Journal of Virology最新文献

Given that ducks serve as critical reservoirs for avian influenza viruses, achieving high immune coverage in duck flocks is essential for preventing the transmission of avian influenza viruses from wild birds to domestic poultry. Duck enteritis is the most important infectious disease that must be prevented with a live-attenuated vaccine in duck breeding. Therefore, we previously constructed a recombinant duck enteritis virus (DEV), rDEV-dH5/H7, which carries the hemagglutinin (HA) genes of two H5 viruses and an H7 influenza virus. It could induce rapid and complete protection against both lethal DEV and H5 and H7 viruses as early as 7 days post-prime vaccination, although almost no antibodies were detected in ducks at this time. In the present study, we demonstrated that rDEV-dH5/H7 immediately initiated innate immune responses and T-cell proliferation in ducks. The expression levels of IFN-γ and granzyme A were markedly upregulated at 5-7 days post-prime vaccination. The percentages of CD3⁺ and CD3⁺CD8⁺ T cells in peripheral blood mononuclear cells significantly increased from 7 days post-prime vaccination. Moreover, HA-specific CD8⁺ and CD4⁺ T cells, as well as those induced by DEV virion, were significantly higher than those in control animals from 7 days post-prime vaccination. Together, the quick response of specific T cells and the innate immune response induced by rDEV-dH5/H7 may confer rapid immune protection against lethal influenza virus and DEV.IMPORTANCEPreviously, we found that recombinant duck enteritis virus, rDEV-dH5/H7, could induce rapid and robust dual protection against lethal DEV and highly pathogenic avian influenza viruses as early as day 7 post-prime vaccination, although almost no antibodies could be detected at this time. In the present study, we reveal that cell-mediated immunity plays a critical role through early upregulation of IFN-γ/granzyme A pathways and robust hemagglutinin (HA)-specific T-cell responses and drives protection even in the absence of detectable antibodies. This is the first mechanistic evidence showing DEV-vectored vaccines activate the robust proliferation of T lymphocytes and HA-specific T-cell responses. Our findings fundamentally advance the understanding of DEV-vectored vaccines, offering new insight for recombinant DEV vaccine design.

Human noroviruses (HuNoVs) are the leading cause of viral gastroenteritis, with ≥80% of infections caused by the GII genogroup. HuNoVs are non-enveloped, with an icosahedral capsid composed of 90 dimers of the major capsid protein VP1, which encloses a minor structural protein, VP2, and a VPg-linked, positive-sense ssRNA genome. Although the atomic structure of the icosahedral capsid formed by VP1 is well characterized using crystallography and cryo-electron microscopy analyses of HuNoV virus-like particles, the structures and the localization of VP2 and VPg inside the capsid, how they are incorporated into the capsid, and whether this process requires interactions between them remain unresolved. Herein, we show that VP2 is the molecular bridge for assembly of particles containing VP1, VP2, and VPg. We used deletion constructs and mutational analyses in transfected HEK293FT cells, guided by bioinformatics, to determine the interaction site on VP2 for VP1 of the pandemic-causing GII.4 Sydney HuNoV. GII.4 HuNoV VP2 contains a unique insertion site at amino acids (AAs) 43-53, relative to VP2s of other GII HuNoV genotypes. We identified that AAs 40-43 on VP2 are required for interaction with VP1, and mutation of VP2 AAs 40-43 abrogates VP2-VP1 interactions. Computational analyses predicted that VP2 has a highly conserved N-terminal α-helical domain and an intrinsically disordered C-terminal domain that exhibits significant sequence diversity. We identified that VP2, not VP1, pulls down VPg; the VP2 C-terminal domain is sufficient to interact with VPg. These findings reveal domain-specific functions of VP2 that are essential for coordinating capsid protein interactions for HuNoV assembly.

Importance: Human noroviruses (HuNoVs) are the leading cause of epidemic and sporadic gastroenteritis in all age groups worldwide. Yet, we lack vaccines or therapeutics for HuNoVs. Knowledge of the fundamental mechanisms governing HuNoV particle assembly is limited. Modern structural techniques have not resolved the complete structure of pandemic GII.4 norovirus or the localization of interior capsid proteins VP2 and VPg. Furthermore, VP2's functional roles during infection remain obscure. Studies of feline and murine caliciviruses show that VP2 may help deliver the viral genome into cells, suggesting that VP2 synergizes with VP1 and VPg. We identified a motif within the N-terminal α-helical domain of VP2, adjacent to a unique insertion site, that is essential for interaction with the major capsid protein VP1. We show VP2 uniquely interacts with the translation initiation protein VPg via its disordered C-terminus. These findings reveal principles of HuNoV capsid protein interactions and highlight VP2 as a bridge facilitating capsid assembly.

