A potent pan-sarbecovirus neutralizing antibody resilient to epitope diversification.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) evolution has resulted in viral escape from clinically authorized monoclonal antibodies (mAbs), creating a need for mAbs that are resilient to epitope diversification. Broadly neutralizing coronavirus mAbs that are sufficiently potent for clinical development and retain activity despite viral evolution remain elusive. We identified a human mAb, designated VIR-7229, which targets the viral receptor-binding motif (RBM) with unprecedented cross-reactivity to all sarbecovirus clades, including non-ACE2-utilizing bat sarbecoviruses, while potently neutralizing SARS-CoV-2 variants since 2019, including the recent EG.5, BA.2.86, and JN.1. VIR-7229 tolerates extraordinary epitope variability, partly attributed to its high binding affinity, receptor molecular mimicry, and interactions with RBM backbone atoms. Consequently, VIR-7229 features a high barrier for selection of escape mutants, which are rare and associated with reduced viral fitness, underscoring its potential to be resilient to future viral evolution. VIR-7229 is a strong candidate to become a next-generation medicine.

N-terminal domain antigenic mapping reveals a site of vulnerability for SARS-CoV-2.

The SARS-CoV-2 spike (S) glycoprotein contains an immunodominant receptor-binding domain (RBD) targeted by most neutralizing antibodies (Abs) in COVID-19 patient plasma. Little is known about neutralizing Abs binding to epitopes outside the RBD and their contribution to protection. Here, we describe 41 human monoclonal Abs (mAbs) derived from memory B cells, which recognize the SARS-CoV-2 S N-terminal domain (NTD) and show that a subset of them neutralize SARS-CoV-2 ultrapotently. We define an antigenic map of the SARS-CoV-2 NTD and identify a supersite (designated site i) recognized by all known NTD-specific neutralizing mAbs. These mAbs inhibit cell-to-cell fusion, activate effector functions, and protect Syrian hamsters from SARS-CoV-2 challenge, albeit selecting escape mutants in some animals. Indeed, several SARS-CoV-2 variants, including the B.1.1.7, B.1.351, and P.1 lineages, harbor frequent mutations within the NTD supersite, suggesting ongoing selective pressure and the importance of NTD-specific neutralizing mAbs for protective immunity and vaccine design.

Circulating SARS-CoV-2 spike N439K variants maintain fitness while evading antibody-mediated immunity.

SARS-CoV-2 can mutate and evade immunity, with consequences for efficacy of emerging vaccines and antibody therapeutics. Here, we demonstrate that the immunodominant SARS-CoV-2 spike (S) receptor binding motif (RBM) is a highly variable region of S and provide epidemiological, clinical, and molecular characterization of a prevalent, sentinel RBM mutation, N439K. We demonstrate N439K S protein has enhanced binding affinity to the hACE2 receptor, and N439K viruses have similar in vitro replication fitness and cause infections with similar clinical outcomes as compared to wild type. We show the N439K mutation confers resistance against several neutralizing monoclonal antibodies, including one authorized for emergency use by the US Food and Drug Administration (FDA), and reduces the activity of some polyclonal sera from persons recovered from infection. Immune evasion mutations that maintain virulence and fitness such as N439K can emerge within SARS-CoV-2 S, highlighting the need for ongoing molecular surveillance to guide development and usage of vaccines and therapeutics.

Mapping Neutralizing and Immunodominant Sites on the SARS-CoV-2 Spike Receptor-Binding Domain by Structure-Guided High-Resolution Serology.

