Unlike Ff, the packaging signal (PS) and the mechanism of integrative filamentous phage assembly remains largely unknown. Here we revived two Inoviridae prophage sequences, ϕLf2 and ϕLf-UK, as infectious virions that lysogenize black rot pathogen Xanthomonas campestris pv. campestris. The genomes of ϕLf2 and ϕLf-UK consist of 6363 and 6062 nucleotides each, and share 85.8% and 98.7% identity with ϕLf, respectively. To explore integrative filamentous phage assembly, we first identified 20-26-nucleotide long PS sequences of 10 Xanthomonas phages. These PS consist of a DNA hairpin with the consensus GGX(A/-)CCG(C/T)G sequence in the stem and C/T nucleotides in the loop, both of which are conserved and essential for PS activity. In contrast to Ff, the 5' to 3' orientation of the PS sequence is not conserved or critical for viral competence. This is the first report to offer insights into the structure and function of the integrative phage PS, revealing the diversity of filamentous phage encapsidation.

The adaptive immune response during BCoV infection of peripheral blood mononuclear cells (PBMCs), the bovine spleen cells, and their isolated T lymphocytes was not studied well. Our study confirmed successful BCoV infection in PBMCs and spleen T cells. The BCoV replication was evidenced by measuring genome copy numbers using real-time PCR and expression levels of BCoV spike and nucleocapsid proteins via western blot and immunofluorescence assays. In infected PBMCs, CD4 T-cell levels were 1.45-fold higher, and CD8 T-cell levels were 1.6-fold lower compared to sham-infected cells. Conversely, infected splenocytes showed a 0.88-fold decrease in CD4 T-cells and a 1.88-fold increase in CD8 T-cells. The cytokine gene expression analysis revealed that BCoV infection activated type I interferon and upregulated IL-6 expression in PBMCs and splenocytes. These findings demonstrate that BCoV successfully infects immune cells from PBMCs and spleen, inducing differential host cytokine gene expression that favors virus replication.

Chronic viral infections are characterized by exhausted virus-specific T cells. Exhaustion is associated with mitochondrial dysfunction, revealing a possible target for treatment. Targeting these metabolic processes may interfere with the exhaustion process of immune cells during infection. It has been shown that the mitochondria-targeted antioxidant MitoTempo could restore hepatitis-B-virus-specific T cells in vitro. Thus, we investigated MitoTempo as a treatment option using the chronic lymphocytic choriomeningitis virus (LCMVcl13) mouse model. MitoTempo treatment of chronically LCMVcl13 infected mice resulted in a transient reduction of LCMV titer. However, no obvious restoration of functional LCMV-specific T cells was observed, beside subtle changes in phenotype of GP33- and NP205-specific T cells. However, these changes did not translate into significantly more functional responses. Our study showed a transient antiviral effect of MitoTempo, but no profound effect on exhausted T cell responses, although further studies are needed to further elucidate the mechanism and use of MitoTempo.

Feline immunodeficiency virus (FIV) is a retrovirus of worldwide distribution that can cause an acquired immunodeficiency disease in domestic cats. FIV and the primate lentiviruses, human and simian immunodeficiency viruses (HIV and SIV, respectively) share structural and biological features but also exhibit important differences, which reflect both their evolutionary relationship and divergence. Given that FIV is not only an important cat pathogen but also a useful model for certain aspects of HIV-1 infections in humans, the study of FIV biology is highly relevant. In this review we provide an updated description of the molecular mechanisms involved in each stage of the FIV life cycle.

Arteriviruses in the Nidovirales order are single-stranded positive-sense RNA viruses infecting mammals. Arteriviruses are recognized for causing various clinical diseases, ranging from asymptomatic infections to severe conditions like respiratory syndromes and viral hemorrhagic fever. Notably, arteriviruses exhibit a high frequency of RNA recombination, and their robust recombination rates are a crucial factor in recurrent outbreaks. The recombination events also shape the countermeasures employed by arteriviruses during virus-host co-evolution and confer specific evolutionary benefits to viruses, implicating a role as a selective advantage in viral adaptation. This review delves into the molecular basis of RNA recombination in arteriviruses, the bioinformatics tools and methodologies used to visualize evolutionary relationships, and the identification of recombination breakpoints. Significant recombination events are highlighted for PRRSV and other arteriviruses, illustrating the profound implications of recombination for viral evolution and pathogenesis. Recombination between field viruses and between field viruses and vaccine strains can generate new variants with altered antigenic profiles and virulence, leading to diagnostic failure, severe clinical outcomes, and reduced vaccine efficacy. Despite the advances, further research is needed to understand recombination rates and hotspots, as well as to develop potential antiviral strategies and diagnostic approaches for arteriviruses.

