Viruses — Open Access Journal

New technologies enable viral discovery in a diversity of hosts, providing insights into viral evolution. We used one such approach, the virome capture sequencing for vertebrate viruses (VirCapSeq-VERT) platform, on 21 samples originating from six dead Maxwell’s duikers (Philantomba maxwellii) from Taï National Park, Côte d’Ivoire. We detected the presence of an orthohepadnavirus in one animal and characterized its 3128 bp genome. The highest viral copy numbers were detected in the spleen, followed by the lung, blood, and liver, with the lowest copy numbers in the kidney and heart; the virus was not detected in the jejunum. Viral copy numbers in the blood were in the range known from humans with active chronic infections leading to liver histolytic damage, suggesting this virus could be pathogenic in duikers, though many orthohepadnaviruses appear to be apathogenic in other hosts, precluding a formal test of this hypothesis. The virus was not detected in 29 other dead duiker samples from the Côte d’Ivoire and Central African Republic, suggesting either a spillover event or a low prevalence in these populations. Phylogenetic analysis placed the virus as a divergent member of the mammalian clade of orthohepadnaviruses, though its relationship to other orthohepadnaviruses remains uncertain. This represents the first orthohepadnavirus described in an artiodactyl. We have tentatively named this new member of the genus Orthohepadnavirus (family Hepadnaviridae), Taï Forest hepadnavirus. Further studies are needed to determine whether it, or some close relatives, are present in a broader range of artiodactyls, including livestock. Full article

We previously determined that virions of Sclerotinia sclerotiorum hypovirulence associated DNA virus 1 (SsHADV-1) could directly infect hyphae of Sclerotinia sclerotiorum, resulting in hypovirulence of the fungal host. However, the molecular mechanisms of SsHADV-1 virions disruption of the fungal cell wall barrier and entrance into the host cell are still unclear. To investigate the early response of S. sclerotiorum to SsHADV-1 infection, S. sclerotiorum hyphae were inoculated with purified SsHADV-1 virions. The pre- and post-infection hyphae were collected at one–three hours post-inoculation for transcriptome analysis. Further, bioinformatic analysis showed that differentially expressed genes (DEGs) regulated by SsHADV-1 infection were identified in S. sclerotiorum. In total, 187 genes were differentially expressed, consisting of more up-regulated (114) than down-regulated (73) genes. The identified DEGs were involved in several important pathways. Metabolic processes, biosynthesis of antibiotics, and secondary metabolites were the most affected categories in S. sclerotiorum upon SsHADV-1 infection. Cell structure analysis suggested that 26% of the total DEGs were related to membrane tissues. Furthermore, 10 and 27 DEGs were predicted to be located in the cell membrane and mitochondria, respectively. Gene ontology enrichment analyses of the DEGs were performed, followed by functional annotation of the genes. Interestingly, one third of the annotated functional DEGs could be involved in the Ras-small G protein signal transduction pathway. These results revealed that SsHADV-1 virions may be able to bind host membrane proteins and influence signal transduction through Ras-small G protein-coupled receptors during early infection, providing new insight towards the molecular mechanisms of virions infection in S. sclerotiorum. Full article

In agriculture, the prevention and treatment of bacterial infections represents an increasing challenge. Traditional (chemical) methods have been restricted to ensure public health and to limit the occurrence of resistant strains. Bacteriophages could be a sustainable alternative. A major hurdle towards the commercial implementation of phage-based biocontrol strategies concerns aspects of regulation and intellectual property protection. Within this study, two datasets have been composed to analyze both scientific publications and patent documents and to get an idea on the focus of research and development (R&D) by means of an abstract and claim analysis. A total of 137 papers and 49 patent families were found from searching public databases, with their numbers increasing over time. Within this dataset, the majority of the patent documents were filed by non-profit organizations in Asia. There seems to be a good correlation between the papers and patent documents in terms of targeted bacterial genera. Furthermore, granted patents seem to claim rather broad and cover methods of treatment. This review shows that there is indeed growing publishing and patenting activity concerning phage biocontrol. Targeted research is needed to further stimulate the exploration of phages within integrated pest management strategies and to deal with bacterial infections in crop production. Full article