Analysis of the specificity and kinetics of neutralizing antibodies (nAbs) elicited by SARS-CoV-2 infection is crucial for understanding immune protection and identifying targets for vaccine design. In a cohort of 647 SARS-CoV-2-infected subjects, we found that both the magnitude of Ab responses to SARS-CoV-2 spike (S) and nucleoprotein and nAb titers correlate with clinical scores. The receptor-binding domain (RBD) is immunodominant and the target of 90% of the neutralizing activity present in SARS-CoV-2 immune sera. Whereas overall RBD-specific serum IgG titers waned with a half-life of 49 days, nAb titers and avidity increased over time for some individuals, consistent with affinity maturation. We structurally defined an RBD antigenic map and serologically quantified serum Abs specific for distinct RBD epitopes leading to the identification of two major receptor-binding motif antigenic sites. Our results explain the immunodominance of the receptor-binding motif and will guide the design of COVID-19 vaccines and therapeutics.

Clonal structure, stability and dynamics of human memory B cells and circulating plasmablasts.

Memory B cells persist for a lifetime and rapidly differentiate into antibody-producing plasmablasts and plasma cells upon antigen re-encounter. The clonal relationship and evolution of memory B cells and circulating plasmablasts is not well understood. Using single-cell sequencing combined with isolation of specific antibodies, we found that in two healthy donors, the memory B cell repertoire was dominated by large IgM, IgA and IgG2 clonal families, whereas IgG1 families, including those specific for recall antigens, were of small size. Analysis of multiyear samples demonstrated stability of memory B cell clonal families and revealed that a large fraction of recently generated plasmablasts was derived from long-term memory B cell families and was found recurrently. Collectively, this study provides a systematic description of the structure, stability and dynamics of the human memory B cell pool and suggests that memory B cells may be active at any time point in the generation of plasmablasts.

Neutralization, effector function and immune imprinting of Omicron variants.

Currently circulating SARS-CoV-2 variants have acquired convergent mutations at hot spots in the receptor-binding domain1 (RBD) of the spike protein. The effects of these mutations on viral infection and transmission and the efficacy of vaccines and therapies remains poorly understood. Here we demonstrate that recently emerged BQ.1.1 and XBB.1.5 variants bind host ACE2 with high affinity and promote membrane fusion more efficiently than earlier Omicron variants. Structures of the BQ.1.1, XBB.1 and BN.1 RBDs bound to the fragment antigen-binding region of the S309 antibody (the parent antibody for sotrovimab) and human ACE2 explain the preservation of antibody binding through conformational selection, altered ACE2 recognition and immune evasion. We show that sotrovimab binds avidly to all Omicron variants, promotes Fc-dependent effector functions and protects mice challenged with BQ.1.1 and hamsters challenged with XBB.1.5. Vaccine-elicited human plasma antibodies cross-react with and trigger effector functions against current Omicron variants, despite a reduced neutralizing activity, suggesting a mechanism of protection against disease, exemplified by S309. Cross-reactive RBD-directed human memory B cells remained dominant even after two exposures to Omicron spikes, underscoring the role of persistent immune imprinting.

Broadly neutralizing antibodies overcome SARS-CoV-2 Omicron antigenic shift.

The recently emerged SARS-CoV-2 Omicron variant encodes 37 amino acid substitutions in the spike protein, 15 of which are in the receptor-binding domain (RBD), thereby raising concerns about the effectiveness of available vaccines and antibody-based therapeutics. Here we show that the Omicron RBD binds to human ACE2 with enhanced affinity, relative to the Wuhan-Hu-1 RBD, and binds to mouse ACE2. Marked reductions in neutralizing activity were observed against Omicron compared to the ancestral pseudovirus in plasma from convalescent individuals and from individuals who had been vaccinated against SARS-CoV-2, but this loss was less pronounced after a third dose of vaccine. Most monoclonal antibodies that are directed against the receptor-binding motif lost in vitro neutralizing activity against Omicron, with only 3 out of 29 monoclonal antibodies retaining unaltered potency, including the ACE2-mimicking S2K146 antibody1. Furthermore, a fraction of broadly neutralizing sarbecovirus monoclonal antibodies neutralized Omicron through recognition of antigenic sites outside the receptor-binding motif, including sotrovimab2, S2X2593 and S2H974. The magnitude of Omicron-mediated immune evasion marks a major antigenic shift in SARS-CoV-2. Broadly neutralizing monoclonal antibodies that recognize RBD epitopes that are conserved among SARS-CoV-2 variants and other sarbecoviruses may prove key to controlling the ongoing pandemic and future zoonotic spillovers.

