Recapitulating memory B cell responses in a Lymphoid Organ-Chip to evaluate mRNA vaccine boosting strategies (preprint)

Predicting the immunogenicity of candidate vaccines in humans remains a challenge. To address this issue, we developed a Lymphoid Organ-Chip (LO chip) model based on a microfluidic chip seeded with human PBMC at high density within a 3D collagen matrix. Perfusion of the SARS-CoV-2 Spike protein mimicked a vaccine boost by inducing a massive amplification of Spike-specific memory B cells, plasmablast differentiation, and Spike-specific antibody secretion. Features of lymphoid tissue, including the formation of activated CD4+ T cell/B cell clusters and the emigration of matured plasmablasts, were recapitulated in the LO chip. Importantly, myeloid cells were competent at capturing and expressing mRNA vectored by lipid nanoparticles, enabling the assessment of responses to mRNA vaccines. Comparison of on-chip responses to Wuhan monovalent and Wuhan/Omicron bivalent mRNA vaccine boosts showed equivalent induction of Omicron neutralizing antibodies, pointing at immune imprinting as reported in vivo. The LO chip thus represents a versatile platform suited to the preclinical evaluation of vaccine boosting strategies.

Distinct evolution of SARS-CoV-2 Omicron XBB and BA.2.86 lineages combining increased fitness and antibody evasion (preprint)

The unceasing circulation of SARS-CoV-2 leads to the continuous emergence of novel viral sublineages. Here, we isolated and characterized XBB.1, XBB.1.5, XBB.1.9.1, XBB.1.16.1, EG.5.1.1, EG.5.1.3, XBF, BA.2.86.1 and JN.1 variants, representing >80% of circulating variants in November 2023. The XBB subvariants carry few but recurrent mutations in the spike, whereas BA.2.86.1 and JN.1 harbor >30 additional changes. These variants replicated in IGROV-1 but no longer in Vero E6 and were not markedly fusogenic. They potently infected nasal epithelial cells, with EG.5.1.3 exhibiting the highest fitness. Antivirals remained active. Neutralizing antibody (NAb) responses from vaccinees and BA.1/BA.2-infected individuals were markedly lower compared to BA.1, without major differences between variants. An XBB breakthrough infection enhanced NAb responses against both XBB and BA.2.86 variants. JN.1 displayed lower affinity to ACE2 and higher immune evasion properties compared to BA.2.86.1. Thus, while distinct, the evolutionary trajectory of these variants combines increased fitness and antibody evasion.

High fusion and cytopathy of SARS-CoV-2 variant B.1.640.1 (preprint)

SARS-CoV-2 variants with undetermined properties have emerged intermittently throughout the COVID-19 pandemic. Some variants possess unique phenotypes and mutations which allow further characterization of viral evolution and spike functions. Around 1100 cases of the B.1.640.1 variant were reported in Africa and Europe between 2021 and 2022, before the expansion of Omicron. Here, we analyzed the biological properties of a B.1.640.1 isolate and its spike. Compared to the ancestral spike, B.1.640.1 carried 14 amino acid substitutions and deletions. B.1.640.1 escaped binding by some anti-NTD and -RBD monoclonal antibodies, and neutralization by sera from convalescent and vaccinated individuals. In cell lines, infection generated large syncytia and a high cytopathic effect. In primary airway cells, B.1.640.1 replicated less than Omicron BA.1 and triggered more syncytia and cell death than other variants. The B.1.640.1 spike was highly fusogenic when expressed alone. This was mediated by two poorly characterized and infrequent mutations located in the spike S2 domain, T859N and D936H. Altogether, our results highlight the cytopathy of a hyper-fusogenic SARS-CoV-2 variant, supplanted upon the emergence of Omicron BA.1.

