Vaccine Development Research Articles

Divergent Synthesis and Antigenicity Evaluation of Core Oligosaccharides of the Lipopolysaccharides from Acinetobacter baumannii SMAL and ATCC 19606.

Acinetobacter baumannii poses a serious threat to human health. Pathogenic bacterial lipopolysaccharides (LPSs) are potent immunogens for the development of antibacterial vaccines. To investigate the antigenic properties of A. baumannii LPS, five well-defined core oligosaccharide fragments from the LPS of A. baumannii SMAL and ATCC 19606 were synthesized. A divergent synthesis strategy based on orthogonally protected α-(2 → 5)-linked Kdo dimer 6 was developed. Selective exposure of different positions in this key precursor and then elongation of sugar chains via stereocontrolled formation of both 1,2-trans and 1,2-cis-2-aminoglycosidic linkages permitted the efficient synthesis of the targets. The synthetic route also highlights a 4-O and then 7-O glycosylation sequence for assembly of the novel 4,7-branched Kdo framework. Antigenicity assay using the glycan microarray technique disclosed that tetrasaccharide 3 featuring both 4,7-branch and α-(2 → 5)-Kdo-Kdo structural elements was a potential antigenic determinant.

Tyrosinase-Mediated Conjugation for Antigen Display on Ferritin Nanoparticles.

Ferritin (Ft) nanoparticles have become versatile platforms for displaying antigens, being a promising technology for vaccine development. While genetic fusion has traditionally been the preferred method for antigen display, concerns about improper folding and steric hindrance that may compromise vaccine efficacy or stability have prompted alternative approaches. Bioconjugation offers the advantage of preserving native protein structure and function, with recent advancements improving efficiency and specificity. In this study, we used tyrosinase (TYR) to bioconjugate the receptor binding domain of the SARS-CoV-2 spike protein, tagged with a tyrosine (RBD-Y), to native cysteines on Ft, resulting in RBD-Y-Ft nanoparticles. We quantified available cysteines on ferritin using Ellman's assay and monitored their reduction during the reactions. Denaturing analytics (via SDS-PAGE, Western blot, and LC-TOF-MS) confirmed the formation of RBD-Y-Ft monomers with an expected molecular weight of 46 kDa. Mass photometry and HPLC estimated a molecular weight of RBD-Y-Ft nanoparticles of 680 kDa, which was higher than that of nonfunctionalized ferritin (480 kDa), indicating successful binding of up to eight RBD-Y antigens per 24-mer Ft nanoparticle. This work enhances our understanding of how Ft nanoparticles can be engineered to present antigens, leveraging them as a robust scaffold for producing tailored-made candidate vaccines in a timely manner.

T Cell Peptide Prediction, Immune Response, and Host-Pathogen Relationship in Vaccinated and Recovered from Mild COVID-19 Subjects.

COVID-19 remains a significant threat, particularly to vulnerable populations. The emergence of new variants necessitates the development of treatments and vaccines that induce both humoral and cellular immunity. This study aimed to identify potentially immunogenic SARS-CoV-2 peptides and to explore the intricate host-pathogen interactions involving peripheral immune responses, memory profiles, and various demographic, clinical, and lifestyle factors. Using in silico and experimental methods, we identified several CD8-restricted SARS-CoV-2 peptides that are either poorly studied or have previously unreported immunogenicity: fifteen from the Spike and three each from non-structural proteins Nsp1-2-3-16. A Spike peptide, LA-9, demonstrated a 57% response rate in ELISpot assays using PBMCs from 14 HLA-A*02:01 positive, vaccinated, and mild-COVID-19 recovered subjects, indicating its potential for diagnostics, research, and multi-epitope vaccine platforms. We also found that younger individuals, with fewer vaccine doses and longer intervals since infection, showed lower anti-Spike (ELISA) and anti-Wuhan neutralizing antibodies (pseudovirus assay), higher naïve T cells, and lower central memory, effector memory, and CD4hiCD8low T cells (flow cytometry) compared to older subjects. In our cohort, a higher prevalence of Vδ2-γδ and DN T cells, and fewer naïve CD8 T cells, seemed to correlate with strong cellular and lower anti-NP antibody responses and to associate with Omicron infection, absence of confusional state, and habitual sporting activity.