Sensitivity of SARS-CoV-2 B.1.1.7 to mRNA vaccine-elicited antibodies.

Transmission of SARS-CoV-2 is uncontrolled in many parts of the world; control is compounded in some areas by the higher transmission potential of the B.1.1.7 variant1, which has now been reported in 94 countries. It is unclear whether the response of the virus to vaccines against SARS-CoV-2 on the basis of the prototypic strain will be affected by the mutations found in B.1.1.7. Here we assess the immune responses of individuals after vaccination with the mRNA-based vaccine BNT162b22. We measured neutralizing antibody responses after the first and second immunizations using pseudoviruses that expressed the wild-type spike protein or a mutated spike protein that contained the eight amino acid changes found in the B.1.1.7 variant. The sera from individuals who received the vaccine exhibited a broad range of neutralizing titres against the wild-type pseudoviruses that were modestly reduced against the B.1.1.7 variant. This reduction was also evident in sera from some patients who had recovered from COVID-19. Decreased neutralization of the B.1.1.7 variant was also observed for monoclonal antibodies that target the N-terminal domain (9 out of 10) and the receptor-binding motif (5 out of 31), but not for monoclonal antibodies that recognize the receptor-binding domain that bind outside the receptor-binding motif. Introduction of the mutation that encodes the E484K substitution in the B.1.1.7 background to reflect a newly emerged variant of concern (VOC 202102/02) led to a more-substantial loss of neutralizing activity by vaccine-elicited antibodies and monoclonal antibodies (19 out of 31) compared with the loss of neutralizing activity conferred by the mutations in B.1.1.7 alone. The emergence of the E484K substitution in a B.1.1.7 background represents a threat to the efficacy of the BNT162b2 vaccine.

SARS-CoV-2 RBD antibodies that maximize breadth and resistance to escape.

An ideal therapeutic anti-SARS-CoV-2 antibody would resist viral escape1-3, have activity against diverse sarbecoviruses4-7, and be highly protective through viral neutralization8-11 and effector functions12,13. Understanding how these properties relate to each other and vary across epitopes would aid the development of therapeutic antibodies and guide vaccine design. Here we comprehensively characterize escape, breadth and potency across a panel of SARS-CoV-2 antibodies targeting the receptor-binding domain (RBD). Despite a trade-off between in vitro neutralization potency and breadth of sarbecovirus binding, we identify neutralizing antibodies with exceptional sarbecovirus breadth and a corresponding resistance to SARS-CoV-2 escape. One of these antibodies, S2H97, binds with high affinity across all sarbecovirus clades to a cryptic epitope and prophylactically protects hamsters from viral challenge. Antibodies that target the angiotensin-converting enzyme 2 (ACE2) receptor-binding motif (RBM) typically have poor breadth and are readily escaped by mutations despite high neutralization potency. Nevertheless, we also characterize a potent RBM antibody (S2E128) with breadth across sarbecoviruses related to SARS-CoV-2 and a high barrier to viral escape. These data highlight principles underlying variation in escape, breadth and potency among antibodies that target the RBD, and identify epitopes and features to prioritize for therapeutic development against the current and potential future pandemics.

Broad sarbecovirus neutralization by a human monoclonal antibody.

