npj Vaccines

Suboptimal rotavirus vaccine effectiveness in low- and middle-income countries (LMICs) highlights the need for next-generation vaccines, such as the neonatal RV3-BB vaccine. However, there is uncertainty in the duration of protection and future price of vaccines in development. We aim to identify the conditions under which switching to RV3-BB is optimal in Malawi and Ghana, where the current immunization programs use 2-dose Rotarix and 3-dose Rotavac schedules, respectively. A full incremental cost-effectiveness analysis was performed using a validated transmission model calibrated to country-specific rotavirus data. Over 2025-2034, introducing RV3-BB resulted in the largest rotavirus-related burden reduction compared with the current country-specific programs. At moderate willingness-to-pay (~0.5 time Gross Domestic Product per capita), RV3-BB was preferred over Rotavac if price per dose was <$1.2 in Malawi and <$2.5 in Ghana, and/or if the average duration of protection exceeded 40 weeks in Malawi. The RV3-BB vaccine is likely to be cost-effective in Malawi and Ghana, as well as other LMICs, based on expected pricing and duration of protection.

Adenoviruses (Ads) are widely used as vaccine vectors. However, pre-existing immunity to commonly used serotypes, like Ad5, can reduce vaccine immunogenicity, with neutralizing antibody titers >200 previously shown to impact vaccine efficacy. Gorilla adenovirus (GRAd) vectors have been developed to evade pre-existing anti-vector responses, but their seroprevalence in southern Africa is poorly defined. Here, we assessed seroprevalence to GRAd32, Ad26 and Ad5 before (baseline) and after COVID-19 vaccination, in cohorts from South Africa and Zimbabwe. Sera from South African participants enrolled in the Sisonke sub-study (n=100, prior to Ad26.COV2.S vaccination) and the follow-up "Booster after Sisonke" (BaSiS) study (n=226) were tested for neutralizing antibodies to Ad5, Ad26, and GRAd32. These samples included paired pre/post boost samples for 27 donors. We also tested sera from the Zimbabwean Mutala cohort (n=131, of which 44 were unvaccinated, and 87 vaccinated with inactivated vaccines). Participants living with HIV (PLWH) comprised 30-50% of each cohort. In the pre-vaccination samples from the Sisonke cohort, geometric mean titers (GMT) for anti-GRAd32, Ad26, and Ad5 antibodies were 78, 142, and 459, with neutralization titers >200 observed in 14%, 32%, and 68% of participants, respectively. Similarly, in the unvaccinated participants in the Mutala cohort, GMTs for GRAd32, Ad26, and Ad5 were 117, 245, and 536, with neutralizing titers >200 in 22%, 42%, and 69% of participants. We observed no significant difference in Ad antibody titers between PLWH and those living without HIV. We next assessed the impact of COVID-19 vaccination on titers. Vaccination with inactivated COVID-19 vaccines (Sinopharm/Sinovac) did not significantly affect Ad5, Ad26 or GRAd32 titers in an unpaired analysis. In contrast, [~]9 months after Ad26.COV2.S vaccination, anti-Ad26 titers for longitudinally sampled participants (n=27) increased 10-fold from a GMT of 141 to 1,426. By comparison, GRAd32 responses were not significantly altered by Ad26.COV2.S vaccination, while anti-Ad5 responses showed a modest <2-fold increase. Our data support previous findings that, whereas anti-Ad5 neutralizing antibody responses are commonly detected globally, GRAd32 responses are less frequent. Importantly, GRAd32 neutralizing responses remained unchanged after Ad26.COV2.S vaccination. HIV status had no significant effect on antibody titers. These results support the use of the GRAd32 vector in upcoming HIV vaccine trials, including in regions where Ad26-based COVID-19 vaccines were widely deployed.

