Rabies is a deadly viral disease caused by the rabies virus (RABV), transmitted through a bite of an infected host, resulting in irreversible neurological symptoms and a 100% fatality rate in humans. Despite many aspects describing rabies neuropathogenesis, numerous hypotheses remain unanswered and concealed. Observations obtained from infected primary neurons or mouse brain samples are more relevant to human clinical rabies than permissive cell lines; however, limitations regarding the ethical issue and sample accessibility become a hurdle for discovering new insights into virus–host interplays. To better understand RABV pathogenesis in humans, we generated human-induced pluripotent stem cell (hiPSC)-derived neurons to offer the opportunity for an inimitable study of RABV infection at a molecular level in a pathologically relevant cell type. This study describes the characteristics and detailed proteomic changes of hiPSC-derived neurons in response to RABV infection using LC-MS/MS quantitative analysis. Gene ontology (GO) enrichment of differentially expressed proteins (DEPs) reveals temporal changes of proteins related to metabolic process, immune response, neurotransmitter transport/synaptic vesicle cycle, cytoskeleton organization, and cell stress response, demonstrating fundamental underlying mechanisms of neuropathogenesis in a time-course dependence. Lastly, we highlighted plausible functions of heat shock cognate protein 70 (HSC70 or HSPA8) that might play a pivotal role in regulating RABV replication and pathogenesis. Our findings acquired from this hiPSC-derived neuron platform help to define novel cellular mechanisms during RABV infection, which could be applicable to further studies to widen views of RABV-host interaction.

Porcine circovirus type 2 (PCV2) is the main causative agent of porcine circovirus-associated disease (PCVAD) that profoundly impacts the swine industry worldwide. While most of the commercial PCV vaccines are developed based on PCV genotype 2a (PCV2a), PCV genotype 2b (PCV2b) has become predominant since 2003. In this study, we developed and evaluated DNA-based bivalent vaccines covering both PCV2a and PCV2b. We generated a new immunogen, PCV2b-2a, by combining consensus sequences of the PCV2a and PCV2b capsid proteins (Cap2a and Cap2b) in a form of fusion protein. We also examined whether modifications of the PCV2b-2a fusion protein with a signal sequence (SS) and granulocyte macrophage-colony stimulating factor (GM-CSF) fusing with interleukine-4 (IL-4) (GI) could further improve the vaccine immunogenicity. An immunogenicity study of BALB/cAJcl mice revealed that the DNA vector pVAX1 co-expressing PCV2b-2a and GI (pVAX1.PCV2b-2a-GI) was most potent at inducing both antibody and cellular immune responses against Cap2a and Cap2b. Interestingly, the vaccines skewed the immune response towards Th1 phenotype (IgG2a > IgG1). By performing ELISA and ELISpot with predicted epitope peptides, the three most immunogenic B cell epitopes and five putative T cell epitopes were identified on Cap2a and Cap2b. Importantly, our DNA vaccines elicited broad immune responses recognizing both genotype-specific and PCV2-conserved epitopes. Sera from mice immunized with the DNAs expressing PCV2b-2a and PCV2b-2a-GI significantly inhibited PCV2a cell entry at serum dilution 1:8. All these results suggest a great potential of our PCV2b-2a-based vaccines, which can be further developed for use in other vaccine platforms to achieve both vaccine efficacy and economical production cost.