In our previous study, we established a method for purifying bacterially expressed HCV core 1-164 under non-denaturing conditions. In the present study, we elucidated the characteristics of the purified core. The purified HCV core exhibited a notable affinity for HCV RNA, with a Kd of 3 nM, as determined by a filter binding assay. Electron microscopy analysis revealed that the purified HCV core self-assembled with RNA into spherical structures approximately 50 nm in diameter. Additionally, we demonstrated the direct binding of the purified HCV core to the purified endoplasmic reticulum (ER). Moreover, lipid strip assays revealed specific binding of the purified HCV core to specific phospholipids, suggesting that the core plays a role in specific ER membrane domains. These studies reveal the biochemical characteristics of the HCV core that significantly advance our understanding of its role as a capsid protein in the viral lifecycle and pathogenesis.

Virus-like particles (VLPs) resemble the parent virus but lack the viral genome, providing a safe and efficient platform for the analysis of virus assembly and budding as well as the development of vaccines and drugs. During the COVID-19 pandemic, caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the formation of SARS-CoV-2 VLPs was investigated as an alternative to authentic virions because the latter requires biosafety level 3 (BSL-3) facilities. This allowed researchers to model its assembly and budding processes, examine the role of mutations in variants of concern, and determine how the structural proteins interact with each other. Also, the absence of viral genome in VLPs circumvents worries of gains in infectivity via mutagenesis. This review summarizes the strengths and limitations of several SARS-CoV-2 VLP systems and details some of the strides that have been made in using these systems to study virus assembly and budding, viral entry, and antibody and vaccine development.

Severe fever with thrombocytopenia syndrome (SFTS) is a significant public health concern, with a high fatality rate in humans and cats. In this study, we explored the genetic determinants that contribute to the different virulence of SFTS virus (SFTSV) based on Tk-F123 and Ng-F264 strains isolated from cats. Tk-F123 was 100% lethal in type I interferon receptor-knockout mice, whereas Ng-F264 exhibited no fatality. We identified a pair of amino acid residues in the Gc protein, glycine and serine, at residues 581 and 934, respectively, derived from Tk-F123, leading to a fatal infection. Those in Ng-F264 were arginine and asparagine. These results suggest that this pair of residues affects the Gc protein function and regulates SFTSV virulence. Our findings provide useful clues for the elucidation of viral pathogenicity and the development of effective live-attenuated vaccines and antiviral strategies.

Kaposi sarcoma herpesvirus (KSHV) is an oncogenic DNA virus associated with various malignancies, including tumours like Kaposi sarcoma and Primary effusion lymphoma. Recently, the importance of the tumour microenvironment in KSHV-associated tumours is being studied. New studies utilizing human primary cells, co-culture experiments with KSHV-infected cells, and modern techniques like time-resolved single cell analysis, have significantly advanced the understanding of KSHV interactions with monocytes and macrophages. These cells play key roles in shaping the tumour microenvironment. It has become clear that KSHV-infected endothelial cells regulate the growth and the differentiation of monocytes and macrophages. Monocytes and macrophages, in turn, can regulate KSHV-infected cells in tumorigenesis and cytokine secretion, leading to the pro-tumour microenvironment. Further investigations into the viral regulation of monocytes and macrophages thus have potential to lead to the discovery of novel antitumour therapeutics.

Viral proteins with intrinsic disorder, such as the p26 movement protein from Pea enation mosaic virus 2 (PEMV2), can phase separate and form condensates that aid specific stages of virus replication. However, little is known about the impact of viral condensate formation on essential cellular processes, like translation. In this study, we performed mass spectrometry on affinity-purified p26 condensates and found an enrichment of RNA-binding proteins involved in translation and ribosome biogenesis. Puromycin assays and polysome profiling show that ectopic p26 expression suppresses ribosome assembly and translation in Nicotiana benthamiana, mirroring defects in late-stage PEMV2 infection. Despite interactions with the 2'-O-methyltransferase fibrillarin, p26 does not inhibit translation by altering rRNA methylation but instead binds directly to rRNAs and decreases their solubility. Disruption of ribosome assembly and translation by p26 during late PEMV2 infection may promote stages of the virus replication cycle that are incompatible with translation, including systemic movement.