Background: The isolation of many HIV broadly neutralizing antibodies (bnAbs) from people living with HIV (PLWH) and rigorous characterization of their ontogeny has promoted the goal of reverse engineering their natural development as a strategy for achieving an effective preventive HIV vaccine. We previously described the developmental process of CH103, a CD4-binding site (CD4bs)-specific monoclonal antibody, and the associated evolution of HIV Envelopes (Envs) within the person (CH505) from whom it was isolated. A series of monomeric gp120 protein subunit immunogens representing the transmitted founder (TF) and Envs that evolved during infection and optimally reacted with lineage members at each step of the CH103 clone maturation path were evaluated in this placebo controlled randomized vaccine trial to test for the first time in humans the concept of whether sequential immunization with gp120 monomeric proteins can recapitulate the development of CD4bs B-cell clonal lineages, including CH103. Methods: HIV Vaccine Trials Network 115 (HVTN 115) was a randomized placebo-controlled vaccine trial at US clinical research sites. We tested the safety and immunogenicity of CH505TF gp120 + GLA-SE (Part A), and then the ability of sequential CH505 gp120 proteins (corresponding to CH505s weeks 53 and 78 Envs) + GLA-SE immunizations to induce CD4bs-specific neutralizing antibodies (Part B). We assessed binding and neutralizing antibody responses, antibody dependent cellular cytotoxicity, antibody dependent cellular phagocytosis, T-cell responses and B-cell phenotyping. Results: We enrolled 42 participants between October 2017 and May 2018 for Part A, and 65 participants from December 2020 to October 2022 for Part B. Immunization with the CH505 gp120 proteins adjuvanted with GLA-SE was well tolerated and induced CD4bs-specific B cells and Env-specific plasma antibodies. The plasma neutralizing antibody response was limited to primarily tier 1 autologous and heterologous HIV-1 strains. Blood-derived B-cell repertoire analyses identified CD4bs antibodies that preferentially bound to open-occluded trimeric Envs that exist in an intermediate state between prefusion-closed to CD4-bound open confirmations, consistent with tier 1 HIV neutralizing activity. Conclusions: Together, these results suggest that the low-affinity CH505TF gp120 monomer elicited CD4bs antibodies in the sera and B-cell repertoires of humans. However, our findings also indicate that gp120 monomers are insufficient to induce detectable bnAb precursors to epitopes on native Env trimers. Nonetheless, our data provide a benchmark for comparison with ongoing clinical trials testing high-affinity CH505 Env trimers for induction of CD4bs bnAb precursors.

Human adenovirus serotype 4 (Ad4) is used as a replication-competent oral vaccine that safely and effectively prevents Ad4 respiratory illness in US military personnel. Recombinant Ad4 vaccine candidates elicit mucosal and systemic immune responses against respiratory viruses in hamsters, nonhuman primates, and humans. Although evaluation of Ad4 vaccine candidates in mice would be extremely useful given the large number of immunologic tools available, this has been limited by concerns about a lack of viral replication in these animals. Here we generated recombinant Ad4 vectors that express either luciferase (Ad4-Luc) or herpes simplex virus type 2 (HSV-2) glycoprotein D (Ad4-gD2) to identify transgene expression kinetics, the presence of Ad4 vector replication, and HSV-2 immune responses and protection against HSV-2 infection. Local luciferase activity was observed from 7 hours to 20 days after intranasal inoculation of BALB/c and humanized mice. Subsequent inoculations with Ad4-Luc showed reduced luciferase expression in BALB/c mice, but robust expression in humanized mice, suggesting an immune response to the vector in wild-type mice. Ad4 DNA, but not luciferase activity, was reduced in the lungs of BALB/c mice treated with cidofovir before inoculation with Ad4, implying that Ad4 replicated, albeit at a low level, in the lungs. Intranasal vaccination of mice with Ad4-gD2 resulted in HSV-2 neutralizing antibody in the serum, and after HSV-2 intravaginal challenge reduced disease scores, increased survival, and reduced shedding. Overall, the BALB/c mouse model is semi-permissive to Ad4 mucosal infection, but transgene expression is sufficient for the study of Ad4-based vaccine candidates. ImportanceMucosal surfaces serve as the primary site of infection and shedding for many viral pathogens. Immune responses at mucosal sites provide protection, but few mucosal vaccines are licensed. The oral replication-competent adenovirus serotype 4 (Ad4) vaccine is used to prevent respiratory illness in military recruits, has an extraordinary record of safety and efficacy and has been tested as a recombinant platform for other viruses. Further development of this vaccine platform has been partially hindered by the perceived inability to evaluate vaccine candidates in mice. Here we characterize recombinant Ad4 transgene expression kinetics and viral replication after inoculation at various sites and show protection against herpes simplex virus type 2 (HSV-2) genital disease in mice after intranasal vaccination. We show that Ad4 can induce protective efficacy, even in a semi-permissive mouse model, suggesting this is a promising vector for HSV-2 and potentially other viral pathogens.

