npj Vaccines

Vaccines targeting outer surface protein A (OspA) of Borrelia burgdorferi protect against Lyme disease by inducing antibodies in the host that neutralize spirochetes in the Ixodes scapularis tick midgut during engorgement before transmission occurs. We evaluated whether heterologous vaccination enhances protection compared to homologous delivery of the immunogen. C3H-HeN mice were immunized with a parainfluenza virus 5 vector (PIV5) containing a modified OspA protein (OspABPBPk) using three prime-boost immunization regimens: homologous PIV5 intranasal/intranasal (IN/IN), homologous rOspABPBPk (protein) subcutaneous/subcutaneous (SC/SC), or heterologous intranasal PIV5-ABPBPk/subcutaneous rOspABPBPk (IN/SC). Immunized mice were then challenged with nymphal I. scapularis ticks infected with 19 strains of B. burgdorferi three months post-prime vaccination. Three weeks after the last day of tick challenge, blood and tissues were collected from euthanized mice. All OspA-containing regimens elicited strong systemic IgG antibody responses that exceeded established protective thresholds. Vaccination markedly reduced B. burgdorferi loads in engorged nymphal ticks. Homologous IN/IN and SC/SC regimens produced the lowest geometric mean flaB burdens in nymphs (2.6 x 103 and 1.8 x 103 copies, respectively), corresponding to [~]1.8-2.0 log10 reductions relative to controls; the heterologous IN/SC regimen produced a more modest reduction ([~]1.7 log10; p = 0.0071 vs IN/IN, p = 0.0003 vs SC/SC). Across all vaccinated groups, no systemic infection with B. burgdorferi was observed as evidenced by absence of motile spirochetes in cultures from tissues, although one mouse (1/9, 11%) in the heterologous IN/SC regimen, had evidence of increased pepVF seroconversion and low-level flaB DNA in culture. Thus, homologous regimens yielded more consistent protective immunity with absent of signs of B. burgdorferi dissemination, suggesting that high systemic anti-OspABPBPk IgG antibody titers, rather than alternate immunization routes, were associated with the most consistent protection outcomes. PIV5-ABPBPk is a promising vaccine candidate for development of next-generation homologous or heterologous human Lyme disease vaccines.

An efficacious blood-stage malaria vaccine would serve as a highly useful public health tool alongside licensed vaccines targeting the pre-erythrocytic life cycle stage of the Plasmodium falciparum parasite. RH5 is the leading blood-stage malaria vaccine candidate antigen due to its highly-conserved sequence and non-redundant role in merozoite invasion of red blood cells. Following encouraging immunogenicity data in UK and Tanzanian Phase Ia/b vaccine trials, RH5-based vaccines have progressed to Phase IIb evaluation in Burkina Faso in recent years. Here, we report a Phase Ia clinical trial in malaria-naive UK adults to assess the safety and immunogenicity of the malaria vaccine candidate RH5.1 soluble protein with Matrix-M adjuvant using two different booster dosing regimens: 10-10-10 micrograms versus 50-50-10 micrograms RH5.1, both delivered in a 0-1-6-month schedule with 50 micrograms Matrix-M adjuvant per dose (ClinicalTrials.gov NCT06141057). A total of n=24 participants were recruited to this study, with n=23 completing all follow-up visits through to 1 year following final vaccination. The RH5.1/Matrix-M formulation was well-tolerated in this population, with injection site pain, myalgia and fatigue being the most commonly reported symptoms up to 7 days post-vaccination. There were no serious adverse events, adverse events of special interest, or suspected unexpected serious adverse reactions reported over the course of the trial. Both vaccination regimens were similarly immunogenic; no differences were observed in peak anti-RH5.1 serum IgG concentrations, in vitro functional anti-parasitic activity, avidity, or durability. Our findings build on other observations from clinical trials of adjuvanted RH5.1 indicating that humoral immunogenicity can be enhanced by delaying the final booster vaccination, but that there is limited impact of fractionation of the final dose. These insights can help to guide the next steps of multi-antigen, multi-stage malaria vaccine development in malaria-endemic settings.