The recent emergence of SARS-CoV-2 variants of concern1-10 and the recurrent spillovers of coronaviruses11,12 into the human population highlight the need for broadly neutralizing antibodies that are not affected by the ongoing antigenic drift and that can prevent or treat future zoonotic infections. Here we describe a human monoclonal antibody designated S2X259, which recognizes a highly conserved cryptic epitope of the receptor-binding domain and cross-reacts with spikes from all clades of sarbecovirus. S2X259 broadly neutralizes spike-mediated cell entry of SARS-CoV-2, including variants of concern (B.1.1.7, B.1.351, P.1, and B.1.427/B.1.429), as well as a wide spectrum of human and potentially zoonotic sarbecoviruses through inhibition of angiotensin-converting enzyme 2 (ACE2) binding to the receptor-binding domain. Furthermore, deep-mutational scanning and in vitro escape selection experiments demonstrate that S2X259 possesses an escape profile that is limited to a single substitution, G504D. We show that prophylactic and therapeutic administration of S2X259 protects Syrian hamsters (Mesocricetus auratus) against challenge with the prototypic SARS-CoV-2 and the B.1.351 variant of concern, which suggests that this monoclonal antibody is a promising candidate for the prevention and treatment of emergent variants and zoonotic infections. Our data reveal a key antigenic site that is targeted by broadly neutralizing antibodies and will guide the design of vaccines that are effective against all sarbecoviruses.

SARS-CoV-2 B.1.617.2 Delta variant replication and immune evasion.

The B.1.617.2 (Delta) variant of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was first identified in the state of Maharashtra in late 2020 and spread throughout India, outcompeting pre-existing lineages including B.1.617.1 (Kappa) and B.1.1.7 (Alpha)1. In vitro, B.1.617.2 is sixfold less sensitive to serum neutralizing antibodies from recovered individuals, and eightfold less sensitive to vaccine-elicited antibodies, compared with wild-type Wuhan-1 bearing D614G. Serum neutralizing titres against B.1.617.2 were lower in ChAdOx1 vaccinees than in BNT162b2 vaccinees. B.1.617.2 spike pseudotyped viruses exhibited compromised sensitivity to monoclonal antibodies to the receptor-binding domain and the amino-terminal domain. B.1.617.2 demonstrated higher replication efficiency than B.1.1.7 in both airway organoid and human airway epithelial systems, associated with B.1.617.2 spike being in a predominantly cleaved state compared with B.1.1.7 spike. The B.1.617.2 spike protein was able to mediate highly efficient syncytium formation that was less sensitive to inhibition by neutralizing antibody, compared with that of wild-type spike. We also observed that B.1.617.2 had higher replication and spike-mediated entry than B.1.617.1, potentially explaining the B.1.617.2 dominance. In an analysis of more than 130 SARS-CoV-2-infected health care workers across three centres in India during a period of mixed lineage circulation, we observed reduced ChAdOx1 vaccine effectiveness against B.1.617.2 relative to non-B.1.617.2, with the caveat of possible residual confounding. Compromised vaccine efficacy against the highly fit and immune-evasive B.1.617.2 Delta variant warrants continued infection control measures in the post-vaccination era.

Cross-neutralization of SARS-CoV-2 by a human monoclonal SARS-CoV antibody.

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is a newly emerged coronavirus that is responsible for the current pandemic of coronavirus disease 2019 (COVID-19), which has resulted in more than 3.7 million infections and 260,000 deaths as of 6 May 20201,2. Vaccine and therapeutic discovery efforts are paramount to curb the pandemic spread of this zoonotic virus. The SARS-CoV-2 spike (S) glycoprotein promotes entry into host cells and is the main target of neutralizing antibodies. Here we describe several monoclonal antibodies that target the S glycoprotein of SARS-CoV-2, which we identified from memory B cells of an individual who was infected with severe acute respiratory syndrome coronavirus (SARS-CoV) in 2003. One antibody (named S309) potently neutralizes SARS-CoV-2 and SARS-CoV pseudoviruses as well as authentic SARS-CoV-2, by engaging the receptor-binding domain of the S glycoprotein. Using cryo-electron microscopy and binding assays, we show that S309 recognizes an epitope containing a glycan that is conserved within the Sarbecovirus subgenus, without competing with receptor attachment. Antibody cocktails that include S309 in combination with other antibodies that we identified further enhanced SARS-CoV-2 neutralization, and may limit the emergence of neutralization-escape mutants. These results pave the way for using S309 and antibody cocktails containing S309 for prophylaxis in individuals at a high risk of exposure or as a post-exposure therapy to limit or treat severe disease.

