Quantifying how single dose Ad26.COV2.S vaccine efficacy depends on Spike sequence features (preprint)

It is of interest to pinpoint SARS-CoV-2 sequence features defining vaccine resistance. In the ENSEMBLE randomized, placebo-controlled phase 3 trial, estimated single-dose Ad26.COV2.S vaccine efficacy (VE) was 56% against moderate to severe–critical COVID-19. SARS-CoV-2 Spike sequences were measured from 484 vaccine and 1,067 placebo recipients who acquired COVID-19 during the trial. In Latin America, where Spike diversity was greatest, VE was significantly lower against Lambda than against Reference and against all non-Lambda variants [family-wise error rate (FWER) p < 0.05]. VE also differed by residue match vs. mismatch to the vaccine-strain residue at 16 amino acid positions (4 FWER p < 0.05; 12 q-value ≤ 0.20). VE significantly decreased with physicochemical-weighted Hamming distance to the vaccine-strain sequence for Spike, receptor-binding domain, N-terminal domain, and S1 (FWER p < 0.001); differed (FWER ≤ 0.05) by distance to the vaccine strain measured by 9 different antibody-epitope escape scores and by 4 NTD neutralization-impacting features; and decreased (p = 0.011) with neutralization resistance level to vaccine recipient sera. VE against severe–critical COVID-19 was stable across most sequence features but lower against viruses with greatest distances. These results help map antigenic specificity of in vivo vaccine protection.

Immune Correlates Analysis of a Single Ad26.COV2.S Dose in the ENSEMBLE COVID-19 Vaccine Efficacy Clinical Trial (preprint)

Anti-spike IgG binding antibody, anti-receptor binding domain IgG antibody, and pseudovirus neutralizing antibody measurements four weeks post-vaccination were assessed as correlates of risk of moderate to severe-critical COVID-19 outcomes through 83 days post-vaccination and as correlates of protection following a single dose of Ad26.COV2.S COVID-19 vaccine in the placebo-controlled phase of ENSEMBLE, an international, randomized efficacy trial. Each marker had evidence as a correlate of risk and of protection, with strongest evidence for 50% inhibitory dilution (ID50) neutralizing antibody titer. The outcome hazard ratio was 0.49 (95% confidence interval 0.29, 0.81; p=0.006) per 10-fold increase in ID50; vaccine efficacy was 60% (43, 72%) at nonquantifiable ID50 (< 2.7 IU50/ml) and rose to 89% (78, 96%) at ID50 = 96.3 IU50/ml. Comparison of the vaccine efficacy by ID50 titer curves for ENSEMBLE-US, the COVE trial of the mRNA-1273 vaccine, and the COV002-UK trial of the AZD1222 vaccine supported consistency of the ID50 titer correlate of protection across trials and vaccine types.

Humoral immune response of patients infected by earlier lineages of SARS-CoV-2 neutralizes wild types of the most prevalent variants in Brazil (preprint)

In the present study, serum samples of 20 hospitalized COVID-19 patients from Brazil who were infected by the earlier SARS-CoV-2 lineages B.1.1.28 and B.1.1.33, and by the variant of concern (VOC) Gamma (P.1) were tested by plaque reduction neutralization test (PRNT90) with wild isolates of a panel of SARS-CoV-2 lineages, including B.1, Zeta, N.10, and the VOCs Gamma, Alpha, and Delta that emerged in different timeframes of the pandemic. The main objectives of the present study were to evaluate if serum of COVID-19 patients infected by earlier lineages of SARS-CoV-2 were capable to neutralize recently emerged VOCs, and if PRNT90 is a reliable serologic method to distinguish infections caused by different SARS-CoV-2 lineages. Overall, sera collected from the day of admittance to the hospital to 21 days after diagnostic of patients infected by the two earlier lineages B.1.1.28 and B.1.1.33 presented neutralizing capacity for all challenged VOCs, including Gamma and Delta, that were the most prevalent VOCs in Brazil. Among all variants tested, Delta and N.10 presented the lowest mean of neutralizing antibody titers, and B.1.1.7, presented the highest titers. Four patients infected with Gamma, that emerged in December 2020, presented neutralizing antibodies for B.1, B.1.1.33 and B.1.1.28, its ancestor lineage. All of them had neutralizing antibodies under the level of detection for the VOC Delta. Interestingly, patients infected by B.1.1.28 presented very similar mean of neutralizing antibody titers for both B.1.1.33 and B.1.1.28. Findings presented here indicate that most patients infected in early stages of COVID-19 pandemic presented neutralizing antibodies up to 21 days after diagnostic capable to neutralize wild types of all recently emerged VOCs in Brazil, and that the PRNT90 it is not a reliable serologic method to distinguish natural infections caused by different SARS-CoV-2 lineages.

