Neutralizing antibody responses to SARS-CoV-2 variants in vaccinated Ontario long-term care home residents and workers

Prioritizing Ontarios long-term care home (LTCH) residents for vaccination against severe acute respiratory syndrome coronavirus 2 has drastically reduced their disease burden; however, recent LTCH outbreaks of variants of concern (VOCs) have raised questions regarding their immune responses. In 198 residents, mRNA vaccine dose 1 elicited partial spike and receptor binding domain antibody responses, while the second elicited a response at least equivalent to convalescent individuals in most residents. Residents administered mRNA-1273 (Moderna) mounted stronger total and neutralizing antibody responses than those administered BNT162b2 (Pfizer-BioNTech). Two to four weeks after dose 2, residents (n = 119, median age 88) produced 4.8-6.3-fold fewer neutralizing antibodies than staff (n = 78; median age 47) against wild-type (with D614G) pseudotyped lentivirus, and residents administered BNT162b2 produced 3.89-fold fewer neutralizing antibodies than those who received mRNA-1273. These effects were exacerbated upon serum challenge with pseudotyped VOC spike, with up to 7.94-fold reductions in B.1.351 (Beta) neutralization. Cumulatively, weaker vaccine stimulation, age/comorbidities, and the VOC produced an [~]130-fold reduction in apparent neutralization titers in LTCH residents and 37.9% of BNT162b2-vaccinated residents had undetectable neutralizing antibodies to B.1.351. Continued immune response surveillance and additional vaccine doses may be required in this population with known vulnerabilities.

A mucosal antibody response is induced by intra-muscular SARS-CoV-2 mRNA vaccination

Although SARS-CoV-2 infects the upper respiratory tract, we know little about the amount, type, and kinetics of antibodies (Ab) generated at this site in response to intramuscular COVID-19 vaccination, and whether these Ab protect against subsequent "breakthrough" infections. We collected longitudinal serum and saliva samples from participants receiving two doses of mRNA COVID-19 vaccines over a 6-month period and measured the relative level of anti-Spike and anti-Receptor Binding Domain (RBD) Ab. We detected anti-Spike/RBD IgG and IgA and associated secretory component in the saliva of most participants receiving 1 dose of mRNA vaccine. Administration of a second dose of mRNA boosted the IgG but not the IgA response, with only 30% of participants remaining positive for IgA at this timepoint. At 6 months post-dose 2, these participants exhibited greatly diminished anti-Spike/RBD IgG and IgA levels concomitant with a reduction in neutralizing activity in the saliva, although the level of secretory component associated anti-Spike was less susceptible to decay. Examining two prospective cohorts of subjects that were monitored for infections post-vaccination, we found that participants who were subsequently infected with SARS-CoV-2 had lower levels of vaccine-induced serum anti-Spike/RBD IgA at 2-4 weeks post-dose 2 compared to participants who did not experience an infection, whereas IgG levels were comparable between groups. These data emphasize the importance of developing COVID-19 vaccines that elicit a durable IgA response. One-Sentence SummaryOur study delves into whether intra-muscular mRNA vaccination regimes confer a local IgA response in the oral cavity and whether the IgA response is associated with protection against breakthrough infection.

Ultrasensitive assay for saliva-based SARS-CoV-2 antigen detection.

Widespread SARS-CoV-2 testing is highly valuable for identifying asymptomatic/pre-symptomatic individuals to slow community disease transmission. However, there remains a technological gap for highly reliable, easy, and quick SARS-CoV-2 diagnostic tests that are suitable for frequent mass testing. Compared to the conventional nasopharyngeal (NP) swab-based tests, saliva-based methods are attractive due to easier and safer sampling protocols. Despite its merits in rapid turn-around-time and high throughput compared to traditional PCR-based technologies, the widespread use of saliva-based SARS-CoV-2 rapid antigen tests is hindered by limited analytical sensitivity of current methods. Here, we report the first ultrasensitive, saliva-based SARS-CoV-2 antigen assay with an analytical sensitivity of < 0.32 pg/ml, corresponding to 4 viral RNA copies/{micro}l, which is comparable to that of PCR-based tests. Using the novel electrochemiluminescence (ECL)-based S-PLEX immunoassay, we measured the SARS-CoV-2 nucleocapsid (N) antigen concentration in 105 saliva samples obtained from non-COVID-19 and COVID-19 patients. Our assay displayed absolute specificity and high sensitivity (90.2%), where it correctly identified samples with viral loads up to 35 CT cycles by saliva-based PCR. Paired NP swab-based PCR results were also obtained for 86 cases for comparison. Our assay showed high concordance with saliva-based and NP swab-based PCR in samples with negative (< 0.32 pg/ml) and strongly positive (> 2 pg/ml) N antigen concentrations. Our study unveiled the ultrasensitivity and specificity of the saliva-based S-PLEX assay, demonstrating its clinical value as a high throughput, complementary alternative to PCR-based techniques. The novel technique is especially valuable in cases where compliance to frequent swabbing may be problematic (e.g. schools, nursing homes, etc.).

Evidence for sustained mucosal and systemic antibody responses to SARS-CoV-2 antigens in COVID-19 patients

While the antibody response to SARS-CoV-2 has been extensively studied in blood, relatively little is known about the mucosal immune response and its relationship to systemic antibody levels. Since SARS-CoV-2 initially replicates in the upper airway, the antibody response in the oral cavity is likely an important parameter that influences the course of infection, but how it correlates to the antibody response in serum is not known. Here, we profile by enzyme linked immunosorbent assays (ELISAs) IgG, IgA and IgM responses to the SARS-CoV-2 spike protein (full length trimer) and its receptor binding domain (RBD) in serum (n=496) and saliva (n=90) of acute and convalescent patients with laboratory-diagnosed COVID-19 ranging from 3-115 days post-symptom onset (PSO), compared to negative controls. Anti-CoV-2 antibody responses were readily detected in serum and saliva, with peak IgG levels attained by 16-30 days PSO. Whereas anti-CoV-2 IgA and IgM antibodies rapidly decayed, IgG antibodies remained relatively stable up to 105 days PSO in both biofluids. In a surrogate neutralization ELISA (snELISA), neutralization activity peaks by 31-45 days PSO and slowly declines, though a clear drop is detected at the last blood draw (105-115 days PSO). Lastly, IgG, IgM and to a lesser extent IgA responses to spike and RBD in the serum positively correlated with matched saliva samples. This study confirms that systemic and mucosal humoral IgG antibodies are maintained in the majority of COVID-19 patients for at least 3 months PSO. Based on their correlation with each other, IgG responses in saliva may serve as a surrogate measure of systemic immunity. One Sentence SummaryIn this manuscript, we report evidence for sustained SARS-CoV-2-specific IgG and transient IgA and IgM responses both at the site of infection (mucosae) and systemically in COVID-19 patients over 3 months and suggest that saliva could be used as an alternative biofluid for monitoring IgG to SARS-CoV-2 spike and RBD antigens.

Sensitivity of nasopharyngeal swabs and saliva for the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)

We enrolled 53 consecutive in-patients with COVID-19 at six hospitals in Toronto, Canada, and tested one nasopharyngeal swab/saliva sample pair from each patient for SARS-CoV-2. Overall, sensitivity was 89% for nasopharyngeal swabs and 77% for saliva (p=NS); difference in sensitivity was greatest for sample pairs collected later in illness.