The characterization of the function of conserved viral genes is central to developing a greater understanding of important aspects of viral replication or pathogenesis. A comparative genomic analysis of the iridoviral genomes identified 26 core genes conserved across the family Iridoviridae. Three of those conserved genes have no defined function; these include the homologs of frog virus 3 (FV3) open reading frames (ORFs) 88R, 91R, and 94L. Conserved viral genes that have been previously identified are known to participate in a number of viral activities including: transcriptional regulation, DNA replication/repair/modification/processing, protein modification, and viral structural proteins. To begin to characterize the conserved FV3 ORFs 88R, 91R, and 94L, we cloned the genes and determined their intracellular localization. We demonstrated that 88R localizes to the cytoplasm of the cell while 91R localizes to the nucleus and 94L localizes to the endoplasmic reticulum (ER). Full article

Filoviruses infect a wide range of cell types with the exception of lymphocytes. The intracellular proteins cathepsin B and L, two-pore channel 1 and 2, and bona fide receptor Niemann–Pick Disease C1 (NPC1) are essential for the endosomal phase of cell entry. However, earlier steps of filoviral infection remain poorly characterized. Numerous plasma membrane proteins have been implicated in attachment but it is still unclear which ones are sufficient for productive entry. To define a minimal set of host factors required for filoviral glycoprotein-driven cell entry, we screened twelve cell lines and identified the nonlymphocytic cell line SH-SY5Y to be specifically resistant to filovirus infection. Heterokaryons of SH-SY5Y cells fused to susceptible cells were susceptible to filoviruses, indicating that SH-SY5Y cells do not express a restriction factor but lack an enabling factor critical for filovirus entry. However, all tested cell lines expressed functional intracellular factors. Global gene expression profiling of known cell surface entry factors and protein expression levels of analyzed attachment factors did not reveal any correlation between susceptibility and expression of a specific host factor. Using binding assays with recombinant filovirus glycoprotein, we identified cell attachment as the step impaired in filovirus entry in SH-SY5Y cells. Individual overexpression of attachment factors T-cell immunoglobulin and mucin domain 1 (TIM-1), Axl, Mer, or dendritic cell-specific intercellular adhesion molecule-3-grabbing non-integrin (DC-SIGN) rendered SH-SY5Y cells susceptible to filovirus glycoprotein-driven transduction. Our study reveals that a lack of attachment factors limits filovirus entry and provides direct experimental support for a model of filoviral cell attachment where host factor usage at the cell surface is highly promiscuous. Full article

Ebola Virus Disease (EVD) is one of the most lethal transmissible infections, characterized by a high fatality rate, and caused by a member of the Filoviridae family. The recent large outbreak of EVD in Western Africa (2013–2016) highlighted the worldwide threat represented by the disease and its impact on global public health and the economy. The development of highly needed anti-Ebola virus antivirals has been so far hampered by the shortage of tools to study their life cycle in vitro, allowing to screen for potential active compounds outside a biosafety level-4 (BSL-4) containment. Importantly, the development of surrogate models to study Ebola virus entry in a BSL-2 setting, such as viral pseudotypes and Ebola virus-like particles, tremendously boosted both our knowledge of the viral life cycle and the identification of promising antiviral compounds interfering with viral entry. In this context, the combination of such surrogate systems with large-scale small molecule compounds and haploid genetic screenings, as well as rational drug design and drug repurposing approaches will prove priceless in our quest for the development of a treatment for EVD. Full article

Viruses are parasites that depend on the host cell’s metabolic resources to provide the energy and molecular building blocks necessary for the production of viral progeny. It has become increasingly clear that viruses extensively modulate the cellular metabolic network to support productive infection. Here, we review the numerous ways through which human cytomegalovirus (HCMV) modulates cellular metabolism, highlighting known mechanisms of HCMV-mediated metabolic manipulation and identifying key outstanding questions that remain to be addressed. Full article

Infection is restrained by the concerted activation of tissue-resident and circulating immune cells. Recent discoveries have demonstrated that tissue-resident lymphocyte subsets, comprised of innate lymphoid cells (ILCs) and unconventional T cells, have vital roles in the initiation of primary antiviral responses. Via direct and indirect mechanisms, ILCs and unconventional T cell subsets play a critical role in the ability of the immune system to mount an effective antiviral response through potent early cytokine production. In this review, we will summarize the current knowledge of tissue-resident lymphocytes during initial viral infection and evaluate their redundant or nonredundant contributions to host protection or virus-induced pathology. Full article

