Low hanging fruit for combatting SARS-CoV-2?

Entry of SARS-CoV-2 into human respiratory cells, mediated by the spike protein, is absolutely dependent on the cellular receptor ACE2 (angiotensin-converting enzyme-2). This makes ACE2 an attractive target for therapeutic intervention in COVID-19. In this issue, Zuo et al. discover that vitamin C, an essential nutrient and common dietary supplement, can target ACE2 for ubiquitin-dependent degradation, resulting in the inhibition of SARS-CoV-2 infection (Zuo et al, 2023). The study identifies novel mechanisms of cellular ACE2 regulation and may inform the design of therapeutics targeting SARS-2 and related coronaviruses.

A Zn2+-triggered two-step mechanism of CLIC1 membrane insertion and activation into chloride channels.

The chloride intracellular channel (CLIC) protein family displays the unique feature of altering its structure from a soluble form to a membrane-bound chloride channel. CLIC1, a member of this family, is found in the cytoplasm or in internal and plasma membranes, with membrane relocalisation linked to endothelial disfunction, tumour proliferation and metastasis. The molecular switch promoting CLIC1 activation remains under investigation. Here, cellular Cl- efflux assays and immunofluorescence microscopy studies have identified intracellular Zn2+ release as the trigger for CLIC1 activation and membrane insertion. Biophysical assays confirmed specific binding to Zn2+, inducing membrane association and enhancing Cl- efflux in a pH-dependent manner. Together, our results identify a two-step mechanism with Zn2+ binding as the molecular switch promoting CLIC1 membrane insertion, followed by pH-mediated activation of Cl- efflux.

Evolutionary remodelling of N-terminal domain loops fine-tunes SARS-CoV-2 spike.

The emergence of SARS-CoV-2 variants has exacerbated the COVID-19 global health crisis. Thus far, all variants carry mutations in the spike glycoprotein, which is a critical determinant of viral transmission being responsible for attachment, receptor engagement and membrane fusion, and an important target of immunity. Variants frequently bear truncations of flexible loops in the N-terminal domain (NTD) of spike; the functional importance of these modifications has remained poorly characterised. We demonstrate that NTD deletions are important for efficient entry by the Alpha and Omicron variants and that this correlates with spike stability. Phylogenetic analysis reveals extensive NTD loop length polymorphisms across the sarbecoviruses, setting an evolutionary precedent for loop remodelling. Guided by these analyses, we demonstrate that variations in NTD loop length, alone, are sufficient to modulate virus entry. We propose that variations in NTD loop length act to fine-tune spike; this may provide a mechanism for SARS-CoV-2 to navigate a complex selection landscape encompassing optimisation of essential functionality, immune-driven antigenic variation and ongoing adaptation to a new host.

Human seasonal coronavirus neutralization and COVID-19 severity.

The virus severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), responsible for the global coronavirus disease-2019 (COVID-19) pandemic, spread rapidly around the world causing high morbidity and mortality. However, there are four known, endemic seasonal coronaviruses in humans (HCoVs), and whether antibodies for these HCoVs play a role in severity of COVID-19 disease has generated a lot of interest. Of these seasonal viruses NL63 is of particular interest as it uses the same cell entry receptor as SARS-CoV-2. We use functional, neutralizing assays to investigate cross-reactive antibodies and their relationship with COVID-19 severity. We analyzed the neutralization of SARS-CoV-2, NL63, HKU1, and 229E in 38 COVID-19 patients and 62 healthcare workers, and a further 182 samples to specifically study the relationship between SARS-CoV-2 and NL63. We found that although HCoV neutralization was very common there was little evidence that these antibodies neutralized SARS-CoV-2. Despite no evidence in cross-neutralization, levels of NL63 neutralizing antibodies become elevated after exposure to SARS-CoV-2 through infection or following vaccination.

Localization and functional characterization of the alternative oxidase in Naegleria.

