Longitudinal Assessment of Nasopharyngeal Biomarkers Post-COVID-19: Unveiling Persistent Markers and Severity Correlations

Abstract

SARS-CoV-19 infection provokes a variety of symptoms; most patients present mild/moderate symptoms, whereas a small proportion of patients progress to severe illness with multiorgan failure accompanied by metabolic disturbances requiring ICU-level care. Given the importance of the disease, researchers focused on identifying severity-associated biomarkers in infected patients as well as markers associated with patients suffering long-COVID. However, little is known about the presence of biomarkers that remain a few years after SARS-CoV-2 infection once the patients fully recover of the symptoms. In this study, we evaluated the presence of persistent biomarkers in the nasopharyngeal tract two years after SARS-Cov-2 infection in fully asymptomatic patients, taking into account the severity of their infection (mild/moderate and severe infections). In addition to the direct identification of several components of the Coronavirus Infection Pathway in those individuals that suffered severe infections, we describe herein 371 proteins and their associated canonical pathways that define the different adverse effects of SARS-CoV-2 infections. The persistence of these biomarkers for up to two years after infection, along with their ability to distinguish the severity of the infection endured, highlights the surprising presence of persistent nasopharyngeal exudate changes in fully recovered patients.

Keywords: COVID-19; SARS-CoV-19; biomarkers; proteomics.

Figure 1

Summary of experiments and proteomic conditions. (a) graphical summary. (b) PCA of the data set of the three experimental conditions. (c) Volcano plots of the comparisons between the three conditions. For biomarker discovery, only proteins are identified in all groups (threshold, q value < 0.05). (d) Heatmap.

Figure 2

Proteins that were significantly different between the three experimental conditions (severe, mild, and non-COVID samples). Violin plots of the intensity of the different proteins (peptides) within the different conditions. Horizontal black and dash lines indicate the median and interquartile range of the protein abundance. (a) Proteins that appear increased with the severity of the infection. (b) Unique protein that was found to decrease expression with the severity of COVID-19. Proteins that increase (c) or decrease (d) in the mild group.

Figure 3

Biomarkers of SARS-CoV-2 severe infections. (a) Representative violin plots of 4 out of 280 proteins that were found overexpressed in severe conditions. (b) Representative violin plots of 4 out of 26 proteins that were down-regulated in the severe group. (c,d) Bioinformatics of the results allowed for the identification of several canonical pathways that were predicted to be increased (orange; positive z-score) or repressed (blue; negative z-score), −log (P value) > 4.5. Fisheŕs exact test p-value. (e) When infectious diseases were investigated using Reactome, SARS-CoV-2 modulation of the immune response was identified. Statistical data can be found in Table S2.

Figure 4

Coronavirus infection pathway (IPA). Proteins up or down-regulated in the severe group and non-COVID that participate in the coronavirus infection pathway. Legends are embedded in the figure. Gene and protein names are included in the pathway.

Figure 5

Violin plots of 4 representative proteins found overexpressed (a) or reduced (b) in mild and severe SARS-CoV-2 infections vs the non-COVID group. (c,d) Canonical pathways identified using IPA analysis are shown. Positive z-score in orange and negative in blue. Statistical data can be found in Table S2.