Background/Objectives: African swine fever (ASF) is a devastating disease affecting the swine industry globally. Development of safe and effective vaccines is an urgent need. This study aimed to evaluate, caASFV001-MA52, a cell-adapted ASFV strain derived from serial passaging in MA-104 cells, as a promising live-attenuated vaccine (LAV) candidate against virulent ASFV infection. Methods: Seven-week-old, crossbred pigs were immunized with caASFV001-MA52 (at a dose of 10⁵ TCID₅₀) and subsequently challenged with a lethal dose of virulent ASFV. Vaccine efficacy was measured through clinical monitoring, immunological and virological analyses, and pathological assessments of tissue protection and viral load reduction. Safety was critically assessed, particularly regarding its profile in animals with concurrent endemic porcine infections, including PCV, PRRSV and S. suis. Results: caASFV001-MA52 exhibits partial protection (70–80%) against the lethal challenge. Vaccinated and challenged survivors exhibited reduced viral loads and significantly alleviated pathological lesions compared to controls. Safety evaluations revealed that the vaccine’s profile is susceptible to concurrent infection. Pigs co-infected with endemic porcine pathogens showed increased virulence and mortality following vaccination. Although vaccination temporarily reactivated latent viral infections (PCV2, PCV3, and PRRSV), most surviving pigs effectively controlled and eliminated these co-infections. Conclusions: The caASFV001-MA52 strain demonstrates promising immunogenicity and protection against lethal challenges, supporting its continued development as an LAV candidate. However, the observed safety concerns regarding concurrent infections emphasize the critical need for veterinary health surveillance during its future practical application.

Background: Middle East Respiratory Syndrome Coronavirus (MERS-CoV) causes severe respiratory illness in humans and currently lacks an approved vaccine. The Newcastle disease virus (NDV) vector is a well-established, safe, and effective platform for vaccine development. With recent advancements in stabilizing coronavirus spike proteins to enhance their antigenicity, this study aimed to determine whether modifications to the MERS-CoV spike protein could improve its presentation on NDV particles, allowing the resulting virus to be used as an inactivated vaccine. Methods: We codon-optimized the gene encoding the ectodomain of the MERS-CoV spike protein and incorporated modifications at the S1/S2 and S2’ cleavage sites, along with a proline substitution at residues V1060-L1061. This modified spike gene was inserted into the NDV genome to create the NDV-S_MERS virus. After purification and inactivation, the vaccine’s immunogenicity was assessed in mice. Results: Mice immunized with the inactivated NDV-S_MERS vaccine developed robust anti-spike IgGs, neutralizing antibodies, and cellular immune responses. The study demonstrated that modifications to the MERS-CoV spike protein were essential for its effective presentation on NDV particles. Additionally, the spike gene insert remained stable through five egg passages, confirming the vector’s stability. Conclusions: Engineering the MERS-CoV spike protein is crucial for its successful display on NDV particles. The strong immune responses elicited by the NDV-S_MERS vaccine in mice highlight that NDV is a promising, safe, and effective platform for MERS-CoV vaccination.

Infectious diseases in swine demand affordable, broad-spectrum antiviral strategies, such as recombinant interferons (IFNs). However, commercial yeast expression platforms often present financial barriers through licensing royalties. To provide a cost-effective regional alternative, we evaluated the domestic, thermotolerant yeast Ogataea polymorpha OP39 for producing recombinant porcine IFNs. Recombinant OP39 strains carrying expression cassettes for types I (poIFN-α1), II (poIFN-γ), and III (poIFN-λ3) IFNs were constructed and evaluated for their recombinant protein production performance. Following initial screening, the highest-yielding candidate, poIFN-α1, was selected for bioprocess optimization in shake flasks and a 5 L fermenter. Results showed that OP39 successfully secreted all three IFN types with a relatively low level of hyperglycosylation. Batch fermentation scale-up of r-poIFN-α1 achieved a final protein concentration of 0.45 ± 0.10 mg/mL. Crucially, the OP39-produced r-poIFN-α1 exhibited antiviral efficacy in vitro against economically important porcine viruses including African Swine Fever Virus, Porcine Epidemic Diarrhea Virus and Porcine Reproductive and Respiratory Syndrome Virus. Based on the proof-of-concept results in this study, the OP39 platform can be further developed into a viable, full freedom-to-operate system for local or regional biomanufacturing of biologically active veterinary therapeutics to support livestock health management.