A case series of SARS-CoV-2 reinfections caused by the variant of concern Gamma in Brazil (preprint)

The rapid spread of the SARS-CoV-2 Variant of Concern (VOC) Gamma during late 2020 and early 2021 in Brazilian settings with high seroprevalence raised some concern about the potential role of reinfections in driving the epidemic. Very few cases of reinfection associated with the VOC Gamma, however, have been reported. Here we describe 25 cases of SARS-CoV-2 reinfection confirmed by real-time RT-PCR twice within months apart in Brazil. SARS-CoV-2 genomic analysis confirmed that individuals were primo-infected between March and December 2020 with distinct viral lineages, including B.1.1, B.1.1.28, B.1.1.33, B.1.195 and P.2, and then reinfected with the VOC Gamma between 3 to 12 months after primo-infection. The overall mean cycle threshold (Ct) value of the first (25.7) and second (24.5) episodes were roughly similar for the whole group and 14 individuals displayed mean Ct values < 25.0 at reinfection. Sera of 14 patients tested by plaque reduction neutralization test after reinfection displayed detectable neutralizing antibodies against Gamma and other SARS-CoV-2 variants (B.1.33, B.1.1.28 and Delta). All individuals have milder or no symptoms after reinfection and none required hospitalization. The present study demonstrates that the VOC Gamma was associated with reinfections during the second Brazilian epidemic wave in 2021 and raised concern about the potential infectiousness of reinfected subjects. Although individuals here analyzed failed to mount a long-term sterilizing immunity, they developed a high anti-Gamma neutralizing antibody response after reinfection that may provide some protection against severe disease.

Nlrp3 Inflammasome Genetic Variants Are Associated With Risk And/Or Protection To Hospitalization Among Covid-19 Patients From Rio De Janeiro, Brazil (preprint)

Background: COVID-19 has a broad spectrum of clinical manifestations, from asymptomatic to mild or moderate symptoms, reaching the most severe forms and death. The mechanisms underlying the SARS-CoV-2 infection and its clinical evolution are still unclear. Once SARS-CoV-2 infects individuals, host factors are activated by the presence of the virus inside the cells, such as the inflammasome system. The search of risk factors for COVID-19 is of relevance for clinical management. In this study, we investigated the impact of 11 single-base polymorphisms (SNPs) in the NLRP3, CARD8, AIM2, CASP-1, IFI16, and IL-1β inflammasome genes in SARS-CoV-2 infected individuals with distinct disease outcomes. Methods Patients were divided into two groups: (1) inpatients, with severe/critical disease (Hospitalized group, n=451), and (2) convalescent volunteers with prior SARS-CoV-2 infection and a history of asymptomatic to mild symptoms (Mild group, n=43). Patients hospitalized were followed up at a Hospital Center for COVID-19 Pandemic – National Institute of Infectology (INI)/FIOCRUZ, Rio de Janeiro, Brazil, from June 2020 to March 2021. The Mild group was recruited at Oswaldo Cruz Institute (IOC)/FIOCRUZ, Rio de Janeiro, Brazil, in 2020. Genotyping of the SNPs was determined by Real-Time PCR. Protection and risk estimations were performed by unconditional logistic regression models. Results Among the genotyped SNPs, significant differences in the NLRP3 rs1539019 and rs10754558 frequencies were observed between the groups. The C/C genotype (OR adj =6.31; P adj =0.026) or allele C (OR adj =1.05; P adj =0.002) in rs1539019 polymorphism were associated with the risk for hospitalization, while the C/G genotype (OR adj =0.16; P adj =0.016) or carrier-G (OR adj =0.2; P adj =0.028) in rs10754558 polymorphism were associated with protection for hospitalization. Regarding the NLRP3 genetic variants, the A-C-G-C-G haplotype (OR adj =0.14; P adj = 0.030) was associated with protection for hospitalization. No significant association was observed for the other polymorphisms. Conclusions As of our knowledge, this is the first study demonstrating the association of inflammasome NLRP3 variants with risk and/or protection for hospitalization in COVID-19. Studies linking the NLRP3 inflammasome and SARS-CoV-2 infection are still scarce due to the recent emergence of this pathogen. Our results contribute to the discussion of the impact of inflammasomes in the clinical evolution of COVID-19.

Efficacy of Lopinavir-Ritonavir Prophylaxis for Individuals Exposed to SARS-CoV-2: The COPEP Pragmatic Open-Label, Cluster Randomized Trial (preprint)