Activation of innate immunity enhances adaptive immune responses and underlies the mechanism of action of vaccine adjuvants. First-generation mRNA vaccines lack a dedicated adjuvant, relying instead on the self-adjuvanting properties of lipid nanoparticles (LNPs), and are thus expected to benefit from additional engagement of innate immune pathways not activated by LNPs. Among innate immune modulators, TLR5 agonists are particularly promising adjuvants due to their ability to induce a balanced Th1/Th2 immune response and their relatively favorable safety profile. Here we tested whether supplementing mRNA vaccines with mRNA encoding a TLR5 agonist could enhance immunization efficacy by induction of TLR5 signaling coordinated with vaccine antigen expression. We designed FL711, an mRNA encoding a derivative of Salmonella flagellin optimized for mammalian expression, functionally active in TLR5 signaling, deimmunized for pre-existing human T and B cell epitopes, and engineered for secretion to stimulate the TLR5 pathway in the local tissue microenvironment. We characterized FL711 in vitro and in vivo for functional and pharmacological parameters and assessed its adjuvant effect as a component of experimental anti-influenza and anti-SARS-CoV-2 mRNA vaccines. Supplementation with small amounts of FL711 mRNA (up to 30-fold less than antigen-encoding mRNA) significantly enhanced vaccine immunogenicity and protective efficacy, stimulating local NF-{kappa}B induction, boosting antibody production and T cell activation, and prolonging the durability of the response -- while enabling a marked reduction in mRNA dose per vaccine. These findings support the potential of FL711 as a broadly applicable mRNA-encoded adjuvant to improve the potency, durability, and dose efficiency of next-generation mRNA vaccines.

Background Despite the significant impact of longstanding paediatric pneumococcal conjugate vaccine (PCV) use in the United Kingdom (UK), pneumococcal disease burden remains substantial and is primarily driven by nonPCV13 serotypes. Higher valent vaccines such as the 20 valent PCV (PCV20) may provide additional public health and economic benefits, yet their value in the contemporary UK setting has not been fully assessed using recent data. Methods We updated an age structured dynamic transmission model using post COVID 19 UK epidemiology (2001 to 2023) to compare pediatric PCV20 with PCV13 and PCV15. Over a 10 year horizon, we assessed cost effectiveness and number needed to vaccinate (NNV), capturing invasive and non invasive disease cases, deaths, costs, quality adjusted life years, and incremental cost effectiveness ratios. PCV20 was evaluated under 1+1 and 2+1 schedules; PCV13 and PCV15 were assessed under 1+1. Scenario analyses examined key uncertainties. Results PCV20 was estimated to avert more cases and deaths than PCV13 or PCV15, driven by broader serotype coverage and indirect effects. Both PCV20 schedules were dominant or cost saving versus lower valent comparators, with lower NNVs. PCV20s higher vaccination costs were offset by reductions in downstream healthcare expenditures. Conclusion Paediatric PCV20 implementation in the UK could deliver substantial health gains while improving economic efficiency, supporting timely adoption.

Vaccines to prevent infant group B streptococcus (GBS) disease are advancing, with licensure likely based on safety and immunologic endpoints rather than clinical efficacy data. This approach requires robust, generalisable serological thresholds of risk reduction (SToRRs). We combined data from six case-control studies in Europe and Africa to define SToRRs for early-onset (EOD) and late-onset (LOD) GBS disease. Across diverse epidemiological and healthcare settings, anti-capsular polysaccharide IgG concentrations were consistently higher in infants who remained disease free than in those who developed disease. Higher antibody concentrations were required to reduce the risk of EOD than LOD, and higher concentrations were required for serotype Ia than for serotype III. This study provides a quantitative framework to support correlates-based evaluation and potential licensure of maternal GBS vaccines.

Tuberculosis (TB) remains a leading cause of death from infectious disease worldwide, underscoring the urgent need for vaccines with greater and more consistent efficacy than Bacille Calmette-Guerin (BCG). We previously showed that mucosal vaccination with an isogenic Mycobacterium tuberculosis (Mtb) mutant lacking the stress-response transcription factor SigH ({Delta}sigH) prevents pulmonary TB in two macaque species. In the absence of SigH, Mtb is unable to effectively counter host oxidative stress. Vaccinated macaques were notably protected from disease, exhibiting an absence of granulomatous pathology together with the formation of lymphoid follicles and robust antigen-specific CD4+ and CD8+ T cell responses, identifying{Delta} sigH as a promising live-attenuated TB vaccine candidate. The{Delta} sigH mutant used in prior studies was generated in the Mtb CDC1551 background, a commonly used laboratory strain for challenge studies. Here, we evaluated whether{Delta} sigH-mediated protection extends to heterologous challenge with the more virulent Mtb Erdman strain. Mucosal{Delta} sigH vaccination conferred significant protection against heterologous challenge, markedly reducing pulmonary bacterial burden and TB-associated pathology. Longitudinal high-resolution PET/CT imaging demonstrated that aerosol{Delta} sigH vaccination induced robust inducible bronchus-associated lymphoid tissue (iBALT) responses in the lung. Unlike granulomas, these iBALT structures resolved over time while remaining associated with protection against subsequent Mtb challenge. Protection of highly susceptible rhesus macaques against virulent heterologous Mtb challenge following aerosol{Delta} sigH vaccination supports the further preclinical development of{Delta} sigH-based live-attenuated TB vaccines and highlights iBALT induction as a potential correlate and mechanistic driver of protective immunity against TB.

