Search Results (439)

Background/Objectives: Post-weaning diarrhea remains a major challenge in pig production worldwide. Enterotoxigenic Escherichia coli (ETEC) encoding fimbriae of the F4 type and producing the heat-stable enterotoxin, STb, are one of the important causes of this disease. The aim of the current study was to evaluate whether vaccination of pregnant sows with a novel capsid virus-like particle (cVLP)-based vaccine against F4 and STb (cVLP-FaeG/cVLP-STb) could enhance performance in piglets born after such vaccinated sows. Methods: A field trial was conducted in a commercial sow-to-finisher pig herd. Thirty-five sows were vaccinated twice with the cVLP-FaeG/cVLP-STb vaccine prior to farrowing, while thirty-five control sows were vaccinated twice with commercial vaccines normally used in the herd. Piglets were followed until eight weeks post-weaning to assess antibody responses, diarrhea and treatment incidences, pathogen shedding, and growth performance. Results: Piglets born from immunized sows receiving the cVLP vaccine showed significantly higher serum antibody levels against ETEC F4 throughout the post-weaning period (p ≤ 0.021). The frequency of pathogen detection was similar between groups, while piglets in the cVLP group exhibited significantly lower diarrhea scores at week 6 (p = 0.047), showed a trend of requiring fewer treatments (p = 0.06) and had significantly higher final body weight (p = 0.048). In addition, the cVLP group showed a significantly greater average daily gain over the study period (p = 0.037). Conclusion: Sow immunization with the cVLP vaccine enhanced passive immune protection of piglets, resulting in reduced antimicrobial treatment 2 weeks post-weaning and improved growth performance. Full article

Background/Objectives: Canine papillomavirus (CPV) is an important viral pathogen associated with papillomatosis in dogs, with canine papillomavirus type 1 (CPV1) and type 2 (CPV2) among the most prevalent and clinically relevant genotypes. The L1 capsid protein is a major immunogenic antigen of papillomaviruses; however, conserved linear B-cell epitopes shared between CPV genotypes remain poorly defined. This study aimed to identify conserved cross-reactive B-cell epitopes within CPV1 and CPV2 L1 proteins and to evaluate their preliminary immunoreactivity. Methods: Conserved linear B-cell epitopes were predicted through integrated bioinformatic and structural analyses based on sequence conservation and surface accessibility. Three candidate epitopes were selected. Recombinant CPV1 and CPV2 L1 proteins were expressed in Escherichia coli (E. coli), purified, used as recombinant L1 antigens, together with BSA-conjugated synthetic epitope peptides for mouse immunization. Antigen-specific IgG responses were assessed by ELISA, antigen-associated IFN-γ responses were evaluated by ELISpot, and cross-reactive antibody recognition was assessed by Western blot. Results: Recombinant L1 proteins induced strong antigen-specific IgG responses in mice. The selected peptides induced detectable but weaker humoral responses compared with the recombinant L1 proteins. Among the three epitopes, TPSGSLV and TVVDNTR elicited antibodies that recognized both CPV1 and CPV2 L1 proteins, while the epitope VIVPKVS showed minimal or no detectable immunoreactivity. ELISpot analysis showed only modest antigen-associated IFN-γ responses, particularly in peptide-immunized groups. Conclusions: This study identified conserved cross-reactive linear B-cell epitope candidates within CPV1 and CPV2 L1 proteins and provided preliminary immunological evidence supporting their potential relevance for CPV antigen design. However, peptide-induced responses were weaker than those induced by recombinant L1 proteins, and VLP formation, antibody neutralizing activity, and protective efficacy were not evaluated. Further studies in dogs, including optimized antigen-display platforms, neutralization assays, and protection studies, are required to determine the practical value of these epitopes for CPV vaccine development. Full article