Evaluating the Compatibility of Three Aluminum Salt-Adjuvanted Recombinant Protein Antigens (Trivalent NRRV) Combined with a Mock Trivalent Sabin-IPV Vaccine: Analytical and Formulation Challenges.

In this work, we describe compatibility assessments of a recombinant, trivalent non-replicating rotavirus vaccine (t-NRRV) candidate with a mock trivalent Sabin inactivated polio vaccine (t-sIPV). Both t-sIPV and t-NRRV are incompatible with thimerosal (TH), a preservative commonly used in pediatric pentavalent combination vaccines (DTwP-Hib-HepB) distributed in low- and middle-income countries (LMICs), preventing the development of a heptavalent combination. The compatibility of t-NRRV with a mock DTwP-Hib-HepB formulation is described in a companion paper. This case study highlights the analytical and formulation challenges encountered when combining a mock t-sIPV vaccine (unadjuvanted) with Alhydrogel® (AH) adjuvanted t-NRRV. Selective and stability-indicating competition ELISAs were implemented to monitor antibody binding to each of the six antigens (±AH). Simple mixing caused the undesired desorption of t-NRRV from AH with the concomitant binding of t-sIPV to AH. Although the former effect was mitigated by dialyzing sIPV bulks, decreased sIPV storage stability was observed at accelerated temperatures in the bivalent combination with a rank-ordering of P[8] > P[6] > P[4] and sIPV3 > sIPV2 > sIPV1. The compatibility of AH-adsorbed t-sIPV with alternative preservatives was evaluated, and parabens (methyl, propyl) were identified for potential use in this multi-dose bivalent formulation. Along with a companion paper, the lessons learned are discussed to facilitate the future formulation development of pediatric combination vaccines with new antigens.

Innate cells and STAT1-dependent signals orchestrate vaccine-induced protection against invasive Cryptococcus infection.

Fungal pathogens are underappreciated causes of significant morbidity and mortality worldwide. In previous studies, we determined that a heat-killed, Cryptococcus neoformans fbp1-deficient strain (HK-fbp1) is a potent vaccine candidate. We determined that vaccination with HK-fbp1 confers protective immunity against lethal Cryptococcosis in an interferon γ (IFNγ)-dependent manner. In this study, we set out to uncover cellular sources and relevant targets of the protective effects of IFNγ in response to the HK-fbp1 vaccine. We found that early IFNγ production peaks at day 3 and that monocytes and neutrophils are important sources of this cytokine after vaccination. Neutralization of IFNγ at day 3 results in impaired CCR2+ monocyte recruitment and reduced differentiation into monocyte-derived dendritic cells (Mo-DC). In turn, depletion of CCR2+ cells prior to immunization results in impaired activation of IFNγ-producing CD4 and CD8 T cells. Thus, monocytes are important targets of innate IFNγ and help promote further IFNγ production by lymphocytes. We employed monocyte-fate mapper and conditional STAT1 knockout mice to uncover that STAT1 activation in CD11c+ cells, including alveolar macrophages, Mo-DCs, and monocyte-derived macrophages (Mo-Mac) is essential for HK-fbp1 vaccine-induced protection. Altogether, our aggregate findings suggest critical roles for innate cells as orchestrators of vaccine-induced protection against Cryptococcus infection.IMPORTANCEThe number of patients susceptible to invasive fungal infections across the world continues to rise at an alarming pace yet current antifungal drugs are often inadequate. Immune-based interventions and novel antifungal vaccines hold the promise of significantly improving patient outcomes. In previous studies, we identified a Cryptococcus neoformans mutant strain (Fbp1-deficient) as a potent, heat-inactivated vaccine candidate capable of inducing homologous and heterologous antifungal protection. In this study, we used a combination of methods together with a cohort of conditional knockout mouse strains to interrogate the roles of innate cells in the orchestration of vaccine-induced antifungal protection. We uncovered novel roles for neutrophils and monocytes as coordinators of a STAT1-dependent cascade of responses that mediate vaccine-induced protection against invasive cryptococcosis. This new knowledge will help guide the future development of much-needed antifungal vaccines.

Development of Glycan-masked SARS-CoV-2 RBD vaccines against SARS-related coronaviruses.

