ETIDIOH - NEW

  Abstract Although seasonal influenza vaccination programs are effective at a population level, our data from inactivated influenza vaccine (IIV) cohorts in years 2015-2022 reveal that 50-60% of individuals do not seroconvert following immunization. The underlying mechanisms of vaccine non-responsiveness are far from understood. In this study, we sought to define key determinants of optimal B cell immune responses elicited by seasonal influenza vaccination, and to explore why some individuals fail to elicit humoral immunity following immunization. Immune responses associated with seroconversion and vaccine failure from individuals immunized with IIVs were compared at cellular and molecular levels using single-cell transcriptomics . We analyzed HA-specific B cell immunity across vaccine-responders, breakthrough infections and patients hospitalized with acute influenza. Droplet-based single-cell RNA sequencing and VDJ-sequencing of influenza-specific B cells from stored PBMCs was pe...

  ABSTRACT Respiratory viruses can infect hosts concurrently or sequentially, potentially influencing each other’s pathogenic trajectory . However, the underlying immune mechanisms governing these interactions remain poorly understood. In this study, we examined whether respiratory syncytial virus (RSV) infection modulates host susceptibility to subsequent SARS-CoV-2 infection using two murine models . We found that prior RSV infection conferred dose- and time-dependent heterologous protection against SARS-CoV-2 . Transcriptomic and immunological analyses revealed that RSV activated lung antigen-presenting cells (APCs) and SARS-CoV-2–reactive mucosal T cells by day 9 post-infection , with responses waning by 1 month . RSV also promoted expansion of pulmonary γδ T cells and upregulation of their metabolic pathways. Notably, RSV-infected TCRδ⁻/⁻ mice , which lack γδ T cells, exhibited diminished SARS-CoV-2–reactive mucosal T cell responses, elevated viral loads, and exacerbated lung ...

  Abstract Currently, there is limited knowledge on the impact of immunity to hemagglutinin (HA) and/or neuraminidase (NA) on the transmission of influenza viruses . Therefore, using intramuscular vaccination , intranasal vaccination , or infection with reassortant viruses , we induced immunity to each antigen alone or both antigens combined in ferrets . We then assessed transmission of the 2009 pandemic H1N1 virus from these ferrets to naïve respiratory contacts . For all strategies used to induce immunity, combined immunity to HA and NA resulted in the largest reductions in transmission . Moreover, immunity to HA and NA conferred additive rather than synergistic reductions in transmission. No escape variants emerged in our transmission studies, and logistical regression showed that the probability of transmission was less than 50% when viral titers in donors were reduced to 101.5 and 102 median tissue culture infectious dose per ml on days 1 and 3 postinfection, respectively. The...

  Abstract Since the West African Ebola virus (EBOV) epidemic in 2014-2016, recurrent outbreaks of the EBOV-Makona variant have been driven by recrudescence and human-to-human transmission emphasizing the need for effective vaccination strategies . A live-attenuated recombinant vesicular stomatitis virus (VSV)-based vaccine expressing the EBOV-Kikwit variant glycoprotein (VSV-Kik) received FDA approval in December 2019 and provides complete, rapid protection against EBOV-Makona as early as 7 days post-vaccination (DPV). During the 2018-2020 Ebola outbreak , the VSV-Kik vaccine, known as ERVEBO , was administered to lower-risk individuals at a 5-fold dose reduction of the standard 2 × 107 PFU to provide broader population protection. Identification of a protective lower dose providing rapid protection would ease supply burdens during future outbreaks and enhance vaccine coverage. We previously generated a VSV-based vaccine expressing the glycoprotein of the Makona variant (VSV-Mak) ...

  Abstract While influenza A virus undergoes rapid antigenic drift in humans, at least some subtypes, such as H3, have relatively stable antigenicity in natural waterfowl reservoirs, despite the presence of immune pressure . However, the underlying mechanisms remain poorly understood. This study identified and characterized 187 antibodies to H3 hemagglutinin from experimentally infected mallard ducks , 18 of which were further analyzed by cryo-EM . Compared with human H3 antibodies , duck H3 antibodies exhibited higher glycan-binding propensity , more balanced immunodominance hierarchy , and targeted distinct epitopes . Other unique features of duck H3 antibodies included a convergent CDR H3-independent heavy chain-only binding mode and an N-glycosylated CDR H3 as decoy receptor . By annotating duck immunoglobulin germline genes , we also demonstrated the importance of gene conversion in duck H3 antibodies. Overall, our findings provide insights into how millennia of coevolution ha...

  Abstract Background :  The H1N1 influenza A virus evades host immunity through continuous antigenic drift, posing a significant challenge to broad-spectrum neutralizing antibody therapies. This study aims to systematically evaluate the neutralizing capacity of the broad-spectrum antibody C12H5 against H1N1 strains from different eras and identify key immune escape mutation sites .  Methods :  Three representative H1N1 virus strains from 2009, 2018, and 2023 were selected. An antigen–antibody binding prediction model based on the ESM-2 large language model was constructed by integrating 48,762 GISAID sequence data and deep mutation scanning data from the Bloom laboratory. Candidate escape sites were screened using SHAP (SHapley Additive exPlanations) value analysis. Mutant viruses were constructed via reverse genetics, and their neutralizing capacity and replication fitness were validated through hemagglutination inhibition assays, microneutralization assays, and vi...

