Journal of General Virology最新文献

Infection with SARS-CoV-2 continues to be a threat to human health. Despite successful immunization campaigns, effective treatment of COVID-19 remains an essential need to help patients and prevent the spread of new virus strains. Viroporins are intracellular ion channels that are essential for virus replication and release, thus presenting promising pharmaceutical targets. Mutations found in variants of concern (VOC) are expected to increase the virulence of the new virus strains. Recognizing the effects of these mutations at the molecular level is essential for the development of improved therapies. Here, we characterized the putative viroporin ORF3a found in VOCs of SARS-CoV-2, using expression constructs containing a myc-tag for identification, and an optional membrane-directing signal peptide. Additionally, constructs containing N-terminal fluorescence protein tags were prepared. Expression and cell surface transport in HEK-293 cells were studied using Western blot and dot blot assays, and the cellular distribution of fluorescent-marked ORF3a was studied using subcellular organelle markers and high-resolution fluorescence microscopy. Viroporin activity of all ORF3a constructs was assessed using cell viability and metabolic assays, as well as patch-clamp recordings of recombinant ORF3a. All ORF3a mutants were expressed well in the recombinant system, and the presence of a signal peptide increased expression on the cellular surface. Intracellular distribution was similar for all variants. The VOC mutants ORF3a-S171L and ORF3a-Q57H showed reduced cytotoxic activity and sensitivity to the viroporin inhibitor rimantadine, respectively, suggesting these positions to be relevant for ORF3a function and a starting point for the search of novel antiviral drugs.

Chikungunya is a febrile infection caused by the Chikungunya virus (CHIKV), an alphavirus which has emerged as a serious public health problem globally. Despite extensive research, our understanding of different host factors facilitating effective CHIKV infection is not clear yet. NEDD4, a member of the E3 ubiquitin ligase, is one such protein. Here, the importance of NEDD4 has been explored during CHIKV infection in vitro. It was observed that the level of NEDD4 is downregulated after CHIKV infection. Interestingly, the CHIKV-nsP3 level and the viral load were decreased significantly when NEDD4 was silenced, while a 93% decrease in the viral load was observed in the case of NEDD4 overexpression, indicating the importance of an optimum level of NEDD4 for effective CHIKV infection. Further study revealed that there was interaction between the NEDD4 and CHIKV-nsP3 proteins through co-immunoprecipitation (CO-IP) during CHIKV infection. Additionally, in silico data illustrated that the WW domain of NEDD4 can bind to the nsP3, as well as the macrodomain of nsP3 (nsp3-MD) of CHIKV. These data were further confirmed by the pull-down assay with purified nsP3-MD. The finding suggested that the host protein NEDD4 might interact directly with nsP3-MD during the CHIKV infection. However, the presence of a faint band of NEDD4 along with nsP3-MD in the pull-down assay may indicate the involvement of some other residues for this interaction. These in silico data were further confirmed by the CO-IP experiments, where all domains of nsP3, MD (macrodomain), AUD (alphavirus unique domain) and HVD (hypervariable domain) were found to interact with NEDD4. Additional experiments with a truncated form of MD, MD1 (1-100 residues of amino acid), revealed that this region is not able to maintain the interaction with NEDD4, indicating the crucial role of the C-terminal region of MD for this binding. In conclusion, these findings offer valuable insights about the importance of NEDD4 during CHIKV infection and the residues of nsP3 for its interaction, which might be useful to design future therapeutics against CHIKV.

Flaviviridae is a family of viruses that are mainly transmitted by mosquito vectors of the genus Aedes, which cause febrile illnesses and, in severe cases, haemorrhagic or neurodegenerative conditions. Over time, these viruses have been reported as emerging pathogens, leading to epidemic outbreaks in various regions worldwide. Additionally, climate change has facilitated the migration of these vectors to regions where they were not previously found. Unfortunately, there are currently no effective treatments or vaccines to prevent or combat Orthoflavivirus infections. Consequently, a deeper understanding of the viral biology and the human host immune response is crucial for advancing the development of therapeutic targets. Amongst the molecules involved in the innate immune response to viral infections are antimicrobial peptides (AMPs), which have been studied for decades. However, their role in Orthoflavivirus infections remains poorly understood. Several researchers have proposed the stimulation or exogenous administration of AMPs during various viral infections, highlighting these molecules as potential innovative therapeutic targets. This study compiles current knowledge on AMPs with a specific focus on Orthoflavivirus infections, emphasizing the importance of these promising therapeutic approaches.

