Journal of Virology最新文献

The outbreak of highly pathogenic avian H5 influenza (HPAI) clade 2.3.4.4b in cattle has spread across the United States. Mice with pre-existing immunity to H1N1 virus or with a live-attenuated influenza vaccine showed protection against a lethal bovine-derived HPAI H5N1 viral challenge. Notably, ferrets with mixed immunity also demonstrated protection against a feline-derived H5N1 virus, independent of cross-reactive neutralization titers, but antibodies to whole virus were observed. To investigate protective factors, we conducted T cell epitope mapping using published H1N1 viral sequences and found high conservation of key T cell epitopes in the bovine HPAI H5N1 strain. Depletion of T cells in mice prior to and during primary H1N1 infection impacted cross-protective antibodies to H5N1 virus, with CD4 depletion increasing mortality and CD8 depletion mildly impacting morbidity upon H5N1 viral challenge. This underscores the need to investigate memory T cell responses alongside antibodies in assessing preexisting cross-protection to HPAI H5N1 viruses.IMPORTANCEThe rapid spread of highly pathogenic avian H5 influenza (HPAI) clade 2.3.4.4b in U.S. cattle represents an urgent and evolving public health threat. Our findings reveal that pre-existing immunity, whether from seasonal H1N1 infection or live-attenuated vaccination, can confer substantial protection against lethal bovine- and feline-derived HPAI H5N1 viruses, even in the absence of strong cross-neutralizing antibody titers. By integrating T cell epitope mapping with mechanistic depletion studies, we demonstrate that conserved CD4 and CD8 T cell epitopes across H1N1 and H5N1 strains underpin this cross-protection. Critically, loss of CD4 T cell help during primary H1N1 infection disrupts the development of cross-reactive antibody responses and markedly worsens outcomes after H5N1 challenge. These results identify memory T cell responses as important determinants of heterosubtypic immunity and highlight the need to incorporate T cell-focused metrics into risk assessment, vaccine evaluation, and preparedness strategies for emerging HPAI H5N1 viruses.

Novel approaches to sensitize latently infected cells to apoptosis may provide additional methods to eliminate latent reservoirs. Prior research identified several retinoids as potential drugs that increase the sensitivity of HIV-infected cells to cell death. Retinoids are derivatives of vitamin A that target retinoid receptors causing antiproliferative and proapoptotic activity. Several are FDA-approved or in clinical trials. The aim of this study was to evaluate the ability of vitamin A, three of its natural metabolites, and nine synthetic derivatives to sensitize HIV-infected CD4 T cells to NK natural cytotoxicity and antibody-dependent cellular cytotoxicity (ADCC). From the retinoids tested, alitretinoin, tazarotene acid, and AM80 significantly enhanced NK natural cytotoxicity in the presence of IL-15. Mechanistically, these retinoids increased NK degranulation upon target recognition in an HLA-F/KIR3DS1-dependent manner. Furthermore, these retinoids enhanced ADCC by transcriptionally increasing CD16 expression on NK cells. In conclusion, our study has identified at least three retinoids capable of enhancing NK natural cytotoxicity and ADCC against HIV-infected cells. These or other retinoids could be used to reduce HIV persistent reservoirs.IMPORTANCEThis study highlights how retinoids, compounds derived from vitamin A, can help the immune system target HIV-infected cells more effectively. HIV often hides in immune cells, making it difficult to fully eliminate the virus. We found that certain retinoids, including alitretinoin, tazarotene acid, and AM80, improve the function of natural killer (NK) cells-key immune cells that target infected cells. These retinoids boost NK cell activity by increasing their ability to release toxic molecules that kill infected cells and by enhancing their response to antibodies targeting HIV. This makes the infected cells more vulnerable to being eliminated. Since some of these retinoids are already approved for medical use, they could offer a promising way to reduce persistent HIV reservoirs in the body and improve efforts to cure the infection.

Epstein-Barr virus (EBV) infects more than 90% of adults worldwide and causes a range of diseases, including multiple malignancies and autoimmune disorders. However, due to a host range restriction, EBV cannot infect commonly used experimental animals, posing a significant obstacle to developing EBV-specific prophylactic and therapeutic agents. Rhesus lymphocryptovirus (rhLCV), an ortholog of EBV, naturally infects rhesus macaques, which is a surrogate model for EBV research. In this study, we demonstrate that cynomolgus macaque (Macaca fascicularis), a primate closely related to rhesus macaque, is susceptible to rhLCV infection. rhLCV can immortalize B cells of cynomolgus macaques to develop cy-LCLs. We developed a high rhLCV-producing cy-LCL cell line, LCL111, and optimized the induction conditions to increase viral production, surpassing the original rhLCV producer LCL8664. Importantly, EBV gHgL-specific monoclonal antibody (mAb) AMMO1 and gB-specific mAb 3A5 can cross-react with rhLCV proteins and block the formation of cy-LCLs. Overall, we established an efficient rhLCV-producing cell line, and rhLCV infection of cynomolgus macaques represents a promising alternative surrogate model for efficiency evaluation of EBV vaccines and mAbs.

