Large Double-stranded DNA Research Articles

African Swine Fever Virus Gene B117L Encodes a Small Protein Endowed with Low-pH-Dependent Membrane Permeabilizing Activity.

African swine fever virus (ASFV) is causing a devastating pandemic in domestic and wild swine in Central Europe to East Asia, resulting in economic losses for the swine industry. The virus contains a large double-stranded DNA genome that contains more than 150 genes, most with no experimentally characterized function. In this study, we evaluate the potential function of the product of ASFV gene B117L, a 115-amino-acid integral membrane protein transcribed at late times during the virus replication cycle and showing no homology to any previously published protein. Hydrophobicity distribution along B117L confirmed the presence of a single transmembrane helix, which, in combination with flanking amphipathic sequences, composes a potential membrane-associated C-terminal domain of ca. 50 amino acids. Ectopic transient cell expression of the B117L gene as a green fluorescent protein (GFP) fusion protein revealed the colocalization with markers of the endoplasmic reticulum (ER). Intracellular localization of various B117L constructs also displayed a pattern for the formation of organized smooth ER (OSER) structures compatible with the presence of a single transmembrane helix with a cytoplasmic carboxy terminus. Using partially overlapping peptides, we further demonstrated that the B117L transmembrane helix has the capacity to establish spores and ion channels in membranes at low pH. Furthermore, our evolutionary analysis showed the high conservation of the transmembrane domain during the evolution of the B117L gene, indicating that the integrity of this domain is preserved by the action of the purifying selection. Collectively our data support a viroporin-like assistant role for the B117L gene-encoded product in ASFV entry. IMPORTANCE ASFV is responsible for an extensively distributed pandemic causing important economic losses in the pork industry in Eurasia. The development of countermeasures is partially limited by the insufficient knowledge regarding the function of the majority of the more than 150 genes present on the virus genome. Here, we provide data regarding the functional experimental evaluation of a previously uncharacterized ASFV gene, B117L. Our data suggest that the B117L gene encodes a small membrane protein that assists in the permeabilization of the ER-derived envelope during ASFV infection.

Functional Profiling and Evolutionary Analysis of a Marine Microalgal Virus Pangenome.

Phycodnaviridae are large double-stranded DNA viruses, which facilitate studies of host-virus interactions and co-evolution due to their prominence in algal infection and their role in the life cycle of algal blooms. However, the genomic interpretation of these viruses is hampered by a lack of functional information, stemming from the surprising number of hypothetical genes of unknown function. It is also unclear how many of these genes are widely shared within the clade. Using one of the most extensively characterized genera, Coccolithovirus, as a case study, we combined pangenome analysis, multiple functional annotation tools, AlphaFold structural modeling, and literature analysis to compare the core and accessory pangenome and assess support for novel functional predictions. We determined that the Coccolithovirus pangenome shares 30% of its genes with all 14 strains, making up the core. Notably, 34% of its genes were found in at most three strains. Core genes were enriched in early expression based on a transcriptomic dataset of Coccolithovirus EhV-201 algal infection, were more likely to be similar to host proteins than the non-core set, and were more likely to be involved in vital functions such as replication, recombination, and repair. In addition, we generated and collated annotations for the EhV representative EhV-86 from 12 different annotation sources, building up information for 142 previously hypothetical and putative membrane proteins. AlphaFold was further able to predict structures for 204 EhV-86 proteins with a modelling accuracy of good-high. These functional clues, combined with generated AlphaFold structures, provide a foundational framework for the future characterization of this model genus (and other giant viruses) and a further look into the evolution of the Coccolithovirus proteome.

African Swine Fever Virus L83L Negatively Regulates the cGAS-STING-Mediated IFN-I Pathway by Recruiting Tollip To Promote STING Autophagic Degradation.