Modelling the association between neutralizing antibody levels and SARS-CoV-2 viral dynamics : implications to define correlates of protection against infection (preprint)

Background While anti-SARS-CoV-2 antibody kinetics have been well described in large populations of vaccinated individuals, we still poorly understand how they evolve during a natural infection and how this impacts viral clearance. Methods For that purpose, we analyzed the kinetics of both viral load and neutralizing antibody levels in a prospective cohort of individuals during acute infection by Alpha variant. Results Using a mathematical model, we show that the progressive increase in neutralizing antibodies leads to a shortening of the half-life of both infected cells and infectious viral particles. We estimated that the neutralizing activity reached 90% of its maximal level within 8 days after symptoms onset and could reduce the half-life of both infected cells and infectious virus by a 6-fold factor, thus playing a key role to achieve rapid viral clearance. Using this model, we conducted a simulation study to predict in a more general context the protection conferred by the existence of pre-existing neutralization, due to either vaccination or prior infection. We predicted that a neutralizing activity, as measured by ED50 >103, could reduce by 50% the risk of having viral load detectable by standard PCR assays and by 99% the risk of having viral load above the threshold of cultivable virus. Conclusions This threshold value for the neutralizing activity could be used to identify individuals with poor protection against disease acquisition.

Resistance of Omicron subvariants BA.2.75.2, BA.4.6 and BQ.1.1 to neutralizing antibodies (preprint)

Convergent evolution of SARS-CoV-2 Omicron BA.2, BA.4 and BA.5 lineages has led to the emergence of several new subvariants, including BA.2.75.2, BA.4.6. and BQ.1.1. The subvariants BA.2.75.2 and BQ.1.1 are expected to become predominant in many countries in November 2022. They carry an additional and often redundant set of mutations in the spike, likely responsible for increased transmissibility and immune evasion. Here, we established a viral amplification procedure to easily isolate Omicron strains. We examined their sensitivity to 6 therapeutic monoclonal antibodies (mAbs) and to 72 sera from Pfizer BNT162b2-vaccinated individuals, with or without BA.1/BA.2 or BA.5 breakthrough infection. Ronapreve (Casirivimab and Imdevimab) and Evusheld (Cilgavimab and Tixagevimab) lost any antiviral efficacy against BA.2.75.2 and BQ.1.1, whereas Xevudy (Sotrovimab) remained weakly active. BQ.1.1 was also resistant to Bebtelovimab. Neutralizing titers in triply vaccinated individuals were low to undetectable against BQ.1.1 and BA.2.75.2, 4 months after boosting. A BA.1/BA.2 breakthrough infection increased these titers, which remained about 18-fold lower against BA.2.75.2 and BQ.1.1, than against BA.1. Reciprocally, a BA.5 breakthrough infection increased more efficiently neutralization against BA.5 and BQ.1.1 than against BA.2.75.2. Thus, the evolution trajectory of novel Omicron subvariants facilitated their spread in immunized populations and raises concerns about the efficacy of most currently available mAbs.

Longitudinal analysis of serum neutralization of SARS-CoV-2 Omicron BA.2, BA.4 and BA.5 in patients receiving monoclonal antibodies (preprint)

The emergence of novel Omicron lineages, such as BA.5, may impact the therapeutic efficacy of anti-SARS-CoV-2 neutralizing monoclonal antibodies (mAbs). Here, we evaluated the neutralization and ADCC activity of 6 therapeutic mAbs against Delta, BA.2, BA.4 and BA.5 isolates. The Omicron sub-variants escaped most of the antibodies but remained sensitive to Bebtelovimab and Cilgavimab. Consistent with their shared spike sequence, BA.4 and BA.5 displayed identical neutralization profiles. Sotrovimab was the most efficient at eliciting ADCC. We also analyzed 121 sera from 40 immunocompromised individuals up to 6 months after infusion of 1200 mg of Ronapreve (Imdevimab + Casirivimab), and 300 or 600 mg of Evusheld (Cilgavimab + Tixagevimab). Sera from Ronapreve-treated individuals did not neutralize Omicron subvariants. Evusheld-treated individuals neutralized BA.2 and BA.5, but titers were reduced by 41- and 130-fold, respectively, compared to Delta. A longitudinal evaluation of sera from Evusheld-treated patients revealed a slow decay of mAb levels and neutralization. The decline was more rapid against BA.5. Our data shed light on the antiviral activities of therapeutic mAbs and the duration of effectiveness of Evusheld pre-exposure prophylaxis.

