Hemagglutinin Research Articles

Molecular epidemiology and vaccine compatibility analysis of seasonal influenza A viruses in the context of COVID-19 epidemic in Wuhan, China.

The COVID-19 pandemic had a significant impact on the global influenza vaccination and the epidemics of seasonal influenza. To further explore the molecular epidemiology of influenza viruses and assess vaccine effectiveness, we collected influenza cases in Wuhan during the 2022-2023 influenza season. Among 1312 clinical samples, 312 samples tested positive for influenza viruses using reverse transcription polymerase chain reaction. These positive samples included 146A/H1N1 subtypes (46.8%), 164A/H3N2 subtypes (52.6%) and 2 influenza B virus types (0.6%). Based on the whole genome sequence information of hemagglutinin (HA) and neuraminidase (NA) from 27A/H1N1 influenza virus strains and 26A/H3N2 influenza virus strains obtained in this study, a phylogenetic analysis was conducted. The analysis revealed that all A/H1N1 strains belonged to the evolutionary branch 6B.1A.5a.2a, and they exhibited specific substitutions at positions K71Q, Q206E, E241A, and R276K. Similarly, all A/H3N2 strains were classified into the 3C.2a1b.2a.1a subclade and displayed amino acid substitutions at positions S172H, N175Y, I176T, K187N, and S214P. Notably, the A/H3N2 strains also acquired a new potential glycosylation site at position N174. Using an epitope model, the predicted vaccine effectiveness was assessed for the A/H1N1 and A/H3N2 strains. The predicted vaccine effectiveness against the Wuhan influenza epidemic strain was over 85% for the A/H1N1 vaccine strain. However, the effectiveness against the A/H3N2 vaccine strain was only 48.7%. To further verify the protection of influenza vaccine against circulating influenza viruses in the region, we conducted in vivo and in vitro animal studies. The results of in vitro neutralization experiment showed that rabbit serum antibodies inoculated with quadrivalent isolated influenza vaccine had neutralization ability against all 24 isolated influenza viruses. In vivo experiments showed that vaccinated mice had fewer lung lesions when infected with the influenza strain circulating in Wuhan, suggesting that vaccination can effectively reduce the occurrence of severe lung damage. These findings emphasize the importance of accurately predicting seasonal influenza strains for effective influenza prevention and control, especially during the co-circulation of SARS-CoV-2 and influenza viruses. This study provides valuable information on the seasonal influenza virus in Wuhan during the COVID-19 pandemic and serves as a basis for vaccine prediction and updates.

Engineering a cleaved, prefusion-stabilized influenza B virus hemagglutinin by identification and locking of all six pH switches.

Vaccine components based on viral fusion proteins require high stability of the native prefusion conformation for optimal potency and manufacturability. In the case of influenza B virus hemagglutinin (HA), the stem's conformation relies on efficient cleavage. In this study, we identified six pH-sensitive regions distributed across the entire ectodomain where protonated histidines assume either a repulsive or an attractive role. Substitutions in these areas enhanced the protein's expression, quality, and stability in its prefusion trimeric state. Importantly, this stabilization enabled the production of a cleavable HA0, which is further processed into HA1 and HA2 by furin during exocytic pathway passage, thereby facilitating correct folding, increased stability, and screening for additional stabilizing substitutions in the core of the metastable fusion domain. Cryo-EM analysis at neutral and low pH revealed a previously unnoticed pH switch involving the C-terminal residues of the natively cleaved HA1. This switch keeps the fusion peptide in a clamped state at neutral pH, averting premature conformational shift. Our findings shed light on new strategies for possible improvements of recombinant or genetic-based influenza B vaccines.

A comprehensive biochemical and epidemiological review of avian influenza: Mechanisms, pharmacology and emerging trends in treatment

Avian influenza viruses, particularly H5N1, H7N9, and H9N2, pose significant threats to avian and human populations through zoonotic transmission, as they have potential to change their genetic material through mutations. Symptoms of Avian Influenza ranges from mild to severe, mainly respiratory problems in both and muscle aches, fatigue in humans. This review highlights the molecular mechanisms by which the virus infects host cells, emphasizing the roles of hemagglutinin (HA) and neuraminidase (NA) in viral entry and release. Biochemical processes involved in viral replication, immune responses, and cytokine production are discussed, with a detailed examination of how antiviral drugs like neuraminidase and polymerase inhibitors disrupt these processes. Outbreak of AIV can cause mass culling results in massive economic loss, trade disruptions, and consumer reductions. The review also addresses the challenges posed by antiviral resistance and outlines novel therapeutic strategies, including combination therapies, vaccine advancements, and host-directed treatments. With an interdisciplinary “One Health” approach, this paper underscores the need for enhanced biosecurity, international cooperation, and continued research to mitigate the global impact of avian influenza.