African swine fever (ASF) is a devastating infectious disease of pigs caused by the African swine fever virus (ASFV), which poses a great danger to the global pig industry. Many viral proteins can suppress with interferon signaling to evade the host's innate immune responses. Therefore, the development of an effective vaccine against ASFV has been dampened. Recent studies have suggested that the L83L gene may be integrated into the host genome, weakening the host immune system, but the underlying mechanism is unknown. Our study found that L83L negatively regulates the cGAS-STING-mediated type I interferon (IFN-I) signaling pathway. Overexpression of L83L inhibited IFN-β promoter and ISRE activity, and knockdown of L83L induced higher transcriptional levels of interferon-stimulated genes (ISGs) and phosphorylation levels of IRF3 in primary porcine alveolar macrophages. Mechanistically, L83L interacted with cGAS and STING to promote autophagy-lysosomal degradation of STING by recruiting Tollip, thereby blocking the phosphorylation of the downstream signaling molecules TBK1, IRF3, and IκBα and reducing IFN-I production. Altogether, our study reveals a negative regulatory mechanism involving the L83L-cGAS-STING-IFN-I axis and provides insights into an evasion strategy involving autophagy and innate signaling pathways employed by ASFV. IMPORTANCE African swine fever virus (ASFV) is a large double-stranded DNA virus that primarily infects porcine macrophages. The ASFV genome encodes a large number of immunosuppressive proteins. Current options for the prevention and control of this pathogen remain pretty limited. Our study showed that overexpression of L83L inhibited the cGAS-STING-mediated type I interferon (IFN-I) signaling pathway. In contrast, the knockdown of L83L during ASFV infection enhanced IFN-I production in porcine alveolar macrophages. Additional analysis revealed that L83L protein downregulated IFN-I signaling by recruiting Tollip to promote STING autophagic degradation. Although L83L deletion has been reported to have little effect on viral replication, its immune evade mechanism has not been elucidated. The present study extends our understanding of the functions of ASFV-encoded pL83L and its immune evasion strategy, which may provide a new basis for developing a live attenuated vaccine for ASF.

The Magic Staff: A Comprehensive Overview of Baculovirus-Based Technologies Applied to Human and Animal Health.

Baculoviruses are enveloped, insect-specific viruses with large double-stranded DNA genomes. Among all the baculovirus species, Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is the most studied. Due to its characteristics regarding biosafety, narrow host range and the availability of different platforms for modifying its genome, AcMNPV has become a powerful biotechnological tool. In this review, we will address the most widespread technological applications of baculoviruses. We will begin by summarizing their natural cycle both in larvae and in cell culture and how it can be exploited. Secondly, we will explore the different baculovirus-based protein expression systems (BEVS) and their multiple applications in the pharmaceutical and biotechnological industry. We will focus particularly on the production of vaccines, many of which are either currently commercialized or in advanced stages of development (e.g., Novavax, COVID-19 vaccine). In addition, recombinant baculoviruses can be used as efficient gene transduction and protein expression vectors in vertebrate cells (e.g., BacMam). Finally, we will extensively describe various gene therapy strategies based on baculoviruses applied to the treatment of different diseases. The main objective of this work is to provide an extensive up-to-date summary of the different biotechnological applications of baculoviruses, emphasizing the genetic modification strategies used in each field.

Molecular Tools to Identify and Characterize Malignant Catarrhal Fever Viruses (MCFV) of Ruminants and Captive Artiodactyla.

The family Herpesviridae includes viruses identified in mammals, birds and reptiles. All herpesviruses share a similar structure, consisting of a large linear double-stranded DNA genome surrounded by a proteic icosahedral capsid further contained within a lipidic bilayer envelope. The continuous rise of genetic variability and the evolutionary selective pressure underlie the appearance and consolidation of novel viral strains. This applies also to several gamma(γ)-herpesviruses, whose role as primary pathogen has been often neglected and, among these to newly emerged viruses or virus variants responsible for the development of Malignant Catarrhal Fever (MCF) or MCF-like disease. The identification of γ-herpesviruses adapted to new zoological hosts requires specific molecular tools for detection and characterization. These viruses can cause MCF in livestock and wild animals, a disease generally sporadic but with serious welfare implications and which, in many cases, leads to death within a few days from the appearance of the clinical signs. In the absence of a vaccine, the first step to improve disease control is based on the improvement of molecular tools to identify and characterize these viruses, their phylogenetic relationships and evolutionary interaction with the host species. A Panherpes PCR-specific test, based on the conserved DNA polymerase gene, employing consensus/degenerate and deoxyinosine-substituted primers followed by sequencing, is still the preferred diagnostic test to confirm and characterize herpesviral infections. The drawback of this test is the amplification of a relatively short sequence, which makes phylogenetic analysis less stringent. Based on these diagnostic requirements, and with a specific focus on γ-herpesviruses, the present review aims to critically analyze the currently available methods to identify and characterize novel MCFV strains, to highlight advantages and drawbacks and to identify the gaps to be filled in order to address research priorities. Possible approaches for improving or further developing these molecular tools are also suggested.