Duration of BA.5 neutralization in sera and nasal swabs from SARS-CoV-2 vaccinated individuals, with or without Omicron breakthrough infection. (preprint)

Since early 2022, Omicron BA.1 has been eclipsed by BA.2, which was in turn outcompeted by BA.5, that displays enhanced antibody escape properties. Here, we evaluated the duration of the neutralizing antibody (Nab) response, up to 16 months after Pfizer BNT162b2 vaccination, in individuals with or without BA.1/BA.2 breakthrough infection. We measured neutralization of the ancestral D614G lineage, Delta and Omicron BA.1, BA.2, BA.5 variants in 291 sera and 35 nasal swabs from 27 individuals. Upon vaccination, serum Nab titers were reduced by 10, 15 and 25 fold for BA.1, BA.2 and BA.5, respectively, compared with D614G. The duration of neutralization was markedly shortened, from an estimated period of 11.5 months post-boost with D614G to 5.5 months with BA.5. After breakthrough, we observed a sharp increase of Nabs against Omicron subvariants, followed by a plateau and a slow decline after 4 or 5 months. In nasal swabs, infection, but not vaccination, triggered a strong IgA response and a detectable Omicron neutralizing activity. Thus, BA.5 spread is partly due to abbreviated vaccine efficacy, particularly in individuals who were not infected with previous Omicron variants.

The spike-stabilizing D614G mutation interacts with S1/S2 cleavage site mutations to promote the infectious potential of SARS-CoV-2 variants (preprint)

ABSTRACT SARS-CoV-2 remained genetically stable during the first three months of the pandemic, before acquiring a D614G spike mutation that rapidly spread worldwide, and then generating successive waves of viral variants with increasingly high transmissibility. We set out to evaluate possible epistatic interactions between the early occurring D614G mutation and the more recently emerged cleavage site mutations present in spike of the Alpha, Delta, and Omicron variants of concern. The P681H/R mutations at the S1/S2 cleavage site increased spike processing and fusogenicity but limited its incorporation into pseudoviruses. In addition, the higher cleavage rate led to higher shedding of the spike S1 subunit, resulting in a lower infectivity of the P681H/R-carrying pseudoviruses compared to those expressing the Wuhan wild-type spike. The D614G mutation increased spike expression at the cell surface and limited S1 shedding from pseudovirions. As a consequence, the D614G mutation preferentially increased the infectivity of P681H/R-carrying pseudoviruses. This enhancement was more marked in cells where the endosomal route predominated, suggesting that more stable spikes could better withstand the endosomal environment. Taken together, these findings suggest that the D614G mutation stabilized S1/S2 association and enabled the selection of mutations that increased S1/S2 cleavage, leading to the emergence of SARS-CoV-2 variants expressing highly fusogenic spikes. AUTHOR SUMMARY The successive emergence of SARS-CoV-2 variants is fueling the COVID pandemic, thus causing a major and persistent public health issue. The parameters involved in the emergence of variants with higher pathogenic potential remain incompletely understood. The first SARS-CoV-2 variant that spread worldwide in early 2020 carried a D614G mutation in the viral spike, making this protein more stable in its cleaved form at the surface of virions, and resulting in viral particles with higher infectious capacity. The Alpha and the Delta variants that spread in late 2020 and early 2021, respectively, proved increasingly transmissible and pathogenic when compared to the original SARS-CoV-2 strain. Interestingly, Alpha and Delta both carried mutations in a spike cleavage site that needs to be processed by cellular proteases prior to viral entry. The cleavage site mutations P681H/R made the Alpha and Delta spikes more efficient at viral fusion, by generating a higher fraction of cleaved spikes subunits S1 and S2. We show here that the early D614G mutation and the late P681H/R mutations act synergistically to increase the fusion capacity of SARS-CoV-2 variants. Specifically, viruses with increased spike cleavage due to P681H/R were even more dependent on the stabilizing effect of D614G mutation, which limited the shedding of cleaved S1 subunits from viral particles. These findings suggest that the worldwide spread of the D614G mutation was a prerequisite to the emergence of more pathogenic SARS-CoV-2 variants with highly fusogenic spikes.

Potent Human Broadly SARS-CoV-2 Neutralizing IgA and IgG Antibodies Effective Against Omicron BA.1 and BA.2 (preprint)