Emerging and recurrent infectious diseases caused by coronaviruses remain a significant public health concern. Here, we present a targeted approach to elicit antibodies capable of neutralizing SARS-CoV-2 variants and other SARS-related coronaviruses. By introducing amino acid mutations at mutation-prone sites, we engineered glycosylation modifications to the Receptor Binding Domain (RBD) of SARS-CoV-2, thereby exposing more conserved, yet less accessible epitopes. We developed both messenger RNA (mRNA) and recombination subunit vaccines using these engineered-RBDs (M1, M2) and the wild-type RBD as immunogens. The engineered-RBD vaccines elicited robust neutralizing responses against various SARS-CoV-2 variants as well as SARS-CoV and WIV1-CoV, and conferred protection in mice challenged with the XBB.1.16 strain. Furthermore, We highlighted that glycan masking is a decisive factor in antibody binding changes and RBD-conserved antibody response. Additionally, the glycan-engineered RBD mRNA vaccines stimulated stronger cell-mediated immune responses. Our glycan modification strategy significantly enhances broad-spectrum neutralizing efficacy and cellular immunity, providing valuable insights for the development of vaccines against a wide range of SARS-related coronaviruses.

Immunogenicity Analysis of Chikungunya Virus DNA Vaccine Based on Mutated Putative N-Linked Glycosylation Sites of the Envelope Protein.

Chikungunya fever is a mosquito-borne infectious disease caused by the chikungunya virus (CHIKV). Recently, CHIKV has spread rapidly worldwide, raising global concerns. However, there is only one approved vaccine is available to prevent CHIKV infection; therefore, different platform vaccines development is a public health priority. The CHIKV genome encodes four non-structural polyproteins (nsP1-4) and one structural polyprotein (capsid, envelope 3, envelope 2, 6 K, and envelope 1). Previous studies have shown that N-linked glycans in viral proteins play important roles in regulating immune responses. Accordingly, in this study, we designed four CHIKV DNA vaccine candidates with mutated N-glycosylation sites in the full-length E and E I/II proteins. Our results indicated that immunization of mice with the vaccine elevated the cytokines levels, including IFN-γ, associated with T cell immune response. Furthermore, the truncated E protein with a deleted E III domain (E I/II) exhibited better immunogenicity than the full-length E protein, and N-linked glycosylation of E I/II protein induced a higher cell-mediated immune response. Overall, our study demonstrates that N-linked glycosylation of the E I/II proteins of CHIKV significantly enhances cell-mediated immune responses, laying the foundation for the development of potential vaccination strategies against CHIKV.

Innate and adaptive immunity gene expression profiles induced by virulent Aeromonas hydrophila infection in the immune-related organs of channel catfish

Aeromonas hydrophila causes motile Aeromonas septicemia (MAS) in freshwater fish. In recent years, MAS outbreaks due to virulent Aeromonas hydrophila (vAh) have been responsible for large-scale losses within commercial catfish farms in Mississippi and Alabama. The aim of this study was to evaluate immune gene expression in catfish immune-competent tissues during infection with vAh strain ML09-119. Specific pathogen-free catfish fingerlings were intraperitoneally infected with vAh strain ML09-119, and relative expression of thirteen immune-related genes was evaluated from head kidney, spleen, and liver. Our results revealed that vAh was detected 2 h post-infection (hpi) in the head kidney, liver, and spleen. The highest concentration of vAh was detected at 12 hpi, from which point concentrations decreased until clearance at 5 days post-infection (dpi). Gene expression analysis revealed upregulation of pro-inflammatory cytokines and innate immune response (TLR 4 and 5) in the first 24 hpi. Adaptive immune-related genes were upregulated at 7 dpi in the spleen and 14 dpi in the head kidney. Furthermore, immunoglobulin M showed significant upregulation at 14 dpi in the head kidney and 21 dpi in the spleen. In summary, vAh ML09-119 infection induced a strong inflammatory response involving multiple innate immunity genes, proinflammatory cytokines, and chemokines. Surviving catfish were able to clear the infection and produce antibodies and memory cells. Assessment of the immunological response to vAh infection is critical for understanding the pathogen's mechanisms of pathogenesis and developing means for MAS control, including vaccine development and improved treatments.

