Biology subjects

Cowling, B. J.

Publications and source records attributed to Cowling, B. J..

9 recordsLinked to original sources

bioRxiv2026

Defining influenza-specific B cells in vaccine responders, non-responders and influenza breakthrough infections

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 performed using 10x Genomics. Our results show that atypical B cells are the major subset of B cell responses in vaccine non-responders on day 28 post-vaccination. Conversely, individuals who seroconvert had diverse B cell phenotypes. The use of recombinant influenza-specific HA probes allowed us to dissect expression patterns on influenza HA-specific B cells. We found that HA-specific B cells of vaccine non-responders for A/H1N1 and A/H3N2 components displayed elevated atypical-like markers (CD11c, FcRL-5) at baseline, compared to responders. Analysis of differentially expressed genes (DEGs) between responders and non-responders identified differential expression of HLA-DR, CD74, CD83, and CXCR3 genes. We subsequently demonstrated reduced frequencies of HLA-DR-, CD74- and CD83-expressing B cells in patients hospitalized with influenza, compared to healthy participants. Hospitalized influenza patients also had significantly higher proportions of atypical CD21-CD27- B cells. Overall, our data demonstrate an association between elevated frequencies of atypical-like B cells with both lack of seroconversion following immunization and severe influenza infection. These findings broaden our understanding of humoral immunity in influenza vaccination and infection, providing novel insights for vaccination strategies and design.

immunology↗

bioRxiv2026

Strain-specific differences in the response to egg-derived versus recombinant protein influenza vaccines

The 2023/2024 influenza vaccine included an updated H1N1 component designed to better match a new clade of H1N1 that had multiple mutations in antigenic epitopes of hemagglutinin. Despite this update, the vaccine trended towards being less effective against the vaccine-matched H1N1 clade than the parental H1N1 clade lacking the new antigenic mutations. Here we measure neutralization titers of serum antibodies from individuals who had received either a recombinant protein or an egg-derived vaccine against a set of viruses with hemagglutinins from 58 H1N1 strains representative of the diversity during the 2023/2024 season. We find that egg-derived vaccine recipients, but not recombinant protein vaccine recipients, had a relatively lower boost in neutralizing titers to the new clade that the updated vaccine was designed to target. We suggest that the difference in the extent that the egg-derived versus recombinant protein vaccines boosted neutralizing titers to the new H1N1 clade is because the seed strain for the egg-derived vaccine strain had acquired a reversion of a key antigenic mutation (K142R) present in that clade. Our results show how egg-derived versus recombinant protein vaccines can elicit different relative titer boosts against different subsets of viral strains, a phenomenon that could impact vaccine effectiveness. ImportanceInfluenza vaccines can be produced from virus grown in eggs, or grown in cells or made with recombinant protein. Egg-derived influenza vaccines often contain egg-adaptive mutations in the viral antigen hemagglutinin (HA) which can impact the antigenicity or immunogenicity of the HA. In this study, we compare neutralization titers from egg-derived and recombinant protein vaccine recipients against recently circulating influenza A(H1N1) strains. We find that the egg-derived vaccine induces less of a boost in titers than the recombinant protein vaccine to the new clade of viral strains that the vaccine was designed to target.

immunology↗

bioRxiv2026

Near real-time data on the human neutralizing antibody landscape to influenza virus as of early 2026 to inform vaccine-strain selection

Twice each year, a decision is made on whether to update the strains included in the seasonal influenza vaccine to better match the most recent circulating viral strains. To characterize the antigenic properties of current seasonal influenza A strains to inform the upcoming decision about which strains to include in the 2026-2027 Northern Hemisphere vaccine, here we perform high-throughput sequencing-based neutralization assays using a library of 57 H3N2 and 34 H1N1 influenza hemagglutinins reflecting the circulating diversity of strains in late 2025 to early 2026. We assay this library against 302 human sera collected in late 2025. The resulting data set encompasses 27,409 titers, and provides a near real-time portrait of the human neutralizing antibody landscape against influenza virus. We find that many human sera have lower titers against the K subclade of H3N2 and the D.3.1.1 subclade of H1N1; these subclades have recently become dominant among their respective subtypes. Our measurements also reveal variability in titers to different subvariants within the K subclade of H3N2, with titers especially low to subclade K strains with additional mutations in antigenic regions D and E. We make all our data and accompanying visualizations publicly available to enable their use in vaccine-strain selection and analyses of influenza evolution and immunity.

