Virological and immunological features of SARS-COV-2 infected children with distinct symptomatology

Abstract

Background: Although SARS-CoV-2 immunizations have started in most countries, children are not currently included in the vaccination programs; thus, it remains crucial to define their anti-SARS-CoV-2 immune response in order to minimize the risk for other epidemic waves. This study sought to provide a description of the virology ad anti-SARS-CoV-2 immunity in children with distinct symptomatology.

Methods: Between March and July 2020, we recruited 15 SARS-CoV-2 asymptomatic (AS) and 51 symptomatic (SY) children, stratified according to WHO clinical classification. We measured SARS-CoV-2 viral load using ddPCR and qPCR in longitudinally collected nasopharyngeal swab samples. To define anti-SARS-CoV-2 antibodies, we measured neutralization activity and total IgG load (DiaSorin). We also evaluated antigen-specific B and CD8+T cells, using a labeled S1+S2 protein and ICAM expression, respectively. Plasma protein profiling was performed with Olink.

Results: Virological profiling showed that AS patients had lower viral load at diagnosis (p = .004) and faster virus clearance (p = .0002) compared with SY patients. Anti-SARS-CoV-2 humoral and cellular response did not appear to be associated with the presence of symptoms. AS and SY patients showed similar titers of SARS-CoV-2 IgG, levels of neutralizing activity, and frequency of Ag-specific B and CD8+ T cells, whereas pro-inflammatory plasma protein profile was found to be associated with symptomatology.

Conclusion: We demonstrated the development of anti-SARS-CoV-2 humoral and cellular response with any regard to symptomatology, suggesting the ability of both SY and AS patients to contribute toward herd immunity. The virological profiling of AS patients suggested that they have lower virus load associated with faster virus clearance.

Keywords: Ag-specific cellular response; SARS-CoV-2; asymptomatic patients; neutralization humoral activity; symptomatic patients.

FIGURE 1

Virological analysis of SARS‐CoV‐2–infected children. (A) Log₂ ddPCR in NP swabs (copies/5 µl) in AS vs SY patients. (B) ET of RdRp gene and (C) NP virus clearance (days) in AS vs SY patients. (D) Association between infectivity (measured as FFU/ml) and ddPCR in NP swabs. (E) AUC for RdRp gene in AS vs SY patients. (F) AUC for N gene in AS vs SY patients. The Mann‐Whitney test was used to define differences in (A–C); Spearman's test was used in (D), AUC was calculated as described in Methods. p values <.05 were considered significant

FIGURE 2

SARS‐CoV‐2 seroconversion and Ag‐specific B and CD8 T cells in children with distinct symptomatology. (A) Left‐hand side represents SARS‐CoV‐2 Ab titers at admission (upper dot plots) and at “post‐acute phase” (lower dot plots). Right‐hand side represents SARS‐CoV‐2 PRNT at admission (upper dot plots) and at “post‐acute phase” (lower dot plots). Neutralization values were identified by dilution factor. Contingency plots show frequency of seronegative, seropositive, and equivocal results in both phases, following the same pattern described for the dot plots. Gating strategy (B) and frequencies of Ag‐specific CD19+IgD‐CD27+ switched B cells (C) are shown in AS and SY patients. Gating strategy (D) and frequencies of Ag‐specific ICAM+CD8+ T cells (E) are shown in AS and SY patients. Box plot in (F) shows results from Boolean gating of intracellular staining analysis from ICAM+CD8 T cells. The Mann‐Whitney test was used to define differences, p values <.05 were considered significant

FIGURE 3

Proteomic profile in AS vs SY patients. PCA in (A) shows distribution according to protein profile between AS and SY patients. (B) PC loading for PC4 is showed in the box plot. The red line indicates the expected values. Violin plots in (C) show differentially expressed proteins among the groups