Background: Since the beginning of the COVID-19 pandemic, no direct antiviral treatment is effective as post-exposure prophylaxis (PEP). Lopinavir/ritonavir (LPV/r) was repurposed as a potential PEP agent against COVID-19.Methods: We conducted a pragmatic open-label, parallel, cluster-randomized superiority trial in four sites in Switzerland and Brazil. Clusters were randomized to receive LPV/r PEP (400/100 mg) twice daily for 5 days or no PEP (surveillance). The primary outcome is the occurrence of COVID-19 within 21 days post-enrollment.Findings: Of 318 participants, 157 (49.4%) were women, median age was 39 (interquartile range, 28-50) years. A total of 209 (179 clusters) participants were randomized to LPV/r PEP and 109 (95 clusters) to surveillance. Baseline characteristics were similar, with the exception of baseline SARS-CoV-2 PCR positivity, which was 3-fold more frequent in the LPV/r arm (34/209 [16.3%] vs 6/109 [5.5%], respectively). During 21-day follow-up, 48/318 (15.1%) participants developed COVID-19: 35/209 (16.7%) in the LPV/r group and 13/109 (11.9%) in the surveillance group (unadjusted hazard ratio 1.44; 95% CI, 0.76 to 2.73). In the primary endpoint analysis adjusted for propensity score to receive LPV/r, the hazard ratio for developing COVID-19 in the LPV/r group vs surveillance was 0.53 (95% CI, 0.23 to 1.23, respectively; P =.14).Interpretation: LPV/r role as PEP for COVID-19 remains unanswered. In this trial, LPV/r over 5 days did not significantly reduce incidence of COVID-19 in exposed individuals. We observed a change in directionality of the effect in favor of LPV/r after adjusting for baseline SARS-CoV-2 PCR results, indicating a potential role of antivirals in COVID-19 prevention.Clinical Trial Registration Details: ClinicalTrials.gov (Identifier: NCT04364022); Swiss National Clinical Trial Portal: SNCTP 000003732.Funding Information: Fondation privée des HUG and Swiss National Fund (project number: 33IC30_166819).Declaration of Interests: None reported.Ethics Approval Statement: The protocol and amendments were approved by Swissmedic and local ethics committees in Switzerland and Brazil. Participants provided written informed consent before study entry.

Identification of SARS-CoV-2 P.1-related lineages in Brazil provides new insights about the mechanisms of emergence of Variants of Concern (preprint)

One of the most remarkable features of the SARS-CoV-2 Variants of Concern (VOC) is the unusually large number of mutations they carry. However, the specific factors that drove the emergence of such variants since the second half of 2020 are not fully resolved. In this study, we described a new SARS-CoV-2 lineage provisionally designated as P.1-like-II that, as well as the previously described lineage P.1-like-I, shares several lineage-defining mutations with the VOC P.1 circulating in Brazil. Reconstructions of P.1 ancestor sequences demonstrate that the entire constellation of mutations that define the VOC P.1 did not accumulate within a single long-term infected individual, but was acquired by sequential addition during interhost transmissions. Our evolutionary analyses further estimate that P.1-ancestors strains carrying half of the P.1-lineage-defining mutations, including those at the receptor-binding domain of the Spike protein, circulated cryptically in the Amazonas state since August 2020. This evolutionary pattern is consistent with the hypothesis that partial human population immunity acquired from natural SARS-CoV-2 infections during the first half of 2020 might have been the major driving force behind natural selection that allowed VOCs' emergence and worldwide spread. These findings also support a long lag-time between the emergence of variants with key mutations of concern and expansion of the VOC P.1 in Brazil.

A randomized, double-blind, controlled trial of convalescent plasma in adults with severe COVID-19 (preprint)

BackgroundAlthough convalescent plasma has been widely used to treat severe coronavirus disease 2019 (COVID-19), data from randomized controlled trials that support its efficacy are limited. ObjectiveTo evaluate the clinical efficacy and safety of convalescent plasma among adults hospitalized with severe and critical COVID-19. DesignRandomized, double-blind, controlled, multicenter, phase 2 trial conducted from April 21st to November 27th, 2020. SettingFive hospitals in New York City (NY, USA) and Rio de Janeiro (Brazil). ParticipantsHospitalized patients aged [≥]18 years with laboratory-confirmed COVID-19, infiltrates on chest imaging and oxygen saturation [≤] 94% on room air or requirement for supplemental oxygen, invasive mechanical ventilation, or extracorporeal membrane oxygenation. InterventionParticipants were randomized 2:1 to a single transfusion of either 1 unit of convalescent or normal control plasma. MeasurementsThe primary outcome was clinical status at 28 days, measured using an ordinal scale and analyzed using a proportional odds model in the intention-to-treat population. ResultsOf 223 participants enrolled, 150 were randomized to receive convalescent plasma and 73 to normal control plasma. At 28 days, no significant improvement in clinical status was observed in participants randomized to convalescent plasma (with an odds ratio (OR) of a 1-point improvement in the scale: 1.50, 95% confidence interval (CI) 0.83-2.68, p=0.180). However, 28-day mortality was significantly lower in participants randomized to convalescent plasma versus control plasma (19/150 [12.6%] versus 18/73 [24.6%], OR 0.44, 95% CI 0.22-0.91, p=0.034). The median titer of anti-SARS-CoV-2 neutralizing antibody in infused convalescent plasma units was 1:160 (IQR 1:80-1:320). In a subset of nasopharyngeal swab samples (n=40) from Brazil that underwent genomic sequencing, no evidence of neutralization-escape mutants was detected. Serious adverse events occurred in 39/147 (27%) participants who received convalescent plasma and 26/72 (36%) participants who received control plasma. LimitationsSome participants did not receive high-titer convalescent plasma. ConclusionIn adults hospitalized with severe COVID-19, use of convalescent plasma was not associated with significant improvement in 28 days clinical status. The significant reduction in mortality associated with convalescent plasma, however, may warrant further evaluation. RegistrationClinicalTrials.gov, NCT04359810 FundingAmazon Foundation Clinical Trial RegistrationClinicalTrials.gov Identifier: NCT04359810