Discriminating North American Swine Influenza Viruses with a Portable, One-Step, Triplex Real-Time RT-PCR Assay, and Portable Sequencing.

Swine harbors a genetically diverse population of swine influenza A viruses (IAV-S), with demonstrated potential to transmit to the human population, causing outbreaks and pandemics. Here, we describe the development of a one-step, triplex real-time reverse transcription-polymerase chain reaction (rRT-PCR) assay that detects and distinguishes the majority of the antigenically distinct influenza A virus hemagglutinin (HA) clades currently circulating in North American swine, including the IAV-S H1 1A.1 (α), 1A.2 (β), 1A.3 (γ), 1B.2.2 (δ1) and 1B.2.1 (δ2) clades, and the IAV-S H3 2010.1 clade. We performed an in-field test at an exhibition swine show using in-field viral concentration and RNA extraction methodologies and a portable real-time PCR instrument, and rapidly identified three distinct IAV-S clades circulating within the N.A. swine population. Portable sequencing is used to further confirm the results of the in-field test of the swine triplex assay. The IAV-S triplex rRT-PCR assay can be easily transported and used in-field to characterize circulating IAV-S clades in North America, allowing for surveillance and early detection of North American IAV-S with human outbreak and pandemic potential.

Broadly cross-reactive immune responses in chickens immunized with chimeric virus-like particles of nodavirus displaying the M2e originated from avian and human influenza A viruses

Avian influenza A viruses (IAVs) pose a persistent threat to poultry industry worldwide, despite the presence of vaccines. Additionally, reverse-zoonosis transmission potentially introduces human-originated IAVs into poultry and complicates the efforts to control the spread of influenza. Current avian influenza vaccines are primarily based upon the rapidly mutating hemagglutinin (HA) and neuraminidase (NA) glycoproteins, which limit their efficacy against diverse strains of IAVs. Hence, the highly conserved ectodomains of matrix 2 protein (M2e) of IAVs are widely studied as alternatives to the HA and NA. However, the differences in the M2e amino acid sequences between avian and human IAVs generate antibodies that do not cross-react reciprocally with IAVs from other origins. To broaden and enhance the immunogenicity of M2e, we fused two copies each of the M2e derived from avian and human IAVs at the C-terminal end of the Macrobrachium rosenbergii nodavirus (MrNV) capsid protein (NvC). Transmission electron microscopic and dynamic light scattering analyses revealed that the chimeric protein self-assembled into virus-like particles (VLPs). Immunization of chickens with the chimeric VLPs demonstrated a robust induction of broadly reactive immune responses against both the M2e of avian and human IAVs. Additionally, the chimeric VLPs elicited the production of cytotoxic T lymphocytes (CTL), macrophages, as well as a well-balanced Th1 and Th2 population, indicating their potential in activating cell-mediated immune responses in chickens. Furthermore, the chimeric VLPs triggered the production of both Th1- and Th2-cytokines, attesting their potential in mounting a robust and balanced immune response in avian species. This study demonstrated the potential of these chimeric VLPs in stimulating and broadening cross-reactive immune responses in chickens against both avian and human IAVs.

Codon optimized influenza H1 HA sequence but not CTLA-4 targeting of HA antigen to enhance the efficacy of DNA vaccines in an animal model

