Virology Journal最新文献

Background: The RNA bacteriophage MS2 is an RNA phage that infects the bacterium E. coli and is one of the most studied and prototypical model phages in molecular biology and microbiology. Previous research revealed complex translational control and fine-tuning for MS2 replication. However, the dynamics of its transcriptional activity and replication during the life cycles within the bacteria remain elusive.

Methods: Here, we employed Nanopore Direct RNA sequencing (DRS) to investigate the transcriptome and epitranscriptome landscape of the MS2 in infected E. coli throughout multiple life cycles.

Results: We discovered that MS2 phages sustain a high level of transcriptional activity required for replication. We found large amounts of subgenomic small transcripts from RNA degradation, Nanopore DRS bias, and transcripts containing the coat-encoding region, required for virion assembly. We found the error-prone activity of the MS2 replicase produced hybrid reads from the template-switching mechanism. We finally evidenced that RNA modification is conserved throughout the entire life cycle in full-length transcripts without the acquisition of new modifications, whereas small transcripts did acquire newly modified sites. The conserved sequence and secondary structure (U-rich hairpin) of Ψ installation sites were the most amenable to RNA modification, from potentially the host RluA-mediated installation.

Conclusions: Overall, our investigation revealed a more complex transcriptional dynamics of MS2 phages than anticipated within E. coli to maintain its growth and replication under host pressure.

Human papillomaviruses (HPVs), particularly types 16 and 18, are major contributors to cervical cancer through the oncogenic activities of the E6 and E7 proteins. These viral proteins inactivate the tumour suppressors p53 and pRB, driving uncontrolled cellular proliferation. In this study, we investigated the interaction between the HPV E7 protein and the R2TP complex, a co-chaperone involved in essential cellular functions, including ribosome biogenesis, transcription, and macromolecular assembly. We identified PIH1D1, a core R2TP subunit, as an interacting partner of HPV16 and HPV18 E7 proteins. Mutagenesis and pull-down assays showed that phosphorylation of HPV E7 by casein kinase 2 (CK2) is critical for this interaction, as mutations of serine residues within the CK2 phospho-acceptor site on E7 disrupted the binding with PIH1D1. Furthermore, PIH1D1 facilitated the association of E7 with the retinoblastoma protein (pRB), forming a complex that likely promotes cancer cell proliferation. Immunohistochemical analysis of cervical cancer tissues revealed overexpression of PIH1D1, RUVBL1, and RPAP3-key components of the R2TP complex. Functional assays confirmed that PIH1D1 is crucial for cervical cancer cell growth and migration, as its silencing reduced E7 stability and impaired proliferation. Collectively, these findings highlight that PIH1D1, and by extension, the R2TP complex, is integral to the HPV-driven malignancy and suggest potential as therapeutic targets in HPV-related cancers. IMPORTANCE: Despite being largely preventable through vaccination, cervical cancer is a significant concern for public health. Research is essential to understand the factors contributing to its high incidence and mortality and to devise effective prevention and treatment strategies. We investigated the functional role of PIH1D1, a core subunit of the R2TP complex, in the HPV-mediated cervical carcinogenesis. The interaction of the R2TP complex, HPV E7, and the tumour suppressor pRB proteins may be essential in driving malignant transformation.

Many viruses express their proteins in the form of large polyproteins comprising structural and non-structural (e.g. enzymatic) units that are released from the precursor through ordered proteolysis. Proteolytic processing of polyproteins is an indispensable regulatory step for virus maturation and replication that is carried out by the virus-encoded and/or cellular proteases. The activity of a viral protease that is expressed as a part of a polyprotein is controlled in part by the self-cleavage (autoprocessing) from the precursor. The mechanism of protease precursor processing has been established at the molecular level for various RNA virus proteases, including human immunodeficiency virus (HIV) and severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Both viral protease precursors are processed via intra- (in cis) and intermolecular (in trans) cleavages at the N- and C-termini, respectively, yielding the mature enzyme. The remarkably similar activation mechanisms of HIV and SARS-CoV-2 PRs suggest that other viral proteases are activated similarly. In this review, we provide a detailed overview on the protease precursor autoprocessing mechanism of HIV-1 and SARS-CoV-2 proteases and compare those to the activation mechanism of non-viral proteases from their zymogens. Also, we review the activation mechanism of other ss(+)RNA viruses that utilize the polyprotein pathway for their replication. Based on such comparison, it appears that the protease activation mechanisms of most enveloped ss(+)RNA viruses from their precursors share many common features, although they do not correlate directly with the evolutionary relationships, the presence or absence of viral envelope or the catalytic mechanism of the viral protease.

