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Intro: Different vaccines against COVID-19 have been approved by the World Health Organization (WHO) at different stages, however, limited data is available on long-term kinetics of antibodies induced by vaccines. This study was performed to investigate the persistence and dynamicity of BBV-152 (Covaxin)- and AZD1222 (Covishield)-induced immunoglobulin-G (IgG) antibodies over the year and neutralizing antibodies' status after the one-month post booster dose. Method(s): This 52-week longitudinal cohort study documented antibody persistence and neutralizing antibody status among 278 health-care workers (HCWs) from four different healthcare and research facilities in Odisha, enrolled in January 2021 and continued until March 2022. An automated chemiluminescence immune assay (CLIA) platform from Abbott Diagnostics was used to quantify IgG antibodies against SARS-CoV-2's spike receptor-binding domain (RBD) and a surrogate virus neutralization test (sVNT) was performed by enzyme-linked immunosorbent assay (ELISA). If any participants developed any symptoms of COVID-19, nasopharyngeal swabs were collected and sent to ICMR- RMRC, Bhubaneswar for RT-PCR confirmation. Finding(s): Among the 243 participants, 119 HCWs (48.97%) were Covaxin recipients and the remaining 124 (51.02%) were Covishield recipients. During the seven follow- ups, 104 participants (42.79%) were identified as vaccine breakthrough cases. In 139 non-infected HCWs, the median antibody titer significantly waned after ten months of double dose, both for Covaxin (342.7 AU/mL at DD1 vs 43.9 AU/mL at DD10) and Covishield (2325.8 AU/mL at DD3 vs 595.2 AU/mL at DD10). No statistically significant differences in antibody titers were observed based on age, gender, comorbidities, and blood groups. The median inhibition activity of sVNT was increased significantly for Covaxin and Covishield booster recipients. Among the booster dose recipients, 24 had breakthrough cases by the Omicron variant. Conclusion(s): Results of this longitudinal cohort study can be used to implement vaccination strategies and could also aid in tracking and designing vaccine mandates to minimize vaccine escape.Copyright © 2023
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Detecting immunity against SARS-CoV-2 is vital for evaluating vaccine response and natural infection, but conventional virus neutralization test (cVNT) requires BSL3 and live viruses, and pseudo-virus neutralization test (pVNT) needs specialized equipment and trained professionals. The surrogate virus neutralization test (sVNT) was developed to overcome these limitations. This study explored the use of angiotensin converting enzyme 2 (ACE2) produced from Nicotiana benthamiana for the development of an affordable neutralizing antibodies detection assay. The results showed that the plant-produced ACE2 can bind to the receptor binding domain (RBD) of the SARS-CoV-2, and was used to develop sVNT with plant-produced RBD protein. The sVNT developed using plant-produced proteins showed high sensitivity and specificity when validated with a group of 30 RBD vaccinated mice sera and the results were correlated with cVNT titer. This preliminary finding suggests that the plants could offer a cost-effective platform for producing diagnostic reagents.
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OBJECTIVES: To assess the long-term humoral immunity induced by booster administration, as well as the ability of binding antibody and surrogate virus neutralization tests (sVNT) to predict neutralizing antibodies (NAbs) against the SARS-CoV-2 Omicron variant. METHODS: A total of 269 sera samples were analyzed from 64 healthcare workers who had received a homologous booster dose of BNT162b2. Neutralizing antibodies assessed by sVNT and anti-RBD IgG measured with the sCOVG assay (Siemens Healthineers®) were analyzed at five timepoints; before and up to 6 months following the booster. Antibody titers were correlated with neutralizing antibodies against the Omicron BA.1 variant obtained by pseudovirus neutralization test (pVNT) as a reference method. RESULTS: While Wild-type sVNT percentage of inhibition (POI) remained above 98.6% throughout the follow-up period after booster administration, anti-RBD IgG and NAbs assessed by Omicron BA.1 pVNT showed respectively a 3.4-fold and 13.3-fold decrease after 6 months compared to the peak reached at day 14. NAbs assessed by Omicron sVNT followed a steady decline until reaching a POI of 53.4%. Anti-RBD IgG and Omicron sVNT assays were strongly correlated (r=0.90) and performed similarly to predict the presence of neutralizing antibodies with Omicron pVNT (area under the ROC: 0.82 for both assays). In addition, new adapted cut-off values of anti-RBD IgG (>1,276 BAU/mL) and Omicron sVNT (POI>46.6%) were found to be better predictors of neutralizing activity. CONCLUSIONS: This study showed a significant drop in humoral immunity 6 months after booster administration. Anti-RBD IgG and Omicron sVNT assays were highly correlated and could predict neutralizing activity with moderate performance.