Infections caused by the influenza virus lead to both epidemic and pandemic outbreaks in humans and animals. Owing to their rapid production, safety, and stability, DNA vaccines represent a promising avenue for eliciting immunity and thwarting viral infections. While DNA vaccines have demonstrated substantial efficacy in murine models, their effectiveness in larger animals remains subdued. This limitation may be addressed by augmenting the immunogenicity of DNA-based vaccines. In the investigation here, protein expression was enhanced via codon optimization and then mouse cytotoxic T-lymphocyte antigen 4 (CTLA-4) was harnessed as a modulatory adjunct to bind directly to antigen-presenting cells. Further, the study evaluated the immunogenicity of two variants of the hemagglutinin (HA) antigen, i.e. the full-length and the C-terminal deletion versions. The study findings revealed that the codon-optimized HA gene (pcHA) led to increased protein synthesis, as evidenced by elevated mRNA levels. Codon optimization also significantly bolstered both cellular and humoral immune responses. In cytokine assays, all plasmid constructs, particularly pCTLA4-cHA, induced robust interferon (IFN)-γ production, while interleukin (IL)-4 levels remained uniformly non-significant. Mice immunized with pcHA displayed an augmented presence of IFNγ+ T-cells, underscoring the enhanced potency of the codon-optimized HA vaccine. Contrarily, CTLA-4-fused DNA vaccines did not significantly amplify the immune response.

Antigenic drift and subtype interference shape A(H3N2) epidemic dynamics in the United States.

Influenza viruses continually evolve new antigenic variants, through mutations in epitopes of their major surface proteins, hemagglutinin (HA) and neuraminidase (NA). Antigenic drift potentiates the reinfection of previously infected individuals, but the contribution of this process to variability in annual epidemics is not well understood. Here, we link influenza A(H3N2) virus evolution to regional epidemic dynamics in the United States during 1997-2019. We integrate phenotypic measures of HA antigenic drift and sequence-based measures of HA and NA fitness to infer antigenic and genetic distances between viruses circulating in successive seasons. We estimate the magnitude, severity, timing, transmission rate, age-specific patterns, and subtype dominance of each regional outbreak and find that genetic distance based on broad sets of epitope sites is the strongest evolutionary predictor of A(H3N2) virus epidemiology. Increased HA and NA epitope distance between seasons correlates with larger, more intense epidemics, higher transmission, greater A(H3N2) subtype dominance, and a greater proportion of cases in adults relative to children, consistent with increased population susceptibility. Based on random forest models, A(H1N1) incidence impacts A(H3N2) epidemics to a greater extent than viral evolution, suggesting that subtype interference is a major driver of influenza A virus infection ynamics, presumably via heterosubtypic cross-immunity.

Antigenic drift and subtype interference shape A(H3N2) epidemic dynamics in the United States

Influenza viruses continually evolve new antigenic variants, through mutations in epitopes of their major surface proteins, hemagglutinin (HA) and neuraminidase (NA). Antigenic drift potentiates the reinfection of previously infected individuals, but the contribution of this process to variability in annual epidemics is not well understood. Here, we link influenza A(H3N2) virus evolution to regional epidemic dynamics in the United States during 1997—2019. We integrate phenotypic measures of HA antigenic drift and sequence-based measures of HA and NA fitness to infer antigenic and genetic distances between viruses circulating in successive seasons. We estimate the magnitude, severity, timing, transmission rate, age-specific patterns, and subtype dominance of each regional outbreak and find that genetic distance based on broad sets of epitope sites is the strongest evolutionary predictor of A(H3N2) virus epidemiology. Increased HA and NA epitope distance between seasons correlates with larger, more intense epidemics, higher transmission, greater A(H3N2) subtype dominance, and a greater proportion of cases in adults relative to children, consistent with increased population susceptibility. Based on random forest models, A(H1N1) incidence impacts A(H3N2) epidemics to a greater extent than viral evolution, suggesting that subtype interference is a major driver of influenza A virus infection ynamics, presumably via heterosubtypic cross-immunity.

Evolutionary Insights from Association Rule Mining of Co-Occurring Mutations in Influenza Hemagglutinin and Neuraminidase.

Seasonal influenza viruses continuously evolve via antigenic drift. This leads to recurring epidemics, globally significant mortality rates, and the need for annually updated vaccines. Co-occurring mutations in hemagglutinin (HA) and neuraminidase (NA) are suggested to have synergistic interactions where mutations can increase the chances of immune escape and viral fitness. Association rule mining was used to identify temporal relationships of co-occurring HA-NA mutations of influenza virus A/H3N2 and its role in antigenic evolution. A total of 64 clusters were found. These included well-known mutations responsible for antigenic drift, as well as previously undiscovered groups. A majority (41/64) were associated with known antigenic sites, and 38/64 involved mutations across both HA and NA. The emergence and disappearance of N-glycosylation sites in the pattern of N-X-[S/T] were also identified, which are crucial post-translational processes to maintain protein stability and functional balance (e.g., emergence of NA:339ASP and disappearance of HA:187ASP). Our study offers an alternative approach to the existing mutual-information and phylogenetic methods used to identify co-occurring mutations, enabling faster processing of large amounts of data. Our approach can facilitate the prediction of critical mutations given their occurrence in a previous season, facilitating vaccine development for the next flu season and leading to better preparation for future pandemics.

