Zika virus (ZIKV)-specific T cells are activated by different peptides derived from virus structural and nonstructural proteins, and contributed to the viral clearance or protective immunity. Herein, we have depicted the profile of CD8+ and CD4+ T cell immunogenicity of ZIKV proteins in C57BL/6 (H-2^b) and BALB/c (H-2^d) mice, and found that featured cellular immunity antigens were variant among different murine alleles. In H-2^b mice, the proteins E, NS2, NS3 and NS5 are recognized as immunodominant antigens by CD8+ T cells, while NS4 is dominantly recognized by CD4+ T cells. In contrast, in H-2^d mice, NS1 and NS4 are the dominant CD8+ T cell antigen and NS4 as the dominant CD4+ T cell antigen, respectively. Among the synthesized 364 overlapping polypeptides spanning the whole proteome of ZIKV, we mapped 91 and 39 polypeptides which can induce ZIKV-specific T cell responses in H-2^b and H-2^d mice, respectively. Through the identification of CD8+ T cell epitopes, we found that immunodominant regions E_294-302 and NS4_2351-2360 are hotspots epitopes with a distinct immunodominance hierarchy present in H-2^b and H-2^d mice, respectively. Our data characterized an overall landscape of the immunogenic spectrum of the ZIKV polyprotein, and provide useful insight into the vaccine development.

Enteroviruses have been a historical concern since the identification of polioviruses in humans. Wild polioviruses have almost been eliminated, while multiple species of non-polio enteroviruses and their variants co-circulate annually. To date, at least 116 types have been found in humans and are grouped into the species Enterovirus A–D and Rhinovirus A–C. However, there are few available antiviral drugs, especially with a universal pharmaceutical effect. Here, we demonstrate that peptide P25 from EV-D68 has broad antiviral activity against EV A–D enteroviruses in vitro. P25, derived from the HI loop and β-I sheet of VP1, operates through a conserved hydrophilic motif -R---K-K--K- and the hydrophobic F near the N-terminus. It could prevent viral infection of EV-A71 by competing for the heparan sulfate (HS) receptor, binding and stabilizing virions by suppressing the release of the viral genome. P25 also inhibited the generation of infectious viral particles by reducing viral protein synthesis. The molecular docking revealed that P25 might bind to the pocket opening area, a potential target for broad-spectrum antivirals. Our findings implicate the multiple antiviral effects of peptide P25, including blocking viral binding to the HS receptor, impeding viral genome release, and reducing progeny particles, which could be a novel universal anti-enterovirus drug candidate.

Yersinia pestis has a broad host range and has caused lethal bubonic and pneumonic plague in humans. With the emergence of multiple resistant strains and the potential for biothreat use, there is an urgent need for new therapeutic strategies that can protect populations from natural or deliberate infection. Targeting F1 has been proven to be the main strategy for developing vaccines and therapeutic antibodies, but data on anti-F1 antibodies, especially in humans, are scarce. To date, three human anti-F1 monoclonal antibodies (m252, αF1Ig2, and αF1Ig8) from naive populations have been reported. Here, we constructed an antibody library from vaccinees immunized with the plague subunit vaccine IIa by phage display. The genetic basis, epitopes, and biological functions of the obtained mAbs were assessed and evaluated in plague-challenged mice. Three human mAbs, namely, F3, F19, and F23, were identified. Their biolayer responses were 0.4, 0.6, and 0.6 nm, respectively. The dissociation constants (K _D ) of the F1 antigen were 1 pM, 0.165 nM, and 1 pM, respectively. Although derived from distinct Ab lineages, that is, VH3-30-D3-10-JH4 (F3&F23) and VH3-43-D6-19-JH4 (F19), these mAbs share similar binding sites in F1 with some overlap with αF1Ig8 but are distinct from αF1Ig2. Each of them provided a significant protective effect for Balb/c mice against a 100 median lethal dose (MLD) challenge of a virulent Y. pestis strain when administered at a dose of 100 µg. No synergistic or antagonistic effects were observed among them. These mAbs are novel and excellent candidates for further drug development and use in clinical practice. IMPORTANCE: In this study, we identified three human monoclonal antibodies with a high affinity to F1 protein of Yersinia pestis . We discovered that they have relatively lower somatic hypermutations compared with antibodies, m252, αF1Ig2, and αF1Ig8, derived from the naive library reported previously. We also observed that these mAbs share similar binding sites in F1 with some overlapping with αF1Ig8 but distinct from that of αF1Ig2. Furthermore, each of them could provide complete protection for mice against a lethal dose of Yersinia pestis challenge. Our data provided new insights into the anti-F1 Ab repertories and their associated epitopes during vaccination in humans. The findings support the additional novel protective human anti-F1Abs for potential therapeutics against plaque.

