Streptococcus pneumoniae colonization associates with impaired adaptive immune responses against SARS-CoV-2

Abstract

BackgroundAlthough recent epidemiological data suggest that pneumococci may contribute to the risk of SARS-CoV-2 disease, cases of coinfection with Streptococcus pneumoniae in patients with coronavirus disease 2019 (COVID-19) during hospitalization have been reported infrequently. This apparent contradiction may be explained by interactions of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and pneumococci in the upper airway, resulting in the escape of SARS-CoV-2 from protective host immune responses.MethodsHere, we investigated the relationship of these 2 respiratory pathogens in 2 distinct cohorts of health care workers with asymptomatic or mildly symptomatic SARS-CoV-2 infection identified by systematic screening and patients with moderate to severe disease who presented to the hospital. We assessed the effect of coinfection on host antibody, cellular, and inflammatory responses to the virus.ResultsIn both cohorts, pneumococcal colonization was associated with diminished antiviral immune responses, which primarily affected mucosal IgA levels among individuals with mild or asymptomatic infection and cellular memory responses in infected patients.ConclusionOur findings suggest that S. pneumoniae impair host immunity to SARS-CoV-2 and raise the question of whether pneumococcal carriage also enables immune escape of other respiratory viruses and facilitates reinfection.Trial registrationISRCTN89159899 (FASTER study) and ClinicalTrials.gov NCT03502291 (LAIV study).

Keywords: Adaptive immunity; Bacterial infections; Immunology; T cells; Virology.

Figure 1. Prevalence of pneumococcal colonization among SARS-CoV-2–positive and –negative HCWs and patients.

(A) Experimental design of the study with the sample type, sample collection schedule, and measurable-per-sample type depicted for both the HCW and patient cohorts. In the patient cohort, day 2 and day 7 samples were collected only for individuals who were hospitalized. (B and C) Doughnut charts showing the percentage of pneumococcal prevalence in (B) HCWs (n = 85) and (C) patients (n = 400) infected or noninfected with SARS-CoV-2. Fisher’s exact test was used to compare percentages. (D) Percentage of the pneumococcal colonization rate detected in the patient cohort during calendar periods with different circulation restrictions: 5% (6 of 119) from April to June; 17.3% (13 of 75) from July to September; 8.5% (13 of 154) from October to December; and 5.8% (3 of 52) in January. (E) Viral load levels expressed as RNA copies/mL, as detected by genesig RT-qPCR in noncolonized (n = 19, light blue) and Spn-colonized (n = 10, yellow) HCWs and noncolonized (n = 73, lilac) and Spn-colonized (n = 19, green) patients. *P < 0.05, ***P < 0.001, and ****P < 0.0001, by Kruskal-Wallis test for comparisons between groups.

Figure 2. Mucosal and systemic antibody responses to SARS-CoV-2 in HCWs and patients.

(A) Salivary IgA titers against SARS-CoV-2 RBD, S1, S2, and N proteins in HCWs, divided into noncolonized (n = 12) and Spn-colonized (n = 9) groups and in unexposed, healthy controls (n = 15). (B) Nasal IgA titers against SARS-CoV-2 RBD, S1, S2, and N proteins in patients, divided into noncolonized (n = 23) and Spn-colonized (n = 15) groups and in unexposed, healthy controls (collected before 2019, n = 12). (C) Serum IgG titers in HCWs (n = 16 noncolonized and n = 10 Spn-colonized), patients (n = 24 noncolonized and n = 14 Spn-colonized), and unexposed, healthy controls (n = 15). Both mucosal and serum antibody titers from SARS-CoV-2–positive participants were measured during the convalescent phase of the viral infection. Antibody levels are expressed as AU. Medians with IQRs are shown for antiviral responses. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by Kruskal-Wallis test for comparisons between groups.

Figure 3. SARS-CoV-2–specific memory B cells in HCWs and patients.

Percentage of (A) S1- and (B) S2-specific memory B cells within CD19⁺CD27⁺ memory B cells in HCWs (n = 12 noncolonized and n = 9 Spn-colonized), recovered patients (n = 23 noncolonized and n = 12 Spn-colonized), and healthy controls (n = 18). Medians with IQRs are shown, and each dot represents an individual. *P < 0.05, **P < 0.01, and ****P < 0.0001, by Kruskal-Wallis test.

Figure 4. SARS-CoV-2–specific T cell responses in HCWs and patients.

Percentage of (A) cytokine-producing (IFN-γ, TNF-α, IL-2) CD4⁺ and (B) CD8⁺ T cells after ex vivo PBMC stimulation with N, S1, and S peptide pools in SARS-CoV-2–positive HCWs (n = 17 noncolonized and n = 8 Spn-colonized), recovered patients (n = 17 colonized and n = 14 Spn-colonized), and healthy controls (n = 16). One peptide pool was used per condition. SEB was used as a positive control and DMSO as the negative control (nonstimulated cell condition – mock). Background (mock) was subtracted from the peptide-stimulated conditions to remove nonspecific signals. Data indicate positivity for any of the 3 measured cytokines. Medians with IQRs are shown, and each dot represents an individual. *P < 0.05, **P < 0.01, ***P < 0.001, and ****P < 0.0001, by Kruskal-Wallis test.

Figure 5. Cytokine concentrations in nasal lining fluid and serum.

(A) Heatmaps showing the median log₂ fold change (FC) in the levels of 30 cytokines in nasal lining fluid and serum of noncolonized and Spn-colonized HCWs and patients during the acute phase of SARS-CoV-2 infection versus unexposed healthy controls. Upregulation (red) and downregulation (blue) are shown compared with cytokine levels in the control group. Cytokines were clustered in active cytokine families. (B) Volcano plots showing the median log₂ fold change versus healthy controls (n = 17) per cytokine in nasal lining fluid and serum of noncolonized HCWs (n = 17), Spn-colonized HCWs (n = 9), noncolonized patients (n = 70), and Spn-colonized patients (n = 19). The horizontal dotted line represents the cutoff of significance (adjusted P = 0.05, after FDR correction of the P value), whereas the vertical dotted line represents a cutoff point for determining whether the levels of cytokines were higher (right, red) or lower (left, blue) compared with those of the healthy control group. Statistical comparisons were applied between each study group and the healthy control group using the Mann-Whitney U test, followed by Benjamini-Hochberg correction for multiple testing. Nonsignif, nonsignificant; Down, downregulation; Up, upregulation.

Figure 6. Nasal and serum inflammatory profiles of SARS-CoV-2 infection and coinfection with Spn in HCWs and patients.

PCA of 30 cytokines in (A) nasal lining fluid and (B) serum of healthy controls (gray), noncolonized HCWs (light blue), Spn-colonized HCWs (yellow), noncolonized patients (lilac), and Spn-colonized patients (green). PC1, principal component 1; PC2, principal component 2.