ETIDIOH - NEW

  ABSTRACT The outbreak of highly pathogenic avian H5 influenza (HPAI) clade 2.3.4.4b in cattle has spread across the United States . Mice with pre-existing immunity to H1N1 virus or with a live-attenuated influenza vaccine showed protection against a lethal bovine-derived HPAI H5N1 viral challenge . Notably, ferrets with mixed immunity also demonstrated protection against a feline-derived H5N1 virus, independent of cross-reactive neutralization titers , but antibodies to whole virus were observed. To investigate protective factors, we conducted T cell epitope mapping using published H1N1 viral sequences and found high conservation of key T cell epitopes in the bovine HPAI H5N1 strain . Depletion of T cells in mice prior to and during primary H1N1 infection impacted cross-protective antibodies to H5N1 virus, with CD4 depletion increasing mortality and CD8 depletion mildly impacting morbidity upon H5N1 viral challenge. This underscores the need to investigate memory T cell responses...

  Abstract Annual influenza vaccination is the cornerstone for seasonal protection, yet antibody responses are highly variable across individuals and over time. To systematically assess the determinants of this heterogeneity, we compiled 20,449 hemagglutination inhibition and neutralization titers from 4,540 participants enrolled in 14 new vaccine studies we conducted and 50 prior studies that collectively span 2010-2023. Seasonal effects dominated , with pre- and post-vaccination titers declining steadily from 2017 onwards, outweighing the influence of age, sex, or repeated vaccination. Titers to B Yamagata remained steady throughout all years examined, suggesting unique durability and offering a reason for lineage extinction . Vaccine timing emerged as a strong and previously underappreciated determinant of immunity, with individuals vaccinated later in the season exhibiting larger post-vaccination titers . Not being vaccinated or receiving the live-attenuated FluMist vaccine in ...

  Abstract Ongoing transmission of influenza A virus (H5N1) in U.S. dairy cattle threatens both animal and human health , underscoring the need to understand the durability of host immunity against reinfection with evolving genotypes . We challenged naive and convalescent cows , infected one year prior with H5N1 genotype B3.13, with either homologous B3.13 or heterologous D1.1 genotype virus . Homologous rechallenge resulted in complete clinical protection with no infectious viral shedding . Conversely, heterologous rechallenge led to transient clinical disease and limited infectious viral shedding . Convalescent cows experienced significantly milder disease than naive cows, which developed severe illness with high viral shedding and required early euthanasia , regardless of the strain. These findings indicate that naturally acquired immunity offers strong protection against severe illness but may allow silent transmission of divergent strains . Therefore, natural herd immunity alo...

  Abstract SARS-CoV-2 immunity and innate immune training may influence influenza vaccine immunogenicity . We investigated this in India . Adult volunteers with hybrid SARS-CoV-2 immunity were administered FluarixTM Tetra (GlaxoSmithKlein) 2022/2023 NH Vaccine in 2022. Significant induction of hemagglutinin inhibition-specific antibodies and polyfunctional central memory CD4+ T-cells (TCM) were observed 1-week post-vaccination with variable induction of CD8+T-cell and innate effectors. Vaccination also expanded Flu-specific regulatory T-cells (Treg), which negatively correlated with CD4 responses , highlighting vaccine immunogenicity may be subject to Treg dampening . FluarixTM did not boost SARS-CoV-2 immunity . However, SARS-CoV-2 -specific T-cell responses correlated positively with vaccine-induced T-cell responses. We evaluated trained immunity post-COVID-19 as a potential regulatory mechanism linking SARS-CoV-2 and heterologous vaccine immunogenicity . We observed, elevated fr...

  ABSTRACT Middle East respiratory syndrome coronavirus (MERS-CoV) causes a highly lethal respiratory infection for which no vaccines or antiviral therapeutics are currently available . Understanding the immune response is critical for designing effective therapeutics . Here, we comprehensively characterized the dynamics of B-cell responses in severely infected MERS-CoV patients and survivors with SARS-CoV-2 exposure history . Infected patients developed robust neutralizing antibody responses within 1 month of illness , with moderate-to-high cross-neutralization activity against SARS-CoV-2 . The enhanced neutralization activity coincided with an increased abundance of specific mutated, class-switched IgG clones. Notably, one such clone was detected at moderate prevalence in both patients, and its expansion was accompanied by high neutralization activity against both viruses . Conversely, MERS-CoV survivors demonstrated higher neutralization activity against MERS-CoV after vaccinati...