Global landmark: 2023 marks the worst year for dengue cases with millions infected and thousands of deaths reported

ObjectivesIn 2023, the world experienced the worst dengue virus (DENV) outbreak on record. The study aimed to identify global regions and continents with high burden of dengue in 2023. DesignWe collected data on the number of DENV cases and deaths reported by various countries to the World Health Organization and World Health Organization regional offices. We estimated DENV cases per million population and case fatality ratio (CFR) among the confirmed cases reported by each country. ResultsOverall, in 2023 more than 6.5 million cases and over 6,800 deaths attributed to DENV were recorded globally, marking a historic milestone. Two distinct hotspots of DENV circulation emerged: South America and South and Southeast Asian regions. South America reported the highest number of cases (3,924,992), and 1,946 deaths, with a CFR of 0.05. In Asia, 1,622,405 cases and 3,637 deaths were reported, with a CFR of 0.22. We observed a statistically significant difference in the number of cases and deaths per million across different continents (P-value <0.001). However, the CFR did not differ significantly across continents (P-value = 0.123). ConclusionsThe increased cases and mortality highlight the urgent need for a comprehensive global approach aimed at DENV infection control, including vaccine development, vector control, public health initiatives, and improved clinical management.

Time-course whole blood transcriptome profiling provides new insights into Microtus fortis natural resistance mechanism to Schistosoma japonicum

Microtus fortis is known as a non-susceptible animal host of S. japonicum. A better understanding of this animal immune defense mechanism during the early stage of infection may offer an alternative route for vaccine development or therapy. Here, we analyzed the whole blood transcriptome of M. fortis using next-generation sequencing (NGS) to identify immune genes of biological relevance that might be involved in the mechanism of its resistance. The blood samples were collected from uninfected animals (control group) and infected animals at different time points (3, 7, 10 and 14 days post-infection). We identified 5310 sequences as unigenes and successfully annotated 4636 of them. The immune response was more intense at 10 dpi. The upregulated genes at this time point were mainly activated in the TNF and NF-kappa B signaling pathways, Th1, Th2and Th17 cell differentiation as well as cytokine-cytokine receptor interaction. Based on the differentially expressed genes analysis, we report that the IF27L2B, RETN, PGRP, IFI35, TYROBP, S100A8, S100A11, CD162, CD88, CYBA, and LBP could play important roles in the mechanism of M. fortis resistance.

The Vaccine Godmother: Dr. Gagandeep Kang's Pioneering Journey in Global Health and Vaccine Development.

Dr. Gagandeep Kang is a distinguished Indian microbiologist and virologist known for her pioneering work in the study of gastrointestinal diseases, diarrheal infections, and vaccine development. This article highlights her career, beginning with her medical education at Christian Medical College (CMC) Vellore, where she embarked on her groundbreaking research in enteric diseases, particularly rotavirus, a major cause of child mortality globally. Over the course of her distinguished career, she has led groundbreaking research on rotavirus, contributing to the development of two WHO-approved vaccines tailored for Indian communities: Rotavac (Bharat Biotech, Hyderabad, India) and Rotasiil (Serum Institute of India, Pune, India). Beyond her research, Dr. Kang has held significant advisory roles on national and global platforms, including WHO's Global Advisory Committee on Vaccine Safety and the Coalition for Epidemic Preparedness Innovations (CEPI). She has authored over 300 scientific publications and coauthored the bestselling book "Till We Win,"which details India's fight against the COVID-19 pandemic. Her exceptional contributions have earned her numerous accolades, including the Infosys Prize in Life Sciences, the Fellowship of the Royal Society, and the Canada Gairdner Global Health Award. Dr. Kang's career exemplifies her dedication to advancing public health, vaccine development, and global health initiatives, making her a trailblazer in the field of microbiology and virology.

Packaging of alphavirus-based self-amplifying mRNA yields replication-competent virus through a mechanism of aberrant homologous RNA recombination.

