In-Ohk Ouh

Background/Objectives: Tuberculosis (TB) remains a major global health challenge, requiring standardized animal models to evaluate vaccine-induced immune responses. This study characterized time-dependent immune responses following Bacillus Calmette–Guérin (BCG) vaccination in BALB/c mice. Methods: BALB/c mice were vaccinated with BCG, and the immune responses and protective efficacy were evaluated at 4, 6, and 8 weeks post-immunization. The cytokine expression in serum, lung, and spleen tissues was analyzed using ELISA, quantitative PCR, and immunohistochemistry. Protective efficacy was assessed via colony-forming unit (CFU) enumeration and the immunohistochemical detection of Mycobacterium TB after aerosol challenge. Results: The BCG vaccination induced time-dependent and tissue-specific cytokine responses. Pulmonary IL-1β and splenic IFN-γ levels were significantly increased four weeks post-vaccination. At 8 weeks, serum IL-2, pulmonary IL-2, and TNF-α were significantly increased, whereas no significant changes in cytokines were observed at 6 weeks. After the challenge, BCG-vaccinated mice exhibited reduced bacterial burdens compared with controls, but the differences among the 4-, 6-, and 8-week groups were modest. Conclusions: Immune responses became detectable starting four weeks after BCG vaccination, with temporal differences observed in cytokine expression. Week 8 may serve as a reference point for monitoring cytokine dynamics rather than as an optimal time for protection.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging tick-borne infectious disease with high case-fatality rates in East Asia. The causative agent, SFTS virus (SFTSV; also known as Dabie bandavirus), exhibits genotype-dependent differences in pathogenicity. However, infection models that recapitulate these variations and can be applied for vaccine and therapeutic evaluation are still lacking. In this study, we assessed the pathogenicity of two Korean SFTSV isolates representing the F and B genotypes in a murine infection model. Wild-type C57BL/6 and IFNAR knockout (IFNAR−/−) mice were intraperitoneally infected with two different doses of SFTSV (2 and 2 × 10⁻¹ FFU). All C57BL/6 mice survived regardless of viral genotype or dose. In IFNAR−/− mice, infection with either F- or B-type virus at the 2 FFU dose resulted in mortality beginning at 5 days post-infection, with all mice succumbing within 6 days. At the higher dose (2 × 10⁻¹ FFU), mortality differed by genotype: B-type infection led to 20% lethality, whereas F-type infection caused 40% lethality by day 5. Infected and deceased mice exhibited body weight loss as a characteristic clinical outcome. Collectively, these findings demonstrate genotype-associated differences in SFTSV pathogenicity in mice and establish a murine challenge model that may be useful for the preclinical evaluation of candidate vaccines and antiviral agents.

Background: Highly pathogenic avian influenza H5N1 viruses of clade 2.3.4.4b have spread globally since 2021, causing extensive outbreaks in avian populations and repeated spillovers into diverse mammalian hosts, including humans. These cross-species transmission events highlight ongoing pandemic risks and underscore the need for vaccine strategies that reflect viral evolution at the human–animal interface. Despite the availability of licensed H5 vaccines and newly recommended World Health Organization (WHO) candidate vaccine viruses (CVVs), the extent to which these vaccines elicit cross-reactive antibody responses against contemporary clade 2.3.4.4b viruses, including mammalian spillover isolates of avian origin, remains incompletely characterized. Method: In this study, ferret antisera were generated using four WHO-recommended H5 CVVs, including a clade 1 strain (A/Vietnam/1194/2004) and three clade 2.3.4.4b strains (A/Astrakhan/3212/2020, A/American wigeon/South Carolina/22-000345-001/2021, and A/Ezo red fox/Hokkaido/1/2022), formulated with alum adjuvant to reflect licensed vaccine formulation used in national preparedness programs. Antibody responses and cross-reactive activity were evaluated using hemagglutination inhibition (HI) and microneutralization (MN) assays against homologous vaccine strains and a feline-origin clade 2.3.4.4b H5N1 field isolate from Korea, A/Feline/Korea/SNU-01/2023. Results: Antisera induced by clade 2.3.4.4b CVVs showed cross-reactive antibody responses against homologous and heterologous clade 2.3.4.4b viruses and demonstrated measurable HI and MN responses against the feline-origin field isolate. In contrast, antisera raised against the clade 1 Vietnam CVV exhibited limited cross-reactivity against clade 2.3.4.4b viruses. Overall, clade 2.3.4.4b CVVs generally showed higher antibody responses than the clade 1 vaccine strain across multiple panels. Conclusions: These findings provide descriptive insights into antigenic differences between clade 1 and clade 2.3.4.4b viruses and support the antigenic relevance of clade 2.3.4.4b CVVs for contemporary H5N1 strains. This study highlights the importance of ongoing antigenic evaluation to inform vaccine strain selection within a One Health framework.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a tick-borne zoonotic pathogen that continues to cause a substantial public health burden in South Korea, yet the molecular epidemiology of SFTSV in dogs, particularly shelter populations, remains poorly characterized. To address this gap, blood samples from 715 dogs, including companion and shelter animals, were collected nationwide in 2024 and screened for SFTSV using RT-PCR targeting the S, M, and L genomic segments, followed by sequencing, phylogenetic analysis, and virus isolation. SFTSV was detected in 16 dogs (2.2%), with significantly higher prevalence in autumn, in the southern region, in shelter dogs, and in younger animals. A localized cluster of six infected dogs was identified in a southern shelter during autumn, and phylogenetic analysis revealed the circulation of three genotypes (B2, D, and F). Live virus was successfully isolated from one shelter dog, and this isolate belonged to genotype F, representing the first isolation of this genotype from dogs in South Korea. These findings demonstrate that dogs are exposed to genetically diverse SFTSV strains circulating in tick populations and support the use of canine surveillance, particularly in shelters, as part of a One Health approach to monitor and mitigate the risk of SFTSV transmission.