Importance: Epstein-Barr virus (EBV) naturally infects only humans, creating a major barrier to evaluating the efficiency of vaccines and therapies in vivo. As an EBV ortholog, rhesus lymphocryptovirus (rhLCV) offers a biologically relevant surrogate system. However, its application has been primarily limited to rhesus macaques. Here, we demonstrate that cynomolgus macaque lymphocytes are also susceptible to rhLCV in vitro, and the newly transformed cy-LCL111 shows superior and sustained rhLCV production ability. rhLCV infection of cynomolgus macaque lymphocytes can be efficiently neutralized by anti-EBV gH/gL nAbs AMMO1 and anti-EBV gB mAbs 3A5, highlighting the potential of cynomolgus macaques as an in vivo model to assess anti-EBV mAb and vaccine efficacy. Our findings support the use of cynomolgus macaques as an additional model for EBV research and offer a useful platform for evaluating EBV-specific prophylactic or therapeutic strategies.

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.

N6-methyladenosine (m⁶A) is the most abundant internal modification in eukaryotic RNA and plays diverse roles in RNA metabolism. Increasing evidence indicates that m⁶A is also present in viral RNAs, where it exerts virus-specific effects. While several studies have shown that m⁶A can facilitate viral replication, its antiviral mechanisms remain less understood. In this study, we used transmissible gastroenteritis virus (TGEV) as a model to investigate the inhibitory role of m⁶A in viral infection. We demonstrated that m⁶A modification is present in the TGEV genome and suppresses viral replication. The m⁶A reader proteins bind to viral RNA and reduce the stability of m⁶A-modified transcripts. Notably, TGEV infection increased global m⁶A levels in host RNA, particularly in interferon (IFN)-associated genes. Inhibition of m⁶A methylation significantly diminished IFN gene expression. Furthermore, compared to other viruses, TGEV genomic RNA displayed an abnormally higher m⁶A ratio, which can be distinguished by RIG-I to promote an immune response. Collectively, our findings reveal that high m⁶A modification enhances RIG-I-mediated sensing of TGEV RNA, leading to the activation of IFN responses and inhibition of viral replication. This study provides new insights into the complex regulatory functions of m⁶A during viral infection and host antiviral defense.IMPORTANCEN6-methyladenosine (m⁶A) is one of the most prevalent RNA modifications in viral genomes, but its functional impact varies widely across viruses. While m⁶A often promotes viral replication, it can exert inhibitory effects in certain viruses, particularly within the Flaviviridae and Coronaviridae families. Despite growing evidence of this antiviral role, the underlying mechanisms remain largely unclear. Here, we used transmissible gastroenteritis virus (TGEV), a swine coronavirus, as a model to explore the inhibitory function of m⁶A. We show that the TGEV genome harbors a relatively high density of m⁶A modification compared to other viruses and host mRNA, which are efficiently detected by the host pattern recognition receptor RIG-I. This interaction enhances innate immune activation and restricts viral replication. Our findings uncover the mechanism by which abnormal m⁶A modification can be sensed to activate antiviral immunity and provide deeper insight into the multifaceted role of m⁶A in host-virus interactions.

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.

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.

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.

Human papillomavirus (HPV) and Merkel cell polyomavirus (MCPyV) are DNA tumor viruses that cause human cancer. The mechanisms by which HPV and MCPyV oncoproteins induce genomic instability are not well defined. This minireview discusses the influence of these oncoproteins on the repertoire of proteins at replicating DNA, known as the host replisome, and discusses how new technologies like isolation of proteins on nascent DNA (iPOND) can drive the discovery of viral dysregulation of the host replisome to enhance our understanding of viral oncogenesis.

Autographa californica multiple nucleopolyhedrovirus (AcMNPV) E18 (AC143, ODV-E18) is an envelope protein common to both occlusion-derived virions (ODVs) and budded virions (BVs). The e18 gene has been demonstrated to be essential for generating infectious BVs. However, its functional role in virion morphogenesis remains unclear. In this study, we constructed an e18 knockout virus and an e18 repair virus to investigate the effects of e18 deletion on virion morphogenesis. Our data indicated that e18 is required for normal intranuclear microvesicle (IMV) formation and accumulation, for intranuclear envelopment and nuclear egress of nucleocapsids, as well as for embedding of ODVs into occlusion bodies (OBs) and BV production. Additionally, we created and characterized a series of recombinant viruses with truncated e18 of varying lengths to identify domains involved in nuclear translocation and virion morphogenesis. We identified two low-complexity domains (LCDs) in E18, in addition to a known transmembrane domain (TM). The AA30-34 sequence within the TM was found to be essential, but not sufficient for nuclear translocation. However, an α-helix structure encompassing the TM domain proved adequate to mediate a fusion protein's trafficking into the nucleus in the context of additional viral factors. Furthermore, we discovered that the TM was required for the accumulation of IMVs, while both the TM and LCD 1 were necessary for intranuclear envelopment, nuclear egress of nucleocapsids, and the embedding of ODVs into OBs; LCD 2 influenced the processing of IMVs and ODV formation. Both the TM and the two LCDs were essential for BV production.IMPORTANCEThe envelope protein E18 is a conserved component common to both ODV and BV virion types of baculoviruses, yet its functional role in virion morphogenesis remains unclear. This study investigated the e18 gene of Autographa californica multiple nucleopolyhedrovirus, determining that it is essential for normal IMV formation and accumulation, intranuclear envelopment and nuclear egress of nucleocapsids, as well as for the embedding of ODVs into occlusion bodies and BV production. The functional roles of the single TM domain and two LCD domains within E18 during virion morphogenesis were identified. Furthermore, it was found that an α-helix structure encompassing the TM domain is sufficient to facilitate the trafficking of a fusion protein into the nucleus in the context of other viral factors, with AA30-34 being critical for the nuclear import of E18.