Frecuencia y conocimiento de tripanosomiasis americana en habitantes de la municipalidad de Vinto, Cochabamba, Bolivia

Introduction: American trypanosomiasis is caused by the blood parasite Trypanosoma cruzi, it affects 21 countries in the America, especially rural populations. Objectives: To determine the frequency and knowledge of American trypanosomiasis, as well as to disaggregate the frequency of American trypanosomiasis reemplazar por by sex and age group in the inhabitants of the municipality of Vinto, Cochabamba - Bolivia. Materials and Methods: A descriptive study with a non-experimental cross-sectional design was carried out. A total of 207 inhabitants of the municipality of Vinto attended the clinical analysis laboratory of the Biochemistry Department of the Adventist University of Bolivia. The sera were processed by indirect hemagglutination (HAI). Knowledge about American trypanosomiasis was determined by means of a structured survey. Pearson’s chi-square was used to relate variables and was considered significant at p<0.05. Results: The frequency of American trypanosomiasis in inhabitants of the municipality of Vinto was 11.6%. There was no relationship between the frequency of American trypanosomiasis and sex (p>0.05) but there was a relationship with the age group (p<0.05). 72.9% claimed to know about American trypanosomiasis. Conclusions: The results show that the frequency of American trypanosomiasis is low and that most of the inhabitants of the municipality of Vinto, Cochabamba, Bolivia have knowledge about the disease. In addition, it was found that there is only a relationship betweenthe frequency of American trypanosomiasis and age group.

Antibody Response of Mice to Bali Isolate of Canine Parvovirus Propagated in Madin-Darby Canine Kidney Cell Culture

Canine parvovirus (CPV) infection is still common among dogs, leading to severe disease with high mortality. The potential of a local isolate of CPV as an effective vaccine to prevent the disease warrants investigation. This study aimed to determine the antibody response in mice against a Bali isolate of CPV propagated in the Madin-Darby Canine Kidney (MDCK) cell culture. The virus was purified using polyethylene glycol (PEG)-6000 and mixed with an Aluminum hydroxide adjuvant. Fifteen 7-week female mice were divided into three treatment groups: treatment group 1 (PEG-purified virus and Adjuvant), treatment group 2 (crude unpurified virus and adjuvant), and treatment group 3 (adjuvant without virus), with five replicates per group. The Bali isolate of CPV was successfully replicated in MDCK cells, achieving a titer of 210-211 hemagglutination (HA) units after eight serial passages through the cell culture. The virus was confirmed as CPV by immunocytochemistry test using a monoclonal antibody and hemagglutination inhibition (HI) test using chicken anti-CPV polyclonal antibody. Following the first immunization, the antibody endpoint titer in mice immunized with PEG-purified CPV (5.6) was significantly higher than those immunized with crude unpurified CPV (4.2) and adjuvant without CPV (1.4). Similarly, after the second immunization, the antibody endpoint titer in mice immunized with PEG-purified CPV (7.6) also remained significantly higher than those immunized with crude unpurified CPV (6.4) and adjuvant without CPV (0.8). Significant increases in antibody endpoint titer were observed after the second immunization in mice immunized with PEG-purified CPV and crude unpurified CPV, but not in those given adjuvant without CPV. The Bali isolate of CPV propagated in MDCK cell culture induced a robust antibody response in mice, suggesting it’s a potential as an alternative vaccine candidate for preventing CPV infection in dogs.

Genetic characteristics of influenza A and B viruses circulating in Russia in 2019-2023.