The alternative oxidase (AOX) is a protein involved in supporting enzymatic reactions of the Krebs cycle in instances when the canonical (cytochrome-mediated) respiratory chain has been inhibited, while allowing for the maintenance of cell growth and necessary metabolic processes for survival. Among eukaryotes, alternative oxidases have dispersed distribution and are found in plants, fungi, and protists, including Naegleria ssp. Naegleria species are free-living unicellular amoeboflagellates and include the pathogenic species of N. fowleri, the so-called "brain-eating amoeba." Using a multidisciplinary approach, we aimed to understand the evolution, localization, and function of AOX and the role that plays in Naegleria's biology. Our analyses suggest that AOX was present in last common ancestor of the genus and structure prediction showed that all functional residues are also present in Naegleria species. Using cellular and biochemical techniques, we also functionally characterize N. gruberi's AOX in its mitochondria, and we demonstrate that its inactivation affects its proliferation. Consequently, we discuss the benefits of the presence of this protein in Naegleria species, along with its potential pathogenicity role in N. fowleri. We predict that our findings will spearhead new explorations to understand the cell biology, metabolism, and evolution of Naegleria and other free-living relatives.

Detection of Serum Cross-Reactive Antibodies and Memory Response to SARS-CoV-2 in Prepandemic and Post-COVID-19 Convalescent Samples.

BACKGROUND: A notable feature of coronavirus disease 2019 (COVID-19) is that children are less susceptible to severe disease. Children are known to experience more infections with endemic human coronaviruses (HCoVs) compared to adults. Little is known whether HCoV infections lead to cross-reactive anti-severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) antibodies. METHODS: We investigated the presence of cross-reactive anti-SARS-CoV-2 IgG antibodies to spike 1 (S1), S1-receptor-binding domain (S1-RBD), and nucleocapsid protein (NP) by enzyme-linked immunosorbent assays, and neutralizing activity by a SARS-CoV-2 pseudotyped virus neutralization assay, in prepandemic sera collected from children (n = 50) and adults (n = 45), and compared with serum samples from convalescent COVID-19 patients (n = 16). RESULTS: A significant proportion of children (up to 40%) had detectable cross-reactive antibodies to SARS-CoV-2 S1, S1-RBD, and NP antigens, and the anti-S1 and anti-S1-RBD antibody levels correlated with anti-HCoV-HKU1 and anti-HCoV-OC43 S1 antibody titers in prepandemic samples (P < .001). There were marked increases of anti-HCoV-HKU1 and - OC43 S1 (but not anti-NL63 and -229E S1-RBD) antibody titers in serum samples from convalescent COVID-19 patients (P < .001), indicating an activation of cross-reactive immunological memory to ß-coronavirus spike. CONCLUSIONS: We demonstrated cross-reactive anti-SARS-CoV-2 antibodies in prepandemic serum samples from children and young adults. Promoting this cross-reactive immunity and memory response derived from common HCoV may be an effective strategy against SARS-COV-2 and future novel coronaviruses.

Identification and characterisation of a cryptic Golgi complex in Naegleria gruberi.

Although the Golgi complex has a conserved morphology of flattened stacked cisternae in most eukaryotes, it has lost the stacked organisation in several lineages, raising the question of what range of morphologies is possible for the Golgi. In order to understand this diversity, it is necessary to characterise the Golgi in many different lineages. Here, we identify the Golgi complex in Naegleria, one of the first descriptions of an unstacked Golgi organelle in a non-parasitic eukaryote, other than fungi. We provide a comprehensive list of Golgi-associated membrane trafficking genes encoded in two species of Naegleria and show that nearly all are expressed in mouse-passaged N. fowleri cells. We then study distribution of the Golgi marker (Ng)CopB by fluorescence in Naegleria gruberi, identifying membranous structures that are disrupted by Brefeldin A treatment, consistent with Golgi localisation. Confocal and immunoelectron microscopy reveals that NgCOPB localises to tubular membranous structures. Our data identify the Golgi organelle for the first time in this major eukaryotic lineage, and provide the rare example of a tubular morphology, representing an important sampling point for the comparative understanding of Golgi organellar diversity.This article has an associated First Person interview with the first author of the paper.