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Introduction: Kidney transplant recipients (KTRs) are at risk for substantial morbidity and mortality during COVID-19 infection. Vaccination for this group of patients is reccommended. However, immunogenicity and safety data after COVID-19 vaccination among KTRs remains limited. Method(s): We conducted an observational prospective trial involving KTRs at Chiang Mai University hospital, Chiang Mai, Thailand. The participants were received homologous ChAdOx1 nCoV-19 (AZ-AZ), or the heterologous prime-boost of CoronaVac,followed by AZ (SV-AZ). The immunogenicity was assessed by measuring antibodies against the S1 receptor-binding domain (anti-RBD), and SARS-CoV-2 surrogate virus neutralization test (sVNT) at specific timepoints. The primary outcome was the seroconversion rate of sVNT at day 28 after complete vaccination. The secondary outcomes were the seroconversion rate of sVNT at day 28 after the first dose of vaccination, the level of sVNT and anti-RBD at specific timepoints, and the adverse events of each vaccine regimen. Result(s): A total of 18 KTRs were recruited. Among those, 13 (72.2%), and 5 (27.8%) patients were received AZ-AZ, and SV-AZ regimen, respectively. The seroconversion rate of sVNT at day 28 after the second dose were 23.1%, and 20.0% for AZ-AZ, and SV-AZ, respectively (P>0.99). The level of sVNT and the level of anti-RBD at day 28 after the first and at day 28 after the second dose were not different between groups (Figure 1). There were no serious adverse events reported in any vaccine groups. However, AZ-AZ showed sign of tubular dysfunction demonstrated by increasing of fractional excretion of magnesium after complete course of vaccination which correlated to the trend of urine albumin and urine protein creatinine ratio (r=0.720, P=0.013;and r=0.726, P=0.011, respectively). [Formula presented] [Formula presented] Figure 1 Percentage of neutralization inhibition (a) and level of anti-RBD antibody (b) at each visit of homologous ChAdOx1 nCoV-19 (AZ-AZ), and heterologous prime-boost of CoronaVac, followed by AZ (SV-AZ) regimen Conclusion(s): Immunogenicity after COVID-19 vaccination with either homologous or heterologous prime-boost regimen among KTRs was compromised. Homologous replication-defective viral vectors vaccine regimen seemed to affect renal tubular function, and further follow-up should be warranted. No conflict of interestCopyright © 2023
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Background: The pandemic unleashed by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has infected more than 500 million people worldwide and caused more than 6 million deaths. Cellular and humoral immunity induced by infection or immunization are key factors in controlling the viral burden and avoiding the recurrence of coronavirus disease. The duration and effectiveness of immunity after infection is relevant to pandemic policy interventions, including the timing of vaccine boosters. Objectives: We sought to evaluate longitudinal binding and functional antibodies against SARS-CoV-2 receptor-binding domain in police officers and health care workers with a history of coronavirus disease 2019 and compare with SARS-CoV-2-naive individuals after vaccination with adenovirus-based ChAdOx1 nCoV-19 (AstraZeneca-Fiocruz) or the inactivated CoronaVac vaccine (Sinovac-Butantan Institute). Methods: A total of 208 participants were vaccinated. Of these, 126 (60.57%) received the ChAdOx1 nCoV-19 vaccine and 82 (39.42%) received the CoronaVac vaccine. Prevaccination and postvaccination blood was collected, and the amount of anti-SARS-CoV-2 IgG and the neutralizing ability of the antibodies to block the interaction between angiotensin-converting enzyme 2 and receptor-binding domain were determined. Results: Subjects with preexisting SARS-CoV-2 immunity and who received a single dose of ChAdOx1 nCoV-19 or CoronaVac have similar or superior antibody levels when compared with levels in seronegative individuals even after 2 doses of the vaccine. Neutralizing antibody titers of seropositive individuals were higher with a single dose of either ChAdOx1 nCoV-19 or CoronaVac compared with those of seronegative individuals. After 2 doses, both groups reached a plateau response. Conclusions: Our data reinforce the importance of vaccine boosters to increase specific binding and neutralizing SARS-CoV-2 antibodies.
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Background: Cancer patients are at high risk of severe COVID infection and recommended at least three doses of SARS-CoV2 mRNA vaccines. Various anti-neoplastic treatments may affect long-term vaccine immunogenicity. Methods: Patients with solid or haematological cancer were recruited from two Singapore hospitals between July 2021 and March 2022. GenScript cPASS surrogate virus neutralisation assays measured antibody responses, which were correlated with clinical outcomes obtained from medical records and national mandatory-reporting databases. Results: In total, 273 patients were recruited (40 with haematological malignancies and the rest solid tumours). Two-hundred and four patients (74.7%) were receiving active cancer therapy: 98 (35.9%) receiving systemic chemotherapy and the rest targeted or immunotherapy. All patients were seronegative at baseline. After receiving one, two and three doses of SARS-CoV-2-mRNA vaccination, seroconversion rate was 35.2%, 79.4% and 92.4% respectively. After three doses, patients on active treatment for haematological malignancies had lower antibodies (57.3%±46.2) as compared to patients on immunotherapy (94.1%±9.56, p<0.05) and chemotherapy (92.8%±18.1, p<0.05). SARS-CoV-2 infection was reported in 77 (28.2%) patients of which 18 were severe. Conclusion: This study demonstrates high immunogenicity of three doses of vaccines and protection against severe infection in cancer patients.