Overview of Mobile Communications in Colombia and Introduction to 5G.

The deployment of 5G around the world continues to progress at a rapid pace, especially in North America and Asia. Its advantages and efficiency as a data transmission network have been widely demonstrated in different fields such as agriculture, education, health, and surveillance. However, this process does not have the same dynamics in Latin America, specifically in Colombia. The country is currently implementing actions aimed at facilitating the deployment of this technology in the short term, including pilot tests for the use of the radio spectrum, spectrum auctions, the planning of future auctions, and the review of spectrum caps. The results of this review allow us to conclude that despite the forecasts and the intentions of the Colombian government and mobile communication service operators, 5G in standalone mode will not be commercially available in Colombia before the end of 2023. The main failures in its deployment are related to the lack of available spectrum to support the ultrahigh-reliability and low-latency, enhanced mobile broadband, and massive machine-type communications scenarios, as well as the delay in the auction processes for its assignment.

Precision Agriculture and Sensor Systems Applications in Colombia through 5G Networks.

The growing global demand for food and the environmental impact caused by agriculture have made this activity increasingly dependent on electronics, information technology, and telecommunications technologies. In Colombia, agriculture is of great importance not only as a commercial activity, but also as a source of food and employment. However, the concept of smart agriculture has not been widely applied in this country, resulting in the high production of various types of crops due to the planting of large areas of land, rather than optimization of the processes involved in the activity. Due to its technical characteristics and the radio spectrum considered in its deployment, 5G can be seen as one of the technologies that could generate the greatest benefits for the Colombian agricultural sector, especially in the most remote rural areas, which currently lack mobile network coverage. This article provides an overview of the current 5G technology landscape in Colombia and presents examples of possible 5G/IoT applications that could be developed in Colombian fields. The results show that 5G could facilitate the implementation of the smart farm in Colombia, improving current production and efficiency. It is useful when designing 5G implementation plans and strategies, since it categorizes crops by regions and products. This is based on budget availability, population density, and regional development plans, among others.

Projections of IoT Applications in Colombia Using 5G Wireless Networks.

Wireless technologies are increasingly relevant in different activities and lines of the economy, as well as in the daily life of people and companies. The advent of fifth generation networks (5G) implies a promising synergy with the Internet of Things (IoT), allowing for more automations in production processes and an increase in the efficiency of information transmission, managing to improve the efficiency in decision-making through tools such as big data and artificial intelligence. This article presents a description of the 5G implementation process in Colombia, as well as a revision of opportunities when combining with IoT in featured sectors of the departmental development plans, such as agriculture, tourism, health, the environment, and industry. Results shows that the startup of 5G in Colombia has been a slow process, but there are comparisons with similar procedures in other developed countries. Additionally, we present examples of 5G and IoT applications which can be promoted in Colombia, aimed at improving the quality of life of their habitants and promoting economic development.

Empirical Model of Radio Wave Propagation in the Presence of Vegetation inside Greenhouses Using Regularized Regressions.

