SARS-CoV-2 Viral RNA Shedding for More Than 87 Days in an Individual With an Impaired CD8+ T Cell Response

Abstract

Prolonged shedding of viral RNA occurs in some individuals following SARS-CoV-2 infection. We perform comprehensive immunologic evaluation of one individual with prolonged shedding. The case subject recovered from severe COVID-19 and tested positive for SARS-CoV-2 viral RNA repeatedly as many as 87 days after the first positive test, 97 days after symptom onset. The subject did not have any associated rise in anti-Spike protein antibody titers or plasma neutralization activity, arguing against re-infection. This index subject exhibited a profoundly diminished circulating CD8+ T cell population and correspondingly low SARS-CoV-2-specific CD8+ T cell responses when compared with a cohort of other recovering COVID-19 subjects. CD4+ T cell responses and neutralizing antibody responses developed as expected in this individual. Our results demonstrate that detectable viral RNA shedding in the upper airway can occur more than 3 months following infection in some individuals with COVID-19 and suggest that impaired CD8+ T cells may play a role in prolonged viral RNA shedding.

Keywords: CD4+ T cell; CD8+ T cell; COVID-19; SARS-CoV-2; cellular immunity.

Conflict of interest statement

Author BP is employed by the company IncellDX. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Copyright © 2021 Turner, Day, Alsoussi, Liu, O’Halloran, Presti, Patterson, Whelan, Ellebedy and Mudd.

Figures

Figure 1

Timeline of study sample collection, administered treatments and clinical SARS-CoV-2 PCR test results for the case subject.

Figure 2

Measurement of cell populations, SARS-CoV-2 viral replication and cytokines over time. (A) Frequency of circulating T cell and B cell populations in the PBMC from WU 350-013 at various time-points post-study enrollment. Inset: in a separate experiment, the frequency of circulating T cell populations is reported in a cohort of recovering COVID-19 subjects, including a sample from WU 350-013 (denoted in red) at day 78 post-study enrollment. (B) WU 350-013 plasma viral load over time. Inset: Plasma viral load quantified in WU 350-013 (red) and a cohort of mechanically ventilated ICU subjects at day 0 and day 14 post-study enrollment. Dashed line represents the limit of detection of the assay. (C) Cycle threshold or Ct values for clinical nasopharyngeal SARS-CoV-2 testing performed on WU 350-013 throughout the course of the study. Lower Ct values represent a higher burden of viral replication. (D) Concentration of select cytokines in plasma from WU 350-013 over time.

Figure 3

Case subject adaptive immune response over time. (A) WU 350-013 SARS-CoV-2-specific CD8+ T cell response to overlapping peptide pools from N, M and S viral proteins as measured by intracellular cytokine staining (ICS) assay at various study time-points. Dotted line is at stimulation index of 1. Inset: CD8+ T cell ICS response to peptide pools from the cohort of recovered COVID-19 subjects, including the day 78 post-study enrollment sample from WU 350-013 (red). (B) WU 350-013 SARS-CoV-2-specific CD4+ T cell response to overlapping peptide pools from N, M and S viral proteins as measured by ICS at various study time-points. Dotted line is at stimulation index of 1. Inset: CD4+ T cell ICS response to peptide pools from recovered COVID-19 subjects, including the day 78 post-study enrollment sample from WU 350-013 (red). (C) Development of plasma spike protein receptor binding domain-binding antibodies (left axis, closed symbols) and pseudotyped VSV (expressing SARS-CoV-2 spike protein) neutralizing antibodies in plasma (right axis, open symbols) over time. Light dotted line represents ELISA area under the curve of SARS-CoV-2 negative control plasma; heavy dotted line represents plasma neutralization limit of detection.