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The receptor binding domain (RBD) plays a pivotal role in the viral entry as it enables the engagement of severe acute respiratory syndrome 2 (SARS-CoV-2) with the human angiotensin-converting enzyme 2 (ACE2) receptor for host cell entry. RBD is the major target for developing viral inhibitors and vaccines. Expression of recombinant RBD in E.coli is highly scalable with a low-cost procedure despite its high expression level compared to expression in mammalian and yeast cells. Using an alternative natural adjuvant system instead of alum adjuvant, increased immunogenicity of RBD antigen in serological assay including direct ELISA and surrogate Virus Neutralization Test (sVNT) was demonstrated with high levels of IgGs and neutralizing antibodies in mice sera immunized with RBD:AlSa (Alum and Sodium alginate) formulation. The sVNT is a simple and fast test that can be used instead of the conventional virus neutralization test requiring live virus and BSL3 laboratory to detect total neutralizing antibodies against RBD. Additionally, results showed a safety profile for sodium alginate which supported using it as an alternative natural adjuvant.
Subject(s)
COVID-19 , SARS-CoV-2 , Animals , Mice , Humans , Antibodies, Blocking , Antibodies, Viral , Antibodies, Neutralizing , Spike Glycoprotein, Coronavirus/chemistry , MammalsABSTRACT
A severe acute respiratory syndrome coronavirus 2(SARS-CoV-2) surrogate virus neutralization test (sVNT) was used to determine the degree of inhibition of binding between human angiotensin converting enzyme 2 (hACE2) and the receptor binding domain (RBD) of spike protein by neutralizing antibodies in a biosafety level 2 facility. Here, to improve the sensitivity and specificity of the commercial sVNT, we developed a new biotin based sVNT using biotinylated RBD and HRP conjugated streptavidin instead of HRP conjugated RBD for direct detection in an ELISA assay that strongly correlated to the FDA approved cPass sVNT commercial kit (R2 = 0.8521) and pseudo virus neutralization test (R2 = 0.9006) (pVNT). The biotin based sVNT was evaluated in 535 postvaccination serum samples corresponding to second and third boosts of AZD1222 and BNT162b2 vaccines of the wild type strain. We confirmed that the neutralizing antibodies against SARS-CoV-2 variants in second vaccination sera decreased after a median of 141.5 days. Furthermore, vaccination sera from BNT162b2-BNT162b2 vaccines maintained neutralizing antibodies for longer than those of AZD1222 only vaccination. In addition, both vaccines maintained high neutralizing antibodies in third vaccination sera against Omicron BA.2 after a median of 27 days, but neutralizing antibodies significantly decreased after a median of 141.5 days. Along with the cPass sVNT commercial kit, biotin based sVNTs may also be suitable for specifically detecting neutralizing antibodies against multiple SARS-CoV-2 variants; however, to initially monitor the neutralizing antibodies in vaccinated sera using high throughput screening, conventional PRNT could be replaced by sVNT to circumvent the inconvenience of a long test time.
Subject(s)
COVID-19 , SARS-CoV-2 , Humans , SARS-CoV-2/genetics , Biotin , BNT162 Vaccine , ChAdOx1 nCoV-19 , Neutralization Tests , COVID-19/prevention & control , Antibodies, Neutralizing , Antibodies, Viral , Spike Glycoprotein, CoronavirusABSTRACT
Background. We compared neutralizing antibodies in healthy healthcare workers against two different coronavirus 2019 disease (Covid-19) vaccines, a messenger RNA (mRNA)-based vaccine (BNT162b2) and a genetically modified organism (virus vector) vaccine (ChAdOx1), to obtain immunity. In our cohort, we tried to compare the level of neutralizing antibody production after a BNT162b2 vaccine booster and to analyze the level of neutralizing antibody titers for delta and omicron variant. Methods. On November 23 to 25, 2021 and December 23 to 24, 2021, a total of 2,133 HCWs at Soonchunhyang University Bucheon Hospital had been vaccinated with BNT162b2 (Pfizer-BioNTech). Among the 115 HCWs who participated in the preceding study, all participants had no history of Covid-19 infection or suspected symptoms at the time of registration. We collected blood samples from participants four weeks after a third dose. All blood samples were analyzed using the commercial virus neutralization test kit (Genscript Biotech Corporation, Piscataway, NJ, USA). Results. Titers were measured 2 months after the initial vaccination and before the booster of the BNT162b2 vaccine (i.e. 6 months after the initial vaccination), and 4 weeks after the booster. Neutralizing antibody level measured by percentage inhibition of surrogate virus neutralization test (sVNT) readings at negative control (unvaccinated) and 4 weeks after booster vaccination. BNT/BNT/BNT group was defined as a booster dose with BNT162b2 (Pfizer) after two doses of BNT162b2 vaccine (Pfizer) 3 weeks apart. ChA/ChA/BNT group was defined as a booster dose with BNT162b2 (Pfizer) after two doses of ChAdOx1 vaccine (AstraZeneca/Oxford) 12 weeks apart. Conclusion. We found that the neutralizing antibody levels against omicron variant were higher among those who received a booster dose of the BNT162b2 vaccine.
