SARS-CoV-2 Infection-and mRNA Vaccine-induced Humoral Immunity among Schoolchildren in Hawassa, Ethiopia

Abstract

Background: With the persisting low vaccination intake, particularly in children of low-and middle-income countries (LMICs), seroepidemiological studies are urgently needed to guide and tailor COVID-19 pandemic response efforts in schools and to put mitigation strategies in place for a future post-pandemic resurgence. However, there is limited data on SARS-CoV-2 infection-induced and vaccine-induced humoral immunity in schoolchildren in LMICs, including Ethiopia.

Methods: As the spike receptor binding domain (RBD) is the major target for neutralization antibodies and useful to predict the correlates of protection, we used an in-house anti-RBD IgG ELISA to assess and compare infection-induced antibody response at two-time points and BNT162b2 (BNT) vaccine-induced antibody response at a one-time point in schoolchildren in Hawassa, Ethiopia. In addition, we measured and compared the levels of binding IgA antibodies to spike RBD of SARS-CoV-2 Wild type, Delta, and Omicron variants in a small subset of unvaccinated and BNT-vaccinated schoolchildren.

Results: When we compare SARS-CoV-2 infection-induced seroprevalences among unvaccinated school children (7-19 years) at the two blood sampling points with a 5-month interval, we observed an over 10% increase, from 51.8% (219/419) in the first week of December 2021 (post-Delta wave) to 67.4% (60/89) by the end of May 2022 (post-Omicron wave). Additionally, we found a significant correlation (p = 0.001) between anti-RBD IgG seropositivity and a history of having COVID-19-like symptoms. Compared to the levels of SARS-CoV-2 infection-induced anti-RBD IgG antibodies before vaccination, higher levels of BNT vaccine-induced anti-RBD IgG antibodies were observed even in SARS-CoV-2 infection-naïve schoolchildren of all age groups (p = 0.0001). Importantly, one dose of the BNT vaccine was shown to be adequate to elicit a strong antibody response in schoolchildren with pre-existing anti-RBD IgG antibodies comparable to that of SARS-CoV-2 infection-naive schoolchildren receiving two doses of BNT vaccine, suggesting a single dose administration of the BNT vaccine could be considered for schoolchildren who had prior SARS-CoV-2 infection when a shortage of vaccine supply is a limiting factor to administer two doses irrespective of their serostatus. Despite the small sample size of study participants, the BNT vaccine is shown to be immunogenic and safe for schoolchildren. Irrespective of schoolchildren's vaccination status, we observed a similar pattern of significantly higher levels of IgA antibodies to Delta-RBD than to Omicron-RBD (p < 0.001) in a randomly selected subset of schoolchildren, yet comparable to Wuhan-RBD, suggesting these schoolchildren were more likely to have had SARS-CoV-2 infection with Delta variant. Additionally, we noted a broader IgA antibody reactivity to SARS-CoV-2 variants in vaccinated schoolchildren with prior SARS-CoV-2 infection, supporting the superiority of hybrid immunity.

Conclusion: Our serological data indicate a significant increase in SARS-CoV-2 seroprevalence in children at a post-Omicron five-month follow-up compared to a post-Delta enrolment. Despite the small sample size of study participants, the BNT vaccine is shown to be immunogenic and safe for schoolchildren. Hybrid immunity would likely provide a broader humoral immunity against Wuhan strain, Delta, and Omicron variants than natural infection or vaccination alone does. However, future longitudinal cohort studies in SARS-CoV-2-naïve and COVID-19-recovered schoolchildren receiving the BNT vaccine are needed for a better understanding of the kinetics, breadth, and durability of BNT vaccine-induced multivariant-cross reactive immunity.

Keywords: BNT162b2 vaccine; COVID-19; RBD; SARS-CoV-2; antibody; dose; schoolchildren; variant.

Figure 1

Study design, time points for the administration of first dose (1d) and second dose (2d) vaccination, and saliva and blood samples collected with the corresponding number (n) of participants. The two time points at a 5-month interval selected for serum and saliva sample collection are indicated on the vertical time axis with blood collection tubes.