microbiology↗

bioRxiv2025

Near real-time data on the human neutralizing antibody landscape to influenza virus to inform vaccine-strain selection in September 2025

The hemagglutinin of human influenza virus evolves rapidly to erode neutralizing antibody immunity. Twice per year, new vaccine strains are selected with the goal of providing maximum protection against the viruses that will be circulating when the vaccine is administered [~]8-12 months in the future. To help inform this selection, here we quantify how the antibodies in recently collected human sera neutralize viruses with hemagglutinins from contemporary influenza strains. Specifically, we use a high-throughput sequencing-based neutralization assay to measure how 188 human sera collected from Oct 2024 to April 2025 neutralize 140 viruses representative of the H3N2 and H1N1 strains circulating in humans as of the summer of 2025. This data set, which encompasses 26,148 neutralization titer measurements, provides a detailed portrait of the current human neutralizing antibody landscape to influenza A virus. The full data set and accompanying visualizations are available for use in vaccine development and viral forecasting.

microbiology↗

bioRxiv2024

A hemagglutinin and neuraminidase biased immunological memory shapes the dynamics of antibody responses to Influenza A virus

Influenza A virus (IAV) infection establishes a more diverse immunological memory to different viral proteins compared to vaccination. We hypothesized that the relative abundance of pre-existing immune memory to different viral antigens could skew post-infection antibody responses. To explore this, we generated mouse models with either an IAV hemagglutinin (HA)- or neuraminidase (NA)-biased immunological memory. We inoculated groups of mice with cocktails of isogenic viruses bearing antigenically-distinct HA (H3v) or NA (N2v) chosen to span the IAV H3N2 human circulation history. We challenged the mice with two H3N2 strains of opposing virulence and antigenic distance (AD) and examined the post-infection antibody landscapes. In both challenges, immune-naive mice seroconverted to both HA and NA whereas in primed mice, antibody response was detected to the antigen for which there is no pre-existing memory. In cases where the homologous antibody response was blunted, there was diversification on the breadth of response to antigenically-related strains with low baseline titers. Our findings clarifies the concept of "original antigenic sin" and demonstrate a mechanism by which the dynamics of antibody responses to HA and NA after infection can be altered by pre-existing immunity.

immunology↗

bioRxiv2024

High-throughput sequencing-based neutralization assay reveals how repeated vaccinations impact titers to recent human H1N1 influenza strains

The high genetic diversity of influenza viruses means that traditional serological assays have too low throughput to measure serum antibody neutralization titers against all relevant strains. To overcome this challenge, we have developed a sequencing-based neutralization assay that simultaneously measures titers against many viral strains using small serum volumes via a workflow similar to traditional neutralization assays. The key innovation is to incorporate unique nucleotide barcodes into the hemagglutinin (HA) genomic segment, and then pool viruses with numerous different barcoded HA variants and quantify infectivity of all of them simultaneously using next-generation sequencing. With this approach, a single researcher performed the equivalent of 2,880 traditional neutralization assays (80 serum samples against 36 viral strains) in approximately one month. We applied the sequencing-based assay to quantify the impact of influenza vaccination on neutralization titers against recent human H1N1 strains for individuals who had or had not also received a vaccine in the previous year. We found that the viral strain specificities of the neutralizing antibodies elicited by vaccination vary among individuals, and that vaccination induced a smaller increase in titers for individuals who had also received a vaccine the previous year--although the titers six months after vaccination were similar in individuals with and without the previous-year vaccination. We also identified a subset of individuals with low titers to a subclade of recent H1N1 even after vaccination. This study demonstrates the utility of high-throughput sequencing-based neutralization assays that enable titers to be simultaneously measured against many different viral strains. We provide a detailed experimental protocol (DOI: https://dx.doi.org/10.17504/protocols.io.kqdg3xdmpg25/v1) and a computational pipeline (https://github.com/jbloomlab/seqneut-pipeline) for the sequencing-based neutralization assays to facilitate the use of this method by others.

microbiology↗

bioRxiv2022

Environment-wide and epigenome-wide association study of adiposity in "Children of 1997" birth cohort