Relevance. Influenza viruses have a high potential for genetic change. These viruses are monitored annually around the world, including Russia, to determine the dominant genetic groups and select the strains to be included in influenza vaccines. Aims of the study include: analysis of the circulation of influenza viruses in Russia in 2019-2023, phylogenetic and molecular analysis of hemagglutinin (HA) sequences of influenza virus strains, detection of drug resistance mutations to neuraminidase (NA) inhibitors and M2-protein (M2) ion channel inhibitors. Materials and methods. Biological samples containing RNA of influenza viruses were studied: 417 A(H1N1)pdm09, 147 A(H3N2) and 167 B(Victoria). Sequencing of the PCR fragments was performed on the 3500xL Genetic Analyzer (Applied Biosytems). Data processing and analysis were carried out using DNASTAR, Nextclade, FluSurver and BioNumerics v.6.6 software. Results. Influenza A(H1N1)pdm09, A(H3N2), B(Victoria) viruses circulating in 2019-2023 were investigated. The highest variability of HA was observed in A(H3N2) viruses. 3% of A(H1N1)pdm09 viruses had the D222N mutation in the receptor-binding site of HA, which is associated with more severe disease. The oseltamivir and zanamivir resistance mutation H275Y in NA was detected in 2% of influenza A(H1N1)pdm09 viruses. No oseltamivir and zanamivir resistance mutations in NA were detected in all influenza A(H3N2) and B viruses tested. In all investigated influenza A(H1N1)pdm09 and A(H3N2) viruses the adamantane resistance mutation S31N in M2 was identified. Conclusions. The detection of amino acid substitutions in HA antigenic sites and resistance mutations in NA and M2 confirms the evolution of influenza viruses and the need for continuous genetic surveillance.

Hemagglutinin and neuraminidase of a non-pathogenic H7N7 avian influenza virus coevolved during the acquisition of intranasal pathogenicity in chickens.

Polybasic amino acid residues at the hemagglutinin (HA) cleavage site are insufficient to induce the highly pathogenic phenotype of avian influenza viruses in chickens. In our previous study, an H7N7 avian influenza virus named "Vac2sub-P0", which is nonpathogenic despite carrying polybasic amino acids at the HA cleavage site, was passaged in chick air sacs, and a virus with high intravenous pathogenicity, Vac2sub-P3, was obtained. Intranasal infection with Vac2sub-P3 resulted in limited lethality in chickens; therefore, in this study, this virus was further passaged in chicken lungs, and the resultant virus, Vac2sub-P3L4, acquired high intranasal pathogenicity. Experimental infection of chickens with recombinant viruses demonstrated that mutations in HA and neuraminidase (NA) found in consecutive passages were responsible for the increased pathogenicity. The HA and NA functions of Vac2sub-P3L4 were compared with those of the parental virus in vitro; the virus growth at 40 °C was faster, the binding affinity to a sialic acid receptor was lower, and the rate of release by NA from the cell surface was lower, suggesting that these changes enabled the virus to replicate efficiently in chickens with high intranasal pathogenicity. This study demonstrates that viruses that are highly pathogenic when administered intranasally require additional adaptations for increased pathogenicity to be highly lethal to intranasally infected chickens.

Isolation and characterization of IgG3 glycan-targeting antibodies with exceptional cross-reactivity for diverse viral families.

Broadly reactive antibodies that target sequence-diverse antigens are of interest for vaccine design and monoclonal antibody therapeutic development because they can protect against multiple strains of a virus and provide a barrier to evolution of escape mutants. Using LIBRA-seq (linking B cell receptor to antigen specificity through sequencing) data for the B cell repertoire of an individual chronically infected with human immunodeficiency virus type 1 (HIV-1), we identified a lineage of IgG3 antibodies predicted to bind to HIV-1 Envelope (Env) and influenza A Hemagglutinin (HA). Two lineage members, antibodies 2526 and 546, were confirmed to bind to a large panel of diverse antigens, including several strains of HIV-1 Env, influenza HA, coronavirus (CoV) spike, hepatitis C virus (HCV) E protein, Nipah virus (NiV) F protein, and Langya virus (LayV) F protein. We found that both antibodies bind to complex glycans on the antigenic surfaces. Antibody 2526 targets the stem region of influenza HA and the N-terminal domain (NTD) region of SARS-CoV-2 spike. A crystal structure of 2526 Fab bound to mannose revealed the presence of a glycan-binding pocket on the light chain. Antibody 2526 cross-reacted with antigens from multiple pathogens and displayed no signs of autoreactivity. These features distinguish antibody 2526 from previously described glycan-reactive antibodies. Further study of this antibody class may aid in the selection and engineering of broadly reactive antibody therapeutics and can inform the development of effective vaccines with exceptional breadth of pathogen coverage.