Severe fever with thrombocytopenia syndrome virus (SFTSV) is a tick-borne zoonotic pathogen of significant public health concern in South Korea, where human cases continue to occur at high levels; however, information on the molecular epidemiology and genotype diversity of SFTSV in goats—an increasingly important livestock species—remains limited. In this study, blood samples were collected from 750 clinically healthy goats during nationwide surveillance in 2024. Viral RNA was detected by RT-PCR targeting the S and M genomic segments. Epidemiological characteristics were analyzed according to season, region, farm size, breed, and sex. Positive samples were subjected to sequencing and phylogenetic analysis to determine SFTSV genotypes. SFTSV RNA was detected in 10 of 750 goats (1.3%), with significantly higher detection rates in autumn compared with summer, in southern regions compared with northern regions, and in female goats compared with males, while no significant association was observed with farm size or breed. Phylogenetic analysis showed that goat-derived SFTSV strains belonged to genotypes B2, D, and F; notably, genotypes D and F were identified in goats for the first time in South Korea. These findings indicate that goats are exposed to genetically diverse SFTSV strains circulating in tick populations and exhibit epidemiological patterns consistent with tick ecology and human SFTS incidence, supporting the role of goats as incidental or sentinel hosts. Continuous molecular surveillance of goats, integrated with vector monitoring programs, may enhance understanding of regional SFTSV transmission dynamics and facilitate early detection of emerging genotypes with public health implication.

Background: Nipah virus (NiV), a zoonotic paramyxovirus with high case fatality and pandemic potential, remains without a licensed vaccine for humans to date. Although there has been progress in vaccine development, it remains limited, and peptide vaccines have rarely been validated in vivo. Methods: Here, we report the rational antigen selection, synthesis, and preliminary immunogenicity evaluation of NiV fusion glycoprotein (NiV-F)-derived linear peptides as vaccine candidates. Candidate epitopes were identified by in silico, and a total of 18 B- and T-cell epitope-derived peptides were shortlisted for synthesis and antigenicity validation by ELISA. Results: Antigenicity evaluation showed that 9 of the synthesized peptides have A_450nm of over 1 (8 from the F11 group, A_450nm: 1.13–3.6; 1 from the F18 group, A_450nm: 1.51), with the peptide constructs F11-3 (A_450nm: 3.5) and F11-4 (A_450nm: 3.6) showing the highest antigenicity. Interestingly, peptides from F11 with amidation increased antibody binding (F11-4-NH2, A_450nm: 3.05; F11-4-9mer-1-NH2, A_450nm: 0.87). The lead peptide candidates, F11-3 and F11-4, were subsequently used for the immunization experiment, and mouse sera were assessed against their homologous peptide antigens or recombinant NiV-F protein. ELISA result showed detectable antibody reactivity against their homologous antigen for the intramuscular (IM) F11-3 vaccinated group (A_450nm: 0.30 ± 0.35), whereas increased binding was observed for both IM-administered F11-3 (A_450nm: 1.62 ± 0.97) and F11-4 (A_450nm: 2.0 ± 0.77) against NiV-F protein, albeit without statistical significance compared to the negative control (NC, p > 0.05), and were markedly lower compared to mice immunized with NiV-F recombinant protein (PC, p < 0.01), underscoring the need for further optimization procedures. Conclusions: Collectively, these results support an exploratory antigen discovery and prioritization framework for NiV-F-derived peptide candidates and provide a foundation for future studies aimed at optimizing immunogenicity and evaluating protective relevance in appropriate preclinical models.