Figure 2

Comparison of the natural infection- and BNT vaccine-induced humoral immunity response in vaccinated and unvaccinated schoolchildren with and without baseline pre-existing anti-RBD IgG antibodies against Wuhan-RBD, A-F. Levels of anti-RBD IgG comparison between (A) schoolchildren (n = 80) who received one dose versus those schoolchildren (n = 21) who received two-doses of the BNT vaccine; (B) schoolchildren who received one dose of the BNT vaccine with and without a baseline pre-existing anti-RBD IgG antibodies; (C): schoolchildren who received two doses of the BNT vaccine with and without a baseline pre-existing anti-RBD IgG antibodies; (D): schoolchildren (n = 26) who received one dose of BNT vaccine with a baseline pre-existing anti-RBD IgG antibodies versus SARS-CoV-2-infection-naïve schoolchildren (n = 12) who received two doses; (E): schoolchildren (n = 38) who received one dose of the BNT vaccine with baseline pre-existing anti-RBD IgG and schoolchildren (n = 12) who received two doses of the BNT vaccine without a baseline pre-existing anti-RBD IgG antibodies: (F): unvaccinated schoolchildren with and without anti-RBD IgG antibodies at baseline or 5-month follow-up versus schoolchildren who received one or two doses of the BNT vaccine with or without a baseline pre-existing anti-RBD IgG. The levels of serum anti-RBD IgG antibodies were measured using serum samples diluted at 1:200 in an in-house indirect ELISA. The OD₄₅₀ S/C ratio value on the y-axis represents the ratio of the sample OD₄₅₀ nm to the average mean OD₄₅₀ nm of the negative controls. The horizontal broken red line shows the cut-off value (= 2.5) for the in-house ELISA test. A Wilcoxon-Mann-Whitney (WMW) matched or unmatched pairs test was performed to compare differences between the two groups. A p-value of < 0.05 indicates the occurrence of a statistically significant difference in anti-RBD IgG levels between any two comparison groups while ns or p > 0.05 indicates a statistically insignificant difference. 1d, 2d, and NonVacc represent schoolchildren who received a single dose, two doses of the BNT vaccine, and neither a first dose nor a second dose, respectively; BL, PV, and 5-month indicate that serum samples were collected before schoolchildren’s vaccination (baseline), postvaccination (after receiving one or two doses of the BNT vaccine), and at the 5-month follow-up visit from unvaccinated children, respectively; Na and Pi indicate being SARS-CoV-2 infection naïve (negative for anti-RBD IgG serological test at baseline) and having a prior SARS-CoV-2 infection (positive for anti-RBD IgG serological test at the baseline), respectively.

Figure 3

Comparison of the levels of infection-induced and BNT vaccine-induced anti-RBD IgG responses by the self-reported history of COVID-19-defining symptoms, sex, and age. The levels of anti-RBD IgG in (A): vaccinated schoolchildren with COVID-19-defining symptoms versus vaccinated schoolchildren without COVID-19-defining symptoms; (B): unvaccinated schoolchildren with COVID-19-defining symptoms versus unvaccinated schoolchildren without COVID-19-defining symptoms; (C): male vaccinated schoolchildren versus female vaccinated schoolchildren; (D): male unvaccinated versus female unvaccinated schoolchildren; (E): schoolchildren of age < 13 years at prevaccination versus schoolchildren of age ≥ 13 years at prevaccination; (F): schoolchildren of age < 13 years at postvaccination versus schoolchildren of age ≥ 13 years at postvaccination; (G). unvaccinated schoolchildren of age < 13 years at baseline versus unvaccinated schoolchildren of age ≥ 13 years at baseline; (H): unvaccinated schoolchildren of age < 13 years at 5-month follow-up versus unvaccinated schoolchildren of age ≥ 13 years at 5-month follow-up. A Wilcoxon-Mann-Whitney (WMW) matched or unmatched pairs test was performed to compare differences between the two groups. For all panels A-H, the p-value> 0.05. ns = p > 0.05 indicates a statistically insignificant difference. The OD₄₅₀ S/C ratio value on the y-axis represents the ratio of the sample OD₄₅₀ nm to the average mean OD₄₅₀ nm of the negative controls. The broken black line represents the cut-off value (2.5).

Figure 4

Comparison of the levels of binding IgA antibodies to the RBD SARS-CoV-2 of Wuhan strain, Delta (AY.2 lineage) variant, and Omicron (B.1.1.529 BA.2 lineage) variant. (A) SARS-CoV-2-infection-naïve unvaccinated schoolchildren (n = 13); (B) unvaccinated schoolchildren with prior (baseline) SARS-CoV-2 infection (n = 8); (C) SARS-CoV-2-infection-naïve BNT-vaccinated schoolchildren (n = 11) and (D) BNT-vaccinated schoolchildren with prior (baseline) SARS-CoV-2 infection (n = 6). The antibody levels were determined in serum samples collected at the 5-month follow-up using an in-house indirect ELISA. The OD₄₅₀ value on the y-axis was calculated as the ratio of the optical density (OD) of the sample to the mean ODs of negative controls. An antibody level (OD₄₅₀ nm S/C ratio) below 2.5 was interpreted as negative. A Wilcoxon-Mann-Whitney (WMW) matched or unmatched pairs test was performed to compare the difference between the two groups. p-value < 0.05 indicates the presence of a statistically significant difference while p > 0.05 indicates a statistically insignificant difference. The broken line represents the cut-off value, of 2.5.