Viruses, Volume 7, Issue 9 (September 2015) – 18 articles

Hepatitis C virus (HCV) infection is one of the leading causes of end-stage liver disease and the main indication for liver transplantation (LT) in most countries. All patients who undergo LT with detectable serum HCV RNA experience graft reinfection progressing to cirrhosis within five years in 20% to 30% of them. Obtaining a sustained virological response (SVR) greatly improves overall and graft survival. Until 2011, standard antiviral therapy using PEGylated interferon (PEG-IFN) and ribavirin (RBV) was the only effective therapy, with an SVR rate around 30% in this setting. For patients infected with genotype 1, first generation NS3/4A protease inhibitors (PIs), boceprevir (BOC) or telaprevir (TVR), associated with PEG-IFN and RBV for 48 weeks have increased the SVR rates to 60% in non-transplant patients. However, tolerability and drug-drug interactions with calcineurin inhibitors (CNI) are both limiting factors of their use in the liver transplant setting. Over recent years, the efficacy of antiviral C therapy has improved dramatically using new direct-acting antiviral (DAA) agents without PEG-IFN and/or RBV, leading to SVR rates over 90% in non-transplant patients. Results available for transplant patients showed a better efficacy and tolerability and less drug-drug interactions than with first wave PIs. However, some infrequent cases of viral resistance have been reported using PIs or NS5A inhibitors pre- or post-LT that can lead to difficulties in the management of these patients. Full article

Tight junctions (TJs) are highly specialized membrane domains involved in many important cellular processes such as the regulation of the passage of ions and macromolecules across the paracellular space and the establishment of cell polarity in epithelial cells. Over the past few years there has been increasing evidence that different components of the TJs can be hijacked by viruses in order to complete their infectious cycle. Viruses from at least nine different families of DNA and RNA viruses have been reported to use TJ proteins in their benefit. For example, TJ proteins such as JAM-A or some members of the claudin family of proteins are used by members of the Reoviridae family and hepatitis C virus as receptors or co-receptors during their entry into their host cells. Reovirus, in addition, takes advantage of the TJ protein Junction Adhesion Molecule-A (JAM-A) to achieve its hematogenous dissemination. Some other viruses are capable of regulating the expression or the localization of TJ proteins to induce cell transformation or to improve the efficiency of their exit process. This review encompasses the importance of TJs for viral entry, replication, dissemination, and egress, and makes a clear statement of the importance of studying these proteins to gain a better understanding of the replication strategies used by viruses that infect epithelial and/or endothelial cells. Full article

The antiviral effect of a catalytic RNA-hydrolyzing antibody, 3D8 scFv, for intranasal administration against avian influenza virus (H1N1) was described. The recombinant 3D8 scFv protein prevented BALB/c mice against H1N1 influenza virus infection by degradation of the viral RNA genome through its intrinsic RNA-hydrolyzing activity. Intranasal administration of 3D8 scFv (50 μg/day) for five days prior to infection demonstrated an antiviral activity (70% survival) against H1N1 infection. The antiviral ability of 3D8 scFv to penetrate into epithelial cells from bronchial cavity via the respiratory mucosal layer was confirmed by immunohistochemistry, qRT-PCR, and histopathological examination. The antiviral activity of 3D8 scFv against H1N1 virus infection was not due to host immune cytokines or chemokines, but rather to direct antiviral RNA-hydrolyzing activity of 3D8 scFv against the viral RNA genome. Taken together, our results suggest that the RNase activity of 3D8 scFv, coupled with its ability to penetrate epithelial cells through the respiratory mucosal layer, directly prevents H1N1 virus infection in a mouse model system. Full article