Changes associated with Ebola virus adaptation to novel species.

MOTIVATION: Ebola viruses are not pathogenic but can be adapted to replicate and cause disease in rodents. Here, we used a structural bioinformatics approach to analyze the mutations associated with Ebola virus adaptation to rodents to elucidate the determinants of host-specific Ebola virus pathogenicity. RESULTS: We identified 33 different mutations associated with Ebola virus adaptation to rodents in the proteins GP, NP, L, VP24 and VP35. Only VP24, GP and NP were consistently found mutated in rodent-adapted Ebola virus strains. Fewer than five mutations in these genes seem to be required for the adaptation of Ebola viruses to a new species. The role of mutations in GP and NP is not clear. However, three VP24 mutations located in the protein interface with karyopherin α5 may enable VP24 to inhibit karyopherins and subsequently the host interferon response. Three further VP24 mutations change hydrogen bonding or cause conformational changes. Hence, there is evidence that few mutations including crucial mutations in VP24 enable Ebola virus adaptation to new hosts. Since Reston virus, the only non-human pathogenic Ebolavirus species circulates in pigs in Asia, this raises concerns that few mutations may result in novel human pathogenic Ebolaviruses. CONTACT: m.n.wass@kent.ac.uk , m.michaelis@kent.ac.uk or j.s.rossman@kent.ac.uk. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

Novel intranasal vaccine targeting SARS-CoV-2 receptor binding domain to mucosal microfold cells and adjuvanted with TLR3 agonist Riboxxim™ elicits strong antibody and T-cell responses in mice.

SARS-CoV-2 continues to circulate in the human population necessitating regular booster immunization for its long-term control. Ideally, vaccines should ideally not only protect against symptomatic disease, but also prevent transmission via asymptomatic shedding and cover existing and future variants of the virus. This may ultimately only be possible through induction of potent and long-lasting immune responses in the nasopharyngeal tract, the initial entry site of SARS-CoV-2. To this end, we have designed a vaccine based on recombinantly expressed receptor binding domain (RBD) of SARS-CoV-2, fused to the C-terminus of C. perfringens enterotoxin, which is known to target Claudin-4, a matrix molecule highly expressed on mucosal microfold (M) cells of the nasal and bronchial-associated lymphoid tissues. To further enhance immune responses, the vaccine was adjuvanted with a novel toll-like receptor 3/RIG-I agonist (Riboxxim™), consisting of synthetic short double stranded RNA. Intranasal prime-boost immunization of mice induced robust mucosal and systemic anti-SARS-CoV-2 neutralizing antibody responses against SARS-CoV-2 strains Wuhan-Hu-1, and several variants (B.1.351/beta, B.1.1.7/alpha, B.1.617.2/delta), as well as systemic T-cell responses. A combination vaccine with M-cell targeted recombinant HA1 from an H1N1 G4 influenza strain also induced mucosal and systemic antibodies against influenza. Taken together, the data show that development of an intranasal SARS-CoV-2 vaccine based on recombinant RBD adjuvanted with a TLR3 agonist is feasible, also as a combination vaccine against influenza.

Differential T-cell and antibody responses induced by mRNA versus adenoviral vectored COVID-19 vaccines in patients with immunodeficiencies.