Porcine circovirus type 3 (PCV3) capsid protein (Cap) is a key antigen for immunological studies and vaccine development. Different optimized PCV3 ORF2 sequences were used to construct six baculovirus transfer plasmids, with the pOET1.1-based design yielding the highest Cap level. Cap expression was confirmed by Western blot, IPMA and IFA. Recombinant baculovirus amplification was optimized, achieving the highest titer at an MOI of 0.1 with a 72 h harvest to 10^7.5TCID₅₀/0.1 mL, while maximal Cap production was obtained at an MOI of 0.1 with a 96 h harvest. PCV3 Cap virus-like particles (VLPs) were purified by sucrose density-gradient ultracentrifugation and cation-exchange chromatography, and TEM revealed spherical particles of approximately 17–20 nm. In mice, VLP immunization induced increasing antigen-specific IgG from day 14. Immunization increased both IgG1 and IgG2a without a significant difference, and post-immunization serum specifically recognized PCV3-positive passaged PK-15 cells in an indirect immunofluorescence assay. In splenic lymphocytes, IFN-γ, TNF-α, IL-4, and IL-10 mRNA levels were significantly upregulated (p < 0.01). Moreover, pig challenge data supported the protective potential of PCV3 Cap VLPs in the natural host. In our study, Cap assembled into VLPs and induced immune responses, providing a basis for PCV3 subunit vaccine development. Full article

Background/Objectives: Recombinant poliovirus (PV) virus-like particle (VLP) antigens mimic the conformation of the surface proteins in native PVs (i.e., serotype-specific D-antigen epitopes). Since they lack genomes and are non-infectious, PV-VLPs offer the promise of a safer, next-generation polio vaccine compared to traditional inactivated (IPV) or attenuated live (OPV) vaccines. Sandwich D-antigen ELISA formats are commonly used to measure the in vitro potency values (relative D-antigen content, DU/mL) of unadjuvanted trivalent IPV antigens. If IPV is formulated with aluminum-salt adjuvants, however, a pretreatment step (i.e., adjuvant dissolution or antigen desorption) is required, which may compromise antigen integrity during sample handling. Methods: This work describes the development of three competitive ELISAs to measure the relative D-antigen content of aluminum-salt adjuvanted PV-VLPs (Types 1, 2, 3) without the need for pretreatment. Results: First, key assay parameters were established, including specificity, accuracy, precision, linearity, limit of quantification, and stability-indication. Next, preformulation characterization studies were performed with these methods including (1) rank-ordering the inherent thermal stability profiles of the PV-VLPs (Types 1 > 3 > 2) in-solution and adsorbed to an aluminum phosphate adjuvant (AdjuPhos™, AP) and (2) determining the effect of formulation variables on the thermal stability profiles of AP-adsorbed PV-VLPs including antimicrobial preservatives (thimerosal, 2-PE) and five different antigens present in pediatric combination vaccines (D, T, wP, Hib, Hep B). Conclusions: The development and application of three competitive D-antigen ELISAs were demonstrated, and future use in formulation and storage stability studies with the AP-adjuvanted, trivalent PV-VLPs (Types 1, 2, 3) is discussed with the long-term goal to develop a stable, efficacious, multi-dose, hexavalent combination vaccine presentation. Full article