BackgroundIncreasing childhood adiposity is a global issue requiring potentially local solutions to ensure it does not continue into adulthood. We systematically identified potentially modifiable targets of adiposity at the onset and end of puberty in Hong Kong the most economically developed major Chinese city. MethodsWe conducted an environment-wide association study (EWAS) and an epigenome-wide association study of adiposity to systematically assess associations with body mass index (BMI) and waist-hip ratio (WHR) in Hong Kongs population-representative "Children of 1997" birth cohort. Univariable linear regression was used to select exposures related to adiposity at ~11.5 years (BMI n[≤]7,119, WHR n=5,691) and ~17.6 years (n = 3,618) at Bonferroni-corrected significance, and multivariable linear regression to adjust for potential confounders followed by replication (n=308) and CpG by CpG analysis (n=286) at ~23 years. Findings were compared with evidence from randomized controlled trials (RCTs) and Mendelian randomization (MR) studies. ResultsAt ~11.5 and ~17.6 years the EWAS identified 14 and 37 exposures associated with BMI, as well as seven and 12 associated with WHR respectively. Most exposures had directionally consistent associations at ~23 years. Maternal second-hand smoking, maternal weight, and birth weight were consistently associated with adiposity. Diet (including dairy intake and artificially sweetened beverages), physical activity, snoring, binge eating, and earlier puberty were positively associated with BMI at ~17.6 years, while eating before sleep was inversely associated with BMI at ~17.6 years. Findings for birth weight, dairy intake, binge eating, and possibly earlier puberty are consistent with available evidence from RCTs or MR studies We found 21 CpGs related to BMI and 18 to WHR. ConclusionsThese novel insights into potentially modifiable factors associated with adiposity at the outset and the end of puberty could, if causal, inform future interventions to improve population health in Hong Kong and similar Chinese settings. FundingThis study was supported by the Health and Medical Research Fund Research Fellowship, Food and Health Bureau, Hong Kong SAR Government (#04180097). The DNA extraction was supported by CFS-HKU1.

genetics↗

bioRxiv2018

Age-specific differences in the dynamics of protective immunity to influenza

Influenza A viruses evolve rapidly to escape host immunity, such that individuals can be infected multiple times with the same subtype. The form and duration of protective immunity after each influenza infection are poorly understood. Here, we quantify the dynamics of protective immunity against influenza A virus infections by fitting individual-level mechanistic models to longitudinal serology from children and adults in a household cohort study. We find that most protection in children is explained by antibody titers measured by the hemagglutination inhibition (HI) assay. In contrast, in adults, HI antibody titers explain a smaller fraction of protection. Protection against circulating strains wanes to approximately 50% of peak levels 3.5-7 years after infection in both age groups, and wanes faster against influenza A(H3N2) than A(H1N1)pdm09. Protection against H3N2 lasts longer in adults than in children. Our results suggest that the focus of influenza antibody responses changes over time from the highly mutable hemagglutinin head to other epitopes, consistent with the immunological theory of original antigenic sin, and that this change of focus might affect protection. Additionally, we estimate that imprinting, or primary infection with a subtype of one phylogenetic group, has little to no effect on the risk of non-medically attended influenza infections in adults. We also find no evidence of long-term cross-protection between subtypes. This work underscores the need for longitudinal data on multiple components of the immune response to better understand the development of immunity and differences in susceptibility within populations.

epidemiology↗

bioRxiv2017

Quantifying biases in test-negative studies of vaccine effectiveness

Test-negative designs have become commonplace in assessments of seasonal influenza vaccine effectiveness. Vaccine effectiveness is measured from the exposure odds ratio (OR) of vaccination among individuals seeking treatment for acute respiratory illness and receiving a laboratory test for influenza infection. This approach is widely believed to correct for differential healthcare-seeking behavior among vaccinated and unvaccinated persons. However, the relation of the measured OR to true vaccine effectiveness is poorly understood. We derive the OR under circumstances of real-world test-negative studies. The OR recovers the true vaccine direct effect when two conditions are met: (1) that individuals vaccination decisions are uncorrelated with exposure or susceptibility to infection, and (2) that vaccination confers \"all-or-nothing\" protection (whereby certain individuals have no protection while others are perfectly protected). Biased effect size estimates arise if either condition is unmet. Such bias may suggest misleading associations of the OR with time since vaccination or the force of infection of influenza. The test-negative design may also fail to correct for differential healthcare-seeking behavior among vaccinated and unvaccinated persons without stringent criteria for enrollment and testing. Our findings demonstrate a need to reassess how data from test-negative studies are interpreted for policy decisions conventionally based on causal inferences.

epidemiology↗