Typhoid fever remains a significant public health challenge in low- and middle-income countries. In 2018, The World Health Organization recommended a single dose typhoid conjugate vaccine (TCV) for routine immunization in endemic settings; however, evidence guiding booster doses remains limited. Homologous TCV booster doses have demonstrated immune boosting. This study assessed the immunogenicity and safety of a heterologous booster using a Vi capsular polysaccharide-CRM197 TCV (Vi-CRM) administered 5-6 years after primary vaccination with a Vi capsular polysaccharide tetanus toxoid TCV (Vi-TT) in children. Children previously enrolled in a Phase 2 trial were recruited. Participants who had received TCV at 9-11 or 15-23 months were given a Vi-CRM booster at 6-7 years of age (Booster-TCV group), and controls received their first TCV dose at the same age (1st-TCV group). Serum anti-Vi IgG concentrations were measured at baseline and 28 days post-vaccination. Solicited and unsolicited adverse events (AEs) and serious adverse events (SAEs) were recorded. Among 147 children enrolled, 87 received a second and 60 received a first TCV dose. Baseline anti-Vi IgG geometric mean titers (GMT) were higher in the Booster-TCV group (21.5 EU/mL; 95% CI: 17.2-26.8) than in the 1st-TCV group (5.5 EU/mL; 95% CI: 4.5-6.7). At day 28, GMTs rose markedly in both groups: 5140.0 EU/mL (95% CI: 4302.0-6141.3) in the Booster-TCV group and 2084.8 EU/mL (95% CI: 1724.4-2520.5) in the 1st-TCV group. Local reactions and systemic AEs were mild. No SAEs were observed. Vi-TT-induced immunity persisted for at least 5-6 years, and a heterologous booster triggered a strong immune response with universal seroconversion. These findings support heterologous prime-boost strategies to maintain protection in school-age children and inform optimization of TCV schedules in endemic regions.

IntroductionMalaria is a leading health problem with high disease burden and mortality rates worldwide. Currently approved vaccines target the sporozoite form of Plasmodium falciparum that initially infects the liver, but only provide modest protection against malaria in young children. There is an urgent need to develop next-generation malaria vaccines that target multiple parasite developmental stages for greater efficacy. Antibodies to merozoites, which are involved in blood-stage replication, and are associated with clinical illness, have multiple functional activities and can protect against malaria. A promising merozoite vaccine candidate is Merozoite Surface Protein 2 (PfMSP2). Antibodies to PfMSP2 can promote multiple antibody Fc-mediated functional activities to clear merozoites. MethodsWe developed and evaluated monovalent and bivalent (3D7 and FC27 variants) PfMSP2-based mRNA vaccines. We designed and codon-optimised mRNA, which was validated for in vitro expression in mammalian cells, and subsequently formulated as lipid nanoparticles for vaccination of mice in a 3-dose regimen. Vaccination with recombinant PfMSP2 protein with adjuvant was performed for comparison. We evaluated the induction of antibodies and functional activities relevant to protective immunity. ResultsmRNA vaccines induced prominent IgG responses using monovalent (3D7 allele) and bivalent (3D7 and FC27 alleles) vaccines encoding near full-length PfMSP2, and antibodies recognised the surface of whole merozoites. Vaccine responses were equivalent to, or superior than, a recombinant protein-based PfMSP2 vaccine. The bivalent vaccine induced equivalent antibodies to the two PfMSP2 alleles. Vaccination induced cytophilic IgG subclasses with multiple functional activities, including complement fixation, binding of human Fc{gamma}-receptors I and IIa, and opsonic phagocytosis. ConclusionsPfMSP2 is highly immunogenic using the mRNA vaccine platform and induces antibodies with multiple functional activities associated with protective immunity in humans. Combining PfMSP2 with other merozoite and sporozoite antigens is a promising strategy to develop highly efficacious vaccines to achieve malaria control and elimination goals.