Small RNA mediated responses are essential for antiviral defence in mosquitoes, however, they appear to differ per virus-vector combination. To further investigate the diversity of small RNA responses against viruses in mosquitoes, we applied a small RNA deep sequencing approach on five mosquito cell lines: Culex tarsalis CT cells, Aedes albopictus U4.4 and C6/36 cells, Ae. aegypti Aag2 cells (cleared from cell fusing agent virus and Culex Y virus (CYV) by repetitive dsRNA transfections) and Ae. pseudoscutellaris AP-61 cells. De novo assembly of small RNAs revealed the presence of Phasi Charoen-like virus (PCLV), Calbertado virus, Flock House virus and a novel narnavirus in CT cells, CYV in U4.4 cells, and PCLV in Aag2 cells, whereas no insect-specific viruses (ISVs) were detected in C6/36 and AP-61 cells. Next, we investigated the small RNA responses to the identified ISVs and to acute infection with the arthropod-borne West Nile virus (WNV). We demonstrate that AP-61 and C6/36 cells do not produce siRNAs to WNV infection, suggesting that AP-61, like C6/36, are Dicer-2 deficient. CT cells produced a strong siRNA response to the persistent ISVs and acute WNV infection. Interestingly, CT cells also produced viral PIWI-interacting (pi)RNAs to PCLV, but not to WNV or any of the other ISVs. In contrast, in U4.4 and Aag2 cells, WNV siRNAs, and pi-like RNAs without typical ping-pong piRNA signature were observed, while this signature was present in PCLV piRNAs in Aag2 cells. Together, our results demonstrate that mosquito small RNA responses are strongly dependent on both the mosquito cell type and/or the mosquito species and family of the infecting virus. Full article

The compartmentalization of small ruminant lentiviruses (SRLVs) subtype A17 was analyzed in colostrum and peripheral blood leukocyte cells of three naturally infected goats. This study aimed to analyze heterogeneity of the SRLV env (V4V5) gene, which encodes neutralizing epitopes of SU glycoprotein, the gag gene encoding capsid protein (CA), and LTR, a noncoding region, responsible for determination of cell tropism. Compartmentalization was assessed using six established tree or distance-based methods, including permutation test to determine statistical significance. We found statistical evidence of compartmentalization between blood and colostrum in all infected goats although phylogenetic evidence of such compartmentalization was not obvious. Our study demonstrated that compartmentalization is not exclusively specific to the env gene, as we revealed that gag and LTR sequences are also compartmentalized between blood and colostrum. The work also confirms the combined use of different methods as essential for reliable determination of intrahost viral compartmentalization. Identifying and characterizing distinct viral subpopulations and the genetic evolution of SRLV in specific anatomical sites enhances our overall understanding of SRLV pathogenesis, immune control, and particularly virus transmission. Full article

There is broad scientific and societal consensus that finding a cure for HIV infection must be pursued. The major barrier to achieving a cure for HIV/AIDS is the capacity of the HIV virus to avoid both immune surveillance and current antiretroviral therapy (ART) by rapidly establishing latently infected cell populations, termed latent reservoirs. Here, we provide an overview of the rapidly evolving field of HIV cure/remission research, highlighting recent progress and ongoing challenges in the understanding of HIV reservoirs, the role of HIV transcription in latency and immune evasion. We review the major approaches towards a cure that are currently being explored and further argue that small molecules that inhibit HIV transcription, and therefore uncouple HIV gene expression from signals sent by the host immune response, might be a particularly promising approach to attain a cure or remission. We emphasize that a better understanding of the game of “cat and mouse” between the host immune system and the HIV virus is a crucial knowledge gap to be filled in both cure and vaccine research. Full article

Despite the successful use of antibacterials, the emergence of multidrug-resistant bacteria has become a serious threat to global healthcare. In this era of antibacterial crisis, bacteriophages (phages) are being explored as an antibacterial treatment option since they possess a number of advantages over conventional antibacterials, especially in terms of specificity and biosafety; phages specifically lyse target bacteria while not affecting normal and/or beneficial bacteria and display little or no toxicity in that they are mainly composed of proteins and nucleic acids, which consequently significantly reduces the time and cost involved in antibacterial development. However, these benefits also create potential issues regarding antibacterial spectra and host immunity; the antibacterial spectra being very narrow when compared to those of chemicals, with the phage materials making it possible to trigger host immune responses, which ultimately disarm antibacterial efficacy upon successive treatments. In addition, phages play a major role in horizontal gene transfer between bacterial populations, which poses serious concerns for the potential of disastrous consequences regarding antibiotic resistance. Fortunately, however, recent advancements in synthetic biology tools and the speedy development of phage genome resources have allowed for research on methods to circumvent the potentially disadvantageous aspects of phages. These novel developments empower research which goes far beyond traditional phage therapy approaches, opening up a new chapter for phage applications with new antibacterial platforms. Herein, we not only highlight the most recent synthetic phage engineering and phage product engineering studies, but also discuss a new proof-of-concept for phage-inspired antibacterial design based on the studies undertaken by our group. Full article