Porcine reproductive and respiratory syndrome virus (PRRSV), a major immunosuppressive pathogen, inflicts substantial economic losses on the global swine industry. Despite extensive research into PRRSV pathogenesis, the mechanisms by which PRRSV induces immune dysfunction in vivo remain incompletely understood. Here, we performed single-cell RNA sequencing on cells isolated from the lung of PRRSV-infected piglets, generating transcriptomic profiles for 46,922 single cells encompassing 15 major cell types. We observed a significant reduction in the number of macrophages in lung tissues, which was primarily attributed to the extensive apoptosis of macrophages induced by PRRSV infection. PRRSV infection triggered aberrant differentiation of macrophage, and the SPP1^high macrophage subpopulation was identified as the primary target cells for PRRSV infection. Cell-cell communication analysis revealed that PRRSV infection enhanced ligand-receptor interactions between macrophages and other cell types, associated with inflammatory responses, activation of T cells and B cells, and cell adhesion. In addition, monocytes exhibited a tendency to differentiate into macrophages, potentially compensating for the depletion of macrophages caused by PRRSV infection. Moreover, PRRSV infection caused abnormal development of B cells and incomplete activation of cytotoxic T lymphocytes in the lungs. This study provides a comprehensive characterization of how PRRSV perturbs pulmonary immune cell populations, offering valuable insights into the mechanisms underlying PRRSV-induced lung injury.IMPORTANCEPorcine reproductive and respiratory syndrome virus (PRRSV) has consistently posed a significant and enduring threat to the swine industry. However, the virus-host interactions during in vivo infection in vivo remain poorly understood. In this study, we applied single-cell RNA sequencing to characterize the cellular heterogeneity of lung tissues from PRRSV-infected piglets. Through intracellular viral RNA tracking, we identified SPP1^high macrophages as the primary reservoir of PRRSV. Furthermore, we analyzed the cell-cell communication between macrophages and other cell types and investigated the immune responses and heterogeneity of monocytes, T cells, and B cells upon PRRSV infection. Our findings provide a comprehensive single-cell landscape of the complex host-pathogen interplay during PRRSV infection.

Enterovirus D68 (EV-D68) is a globally reemerging respiratory pathogen of notable clinical concern due to its association with severe respiratory disease and the paralytic complication acute flaccid myelitis (AFM). Viral tropism and pathogenesis are critically dictated by interactions with host cell receptors. Our understanding of this process has evolved from a simple model of sialic acid dependence to a dynamic paradigm involving a repertoire of attachment factors and proteinaceous entry receptors. This review synthesizes the evolving landscape of EV-D68 receptor usage. We detail the well-established role of α2,6-linked sialic acid as an attachment factor and uncoating trigger for historical strains. We further discuss the discovery of intracellular adhesion molecule-5 (ICAM-5) as a neuron-specific receptor that provides a molecular explanation for neurotropism in AFM. A pivotal recent advance is the identification of major facilitator superfamily domain-containing 6 (MFSD6) as an essential entry receptor for a broad range of EV-D68 strains in both respiratory and neuronal cells. We explore the implications of this receptor versatility, whereby the virus can switch between or co-opt sialic acid, ICAM-5, and MFSD6, a plasticity that influences tissue tropism and viral evolution. Finally, we highlight how these mechanistic insights, particularly the characterization of the MFSD6 interface, are paving the way for novel therapeutic strategies, such as engineered decoy receptors, and outline key future directions in the field.