Diversity and genomics of giant viruses in the North Pacific Subtropical Gyre.

Large double-stranded DNA viruses of the phylum Nucleocytoviricota, often referred to as "giant viruses," are ubiquitous members of marine ecosystems that are important agents of mortality for eukaryotic plankton. Although giant viruses are known to be prevalent in marine systems, their activities in oligotrophic ocean waters remain unclear. Oligotrophic gyres constitute the majority of the ocean and assessing viral activities in these regions is therefore critical for understanding overall marine microbial processes. In this study, we generated 11 metagenome-assembled genomes (MAGs) of giant viruses from samples previously collected from Station ALOHA in the North Pacific Subtropical Gyre. Phylogenetic analyses revealed that they belong to the orders Imitervirales (n = 6), Algavirales (n = 4), and Pimascovirales (n = 1). Genome sizes ranged from ~119-574 kbp, and several of the genomes encoded predicted TCA cycle components, cytoskeletal proteins, collagen, rhodopsins, and proteins potentially involved in other cellular processes. Comparison with other marine metagenomes revealed that several have broad distribution across ocean basins and represent abundant viral constituents of pelagic surface waters. Our work sheds light on the diversity of giant viruses present in oligotrophic ocean waters across the globe.

Identification and analysis of the interaction network of African swine fever virus D1133L with host proteins.

African swine fever (ASF) is a contagious and lethal hemorrhagic disease in pigs; its spread results in huge economic losses to the global pig industry. ASF virus (ASFV) is a large double-stranded DNA virus encoding >150 open reading frames. Among them, ASFV-encoded D1133L was predicted to be a helicase but its specific function remains unknown. Since virus-host protein interactions are key to understanding viral protein function, we used co-immunoprecipitation combined with liquid chromatography-mass spectrometry to investigate D1133L. This study describes the interaction network of ASFV D1133L protein in porcine kidney PK-15 cells. Overall, 1,471 host proteins that potentially interact with D1133L are identified. Based on these host proteins, a protein-protein network was constructed. Gene ontology and Kyoto Encyclopedia of Genes and Genomes enrichment analyses showed that cellular D1133L-interacted proteins are involved in the ribosome, spliceosome, RNA transport, oxidative phosphorylation, proteasome, and DNA replication. Vimentin (VIM), tripartite motif-containing protein 21 (TRIM21), and Tu translation elongation factor (TUFM) were confirmed to interact with D1133L in vitro. VIM or TRIM21 overexpression significantly promoted ASFV replication, but TUFM overexpression significantly inhibited ASFV replication. These results help elucidate the specific functions of D1133L and the potential mechanisms underlying ASFV replication.

In vitro and in vivo antiviral activity of nucleoside analogue cHPMPC against African swine fever virus replication

African swine fever virus (ASFV) causes a haemorrhagic disease affecting wild boar and domestic pigs which can result in morbidity and fatality rates of up to 100%. ASFV is a large double-stranded DNA virus which replicates predominantly in the cell cytoplasm and codes for its replication and transcription machinery. No vaccine is widely available and control depends on early detection, culling of infected herds and adherence to biosecurity measures. In this study the small molecule nucleoside analogue, cyclic cidofovir (cHPMPC), was evaluated for its ability to inhibit replication of four different ASFV genotypes in primary porcine macrophages. Time of addition studies demonstrated that cHPMPC effectively inhibits ASFV replication and late gene expression when added pre-infection or early post-infection but not when added at late times, suggesting the drug target may be the virus DNA polymerase, or the RNA polymerase involved in late transcription. Oral administration of cHPMPC delayed onset of clinical signs and significantly reduced viral titres in blood and tissues of treated pigs. These results indicate that cHPMPC is a promising compound for further development to control ASFV outbreaks.

