2019-nCoV assay, a single-tube assay targeting three viral genes (E, RdRP, and N). Total viral RNA quantification was performed using the Quanty COVID-19 assay (Clonit S.r.l, Milan, Italy). Serum samples were tested with a commercial assay (Euroimmun; Lübeck, Germany), an enzyme-linked immunosorbent assay (ELISA) that uses the recombinant S1 domain of the SARS-CoV-2 spike (S) protein as antigen, for semi-quantitative detection of anti–SARS-CoV-2 IgG and IgA antibodies.

The investigators reported that 32 of 176 NOS samples (18.2%) tested positive for total SARS-CoV-2 RNA, with viral loads ranging from 1.6 × 10¹ to 1.3 × 10⁴ SARS-CoV-2 RNA copies per mL. One of the 32 samples (3.1%) had replicative SARS-CoV-2 RNA. Samples from the 32 patients at the time of COVID-19 diagnosis were also tested and, expectedly, had replicative SARS-CoV-2 RNA. All but 1 of 32 patients had a positive serology result against SARS-CoV-2, as well as 139 of remaining 144 patients, at COVID-19 follow-up. The patient who tested serologically negative was not the one with a positive test result for replicative SARS-CoV-2 RNA. The mean ±SD time from COVID-19 diagnosis to follow-up was 48.6 ± 13.1 days in 32 patients and 57.7 ±16.9 days in 144 patients.

The authors concluded that their study highlights that many patients who recovered from COVID-19 may be still positive (albeit at lower levels) for SARS-CoV-2 RNA, but only a minority of the patients may carry a replicating SARS-CoV-2 in the respiratory tract. The study was published on November 12, 2020 in the journal JAMA Internal Medicine.

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