Messenger (m)RNA has taken center stage in vaccine development, gene therapy, and cancer immunotherapy. A next-generation of mRNA is the self-amplifying (sa)mRNA, which induces broad and long-lasting immunity at a lower dose which provides better clinical outcomes in conjunction with fewer adverse effects. SamRNA, also known as "replicon" RNA, encodes the replication machinery of an alphavirus together with an antigen. Efficient delivery of replicon RNA to target tissues can be accomplished by packaging the replicon RNA in virus-like replicon particles (VRPs) via co-transfection of producer cells with defective helper RNA(s) encoding the alphavirus structural proteins. During the manufacture of VRPs, however, there is a potential risk of RNA recombination, which may lead to the formation of replication-competent virus (RCV). To investigate the factors influencing the unwanted RCV formation, we evaluated how sequence homology orchestrates alphavirus RNA recombination. Several combinations of complementing alphavirus replicon and helper RNAs varying in length of sequences overlap were co-transfected in mammalian cells. The culture fluid was serially passaged to detect RCV. Nanopore sequencing of cells after the first passage in combination with amplicon-based Sanger sequencing of RCV in the culture fluid after four passages led to the detection of RNA recombination. RCV was generated between replicon and helper RNAs with sequence homology in either the non-structural or structural genes, whereas RNAs without overlapping gene regions did not generate RCV. Remarkably, no sequence overlap was detected at the recombination junction sites in the RCV genome, suggesting a mechanism of "aberrant homologous RNA recombination." Accordingly, we conclude that the alphavirus RNA recombination process leading to the formation of RCV is homology-assisted and can be prevented by avoiding sequence homology between replicon and helper RNAs.IMPORTANCEThere is a growing interest in the use of self-amplifying (sa)mRNA vectors for next-generation vaccine development, gene therapy, and cancer immunotherapy. The delivery of samRNA in the form of virus-like replicon particles (VRPs) enables efficient delivery of samRNA to target tissue. The production of these VRPs, however, suffers from contamination with replication-competent virus (RCV) that is thought to arise from recombination events between samRNA and helper RNAs for VRP packaging. The presence of RCV in samRNA in the clinical product is undesirable as alphaviruses may cause serious disease in humans. However, the underlying recombination mechanism leading to RCV is currently unknown. In our work, we demonstrate a detailed evaluation of the recombination sites, which indicates that RCV is formed through an unusual mechanism of "aberrant homologous RNA recombination." The results are useful for researchers in the field of RNA vaccine manufacture and delivery.

Buffer Screening of Protein Formulations Using a Coarse-Grained Protocol Based on Medicinal Chemistry Interactions.

In drug and vaccine development, the designed protein formulation should be highly stable against the temperature, pH, buffer, excipients, and other environmental settings. Similarly, in a sensing unit, one needs to know how strongly two biomolecules bind to guide the design of the biorecognition unit accordingly. Typically, the community performs a series of experiments to thoroughly examine the parameter space, the so-called design-of-experiment (DoE) method, to identify the optimal formulation conditions. Unfortunately, extensive physical testing entails high costs, repeatability issues, and a lack of in-depth knowledge of the underlying mechanisms that affect the final outcome. To address these challenges, we developed a physics-based simulation protocol for buffer screening of protein formulations. We are introducing a coarse-grained molecular simulation protocol that consists of six different interactions. The so-called medicinal chemistry interactions (electrostatics, hydrophobicity, hydrogen bonding propensity, disulfide bonding, and water-water) are based on the physical nature of the protein's amino acid and the partitioning/polarity of any other chemical constituent. The protocol is applied in immunoglobulin-based monoclonal antibodies. We have analyzed the protein behavior as a function of acidity (pH) to discover the isoelectric point by solving the Poisson-Boltzmann equation in a mesoscale grid. To identify the conditions under which the protein oligomerizes in a given buffer, pH, temperature, and ionic strength, we are performing dissipative particle dynamics (DPD) simulations. The protocol allows researchers to reach the high time/space scales required to study protein formulations in their full complexity. Combined with the disruptive protein folding artificial intelligence (AI) algorithms that have been recently developed, the protocol creates a powerful digital framework for cultivating advanced pharmaceutical and biological applications.

Isolation of Virulent Leptospira Serogroup Australis Field Strains from Symptomatic Dogs for Canine Leptospiral Vaccine Development.