Background: Induction of HIV envelope (Env)-specific broadly neutralizing antibodies (bnAbs) is considered a key objective for HIV-1 vaccine development. One approach is to vaccinate with HIV Env immunogens that initially target the naive B cell receptors of a bnAb type and boost with a series of HIV Env variants. We chose a priming immunogen, the CH505 transmitted/founder Env with high affinity for the naive B cell receptor of the prototype CD4 binding site (bs) bnAb lineage, CH103, as a candidate priming immunogen to induce the initial critical step in CD4bs bnAb development. Methods: HVTN 300 is a first-in-human, open-label Phase 1 study evaluating the safety and immunogenicity of a CH505 TF chimeric (ch) Trimer adjuvanted with 3M-052-AF (a TLR7/8 agonist) + Alum. The immunogen is a recombinant, stabilized chimeric Env trimer protein with the N-terminal sequence of CH505 TF gp120 Env transplanted into the BG505 SOSIP sequence. Participants received the adjuvanted protein administered in both deltoid muscles at months 0, 2, 4, 8, and 12. Results: Adults (n=18) aged 18 to 55 were screened at a single site in Boston, USA, and 13 were enrolled. Local and systemic reactogenicity was typically mild to moderate. One participant had severe pain/tenderness, and five participants reported transient severe systemic symptoms at least once. Five participants chose to stop further vaccination due to reactogenicity. No vaccine-related SAEs occurred. Vaccine-specific B-cell response rates reached 100% two weeks post third and fifth vaccinations. Antibody blocking experiments with monoclonal antibodies demonstrated that most participants had antibodies directed to the CD4bs. Four out of 11 participants had serum neutralization signatures for CD4bs bnAb precursors. Conclusions: No safety concerns were identified. The adjuvanted CH505 TF chTrimer elicited serum antibodies capable of CD4bs-mediated neutralization against strains designed to detect early precursors of the CD4bs B-cell lineages. Trial Registration: NCT04915768 Disclosure: Presented in part at HIVR4P 2024, Lima, Peru, October 6-10, 2024

A next-generation SARS-CoV-2 vaccine must address the currently inadequate prevention of virus transmission, particularly against emerging variants of concern, a challenge that none of the licensed commercial vaccines fully meet. Effective control of respiratory pandemics necessitates vaccines that 1) can be rapidly adapted, 2) have high patient compliance with simple and non-invasive administration, and 3) block transmission in a virus challenge. We describe here the characterization of an updated single-cycle SARS-CoV-2 vaccine candidate (scVac), engineered as a replication-defective virus with targeted deletions of the E gene and ORF6 and ORF7a, along with a truncation of ORF3a. The candidate carries an Omicron XBB.1.5 Spike (scVacXBB), maintaining all essential antigenic properties. The vaccine demonstrated an excellent safety profile in K18-hACE2 transgenic mice, the most sensitive virulence model, with no clinical signs or adverse events observed. In the Syrian hamster model, potent systemic and mucosal immune responses were induced, along with a strong neutralizing antibody response. Notably, there was no virus transmission to co-housed naive animals, which outperforms a bivalent Omicron mRNA vaccine reference. Our results demonstrate that scVacXBB-induced immunity not only prevents disease but also effectively blocks transmission. Furthermore, the successful introduction of the XBB.1.5 Spike protein into the scVac platform demonstrates the pipelines ability to adapt quickly to any emerging variant. These findings highlight the potential of this single-cycle concept as a next-generation COVID-19 vaccine, offering robust protection with a strong safety profile.