Memory B-cell and antibody responses to the SARS-CoV-2 spike protein contribute to long-term immune protection against severe COVID-19, which can also be prevented by antibody-based interventions. Here, wide SARS-CoV-2 immunoprofiling in COVID-19 convalescents combining serological, cellular and monoclonal antibody explorations, revealed humoral immunity coordination. Detailed characterization of a hundred SARS-CoV-2 spike memory B-cell monoclonal antibodies uncovered diversity in their repertoire and antiviral functions. The latter were influenced by the targeted spike region with strong Fc-dependent effectors to the S2 subunit and potent neutralizers to the receptor binding domain. Amongst those, Cv2.1169 and Cv2.3194 antibodies cross-neutralized SARS-CoV-2 variants of concern including Omicron BA.1 and BA.2. Cv2.1169, isolated from a mucosa-derived IgA memory B cell, demonstrated potency boost as IgA dimers and therapeutic efficacy as IgG antibodies in animal models. Structural data provided mechanistic clues to Cv2.1169 potency and breadth. Thus, potent broadly neutralizing IgA antibodies elicited in mucosal tissues can stem SARS-CoV-2 infection, and Cv2.1169 and Cv2.3194 are prime candidates for COVID-19 prevention and treatment.

SARS-CoV-2 Omicron neutralization and risk of infection among elderly after a booster dose of Pfizer vaccine (preprint)

Background: The protective immunity against Omicron following a BNT162b2 Pfizer booster dose among elderly is not well characterized. Methods: Thirty-eight residents from three nursing homes were recruited for the study. Antibodies targeting the Spike protein of SARS-CoV-2 were measured with the S-Flow assay. Neutralizing activities in sera were measured as effective dilution 50% (ED50) with the S-Fuse assay using authentic isolates of Delta and Omicron. Results: Among the 38 elderly included in the study, with median (inter-quartile range, IQR) age of 88 (81-92) years, 30 (78.9%) had been previously infected. The ED50 of neutralization were lower against Omicron than Delta, and higher among convalescent compared to naive residents. During an Omicron epidemic affecting two of the three nursing homes in December 2021-January 2022, 75% (6/8) of naive residents got infected, compared to 25% (6/24) of convalescents (P=0.03). Antibody levels to Spike and ED50 of neutralization against Omicron after the BNT162b2 booster dose were lower in those with breakthrough infection (n=12) compared to those without (n=20): median of 1256 vs 2523 BAU/mL (P=0.02) and median ED50 of 234 vs 1298 (P=0.0004), respectively. Conclusion: This study confirmed the importance of receiving at least three antigenic exposures to the SARS-CoV-2 Spike protein for achieving satisfactory neutralizing antibody levels. In this population, protection against Omicron infection was increased in individuals who had been previously infected in addition to the three vaccine doses. Thus, a fourth antigenic exposure may be useful in the elderly population to prevent infection with Omicron, a variant known for its high escape immunity properties.

Seroneutralization of Omicron BA.1 and BA.2 in patients receiving anti-SARS-CoV-2 monoclonal antibodies. (preprint)

The SARS-CoV-2 Omicron BA.1 variant has been supplanted in many countries by the BA.2 sub-lineage. BA.2 differs from BA.1 by about 21 mutations in its spike. Human anti-spike monoclonalantibodies(mAbs)areusedforpreventionortreatmentofCOVID-19. However, the capacity of therapeutic mAbs to neutralize BA.1 and BA.2 remains poorly characterized. Here, we first compared the sensitivity of BA.1 and BA.2 to neutralization by 9 therapeutic mAbs. In contrast to BA.1, BA.2 was sensitive to Cilgavimab, partly inhibited by Imdevimab and resistant to Adintrevimab and Sotrovimab. Two combinations of mAbs, Ronapreve (Casirivimab + Imdevimab) and Evusheld (Cilgavimab + Tixagevimab), are indicated as a pre-exposure prophylaxis in immunocompromised persons at risk of severe disease. We analyzed sera from 29 such individuals, up to one month after administration of Ronapreve and/or Evusheld. After treatment, all individuals displayed elevated antibody levels in their sera and neutralized Delta with high titers. Ronapreve recipients did not neutralize BA.1 and weakly impaired BA.2. With Evusheld, neutralization of BA.1 and BA.2 was detected in 19 and 29 out of 29 patients, respectively. As compared to Delta, titers were more severely decreased against BA.1 (344-fold) than BA.2 (9-fold). We further report 4 breakthrough Omicron infections among the 29 participants. Therefore, BA.1 and BA.2 exhibit noticeable differences in their sensitivity to therapeutic mAbs. Anti-Omicron activity of Ronapreve, and to a lesser extent that of Evusheld, is reduced in patients sera, a phenomenon associated with decreased clinical efficacy.