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Purpose: Prior studies suggest that two doses of mRNA vaccine in SOTR may result in lower antibody and T-cell responses relative to levels seen following natural SARS-CoV-2 infection. In this study, we evaluated whether three doses of mRNA-1273 vaccine result in immune responses more comparable to, or greater than, natural infection. Method(s): Serum was collected 4-6 weeks from symptom onset in n=74 SOTR recovered from SARS-CoV-2 infection, and in n=60 SOTR receiving a third dose of mRNA-1273. Disease severity in the infection cohort ranged from mild to severe, but no deaths were reported. Vaccinated SOTR all had negative anti-nucleoprotein antibody results to confirm absence of infection. SARS-CoV-2 serology was assessed using an anti-spike (S) receptor binding domain (RBD) immunoassay (Roche). Neutralizing antibodies (nAb) were assessed using a commercial surrogate virus neutralization test (SVNT) targeting wildtype (WT), alpha, beta and delta strains (GenScript). A subset of participants underwent spike-specific T-cell testing (infection n=50, three doses n=34). PBMCs were stimulated overnight with overlapping peptides and frequencies of S-specific polyfunctional CD4+ and CD8+ T-cells (expressing IFN-gamma and IL-2) were measured by intracellular cytokine staining. Mann Whitney U, and Chi-square tests were used for statistical comparisons;significance was defined at p<0.05. Result(s): Anti-S RBD antibodies in SOTR recovered from infection were similar to levels in those receiving three doses of mRNA-1273 (median U/mL [IQR]: 73.5 [14.9-240.1] vs. 313.8 [313.8-2191.0];p=0.17). Relative to SOTR recovered from infection, the proportion of SOTR positive for nAb after three doses of vaccine was significantly lower. This was true for WT (93.2% vs. 60.0%, p<0.0001) and all variants tested - alpha: 90.5% vs. 56.7%, p<0.0001;beta: 67.6% vs. 50%, p=0.039;and delta: 85.1% vs. 55%, p=0.0001. Spike-specific polyfunctional CD4+ T-cell frequencies were similar between infection and three doses of vaccine (median cell frequency [IQR]: 241.7 [50-539.7] vs. 432.4 [50-1226];p>0.05). Spike-specific polyfunctional CD8+ T-cells were uncommonly detected following infection or vaccination. Vaccinated participants were significantly older than infected SOTR (p<0.001), and some differences in type of transplant were found between groups. However, sex and type of immunosuppressive medications were similar between infected and vaccinated SOTR cohorts (p>0.05). Conclusion(s): Three doses of mRNA vaccine may be required to optimize binding antibody, and to a lesser extent, CD4+ T-cell immunity, to levels similar to natural infection. However, nAb responses to wild-type virus and variants of concern were highest in SOTR recovered from infection when compared to vaccinated patients. These data provide further evidence of impaired SARS-CoV-2 vaccine responses in SOTR.