The mechanism of antibody-mediated protection is a major focus of HIV-1 vaccine development and a significant issue in the control of viremia. Virus neutralization, Fc-mediated effector function, or both, are major mechanisms of antibody-mediated protection against HIV-1, although other mechanisms, such as virus aggregation, are known. The interplay between virus neutralization and Fc-mediated effector function in protection against HIV-1 is complex and only partially understood. Passive immunization studies using potent broadly neutralizing antibodies (bnAbs) show that both neutralization and Fc-mediated effector function provides the widest dynamic range of protection; however, a vaccine to elicit these responses remains elusive. By contrast, active immunization studies in both humans and non-human primates using HIV-1 vaccine candidates suggest that weakly neutralizing or non-neutralizing antibodies can protect by Fc-mediated effector function, albeit with a much lower dynamic range seen for passive immunization with bnAbs. HIV-1 has evolved mechanisms to evade each type of antibody-mediated protection that must be countered by a successful AIDS vaccine. Overcoming the hurdles required to elicit bnAbs has become a major focus of HIV-1 vaccine development. Here, we discuss a less studied problem, the structural basis of protection (and its evasion) by antibodies that protect only by potent Fc-mediated effector function. Full article

Alphaherpesviruses like herpes simplex virus are large DNA viruses characterized by their ability to establish lifelong latent infection in neurons. As for all herpesviruses, alphaherpesvirus virions contain a protein-rich layer called “tegument” that links the DNA-containing capsid to the glycoprotein-studded membrane envelope. Tegument proteins mediate a diverse range of functions during the virus lifecycle, including modulation of the host-cell environment immediately after entry, transport of virus capsids to the nucleus during infection, and wrapping of cytoplasmic capsids with membranes (secondary envelopment) during virion assembly. Eleven tegument proteins that are conserved across alphaherpesviruses have been implicated in the formation of the tegument layer or in secondary envelopment. Tegument is assembled via a dense network of interactions between tegument proteins, with the redundancy of these interactions making it challenging to determine the precise function of any specific tegument protein. However, recent studies have made great headway in defining the interactions between tegument proteins, conserved across alphaherpesviruses, which facilitate tegument assembly and secondary envelopment. We summarize these recent advances and review what remains to be learned about the molecular interactions required to assemble mature alphaherpesvirus virions following the release of capsids from infected cell nuclei. Full article

Exosomes are extracellular vesicles released upon fusion of multivesicular bodies(MVBs) with the cellular plasma membrane. They originate as intraluminal vesicles (ILVs) duringthe process of MVB formation. Exosomes were shown to contain selectively sorted functionalproteins, lipids, and RNAs, mediating cell-to-cell communications and hence playing a role in thephysiology of the healthy and diseased organism. Challenges in the field include the identificationof mechanisms sustaining packaging of membrane-bound and soluble material to these vesicles andthe understanding of the underlying processes directing MVBs for degradation or fusion with theplasma membrane. The investigation into the formation and roles of exosomes in viral infection is inits early years. Although still controversial, exosomes can, in principle, incorporate any functionalfactor, provided they have an appropriate sorting signal, and thus are prone to viral exploitation.This review initially focuses on the composition and biogenesis of exosomes. It then explores theregulatory mechanisms underlying their biogenesis. Exosomes are part of the endocytic system,which is tightly regulated and able to respond to several stimuli that lead to alterations in thecomposition of its sub-compartments. We discuss the current knowledge of how these changesaffect exosomal release. We then summarize how different viruses exploit specific proteins ofendocytic sub-compartments and speculate that it could interfere with exosome function, althoughno direct link between viral usage of the endocytic system and exosome release has yet beenreported. Many recent reports have ascribed functions to exosomes released from cells infectedwith a variety of animal viruses, including viral spread, host immunity, and manipulation of themicroenvironment, which are discussed. Given the ever-growing roles and importance of exosomesin viral infections, understanding what regulates their composition and levels, and defining theirfunctions will ultimately provide additional insights into the virulence and persistence of infections. Full article

The rise in human papillomavirus (HPV)-associated head and neck squamous cell carcinoma (HNSCC) has elicited significant interest in the role of high-risk HPV in tumorigenesis. Because patients with HPV-positive HNSCC have better prognoses than do their HPV-negative counterparts, current therapeutic strategies for HPV⁺ HNSCC are increasingly considered to be overly aggressive, highlighting a need for customized treatment guidelines for this cohort. Additional issues include the unmet need for a reliable screening strategy for HNSCC, as well as the ongoing assessment of the efficacy of prophylactic vaccines for the prevention of HPV infections in the head and neck regions. This review also outlines a number of emerging prospects for therapeutic vaccines, as well as for targeted, molecular-based therapies for HPV-associated head and neck cancers. Overall, the future for developing novel and effective therapeutic agents for HPV-associated head and neck tumors is promising; continued progress is critical in order to meet the challenges posed by the growing epidemic. Full article