Blocking transmission of Borrelia burgdorferi (Bb), the causative agent of Lyme disease (LD), from reservoir hosts to humans via Ixodes scapularis ticks represents an alternative strategy to reduce LD incidence. Here, we evaluated Purified Borrelial Lipoproteins (PBL) with a combination of adjuvants, for their ability to limit Bb transmission using C3H/HeN mice and Peromyscus leucopus reservoir models. Immunization with PBL as oral gavage, either alone or nanoparticle-encapsulated, elicited increased antibody responses and reduced pathogen burden in fed larvae and select host tissues. A formulation combining PBL with a recombinant fusion protein adjuvant consisting of Cholera Toxin B subunit, Outer surface protein A, and two-tandem repeats of an M-cell-targeting peptide (rCOM) induced durable protective immunity for up to 10 months in C3H/HeN mice. This oral regimen significantly reduced Bb burden in host tissues, in fed larvae from vaccinated hosts, molted nymphs, and nymph-challenged naive mice. Immunization with PBL+rCOM elevated peripheral levels of Bb-specific IgG isotypes and increased antigen-specific T cell responses producing IFN-{gamma} and IL-4 at days 28 and 65 post-immunization. Significant protective responses were observed in P. leucopus, including strong antibody responses, reduced Bb burden in tissues and reduced Bb transmission to naive larvae, independent of sex but influenced by challenge dose. Sodium chloride content in oral formulation modulated vaccine induced protective responses. Notably, Bb burden in infected nymphs was reduced during the bloodmeal on vaccinated hosts with decreased pathogen transmission to both vertebrate hosts. These findings support PBL+rCOM as a promising oral, reservoir-targeted, transmission-blocking biologic for controlling Lyme disease. Lay AbstractNumerous vertebrate hosts serve as reservoirs of pathogens that are transmitted to humans via the bite of blood feeding vectors such as ticks. Lyme disease, caused by Borrelia burgdorferi (Bb), is the most common tick-borne disease in the US. Bb is transmitted to humans following the bite of infected Ixodes scapularis ticks. In nature, ticks acquire Bb and other pathogens from a variety of reservoir hosts, notably Peromyscus leucopus. Therefore, strategies that limit pathogen burden in reservoir hosts or block their transmission via ticks are options to prevent human infectious diseases, circumventing need for human vaccines and therapeutics. An oral, reservoir host-targeted, pathogen-derived, biologic prepared by extracting immunogenic lipoproteins (Purified Borrelial Lipoproteins) from Bb and combining them with a mucosal adjuvant derived by fusing Cholera-Toxin B subunit, Outer surface protein A of Bb and 2 repeats of an M-cell targeting peptide was tested in C3H/HeN mice and Peromyscus leucopus hosts. Single or two dose regimens via the oral route resulted in significant increases in peripheral Bb specific antibody responses, select T cell responses, blocking the transmission of Bb to naive Is larvae, reducing pathogen burden in vaccinated hosts, and interfering with the infectious cycle of the agent of Lyme disease.

Neonatal sepsis caused by Klebsiella pneumoniae is a major cause of under-five mortality in sub-Saharan Africa, and the rapid increase of infections caused by bacteria resistant to most or all available antimicrobials severely limits treatment options. An effective, maternally-administered vaccine could make a substantial reduction in neonatal sepsis and associated negative outcomes, as well as reduce the overall need for antimicrobials, a key driver of antimicrobial resistance. This study explored the potential for a maternally administered protein-based vaccine to provide neonatal protection via antibodies transferred transplacentally and through breastfeeding. A case-control study of mother and baby dyads was designed with 20 neonates developing K. pneumoniae sepsis and 80 uninfected control neonates to analyse breastmilk IgA, cord blood IgG and maternal serum IgA and IgG antibodies on a protein microarray with 161 selected K. pneumoniae proteins representing 152 unique genes. This analysis identified a set of proteins eliciting antibody responses, some associated with lack of K. pneumoniae sepsis, that indicate the presence of potentially protective antibodies. This is an essential first step in exploring surface protein accessibility, despite the large capsule. We highlight fimbrial structures, conjugative pili, and small lipoproteins associated with large outer membrane complexes as potential protein vaccine targets.