Background: Immunodeficient patients (IDPs) are at higher risk of contracting severe coronavirus disease 2019 (COVID-19). Targeted vaccination strategies have been implemented to enhance vaccine-induced protection. In this population, however, clinical effectiveness is variable and the duration of protection unknown. Objective: We sought to better understand the cellular and humoral immune responses to mRNA and adenoviral vectored COVID-19 vaccines in patients with immunodeficiency. Methods: Immune responses to severe acute respiratory syndrome coronavirus 2 spike were assessed after 2 doses of homologous ChAdOx1-nCoV-19 or BNT162b2 vaccines in 112 infection-naive IDPs and 131 healthy health care workers as controls. Predictors of vaccine responsiveness were investigated. Results: Immune responses to vaccination were low, and virus neutralization by antibody was not detected despite high titer binding responses in many IDPs. In those exhibiting response, the frequency of specific T-cell responses in IDPs was similar to controls, while antibody responses were lower. Sustained vaccine specific differences were identified: T-cell responses were greater in ChAdOx1-nCoV-19- compared to BNT162b2-immunized IDPs, and antibody binding and neutralization were greater in all cohorts immunized with BNT162b2. The positive correlation between T-cell and antibody responses was weak and increased with subsequent vaccination. Conclusion: Immunodeficient patients have impaired immune responses to mRNA and viral vector COVID-19 vaccines that appear to be influenced by vaccine formulation. Understanding the relative roles of T-cell- and antibody-mediated protection as well as the potential of heterologous prime and boost immunization protocols is needed to optimize the vaccination approach in these high-risk groups.

Correlation between pseudotyped virus and authentic virus neutralisation assays, a systematic review and meta-analysis of the literature.

Background: The virus neutralization assay is a principal method to assess the efficacy of antibodies in blocking viral entry. Due to biosafety handling requirements of viruses classified as hazard group 3 or 4, pseudotyped viruses can be used as a safer alternative. However, it is often queried how well the results derived from pseudotyped viruses correlate with authentic virus. This systematic review and meta-analysis was designed to comprehensively evaluate the correlation between the two assays. Methods: Using PubMed and Google Scholar, reports that incorporated neutralisation assays with both pseudotyped virus, authentic virus, and the application of a mathematical formula to assess the relationship between the results, were selected for review. Our searches identified 67 reports, of which 22 underwent a three-level meta-analysis. Results: The three-level meta-analysis revealed a high level of correlation between pseudotyped viruses and authentic viruses when used in an neutralisation assay. Reports that were not included in the meta-analysis also showed a high degree of correlation, with the exception of lentiviral-based pseudotyped Ebola viruses. Conclusion: Pseudotyped viruses identified in this report can be used as a surrogate for authentic virus, though care must be taken in considering which pseudotype core to use when generating new uncharacterised pseudotyped viruses.

Serological evidence of virus infection in Eidolon helvum fruit bats: implications for bushmeat consumption in Nigeria.

Introduction: The Eidolon helvum fruit bat is one of the most widely distributed fruit bats in Africa and known to be a reservoir for several pathogenic viruses that can cause disease in animals and humans. To assess the risk of zoonotic spillover, we conducted a serological survey of 304 serum samples from E. helvum bats that were captured for human consumption in Makurdi, Nigeria. Methods: Using pseudotyped viruses, we screened 304 serum samples for neutralizing antibodies against viruses from the Coronaviridae, Filoviridae, Orthomyxoviridae and Paramyxoviridae families. Results: We report the presence of neutralizing antibodies against henipavirus lineage GH-M74a virus (odds ratio 6.23; p < 0.001), Nipah virus (odds ratio 4.04; p = 0.00031), bat influenza H17N10 virus (odds ratio 7.25; p < 0.001) and no significant association with Ebola virus (odds ratio 0.56; p = 0.375) in this bat cohort. Conclusion: The data suggest a potential risk of zoonotic spillover including the possible circulation of highly pathogenic viruses in E. helvum populations. These findings highlight the importance of maintaining sero-surveillance of E. helvum, and the necessity for further, more comprehensive investigations to monitor changes in virus prevalence, distribution over time, and across different geographic locations.

Glycan masking of a non-neutralising epitope enhances neutralising antibodies targeting the RBD of SARS-CoV-2 and its variants.