The detailed mechanisms of replication initiation, termination and segregation events were not yet known in Acanthamoeba polyphaga mimivirus (APMV). Here, we show detailed bioinformatics-based analyses of chromosomal replication in APMV from initiation to termination mediated by proteins bound to specific DNA sequences. Using GC/AT skew and coding sequence skew analysis, we estimated that the replication origin is located at 382 kb in the APMV genome. We performed homology-modeling analysis of the gamma domain of APMV-FtsK (DNA translocase coordinating chromosome segregation) related to FtsK-orienting polar sequences (KOPS) binding, suggesting that there was an insertion in the gamma domain which maintains the structure of the DNA binding motif. Furthermore, UvrD/Rep-like helicase in APMV was homologous to Bacillus subtilis AddA, while the chi-like quartet sequence 5′-CCGC-3′ was frequently found in the estimated ori region, suggesting that chromosomal replication of APMV is initiated via chi-like sequence recognition by UvrD/Rep-like helicase. Therefore, the replication initiation, termination and segregation of APMV are presumably mediated by DNA repair machineries derived from gram-positive bacteria. Moreover, the other frequently observed quartet sequence 5′-CGGC-3′ in the ori region was homologous to the mitochondrial signal sequence of replication initiation, while the comparison of quartet sequence composition in APMV/Rickettsia-genome showed significantly similar values, suggesting that APMV also conserves the mitochondrial replication system acquired from an ancestral genome of mitochondria during eukaryogenesis. Full article

A double-stranded RNA (dsRNA) virus was isolated and characterized from strain EW220 of the phytopathogenic fungus Botryosphaeria dothidea. The full-length cDNAs of the dsRNAs were 6434 bp and 5986 bp in size, respectively. The largest dsRNA encodes a cap-pol fusion protein that contains a coat protein gene and an RNA-dependent RNA polymerase (RdRp) domain, and the second dsRNA encodes a hypothetical protein. Genome sequence analysis revealed that the sequences of the dsRNA virus shared 99% identity with Bipolaris maydis botybirnavirus 1(BmBRV1) isolated from the causal agent of corn southern leaf blight, Bipolaris maydis. Hence, the dsRNA virus constitutes a new strain of BmBRV1 and was named Bipolaris maydis botybirnavirus 1 strain BdEW220 (BmBRV1-BdEW220). BmBRV1-BdEW220 contains spherical virions that are 37 nm in diameter and consist of two dsRNA segments. The structural proteins of the BmBRV1-BdEW220 virus particles were 110 kDa, 90 kDa, and 80 kDa and were encoded by dsRNA1 and 2-ORFs. Phylogenetic reconstruction indicated that BmBRV1 and BmBRV1-BdEW220 are phylogenetically related to the genus Botybirnavirus. Importantly, BmBRV1-BdEW220 influences the growth of B. dothidea and confers hypovirulence to the fungal host. To our knowledge, this is the first report of a botybirnavirus in B. dothidea. Full article

There is a growing interest in phage therapy as a complementary tool against antimicrobial resistant infections. Since 2007, phages have been used sporadically to treat bacterial infections in well-defined cases in the Queen Astrid military hospital (QAMH) in Brussels, Belgium. In the last two years, external requests for phage therapy have increased significantly. From April 2013 to April 2018, 260 phage therapy requests were addressed to the QAMH. Of these 260 requests, only 15 patients received phage therapy. In this paper, we analyze the phage therapy requests and outcomes in order to improve upon the overall capacity for phage therapy at the QAMH. Full article