The emergence of new SARS-CoV-2 variants and breakthrough infections underscores the need for next-generation vaccines capable of protecting from natural infection and/or preventing virus transmission to others. Intranasal vaccination offers a promising approach by eliciting local immune responses in the nasal mucosa, the primary site of infection and reservoir for transmissible virus. We evaluated two live-attenuated, respiratory syncytial virus vectored vaccines in which the RSV F and G surface glycoproteins were replaced with a chimeric SARS-CoV-2 Spike protein from either the ancestral USA/WA-1/2020 strain (MV-014-212) or the Delta variant (MV-014-212-delta). A single intranasal dose of either vaccine elicited systemic and mucosal immunity in K18-hACE2 mice, including serum neutralizing antibodies, Spike-specific memory B cells and plasmablasts, and Spike-specific CD8 lung-resident memory T cells. Although MV-014-212-delta vaccination provided the best protection against Delta variant virus challenge, both vaccines decreased viral loads in nasal discharge, lung and brain, and reduced weight loss and mortality. In naturally acquired infection studies, vaccinated hamsters exposed to infected cagemates exhibited minimal weight loss, limited viral replication within the nasal mucosa, and attenuated lung pathology. Therefore, intranasal RSV-vectored vaccines can elicit broad protective respiratory immunity, suggesting that this platform could be leveraged for other respiratory pathogens.

West Nile virus (WNV) is the causative agent of fatal West Nile encephalitis. To date, no human vaccine against WNV has been approved. Adjuvants are important for developing effective and affordable vaccines that enhance the immunogenicity and decrease the required antigen doses. In this study, we assessed the efficacy of AddaS03, a synthetic adjuvant analogous to AS03, in a WNV subunit vaccine composed of soluble recombinant envelope protein (sEnv). Using a passive immunization mouse model, we defined the neutralizing antibody titer threshold required for protection against lethal WNV infections and applied this threshold as a surrogate marker to evaluate adjuvant efficacy. AddaS03-adjuvanted formulations elicited markedly higher neutralizing antibody titers compared to Alhydrogel adjuvant 2% (Alhydrogel), even at suboptimal antigen doses, and consistently exceeded the defined protective threshold titer. Moreover, in a sequential challenge mouse model, AddaS03-adjuvanted vaccines completely protected mice from symptomatic WNV infections, whereas Alhydrogel-adjuvanted vaccines failed to confer full protection. Collectively, these findings demonstrate that AddaS03 is a promising adjuvant for WNV subunit vaccine development and highlights the utility of a passive immunization model for defining protective antibody thresholds as a surrogate marker for vaccine evaluation.

Urinary tract infections, caused primarily by uropathogenic E. coli, are a significant public health burden, affecting approximately 50% of women worldwide. The adhesin FimH is responsible for host receptor binding and is therefore a promising vaccine candidate, but prior studies showed that recombinant monomeric FimH is poorly immunogenic. Here we displayed FimH antigens on the two-component protein nanoparticle I53-50 to generate nanoparticle immunogens that elicit robust levels of receptor-blocking antibodies in mice and non-human primates. We produced nanoparticle immunogens displaying either the FimH lectin domain or a recently reported conformationally stabilized antigen, FimH-DSG, comprising both the lectin and pilin domains. When formulated on aluminum hydroxide, both nanoparticle immunogens elicited similar levels of receptor-blocking activity as a ten-fold higher dose of monomeric FimH-DSG formulated with a potent adjuvant. The improved manufacturability of the stabilized antigen, combined with the ability of nanoparticle display to obviate the need for complex adjuvants, provides important preclinical data for FimH-based vaccines intended to prevent urinary tract infections. More broadly, our data extend the applicability of the I53-50 nanoparticle platform, which to date has been mainly used for displaying viral and protozoan antigens, to bacterial indications.