Objectives: Membrane-enveloped virus-like particles (VLPs) constitute a versatile vaccine platform allowing for the display of heterologous viral surface antigens. The density of displayed antigens is paramount for the efficient elicitation of a strong cellular and humoral immune response. SARS-CoV-2 spike protein variants with engineered cytoplasmic tails (CTs) were generated to enhance decoration efficiency on the surface of VLPs formed by the HIV core protein Gag. These HIV (SARS-CoV-2) chimeric particles serve as a vaccine component prototype. Methods: Spike variants were first analyzed for cellular and surface expression as well as incorporation into extracellular vesicles (EVs) and VLPs using flow cytometric analysis and Western blot analysis. Receptor binding, fusogenicity, i.e., mediating the fusion of spike-positive with receptor-containing membranes, and the proteins’ potential to mediate lentiviral vector gene transduction into susceptible target cells was examined by employing syncytia-formation assays and vector titration experiments. The display of a neutralization-sensitive epitope was examined utilizing immuno-precipitation using a neutralizing antibody. Results: All four variants were shown to be cell-surface expressed, to recruit the cognate receptor, to mediate membrane fusion and cell entry of lentiviral pseudotype vector particles and to decorate VLPs and EVs. However, the spike variant encompassing a truncated CT derived from the gibbon ape leukemia virus (GaLV) transmembrane (TM) envelope protein was most efficiently incorporated into HIV Gag-formed VLPs. All variants exposed a neutralization-sensitive epitope in the receptor binding domain. Conclusions: Engineering of the CTs of viral surface antigens can enhance VLP decoration, while required functionality of the ecto-domain such as receptor recognition, fusogenicity and neutralization-sensitive epitope presentation are not abrogated. This indicates the preservation of the structural integrity of the antigen required to elicit a neutralizing humoral immunity upon vaccination. The identified truncated CT of GaLV TM may be of utility to improve the incorporation of other viral surface antigens into a variety of membrane-enveloped VLPs derived from a range of different parental viruses. Full article

This review systematically evaluates research on self-assembling protein nanoparticles (SPNPs)-based vaccine, focusing on three major categories: virus-like particles (VLPs), natural protein nanoparticles (e.g., ferritin and encapsulin), and computationally designed protein nanoparticles—with comparative analyses of their design strategies, immunogenicity, and applicability in next-generation vaccines. VLPs can elicit robust humoral and cellular immune responses due to their virus-mimetic structures. Natural protein nanoparticles provide excellent biocompatibility and controllable self-assembly for multivalent antigen presentation. Artificially designed protein nanoparticles allow precise structural optimization to facilitate tailored antigen display and immune modulation. Overall, SPNPs constitute a versatile and powerful platform for vaccine development. By integrating natural and engineered design principles, they offer new opportunities for rational vaccine design and the development of safer and more effective immunization strategies. Full article

Background: Prophylactic human papillomavirus (HPV) vaccines are crucial for preventing HPV-related cancers. This study aimed to preclinically evaluate a novel recombinant 9-valent HPV vaccine produced in Escherichia coli (E. coli), which targets HPV types 6, 11, 16, 18, 31, 33, 45, 52, and 58, and is based on virus-like particles (VLPs) of the HPV major capsid protein L1. Methods: The molecular weight and purity of HPV L1 protein bands were assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) with Coomassie Brilliant Blue staining. The morphology and size distribution of VLPs were characterized using cryo-electron microscopy and DLS. The immunogenicity and durability of the recombinant 9-valent HPV vaccine were evaluated in BALB/c mice and Wistar rats. Mice received single or triple immunizations (2-week intervals) of two vaccine batches or Gardasil^®9 (MSD, USA) control at 1/20 human dose. Antibody responses were monitored via ELISA and pseudovirus neutralization assays over 24 weeks. Rats were administered single or triple immunizations (2-week intervals) of high- (1/10), medium- (1/20), or low-dose (1/40) vaccine or Gardasil^®9 control (1/20), with neutralizing antibodies tracked for 16 weeks. Results: Cryo-electron microscopy and DLS revealed that VLPs of each type appeared as uniformly distributed, spherical or ellipsoidal hollow intact particles with a diameter of approximately 45–65 nm. This vaccine demonstrated robust immunogenicity and long-lasting efficacy in BALB/c mice and Wistar rats, with effects comparable to those of the commercially available vaccine Gardasil^®9. Conclusions: The 9-valent HPV vaccine induces robust and persistent immune responses in mice and rats, strongly supporting further clinical trials. It is expected to be an alternative to marketed vaccines and ease the global supply shortage of 9-valent HPV vaccines. Full article

