Yanan Zhou

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: Nipah virus (NiV) is a highly pathogenic zoonotic agent that causes fatal respiratory and neurological diseases, for which no approved vaccines are currently available. To address this unmet need, we developed and evaluated novel mRNA-LNP vaccine candidates. These vaccines employ an epitope- and structure–guided “immune-focusing” strategy, aiming to maximize the exposure of critical neutralizing epitopes on the viral attachment (G) and fusion (F) glycoproteins. Methods: Head-to-head dimeric antigens (2Gs and 2Fs) were engineered by removing subdominant stalk regions and tandemly duplicating epitope-rich globular head domains. Three LNP-encapsulated candidates-NV1 (encoding 2Gs), NV2 (encoding 2Fs), and NV3 (a 1:1 mixture of NV1 and NV2)-were evaluated in BALB/c mice and Syrian hamsters to systematically assess humoral responses, cytokine secretion, BCR/TCR repertoires, and systemic safety. Results: NV1 and NV3 induced potent binding antibodies and strong neutralizing activity against NiV pseudoviruses. Meanwhile, although NV2 lacked neutralizing activity, it induced superior Th1-biased cellular immunity, accompanied by extensive T-cell clonal proliferation. BCR/TCR sequencing revealed unique adaptive immune characteristics that perfectly align with the distinct immunogenic properties described above. Preliminary safety assessments found no toxicity to vital organs, while active germinal center formation confirmed the success of immune mobilization. Conclusions: An epitope-centered dimer design effectively shapes the unique defense mechanisms of the adaptive immune system. NV3 offers a balanced and synergistic strategy that combines potent humoral and cellular immune defenses, providing a highly promising platform for the development of a Nipah virus (NiV) vaccine.

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.

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.

Soluble host factors in the upper respiratory tract can serve as the first line of defense against SARS-CoV-2 infection. In this study, we described the identification and function of a human airway trypsin–like protease (HAT), capable of reducing the infectivity of ancestral SARS-CoV-2. Further, in mouse models, HAT analogue expression was upregulated by SARS-CoV-2 infection. The antiviral activity of HAT functioned through the cleavage of the SARS-CoV-2 spike glycoprotein at R682. This cleavage resulted in inhibition of the attachment of ancestral spike proteins to host cells, which inhibited the cell-cell membrane fusion process. Importantly, exogenous addition of HAT notably reduced the infectivity of ancestral SARS-CoV-2 in vivo. However, HAT was ineffective against the Delta variant and most circulating Omicron variants, including the BQ.1.1 and XBB.1.5 subvariants. We demonstrate that the P681R mutation in Delta and P681H mutation in the Omicron variants, adjacent to the R682 cleavage site, contributed to HAT resistance. Our study reports what we believe to be a novel soluble defense factor against SARS-CoV-2 and resistance of its actions in the Delta and Omicron variants.