Fusogenicity and neutralization sensitivity of the SARS-CoV-2 Delta sublineage AY.4.2 (preprint)

SARS-CoV-2 lineages are continuously evolving. As of December 2021, the AY.4.2 Delta sub-lineage represented 20 % of sequenced strains in UK and has been detected in dozens of countries. It has since then been supplanted by the Omicron variant. AY.4.2 displays three additional mutations (T95I, Y145H and A222V) in the N-terminal domain (NTD) of the spike when compared to the original Delta variant (B.1.617.2) and remains poorly characterized. Here, we analyzed the fusogenicity of the AY.4.2 spike and the sensitivity of an authentic AY.4.2 isolate to neutralizing antibodies. The AY.4.2 spike exhibited similar fusogenicity and binding to ACE2 than Delta. The sensitivity of infectious AY.4.2 to a panel of monoclonal neutralizing antibodies was similar to Delta, except for the anti-RBD Imdevimab, which showed incomplete neutralization. Sensitivity of AY.4.2 to sera from individuals having received two or three doses of Pfizer or two doses of AstraZeneca vaccines was reduced by 1.7 to 2.1 fold, when compared to Delta. Our results suggest that mutations in the NTD remotely impair the efficacy of anti-RBD antibodies. The temporary spread of AY.4.2 was not associated with major changes in spike function but rather to a partially reduced neutralization sensitivity.

Considerable escape of SARS-CoV-2 variant Omicron to antibody neutralization (preprint)

The SARS-CoV-2 Omicron variant was first identified in November 2021 in Botswana and South Africa 1,2 . It has in the meantime spread to many countries and is expected to rapidly become dominant worldwide. The lineage is characterized by the presence of about 32 mutations in the Spike, located mostly in the N-terminal domain (NTD) and the receptor binding domain (RBD), which may enhance viral fitness and allow antibody evasion. Here, we isolated an infectious Omicron virus in Belgium, from a traveller returning from Egypt. We examined its sensitivity to 9 monoclonal antibodies (mAbs) clinically approved or in development 3 , and to antibodies present in 90 sera from COVID-19 vaccine recipients or convalescent individuals. Omicron was totally or partially resistant to neutralization by all mAbs tested. Sera from Pfizer or AstraZeneca vaccine recipients, sampled 5 months after complete vaccination, barely inhibited Omicron. Sera from COVID-19 convalescent patients collected 6 or 12 months post symptoms displayed low or no neutralizing activity against Omicron. Administration of a booster Pfizer dose as well as vaccination of previously infected individuals generated an anti-Omicron neutralizing response, with titers 5 to 31 fold lower against Omicron than against Delta. Thus, Omicron escapes most therapeutic monoclonal antibodies and to a large extent vaccine-elicited antibodies.

Absence of severe COVID-19 in patients with clonal mast cells activation disorders: effective anti-SARS-CoV-2 immune response. (preprint)

Mast cells are key actors of innate immunity and Th2 adaptive immune response which counterbalance Th1 response, critical for anti-viral immunity. Clonal Mast Cells Activation Disorders (cMCADs) such as mastocytosis and clonal mast cells activation syndrome are characterized by an abnormal mast cells accumulation and/or activation. No data have been published on the anti-viral immune response of patients with cMCADs. The aims of the study were to collected, in a comprehensive way, outcomes of cMCADs patients who experienced a biologically-proven COVID-19 and to characterize both anti-endemic coronaviruses and specific anti-SARS-CoV-2 immune responses in these patients. Clinical follow-up and outcome data were collected prospectively for one year within the French rare disease network CEREMAST encompassing patients from all over the country. Anti-SARS-CoV-2 and anti-endemic coronaviruses specific T-cells were assessed with an enzyme-linked immunospot assay (EliSpot) and anti-SARS-CoV-2 humoral response with dosage of circulating levels of specific IgG, IgA and neutralizing antibodies. Overall, 32 cMCADs patients were identified. None of them required non-invasive or mechanical ventilation; two patients were hospitalized to receive oxygen and steroid therapy. In 21 patients, a characterization of the SARS-CoV-2-specific immune response has been performed. A majority of patients showed a high proportion of circulating SARS-CoV-2-specific interferon (IFN)-{gamma} producing T-cells and high levels of anti-Spike IgG antibodies with neutralizing activity. In addition, no defects in anti-endemic coronaviruses responses were found in patients with cMCADs compared to non-cMCADs controls. Patients with cMCADs frequently showed a spontaneous IFN-{gamma} T-cell production in absence of any stimulation that correlated with circulating basal tryptase levels, a marker of mast cells burden. These findings underscore that patients with cMCADs might be not at risk of severe COVID-19 and the spontaneous IFN-{gamma} production might explain this observation. Author SummaryMast cells are immune cells involved in many biological processes including the anti-microbial response. However, previous studies suggest that mast cells may have a detrimental role in the response against viruses such as SARS-CoV-2, responsible for COVID-19. When a mutation occurs in mast cells, it can lead to a group of diseases called clonal mast cells activation disorders (cMCADs), characterized by deregulated activation of these cells. Hence, patients with cMCADs might be more susceptible to severe COVID-19 than general population. We therefore conducted a 1-year study in France to collect data from all cMCADs patients included in the CEREMAST rare disease French network and who experienced COVID-19. Interestingly, we did not find any severe COVID-19 (i.e. requiring non-invasive or mechanical ventilation) in spite of well-known risk factors for severe COVID-19 in a part of cMCADs patients. We then have studied the immune response against SARS-CoV-2 and other endemic coronaviruses in these patients. We did not observe any abnormalities in the immune response either at the level of T and B lymphocytes. These findings underscore that these patients might not be at risk of severe COVID-19 as one might have feared.