  Abstract The development of T-cell-based influenza vaccines relies on eliciting broad CD8+ T-cell immunity, wherein T stem cell-like memory (TSCM) cells serve as the ultimate long-lived reservoir for immune memory, thereby unlocking the potential for durable protection against viral drift and shift. However, the specific immunological cues that drive the robust expansion and functional preservation of this self-renewing, multipotent subset remain unknown. Here, utilizing multi-omic systems immunology in a pediatric cohort immunized with live attenuated influenza vaccine , we identified the determinants governing the expansion of influenza virus-reactive TSCM cells . We show that a pre-existing state of innate antiviral readiness , defined by a plasmacytoid dendritic cell-associated type I interferon signature , is the requisite condition for a robust TSCM expansion. Mechanistically, this baseline innate state enhances antigen priming and enforces a qualitative divergence in T-cel...

  Abstract The unprecedented 2.3.4.4b. A(H5N1) outbreak in dairy cattle, poultry, and spillover to humans in the United States (US) poses a major public health threat. Population immunity is a critical component of influenza pandemic risk assessment . We assessed the pre-existing cross-reactive immunity to 2.3.4.4b A(H5N1) viruses and analyzed 1794 sera from 723 people (0.5–88 yrs) in multiple US geographic regions during 2021–2024. Pre-existing neutralizing and hemagglutinin (HA)-head- binding antibodies to A(H5N1) were low , but there were substantial cross-reactive binding antibodies to N1 neuraminidase (NA) of 2.3.4.4b A(H5N1). Antibodies to group 1 HA stalk were also prevalent and increased with age . A(H1N1)pdm09 infection and influenza vaccination did not induce neutralizing antibodies to A(H5N1) viruses but induced significant rise of functional NA inhibition (NAI) antibodies to N1 of 2.3.4.4b A(H5N1), and group 1 HA stalk antibodies . Moreover, pre-pandemic stockpiled 2.3....

  Abstract Influenza viruses cause hundreds of thousands of infections globally every year. In the past century, seasonal influenza viruses have included H1N1, H2N2 or H3N2 strains . H2N2 influenza viruses circulated in the human population between 1957-1968 . Previously, our group demonstrated a lack of H2N2 influenza virus immunity in individuals born after 1968 , as well as the effectiveness of hemagglutinin (HA) based vaccines for multiple influenza virus subtypes. In this study, H2 antigenic maps and radial graphs were generated using previously published data from H2 HA vaccinations of ferrets and seasonal influenza vaccinations of humans . The antigenic maps revealed a stark difference in clustering of HA antigens between the ferrets and humans, and the radial graphs showed specific antigen recognition varies greatly between different influenza preimmune ferrets . These maps also revealed the significant impact that different pre-existing immunities have on antigenic recogni...

  Significance Since pigs serve as intermediate hosts between humans and the natural reservoir of influenza viruses in wild birds, they play a key role in the emergence of influenza strains with pandemic potential , as demonstrated by the 2009 pandemic. Therefore, influenza pandemic preparedness will benefit from the development of vaccines that broadly protect pigs against diverse influenza A strains. However, progress is limited by our poor molecular understanding of porcine antibody responses to influenza virus. This study isolates and characterizes a panel of broadly neutralizing influenza antibodies from pigs . Our findings not only have significant implications for the development of broadly protective influenza vaccines for pigs, but also reveal the molecular differences in the antibody responses between pigs and humans. Abstract Antibody responses to the influenza virus hemagglutinin (HA) stem, a major target for broadly protective vaccine development , have been extensivel...

  Abstract Human influenza viruses rapidly acquire mutations in their hemagglutinin (HA) protein that erode neutralization by antibodies from prior exposures. Here, we use a sequencing-based assay to measure neutralization titers for 78 recent H3N2 HA strains against a large set of children and adult sera , measuring ~10,000 total titers . There is substantial person-to-person heterogeneity in the titers against different viral strains, both within and across age cohorts. The growth rates of H3N2 strains in the human population in 2023 are highly correlated with the fraction of sera with low titers against each strain. Notably, strain growth rates are less correlated with neutralization titers against pools of human sera, demonstrating the importance of population heterogeneity in shaping viral evolution . Overall, these results suggest that high-throughput neutralization measurements of human sera against many different viral strains can help explain the evolution of human influen...