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Purpose: An additional dose of the COVID-19 vaccine following a primary series is recommended for solid organ transplant recipients. However, a comparison of the immunogenicity and safety of an additional dose of COVID-19 vaccine among different platforms has not been investigated. Method(s): Eligible adult KT recipients were randomized using a stratified (by a previous vaccine regimen) block randomization approach to receive either ChAdOx1 nCoV-19 vaccine (AstraZeneca);V group, or mRNA vaccine (Pfizer-BioNTech or mRNA-1273);M group. Two weeks later, humoral immunity (HMI) was evaluated by anti-RBD IgG level, and percentages of neutralizing antibody inhibition using surrogate viral neutralization test (%SVNT), and cell-mediated immunity (CMI) was investigated by the ELISpot assays. Result(s): A total of 85 KT recipients were included. Of those, 62% were male with the median (IQR) age of 50 (43-59) years. The median (IQR) duration after transplantation was 46 (26-82) months. Twenty-six (34%) and 51 (66%) of those received two-dose of CoronaVac followed by one dose of ChAdOx1 nCoV-19 vaccine and two-dose of ChAdOx1 nCoV-19 vaccine, respectively. Of all, 42 and 43 patients were assigned to V and M groups, respectively. At two weeks after an additional dose, KT recipients in the M group elicited a greater trend of the median (IQR) anti-RBD antibody levels compared to those in the V group (51.8 [5.1-591] vs. 28.5 [2.9-119.3] BAU/mL,p=0.18), which resulted in significantly higher seroconversion rate (anti-RBD antibody > 7.1 BAU/mL) in M group than those in V group (83%vs.51%, p<0.01). Additionally, sVNT positivity rate (%SVNT > 35%) were also significantly greater in M group (58%vs32%, p=0.03). However, there was no difference in S1-specific T cell and RBD-specific B cell responses between M and V groups (230 [41-420] vs. 268 [118-510] SFUs/106 PMBCs, p=0.65 and 2 [0-10] vs. 2 [0-13] SFUs/106 PMBCs, p=0.60). Adverse events were mild and similar between groups (p=NS). Conclusion(s): KT recipients who received an additional dose of mRNA COVID-19 vaccine could elicit HMI greater than a viral vector COVID-19 vaccine along with comparable CMI and safety profile.
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Purpose: Emerging SARS-CoV-2 variants may be associated with a higher risk of breakthrough infections compared to wild-type (WT) virus in kidney transplant recipients (KTRs). The purpose of this study was to evaluate antiviral immune responses against WT and Delta (B.1.617.2) variant of SARS-CoV-2 after 3 doses of SARS-CoV-2 mRNA vaccines in KTRs. Method(s): We conducted a multicenter prospective cohort study of adult KTRs who received 3 doses of BNT162b2 or mRNA-1273. Blood samples were collected from KTRs before and 4 weeks after the 3rd vaccine dose. Sera from pre-pandemic healthy controls (HCs) and pre-pandemic kidney transplant control patients (KCs) were used for comparison. A Luminex-based multiplex assay was used to measure anti-spike antibodies for the WT, Alpha, Beta, Gamma and Delta variants of SARSCoV- 2. A surrogate virus neutralization test was used to assess neutralization against the WT and Delta variant. Patients were also monitored for rejection using several non-invasive biomarkers. Result(s): 54 KTRs were enrolled in the study. The median age was 63, 44% were female and the median time post-transplantation was 42 months. 94% received BNT162b2 vaccine. After the 3rd vaccine dose, there was a significant increase in anti-spike antibody MFIs against the WT, Alpha, Beta, Gamma and Delta variants (Fig. 1A, p<0.0001 for all). For comparison, all pre-pandemic HCs and KCs had a negative result for anti-spike antibody levels (Fig. 1B). Prior to the 3rd vaccine dose, 29% of KTRs had anti-spike antibodies against the WT compared to only 2% against the Delta variant (Fig. 1C, p=0.0001). After the 3rd vaccine dose, 67% of KTRs had anti-spike antibodies against the WT compared to 25% against the Delta variant (p<0.0001, Fig. 1D). Differences between WT and other variants are shown in Figure 1C-D. After the 3rd vaccine dose, there was a 2.1-fold and 2.5-fold increase in the percentage of KTRs with neutralizing responses against the WT and Delta variant respectively (p<0.0001 for both). There was no significant change in serum creatinine, proteinuria, or donor-derived cell-free DNA levels after vaccination. No episodes of rejection occurred during follow-up. Conclusion(s): Two doses of SARS-CoV-2 mRNA vaccines in KTRs are associated with minimal anti-spike antibody response directed against the Delta variant of SARS-CoV-2. After the third dose, a quarter of KTRs developed anti-spike antibodies directed against the Delta variant of SARS-CoV-2.