Monkeypox: Virology, Pathophysiology, Clinical Characteristics, Epidemiology, Vaccines, Diagnosis, and Treatments.

The World Health Organization, has declared the recent multiregional outbreak of monkeypox, a global public health emergency. Monkeypox is a zoonotic viral infection endemic to the west and central Africa. It belongs to the Poxviridae family, the Chordopoxvirinae subfamily, and the Orthopoxvirus genus. The Poxviridae family generally consists of complex, large, enveloped, and linear double-stranded DNA viruses. The initial clinical symptoms of monkeypox are often fever, severe headache, lymphadenopathy, myalgia, and fatigue. The skin lesions typically erupt within 1-3 days of the onset of fever. The rash tends to be more localized on the face and extremities than on the trunk. Monkeypox is often a self-limiting infection, and symptoms last from 2 to 4 weeks. It is isolated from various species, but the exact natural host is uncertain. Monkeypox is transmitted by close contact with infected humans or animals. Currently, no specific medication is available for monkeypox, and the existing therapeutics are the anti-viral agents approved for smallpox infection, including tecovirimat, cidofovir, and brincidofovir. Additionally, the U.S. Food and Drug Administration has approved Vaccinia Immune Globulin Intravenous for treating vaccination complications. It is diagnosed by PCR. There are currently two vaccines licensed by the U.S. Food and Drug Administration. According to the WHO guidance, the first-generation smallpox vaccines held in national reserves of some countries are not recommended as they do not meet the current safety and manufacturing standards. The interim guidance indicates that new and safer (second- and third generation) vaccines for smallpox, may be beneficial for monkeypox prevention, including JYNNEOS, which has been approved for the prevention of monkeypox. Human monkeypox was first reported in 1970. Since then, it has caused several outbreaks, mainly in central and west Africa. The first monkeypox outbreak outside of Africa occurred in the United States in 2003, linked to contact with infected pet prairie dogs. More recently (2018-2021), monkeypox cases have been reported in travelers from Nigeria to the United Kingdom, Israel, Singapore, and the US. Since May 2022, multiple monkeypox cases have been confirmed in several non-endemic countries, raising the concern of an emerging global pandemic. This review is an updated overview of our current state of knowledge regarding monkeypox virology, pathophysiology, clinical characteristics, epidemiology, vaccines, diagnosis, and treatment options.

Discovery of the first PD-1 ligand encoded by a pathogen.

Large double-stranded DNA viruses deploy multiple strategies to subvert host immune defenses. Some of these tactics are mediated by viral gene products acquired by horizontal gene transfer from the corresponding hosts and shaped throughout evolution. The programmed death-1 (PD-1) receptor and its ligands, PD-L1 and PD-L2, play a pivotal role attenuating T-cell responses and regulating immune tolerance. In this study, we report the first functional PD-L1 homolog gene (De2) found in a pathogen. De2, captured by a γ-herpesvirus from its host during co-evolution around 50 million years ago, encodes a cell-surface glycoprotein that interacts with high affinity and stability with host PD-1. We also find that mutations evolved by the viral protein result in a significant loss of its ability to interact in cis with CD80, an interaction that for PD-L1:CD80 has been reported to block PD-1 inhibitory pathways. Furthermore, we demonstrate that the viral protein strongly inhibits T-cell signaling. Our observations suggest that PD-L1 homologs may enable viruses to evade T cell responses, favor their replication, and prevent excessive tissue damage. Altogether, our findings reveal a novel viral immunosuppressive strategy and highlight the importance of the modulation of the PD-1/PD-L1 axis during viral infections.