The accelerated development of the first generation COVID-19 vaccines has saved millions of lives, and potentially more from the long-term sequelae of SARS-CoV-2 infection. The most successful vaccine candidates have used the full-length SARS-CoV-2 spike protein as an immunogen. As expected of RNA viruses, new variants have evolved and quickly replaced the original wild-type SARS-CoV-2, leading to escape from natural infection or vaccine induced immunity provided by the original SARS-CoV-2 spike sequence. Next generation vaccines that confer specific and targeted immunity to broadly neutralising epitopes on the SARS-CoV-2 spike protein against different variants of concern (VOC) offer an advance on current booster shots of previously used vaccines. Here, we present a targeted approach to elicit antibodies that neutralise both the ancestral SARS-CoV-2, and the VOCs, by introducing a specific glycosylation site on a non-neutralising epitope of the RBD. The addition of a specific glycosylation site in the RBD based vaccine candidate focused the immune response towards other broadly neutralising epitopes on the RBD. We further observed enhanced cross-neutralisation and cross-binding using a DNA-MVA CR19 prime-boost regime, thus demonstrating the superiority of the glycan engineered RBD vaccine candidate across two platforms and a promising candidate as a broad variant booster vaccine.

Evolutionary analysis of cellular reduction and anaerobicity in the hyper-prevalent gut microbe Blastocystis.

Blastocystis is the most prevalent microbial eukaryote in the human and animal gut, yet its role as commensal or parasite is still under debate. Blastocystis has clearly undergone evolutionary adaptation to the gut environment and possesses minimal cellular compartmentalization, reduced anaerobic mitochondria, no flagella, and no reported peroxisomes. To address this poorly understood evolutionary transition, we have taken a multi-disciplinary approach to characterize Proteromonas lacertae, the closest canonical stramenopile relative of Blastocystis. Genomic data reveal an abundance of unique genes in P. lacertae but also reductive evolution of the genomic complement in Blastocystis. Comparative genomic analysis sheds light on flagellar evolution, including 37 new candidate components implicated with mastigonemes, the stramenopile morphological hallmark. The P. lacertae membrane-trafficking system (MTS) complement is only slightly more canonical than that of Blastocystis, but notably, we identified that both organisms encode the complete enigmatic endocytic TSET complex, a first for the entire stramenopile lineage. Investigation also details the modulation of mitochondrial composition and metabolism in both P. lacertae and Blastocystis. Unexpectedly, we identify in P. lacertae the most reduced peroxisome-derived organelle reported to date, which leads us to speculate on a mechanism of constraint guiding the dynamics of peroxisome-mitochondrion reductive evolution on the path to anaerobiosis. Overall, these analyses provide a launching point to investigate organellar evolution and reveal in detail the evolutionary path that Blastocystis has taken from a canonical flagellated protist to the hyper-divergent and hyper-prevalent animal and human gut microbe.

A computationally designed antigen eliciting broad humoral responses against SARS-CoV-2 and related sarbecoviruses.

The threat of spillovers of coronaviruses associated with the severe acute respiratory syndrome (SARS) from animals to humans necessitates vaccines that offer broader protection from sarbecoviruses. By leveraging a viral-genome-informed computational method for selecting immune-optimized and structurally engineered antigens, here we show that a single antigen based on the receptor binding domain of the spike protein of sarbecoviruses elicits broad humoral responses against SARS-CoV-1, SARS-CoV-2, WIV16 and RaTG13 in mice, rabbits and guinea pigs. When administered as a DNA immunogen or by a vector based on a modified vaccinia virus Ankara, the optimized antigen induced vaccine protection from the Delta variant of SARS-CoV-2 in mice genetically engineered to express angiotensin-converting enzyme 2 and primed by a viral-vector vaccine (AZD1222) against SARS-CoV-2. A vaccine formulation incorporating mRNA coding for the optimized antigen further validated its broad immunogenicity. Vaccines that elicit broad immune responses across subgroups of coronaviruses may counteract the threat of zoonotic spillovers of betacoronaviruses.