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Background: Heterologous prime-boost SARS-CoV-2 vaccination is a widely accepted strategy during the COVID-19 pandemic, which generated a superior immune response than homologous vaccination strategy. Objective: To describe immunogenicity of heterologous prime-boost vaccination with inactivated vaccine, CoronaVac, followed by BNT162b2 and 5-month booster dose with BNT162b2 in healthy Thai adolescents. Methods: Adolescents aged 12-18 years were randomized 1:1:1:1 to receive CoronaVac (SV) followed by BNT162b2 (PZ) 30 or 20 µg at either 3- or 6-week interval (SV3w/PZ30µg, SV3w/PZ20µg, SV6w/PZ30µg or SV6w/PZ20µg). During the Omicron-predominant period, participants were offered a BNT162b2 booster dose 30, 15, or 10 µg. Immunogenicity was determined using IgG antibody against spike-receptor-binding domain of wild type(anti-S-RBD IgG) and surrogate virus neutralization test(sVNT) against Delta variant at 14 days and 5 months after the 2nd dose. Neutralization tests(sVNT and pseudovirus neutralization test; pVNT) against Omicron strain were tested pre- and 14 days post-booster dose. Results: In October 2021, 76 adolescents with a median age of 14.3 years (IQR 12.7-16.0) were enrolled: 20 in SV3w/PZ30µg; 17 in SV3w/PZ20µg; 20 in SV6w/PZ30µg; 19 in SV6w/PZ20µg. At day 14, the geometric mean(GM) of anti-S-RBD IgG in SV3w/PZ30µg was 4713 (95 %CI 4127-5382) binding-antibody unit (BAU)/ml, while geometric mean ratio(GMR) was 1.28 (1.09-1.51) in SV6w/PZ30µg. The GMs of sVNT against Delta variants at day 14 among participants in SV3w/PZ30µg and SV6wk/PZ30µg arm were 95.3 % and 99.7 %inhibition, respectively. At 5 months, GMs of sVNT against Delta variants in SV3w/PZ30µg were significantly declined to 47.8 % but remained at 89.0 % inhibition among SV6w/PZ30µg arm. In April 2022, 52 adolescents received a BNT162b2 booster dose. Proportion of participants with sVNT against Omicron strain > 80 %inhibition was significantly increased from 3.8 % pre-booster to 67 % post-booster. Proportion of participants with pVNT ID50 > 185 was 42 % at 14 days post 2nd dose and 88 % post booster, respectively. Conclusions: Heterologous prime-boost vaccination with CoronaVac followed by BNT162b2 induced high neutralizing titer against SARS-CoV-2 Delta strain. After 5-month interval, booster with BNT162b2 induced high neutralizing titer against Omicron strain.Thai Clinical Trials Registry (thaiclinicaltrials.org): TCTR20210923012.
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Serological assays are useful in investigating the development of humoral immunity against SARS-CoV-2 in the context of epidemiological studies focusing on the spread of protective immunity. The plaque reduction neutralization test (PRNT) is the gold standard method to assess the titer of protective antibodies in serum samples. However, to provide a result, the PRNT requires several days, skilled operators, and biosafety level 3 laboratories. Therefore, alternative methods are being assessed to establish a relationship between their outcomes and PRNT results. In this work, four different immunoassays (Roche Elecsys® Anti SARS-CoV-2 S, Snibe MAGLUMI® SARS-CoV-2 S-RBD IgG, Snibe MAGLUMI® 2019-nCoV IgG, and EUROIMMUN® SARS-CoV-2 NeutraLISA assays, respectively) have been performed on individuals healed after SARS-CoV-2 infection. The correlation between each assay and the reference method has been explored through linear regression modeling, as well as through the calculation of Pearson's and Spearman's coefficients. Furthermore, the ability of serological tests to discriminate samples with high titers of neutralizing antibodies (>160) has been assessed by ROC curve analyses, Cohen's Kappa coefficient, and positive predictive agreement. The EUROIMMUN® NeutraLISA assay displayed the best correlation with PRNT results (Pearson and Spearman coefficients equal to 0.660 and 0.784, respectively), as well as the ROC curve with the highest accuracy, sensitivity, and specificity (0.857, 0.889, and 0.829, respectively).
Subject(s)
COVID-19 , SARS-CoV-2 , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/diagnosis , COVID-19 Testing , Humans , Immunoglobulin G , Sensitivity and Specificity , Serologic Tests/methodsABSTRACT
Background: Limited commercial LFA assays are available to provide a reliable quantitative measurement of the total binding antibody units (BAU/mL) against the receptor-binding domain of the SARS-CoV-2 spike protein (S-RBD). Aim: This study aimed to evaluate the performance of the fluorescence LFA FinecareTM 2019-nCoV S-RBD test along with its reader (Model No.: FS-113) against the following reference methods: (i) the FDA-approved GenScript surrogate virus-neutralizing assay (sVNT); and (ii) three highly performing automated immunoassays: BioMérieux VIDAS®3, Ortho VITROS®, and Mindray CL-900i®. Methods: Plasma from 488 vaccinees was tested by all aforementioned assays. Fingerstick whole-blood samples from 156 vaccinees were also tested by FinecareTM. Results and conclusions: FinecareTM showed 100% specificity, as none of the pre-pandemic samples tested positive. Equivalent FinecareTM results were observed among the samples taken from fingerstick or plasma (Pearson correlation r = 0.9, p < 0.0001), suggesting that fingerstick samples are sufficient to quantitate the S-RBD BAU/mL. A moderate correlation was observed between FinecareTM and sVNT (r = 0.5, p < 0.0001), indicating that FinecareTM can be used for rapid prediction of the neutralizing antibody (nAb) post-vaccination. FinecareTM BAU results showed strong correlation with VIDAS®3 (r = 0.6, p < 0.0001) and moderate correlation with VITROS® (r = 0.5, p < 0.0001) and CL-900i® (r = 0.4, p < 0.0001), suggesting that FinecareTM can be used as a surrogate for the advanced automated assays to measure S-RBD BAU/mL.