  Abstract Highly pathogenic avian influenza A H5N1 has recently expanded its mammalian host range ; in 2024, genotype B3.13 emerged in U.S. dairy cattle with pronounced mammary tropism . In the past, Influenza A virus immunology has been characterized primarily in respiratory infection models , whereas this study delineates immune responses after intramammary infection . An intramammary H5N1 challenge in Jersey cows in the early dry-off period enabled integration of dose- and compartment-resolved (alveoli versus teat cistern) cytokine and chemokine profiles with peripheral leukocyte dynamics and H5/N1-specific antibody responses. Infection-induced quarter-restricted, monophasic inflammatory networks peaking at 3 to 7 days post-infection , coordinated peripheral myeloid expansion and IFN gamma competent lymphocyte activation, and rising antibody titers across quarters. Competing Interest Statement The J.A.R. laboratory received support from Tonix Pharmaceuticals, Xing Technologies,...

  Abstract Antibodies to the influenza hemagglutinin protein (HA) confer the strongest protection against infection . Immunity elicited by endemic, seasonal, human viruses is correlated with diminished disease severity and death caused by antigenically novel viruses. Antibodies to the HA head interface are broadly protective and abundant in human serologic and memory repertoires . Notably, few head interface antibodies from H5 naive donors are reported to bind H5 HAs . We find head interface antibodies engage a wide range of H5 isolates but fail to engage most isolates from the goose Guangdong (GsGd) lineage . We identify a single substitution, P221S, largely dictates antibody binding. Phylogenetic analysis indicates that P221S arose in a Chinese avian reservoir by the year 2000 . Descendants of these viruses have caused the current global panzootic and have achieved sustained mammal-mammal transmission in farmed and wild mammals. Our findings demonstrate that viral evolution in no...

  Highlights •  MERS-CoV structural proteins and ORFs potently induce T cell responses in mice •  MERS-CoV-specific T cell epitope repertoires are identified in C57BL/6 and BALB/c mice •  Airway ORF4b208-CD4+ and ORF5167-CD8+ T cells are optimal effector T cells •  ORF4b208 and ORF5167-specific T cells protect mice against MERS-CoV infection Summary Since its initial emergence in 2012, MERS-CoV has remained endemic and a global health threat . While accessory proteins (ORFs) are known for immune evasion , their role in adaptive immunity is unexplored. This study systematically investigated T cell responses against MERS-CoV ORFs. We mapped epitope repertoires targeting structural proteins and ORFs in C57BL/6 and BALB/c mice , revealing that ORFs potently induced virus-specific T cells . Notably, ORF5 induced the dominant CD8+ T cell responses in BALB/c mice. Further analysis revealed that ORF4b208-specific CD4+ and ORF5167-specific CD8+ T cells in the respiratory...

  Abstract Newly emerging influenza virus strains pose a constant threat as they encounter a population lacking neutralizing antibodies against the new strain . However, cross-reactive non-neutralizing antibodies (nnABs) may be present and assist in mitigating disease symptoms via various effector mechanisms, including antibody-dependent cellular cytotoxicity (ADCC). Although nnABs to influenza virus have received more attention lately , little information is available on their age-related prevalence , steady-state levels, functional properties , and changes in these parameters over time. Using longitudinal samples from adolescents, adults, and older adults , collected before and after the 2009 swine flu pandemic , we comprehensively characterized the specificity and functionality of nnAB responses against H1N1 pandemic 2009 (H1N1pdm09) virus . Remarkably, all participants exhibited cross-reactive antibodies to this virus before having encountered it through infection or vaccinatio...

  {Summary} In summer 2025, the SARS-CoV-2 JN.1 sublineage became dominant with more resistant variants, such as XFG, LP.8.1, and NB.1.8.1 . COVID-19 mRNA boosters were therefore updated for the 2025–2026 season to target the LP.8.1 spike .1 Previous boosters, particularly the WA1/2020+BA.5 bivalent booster , were characterised by substantial boosting of the ancestral strain , a phenomenon known as immune imprinting .2,3 We therefore evaluated whether the phylogenetically more distant LP.8.1 mRNA booster would preferentially boost currently circulating strains. Recent data from European and Asian populations have reported the immunogenicity of the LP.8.1 mRNA booster .4,5 Herein, we report the immunogenicity of the LP.8.1 mRNA booster in a US population with a different exposure history and high population immunity. We show that the LP.8.1 mRNA booster induced neutralising antibody (NAb) and binding antibody responses, primarily to the vaccine-matched L.P.8.1 variant and other curr...

  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...