Vaccination frequently elicits suboptimal immunogenicity in organ transplant recipients, particularly those on long-term immunosuppressive therapy, highlighting the need for improved understanding of immunosuppression mechanisms and optimized vaccination strategies. This study enrolled a cohort of 132 individuals and observed significantly lower antibody levels in kidney transplant recipients (KTRs) compared to non-transplant controls (non-KTRs). Antibody levels were inversely associated with both the dosage and duration of immunosuppressive therapy. Complementary small animal studies demonstrated that immunosuppressive treatment dosage-dependently and reversibly impaired antibody production, primarily by depleting immune cells, notably B cells. A single shot of adenoviral vector-based vaccines demonstrated enhanced immunogenicity relative to two shots of alum-adjuvanted protein vaccines, inducing potent neutralizing antibodies (NAbs) and a Th1-biased T-cell response even under continuous immunosuppression. The enhanced response was driven by reduced interference from pre-existing antibodies, sustained transgene expression, and the reprogramming of lipid metabolism to activate T and B cells. Our findings advocate for tailored vaccination strategies, positioning adenoviral vectors as a candidate modality for this vulnerable population.

Highly pathogenic avian influenza (HPAI) clade 2.3.4.4b H5N1 virus has recently emerged in dairy cattle in the United States. The virus replicates primarily in the mammary gland of infected cattle, leading to dramatic reductions in milk production. It is thought that the virus transmits from animal to animal through viral shedding in milk, and therefore, vaccines that decrease the amount of virus in milk can potentially limit the current outbreak and reduce the risk of H5N1 spillover into humans. Here, we assess the immunogenicity and efficacy of a clade 2.3.4.4b H5 mRNA-LNP vaccine in lactating dairy cows. We found that the H5 mRNA-LNP vaccine elicited robust antibody responses in sera and milk and significantly reduced viral replication and disease caused by clade 2.3.4.4b H5N1 intramammary infection.

In a confirmatory study, we evaluated the immunogenicity and protective efficacy of a heterologous prime-boost vaccination strategy against respiratory syncytial virus (RSV) in non-human primates. Building on prior evidence of protective mucosal immunity induced by intramuscular DNA priming followed by an oropharyngeal adenoviral boost, we conducted a randomized, blinded, dual-centre study across two European primate research facilities. Rhesus macaques received a codon-optimized RSV-F DNA vaccine via electroporation, followed by two mucosal administrations of a recombinant adenovirus serotype 5 vector encoding the same antigen. Control groups included animals vaccinated with irrelevant influenza antigens and a comparator group mimicking natural immunity induced by primary RSV infection. Systemic and mucosal immune responses, including RSV-F-specific antibodies and tissue-resident memory T cells, were monitored longitudinally. Here, we detected robust immune responses, but with some variability between the two centres. However, following experimental RSV challenge performed 22 weeks after the final immunization, RSV-vaccinated animals demonstrated markedly reduced viral replication in both upper and lower respiratory tracts. However, unexpected RSV-specific immunity in the control group at one single study site prevented confirmation of the predefined primary endpoint. Overall, these results support the potential of mucosal adenoviral boosting following DNA priming to induce protective immunity against RSV, while highlighting challenges associated with multi-centre preclinical vaccine studies.

Influenza A virus can cause severe complications in pregnant women and infants, yet no influenza vaccines are approved for infants younger than six months. To address this, novel maternal vaccination strategies are needed to increase global access and coverage in these vulnerable populations. This study evaluated a hemagglutinin (HA) A/California/2009 (H1N1)-based human adenovirus 5 (huAd5) vector vaccine, adjuvanted with a TLR3 agonist, for its ability to induce influenza-specific passive immunity from pregnant and lactating pigs to their piglets following different immunization routes. Influenza naive pregnant dams were vaccinated via oral, intranasal (IN), or intramuscular (IM) routes three weeks prepartum and boosted four weeks later. Serum, colostrum and milk samples were collected longitudinally to assess HA-specific antibody induced by vaccination. H1N1-Ca/09 neutralizing antibodies were evaluated in serum and IFN{gamma} producing cells were assessed in blood, spleen and lymph node cells. IN and IM routes elicited robust serum HA-specific antibody responses when compared to control animals at one- and four-weeks post-boost, whereas the oral route resulted in poor antibody induction across all samples tested. Piglets nursing from IN and IM vaccinated dams showed a significantly higher level of HA-specific antibodies in serum at 2-3 weeks post-partum compared to control piglets. Notably, IN immunized dams and their piglets showed significantly elevated influenza neutralizing antibodies compared to controls. This work demonstrated that both IN and IM immunization with a huAd5-vectored vaccine robustly induced maternal influenza-specific immunity that supported passive transfer to nursing piglets, with IN immunization resulting in superior transfer of neutralizing antibodies.