The emergence of the SARS-CoV-2 Omicron BA.2.86 subvariant, a lineage derived from the BA.2 strain, led to the 2024-2025 COVID-19 vaccine update to include KP.2 or related JN.1-lineage spike antigens. We evaluated the magnitude, breadth, and durability of humoral immune responses following a single KP.2 vaccine dose in a longitudinal cohort of 21 individuals up to six months. KP.2 vaccination increased spike-specific binding and neutralizing antibodies against the ancestral WA1 strain, alongside the BA.5, XBB.1.5, and KP.2 variants. Power law modeling estimated half-lives for WA1- and KP.2-specific IgG responses at 770 and 248 days, respectively. Additionally, the KP.2 dose increased IgG1 and IgG4 subclasses more than IgG2 and IgG3 responses to both spike proteins. Serum depletion experiments using WA1 or KP.2 proteins demonstrated most vaccine-elicited antibodies were cross-reactive. Consequently, KP.2 vaccine-induced antibodies retained broad neutralizing activity against recently circulating Omicron subvariants (BA.2.86, KP.3.1.1, XEC, LP.8.1, LF.7, XFG.3.12, PQ.1, BA.3.2.1, and RE.2). Using a live virus neutralization assay, XFG.3.12 showed the greatest reduction in neutralizing titers relative to KP.2 (4.2-fold). In a small subset, an LP.8.1 vaccine dose increased neutralizing activity against the matched variant while maintaining WA1 and KP.2 cross-reactivity, but only modestly increased antibodies to divergent variants BA.3.2.1 and RE.2. Ultimately, these data indicate the KP.2 mRNA vaccine generates durable, cross-reactive responses against current Omicron subvariants. However, ongoing spike evolution impacts neutralization of emerging lineages, highlighting the need for continued viral monitoring and timely vaccine updates. IMPORTANCESARS-CoV-2 continues to evolve, raising ongoing concerns about how well updated vaccines protect against emerging variants. This study evaluates antibody responses after KP.2 spike mRNA vaccine dose and shows that a single dose induces durable and broadly cross-reactive immunity against both earlier strains and recently circulating Omicron subvariants. Despite this breadth, reduced neutralizing activity against certain emerging variants indicates that ongoing antigenic changes can impact vaccine induced antibody effectiveness. These findings provide insight into how current vaccines perform over time and highlight the need to track viral evolution and update vaccine antigens to maintain broad protection against severe disease, hospitalization, and death.

