Discrepant serological findings in SARS-CoV-2 PCR-negative hospitalized patients with fever and acute respiratory symptoms during the pandemic

Abstract

Coronavirus Disease 2019 (COVID-19) serology has an evolving role in the diagnosis of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection. However, its use in hospitalized patients with acute respiratory symptoms remains unclear. Hospitalized patients with acute respiratory illness admitted to an isolation ward were recruited. All patients had negative nasopharyngeal swab polymerase chain reaction (PCR) for SARS-CoV-2. Serological studies using four separate assays (cPass: surrogate neutralizing enzyme-linked immunosorbent assay [ELISA]; Elecsys: N-antigen based chemiluminescent assay; SFB: S protein flow-based; epitope peptide-based ELISA) were performed on stored plasma collected from patients during the initial hospital stay, and a convalescent visit 4-12 weeks later. Of the 51 patients studied (aged 54, interquartile range 21-84; 62.7% male), no patients tested positive on the Elecsys or cPass assays. Out of 51 patients, 5 had antibodies detected on B-cell Epitope Assay and 3/51 had antibodies detected on SFB assay. These 8 patients with positive serological test to COVID-19 were more likely to have a high-risk occupation (p = 0.039), bacterial infection (p = 0.028), and neutrophilia (p = 0.013) during their initial hospital admission. Discrepant COVID-19 serological findings were observed among those with recent hospital admissions and bacterial infections. The positive serological findings within our cohort raise important questions about the interpretation of sero-epidemiology during the current pandemic.

Keywords: COVID-19; Singapore; serology.

Figure 1

Serological analysis by S protein flow‐based (SFB) and B‐cell epitope assays. Sera from symptomatic patients (n = 51), isolated for evaluation of COVID‐19, were collected at acute and convalescent (between 3 and 12 weeks later) timepoints. Serum samples were screened at 1:100 dilution (A) in an SFB assay for specific total IgG, IgG1, and IgG3 against full‐length SARS‐CoV‐2 S protein expressed on the surface of HEK293T cells, and (B) in a peptide‐based enzyme‐linked immunosorbent assay (ELISA) against four IgG linear B‐cell epitopes of SARS‐CoV‐2: spike S14P5, S20P2 and S21P2, and nucleocapsid N4P5. Sera or plasma samples from healthy donors (n = 22 for SFB; n = 10 for epitope assay), recovered SARS patients (n = 20 for SFB; n = 10 for epitope assay), and COVID‐19 patients (n = 15; median 23 days postillness onset) were included as controls. Data are shown as mean ± SD of two independent experiments, with dotted lines indicating mean + 3 SD of healthy donors. An isotype response was defined as positive by SFB assay when the binding is more than mean + 3 SD of the healthy controls