HIV-1 plasma viral load decays in a biphasic manner during antiretroviral therapy (ART). It was hypothesized that this is due to infection of different cell types, namely CD4+ T cells and macrophages. We studied this possibility directly by modeling the decay of HIV-1 in humanized mice. We utilized previously published data from humanized T-cell only mice (TOM) and myeloid-only mice (MOM) infected with HIV-1 and treated with a potent ART regimen. Viral load decay dynamics were modeled using either a single or a biexponential decay fitted using nonlinear mixed effects techniques. Fits were compared using the corrected Bayesian information criterion (BICc). In TOM, the biphasic model was significantly better than a single-phase decay model (ΔBICc ≈ 16) despite additional parameters. In MOM, the biphasic decay was statistically better, but there was substantial uncertainty because the virus goes below detection very fast. The first-phase half-life was consistent between groups (1.2 days in MOM and 1.3 days in TOM) and similar to the half-life estimated in human infection. The second-phase decay in these mice was minimal likely due to low initial viral loads. Additional analyses with mice containing both CD4+ T cells and macrophages or X4-tropic virus-infected MOM mice confirmed the biphasic pattern, demonstrating the robustness of this result. The biphasic decline in HIV-1 occurs, even with only CD4+ T cells, refuting the hypothesis that distinct cell populations (CD4+ T cells and macrophages) drive each decay phase. These findings support an alternative model in which the observed dynamics arise from intrinsic properties of the viral infection lifecycle rather than from cellular compartmentalization.IMPORTANCEIt is well known that when antiretroviral therapy is started in people infected with HIV, the decay of virus in the periphery is biphasic early on (followed by other slower phases). One possibility for this pattern of decay is infection of two different types of cells (suggested previously to be CD4+ T cells and macrophages), with different turnovers giving rise to the biphasic decline. We addressed this issue directly in a humanized mouse model of HIV, taking advantage of mice reconstituted with just T cells and treated with antiretroviral drugs. We found that the observed decay is biphasic, which eliminates the hypothesis that the biphasic decline is due to the co-existence of the two types of cells. It is possible that integration dynamics, as we previously proposed, are responsible for the observed biphasic decline.

Duck plague virus (DPV) is a highly pathogenic avian herpesvirus that affects ducks, geese, and other anseriform poultry. The primary pathological changes observed in infected animals are mucosal, serosal, and systemic hemorrhages accompanied by exceptionally high fatality rates. While most DPV genes are conserved among herpesviruses, a small subset of genes, including the SORF3 gene, is unique to avian herpesviruses. To date, reports on the function and characteristics of the SORF3 gene are limited. In this study, the use of a polyclonal antibody against SORF3 demonstrated that this open reading frame could encode proteins. Through the use of DNA and protein synthesis inhibitors in infected cells, we delineated the gene's transcriptional and translational timeline, establishing SORF3 as a late gene. To further investigate the role of the protein encoded by the SORF3 gene in the pathogenic mechanism, we constructed a virus lacking the SORF3 gene. The growth kinetics results indicated that the SORF3 protein is not essential for viral replication. In vivo experimental findings revealed that while the SORF3-deleted virus still induced clinical symptoms and pathological changes associated with duck plague upon infection of ducks, its lethality was lower than that of the parental virus. In conclusion, this study revealed that the SORF3 gene, which is specific to avian herpesviruses, encodes a late viral protein in DPV and explored its potential role in DPV pathogenesis.IMPORTANCEDuck plague virus (DPV) has a high incidence rate and mortality rate of up to 90%, resulting in substantial economic losses in poultry farming. Consequently, investigating the temporal transcription and functional characterization of the proteins encoded by each DPV gene is crucial for understanding its complex life cycle and pathogenesis. This study revealed that the SORF3 gene, identified as an avian herpesvirus-specific gene, encodes a protein. Furthermore, the temporal transcription of this gene throughout the virus's life cycle confirmed that the protein encoded by SORF3 significantly influences the pathogenicity of DPV.