A hallmark of the dengue virus (DENV) infection is the manipulation of host cell membranes, lipid trafficking and lipid droplets (LD), all cellular functions that depend on the cytoskeleton and the cytoplasmic streaming system. We previously reported the interaction between the DENV non-structural (NS1) protein and members of the kinesin motor complex in the Aedes albopictus cell line C6/36. In this work, we present evidence indicating that the protein kinesin light chain 1 (KLC1) is indeed a susceptibility factor for the DENV replicative cycle in mosquito cells. The interaction between NS1 and KLC1 was confirmed by proximity ligation and co-immunoprecipitation assays in cells harvested 24 hpi. In addition, transmission immunoelectron microscopy showed KLC1 decorating the surface of vacuoles in association with NS1. Increased levels of KLC1 were observed starting at 6 hpi, suggesting that virus infection stimulates KLC1 synthesis. Silencing KLC1 expression results in a reduction in viral genome synthesis, decreased secretion of NS1 and a reduction of virus progeny by nearly 1 log. In agreement, similar affectations were observed in infected cells transfected with a peptide that competes and interferes with the interaction between KLC1 and its cargo molecules. Of note, both silencing the expression and interfering with the function of KLC1 resulted in a disorganization of LD, which decreased in number and increased in area, in mock or infected cells. These results, taken together, suggest that KLC1 is a host susceptibility factor for DENV in mosquito cells and appears to play an important role in the proper transport and homeostasis of LD required for flavivirus replication. However, modest colocalization was observed between NS1 and LD, and the significance of the KLC1 and NS1 interactions needs to be further investigated.

This article summarises the activities of the International Committee on Taxonomy of Viruses Bacterial Viruses Subcommittee, detailing developments in the classification of bacterial viruses. We provide here an overview of all new, abolished, moved and renamed taxa proposed in 2024, approved by the Executive Committee, and ratified by membership vote in 2025. Through the collective efforts of 74 international contributors of taxonomy proposals in this round, 43 ratified proposals have led to the creation of one new phylum, one class, four orders, 33 families, 14 subfamilies, 194 genera and 995 species. These proposals mark significant progress in refining the taxonomy of bacterial viruses. Key updates include the creation of new orders and families that include existing taxa to better reflect genomic and evolutionary relationships. As sequencing and bioinformatics approaches continue to advance, further expansion and refinements in viral taxonomy can be anticipated in the coming years.

Mpox (formerly known as monkeypox) virus (MPXV) is the zoonotic pathogen of mpox disease in humans. Its increasing emergence outside of its endemic area has heightened the importance of investigating the virus' prevalence and maintenance in sylvatic reservoirs. The common brown rat (Rattus norvegicus) can inhabit almost anywhere in the UK, posing a threat to zoonotic transmission to humans. Two independent studies were carried out; the first investigated the susceptibility of brown rats to MPXV infection with a clade IIb mpox strain via two challenge routes: intranasal and intradermal. The second study considered the transmission of MPXV between challenged and naïve brown rats. All animals were asymptomatic to mpox disease, although enzyme-linked immunosorbent assay (ELISA) confirmed subclinical infection in challenge groups. In the susceptibility study, reverse transcription PCR (RT-PCR) detected mpox DNA in the lung tissue and throat swabs within the intranasally inoculated group, in addition to viable virus observed from the intranasal throat swabs. In contrast, no virus was detected in either tissues or swabs in the intradermally inoculated group or control group. RT-PCR results from the transmission study detected mpox DNA in tissues and throat swabs taken from challenged animals. Viable virus was observed from tissues and swabs of intranasally challenged animals with infectious titres of ~10²-10⁴ TCID₅₀ per millilitre. ELISA assays in the transmission study showed replicable results compared to the first susceptibility study in directly challenged animals alongside evidence of seroconversion in co-housed naïve animals. In conclusion, brown rats are susceptible to MPXV infection, as they have been demonstrated to maintain viable virus in the absence of clinical signs. Viral transmission of MPXV from infected rats to naïve rats was not observed by RT-PCR, although naïve rats did show antibody responses when exposed to infected rats indicating exposure to virus.

