Hao Yang

An AG129 mouse infection model with four DENV isolates was established to systematically characterize the early pathogenic phenotypes of these strains. DENV-1 caused 100% mortality by 15 dpi, with sustained viremia, widespread multi-organ injury, and persistent leukopenia. DENV-4 exhibited peak viremia at 4 dpi and unique neurovirulence, presenting exclusive hind-limb paralysis, severe pulmonary hemorrhage, and characteristic cerebral parenchymal lesions. DENV-2 and DENV-3 induced only transient mild viremia and focal tissue alterations without mortality. Hematological and biochemical alterations, as well as occasional splenic hemosiderin deposition, were noted among infected animals. This preclinical dataset offers insights into serotype-specific DENV pathogenesis and may aid future evaluation of broad-spectrum antivirals and tetravalent vaccine candidates.

Background/Objectives: Recurrent influenza epidemics impose a severe global burden, with conventional vaccines constrained by production time lags and rapid viral mutation. This study aims to explore a novel influenza mRNA vaccine design that balances conserved and mutable antigen regions. By combining hemagglutinin (HA) and neuraminidase (NA) into a dual-target approach, the objective is to simultaneously block viral entry and inhibit progeny release, potentially establishing a proposed “front-blockade, rear-containment” dual protective barrier against multiple H1N1 strains. Methods: We engineered a dual-target tandem mRNA vaccine linking mutated HA with conserved NA, with strategic amino acid mutations introduced into key antigenic sites within the HA head domain. Vaccine efficacy was evaluated in a mouse model. Humoral immunity was assessed by measuring antigen-specific antibody titers, and cellular immunity was evaluated via ELISpot assay. Protective capacity was determined through lethal challenge experiments using diverse H1N1 viral strains. Results: The vaccine successfully expressed the HA-NA tandem antigen at 130 kDa, and the in vitro-expressed antigen exhibited normal neuraminidase activity. Preliminary evidence supported the dual-target concept in model mice: hemagglutination-inhibiting and micro-neutralizing antibodies targeting HA were detected, and serum neuraminidase-inhibiting activity was also observed. In addition to triggering potent cellular immune responses, the vaccine offered total protection against lethal doses of various H1N1 variants. Conclusions: This study suggests a promising dual-target strategy that harmonizes antigen conservation and mutation while potentially establishing a synergistic front-blockade and rear-containment defense. The approach offers a viable pathway for developing improved H1N1 influenza vaccines.

Chikungunya virus (CHIKV) pathogenesis research has long been constrained by the lack of suitable immunocompetent rodent models. Through serial passaging in A129 and C57BL/6 mice, we obtained an adapted strain (CHIKV-Adapt) harboring an E2-K200R substitution along with non-structural protein mutations. Phenotypic analysis in C57BL/6 mice, BALB/c mice, and hamster models demonstrated that compared to the wild-type virus CHIKV-Adapt induced significantly higher and more prolonged viremia, broader tissue tropism, and more severe internal joint inflammation, without exacerbating external swelling. Notably, the K200R mutation did not alter the viral replication kinetics in vitro and was predicted not to affect its binding pattern to the MXRA8 receptor. Furthermore, mice challenged 160 days after primary infection exhibited nearly complete protective immunity. These findings indicate that E2-K200R is a critical adaptive mutation that, together with accompanying non-structural mutations, significantly enhances CHIKV replication capacity and pathogenicity in immunocompetent rodents without changing its in vitro replication ability or predicted receptor-binding mode. The acquisition of this adapted strain provides a new tool for CHIKV pathogenesis research and vaccine evaluation.

Chikungunya virus (CHIKV) is now prevalent in multiple regions worldwide, posing a serious threat to human health. In this study, we have successfully established a rhesus macaque model of Chikungunya virus infection. Notably, while no viral load was detected in liver tissue on day 7 post-infection, significant pathological damage was observed. Single-cell RNA sequencing of liver tissue revealed a reduction in B cells following infection. Among T cells, CD8⁺ T and NKT cells mediated major cytotoxic effects, whereas CD4⁺ T and memory T cells primarily exerted regulatory functions that further enhanced the activation of CD8⁺ T and NKT cells. In macrophages, inflammatory macrophages fc gamma R-mediated phagocytosis upregulated, with multiple key activation-related genes being highly upregulated. Furthermore, we observed that there might be a potential bidirectional activation effect between T cells and macrophages. These results indicate that CHIKV-induced indirect liver injury is likely mediated not only by the virus itself but also, in part, by the activation of hepatic immune cells.

The multiple epidemics of Zika virus (ZIKV) posed a substantial threat to public health. Clinical evidence suggests that ZIKV could break through the blood-brain, blood-placenta, and blood-testis barriers, leading to severe outcomes such as congenital malformations in newborns and Guillain-Barré syndrome in adults. Currently, there are no specific treatments for ZIKV infection. To address the antibody-dependent enhancement (ADE) of dengue virus (DENV) infection induced by ZIKV vaccination, we designed two modified prM-E RNAs (ZA and ZB) with specific mutations either shielding or disrupting the conserved fusion-loop epitope in the E protein. Then, we chose the mRNA-LNP vaccine platform to evaluate the safety and efficacy. After prime-boost immunization, ZA vaccine could induce high levels of T cells secreting IFN-γ and exhibit limited neutralizing ability against Asian-lineage and African-lineage ZIKV. After ZIKV challenge, ZA vaccine could provide complete protection in immunocompromised AG129 mice at low levels of neutralizing antibodies, preventing viral dissemination to the brain, uterus, and testes. Importantly, the ZA vaccine also reduced the ADE effect of DENV infection. Although ZB vaccine exhibited good immunogenicity, it could not achieve complete viral clearance in AG29 mice. Our findings suggested that the ZA vaccine could prevent both lethal ZIKV infection and DENV ADE induced by infection or vaccination.

Clinical evidence has associated H1N1 influenza with liver impairment, yet the underlying mechanisms remain poorly understood. Here, we investigated H1N1-induced liver damage and its potential mechanisms using a BALB/c mouse infection model. Pathological examination and serum aspartate transaminase (AST) and alanine transaminase (ALT) were assessed. Messenger ribonucleic acid-sequence was used to analyze the transcriptomic changes in tissues. Multiple inflammatory cytokines in tissues and inflammatory cells in the blood were detected on the fifth day post-infection. Our results showed that H1N1 infection caused significant liver pathology and elevated serum AST/ALT levels. Transcriptomic analysis revealed significant alterations in liver gene expression profiles following H1N1 infection, particularly in genes associated with inflammatory responses, including those involved in monocyte adhesion/activation and neutrophil/macrophage infiltration. Marked increases in inflammatory mediators were observed in lungs, serum, and liver, accompanied by systemic changes in circulating inflammatory cells, indicating H1N1 triggered a robust systemic inflammatory response. These findings suggest that H1N1-induced liver damage may be associated with the systemic inflammatory response induced by H1N1 and changes in liver gene regulation.