Spain is Europe's leading exporter of tomatoes harvested in greenhouses. The production of tomatoes should be kept and increased, supported by precision agriculture to meet food and commercial demand. The wireless sensor network (WSN) has demonstrated to be a tool to provide farmers with useful information on the state of their plantations due to its practical deployment. However, in order to measure its deployment within a crop, it is necessary to know the communication coverage of the nodes that make up the network. The multipath propagation of radio waves between the transceivers of the WSN nodes inside a greenhouse is degraded and attenuated by the intricate complex of stems, branches, leaf twigs, and fruits, all randomly oriented, that block the line of sight, consequently generating a signal power loss as the distance increases. Although the COST235 (European Cooperation in Science and Technology - COST), ITU-R (International Telecommunications Union-Radiocommunication Sector), FITU-R (Fitted ITU-R), and Weisbberger models provide an explanation of the radio wave propagation in the presence of vegetation in the 2.4 GHz ICM band, some significant discrepancies were found when they are applied to field tests with tomato greenhouses. In this paper, a novel method is proposed for determining an empirical model of radio wave attenuation for vegetation in the 2.4 GHz band, which includes the vegetation height as a parameter in addition to the distance between transceivers of WNS nodes. The empirical attenuation model was obtained applying regularized regressions with a multiparametric equation using experimental signal RSSI measurements achieved by our own RSSI measurement system for our field tests in four plantations. The evaluation parameters gave 0.948 for R2, 0.946 for R2 Adj considering 5th grade polynomial (20 parameters), and 0.942 for R2, and 0.940 for R2 Adj when a reduction of parameters was applied using the cross validation (15 parameters). These results verify the rationality and reliability of the empirical model. Finally, the model was validated considering experimental data from other plantations, reaching similar results to our proposed model.

CXCR7 influences the migration of B cells during maturation.

The atypical chemokine receptor CXCR7 binds the chemokines CXCL12 and CXCL11. The receptor is widely expressed and was shown to tune CXCR12-induced responses of CXCR4. Here, the function of CXCR7 was examined at late stages of human B-cell maturation, when B cells differentiate into Ab-secreting plasmablasts. We identified two populations of CXCR7(+) cells in tonsillar lymphocytes, one being presumably memory B cells or early plasmablasts (FSC(low) CD19(+) CD38(mid) ) and the other being plasmablasts or early plasma cells (FSC(high) CD19(+) CD38(+) ). CXCR7 is expressed on CD19(+) CD27(+) memory B cells, on CD19(+) CD38(+) CD138(-) and intracellular immunoglobulin high plasmablasts, but not on CD19(+) CD138(+) icIg(high) plasma cells. The differential expression pattern suggests a potential contribution of the scavenger receptor in final B-cell maturation. On in vitro differentiating B cells, we found a marked inverse correlation between CXCR7 and CXCR5 cell surface levels, whereas expression of CXCR4 remained almost constant. Migration assays performed with tonsillar mononuclear cells or in vitro differentiated cells revealed that inhibition of CXCR7 markedly increases chemotaxis toward CXCL12, especially at late stages of B-cell maturation. Chemotaxis was attenuated in the presence of CXCR4 antagonists, confirming that migration is CXCR4 mediated. Our findings unequivocally demonstrate a novel role for CXCR7 in regulating the migration of plasmablasts during B-cell maturation.

A functional BCR in human IgA and IgM plasma cells.

Plasma cells (PCs) are terminally differentiated cells of the B-cell lineage that secrete antibodies at a high rate and are thought to lack the expression of the B-cell receptor (BCR). Here, we report that human IgA and IgM, unlike IgG, PCs express a membrane functional BCR associated with the Igα/Igß heterodimer. BCR cross-linking on IgA and IgM PCs led to Ca(2+) mobilization and extracellular signal-regulated kinase 1/2 and AKT phosphorylation and impacted survival of IgA PCs. These findings demonstrate a significant difference between human IgG, IgM, and IgA PCs and suggest that the IgA PC repertoire may be modulated by specific antigens with implications for the regulation of the mucosal immune system.

Dissecting the human immunologic memory for pathogens.

Studies on immunologic memory in animal models and especially in the human system are instrumental to identify mechanisms and correlates of protection necessary for vaccine development. In this article, we provide an overview of the cellular basis of immunologic memory. We also describe experimental approaches based on high throughput cell cultures, which we have developed to interrogate human memory T cells, B cells, and plasma cells. We discuss how these approaches can provide new tools and information for vaccine design, in a process that we define as 'analytic vaccinology'.