  Abstract Influenza continues to cause significant mortality globally and possesses substantial pandemic potential . Assessing pandemic risk requires a clear understanding of existing population immunity . Leveraging a unique large-scale cohort of human sera , we evaluated total and neutralising antibody -mediated immunity to multiple haemagglutinin (HA) proteins, including those from subtypes with high pandemic potential. Our analysis reveals that population immunity is heterogeneous , with distinct age-dependent differences in responses to H5, H7, and H9 avian influenza subtypes. These shifts align with historical circulation patterns of seasonal H1N1 and H3N2 human viruses. Notably, H7 viruses are primarily neutralised through head domain epitopes , while H5 viruses are targeted mainly via stem epitopes , although in both instances some neutralisation occurred via receptor binding site-adjacent epitopes . Furthermore, H7 responses were dominated by non-glycan-targeted IgG2 anti...

  Abstract H5N1 highly pathogenic avian influenza viruses have spread globally and pose a risk for a human pandemic . Prior studies suggest that early life exposures to group 1 influenza viruses (H1N1 and H2N2) prime antibodies that cross-react to the hemagglutinin of H5N1 , which is also a group 1 virus. Less is known about how immune history affects antibody responses against the neuraminidase (NA) of H5N1 viruses. Here, we measured NA inhibition antibodies against multiple H5N1 viruses using sera from 155 individuals born between 1927 and 2016 . We found that individuals primed in childhood with H1N1 viruses were more likely to possess higher levels of antibodies that cross-react with the NA of H5N1 viruses compared to individuals primed in childhood with H2N2 or H3N2 viruses. While young children rarely possessed cross-reactive NA antibodies, we found that childhood infections with contemporary H1N1 , but not H3N2, viruses can elicit them. These data suggest that immune history...

  Abstract As intrinsic differences in humoral immune response to SARS-CoV-2 between children and adults remain unclear , we improved characterisation by defining the kinetics, specificity and function of antibodies to SARS-CoV-2 in children (n = 146, aged 9.4 ± 4.8 years with n = 257 samples) compared to adults (n = 85, aged 39.5 ± 15.2 years with n = 122 samples). We used plasma samples from an infection and vaccination-naive cohort study with RT-PCR confirmed ancestral B.1* SARS-CoV-2 virus infection with asymptomatic or mild disease, collected in Hong Kong between March to December 2020, from acute (0–14 days post infection) to convalescent (15–206 days) timepoints. Children had significantly lower primary antibody responses against SARS-CoV-2 proteins overall, leading to a less isotype switched response. While children had lower OC43 Spike and SARS-CoV-2 S2 IgG and avidity than adults, they exhibited higher avidities for SARS-CoV-2 whole Spike and Nucleocapsid , and higher lev...

  Abstract Reassortment between different influenza strains occurs when they co-infect the same host cell . The emergence of a reassortant virus depends on both its intrinsic fitness and extrinsic factors , including pre-existing humoral immunity . The generation of pandemic strains , such as H2N2 and H3N2 , and zoonotic influenza A viruses, like H5N6, H5N8, and H7N9 , in birds is suggested to be the result of extensive selection by pre-existing antibodies . To further explore the role of humoral immunity in reassortment , we generated two divergent fluorescent protein-expressing viruses and used strain-specific and cross-reactive monoclonal antibodies (mAbs) to assess the impact of cross-immunity on reassortment. Our results indicate that all mAbs altered the genotypic diversity and significantly reduced the release of progeny virions in co-infected cells both in vitro and in vivo. Moreover, antibody transfer studies in mice revealed protection from challenge with divergent pathog...

ABSTRACT Seasonal influenza causes 290,000–650,000 deaths annually, with vaccination efficacy ranging from 10 to 60%. The emergence of drug-resistant and highly pathogenic avian influenza viruses underscores the urgent need for novel protective strategies . Epidemiological observations have long suggested that certain vaccines, such as Bacillus Calmette-Guérin (BCG), can provide protection against diverse pathogens (S. Biering-Sørensen, P. Aaby, N. Lund, et al., Clin Infect Dis 65:1183–1190, 2017, https://doi.org/10.1093/cid/cix525 ; M.-L. Garly, C. L. Martins, C. Balé, et al., Vaccine 21:2782–2790, 2003, https://doi.org/10.1016/s0264-410x(03)00181-6 ; C. A. G. Timmermann, S. Biering‐Sørensen, P. Aaby, et al., Trop Med Int Health 20:1733–1744, 2015, https://doi.org/10.1111/tmi.12614 ). While the cellular and molecular mechanisms underlying such protection remain incompletely understood, emerging research offers critical insights into innate immune system modulation (B. Cirovic, L. C. J...