The yellow fever 17D vaccine is one of the most successful live-attenuated viral vaccines, yet the cellular mechanisms underlying its long-term protection remain not fully understood. This study provides a longitudinal analysis of the human CD4+ T cell and IgG response following de novo yellow fever vaccination by focusing. Peripheral blood mononuclear cells (PBMCs) from 49 vaccinated individuals were stimulated with yellow fever (YF) and control peptide pools across four timepoints: pre-vaccination/baseline (D1), day 22 (D22), day 43 (D43), and one year (D365) post-vaccination. Activation-induced marker (AIM) assays confirmed robust activation of CD4+ T cells following yellow fever peptide stimulation, peaking at day 22 post-vaccination and subsequently declining. T-cell receptor (TCR{beta}) sequencing of AIM-sorted CD4+ T cells showed a transient increase in clonal diversity at D22, consistent with broad epitope targeting and early polyclonal expansion. This was followed by repertoire contraction, which could indicate the persistence of a limited set of dominant clonotypes responsible for immune memory formation. TCR repertoires remained largely private over time, indicating a mostly individualized immune response. Next, serological analyses revealed a robust and highly yellow fever virus (YFV)-specific IgG response. Antibody levels peaked within the D22-D43 window and remained elevated at one year post-vaccination. Cross-reactivity toward other flaviviruses was limited, suggesting an antigen-specific humoral response. Together, these findings characterize the longitudinal dynamics of the CD4+ T cell and IgG response following de novo yellow fever vaccination and provide insights into the mechanisms contributing to durable antiviral immunity.

BackgroundShigellosis morbidity and mortality, combined with the increase in multidrug-resistant infections make Shigella vaccine development a global imperative. Glycoconjugate vaccines that couple immunogenic O-antigen to protein derived from Shigella may provide broader protection across Shigella species and serogroups. Such an approach also circumvents immunotolerance arising from repeated use of the same carrier. Here we use bioconjugation, exploiting an oligosaccharyltransferase (OST) enzyme to couple O-antigen and carrier protein in vivo, to generate a "double-hit" Shigella glycoconjugate vaccine. MethodGlycoconjugates were synthesised in E. coli SDB1 cells expressing S. sonnei O-antigen, the OST PglS, and one of two Shigella carrier proteins. Recombinant glycoconjugate was purified using anion exchange chromatography and then used to immunise mice. Antibody responses were measured and compared by ELISA. ResultsWhen co-produced in E. coli, PglS was able to transfer the cloned S. sonnei O-antigen onto three carrier proteins, modified to accept glycans from the PglS transferase enzymes- the standard bioconjugate carrier ExoA and two immunogenic Shigella-specific outer membrane proteins, EmrK and MdtA. Production of MdtA or ExoA glycoconjugates for immunisation studies utilised successive rounds of anion exchange chromatography, to remove unglycosylated material and obtain highly purified glycoconjugate proteins for us in vaccination. Analysis of murine sera following immunisation revealed an IgG response was raised against both carrier protein and the S. sonnei O-antigen for each glycoconjugate. ConclusionA novel, conserved Shigella protein can be utilised as an effective carrier for the generation of a "double-hit", immunogenic Shigella glycoconjugate vaccine that elicits IgG responses to both carrier protein and S. sonnei O-antigen.

