Detection of anti-SARS-CoV-2 salivary antibodies in vaccinated adults

Abstract

Since the introduction of efficient anti-SARS-CoV-2 vaccines, the detection of antibodies becomes useful for immunological monitoring and COVID-19 control. Therefore, this longitudinal study aimed to evaluate the detection of SARS-CoV-2 antibodies in the serum and saliva of COVID-19-vaccinated adults. The study included 13 not vaccinated and 35 vaccinated participants with two doses of CoronaVac (Sinovac/Butantan) vaccine who subsequently received BNT162b2 (Pfizer-BioNTech) vaccine as a booster dose. Vaccinated participants donated saliva and serum in three different time points. Enzyme-linked immunosorbent assay was used for antibody detection. In our results, the serum neutralizing antibodies (NAb) were detected in 34/35 samples after second dose and in 35/35 samples one and five months after the booster dose. In saliva, NAb were detected in 30/35 samples after second dose and in 35/35 of samples one and five months after the booster dose. IgA was detected in 19/34 saliva samples after second dose, in 18/35 one month after the booster and in 30/35 five months after. IgG in saliva was detected in 1/34 samples after second dose, 33/35 samples one month after the booster dose and in 20/35 five months after. A strong correlation was found between IgG and neutralizing activity in saliva, and salivary IgA would be a sign of recent exposure to the virus. In conclusion, saliva can be suitable for monitoring antibodies anti-SARS-CoV-2 after vaccination. Heterologous vaccination contributed to increase anti-SARS-CoV-2 antibodies in the Brazilian health context. Complementary studies with large groups are mandatory to conclude the interest in following mucosal immunity.

Keywords: COVID-19; COVID-19 vaccines; IgA; IgG; SARS-CoV-2; antibodies; neutralizing antibodies; saliva.

Figure 1

Flow diagram of participants and sample analyzes. IgA, Immunoglobulin A; IgG, Immunoglobulin G; NAb, Neutralizing Antibodies; TAb, Total Antibodies; n, number of participants.

Figure 2

Median concentration of neutralizing antibodies in serum and saliva, the dotted red line indicates the cutt-off to each test. (A) Comparation of neutralizing antibodies (NAb) median in serum of negative control group to vaccinated group in each time point (p<0.0001). Statistical analysis: Kruskal-Wallis test. (B) Comparation of neutralizing antibodies (Nab) median in saliva of negative control group to vaccinated group in each time point (p<0.0001). Statistical analysis: Kruskal-Wallis test. (C) Comparation of neutralizing antibodies (NAb) median in serum for each time point of vaccinated individuals (p<0.0001). Statistical analysis: Friedman test. (D) Comparation of neutralizing antibodies (Nab) median in saliva for each time point of vaccinated individuals (p<0.0001). Statistical analysis: Friedman test. Graphpad Prism, version 9.5.0 (California, USA). NAb, of neutralizing antibodies. Group 0 (G0): without vaccination; T1: 2 doses of CoronaVac; T2: 1 month after 2 doses of CoronaVac and one booster with Pfizer; T3: 5 months after 2 doses of CoronaVac and one booster with Pfizer *p<0.05; ***p<0.0005; ****p<0.0001.

Figure 3

Median of IgA and IgG antibodies detected in saliva by ELISA method (Ratio OD). (A) Comparation of IgA median of negative control group to vaccinated group in each time point (p=0.0013). Statistical analyses: Kruskal-Wallis test. (B) Comparation of IgA median for each time point for vaccinated individuals (p=0.0008). Statistical analyses: Friedman test. (C) Comparation of IgG median of negative control group to vaccinated group in each time point (p<0.0001). Statistical analysis: Kruskal-Wallis test. (D) Comparation of IgG median for each time point for vaccinated individuals (p<0.0001). Statistical analyses: Friedman test. Graphpad Prism, version 9.5.0 (California, USA). IgA, Immunoglobulin A; IgG, Immunoglobulin G; OD, Optical Density. Group 0 (G0): without vaccination; T1: 2 doses of CoronaVac; T2: 1 month after 2 doses of CoronaVac and one booster with Pfizer; T3: 5 months after 2 doses of CoronaVac and one booster with Pfizer *p<0.05; **p<0.005; ***p<0.0005; ****p<0.0001.

Figure 4

Significant correlations between different types of antibodies in saliva and IgA titers of infected participants. (A) Correlation IgG vs NAb in saliva for all the samples, there is a Strong correlation (n=117 pairs, r=0.72, p<0.0001). (B) Correlation IgA vs NAb in saliva for all the samples, the is no significant correlation (n=117 pairs, r=0.048, p=0.61). (C) Compartion of IgA titers of infected participants in T3 before and after infection (n=4), paired t test for parametric data (p=0.011*); (D) Individual evolution of salivary IgA in infected participants (n=13), purple lines are non-recent infected participants and lines in blue are the recent infected participants. Statistical analysis: For non-parametric data Spearman’s correlation. Graphpad Prism, version 9.5.0 (California, USA). Ab, antibodies; IgG, immunoglobin G; IgA, immunoglobin A; NAb, neutralizing antibodies; I, infected; NI, not infected; r, correlation coefficient. T2: 1 month after 2 doses of CoronaVac and one booster with Pfizer; T3: 5 months after 2 doses of CoronaVac and one booster with Pfizer *p<0.05.