Analysis of Antibody Neutralisation Activity against SARS-CoV-2 Variants and Seasonal Human Coronaviruses NL63, HKU1, and 229E Induced by Three Different COVID-19 Vaccine Platforms.

Coronaviruses infections, culminating in the recent severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic beginning in 2019, have highlighted the importance of effective vaccines to induce an antibody response with cross-neutralizing activity. COVID-19 vaccines have been rapidly developed to reduce the burden of SARS-CoV-2 infections and disease severity. Cross-protection from seasonal human coronaviruses (hCoVs) infections has been hypothesized but is still controversial. Here, we investigated the neutralizing activity against ancestral SARS-CoV-2 and the variants of concern (VOCs) in individuals vaccinated with two doses of either BNT162b2, mRNA-1273, or AZD1222, with or without a history of SARS-CoV-2 infection. Antibody neutralizing activity to SARS-CoV-2 and the VOCs was higher in BNT162b2-vaccinated subjects who were previously infected with SARS-CoV-2 and conferred broad-spectrum protection. The Omicron BA.1 variant was the most resistant among the VOCs. COVID-19 vaccination did not confer protection against hCoV-HKU1. Conversely, antibodies induced by mRNA-1273 vaccination displayed a boosting in their neutralizing activity against hCoV-NL63, whereas AZD1222 vaccination increased antibody neutralization against hCoV-229E, suggesting potential differences in antigenicity and immunogenicity of the different spike constructs used between various vaccination platforms. These data would suggest that there may be shared epitopes between the HCoVs and SARS-CoV-2 spike proteins.

Pseudotyped Bat Coronavirus RaTG13 is efficiently neutralised by convalescent sera from SARS-CoV-2 infected patients.

RaTG13 is a close relative of SARS-CoV-2, the virus responsible for the COVID-19 pandemic, sharing 96% sequence similarity at the genome-wide level. The spike receptor binding domain (RBD) of RaTG13 contains a number of amino acid substitutions when compared to SARS-CoV-2, likely impacting affinity for the ACE2 receptor. Antigenic differences between the viruses are less well understood, especially whether RaTG13 spike can be efficiently neutralised by antibodies generated from infection with, or vaccination against, SARS-CoV-2. Using RaTG13 and SARS-CoV-2 pseudotypes we compared neutralisation using convalescent sera from previously infected patients or vaccinated healthcare workers. Surprisingly, our results revealed that RaTG13 was more efficiently neutralised than SARS-CoV-2. In addition, neutralisation assays using spike mutants harbouring single and combinatorial amino acid substitutions within the RBD demonstrated that both spike proteins can tolerate multiple changes without dramatically reducing neutralisation. Moreover, introducing the 484 K mutation into RaTG13 resulted in increased neutralisation, in contrast to the same mutation in SARS-CoV-2 (E484K). This is despite E484K having a well-documented role in immune evasion in variants of concern (VOC) such as B.1.351 (Beta). These results indicate that the future spill-over of RaTG13 and/or related sarbecoviruses could be mitigated using current SARS-CoV-2-based vaccination strategies.

Durable T-cellular and humoral responses in SARS-CoV-2 hospitalized and community patients.

BACKGROUND: Neutralizing antibodies are important for protection against the pandemic SARS-CoV-2 virus, and long-term memory responses determine the risk of re-infection or boosting after vaccination. T-cellular responses are considered important for partial protection against novel variants of concern. METHODS: A prospective cohort of hospitalized (n = 14) and community (n = 38) patients with rt-PCR confirmed SARS-CoV-2 infection were recruited. Blood samples and clinical data were collected when diagnosed and at 6 months. Serum samples were analyzed for SARS-CoV-2-spike specific antibodies using ELISA (IgG, IgA, IgM), pseudotype neutralization and microneutralization assays. Peripheral blood mononuclear cells were investigated for virus-specific T-cell responses in the interferon-γ and interleukin-2 fluorescent-linked immunosorbent spot (FluroSpot) assay. RESULTS: We found durable SARS-CoV-2 spike- and internal protein specific T-cellular responses in patients with persistent antibodies at 6 months. Significantly higher IL-2 and IFN-γ secreting T-cell responses as well as SARS-CoV-2 specific IgG and neutralizing antibodies were detected in hospitalized compared to community patients. The immune response was impacted by age, gender, comorbidity and severity of illness, reflecting clinical observations. CONCLUSIONS: SARS-CoV-2 specific T-cellular and antibody responses persisted for 6 months post confirmed infection. In previously infected patients, re-exposure or vaccination will boost long-term immunity, possibly providing protection against re-infection with variant viruses.