The red swamp crayfish (Procambarus clarkii) is the second most widely cultured crustacean globally. As a highly invasive species with a worldwide distribution, P. clarkii presents a substantial risk for the transmission of viral pathogens to native aquatic organisms. Recently, the emergence of growth retardation disease (GRD) in P. clarkii has led to significant production declines and economic losses. A comprehensive viromic analysis could offer valuable insights into the potential viral pathogens harbored by P. clarkii. Here we systematically examined the RNA viromes of healthy and GRD-affected P. clarkii collected from Qianjiang, China. Our investigation identified a total of 1729 viral species across 21 known viral taxa, with 1603 species being previously unreported. The orders Picornavirales, Tolivirales, and Nodamuvirales were predominant in both species count and relative abundance. Moreover, seven viruses exhibited higher abundance in GRD-affected P. clarkii compared to healthy individuals. Our work uncovers an unexpectedly diverse RNA viral community within P. clarkii and identifies potential viral pathogens associated with GRD in this species.

Mammalian membrane sialic acid is the key receptor for influenza virus. Sialidases, the main type of enzyme that can hydrolyze membrane sialic acids in mammalian cells, have the potential to affect the invasion process of influenza viruses, including H1N1. For the first time, this study focused on the regulation mechanism of sialidase NEU1 expression, and revealed that swine-origin influenza (H1N1) virus infection can promote NEU1 expression through histone H3 acetylation, which is regulated by HDAC2 in host cells. This study not only provides evidence for the regulatory mechanisms of mammalian sialase NEU1 expression, but also provides new insights into the host immune defense response against influenza virus infection.

Coxsackievirus A6 (CVA6) is a primary pathogen associated with hand, foot, and mouth disease (HFMD) and is typified by fever, rashes or herpetic lesions at distinct locations. Although HFMD patients exhibit mild symptoms, a subset of patients may develop severe complications, such as viral encephalitis, myocarditis, pneumonia, and neurological disorders. However, in addition to rodent models, such as the CVA6-infected mouse model, no definitive nonhuman primate animal model or related research or analysis tool is available, which makes the development of suitable nonhuman primate animal models particularly crucial. In this study, 3- to 4-month-old rhesus monkeys were infected via the respiratory or digestive tract, and the pathogenic, pathological, and immunological alterations following CVA6 infection were investigated. The results revealed that the infected rhesus monkeys exhibited symptoms similar to those of patients, including signs of HFMD, blood cell changes, viremia, viral excretion, and inflammatory reactions during the acute phase (1-11 days). Pathological observations revealed inflammatory reactions in the intestinal and lymph node tissues. Notably, the acute symptoms gradually waned in the recovery phase (12-120 days), and a high level of neutralizing antibodies was sustained. Intriguingly, no significant disparity was observed between the infections initiated via the respiratory or digestive tract in terms of clinical symptoms, hemogram results or virus shedding. Overall, this study yielded a comprehensive dataset regarding the physiological, pathological, and immunological outcomes of CVA6 infection in a primate host, enhancing our comprehension of the mechanism of CVA6 infection and providing essential data for related vaccine and drug development.

Vaccine efficacy relies not only on antigens but also on immunomodulatory agents/adjuvants that are often used to stimulate the immune system and enhance the immune response. However, current immunomodulatory agents are used to increase the immune response induced by viral or bacterial inactivated vaccine antigens, bacterial toxoids or polysaccharides but not attenuated live viruses. Based on the immunomodulatory functions of G-CSF and the characteristics of feline herpesvirus-1 (FHV-1) as an expression vector, a recombinant virus expressing feline G-CSF (WH2020-ΔTK/gI/gE-G-CSF) was constructed. The growth dynamics of WH2020-ΔTK/gI/gE-G-CSF were similar to those of WH2020-ΔTK/gI/gE. Compared with kittens vaccinated with WH2020 Δ TK/gI/gE, felines inoculated with WH2020 ΔTK/gI/gE-G-CSF produced more neutralizing antibodies and neutrophils, further alleviating clinical symptoms after FHV-1 infection. Taken together, our results revealed the potential of G-CSF as an ideal immune potentiator that can augment immune responses to FHV-1 and even other attenuated live vaccines.