Background/Objectives: Cervical cancer, predominantly driven by persistent infection with high-risk human papillomaviruses (HPVs), is one of the most common malignancies and an important cause of cancer-related mortality among women worldwide. Although existing licensed prophylactic HPV vaccines confer excellent protection, their global use remains suboptimal due to concentrated manufacturing capacity and high production costs. This study aimed to establish a cost-effective multivalent HPV virus-like particle (VLP) vaccine platform. Specifically, we used an enhanced baculovirus expression vector system to produce a 12-valent HPV VLP vaccine to improve antigen yield, thereby reducing manufacturing costs and ultimately improving affordability and availability in low- and middle-income countries. Methods: Optimized expression cassettes and an insect cell culture process were designed to enhance productivity across 12 HPV L1 genotypes. A scalable purification scheme integrating ion-exchange and adsorption chromatography was developed to produce high-purity VLPs with consistent structural integrity. Immunogenicity was assessed in a murine model. Elicited HPV type-specific IgG antibody responses were compared with those induced by the licensed 9-valent HPV vaccine. Results: The assembled 12-valent VLPs were comprehensively characterized using biophysical and immunochemical analyses, confirming structural stability and correct antigenicity. In vivo immunogenicity studies in mice showed strong and serotype-specific IgG responses, comparable or superior to those induced by the licensed 9-valent commercial vaccine. Conclusions: The enhanced baculovirus expression vector system is a versatile and economically sustainable platform for next-generation HPV vaccine production. This technology offers a promising approach to lowering vaccine manufacturing costs and improving global access, particularly in low- and middle-income regions heavily burdened by HPV-associated diseases. Full article

Virus-like particles (VLPs), nanoscale self-assembled structures lacking viral genetic material, have emerged as a versatile platform for vaccines, targeted delivery systems, and gene-editing applications owing to their strong immunogenicity, favorable biosafety profile, and high engineerability. However, the complex architecture of VLPs, their significant size heterogeneity, and the diversity of process- and product-related impurities generated in different expression systems make downstream purification a major bottleneck limiting product quality, yield, and manufacturability. This review systematically discusses advanced downstream purification strategies for VLPs from the perspective of three major objectives: preservation of structure and biological activity, control of product heterogeneity, and assurance of viral safety. First, strategies for maintaining VLP integrity and function are examined, including optimization of solution conditions, adoption of gentle yet efficient separation operations, and integration of process analytical technology (PAT) to reduce process-induced damage. Second, the review summarizes multi-step purification approaches—spanning clarification, ultrafiltration/diafiltration (UF/DF), chromatography, and disassembly/reassembly—to remove host cell proteins, host cell DNA, and product-related impurities while improving particle homogeneity and stability. Third, viral safety is discussed primarily from the perspective of downstream virus clearance under host-dependent risk, with particular attention to orthogonal clearance steps tailored to VLP properties and expression systems such as CHO cells and insect cell–baculovirus platforms. Overall, this review provides a CQA-oriented framework and practical guidance for the development of robust, scalable, and GMP-compliant downstream purification processes for VLP-based products. Full article

Chikungunya virus (CHIKungunya Virus, CHIKV) is a mosquito-borne plus-stranded RNA virus. Adaptive mutations such as A226V in the E1 envelope protein of CHIKV significantly enhance the transmission efficiency of the virus in Aedes albostriae, leading to multiple rounds of epidemics around the world including the large-scale outbreak in Guangdong Province in 2025. After a viral infection, a significant proportion of patients will progress from acute arthralgia to chronic arthritis that persists. The pathogenesis of the disease involves the persistence of the virus in joint tissues, the persistent inflammatory response with IL-1β, IL-6 and IL-17 as the core mediated by macrophages, possible autoimmune cross-reactions, and individual genetic susceptibility. At present, there is no specific antiviral drug, but important progress has been made in vaccine development against the virus. Vaccines based on live attenuated virus (VLA1553) and virus-like particle (VLP) platforms have been approved for the market and provide a tool to prevent and control this important public health threat. This review synthesizes current knowledge on CHIKV-induced chronic arthritis pathogenesis and recent vaccine advances, providing a framework for understanding disease mechanisms and guiding future prevention strategies. Full article