Leptospirosis is a widespread zoonosis caused by spirochaetes belonging to the pathogenic species of Leptospira, which are classified into more than 25 serogroups and 250 serovars. Vaccination can prevent the disease in dogs but offers incomplete efficacy because of a lack of cross-protection between serogroups. The aim of this study was to validate a robust recruitment and sampling process, with the objectives of isolating and typing circulating Leptospira pathogenic strains and then selecting those of proven virulence and pathogenicity for vaccine development. Blood and urine samples from dogs with clinical syndromes compatible with acute leptospirosis were sterilely collected and transported to a reference laboratory for a micro-agglutination test (MAT), PCR, and bacterial isolation. Isolated strains underwent molecular typing using RNA16S, variable-number tandem repeat (VNTR), and pulsed-field gel electrophoresis (PFGE). Subtyping was performed using core genome multilocus sequence typing (CgMLST). Among 64 included dogs, 41 had MAT and/or PCR results compatible with Leptospira infection, and 14 Leptospira strains were isolated. Based on molecular typing, 11 isolates were classified as L. interrogans serogroup Australis, serovar Bratislava, and 3 as serogroup Icterohaemorrhagiae, serovar Icterohaemorrhagiae. CgMLST subtyping revealed a diversity of clonal groups (CGs) distributed in several regional clusters. Besides validating a robust recruitment and sampling process, this study outlines the value of combining PCR and serological testing when suspecting leptospirosis and the usefulness of implementing molecular typing methods to identify circulating field strains. It also confirms the epidemiological importance of the Australis serogroup and allows for the collection of different highly pathogenic strains for vaccine development.

Production of recombinant cytokines and polyclonal antibodies for analysis of cellular immune response in golden Syrian hamster.

The development of therapies and vaccines for various diseases often necessitates the analysis of cellular immunity. However, unlike other rodents, the limited availability of reagents for Syrian hamsters restricts immunological analysis, particularly in the determination of serum effector molecules such as cytokines. In this study, we aim to produce and characterize the cytokines IFN-γ, TGF-β, IL-6, IL-10, and TNF-α from Syrian hamsters in recombinant form and to generate polyclonal antibodies against them in rats. Cytokine transcript sequences were cloned into expression vectors in E. coli. Recombinant proteins were produced, purified through affinity chromatography, and characterized by Western blot using an anti-6xHis monoclonal antibody. Rats were immunized with the recombinant proteins to generate polyclonal antibodies (pAbs). These pAbs were characterized by Western blot and titrated by indirect ELISA. The recombinant cytokines rTNF-α, rIL-10, rIFN-γ, rTGF-β, and rIL-6 were produced and specifically recognized at their expected molecular weights of 22.3kDa, 19.8kDa, 18.9kDa, 11.8kDa, and 22.9kDa. pAbs were produced and demonstrated the ability to specifically recognize their target proteins with titers of 409,600 (rIL-10), 204,800 (rTNF-α), 102,400 (rIL-10), 51,200 (rTGF-β), and 25,600 (rIFN-ɣ). The reagents produced represent a starting point for developing immunoassays to detect hamster cytokines, facilitating the analysis of cellular immunity in this biomodel.

Dynamics of Spike-Specific Neutralizing Antibodies Across Five-Year Emerging SARS-CoV-2 Variants of Concern Reveal Conserved Epitopes that Protect Against Severe COVID-19.

Since early 2020, several SARS-CoV-2 variants of concern (VOCs) continue to emerge, evading waning antibody mediated immunity produced by the current Spike-alone based COVID-19 vaccines. This caused a prolonged and persistent COVID-19 pandemic that is going to enter its fifth year. Thus, the need remains for innovative next generation vaccines that would incorporate protective Spike-derived B-cell epitopes that resist immune evasion. Towards that goal, in this study we (i) Screened the sequences of Spike among many VOCs and identified conserved and non-conserved linear B-cell epitopes; (ii) Compared titers and neutralization antibodies specific to these conserved and non-conserved B-cell epitopes from serum of symptomatic and asymptomatic COVID-19 patients that were exposed to multiple VOCs across the 5-year COVID-19 pandemic, and (iii) Compared protective efficacy of conserved versus non-conserved B-cell epitopes against the most pathogenic Delta variant in a "humanized" ACE-2/HLA transgenic mouse model. We found robust conserved B-cell epitope-specific antibody titers and neutralization in sera from asymptomatic COVID-19 patients. In contrast, sera from symptomatic patients contained weaker antibody responses specific to conserved B-cell epitopes. A multi-epitope COVID-19 vaccine that incorporated the conserved B-cell epitopes, but not the non-conserved B-cell epitopes, significantly protected the ACE2/HLA transgenic mice against infection and COVID-19 like symptoms caused by the Delta variant. These findings underscore the importance of conserved B-cell epitopes in generating robust protective immunity against severe COVID-19 symptoms caused by various VOCs, providing valuable insights for the development of broad-spectrum next generation Coronavirus vaccines capable of conferring cross-variant protective immunity.