AQUAREOVIRUS: a genus of within the family Spinareoviridae, order Reovirales, infects aquatic animals. Their genomes comprise 11 segments of double-stranded RNA, which function directly as mRNAs upon release into the cytoplasm of infected cells. Here, liquid chromatography-tandem mass spectrometry was employed to annotate small coding ORFs in the Aquareovirus-C genome. Its plus-strand RNA of segment 8 (S8) contains a novel protein-coding frame (NS15), and S5 seems to has an additional reading frame (NS18) with a putative non-AUG initiation codon. Among them, NS15 polypeptide has been proved by immunoblotting assay. Remarkably, the S4 and S11 minus-strand mRNAs may encode polypeptides, suggesting ambisense polarity of the two segmented RNAs. And the newly discovered NS12 ORF in 2019, from viral tricistronic S7 mRNA, was also confirmed by this mass-spectrometry data. Taken together, these identified new ORFs reveal the genome-coding complexity of Aquareovirus-C.

Foamy virus (FV) is a retrovirus with a safer integration profile than other retroviruses, rendering it appealing for gene therapy. Prototype FV (PFV) vector systems have been devised to yield high-titer vectors carrying large transgenes. Subsequent iterations of PFV vectors have been engineered to be replication-incompetent, enhancing their safety. A third generation PFV vector system, composed of four plasmids, has been adapted to accommodate large transgenes. Additionally, a novel dual-vector system shows promise for convenient and efficient gene delivery, particularly with the forthcoming development of stable producer cell lines expressing PFV Env. FVs exhibit a broad host spectrum due to the ubiquitous presence of the host factor, heparan sulfate (HS), on their surface. The receptor-binding domain (RBD) of FV Env proteins plays a crucial role in binding to the host cell HS. The FV vector system has been employed in hematopoietic stem cell (HSC) gene therapy to address monogenic diseases in dog and mouse models. In addition, FV vectors safely and efficiently deliver anti-HIV transgenes to HSCs, and vectors carrying HIV epitopes successfully induce antibodies against HIV, offering the promise of anti-HIV gene therapy and vaccine development. In this review, we delve into the development and utilization of FV vector systems, emphasizing their unique advantages in gene therapy, including their non-pathogenic nature, broad host tropism, large transgene capacity, and persistence in resting cells. Furthermore, we discuss the potential of FV vectors in tackling current challenges in gene therapy and their viability as valuable tools for treating genetic diseases.

Viruses as obligate intracellular parasites are limited by their small genome. They have thus developed various strategies to maximize viral fitness with a limited amount of coding information. Among these strategies is the use of the same viral protein for multiple functions. The μ2 protein of mammalian reovirus is one such example of a multifunctional protein. We will present recent progress in our understanding of some functions and properties of this protein that have been revealed in the last two or three decades, such as its impact on the formation of viral factories or the control of the interferon response. We will also examine the recently established structure of the protein and the most recent data on the protein's enzymatic activities in the context of viral RNA synthesis. Finally, the impact of μ2 in the regulation of host-cell alternative mRNA splicing will be presented and future avenues of research discussed.

CD4 and CD8 tissue resident memory cells (TRM) express many shared anti-viral activities upon re-exposure to virus. CD4 T cells were depleted from HSV-immune guinea pigs to identify CD4-dependent functions in the vaginal mucosa following HSV-2 challenge. The incidence of animals shedding HSV-2 fell rapidly after challenge in control animals but remained significantly higher through day four post infection in CD4-depleted animals. Genes encoding CD14, IFN-γ, CCL2, and CCL5 were up-regulated in the vaginal mucosa of both groups following challenge. However, significantly higher expression of CD107b, IL-15, and TLR9 but lower expression of CD20, IL-21, and CCL5 was detected in CD4-depleted- compared to control-treated animals. Further, antigen stimulation of CD4 TRM increased the expression of IFN-γ, IL-2, IL-21, IL-17A, and CCL5. The impact of these gene expression patterns on the recruitment and maintenance of the cellular milieu of the vaginal mucosa upon virus challenge is discussed.

The ΦX174 reduction effect describes a plasmid-based inhibitory phenomenon that mimics the superinfection inhibition found in wild phage populations. In this effect, when a portion of the ΦX174 genome - the 3' end of the pilot protein gene (H), the 5' end of the replication gene (A), and the H-A intergenic region - is present on a plasmid in the host cell, almost complete protection from phage infection occurs. Here we demonstrate that only the phage pilot protein H portion of the plasmid is sufficient for the observed inhibition, that protein synthesis is necessary for inhibition to occur, that inserting the entire H gene in the plasmid may also impart a blocking effect, and that partial to complete recovery from this inhibition is possible with minimal viral evolution.