Dually-amplified electrochemical aptasensor based on the self-linking AuPt nanoflowers for ultrasensitive determination of hemagglutinin

BackgroundInfluenza virus can spread from person to person and cause epidemics. Therefore, rapid and sensitive diagnosis of virus is essential in controlling influenza outbreaks. Conventional virus diagnostic techniques are time-consuming, labor-intensive and requires large instruments. In this work, a sandwich electrochemical assay by a pair of aptamers was developed for ultrasensitive determination of hemagglutinin (HA) protein, which is one of the two surface glycoproteins of influenza A (H1N1) virus, using dual signal amplification techniques. ResultsHA was captured and magnetically separated by Fe3O4@SiO2–NH2@Au attached to aptamer 1 (Apt1), creating a sandwich structure with AuPt nanoflowers (AuPtNFs) connected to aptamer 2 (Apt2). Herein, AuPtNFs could catalyze H2O2/hydroquinone (HQ) to generate 1,4-benzoquinone (BQ), and achieved amplification of electrochemical signal detection through differential pulse voltammetry (DPV). The constructed aptasensor expressed a wide linear range (10 pg/mL-100 ng/mL) with limit of detection (LOD) of 2.4 pg/mL. Moreover, a novel strategy for dual signal amplification was developed to further enhance sensitivity. The innovative electrochemical aptasensor could achieve secondary amplification of the detection signal with LOD of 0.3 pg/mL and linear concentration range from 0.5 pg/mL to 100 ng/mL. The secondary amplification could be achieved only through the self-linking process, which allowed for the retention of numerous AuPtNFs by simple complementary base pairing to connect more AuPtNFs onto the above-mentioned sandwich structure. SignificanceOverall, the constructed aptasensor exhibited favorable sensitivity and accuracy, indicating the potential expanded application for the clinical detection of numerous viruses.

COBRA N2 NA vaccines induce protective immune responses against influenza viral infection

ABSTRACT Influenza neuraminidase (NA) is a promising target for a broadly protective vaccine. In this study, the Computationally Optimized Broadly Reactive Antigen (COBRA) methodology was used to develop N2 NA vaccine candidates. The unique wild type (WT) N2 sequences of human and swine influenza strains isolated between 1957 and 2019 were used to design the COBRA N2-A NA vaccine, while the unique WT N2 sequences of human influenza strains isolated between 2000 and 2019 were used to design the COBRA N2-B NA vaccine. Sera collected from COBRA N2 NA vaccinated mice showed more broadly reactive antibody responses against a broad panel of H×N2 influenza virus strains than sera collected from mice vaccinated with WT N2 NA vaccines. Antibodies elicited by COBRA or WT N2 NA antigens cross react with recent human H3N2 influenza viruses from different clades, while the antibodies elicited by A/Switzerland/9715293/2013 hemagglutinin (HA) reacted with viruses from the same clade. Furthermore, mice vaccinated with COBRA N2-B NA vaccine had lower viral lung titers compared to mock vaccinated mice when challenged with human H3N2 influenza viruses. Thus, the COBRA N2 NA vaccines elicit broadly protective murine anti-NA antibodies against multiple strains across subtypes and the viral loads were significantly decreased in the lungs of the mice in the COBRA N2 NA vaccine groups, compared to the mice in the mock vaccinated group, indicating that the COBRA-based N2 subtype NA vaccines have a potential to be a component in a universal influenza vaccine.

Influenza virus antibodies inhibit antigen-specific de novo B cell responses in mice.