Class II major histocompatibility complexes (MHC-II) are a highly polymorphic and multigenic family of molecules that present exogenous peptides to CD4⁺ helper T cells, thereby activating the host adaptive immune system. In this study, we systematically analyzed the genomic distribution and tissue-specific expression of MHC-II genes in Carassius gibelio. The Cagi-DDA/DFA molecule was found to be highly expressed in the spleen and head kidney, moderately expressed in the intestine, gills, and trunk kidney, and expressed at low levels in the liver and brain. A polyclonal antibody was generated against the most prevalent Cagi-DDA/DFA allele in the population. Using immunopeptidomics, we identified viral peptides bound to Cagi-DDA/DFA molecules in the head kidney tissues of C. gibelio following Cyprinid herpesvirus 2 (CyHV-2) infection. A total of 276 antigen peptides were identified, originating from 39 viral proteins. Notably, viral proteins with high abundance and early expression profiles, such as ORF88, ORF121, and ORF141 proteins, were more likely to generate antigen peptides. The identified CyHV-2 peptide epitopes presented by C. gibelio MHC-II molecules provide candidate antigens required for anti-CyHV-2 vaccine development.IMPORTANCEVaccination represents a cornerstone in the prevention of infectious diseases, achieving substantial success in disease control. Upon immunization, protein-derived peptides are processed and presented by major histocompatibility complex class II (MHC-II) molecules, activating CD4⁺ T cells and triggering adaptive immune responses. Cyprinid herpesvirus 2 (CyHV-2), a pathogenic virus in crucian carp, poses a serious threat to global aquaculture. However, the absence of a comprehensive antigenic profile for CyHV-2 has hindered the development of effective vaccines. Here, we employed immunoaffinity purification coupled with mass spectrometry to systematically identify CyHV-2-derived peptides presented by MHC-II in Carassius gibelio. We identified 276 antigenic peptides originating from 39 viral proteins, which collectively delineate the antigenic landscape of CyHV-2 and provide a rational basis for the design of a vaccine against CyHV-2.

Secondary bacterial infections can significantly worsen the clinical course of influenza virus infections and are a leading cause of morbidity and mortality during seasonal influenza epidemics. Despite being a vaccine-preventable disease, influenza-related complications from secondary bacterial infections are an important cause of death, particularly among the elderly population. Streptococcus pneumoniae (Spn) is the most common agent responsible for influenza-related secondary bacterial infections. Influenza virus vaccination serves as an effective prophylactic strategy for preventing influenza and reducing the burden of influenza-associated pathology, including secondary bacterial infection. However, whether the protective effects of influenza virus vaccination differ in the context of a secondary Spn infection at the level of the host response remains poorly characterized. Here, we present a preclinical mouse model to examine the impact of influenza vaccination in scenarios involving single infections with influenza A virus H1N1 (NC99) or Spn serotype 1; simultaneous infection with both NC99 and Spn (coinfection), or NC99 infection followed by Spn infection seven days later (superinfection). A single dose of trivalent inactivated Influenza vaccine (TIV) is able to decrease infection lethality in both secondary bacterial infection scenarios. Protection is associated with reduction in both viral and bacterial titers, decreased production of pro-inflammatory cytokines, protection of alveolar macrophages, prevention of exacerbated lung neutrophil recruitment, modulation of neutrophil activation status, and induction of lung eosinophil recruitment and activation. These findings underscore the importance of influenza vaccination in modulating disease progression and preventing morbidity and mortality associated with secondary bacterial infections.

Importance: In this study, we show that a licensed influenza vaccine not only prevents severe disease upon influenza virus infection but also helps protect against enhanced morbidity due to co- or superinfection with Streptococcus pneumoniae in a mouse model. This protection correlates with better control of viral and bacterial titers, as well as with altered host immune responses during bacterial co- and superinfection, characterized by the recruitment of activated granulocytes.

HIV-1 continues to pose a significant global health challenge, requiring ongoing research into effective prevention and treatment strategies. Understanding the B-cell repertoire that can be engaged upon vaccination in humans is crucial for the development of future preventive vaccines. In this study, peripheral blood mononuclear cells from HIV-negative participants in the multivalent HVTN124 human HIV-1 vaccine clinical trial were interrogated for HIV-reactive B cells using LIBRA-seq, a high-throughput B-cell mapping technology. We report the discovery of glycan-reactive antibodies, with one of these being capable of neutralizing diverse heterologous HIV-1 virus strains. Furthermore, isolated antibodies showed broad cross-reactivity against antigens from a variety of other pathogens. The emerging class of glycan-reactive virus-neutralizing antibodies with exceptional breadth of pathogen cross-reactivity may present an effective target for vaccination at the population level.

Importance: Understanding how the human immune system recognizes and combats viruses is crucial for developing better vaccines and treatments. Here, through characterization of the B-cell receptor repertoires of participants in HVTN124, a multivalent HIV-1 vaccine human clinical trial, we discovered antibodies that recognize sugar molecules (glycans) on antigens from a range of unrelated viral families. In addition to their binding breadth, these antibodies can also neutralize multiple diverse strains of HIV-1. Our findings reveal an emerging and underappreciated mechanism for antibodies to counteract virus infection, potentially opening doors for developing vaccines that preferentially elicit glycan-reactive antibody species to broadly protect against different viruses.This study is registered with ClinicalTrials.gov as NCT03409276.