Neutralisation Hierarchy of SARS-CoV-2 Variants of Concern Using Standardised, Quantitative Neutralisation Assays Reveals a Correlation With Disease Severity; Towards Deciphering Protective Antibody Thresholds.

The rise of SARS-CoV-2 variants has made the pursuit to define correlates of protection more troublesome, despite the availability of the World Health Organisation (WHO) International Standard for anti-SARS-CoV-2 Immunoglobulin sera, a key reagent used to standardise laboratory findings into an international unitage. Using pseudotyped virus, we examine the capacity of convalescent sera, from a well-defined cohort of healthcare workers (HCW) and Patients infected during the first wave from a national critical care centre in the UK to neutralise B.1.1.298, variants of interest (VOI) B.1.617.1 (Kappa), and four VOCs, B.1.1.7 (Alpha), B.1.351 (Beta), P.1 (Gamma) and B.1.617.2 (Delta), including the B.1.617.2 K417N, informally known as Delta Plus. We utilised the WHO International Standard for anti-SARS-CoV-2 Immunoglobulin to report neutralisation antibody levels in International Units per mL. Our data demonstrate a significant reduction in the ability of first wave convalescent sera to neutralise the VOCs. Patients and HCWs with more severe COVID-19 were found to have higher antibody titres and to neutralise the VOCs more effectively than individuals with milder symptoms. Using an estimated threshold for 50% protection, 54 IU/mL, we found most asymptomatic and mild cases did not produce titres above this threshold.

Persistent but dysfunctional mucosal SARS-CoV-2-specific IgA and low lung IL-1ß associate with COVID-19 fatal outcome: A cross-sectional analysis.

The role of the mucosal pulmonary antibody response in coronavirus disease 2019 (COVID-19) outcome remains unclear. Here, we found that in bronchoalveolar lavage (BAL) samples from 48 patients with severe COVID-19-infected with the ancestral Wuhan virus, mucosal IgG and IgA specific for S1, receptor-binding domain (RBD), S2, and nucleocapsid protein (NP) emerged in BAL containing viruses early in infection and persist after virus elimination, with more IgA than IgG for all antigens tested. Furthermore, spike-IgA and spike-IgG immune complexes were detected in BAL, especially when the lung virus has been cleared. BAL IgG and IgA recognized the four main RBD variants. BAL neutralizing titers were higher early in COVID-19 when virus replicates in the lung than later in infection after viral clearance. Patients with fatal COVID-19, in contrast to survivors, developed higher levels of mucosal spike-specific IgA than IgG but lost neutralizing activities over time and had reduced IL-1ß in the lung. Altogether, mucosal spike and NP-specific IgG and S1-specific IgA persisting after lung severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) clearance and low pulmonary IL-1ß correlate with COVID-19 fatal outcome. Thus, mucosal SARS-CoV-2-specific antibodies may have adverse functions in addition to protective neutralization. Highlights: Mucosal pulmonary antibody response in COVID-19 outcome remains unclear. We show that in severe COVID-19 patients, mucosal pulmonary non-neutralizing SARS-CoV-2 IgA persit after viral clearance in the lung. Furthermore, low lung IL-1ß correlate with fatal COVID-19. Altogether, mucosal IgA may exert harmful functions beside protective neutralization.