mRNA-lipid nanoparticle (LNP) vaccines are detectable in human blood after vaccination, but platform-specific differences in systemic persistence and transcript integrity remain poorly defined. We analyzed serial blood samples from 73 participants receiving Moderna mRNA-1273 (three formulations), Pfizer/BioNTech BNT162b2, or an investigational receptor-binding domain (RBD) mRNA vaccine (three different doses). Using droplet digital polymerase chain reaction (ddPCR) assays, we quantified total and long-range linked ("intact") vaccine mRNA, and we measured vaccine-specific ionizable lipids by liquid chromatography-mass spectrometry (LC-MS). Across platforms, mRNA decay was fastest for mRNA-1273, intermediate for BNT162b2, and slowest for the RBD vaccine, with ionizable lipid decay following the same rank order. Notably, intact spike mRNA declined two-fold faster after mRNA-1273 than BNT162b2 vaccination. Kinetics modelling revealed platform-dependent coupling of mRNA and lipid kinetics: intact mRNA tracked closely with SM-102 for mRNA-1273, whereas ALC-0315 persisted longer than intact mRNA for BNT162b2. A ten-fragment linkage ddPCR panel spanning the spike transcript showed lower linkage toward 3'-proximal regions that mirrored the administered mRNA-1273 formulation. Together, these data establish a quantitative framework for benchmarking mRNA-LNP platform kinetics and transcript integrity in humans.

BackgroundGiven emerging evidence on the waning of immunity from typhoid conjugate vaccines (TCV), the World Health Organization (WHO) commissioned a multi-model comparison to determine the optimal schedule in terms of health and economic impact to inform updated recommendations for TCV use across different settings. Methods and findingsTo identify optimal vaccination strategies across different incidence settings and vaccine waning assumptions, we compared two agent-based and two compartmental dynamic models of typhoid transmission. All models were fitted to harmonized age-specific incidence data from medium, high, and very high incidence settings. We assessed different TCV schedules under slow- and fast-waning scenarios to evaluate the best age for routine vaccination and the potential need for booster doses and catch-up campaigns. We evaluated the public health and economic impact predicted for each model and scenario using the net-monetary-benefit framework to determine cost-effectiveness under two representative scenarios for the health outcomes and costs of vaccination and treatment. Over a 10-year time horizon, routine vaccination at 9 months with a catch-up campaign to 15 years and a booster dose at 5 years was predicted to have the greatest impact, reducing cases by a median of 48-64% across the incidence settings. Across all four models, TCV introduction with a catch-up campaign was cost-effective at willingness-to-pay (WTP) thresholds >$1,250 per disability-adjusted life-year (DALY) averted in medium incidence settings when costs and case-fatality risk (CFR) are high and in high incidence settings when costs and CFR are low. The optimal strategy was to delay vaccination to 2 or 5 years of age if waning is fast, depending on the age of peak incidence. In very high incidence settings, TCV introduction at 9 months or 2 years of age was cost-saving, and adding a booster dose at 5 years was cost-effective at most WTP values across all scenarios. ConclusionsModel predictions for the impact and cost-effectiveness of different TCV schedules were fairly robust to uncertainty in parameter values and model structure, but the optimal strategy depends on the typhoid incidence rate, CFR, and waning rate of vaccine protection.

The development of broadly protective and dose-sparing influenza vaccines remains a critical challenge, particularly for zoonotic H5N1 strains with pandemic potential. This study evaluates BECC470s, a synthetic TLR4 adjuvant, for its ability to enhance the immunogenicity and protective efficacy of recombinant H5 hemagglutinin (rHA) vaccination in murine models. BECC470s-adjuvanted rHA elicited robust IgG1/IgG2a antibody responses and complete survival following homologous 2004 H5N1 challenge in a prime-boost model. Although BECC470s broadened antibody binding to both variable HA head and conserved stalk domains by ELISA, functional neutralizing antibody responses were restricted to the matched 2004 H5N1 isolate, with no detectable neutralization of H5N1 viruses isolated in 2022 or 2024. These data indicate that BECC470s enhances the magnitude and apparent breadth of binding antibody responses while maintaining strain-specific neutralizing activity, supporting its potential as an adjuvant for next-generation influenza vaccines while underscoring the need for further optimization to achieve true cross-neutralizing protection.