Feline alphaherpesvirus-1 (FHV-1) is generally considered to have a narrow host range restricted to the Felidae family. As a result, FHV-1 has been proposed as a potential vaccine vector to carry foreign pathogen or immunocontraceptive antigens, for use in domestic cat (Felis catus) populations. The species-specificity of FHV-1 has been described previously in the 1970s where several non-feline hosts were assessed for their inability to be infected by FHV-1. However, more recently, evidence of FHV-1 infection in BALB/c mice was reported, furthering the need for additional investigation into the host range potential of FHV-1. This study investigated the species-specificity of FHV-1 and three modified FHV-1 variants containing antigens intended as immunocontraceptive targets. Their ability to replicate in respiratory tissue, cause clinical signs and induce disruptions in female reproductive tissues was studied in an in vivo murine model. Furthermore, a diverse range of non-feline cell types originating from species including domestic animals, wildlife species and a non-human primate were also investigated for their ability to support FHV-1 immunocontraceptive replication in vitro. No evidence of FHV-1 replication was detected in vitro in the non-feline cell lines, nor was any evidence of disruption to female reproductive tissues or viral activity detected in the murine model. The findings contribute towards the global understanding of FHV-1 host range and add support for its use as a feline-specific viral vector.

Dengue virus (DENV) transmission in Bolivia has historically been restricted to tropical lowland regions, where the primary vector Ae. aegypti is endemic. In recent years, however, the vector has expanded into high-altitude areas. In 2024, Cochabamba, located at ~ 2,550 m above sea level, experienced its largest recorded dengue outbreak. We analyzed data from 9,576 suspected dengue cases reported between January and July 2024, of which 5,923 were laboratory-confirmed. A subset of DENV-positive samples underwent whole-genome sequencing and phylogenetic analysis. Dengue cases were detected in municipalities located at elevations up to 2,719 m, with the highest number reported in Cochabamba city (4,104/5,923, 69.3%; 2,558 m) and the highest incidence observed in Capinota (839 cases per 100,000 population; 2,386 m). Molecular analyses indicated predominant circulation of DENV-2 genotype II clade F.1.1.2. Phylogenetic reconstruction identified two distinct clades closely related to viruses previously detected in southern Brazil, suggesting multiple independent introductions into the department in late 2023. However, the limited availability of genomic data from Bolivia and neighboring countries limits precise identification of the outbreak's origin. This study provides the first characterization of the 2024 dengue outbreak in Cochabamba, an exceptional event of substantial DENV circulation at high altitude, likely facilitated by the expanding geographic range of Ae. aegypti, potentially driven by climate change.

Zika virus (ZIKV) infections have impacted public health in Brazil since 2015, primally due to Congenital Zika Virus Syndrome (CZVS) cases, which may lead to microcephaly and other clinical manifestations, such as hearing and visual impairments. More than ten years after the first diagnosis of Zika in Brazil, some progress has been made and the epidemiological scenario has improved considerably. However, despite these advances, more than a thousand ZIKV infections are reported annually in Brazil and new CZVS cases continue to be observed. Herein, we performed a retrospective and prospective analysis to assess the progress made and identify gaps and challenges that still need to be addressed. Overall, we believe that future ZIKV control efforts in Brazil must include: enhanced vector control measures; surveillance for potential vertebrate reservoirs; medical care for pregnant women, including prevention of infection and vertical transmission; sensitive and specific intrauterine CZVS diagnosis; ongoing support for children and families with CZVS cases; expansion of the national diagnostic network for arboviruses, including encouraging healthcare professionals to perform laboratory tests; assessment of the impactof the dengue vaccine, recently implemented in Brazil, on ZIKV infections; and affordable, sensitive and specific multiplex diagnostic strategies adequately validated for cross-reactivity with other arboviruses circulating in Brazil.

Background: Human cytomegalovirus (HCMV) infection is responsible for congenital infections and poses a significant health risk to immunocompromised individuals. To date, there is no effective HCMV vaccine. Salvia miltiorrhiza is a well-known traditional Chinese herb that possesses active pharmacological effects including antiviral activity. However, no studies on the effects of Salvia miltiorrhiza on HCMV have been reported thus far.