Abstract Background .  Highly pathogenic avian influenza A(H5) viruses pose a pandemic threat , with a history of zoonotic spillovers into humans that are presumed immunologically naive. Whether the general population is currently immunologically naive to circulating A(H5) influenza viruses is unknown.  Methods .  To evaluate the presence of cross-reactive immune responses to emerging A(H5) clade 2.3.4.4b influenza viruses in the general population, we conducted comprehensive immune profiling on cross-sectional samples from healthcare workers (n=107). Samples were collected in August and September 2024 in the scope of an ongoing prospective follow-up study: Surveillance of rEspiratory viruses iN healThcare and anImal workers in the NethErLands (SENTINEL).  Findings .  Low-level antibody responses directed against the A(H5) hemagglutinin (HA) head were detected in a limited number of individuals , but without hemagglutination inhibition activity. Nevertheless, we...

Editor’s summary The vast majority of the human population has immunity to influenza A virus (IAV) by prior infection, vaccination, or both . However, protection is generally subtype-specific , and it is not clear whether prior infection against one subtype could confer protection against clade 2.3.4.4b H5N1 IAVs , which are currently circulating in birds and dairy cows . Here, Restori et al. demonstrated that prior infection with the 2009 pandemic H1N1 IAV was protective against subsequent direct infection with H5N1 IAV in ferrets. Moreover, prior immunity reduced susceptibility to infection by transmission from an infected donor ferret. These data suggest that prior immunity to IAV, especially to the 2009 pandemic H1N1 virus, may offer a degree of protection against H5N1 infection. —Courtney Malo Abstract Zoonotic infections with emerging influenza viruses occur in the context of population-wide immunity to seasonal strains . Because of the worldwide spread of highly pathogenic clade...

Abstract The spill-over of Influenza A virus H5N1 clade 2.3.4.4b from cattle to humans highlights the risk of a human H5N1 pandemic . Given the impact of pre-existing immunity on the course and severity of viral infections, we comprehensively assessed the humoral immunity against the H5N1 A/Texas/37/2024 isolate in H5N1- naive individuals . To this end, we performed complementary binding and neutralization assays on 66 subjects and ranked activities among a panel of 76 influenza A virus isolates. We detected low but distinct cross-neutralizing titers against A/Texas/37/2024 with a 3.9 to 15.6-fold reduction compared to selected H1N1 or H3N2 strains. By cloning and evaluating 136 monoclonal antibodies from memory B cells, we identified potent A/Texas/37/2024-neutralizing monoclonal antibodies in five out of six investigated individuals. These antibodies cross-neutralize H1, compete with antibodies targeting the HA stem, and protect mice from lethal H5N1 challenge. Our findings demonstra...

Abstract An increase in the number of human cases of influenza A/H5N1 infection in the US has raised concerns about the pandemic potential of the virus . Preexisting population immunity is a key determinant for risk assessment and pandemic potential for any virus. Antibody responses against the bovine A/H5N1 hemagglutinin (HA) and neuraminidase (NA) proteins were measured among a population of influenza-vaccinated or influenza-infected individuals . Modest titers of bovine A/H5N1 HA-binding antibodies and low to undetectable neutralizing antibody responses were detected in a cohort of 73 individuals . Conversely, bovine A/H5N1 NA binding and neuraminidase-inhibiting antibody responses were comparable to those against a human A/ H1N1 NA at baseline. Seasonal influenza vaccination failed to significantly increase antibody titers against both HA and NA glycoproteins of bovine A/H5N1. Recent infection with human A/H1N1 but not A/H3N2 viruses induced significant increases in bovine A/H5N1 n...

Abstract Highly pathogenic avian influenza of the H5N1 subtype has shown recent unprecedented expansion in its geographic and host range , increasing the pandemic threat . The younger age of H5N1 versus H7N9 avian influenza in humans has previously been attributed to imprinted pre-immunity to hemagglutinin stalk (HA2) epitopes shared with group 1 ( H1N1, H2N2 ) versus group 2 ( H3N2 ) influenza A subtypes predominating in the human population before versus after 1968, respectively. Here we review the complex immuno-epidemiological interactions underpinning influenza risk assessment and extend the imprinting hypothesis to include a potential role for cross-protective neuraminidase (NA) imprinting . We compare H5N1 distributions and case fatality ratios by age and birth cohort (as proxy for HA2 and/or NA imprinting epoch) not only to H7N9 but also H5N6 and H9N2 avian influenza, representing more varied conditions of zoonotic influenza relatedness to human subtypes of the past century. We...