Introduction: Adenoviral vectors such as chimpanzee ChAdOx1 were selected for COVID-19 vaccines due to their low seroprevalence in humans, minimizing the impact of neutralising anti-vector immunity that could attenuate vaccine responses. However, the influence of pre-existing adenoviral immunity on vaccine response remains incompletely understood. We have previously shown that SARS-CoV-2 spike-specific T cells were enhanced in ChAdOx1 nCoV-19 vaccinated immunodeficient patients compared to mRNA-based BNT162b2. Here, we assess immune cross-reactivity between ChAdOx1 and human adenovirus 5 (HuAd5), and test the hypothesis that in antibody-deficient individuals, cross-neutralisation may be impaired, allowing bystander enhancement of SARS-CoV-2 spike-specific T cell responses following ChAdOx1 nCoV-19 vaccination. Methods: We studied healthy healthcare workers (HCWs) and immunodeficient patients (IDPs) who received homologous ChAdOx1 nCoV-19 or BNT162b2 vaccines. HCWs samples were collected pre-vaccination and 4-6 weeks after the second dose, while IDP samples were obtained 4-6 weeks after the second dose. Serum anti-HuAd5 hexon IgG was quantified using a Luminex multiplex assay, and neutralizing antibodies were assessed using a replication-deficient HuAd5-GFP virus neutralization assay with flow cytometry readout. Ex vivo ELISpot and flow cytometry assays were used to measure T cell responses to HuAd5 hexon. These data were compared with previously published ChAdOx1 nCoV-19 vaccine responses in the same cohorts. Results: HuAd5 hexon-binding IgG titres were significantly higher in ChAdOx1 nCoV-19 compared to BNT162b2 vaccine recipients in both HCWs (p = 0.0043) and IDPs (p = 0.0328). Within ChAdOx1 nCoV-19 vaccine group, titres were lower in IDPs than HCWs (p = 0.0015) but not within the BNT162b2 group (p = 0.1261). HuAd5 neutralisation titres did not differ between cohorts or vaccine groups. In ChAdOx1 nCoV-19 vaccinated IDPs and HCWs, there was a significant negative correlation between HuAd5 hexon IgG titres and SARS-CoV-2 spike-specific T cell responses. Similarly, HuAd5 neutralisation titres showed an inverse correlation with spike-specific T cell responses in ChAdOx1 nCoV-19 vaccinated IDPs and HCWs. ChAdOx1 nCoV-19 vaccination induced significantly higher frequencies of HuAd5 hexon-reactive T cells compared with BNT162b2 vaccination in IDPs (p < 0.0001), consistent with cross-reactive adenoviral T cell responses. In IDPs, HuAd5 hexon-specific T cell frequencies positively correlated with SARS-CoV-2 spike-specific T cell responses following ChAdOx1 nCoV-19 vaccination but not following BNT162b2 vaccination. Functional profiling in ChAdOx1 nCoV-19 vaccinated IDPs demonstrated expansion of HuAd5 hexon-specific CD4IFN-{gamma}TNF T cells in high SARS-CoV-2 spike responders (p = 0.0002) compared to low responders, and the frequency of these cells strongly correlated with spike-specific T cell response. Discussion: ChAdOx1 nCoV-19 has been associated with stronger T cell responses than BNT162b2 in certain populations, including immunodeficient and elderly individuals. While this has been attributed to antigen persistence and innate adjuvant effects, our findings support a mechanism whereby heterologous pre-existing adenovirus immunity modulates vaccine-induced responses. Specifically, cross-reactive HuAd5-specific T cells may enhance spike-specific T cell responses via bystander enhancement, while cross-reactive binding antibodies may exert opposing effects. An implication of this study is that vaccine protocols could incorporate therapies that suppress vector-specific or cross-reactive antibodies while preserving T cell responses especially in cases where T cell-specific responses are most desirable. Also, safe vector-based vaccines can be developed for patient groups with predominant antibody deficiency. Targeted vaccination strategy could be implemented for clinical cohorts based on immune competence.

PRV-101 is a multivalent formalin-inactivated Coxsackievirus B (CVB) vaccine developed to prevent CVB infections, which are associated with increased risk of islet autoimmunity. While PRV-101 induces robust neutralizing antibody responses, its T-cell immunogenicity is unknown. We analyzed peripheral blood mononuclear cells from 25 healthy adults receiving three high or low PRV-101 doses or placebo in a Phase I randomized, placebo-controlled trial. CVB-reactive CD8 T-cell responses were assessed using HLA Class I multimers, and CD4 and T follicular helper (Tfh) responses were measured by activation-induced marker assays following stimulation with a CVB peptide library. PRV-101 elicited minimal CVB-reactive CD8 T-cell responses but robust CD4 and Tfh responses, peaking at week 12 and persisting through week 32. Responses were observed in both seronegative and seropositive individuals, consistent with effective immune priming and boosting. Tfh frequencies correlated with neutralizing antibody titers. Female participants exhibited higher peak Tfh responses than males. We conclude that PRV-101 elicits a CVB-protective immune profile, dominated by Tfh responses supporting durable humoral immunity and devoid of potentially diabetogenic cytotoxic T-cell responses. This profile invites further investigations in vaccine trials for type 1 diabetes prevention.

Background: The ongoing Bundibugyo virus disease (BDBV) outbreak in Central Africa highlights the absence of approved vaccines specifically targeting BDBV. Whether licensed Zaire ebolavirus (EBOV) vaccines induce cross-reactive immunity against BDBV remains largely unknown. Methods: We performed an immunogenicity analysis using serum samples from participants enrolled in the PREVAC randomized clinical trial evaluating licensed Ebola vaccine strategies in West Africa. Samples collected at day 28 (D28) and month 3 (M3) following vaccination with rVSV{Delta}G-ZEBOV-GP or Ad26.ZEBOV/MVA-BN-Filo were assessed using a multiplex Luminex assay against glycoproteins from multiple filoviruses, including EBOV Kikwit, EBOV Mayinga, BDBV, Sudan virus, Reston virus, and Marburg virus. Results: A total of 179 samples were analysed. Detectable cross-reactive antibody responses against BDBV were observed across vaccine groups, timepoints, and age categories. However, BDBV responses remained substantially lower than homologous EBOV responses. In rVSV recipients, median BDBV responses (net MFI) reached 282 (IQR 164-644) at D28 compared with 1788 (832-3311) against the homologous Kikwit antigen. Similar patterns were observed following rVSV booster vaccination and Ad26.ZEBOV/MVA-BN-Filo vaccination. The heterologous Ad26/MVA regimen demonstrated increasing BDBV responses between D28 and M3. Conclusions: Licensed EBOV vaccines induced detectable but quantitatively reduced cross-reactive antibody responses against BDBV. Although no direct assessment of vaccine efficacy against BDBV disease was possible, these findings support the plausibility of partial heterologous immunity following EBOV vaccination. In the absence of approved BDBV-specific vaccines, these data support the urgent evaluation of currently available Ebola vaccines during BDBV outbreaks and reinforce the importance of developing broadly protective pan-filovirus vaccines.