Treatment with pan-genotypic direct-acting antivirals, targeting different viral proteins, is the best option for clearing hepatitis C virus (HCV) infection in chronically infected patients. However, the diversity of the HCV genome is a major obstacle for the development of antiviral drugs, vaccines, and genotyping assays. In this large-scale analysis, genome-wide diversity and selective pressure was mapped, focusing on positions important for treatment, drug resistance, and resistance testing. A dataset of 1415 full-genome sequences, including genotypes 1–6 from the Los Alamos database, was analyzed. In 44% of all full-genome positions, the consensus amino acid was different for at least one genotype. Focusing on positions sharing the same consensus amino acid in all genotypes revealed that only 15% was defined as pan-genotypic highly conserved (≥99% amino acid identity) and an additional 24% as pan-genotypic conserved (≥95%). Despite its large genetic diversity, across all genotypes, codon positions were rarely identified to be positively selected (0.23%–0.46%) and predominantly found to be under negative selective pressure, suggesting mainly neutral evolution. For NS3, NS5A, and NS5B, respectively, 40% (6/15), 33% (3/9), and 14% (2/14) of the resistance-related positions harbored as consensus the amino acid variant related to resistance, potentially impeding treatment. For example, the NS3 variant 80K, conferring resistance to simeprevir used for treatment of HCV1 infected patients, was present in 39.3% of the HCV1a strains and 0.25% of HCV1b strains. Both NS5A variants 28M and 30S, known to be associated with resistance to the pan-genotypic drug daclatasvir, were found in a significant proportion of HCV4 strains (10.7%). NS5B variant 556G, known to confer resistance to non-nucleoside inhibitor dasabuvir, was observed in 8.4% of the HCV1b strains. Given the large HCV genetic diversity, sequencing efforts for resistance testing purposes may need to be genotype-specific or geographically tailored. Full article

The emergence of HIV-1 groups M, N, O, and P is the result of four independent cross-species transmissions between chimpanzees (cpz) and gorillas (gor) from central/south Cameroon and humans respectively. Although the first two SIVcpz were identified in wild-born captive chimpanzees in Gabon in 1989, no study has been conducted so far in wild chimpanzees in Gabon. To document the SIVcpz infection rate, genetic diversity, and routes of virus transmission, we analyzed 1458 faecal samples collected in 16 different locations across the country, and we conducted follow-up missions in two of them. We found 380 SIV antibody positive samples in 6 different locations in the north and northeast. We determined the number of individuals collected by microsatellite analysis and obtained an adjusted SIV prevalence of 39.45%. We performed parental analysis to investigate viral spread between and within communities and found that SIVs were epidemiologically linked and were transmitted by both horizontal and vertical routes. We amplified pol and gp41 fragments and obtained 57 new SIVcpzPtt strains from three sites. All strains, but one, clustered together within a specific phylogeographic clade. Given that these SIV positive samples have been collected nearby villages and that humans continue to encroach in ape’s territories, the emergence of a new HIV in this area needs to be considered. Full article

Viral interactions with host nucleus have been thoroughly studied, clarifying molecular mechanisms and providing new antiviral targets. Considering that African swine fever virus (ASFV) intranuclear phase of infection is poorly understood, viral interplay with subnuclear domains and chromatin architecture were addressed. Nuclear speckles, Cajal bodies, and promyelocytic leukaemia nuclear bodies (PML-NBs) were evaluated by immunofluorescence microscopy and Western blot. Further, efficient PML protein knockdown by shRNA lentiviral transduction was used to determine PML-NBs relevance during infection. Nuclear distribution of different histone H3 methylation marks at lysine’s 9, 27 and 36, heterochromatin protein 1 isoforms (HP1α, HPβ and HPγ) and several histone deacetylases (HDACs) were also evaluated to assess chromatin status of the host. Our results reveal morphological disruption of all studied subnuclear domains and severe reduction of viral progeny in PML-knockdown cells. ASFV promotes H3K9me3 and HP1β foci formation from early infection, followed by HP1α and HDAC2 nuclear enrichment, suggesting heterochromatinization of host genome. Finally, closeness between DNA damage response factors, disrupted PML-NBs, and virus-induced heterochromatic regions were identified. In sum, our results demonstrate that ASFV orchestrates spatio-temporal nuclear rearrangements, changing subnuclear domains, relocating Ataxia Telangiectasia Mutated Rad-3 related (ATR)-related factors and promoting heterochromatinization, probably controlling transcription, repressing host gene expression, and favouring viral replication. Full article