Identification and characterization of coiled-coil motifs across Autographa californica multiple nucleopolyhedrovirus genome

Coiled coils (CCs) are protein structural motifs universally found in proteins and mediate a plethora of biological interactions, and thus their reliable annotation is crucial for studies of protein structure and function. Autographa californica multiple nucleopolyhedrovirus (AcMNPV) is a large double-stranded DNA (dsDNA) virus and encodes 154 proteins. In this study, genome-wide scans of previously uncharacterized CC motifs throughout AcMNPV was conducted using CC prediction software. In total, 24 CC motifs in 19 CC proteins with high confidence were identified. The characteristic of viral CC motifs were analyzed. The CC proteins could be divided into 12 viral structural proteins and 7 non-structural proteins, including viral membrane fusion proteins, enzymes, and transcription factors. Moreover, CC motifs are conserved in the baculoviral orthologs of 14 of the 19 proteins. It is noted that five CC proteins, including Ac51, Ac66, Exon0, Ac13, and GP16, were previously identified to function in the nuclear egress of nucleocapsids, and Ac66 contains multiple CC motifs, the longest of which comprises 252 amino acids, suggesting a role of CC motifs in this process. Taken together, the CC motifs identified in this study are valuable resource for studying protein function and protein interaction networks during virus replication.

Structural variation turnovers and defective genomes: key drivers for the in vitro evolution of the large double-stranded DNA koi herpesvirus (KHV)

Structural variation turnovers and defective genomes: key drivers for the in vitro evolution of the large double-stranded DNA koi herpesvirus (KHV)

KSHV Topologically Associating Domains in Latent and Reactivated Viral Chromatin.

ABSTRACTEukaryotic genomes are structurally organized via the formation of multiple loops that create gene expression regulatory units called topologically associating domains (TADs). Here we revealed the KSHV TAD structure at 500 bp resolution and constructed a 3D KSHV genomic structural model with 2 kb binning. The latent KSHV genome formed very similar genomic architectures in three different naturally infected PEL cell lines and in an experimentally infected epithelial cell line. The majority of the TAD boundaries were occupied by structural maintenance of chromosomes (SMC1) cohesin complex and CCCTC-binding factor (CTCF), and the KSHV transactivator was recruited to those sites during reactivation. Triggering KSHV gene expression decreased prewired genomic loops within the regulatory unit, while contacts extending outside of regulatory borders increased, leading to formation of a larger regulatory unit with a shift from repressive to active compartments (B to A). The 3D genomic structural model proposes that the immediate early promoter region is localized on the periphery of the 3D viral genome during latency, while highly inducible noncoding RNA regions moved toward the inner space of the structure, resembling the configuration of a “bird cage” during reactivation. The compartment-like properties of viral episomal chromatin structure and its reorganization during the transition from latency may help facilitate viral gene transcription.IMPORTANCE The 3D architecture of chromatin allows for efficient arrangement, expression, and replication of genetic material. The genomes of all organisms studied to date have been found to be organized through some form of tiered domain structures. However, the architectural framework of the genomes of large double-stranded DNA viruses such as the herpesvirus family has not been reported. Prior studies with Kaposi’s sarcoma-associated herpesvirus (KSHV) have indicated that the viral chromatin shares many biological properties exhibited by the host cell genome, essentially behaving as a mini human chromosome. Thus, we hypothesized that the KSHV genome may be organized in a similar manner. In this report, we describe the domain structure of the latent and lytic KSHV genome at 500 bp resolution and present a 3D genomic structural model for KSHV under each condition. These results add new insights into the complex regulation of the viral life cycle.