Methods: Firstly, the cytotoxicity of five active components derived from Salvia miltiorrhiza was screened, then morphology, western blotting, qPCR, indirect immunofluorescence assay (IFA) and TCID₅₀ assay were used to assess anti-HCMV activity. Time-of-addition experiments were performed to identify the stage at which the active components were active. The mechanism of the anti-HCMV activity of the active component was also illustrated in this study.

Results: Salvianolic acid B, Tanshinone ⅡA, and Cryptotanshinone derived from Salvia miltiorrhiza were found to prevent lytic cytopathic changes, inhibit the expression of viral proteins, suppress the replication of HCMV DNA, and significantly reduce the viral titer in WI-38 cells. Time-of-addition experiments showed that they predominantly act during the Post-entry treatment (Post-T) stage. Moreover, they effectively suppressed HCMV-induced cellular senescence, which was evidenced by a decline in senescence-associated β-galactosidase activity, alleviated senescence-associated heterochromatin foci (SAHF), reduced expression of p16, p21, and p53, and decreased production of reactive oxygen species (ROS).

Conclusion: To our knowledge, this is the first report that salvianolic acid B, tanshinone IIA, and cryptotanshinone derived from Salvia miltiorrhiza exhibit anti-HCMV activity in vitro, and suggest potential for further development.

African swine fever (ASF) is a highly fatal disease caused by the African swine fever virus (ASFV). It infects domestic pigs and wild boars, causing significant economic losses worldwide. However, effective vaccines against this virus remain not be commercialized because of its large genome and high mutation frequency. Thus, antiviral therapies need to be developed urgently. Chloroquine phosphate (CQP) has been demonstrated in previous studies to exert inhibitory effects against a variety of viruses, but its inhibitory effect against the SY-1 strain of ASFV remains unclear. Therefore, we selected CQP as the research subject to investigate its anti-ASFV function. In this study, we confirmed that CQP has a significant inhibitory effect on the ASFV SY-1 strain by RT-qPCR, Western Blot, and HAD50. Transcriptome sequencing and KEGG pathway enrichment analysis showed that CQP treatment significantly affected multiple signaling pathways, including the cytokine-cytokine receptor interaction, Toll-like receptor signaling pathway, tumour necrosis factor (TNF) signaling pathway and IL-17 signaling pathway. Western Blot results further indicated that CQP can inhibit ERK phosphorylation. Treatment with the MAPK agonist C16-PAF reversed the inhibitory effect of CQP, verifying the key role of this pathway in the anti-viral mechanism of CQP. In sum, the results of this study indicate that CQP effectively inhibits ASFV replication by suppressing the MAPK-ERK signaling pathway. This study provides a theoretical basis and technical support for the development of anti-viral strategies targeting ASFV.

West Nile virus (WNV) and Japanese encephalitis virus (JEV) are major mosquito-borne pathogens that cause severe neurological illnesses in humans, including encephalitis and meningitis. Neurons are the primary target of WNV and JEV infections in the brain. However, the pathogenic mechanisms and host immune response associated with WNV and JEV infections in the primary neurons are not fully characterized. Herein, using high-throughput RNA sequencing (RNA-Seq), we performed a comparative transcriptomic analysis of the primary mouse neurons following infection with WNV and JEV. We identified differentially expressed host genes (DEGs) that were commonly induced in neurons by both viruses, as well as those uniquely regulated in a virus-specific manner. We observed pronounced changes in gene expression profiles that indicate activation of a robust antiviral response accompanied by signatures of neuronal dysfunction. Notably, interferon-stimulated genes such as Irf7, Isg15, and Ifnb1 were markedly upregulated in WNV- and JEV-infected neurons. Additionally, we observed the significant upregulation of proinflammatory chemokine and cytokine genes such as Il6, Cxcl2 and Cxcl10 which was further validated by Luminex assay. Ingenuity Pathway Analysis (IPA) revealed the activation of several cell death pathways including pyroptosis and necroptosis. We also showed the significant dysregulation in pattern recognition receptors and neurotransmitter receptors such as glutamatergic and acetylcholine receptors. This dysregulation extended to the synaptic signaling and reduced synaptogenesis in infected neurons. Quantitative RT-PCR confirmed the upregulation of key cytokines like Il6 and Ifna, further establishing the validity of the RNA-Seq approach. Overall, this study provides insights into the pathological changes in the neurons following WNV and JEV infections.