Anti-CD38 therapy impairs SARS-CoV-2 vaccine response in multiple myeloma patients (preprint)

Multiple myeloma (MM) patients are at risk of fatal outcome after SARS-CoV-2 infection. Preliminary data suggest that MM patients have an impaired response to vaccination. This prospective study analyzed the humoral and cellular immune responses to two doses of BNT162b2 in 72 MM patients, including 48 receiving anti-CD38 immunotherapy. Results evidenced that MM patients display lower levels of SARS-CoV-2 specific IgG and IgA antibodies and decreased neutralization of alpha and delta variants when compared to healthy controls. They also showed decreased numbers of circulating IFN{gamma}-producing Spike SARS-CoV-2 specific T lymphocytes. This defective immune response was particularly marked in patients receiving anti-CD38 immunotherapy. Furthermore, a retrospective investigation of MM patients among COVID-19-related death in the Paris area suggested a limited efficacy of BNT162b2 in patients treated with anti-CD38. Overall, these results show a decreased immunogenicity of BNT162b2 in MM patients and stress the need for novel strategies to improve SARS-CoV-2 prophylaxis in immunocompromised individuals.

Immunogenicity of BNT162b2 vaccine Against the Alpha and Delta Variants in Immunocompromised Patients (preprint)

BackgroundThe emergence of strains of SARS-CoV-2 exhibiting increase viral fitness and immune escape potential, such as the Delta variant (B.1.617.2), raises concerns in immunocompromised patients. To what extent Delta evades vaccine-induced immunity in immunocompromised individuals with systemic inflammatory diseases remains unclear. MethodsWe conducted a prospective study in patients with systemic inflammatory diseases (cases) and controls receiving two doses of BNT162b2. Primary end points were anti-spike antibodies levels and cross-neutralization of Alpha and Delta variants after BNT162b2 vaccine. Secondary end points were T-cell responses, breakthrough infections and safety. ResultsSixty-four cases and 21 controls not previously infected with SARS-CoV-2 were analyzed. Kinetics of anti-spike IgG and IgA after BNT162b2 vaccine showed lower and delayed induction in cases, more pronounced with rituximab. Administration of two doses of BNT162b2 generated a neutralizing response against Alpha and Delta in 100% of controls, while sera from only one of rituximab-treated patients neutralized Alpha (5%) and none Delta. Other therapeutic regimens induced a partial neutralizing activity against Alpha, even lower against Delta. All controls and cases except those treated with methotrexate mounted a SARS-CoV-2 specific T-cell response. Methotrexate abrogated T-cell responses after one dose and dramatically impaired T-cell responses after 2 doses of BNT162b2. ConclusionsRituximab and methotrexate differentially impact the immunogenicity of BNT162b2, by impairing B-cell and T-cell responses, respectively. Delta fully escapes the humoral response of individuals treated with rituximab. These findings support efforts to improve BNT162b2 immunogenicity in immunocompromised individuals (Funded by the Fonds IMMUNOV; ClinicalTrials.gov number, NCT04870411).