Background/Objectives: Pathogenic mammarenaviruses cause severe hemorrhagic and neurologic disease in humans. Machupo virus (MACV), a New World (NW) mammarenavirus, causes Bolivian hemorrhagic fever in humans, and there are no approved vaccines. Methods: Here, we describe and compare the immunogenicity of three vaccines expressing the MACV glycoprotein complex (GPC) in C57BL/6 mice: a recombinant vesicular stomatitis virus (rVSV) and two different lipid nanoparticle (LNP)-encapsulated nucleoside-modified mRNA (mRNA-LNP) vaccines. The first mRNA-LNP vaccine, designated MACV mRNA, expresses the full-length MACV GPC. The second mRNA-LNP vaccine, called MACV VLP mRNA, encodes MACV GPC with appended sequences that induce the budding of virus-like particles (VLPs) with MACV GPC on the surface. This is the first description of any mRNA-LNP vaccine for MACV and the first comparison of mRNA and rVSVs as vaccine candidates for MACV. Results: We find that two doses of either MACV mRNA or MACV VLP mRNA are required for the induction of robust humoral and cellular immune responses including total MACV GPC IgG, neutralizing antibodies, cross-reactive antibodies that bind the related Junín virus GPC, and MACV-specific T-cell responses. To further investigate vaccination strategies for MACV, we also evaluated a heterologous prime-boost regimen involving the MACV mRNA vaccine coupled with the rVSV-based MACV vaccine. We find that the highest levels of MACV GPC-specific IgG and neutralizing titers were achieved when heterologous mRNA and rVSV prime-boost regimens were employed. Conclusions: These results elucidate differences in the immune response to different vaccine platforms for MACV and can inform future vaccine development for NW arenaviruses. Full article

Background: African swine fever virus (ASFV) causes a fatal hemorrhagic disease in domestic pigs and wild boars. While live attenuated vaccines (LAVs) provide protection, their use raises safety concerns. Therefore, the aim of the present study was to identify viral B-cell antigens associated with protection and to test their potential using highly immunogenic vaccine delivery platforms. Methods: We employed a microarray of 169 ASFV proteins expressed in a cell-free prokaryotic system to identify immunodominant antigens using sera from immune pigs. Six structural proteins were selected and formulated into AP205 virus-like particles (VLPs). Additionally, replication-defective vesicular stomatitis virus (VSV)-based vaccine candidates expressing glycosylated CD2v and EP153R proteins were generated. Three groups of specific pathogen-free pigs were immunized with either VLP- or VSV-based vaccines and challenged with the virulent ASFV Georgia 2007 strain. Control groups included pigs immunized with the attenuated ASFV Estonia 2014 strain and a naïve group. Results: Most vaccine candidates induced detectable antibody responses against target ASFV proteins. However, neither VLP- nor VSV-based vaccines provided protection, as clinical scores, hematology, cytokine responses, and viremia levels were similar to those in the negative control group. In contrast, only the ASFV Estonia 2014 strain elicited a robust T-cell response and protective immunity. Conclusions: These findings highlight the challenges in identifying protective B-cell antigens of ASFV and emphasize the pivotal role of cellular immunity in mediating protection. Full article