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Introduction: Limited data exist on the immunogenicity of SARS-CoV-2 vaccination in liver transplant (LT) recipients. Homologous vaccination protocol demonstrated suboptimal responses in this population. Several studies showed that heterologous prime-boost approach yielded a better seroconversion rate in healthy individuals. We, therefore, aimed to explore the immunogenicity and safety of heterologous prime-boost immunization with ChAdOx1 nCoV-19 AstraZeneca (AZ) and BNT162b2 Pfizer-BioNTech (BNT) among LT patients. Method: LT recipients receiving SARS-CoV-2 vaccine at King Chulalongkorn Memorial Hospital between April and November 2021 were consecutively enrolled. Patients received either heterologous prime-boost protocol (AZ/BNT) or homologous regimen (AZ/AZ) depending on national policies. Blood samples were collected before vaccine administration and 4 weeks after the second dose. SARS-CoV-2 spike receptor-binding-domain (RBD) IgG was measured using electrochemiluminescence immunoassay (Roche). A titer $100 U/mL was considered as a protective antibody. The neutralizing antibody was detected by enzymelinked immunosorbent assay-based surrogate virus neutralization test (sVNT) (GenScript) with positive cut-off of ≥30% inhibition. Adverse events (AEs) within 7 days following vaccination were recorded by questionnaires. Results: Eighty-nine LT recipients were enrolled. Of these, 64 (71.9%) and 25 (28.1%) patients received AZ/BNT and AZ/AZ, respectively. Majority was male (68.5%) with mean age of 58.3±14.5 years. Median time since transplant was 5.7 years (IQR 2.8-11.8). No patient had a previous diagnosis of SARSCoV- 2 infection. Comparable seroconversion rate of anti-RBD antibody and sVNT after vaccination were observed in both AZ/BNT and AZ/AZ groups (70.3% vs 64.0%, p=0.62 and 77.8% vs 59.1%, p=0.10, respectively). However, median IgG titer was significantly higher in AZ/BNT group compared to AZ/AZ group (842.9 U/mL vs 152.2 U/mL, p=0.011). Patients receiving AZ/BNT also had significantly higher sVNT levels than those receiving AZ/BNT (91.2% vs 35.8%, p=0.003) (Figure 1). In multivariate analysis, use of prednisolone (OR 6.3, 95%CI 1.1-35.4, p=0.036) and LT duration <5 years (OR 3.2, 95%CI 1.1-9.1, p= 0.029) were associated with lack of protective serological response. AZ/BNT regimen trended to have higher reported AEs than AZ/AZ (Figure 2). No graft rejection or serious AEs were found. Conclusion: Heterologous prime-boost regimen with adenoviral vectored and mRNA SARS-CoV-2 vaccine yielded greater humoral response than homologous protocol and was well-tolerated in LT recipients. T-cell response, which might play additional role in protection, and immune durability of this mix-and-match strategy are eagerly awaited. (Figure Presented)
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Determination of neutralizing antibody titers is still considered the gold standard for infection protection. A full virus neutralization test (VNT) with replication-competent, infectious SARS-CoV-2, is labor-intensive and requires Biosafety Level 3 certified laboratories. Therefore, several commercial SARS-CoV-2 surrogate virus neutralization tests (sVNTs) have been developed that aim to detect neutralizing antibodies targeting the receptor binding domain (RBD) of the viral spike glycoprotein (S). Neutralizing antibodies to the RBD block its interaction with the angiotensin-converting enzyme 2 (ACE2) receptor protein. Here, we compared a full virus neutralization test (VNT) with two SARS-CoV-2 surrogate virus neutralization tests (sVNT) and validated them in two cohorts of i) convalescent SARS-CoV-2-infected individuals and ii) COVID vaccinated individuals. The sVNTs showed highly different results both, compared to the VNT-titers and also between the two cohorts. This indicates that currently, sVNT provide a qualitative instead of a quantitative measurement of neutralizing antibodies. The findings in this work show that the cutoff levels for sVNTs might need to be readjusted for convalescent and vaccinated individuals.