Adeno-associated virus (AAV)-delivered anti-HIV-1 broadly neutralizing antibodies (bNAbs) could prevent and treat HIV-1 infection but are limited by host immune responses, specifically anti-drug antibodies (ADA). We tested whether PD-L1-mediated immune shielding could improve the consistency of AAV-delivered bNAb expression from muscle tissue in rhesus macaques. AAV9.PD-L1 co-delivery with AAV9.3BNC117 reduced the occurrence of ADA and T cell responses and improved the durability of 3BNC117 expression for one year post administration. Importantly, 5 of 6 macaques that received co-delivered AAV9.PD-L1 vectors were protected against ten repeated SHIVAD8-EO challenges. Histopathological and spatial transcriptomic profiling showed that AAV9.PD-L1 co-delivery prevented severe local inflammation, muscle injury, and tertiary lymphoid structure formation at the administration site. Thus, immune shielding could serve as a strategy to prolong transgene expression from muscle-directed AAV-delivered biologics.

Despite concerns about the spread and pandemic potential of H5N1, there is no commercial H5N1 vaccine. Seasonal influenza vaccines offer some cross-protection against H5N1, but to date there has been no consideration of whether protection differs between the sexes. We investigated immune responses and protection in adult male and female C57BL/6 mice following vaccination with either inactivated H1N1 or H5N1 (LAIV backbone) virus vaccines. Vaccination induced strong homologous antibody responses, with females generating greater total IgG than males against both H1N1 and H5N1 vaccine, which was primarily mediated by greater IgG responses to neuraminidase (NA) than hemagglutinin (HA) protein. IgG cross-recognition of H1N1 also was greater among H5N1 vaccinated females and was primarily caused by greater IgG responses to N1. IgG2b and IgG2c were the primary isotypes generated in response to these vaccines, with females having greater IgG2b responses and greater binding to Fc{gamma}RIV for avian and human NA than males in response to both homologous and heterologous vaccination. Antibody-dependent complement deposition was measured as an FcR-mediated non-neutralizing response against HA and NA and was robust in both sexes. Vaccinated females had greater neutralizing antibody titers than males against the homologous vaccine virus, with limited cross-neutralizing antibodies detected in either sexes. Neuraminidase inhibition titers were greater in vaccinated females than males against the heterologous virus following H1N1 vaccination and against both the vaccine and heterologous viruses following H5N1 vaccination. When H1N1 and H5N1 vaccinated mice were challenged with a lethal dose of A/Texas/37/2024 H5N1, all H5N1 vaccinated mice were protected, regardless of sex. Among H1N1 vaccinated mice, while both sexes were protected against disease, H1N1 vaccinated females cleared virus faster than their male counterparts. These findings highlight that female-biased NA-specific antibodies result in greater cross-protection and should be considered in studies of influenza vaccines. HighlightsO_LIFemales mount stronger IgG responses than males to both H1N1 and H5N1 vaccines C_LIO_LISex differences in vaccine responses are driven by immunity to neuraminidase (NA) C_LIO_LINA inhibition titers are greater in females, supporting broader cross-protection C_LIO_LIH5N1 vaccination confers full protection in both sexes against lethal H5N1 challenge C_LIO_LIH1N1-vaccinated females clear H5N1virus faster than males after lethal challenge C_LI