Marek’s disease virus (MDV) is an oncogenic alphaherpesvirus that infects chickens and poses a serious threat to poultry health. In infected animals, MDV efficiently replicates in B cells in various lymphoid organs. Despite many years of research, the viral transcriptome in primary target cells of MDV remained unknown. In this study, we uncovered the transcriptional landscape of the very virulent RB1B strain and the attenuated CVI988/Rispens vaccine strain in primary chicken B cells using high-throughput RNA-sequencing. Our data confirmed the expression of known genes, but also identified a novel spliced MDV gene in the unique short region of the genome. Furthermore, de novo transcriptome assembly revealed extensive splicing of viral genes resulting in coding and non-coding RNA transcripts. A novel splicing isoform of MDV UL15 could also be confirmed by mass spectrometry and RT-PCR. In addition, we could demonstrate that the associated transcriptional motifs are highly conserved and closely resembled those of the host transcriptional machinery. Taken together, our data allow a comprehensive re-annotation of the MDV genome with novel genes and splice variants that could be targeted in further research on MDV replication and tumorigenesis. Full article

Backgrounds and Aims: We previously demonstrated that serum hepatitis B virus (HBV) DNA in HBV infected humanized mice exhibited a highly dynamic multiphasic kinetic pattern from infection initiation to steady-state. Here, we investigated whether this pattern is consistent across different HBV clones or in the presence of hepatitis D virus (HDV) co-infection. Methods: We analyzed early serum viral kinetics using 26 HBV genotype C (GtC) mono-infected mice [clones: PXB, Hiroshima GtC CL4 (CL4) and Hiroshima GtC CL5 (CL5)] and four HBV CL4/HDV genotype one co-infected mice. Results: The HBV kinetics observed with clones CL4 and CL5 were similar to that previously defined in HBV PXB infected mice. Additionally, no significant differences in HBV DNA levels were observed between HBV mono-infected and HBV/HDV co-infected mice through 4 weeks post-inoculation (p.i.). However, HBV DNA levels at 6 weeks p.i. in HBV/HDV co-infected mice were significantly lower than those in HBV mono-infected mice (P = 0.002), consistent with HDV suppression of chronic HBV. Conclusions: HBV infection initiation is multiphasic across multiple viral clones and is not altered by HDV co-infection. The latter suggests that higher HDV titers (>8 log IU/mL) and/or longer duration of HDV infection might be needed to trigger HDV-induced suppression on HBV. Full article

Serotype 4 fowl adenovirus (FAdV-4) is the main pathogen for hydropericardium syndrome (HPS) in chickens. It has caused major economic losses in the global poultry industry. Currently, FAdV-4′s transmission routes in chickens remain unclear. Here we investigate the airborne transmission routes of FAdV-4 in chickens. A total of 45 ten-day-old chickens were equally divided into three groups (infected group/isolator A, airborne group/isolator B, and control group/isolator C). Of note, isolators A and B were connected by a leak-free pipe. The results showed that the virus could form a viral aerosol, detected in isolators two days post infection (dpi). The viral aerosol reached a peak at 4 dpi in the infected group. Healthy chickens in the airborne group were infected by the virus at 8 dpi. The chickens of the airborne group demonstrated subclinical symptoms capable of shedding the virus for some time. This finding suggests that FAdV-4 can be efficiently transmitted among chickens by aerosol transmission. These findings have significant implications for developing strategies to control this infectious disease epidemic. Full article

Increasing evidence suggests that neurodegenerative disorders share a common pathogenic feature: the presence of deposits of misfolded proteins with altered physicochemical properties in the Central Nervous System. Despite a lack of infectivity, experimental data show that the replication and propagation of neurodegenerative disease-related proteins including amyloid-β (Aβ), tau, α-synuclein and the transactive response DNA-binding protein of 43 kDa (TDP-43) share a similar pathological mechanism with prions. These observations have led to the terminology of “prion-like” to distinguish between conditions with noninfectious characteristics but similarities with the prion replication and propagation process. Prions are considered to adapt their conformation to changes in the context of the environment of replication. This process is known as either prion selection or adaptation, where a distinct conformer present in the initial prion population with higher propensity to propagate in the new environment is able to prevail over the others during the replication process. In the last years, many studies have shown that prion-like proteins share not only the prion replication paradigm but also the specific ability to aggregate in different conformations, i.e., strains, with relevant clinical, diagnostic and therapeutic implications. This review focuses on the molecular basis of the strain phenomenon in prion and prion-like proteins. Full article