Background: Cervical carcinoma remains a widespread cancer worldwide, primarily caused by persistent infection with high-risk human papillomavirus (HPV). HPV types 16 and 18 account for approximately 70% of cervical cancer cases. Although prophylactic HPV vaccines are commercially available, their high cost and reliance on expensive expression platforms limit their accessibility in developing countries. Objectives: This study aimed to develop a cost-effective, plant-based HPV vaccine candidate by expressing capsomeric HPV-16 and HPV-18 L1 antigens in Brassica oleracea (broccoli). Methods: Modified L1 from HPV types 16 and 18 were designed to retain capsomeric assembly and fused with heat-labile enterotoxin B subunit (LTB). Immunoinformatics analyses were used to assess antigenicity, epitope distribution, and structural characteristics. Codon-optimized genes were cloned using Gateway^® technology and expressed in broccoli via Agrobacterium-mediated transformation. Transgenic plants were validated by PCR and qRT-PCR. Protein accumulation was quantified, and immunogenicity was evaluated in mice. Results: PCR and qRT-PCR confirmed the stable integration of two copies of the LTB-L1 transgenes in broccoli plants. Western blotting detected L1 protein at ~56.5 kDa, indicating the cleavage of the LTB-L1 fusion protein. The correct folding of L1 capsomeres was verified by antigen-capture ELISA. The recombinant proteins accumulated to approximately 0.33% and 0.35% of total soluble protein for HPV-16 and HPV-18, respectively. The immunization of mice with transgenic L1 induced significant humoral immune responses, comparable to those elicited by purified VLPs. Conclusions: The results demonstrate broccoli as a promising platform for the expression of immunogenic HPV L1 capsomeres and highlight its potential for the development of affordable, plant-based HPV vaccines. Full article

As SARS-CoV-2 continues to evolve with increased transmissibility and immune evasion, the need for vaccines that provide broader and more durable protection has become increasingly urgent. The extensive research spurred by the pandemic has accelerated the development of diverse vaccine platforms, including mRNA, DNA, virus-like particles (VLPs), recombinant proteins, and mosaic mono- and polyvalent vaccines. While several of these platforms have reached regulatory approval and widespread clinical employment, others remain under evaluation or in various stages of clinical development. These vaccines have significantly reduced infection rates, severe disease, and hospitalizations, particularly among high-risk group. Nevertheless, the ongoing emergence of novel variants and subvariants has challenged the efficacy of both existing and newly developed vaccines. This evolving landscape underscores the urgent need for a universal SARS-CoV-2 vaccine platform capable of providing comprehensive and long-lasting immunity. In this review, we evaluate current and emerging strategies for SARS-CoV-2 universal vaccine development, with a focus on antigen design, breadth of immune protection, and clinical feasibility. Attention is given to various universal vaccine platforms such as the mosaic polyvalent spike construct, multi-epitope vaccines targeting the receptor-binding domain (RBD), and approaches centered on the conserved S2 subunit of the spike protein. We also discuss strategies leveraging additional conserved viral proteins and T helper (Th) and cytotoxic T lymphocyte (CTL) epitopes from across coronaviruses. By highlighting the advances in these areas, this review provides a framework to guide the rational design of next-generation universal vaccines capable of delivering broad and durable protection against SARS-CoV-2 variants. Full article

Poliovirus (PV), a historically significant enterovirus responsible for severe paralytic diseases, has seen its incidence dramatically reduced through widespread vaccination efforts, propelling global eradication initiatives. Despite the success of traditional oral poliovirus vaccines (OPVs) and inactivated poliovirus vaccines (IPVs), challenges such as vaccine-derived virus reversion and biosafety concerns during vaccine production persist. Virus-like particle (VLP) vaccines, which mimic native viral structures without containing viral genomes, offer enhanced safety profiles and robust immunogenicity, positioning them as promising candidates for next-generation poliovirus vaccines, especially in the post-certification era. This review systematically summarizes current progress in poliovirus VLP vaccine research, including the diverse expression systems employed for VLP production, strategies for peptide assembly and stabilization, and evaluations of antigenicity and immunogenicity. Additionally, it highlights structural analyses utilizing cutting-edge cryo-electron microscopy. By integrating recent developments in genetic engineering, structural biology, and immunology, this article discusses the advantages and challenges associated with poliovirus VLP vaccines and explores future directions aimed at supporting the global goal of a poliovirus-free world. This comprehensive overview aims to provide a theoretical foundation and technical guidance to facilitate the development and deployment of safer and more effective poliovirus vaccines. Full article