Approximately 240 million people worldwide are chronically infected with hepatitis B virus (HBV), which represents a significant challenge to public health. The current goal in treating chronic HBV infection is to block progression of HBV-related liver injury and inflammation to end-stage liver diseases, including cirrhosis and hepatocellular carcinoma, because we are unable to eliminate chronic HBV infection. Available therapies for chronic HBV infection mainly include nucleos/tide analogues (NAs), non-NAs, and immunomodulatory agents. However, none of them is able to clear chronic HBV infection. Thus, a new generation of anti-HBV drugs is urgently needed. Progress has been made in the development and testing of new therapeutics against chronic HBV infection. This review aims to summarize the state of the art in new HBV drug research and development and to forecast research and development trends and directions in the near future. Full article

Tomato yellow leaf curl China virus (TYLCCNV) is a monopartite begomovirus associated with different betasatellites. In this study, we investigate two different isolates of Tomato yellow leaf curl China betasatellite (TYLCCNB) to determine what features of the viral genome are required for induction of characteristic phenotypic differences between closely-related betasatellite. When co-agroinoculated with TYLCCNV into Nicotiana spp. and tomato plants, TYLCCNB-Y25 induced only leaf curling on all hosts, while TYLCCNB-Y10 also induced enations, vein yellowing, and shoot distortions. Further assays showed that βC1 of TYLCCNB-Y25 differs from that of TYLCCNB-Y10 in symptom induction and transcriptional modulating. Hybrid satellites were constructed in which the βC1 gene or 200 nt partial promoter-like fragment upstream of the βC1 were exchanged. Infectivity assays showed that a TYLCCNB-Y25 hybrid with the intact TYLCCNB-Y10 βC1 gene was able to induce vein yellowing, shoot distortions, and a reduced size and number of enations. A TYLCCNB-Y10 hybrid with the intact TYLCCNB-Y25 βC1 gene produced only leaf curling. In contrast, the TYLCCNB-Y25 and TYLCCNB-Y10 hybrids with swapped partial promoter-like regions had little effect on the phenotypes induced by wild-type betasatellites. Further experiments showed that the TYLCCNB-Y25 hybrid carrying the C-terminal region of TYLCCNB-Y10 βC1 induced TYLCCNB-Y10-like symptoms. These findings indicate that the βC1 protein is the major symptom determinant and that the C-terminal region of βC1 plays an important role in symptom induction. Full article

Influenza is a major cause of severe respiratory infections leading to excessive hospitalizations and deaths globally; annual epidemics, pandemics, and sporadic/endemic avian virus infections occur as a result of rapid, continuous evolution of influenza viruses. Emergence of antiviral resistance is of great clinical and public health concern. Currently available antiviral treatments include four neuraminidase inhibitors (oseltamivir, zanamivir, peramivir, laninamivir), M2-inibitors (amantadine, rimantadine), and a polymerase inhibitor (favipiravir). In this review, we focus on resistance issues related to the use of neuraminidase inhibitors (NAIs). Data on primary resistance, as well as secondary resistance related to NAI exposure will be presented. Their clinical implications, detection, and novel therapeutic options undergoing clinical trials are discussed. Full article