Enterovirus D68 (EV-D68) is a significant global pathogen associated with severe respiratory infections and acute flaccid myelitis in children. Currently, there are no vaccines or antiviral drugs available for EV-D68, and a robust model to elucidate the pathogenesis of EV-D68 and evaluate treatment methods is lacking. We developed a mouse-adapted EV-D68 strain that caused progressive limb paralysis after intramuscular inoculation in 7-day-old mice. Viral load analysis showed that the skeletal muscle and spinal cord had the highest titers and most severe injuries. RNA sequencing of the infected muscle, brain, spinal cord, and lung tissues revealed differentially expressed genes (DEGs) associated with viral infection and pathogenesis. DEGs were significantly enriched in various pathways associated with antiviral immunity, interferon responses, and cytokine signaling. In the spinal cord, DEGs highlighted mitochondrial dysfunction and oxidative stress as crucial contributors to neural damage. Flow cytometry analysis of spinal cord cells showed that EV-D68 activates the immune system, leading to systemic inflammation and significant increases in CD8⁺ and CD4⁺ T cells, but limited neutrophil and monocyte infiltration. This mouse model provides a valuable tool for studying EV-D68 pathogenesis and evaluating antiviral and vaccine efficacy, thereby advancing the understanding of its neuropathological mechanisms. Importance: We developed a novel mouse model of EV-D68 that provides a valuable tool for studying its pathogenesis and evaluating antiviral and vaccine efficacy, deepening the understanding of its neuropathological basis.

Coxsackievirus A10 (CVA10) is an enterovirus that has caused global outbreaks of hand, foot, and mouth disease (HFMD), accompanied by neurological and systemic complications. Animal models that can simulate natural infections are necessary for studying viral pathogenesis. In this study, our objective was to establish a mouse-adapted strain (CVA10-P8) through serial passaging, which was capable of orally infecting 14-day-old ICR mice, leading to hind-limb paralysis and death. Viral titers in various tissues indicated tropism in muscle tissue, with significant increases observed in the brain, lung, and intestinal tissues as the infection progressed. Pathological examination revealed tissue damage in the muscles, brain, lungs, and intestines accompanied by neutrophil infiltration of the brain. Furthermore, flow cytometry and transcriptome analysis revealed metabolic abnormalities, immune system activation, and the promotion of systemic inflammatory responses in the brain, muscle, and lung tissues. In summary, we successfully developed a CVA10 mouse-adapted strain and a corresponding mouse model, providing valuable tools for studying CVA10 pathogenesis and evaluating the efficacy of antiviral interventions. IMPORTANCE: CVA10 has emerged as a predominant pathogen in the etiology of HFMD, with the potential to elicit neurological manifestations and systemic complications. In this study, we successfully established a novel murine model of CVA10 infection by serially propagating a clinical isolate of CVA10, which enabled oral infection in 14-day-old ICR mice. This model facilitated the investigation of the pathogenesis of CVA10-induced disease. Utilizing this infection model, we employed flow cytometry and transcriptome analysis to elucidate the central nervous system (CNS) inflammatory responses elicited by CVA10 in mice, which closely mimic the natural route of infection. Our findings provide novel insights into the pathophysiological mechanisms underlying CVA10-induced neuroinflammation and pave the way for further research into targeted therapeutic interventions for HFMD associated with CVA10.

Enteroviruses (EVs and RVs) are prevalent worldwide and cause various diseases in humans, of which the VP1-pocket is a target of antivirals, with a lipid molecule as a pocket factor to stabilize the virion. However, the characterization of the structure of the VP1-pocket in EVs is poor. Here, we compared the published capsid crystals of EVs and RVs and proposed a structural framework for the VP1-pocket: Frame 1–4, which is located at the CD loop, GH loop, and C-terminus, presenting with an outward opening appearance or not. The non-outward viral strains—CVB3, Echo 11, RV-A81, and RV-B70—are more thermally stable, with a breakpoint temperature (B.T.) of 51~62 °C for genome releasing, which is 4~10 °C higher than its outward temperature of 41~47 °C, and infectivity preservation when treated at 50 °C for 3 min. Its outward versus non-outward opening is correlated significantly with the B.T. for genome release (r = −0.90; p = 0.0004) and infectivity (r = −0.82, p = 0.0039). The energy of Frames 1, 2, and 4, including Van der Waals attractive and repulsive interactions and hydrogen bonds, showed significant correlations with the B.T. (r = −0.67, 0.75, and −0.8; p = 0.034, 0.013, and 0.006, respectively). These characters of the VP1-pocket could be predictors for virion thermostability and aid in the development of vaccines or antivirals.