Chronic hepatitis B cure requires the inactivation and/or elimination of covalently closed circular DNA (cccDNA), together with silencing of integrated viral genomes and restoration of HBV-specific immune responses. The TLR1/2 agonist Pam3CSK4 has previously been identified as a potent direct anti-HBV agent in vitro. In the present study, we engineered a liver-targeting polymeric nanoparticle formulation of Pam3CSK4 to enhance its in vivo immunostimulatory and antiviral activity. We evaluated the antiviral efficacy of this novel nanoformulation carrying the TLR1/2 agonist (NP-Pam3CSK4) in monotherapy and started to investigate its mechanism of action through immunological correlates in an immune-competent AAV-HBV mouse model. AAV-HBV-infected mice received intravenous administrations of NP-Pam3CSK4 at doses of 5 or 20 g twice per treatment cycle over four cycles, followed by a 2-week follow-up period. Soluble Pam3CSK4 was administered at substantially higher doses (100 g). Serial blood samples were regularly collected to monitor virological and host immune parameters. At study completion, liver tissues were harvested for intrahepatic quantification of viral and immunological markers using immunoassays, quantitative PCR, and histological analyses. The most pronounced antiviral effects were observed in mice treated with NP-Pam3CSK4 formulations, which achieved greater viral suppression than free Pam3CSK4 despite markedly lower administered doses. Histological examination of liver biopsies from treated animals revealed prominent immune cell infiltration, including macrophages, monocytes, and T cells, organized in dense cluster-like structures. These findings support the induction of coordinated innate and adaptive immune responses contributing to HBV control and clearance. Collectively, our results demonstrate that nanoparticle-based delivery of TLR1/2 agonist represents a promising therapeutic strategy for chronic HBV infection and may improve the likelihood of achieving functional cure. Further mechanistic and translational studies (combination) are warranted to support clinical development.

To better understand how pregnancy impacts humoral immunity, we conducted an in-depth longitudinal analysis of the kinetics and characteristics of vaccine responses in a prospective cohort of pregnant and non-pregnant women. Humoral immune responses observed among pregnant participants who received the mRNA-delivered SARS-CoV-2 vaccination, including their effector functions, were in some cases marginally lower than those among non-pregnant controls, while prior infection was associated with some potentiation in humoral responses. Importantly, vaccine-induced antibodies were efficiently transferred across the placenta, providing the fetus with passive immunity and underscoring the dual benefit of maternal vaccination for both mother and neonate against COVID-19. Delayed induction of spike-specific IgG4 following the primary two-dose vaccination series was observed in vaccine recipients, independent of pregnancy status. In a subset (n=6) of pregnant women whose spike-specific serum IgG repertoires were extensively profiled at the clonotypic level over time as part of another study, we proteomically identified secreted IgG clonotypes that had class-switched to IgG4. Matching of these clonotypes detected as IgG4 to those defined as SARS-CoV-2 spike-specific revealed that, while a minority of total clonotypes, they were elicited early in the immunization series and tended to be more highly mutated, more prevalent, and more persistent than clonotypes in the serological repertoire that were not detected as IgG4. Consistent with the increase in secreted vaccine-specific IgG4 over time, but its poorer placental transfer, these clonotypes were detected at greater levels in maternal but not cord blood at the time of delivery as compared to 28 days post the second vaccine dose. These findings indicate some impact in the kinetics, characteristics, and functions of the humoral response that may be associated with pregnancy-related immune modulation. Conservation of the late class-switch recombination to IgG4 that has previously been associated with mRNA-based SARS-CoV-2 vaccines raises questions about how different immunological states and vaccine components influence short- and long-term characteristics of the humoral immune response.

Ebola virus disease (EVD), caused by the Ebola virus (EBOV), is characterized by high morbidity and mortality, with 16 distinct EVD outbreaks reported in the Democratic Republic of the Congo (DRC), alone. As part of the formal response to the 2018 outbreaks in Equateur and North Kivu provinces, a recombinant vesicular stomatitis virus-Zaire Ebolavirus envelope glycoprotein vaccine (rVSV-ZEBOV-GP) vaccine was deployed under emergency use. While clinical trials have evaluated vaccine safety and efficacy, there is a paucity of real-world data documenting antibody durability for longer periods post-vaccination. Here, we present serologic data from 1081 individuals in Beni, North Kivu (n = 599) and Mbandaka, Equateur (n = 482) who were vaccinated during the outbreaks--with samples from baseline up to five-years following vaccination. Using a multiplexed immunoassay, we show sustained anti-EBOV GP reactivity: at year-5 collection, 72% of individuals naive at time of vaccination remained seroreactive to EBOV GP. Stratifying by site, antibody titers remained significantly elevated after baseline across all post-vaccination timepoints in both linear and logistic mixed-effects models. Pre-existing EBOV GP reactivity at baseline was the strongest independent predictor of antibody response in Mbandaka, associated with higher titers and greater odds of seropositivity (OR = 3.87, 95% CI: 2.50-6.01, p-value < 0.001), consistent with a boosting effect among previously exposed individuals. However, this was not replicated in Beni (OR: 0.66, 95% CI: 0.27-1.58, p-value = 0.348). In Mbandaka, among those recipients who reported receiving a booster dose, the odds of seroreactivity were 12.75-fold (p-value < 0.001) and 3.68-fold higher (p-value = 0.04) at 4.2 and 5-years post-vaccination, respectively, in comparison to odds of reactivity at three weeks following administration of the initial dose. Occupational groups with zoonotic or community-level exposure had trending lower odds of seroreactivity relative to healthcare workers, most consistently in Beni. Ultimately, these data indicate that five years following administration of the rVSV-ZEBOV-GP vaccine, most vaccinated individuals retain detectable anti-EBOV GP antibodies. While correlates of protection for EVD are not well established, sustained IgG seroreactivity to EBOV GP may serve as a marker for future understandings of the durability of and variation in immune responses to this high-consequence pathogen.