Arthropod-borne viruses (arboviruses), especially those transmitted by mosquitoes, are a significant cause of morbidity and mortality in humans and animals worldwide. Recent discoveries indicate that mosquitoes are naturally infected with a wide range of other viruses, many within taxa occupied by arboviruses that are considered insect-specific. Over the past ten years there has been a dramatic increase in the literature describing novel insect-specific virus detection in mosquitoes, which has provided new insights about viral diversity and evolution, including that of arboviruses. It has also raised questions about what effects the mosquito virome has on arbovirus transmission. Additionally, the discovery of these new viruses has generated interest in their potential use as biological control agents as well as novel vaccine platforms. The arbovirus community will benefit from the growing database of knowledge concerning these newly described viral endosymbionts, as their impacts will likely be far reaching. Full article

Dengue is the most widespread arbovirus infection and poses a serious health and economic issue in tropical and subtropical countries. Currently no licensed vaccine or compounds can be used to prevent or manage the severity of dengue virus (DENV) infection. Honokiol, a lignan biphenol derived from the Magnolia tree, is commonly used in Eastern medicine. Here we report that honokiol has profound antiviral activity against serotype 2 DENV (DENV-2). In addition to inhibiting the intracellular DENV-2 replicon, honokiol was shown to suppress the replication of DENV-2 in baby hamster kidney (BHK) and human hepatocarcinoma Huh7 cells. At the maximum non-toxic dose of honokiol treatment, the production of infectious DENV particles was reduced >90% in BHK and Huh7 cells. The underlying mechanisms revealed that the expression of DENV-2 nonstructural protein NS1/NS3 and its replicating intermediate, double-strand RNA, was dramatically reduced by honokiol treatment. Honokiol has no effect on the expression of DENV putative receptors, but may interfere with the endocytosis of DENV-2 by abrogating the co-localization of DENV envelope glycoprotein and the early endosomes. These results indicate that honokiol inhibits the replication, viral gene expression, and endocytotic process of DENV-2, making it a promising agent for chemotherapy of DENV infection. Full article

Poxviruses encode a broad array of proteins that serve to undermine host immune defenses. Structural analysis of four of these seemingly unrelated proteins revealed the recurrent use of a conserved beta-sandwich fold that has not been observed in any eukaryotic or prokaryotic protein. Herein we propose to call this unique structural scaffolding the PIE (Poxvirus Immune Evasion) domain. PIE domain containing proteins are abundant in chordopoxvirinae, with our analysis identifying 20 likely PIE subfamilies among 33 representative genomes spanning 7 genera. For example, cowpox strain Brighton Red appears to encode 10 different PIEs: vCCI, A41, C8, M2, T4 (CPVX203), and the SECRET proteins CrmB, CrmD, SCP-1, SCP-2, and SCP-3. Characterized PIE proteins all appear to be nonessential for virus replication, and all contain signal peptides for targeting to the secretory pathway. The PIE subfamilies differ primarily in the number, size, and location of structural embellishments to the beta-sandwich core that confer unique functional specificities. Reported ligands include chemokines, GM-CSF, IL-2, MHC class I, and glycosaminoglycans. We expect that the list of ligands and receptors engaged by the PIE domain will grow as we come to better understand how this versatile structural architecture can be tailored to manipulate host responses to infection. Full article

Posttranslational modifications (PTMs) of proteins include enzymatic changes by covalent addition of cellular regulatory determinants such as ubiquitin (Ub) and small ubiquitin-like modifier (SUMO) moieties. These modifications are widely used by eukaryotic cells to control the functional repertoire of proteins. Over the last decade, it became apparent that the repertoire of ubiquitiylation and SUMOylation regulating various biological functions is not restricted to eukaryotic cells, but is also a feature of human virus families, used to extensively exploit complex host-cell networks and homeostasis. Intriguingly, besides binding to host SUMO/Ub control proteins and interfering with the respective enzymatic cascade, many viral proteins mimic key regulatory factors to usurp this host machinery and promote efficient viral outcomes. Advanced detection methods and functional studies of ubiquitiylation and SUMOylation during virus-host interplay have revealed that human viruses have evolved a large arsenal of strategies to exploit these specific PTM processes. In this review, we highlight the known viral analogs orchestrating ubiquitin and SUMO conjugation events to subvert and utilize basic enzymatic pathways. Full article