Molecular mechanism and structure-guided humanization of a broadly neutralizing antibody against SFTSV.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a novel tick-borne bunyavirus that causes severe fever with thrombocytopenia syndrome (SFTS), with a high mortality rate of up to 30%. The envelope glycoproteins of SFTSV, glycoprotein N (Gn) and glycoprotein C (Gc), facilitate the recognition of host receptors and the process of membrane fusion, allowing the virus to enter host cells. We previously reported a monoclonal antibody, mAb 40C10, capable of neutralizing different genotypes of SFTSV and SFTSV-related viruses. However, the specific neutralization mechanism is poorly understood. In this study, we elucidated the high-resolution structure of the SFTSV Gn head domain in complex with mAb 40C10, confirming that the binding epitope in the domain I region of SFTSV Gn, and it represented that a novel binding epitope of SFTSV Gn was identified. Through in-depth structural and sequence analyses, we found that the binding sites of mAb 40C10 are relatively conserved among different genotypes of SFTSV and SFTSV-related Heartland virus and Guertu virus, elucidating the molecular mechanism underlying the broad-spectrum neutralizing activity of mAb 40C10. Furthermore, we humanized of mAb 40C10, which is originally of murine origin, to reduce its immunogenicity. The resulting nine humanized antibodies maintained potent affinity and neutralizing activity. One of the humanized antibodies exhibited neutralizing activity at picomolar IC50 values and demonstrated effective therapeutic and protective effects in a mouse infection model. These findings provide a novel target for the future development of SFTSV vaccines or drugs and establish a foundation for the research and development of antibody therapeutics for clinical applications.

Carbapenem-Resistant Enterobacter hormaechei Uses Mucus Metabolism to Facilitate Gastrointestinal Colonization.

Bloodstream infections caused by Enterobacter species pose a significant clinical threat. The intestine acts as the primary site for colonization and serves as a reservoir for infection. To combat this pathogen, it is crucial to understand how carbapenem-resistant Enterobacter species colonize the gut, as such knowledge can pave the way for alternative therapeutic targets. In this study, we developed a novel neonatal mouse model for gastrointestinal colonization by Enterobacter species and discovered that mucus plays a key role as a carbon source during colonization. Additionally, we identified two mucus catabolism pathways that contribute to intestinal colonization by carbapenem-resistant E. hormaechei. This new mouse model offers valuable insights into host-pathogen interactions and helps identify critical gastrointestinal fitness factors of Enterobacter, potentially guiding the development of vaccines and alternative therapeutic strategies to minimize intestinal carriage in patient populations at risk for infection with Enterobacter species.

Rabies virus as vector for development of vaccine: pros and cons.

Rabies virus as vector for development of vaccine: pros and cons.

Evolutionary Insights from Association Rule Mining of Co-Occurring Mutations in Influenza Hemagglutinin and Neuraminidase.

Seasonal influenza viruses continuously evolve via antigenic drift. This leads to recurring epidemics, globally significant mortality rates, and the need for annually updated vaccines. Co-occurring mutations in hemagglutinin (HA) and neuraminidase (NA) are suggested to have synergistic interactions where mutations can increase the chances of immune escape and viral fitness. Association rule mining was used to identify temporal relationships of co-occurring HA-NA mutations of influenza virus A/H3N2 and its role in antigenic evolution. A total of 64 clusters were found. These included well-known mutations responsible for antigenic drift, as well as previously undiscovered groups. A majority (41/64) were associated with known antigenic sites, and 38/64 involved mutations across both HA and NA. The emergence and disappearance of N-glycosylation sites in the pattern of N-X-[S/T] were also identified, which are crucial post-translational processes to maintain protein stability and functional balance (e.g., emergence of NA:339ASP and disappearance of HA:187ASP). Our study offers an alternative approach to the existing mutual-information and phylogenetic methods used to identify co-occurring mutations, enabling faster processing of large amounts of data. Our approach can facilitate the prediction of critical mutations given their occurrence in a previous season, facilitating vaccine development for the next flu season and leading to better preparation for future pandemics.