Background: The emergence of several paramyxoviruses, including Nipah virus (NiV), makes continued efforts in vaccine development as part of pandemic preparedness efforts necessary. Although NiV is a zoonotic pathogen with high case fatality, there is still no licensed vaccine. Methods: Herein, NiV attachment glycoprotein G (NiV-G), which is crucial to host cell receptor binding, was used to develop Nipah epitope-based peptide vaccines. A total of 39 B- and T-cell epitopes of NiV-G were shortlisted for peptide synthesis and evaluation using in silico analysis. Results: The in vitro antigenicity evaluation of the peptide candidates showed eight synthesized peptides (G7, stalk-domain epitopes) with relatively high binding to NiV-G antibody-positive serum (A_450nm: 1.39–3.78). Moreover, nine-mer (9-mer) peptides were found to be less reactive than their longer peptide counterparts (15–30 aa, G7-1, and G7-4), but 9-mer activity was enhanced with cyclization (NPLPFREYK, A_450nm: 2.66) and C-terminal amidation modification (NPLPFREYK-NH2, A_450nm: 1.39). Subsequently, in vivo validation in immunized mice revealed the immunogenicity potential of the G7-1 peptide vaccine (30 aa, NENVNEKCKFTLPPLKIHECNISCPNPLPF) to elicit a strong antigen-specific antibody response against their homologous peptide antigen (I.V., A_450nm: 1.48 ± 0.78; I.M., A_450nm: 1.66 ± 0.66). However, antibody binding to recombinant NiV-G protein remained low, suggesting limited recognition to the native antigen. Conclusions: This study focused on the preliminary screening and validation of peptide vaccines using single formulations with minimal modifications in the peptide candidates. Our findings collectively show the immunogenic potential of the NiV-G stalk-based epitope peptide vaccine as a novel therapeutic for NiV and underscores the need for strategic design, delivery, and formulation optimization to enhance its protective efficacy and translational application.

Background/Objectives: Tuberculosis (TB), caused by Mycobacterium tuberculosis H37Rv (M. tuberculosis), primarily affects the lungs. The Bacillus Calmette–Guérin (BCG) vaccine is the only available TB vaccine. Guinea pigs serve as an excellent preclinical model due to the similarity to human Tuberculosis pathology. However, the lack of a standardized vaccination protocol in guinea pigs causes inconsistencies in efficacy assessments, limiting precise evaluation and its application in vaccine studies. This study aims to address this gap by establishing a consistent and reliable protocol for evaluating the immunological efficacy of BCG vaccination. Methods: Guinea pigs were divided into control, M. tuberculosis-infected, and BCG-vaccinated groups. Four weeks post-vaccination, the infected and vaccinated groups were challenged with M. tuberculosis. The bacterial burden in the lungs and spleen was measured, histopathological changes were analyzed using hematoxylin and eosin (H&E) staining, and the infection levels of M. tuberculosis, as well as the presence of interleukin-2 (IL-2), tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ) positive cells, were evaluated through immunohistochemical (IHC) staining. Results: BCG vaccination reduced the bacterial load to 3.60 × 10⁴ CFU/lung and 5.52 × 10³ CFU/spleen compared to 3.78 × 10⁵ CFU/lung and 1.54 × 10⁴ CFU/spleen in the infected group. The mean histopathological score for lungs was 1.67 compared to 2.67 in the infected group. Similarly, the mean histopathological score for the spleen was 1.33 compared to 2.33 in the infected group. IHC analysis showed a notable reduction in M. tuberculosis and inflammatory cytokine-positive cells in the vaccinated group. The TNF-α, IL-2, and IFN-γ staining intensity decreased by 9.3, 4.8, and 11, respectively, compared to the infected group. Conclusions: This protocol enhances consistency in vaccine assessments, providing a reliable benchmark for the development of safer, more effective, and accessible TB vaccines.

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.

Nipah virus (NiV), of the Paramyxoviridae family, causes highly fatal infections in humans and is associated with severe neurological and respiratory diseases. Currently, no commercial vaccine is available for human use. Here, eight structure-based mammalian-expressed recombinant proteins harboring the NiV surface proteins, fusion glycoprotein (F), and the major attachment glycoprotein (G) were produced. Specifically, prefusion NiV-F and/or NiV-G glycoproteins expressed in monomeric, multimeric (trimeric F and tetra G), or chimeric forms were evaluated for their properties as sub-unit vaccine candidates. The antigenicity of the recombinant NiV glycoproteins was evaluated in intramuscularly immunized mice, and the antibodies in serum were assessed. Predictably, all homologous immunizations exhibited immunogenicity, and neutralizing antibodies to VSV-luciferase-based pseudovirus expressing NiV-GF glycoproteins were found in all groups. Comparatively, neutralizing antibodies were highest in vaccines designed in their multimeric structures and administered as bivalent (GMYtet + GBDtet) and trivalent (Ftri + GMYtet + GBDtet). Additionally, while all adjuvants were able to elicit an immunogenic response in vaccinated groups, bivalent (GMYtet + GBDtet) and trivalent (Ftri + GMYtet + GBDtet) induced more potent neutralizing antibodies when administered with oil-in-water nano-emulsion adjuvant, AddaS03. For all experiments, the bivalent GMYtet + GBDtet was the most immunogenic vaccine candidate. Results from this study highlight the potential use of these mammalian-expressed recombinant NiV as vaccine candidates, deserving further exploration.