Reduced sensitivity of infectious SARS-CoV-2 variant B.1.617.2 to monoclonal antibodies and sera from convalescent and vaccinated individuals (preprint)

The SARS-CoV-2 B.1.617 lineage emerged in October 2020 in India. It has since then become dominant in some indian regions and further spread to many countries. The lineage includes three main subtypes (B1.617.1, B.1617.2 and B.1.617.3), which harbour diverse Spike mutations in the N-terminal domain (NTD) and the receptor binding domain (RBD) which may increase their immune evasion potential. B.1.617.2 is believed to spread faster than the other versions. Here, we isolated infectious B.1.617.2 from a traveller returning from India. We examined its sensitivity to monoclonal antibodies (mAbs) and to antibodies present in sera from COVID-19 convalescent individuals or vaccine recipients, in comparison to other viral lineages. B.1.617.2 was resistant to neutralization by some anti-NTD and anti-RBD mAbs, including Bamlanivimab, which were impaired in binding to the B.1.617.2 Spike. Sera from convalescent patients collected up to 12 months post symptoms and from Pfizer Comirnaty vaccine recipients were 3 to 6 fold less potent against B.1.617.2, relative to B.1.1.7. Sera from individuals having received one dose of AstraZeneca Vaxzevria barely inhibited B.1.617.2. Thus, B.1.617.2 spread is associated with an escape to antibodies targeting non-RBD and RBD Spike epitopes.

Release of infectious virus and cytokines in nasopharyngeal swabs from individuals infected with non-B.1.1.7 or B.1.1.7 SARS-CoV-2 variants. (preprint)

The mechanisms that allowed for the SARS-CoV-2 B.1.1.7 variant to rapidly outcompete pre-existing variants in many countries remain poorly characterized. Here, we analyzed viral release, anti-SARS-CoV-2 antibodies and cytokine production in a retrospective series of 427 RTqPCR+ nasopharyngeal swabs collected in COVID-19 patients harbouring either non-B.1.1.7 or B.1.17 variants. We utilized a novel rapid assay, based on S-Fuse-T reporter cells, to quantify infectious SARS-CoV-2. With both non-B.1.1.7 and B.1.1.7 variants, viral titers were highly variable, ranging from 0 to >106 infectious units, and correlated with viral RNA levels. Lateral flow antigenic rapid diagnostic tests (RDTs) were positive in 96% of the samples harbouring infectious virus. About 67 % of individuals carried detectable infectious virus within the first two days after onset of symptoms. This proportion decreased overtime, and viable virus was detected up to 14 days. Samples containing anti-SARS-CoV-2 IgG or IgA did not generally harbour infectious virus. The proportion of individuals displaying viable virus or being RDT-positive was not higher with B.1.1.7 than with non- B.1.1.7 variants. Ct values were slightly but not significantly lower with B.1.1.7. The variant was characterized by a fast decrease of infectivity overtime and a marked release of 17 cytokines (including IFN-b, IP-10, IL-10 and TRAIL). Our results highlight differences between non-B.1.1.7 and B.1.1.7 variants. B.1.1.7 is associated with modified viral decays and cytokine profiles at the nasopharyngeal mucosae during symptomatic infection.

Sensitivity of infectious SARS-CoV-2 B.1.1.7 and B.1.351 variants to neutralizing antibodies (preprint)

SARS-CoV-2 B.1.1.7 and B.1.351 variants emerged respectively in United Kingdom and South Africa and spread in many countries. Here, we isolated infectious B.1.1.7 and B.1.351 strains and examined their sensitivity to anti-SARS-CoV-2 antibodies present in sera and nasal swabs, in comparison with a D614G reference virus. We established a novel rapid neutralization assay, based on reporter cells that become GFP+ after overnight infection. B.1.1.7 was neutralized by 79/83 sera from convalescent patients collected up to 9 months post symptoms, almost similar to D614G. There was a mean 6-fold reduction in titers and even loss of activity against B.1.351 in 40% of convalescent sera after 9 months. Early sera from 19 vaccinated individuals were almost as potent against B.1.1.7 but less efficacious against B.1.351, when compared to D614G. Nasal swabs from vaccine recipients were not neutralizing, except in individuals who were diagnosed COVID-19+ before vaccination. Thus, faster-spreading variants acquired a partial resistance to humoral immunity generated by natural infection or vaccination, mostly visible in individuals with low antibody levels.