Recent studies have reported a genotype 4 (G4) reassortant Eurasian avian-like (EA) H1N1 virus in swine, demonstrating a potential pandemic threat in humans. Here, we have compared the tropism, replication competence and pro-inflammatory cytokine and chemokine induction of the two G4 EA H1N1 strains in parallel with 2009 pandemic H1N1 (H1N1/pdm/09) and A/Quail/HK/G1/1997 H9N2 (G1) using ex vivo culture of the human respiratory tract and in vitro culture of human peripheral blood-derived macrophages. Our results showed that G4 strains could replicate in ex vivo cultures of human lung and bronchus with a similar replication competence to H1N1/pdm/09. The cytokine induction levels of G4 were similar to H1N1/pdm/09 in macrophages. Taken together, we could extrapolate that the G4 EA H1N1 swine influenza may pose a notable public health threat towards human and should not underestimate this threat.

The Fungal and Protist Viruses Subcommittee (SC) of the International Committee on Taxonomy of Viruses (ICTV) has received a total of eight taxonomic proposals for the 2024 annual cycle. The extent of proposed changes varied, including nomenclatural updates, creation of new taxa and reorganization of established taxa. Following the ICTV procedures, all proposals were reviewed and voted upon by the members of the Executive Committee with ratification in March 2025. As a result, a total of 52 species in the families Botourmiaviridae and Marnaviridae were renamed to comply with the mandated binomial format. A new genus has been added to the dsRNA virus family Amalgaviridae, while two new families, Splipalmiviridae (Wolframvirales) and Mycoalphaviridae (Hepelivirales), were created to classify new groups of positive-sense (+) RNA mycoviruses. The class Arfiviricetes (Cressdnaviricota) was expanded by a new order Lineavirales and a new family Oomyviridae of ssDNA viruses. Additionally, a new class Orpoviricetes was created in the kingdom Orthornavirae to classify a group of bisegmented (+)RNA viruses reported from fungi and oomycetes. Finally, the order Pimascovirales was reorganized to better depict evolutionary relationships of pithoviruses and related viruses with large dsDNA genomes. The summary of updates in the taxonomy of fungal and protist viruses presented here is limited to taxa within the remit of this Subcommittee. For information on taxonomy changes on other fungal viruses closely related to animal and/or plant viruses, please see reports from sister ICTV Subcommittees (i.e. Plant Virus SC and Animal dsRNA and ssRNA(-) Viruses SC).

As intracellular parasites, viruses must hijack and often rewire organelles, signalling pathways and the bioenergetics machinery of the infected cell to replicate their genome, produce viral proteins and assemble new viral particles. Mitochondria are key eukaryotic organelles often referred to as the cell's powerhouse. They control many fundamental cellular processes, from metabolism and energy production to calcium homeostasis and programmed cell death. Importantly, mitochondrial membranes are also critical sites for the integration and amplification of antiviral innate immune responses. Overall, mitochondria are therefore both supporting the virus life cycle by sustaining energy production, metabolism and synthesis of macromolecules and part of the cell's first line of defence against viruses. This review summarizes recent findings on viral manipulations of mitochondria and their functions. We explore the evolving understanding of how mitochondrial dynamics is targeted to regulate innate immunity, evasion strategies used to avoid mitochondrial-associated mechanisms that impair replication and the role of mitochondrial functions such as generating reactive oxygen species or regulating the electron transport chain during infection. Overall, we provide a comprehensive view of how viruses modulate mitochondrial function to promote replication.

During the 56th annual meeting of the International Committee on Taxonomy of Viruses (ICTV), held in Bari, Italy, in August 2024, two technical proposals were presented. The first called for amended versions of accepted taxonomic proposals to be named in such a way to ensure that they are readily accessible on the ICTV website (2024.001G). The second proposed a substantial reformatting of the ICTV statutes and codes to produce a more unified text after the numerous changes made to both documents in previous years (2024.002G). Finally, the ICTV Executive Committee (EC) nominated Professor Stuart Siddell as a Life Member of the ICTV for his work over four decades on virus taxonomy, including 16 years as a member of the EC (2024.003G).