(Invited) Carbon Nanotubes for Plant Genetic Engineering

Plants will benefit from genetic engineering to feed our growing global population in the midst of climate change, to provide our most clinically-relevant drugs, and for our environmental remediation efforts. New generation genome engineering technologies, such as CRISPR/Cas, hold the potential to alter plant genes in a precise and targeted manner. The first step of any plant genetic engineering application is to deliver important biomolecular cargoes into the desired subcellular locations of plant cells. However, the limitations of traditional plant gene delivery methods, i.e. Agrobacterium and gene gun, prevent plant genetic engineering from reaching its full potential.Given the lack of efficient biomolecule delivery vehicles to plants and potential impact of plant biotechnology on food security and sustainability, we developed nanoparticle-based platforms to efficiently deliver small interfering RNA1,2 and plasmid DNA3,4 cargoes into plant cells in a species-independent manner, which eliminates the use of gene gun and addresses the main delivery bottlenecks of plant genetic engineering. We show that pristine single walled carbon nanotubes (SWCNTs) can adsorb small interfering RNA molecules through pi-pi stacking for plant delivery, whereas a covalently modified and positively charged SWCNT platform is more efficient for large double-stranded DNA delivery to mature plant leaf cells.This talk will first briefly summarize the development, characterization and biological verification of each nanoparticle platform for plant delivery applications. Next, I will present our recent results of CRISPR/Cas gene editing in plant leaves using the SWCNT DNA delivery platform5. The talk will be concluded by discussing the several assays performed to demonstrate the effect of nanoparticles on plant health, including an RNA-sequencing analysis of nanoparticle-treated plant leaves6. We believe the SWCNT delivery technologies that are discussed in this talk will have an immense impact on food security and sustainability by enabling the generation of improved plants that can rapidly reach the market in a cost-effective manner. Demirer G.S.*, Zhang H.* et al., DNA Nanostructures Coordinate Gene Silencing in Mature Plants. PNAS (2019). Demirer G.S. et al., Carbon Nanocarriers Deliver siRNA to Intact Plant Cells for Efficient Gene Knockdown. Science Advances (2020). Demirer G.S. et al., High Aspect Ratio Nanomaterials Enable Delivery of Functional Genetic Material Without Transgenic DNA Integration in Mature Plants. Nature Nanotechnology (2019). Demirer G.S. et al., Carbon nanotube–mediated DNA delivery without transgene integration in intact plants. Nature Protocols (2019). Demirer G.S. et al., Nanotechnology to advance CRISPR–Cas genetic engineering of plants. Nature Nanotechnology (2021).González-Grandío E., Demirer, G.S. et al. Carbon nanotube biocompatibility in plants is determined by their surface chemistry. bioRxiv (2021).

Insights on the cGAS-STING Signaling Pathway During Herpesvirus Infections.

Herpesviruses belong to large double-stranded DNA viruses. They are under a wide range of hosts and establish lifelong infection, which creates a burden on human health and animal health. Innate immunity is the host’s innate defense ability. Activating the innate immune signaling pathway and producing type I interferon is the host’s first line of defense against infectious pathogens. Emerging evidence indicates that the cGAS-STING signaling pathway plays an important role in the innate immunity in response to herpesvirus infections. In parallel, because of the constant selective pressure imposed by host immunity, herpesvirus also evolves to target the cGAS-STING signaling pathway to inhibit or escape the innate immune responses. In the current review, we insight on the classical cGAS-STING signaling pathway. We describe the activation of cGAS-STING signaling pathway during herpesvirus infections and strategies of herpesvirus targeting this pathway to evade host antiviral response. Furthermore, we outline the immunotherapy boosting cGAS-STING signaling pathway.

Genomes of six viruses that infect Asgard archaea from deep-sea sediments.

Asgard archaea are globally distributed prokaryotic microorganisms related to eukaryotes; however, viruses that infect these organisms have not been described. Here, using metagenome sequences recovered from deep-sea hydrothermal sediments, we characterize six relatively large (up to 117 kb) double-stranded DNA (dsDNA) viral genomes that infected two Asgard archaeal phyla, Lokiarchaeota and Helarchaeota. These viruses encode Caudovirales-like structural proteins, as well as proteins distinct from those described in known archaeal viruses. Their genomes contain around 1-5% of genes associated with eukaryotic nucleocytoplasmic large DNA viruses (NCLDVs) and appear to be capable of semi-autonomous genome replication, repair, epigenetic modifications and transcriptional regulation. Moreover, Helarchaeota viruses may hijack host ubiquitin systems similar to eukaryotic viruses. Genomic analysis of these Asgard viruses reveals that they contain features of both prokaryotic and eukaryotic viruses, and provides insights into their potential infection and host interaction mechanisms.

Characterization of an alloherpesvirus from wild lake sturgeon Acipenser fulvescens in Wisconsin (USA).