Subject(s)
COVID-19 , SARS-CoV-2 , Antibodies, Neutralizing , Antibodies, Viral , COVID-19/diagnosis , Humans , Neutralization Tests , Spike Glycoprotein, CoronavirusABSTRACT
The coronavirus disease 2019 (COVID-19) pandemic has progressed over 2 years since its onset causing significant health concerns all over the world and is currently curtailed by mass vaccination. Immunity acquired against severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) can be following either infection or vaccination. However, one can never be sure whether the acquired immunity is adequate to protect the individual from subsequent infection because of three important factors: individual variations in humoral response dynamics, waning of protective antibodies over time, and the emergence of immune escape mutants. Therefore, a test that can accurately differentiate the protected from the vulnerable is the need of the hour. The plaque reduction neutralization assay is the conventional gold standard test for estimating the titers of neutralizing antibodies that confer protection. However, it has got several drawbacks, which hinder the practical application of this test for wide-scale usage. Hence, various tests have been developed to detect protective immunity against SARS-CoV-2 that directly or indirectly assess the presence of neutralizing antibodies to SARS-CoV-2 in a lower biosafety setting. In this review, the pros and cons of the currently available assays are elaborated in detail and special focus is put on the scope of the novel split nanoluciferase technology for detecting SARS-CoV-2 neutralizing antibodies.
Subject(s)
COVID-19 , SARS-CoV-2 , Antibodies, Neutralizing , Antibodies, Viral , Humans , Neutralization Tests , TechnologyABSTRACT
OBJECTIVES: Evaluating anti-SARS-CoV-2 antibody levels is a current priority to drive immunization, as well as to predict when a vaccine booster dose may be required and for which priority groups. The aim of our study was to investigate the kinetics of anti-SARS-CoV-2 Spike S1 protein IgG (anti-S1 IgG) antibodies and neutralizing antibodies (NAbs) in an Italian cohort of healthcare workers (HCWs), following the Pfizer/BNT162b2 mRNA vaccine, over a period of up to six months after the second dose. METHODS: We enrolled 57 HCWs, without clinical history of COVID-19 infection. Fluoroenzyme-immunoassay was used for the quantitative anti-S1 IgG antibodies at different time points T1 (one month), T3 (three months) and T6 (six months) following the second vaccine shot. Simultaneously, a commercial surrogate virus neutralization test (sVNT) was used for the determination of NAbs, expressed as inhibition percentage (% IH). RESULTS: Median values of anti-S1 IgG antibodies decreased from T1 (1,452 BAU/mL) to T6 (104 BAU/mL) with a percent variation of 92.8% while the sVNT showed a percent variation of 34.3% for the same time frame. The decline in anti-S1 IgG antibodies from T1 to T6 was not accompanied by a loss of the neutralizing capacity of antibodies. In fact at T6 a neutralization percentage <20% IH was observed only in 3.51% of HCWs. CONCLUSIONS: Our findings reveal that the decrease of anti-S1 IgG levels do not correspond in parallel to a decrease of NAbs over time, which highlights the necessity of using both assays to assess vaccination effectiveness.
Subject(s)
Antibodies, Neutralizing , COVID-19 , Antibodies, Viral , BNT162 Vaccine , COVID-19/prevention & control , COVID-19 Vaccines , Health Personnel , Humans , Immunoglobulin G , SARS-CoV-2 , Serologic Tests , Vaccines, Synthetic , mRNA VaccinesABSTRACT
BACKGROUND: Several commercial surrogate Virus Neutralization Tests (sVNTs) have been developed in the last year. Neutralizing anti-SARS-CoV-2 antibodies through interaction with Spike protein Receptor Binding Domain (S-RBD) can block the virus from entering and infecting host cells. However, there is a lack of information about the functional activity of SARS-CoV-2 antibodies that may be associated with protective responses. For these reasons, to counteract viral infection, the conventional virus neutralization test (VNT) is still considered the gold standard. The aim of this study was to contribute more and detailed information about sVNTs' performance, by determining in vitro the anti-SARS-CoV-2 neutralizing antibody concentration using four different commercial assays and then comparing the obtained data to VNT. METHODS: Eighty-eight samples were tested using two chemiluminescence assays (Snibe and Mindray) and two ELISA assays (Euroimmun and Diesse). The antibody titers were subsequently detected and quantified by VNT. RESULTS: The overall agreement between each sVNT and VNT was 95.45% for Euroimmun and 98.86% for Diesse, Mindray and Snibe. Additionally, we investigated whether the sVNTs were closer to the gold standard than traditional anti-SARS-CoV-2 antibody assays S-RBD or S1 based, finding a higher agreement mean value for sVNTs (98.01 ± 1.705% vs 95.45 ± 1.921%; p < 0.05). Furthermore, Spearman's statistical analysis for the correlation of sVNT versus VNT showed r = 0.666 for Mindray; r = 0.696 for Diesse; r = 0.779 for Mindray and r = 0.810 for Euroimmun. CONCLUSIONS: Our data revealed a good agreement between VNT and sVNTs. Despite the VNT still remains the gold standard, the sVNT might be a valuable tool for screening wider populations.