Adeno-associated virus (AAV)-delivered anti-HIV-1 broadly neutralizing antibodies (bNAbs) have demonstrated promise for preventing and treating HIV-1 infection in preclinical models. However, host immune responses, specifically anti-drug antibodies (ADA), limit sustained bNAb expression. We have previously shown that PD-L1-mediated immune shielding improves the consistency of AAV-delivered bNAb 3BNC117 expression from muscle tissue in rhesus macaques. Here, we test the breadth of this approach with another bNAb, 10-1074. We show that AAV9.PD-L1 co-delivery with AAV9.10-1074 reduced the occurrence of ADA responses and improved the durability of bNAb expression for one year post administration. Notably 12 of 12 macaques that received AAV9.10-1074 vectors were protected against ten repeated SHIVAD8-EO challenges. Histopathological profiling showed that AAV9.PD-L1 co-delivery prevented severe local inflammation and tertiary lymphoid structure formation at the administration site. Thus, immune shielding could serve as a broad strategy to prolong transgene expression from muscle-directed AAV-delivered biologics.

Background: Malaria exposure has been hypothesized to alter immune responses to childhood vaccines, but evidence is inconsistent. We evaluated whether early-life malaria exposure and perennial malaria chemoprevention (PMC) modify antibody responses to the 10-valent pneumococcal conjugate vaccine (PCV-10) among infants in a high malaria transmission setting in eastern Uganda. Methods: This study was nested within the MIC-DroP trial (NCT04978272) whereby 202 infants were selected for inclusion. Serotype-specific IgG concentrations were measured using an in-house multiplex seroassay from samples obtained at 8 and 24 weeks of age. Immunogenicity was quantified as the log10 fold-change in IgG concentration between the 8 and 24-week timepoints, and seroconversion as [&ge;]0.35 g/mL at week 24 (i.e., seropositive). Generalized estimating equation models were used to assess associations of PCV-10 immunogenicity and seroconversion with malaria exposure, malaria chemoprevention and birth outcomes. Results: Among the 195 of 202 infants who completed the three-dose PCV-10 series, neither infant PMC nor malaria exposure from study enrollment to 14 weeks were associated with PCV-10 immunogenicity or seroconversion. In contrast, low birthweight (<2500g) was associated with lower immunogenicity (82% lower antibody fold-change, p=0.003) and reduced odds of seroconversion (OR=0.19, p=0.003); preterm birth (<37 weeks) showed similar associations (79% lower antibody fold-change, p=0.018; OR=0.181, p=0.009). Conclusion: In this malaria-endemic setting, early-life malaria exposure and chemoprevention did not measurably alter PCV-10 antibody responses. However, low birthweight and prematurity were associated with reduced vaccine immunogenicity.

Age is a major determinant of disease severity following La Crosse virus (LACV) infection, yet the immunological mechanisms underlying heightened susceptibility in children remains poorly defined. Here, we show that acute LACV infection in weanling mice induces T cell dysfunction characterized by early PD-1 upregulation and impaired effector differentiation despite evidence of activation. This state is associated with reduced IL-2-dependent STAT5 signaling, indicating a failure to respond to available cytokine cues. Although regulatory T cells expand and exhibit elevated CD25 expression, their depletion increases IL-2 levels without restoring antiviral T cell responses or viral control. In contrast, PD-1 blockade partially restores T cell activation, and combined PD-1 blockade with CD25 targeting enables robust effector differentiation and improved viral control. These findings demonstrate that checkpoint signaling limits T cell responsiveness to IL-2, uncoupling activation from differentiation and driving age-dependent susceptibility to LACV infection.