In the spring of 2017, 2 adult lake sturgeon (LS) Acipenser fulvescens captured from the Wolf River, Wisconsin (USA), presented with multiple cutaneous plaques that, upon microscopic examination, indicated proliferative epidermitis. Ultrastructural examination of affected keratinocytes revealed particles in the nucleus having a morphology typical of herpesviruses. A degenerate PCR assay targeting the DNA polymerase catalytic subunit (pol) gene of large double-stranded DNA viruses generated amplicons of the anticipated size from skin samples, and sequences of amplicons confirmed the presence of a novel alloherpesvirus (lake sturgeon herpesvirus, LSHV) related to acipenserid herpesvirus 1 (AciHV1). The complete genome (202660 bp) of this virus was sequenced using a MiSeq System, and phylogenetic analyses substantiated the close relationship to AciHV1. A PCR assay targeting the LSHV DNA packaging terminase subunit 1 (ter1) gene demonstrated the presence of the virus in 39/42 skin lesion samples collected from wild LS captured in 2017-2019 and 2021 in 4/4 rivers in Wisconsin. Future efforts to isolate LSHV in cell culture would facilitate challenge studies to determine the disease potential of the virus.

Early in vivo transcriptome of Trichoplusia ni ascovirus core genes.

Ascoviruses are large double-stranded DNA insect viruses that destroy the nucleus and transform each cell into 20 or more viral vesicles for replication. In the present study we used RNA-sequencing to compare the expression of Trichoplusia ni ascovirus 6a1 (TnAV-6a1) core genes during the first week of infection, with emphasis on the first 48 h, comparing transcript levels in major somatic tissues (epidermis, tracheal matrix and fat body), the sites infected initially, with those of the haemolymph, where viral vesicles circulate and most replication occurs. By 48 h post-infection (p.i.), only 26 genes were expressed in somatic tissues at ≥5 log2 reads per kilobase per million, whereas in the haemolymph 48 genes were expressed at a similar level by the same time. Early and high expression of TnAV caspase-2-like gene occurred in all tissues, implying it is required for replication, but that it is probably not associated with apoptosis induction, which occurs in infections of Spodoptera frugiperda ascovirus 1 a (SfAV-1a), the ascovirus type species. Other highly expressed viral genes at 48 h p.i. in viral vesicles included a dynein-like beta chain and lipid-modifying enzymes, suggesting their importance to vesicle formation and growth as well as virion synthesis. Finally, as occurs in SfAV expression, we found bicistronic and tricistronic mRNA messages produced by TnAV.

Complete Genome Sequence of Emiliania huxleyi Virus Strain M1, Isolated from an Induced E. huxleyi Bloom in Bergen, Norway.

ABSTRACTEmiliania huxleyi virus strain M1 (EhVM1), a large double-stranded DNA virus from the family Phycodnaviridae, was isolated from an Emiliania huxleyi bloom during a mesocosm experiment in Raunefjorden, Bergen, Norway. Here, we report its complete genome, composed of one full contig.

The African swine fever virus protease pS273R inhibits DNA sensing cGAS-STING pathway by targeting IKKε

ABSTRACT African swine fever virus (ASFV), a large and complex cytoplasmic double-stranded DNA virus, has developed multiple strategies to evade the antiviral innate immune responses. Cytosolic DNA arising from invading ASFV is mainly detected by the cyclic GMP-AMP synthase (cGAS) and then triggers a series of innate immune responses to prevent virus invasion. However, the immune escape mechanism of ASFV remains to be fully clarified. The pS273R of ASFV is a member of the SUMO-1-specific protease family and is crucial for valid virus replication. In this study, we identified pS273R as a suppressor of cGAS-STING pathway mediated type I interferon (IFN) production by ASFV genomic open reading frame screening. The pS273R was further confirmed as an inhibitor of IFN production as well as its downstream antiviral genes in cGAS-STING pathway. Mechanistically, pS273R greatly decreased the cGAS-STING signaling by targeting IKKε but not TBK1, and pS273R was found to disturb the interaction between IKKε and STING through its interaction with IKKε. Further, mutational analyses revealed that pS273R antagonized the cGAS-STING pathway by enzyme catalytic activity, which might affect the IKKε sumoylation state required for the interaction with STING. In summary, our results revealed for the first time that pS273R acts as an obvious negative regulator of cGAS-STING pathway by targeting IKKε via its enzymatic activity, which shows a new immune evasion mechanism of ASFV.