Antibody responses to influenza vaccines tend to be focused on epitopes encountered during prior influenza exposures, with little production of de novo responses to novel epitopes. To examine the contribution of circulating antibodies to this phenomenon, we passively transferred a hemagglutinin (HA)-specific monoclonal antibody (mAb) into mice before immunizing with whole inactivated virions. The HA mAb inhibited de novo HA-specific antibodies, plasmablasts, germinal center B cells, and memory B cells, while responses to a second antigen in the vaccine, neuraminidase (NA), were uninhibited. The HA mAb potently inhibited de novo antibody responses against epitopes near the HA mAb binding site. The HA mAb also promoted IgG1 class switching, an effect that, unlike the inhibition of HA responses, relied on signaling through Fc-gamma receptors. These studies suggest that circulating antibodies inhibit de novo B cell responses in an antigen-specific manner, which likely contributes to differences in antibody specificities elicited during primary and secondary influenza virus exposures.IMPORTANCEMost humans are exposed to influenza viruses in childhood and then subsequently exposed to antigenically drifted influenza variants later in life. It is unclear if antibodies elicited by earlier influenza virus exposures impact immunity against new influenza virus strains. Here, we used a mouse model to investigate how an anti-hemagglutinin (HA) monoclonal antibody (mAb) affects de novo B cell and antibody responses to the protein targeted by the monoclonal antibody (HA) and a second protein not targeted by the monoclonal antibody [neuraminidase (NA)]. Collectively, our studies suggest that circulating anti-influenza virus antibodies can potently modulate the magnitude and specificity of antibody responses elicited by secondary influenza virus exposures.

Phase 1 dose-escalation trial evaluating a group 2 influenza hemagglutinin stabilized stem nanoparticle vaccine

The relative conservation of the influenza hemagglutinin (HA) stem compared to that of the immunodominant HA head makes the HA stem an attractive target for broadly protective influenza vaccines. Here we report the first-in-human, dose-escalation, open-label trial (NCT04579250) evaluating an unadjuvanted group 2 stabilized stem ferritin nanoparticle vaccine based on the H10 A/Jiangxi-Donghu/346/2013 influenza HA, H10ssF, in healthy adults. Participants received a single 20 mcg dose (n = 3) or two 60 mcg doses 16 weeks apart (n = 22). Vaccination with H10ssF was safe and well tolerated with only mild systemic and local reactogenicity reported. No serious adverse events occurred. Vaccination significantly increased homologous H10 HA stem binding and neutralizing antibodies at 2 weeks after both first and second vaccinations, and these responses remained above baseline at 40 weeks. Heterologous H3 and H7 binding antibodies also significantly increased after each vaccination and remained elevated throughout the study. These data indicate that the group 2 HA stem nanoparticle vaccine is safe and induces stem-directed binding and neutralizing antibodies.

Structure-guided assembly of an influenza spike nanobicelle vaccine provides pan H1 intranasal protection.

Development of intranasal vaccines for respiratory viruses has gained popularity. However, currently only a live-attenuated influenza vaccine is FDA-approved for intranasal administration. Here, we focused on influenza virus as it circulates seasonally, has pandemic potential, and has vaccine formulations that present hemagglutinin (HA) in different structural arrangements. These display differences have not been correlated with induction of pan-H1 antibodies or shown to provide intranasal protection. Using electron microscopy, biochemistry and animal studies, we identified HA complexes arranged as lipid discs with multiple trimeric HAs displayed along the perimeter, termed spike nanobicelles (SNB). We utilized a structure-guided approach to synthesize in vitro assembled spiked nanobicelles (IA-SNB) from a classical 1934 H1N1 influenza virus. IA-SNBs elicited pan-H1 antibodies and provided protection against antigenically divergent H1N1 viruses via intranasal immunizations. Viral glycoprotein spikes displayed as SNBs could aid in combating antigenic variation and provide innovative intranasal vaccines to aid universal influenza vaccine development.

Red Blood Cell‐Derived Small Extracellular Vesicles Inhibit Influenza Virus through Surface‐Displayed Sialic Acids

Disrupting the conserved multivalent binding of hemagglutinin (HA) on influenza A virus (IAV) to sialic acids (SAs) on the host cell membrane offers a robust strategy to block viral attachment and infection, irrespective of antigenic evolution or drug resistance. In this study, we exploit red blood cell‐derived small extracellular vesicles (RBC sEVs) as nanodecoys by harnessing their high abundance of surface‐displayed SAs to interact with IAV through multivalent HA‐SA interactions. This high‐avidity binding inhibits viral adhesion to the cell surface, effectively preventing both attachment and infection in a dose‐dependent manner. Notably, enzymatic removal of SAs from RBC sEVs significantly diminishes their anti‐IAV efficacy. Our findings indicate that RBC sEVs possess intrinsic anti‐IAV properties due to their native multivalent SAs and hold considerable promise as antiviral therapeutics.