Viruses

22 pages, 3974 KiB

Open AccessArticle

Glutamic Acid Residues in HIV-1 p6 Regulate Virus Budding and Membrane Association of Gag

by Melanie Friedrich¹, Christian Setz¹, Friedrich Hahn¹, Alina Matthaei¹, Kirsten Fraedrich¹

, Pia Rauch¹, Petra Henklein², Maximilian Traxdorf³, Torgils Fossen⁴ and Ulrich Schubert^1,*

¹ Institute of Virology, Friedrich-Alexander University Erlangen-Nürnberg (FAU), Erlangen 91054, Germany

² Institute of Biochemistry, Charité Universitätsmedizin-Berlin, Berlin 10117, Germany

³ Department of Otorhinolaryngology, Head and Neck Surgery, Friedrich-Alexander University Erlangen-Nürnberg (FAU), Erlangen 91054, Germany

⁴ Department of Chemistry and Center for Pharmacy, University of Bergen, Bergen N-5007, Norway

Viruses 2016, 8(4), 117; https://doi.org/10.3390/v8040117 - 25 Apr 2016

Cited by 11 | Viewed by 8171

Abstract

The HIV-1 Gag p6 protein regulates the final abscission step of nascent virions from the cell membrane by the action of its two late (l-) domains, which recruit Tsg101 and ALIX, components of the ESCRT system. Even though p6 consists of [...] Read more.

The HIV-1 Gag p6 protein regulates the final abscission step of nascent virions from the cell membrane by the action of its two late (l-) domains, which recruit Tsg101 and ALIX, components of the ESCRT system. Even though p6 consists of only 52 amino acids, it is encoded by one of the most polymorphic regions of the HIV-1 gag gene and undergoes various posttranslational modifications including sumoylation, ubiquitination, and phosphorylation. In addition, it mediates the incorporation of the HIV-1 accessory protein Vpr into budding virions. Despite its small size, p6 exhibits an unusually high charge density. In this study, we show that mutation of the conserved glutamic acids within p6 increases the membrane association of Pr55 Gag followed by enhanced polyubiquitination and MHC-I antigen presentation of Gag-derived epitopes, possibly due to prolonged exposure to membrane bound E3 ligases. The replication capacity of the total glutamic acid mutant E0A was almost completely impaired, which was accompanied by defective virus release that could not be rescued by ALIX overexpression. Altogether, our data indicate that the glutamic acids within p6 contribute to the late steps of viral replication and may contribute to the interaction of Gag with the plasma membrane. Full article

(This article belongs to the Special Issue Host Membranes and the Viral Infection Cycle)

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12 pages, 3605 KiB

Open AccessArticle

Development of a Triple-Color Pseudovirion-Based Assay to Detect Neutralizing Antibodies against Human Papillomavirus

by Jianhui Nie^†, Yangyang Liu^†, Weijin Huang and Youchun Wang^*

¹ Division of HIV/AIDS and Sexually Transmitted Virus Vaccines, National Institutes for Food and Drug Control (NIFDC), No. 2 Tiantanxili, Beijing 100050, China

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 107; https://doi.org/10.3390/v8040107 - 25 Apr 2016

Cited by 40 | Viewed by 7265

Abstract

Pseudovirion-based neutralization assay is considered the gold standard method for evaluating the immune response to human papillomavirus (HPV) vaccines. In this study, we developed a multicolor neutralization assay to simultaneously detect the neutralizing antibodies against different HPV types. FluoroSpot was used to interpret [...] Read more.

Pseudovirion-based neutralization assay is considered the gold standard method for evaluating the immune response to human papillomavirus (HPV) vaccines. In this study, we developed a multicolor neutralization assay to simultaneously detect the neutralizing antibodies against different HPV types. FluoroSpot was used to interpret the fluorescent protein expression instead of flow cytometry. The results of FluoroSpot and flow cytometry showed good consistency, with R² > 0.98 for the log-transformed IC₅₀ values. Regardless of the reporter color, the single-, dual-, and triple-color neutralization assays reported identical results for the same samples. In low-titer samples from naturally HPV-infected individuals, there was strong agreement between the single- and triple-color assays, with kappa scores of 0.92, 0.89, and 0.96 for HPV16, HPV18, and HPV58, respectively. Good reproducibility was observed for the triple-color assay, with coefficients of variation of 2.0%–41.5% within the assays and 8.3%–36.2% between the assays. Three triple-color systems, HPV16-18-58, HPV6-33-45, and HPV11-31-52, were developed that could evaluate the immunogenicity of a nonavalent vaccine in three rounds of the assay. With the advantages of an easy-to-use procedure and less sample consumption, the multiple-color assay is more suitable than classical assays for large sero-epidemiological studies and clinical trials and is more amenable to automation. Full article

(This article belongs to the Special Issue Replication-Competent Reporter-Expressing Viruses)

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16 pages, 3113 KiB

Open AccessArticle

The Correlation between the Virus- and Brain Antigen-Specific B Cell Response in the Blood of Patients with Multiple Sclerosis

by Marie Wunsch^1,*, Christopher Hohmann², Bianca Milles², Christina Rostermund¹, Paul V. Lehmann³, Michael Schroeter⁴, Antonios Bayas⁵, Jochen Ulzheimer⁶, Mathias Mäurer⁶, Süleyman Ergün¹ and Stefanie Kuerten¹

¹ Department of Anatomy and Cell Biology, University of Wuerzburg, Koellikerstr. 6, 97070 Wuerzburg, Germany

² Department of Anatomy I, University of Cologne, Joseph-Stelzmann-Str. 9, 50931 Cologne, Germany

³ Cellular Technology Limited, 20521 Chagrin Blvd, Shaker Heights, OH 44122, USA

⁴ Department of Neurology, University Hospitals of Cologne, Kerpener Straße 62, 50937 Cologne, Germany

⁵ Department of Neurology, Klinikum Augsburg, Stenglinstraße 2, 86156 Augsburg, Germany

⁶ Department of Neurology, Caritas-Krankenhaus Bad Mergentheim, Uhlandstraße 7, 97980 Bad Mergentheim, Germany

Viruses 2016, 8(4), 105; https://doi.org/10.3390/v8040105 - 23 Apr 2016

Cited by 5 | Viewed by 6802

Abstract

There is a largely divergent body of literature regarding the relationship between Epstein-Barr virus (EBV) infection and brain inflammation in multiple sclerosis (MS). Here, we tested MS patients during relapse (n = 11) and in remission (n = 19) in addition [...] Read more.

There is a largely divergent body of literature regarding the relationship between Epstein-Barr virus (EBV) infection and brain inflammation in multiple sclerosis (MS). Here, we tested MS patients during relapse (n = 11) and in remission (n = 19) in addition to n = 22 healthy controls to study the correlation between the EBV- and brain-specific B cell response in the blood by enzyme-linked immunospot (ELISPOT) and enzyme-linked immunosorbent assay (ELISA). Cytomegalovirus (CMV) was used as a control antigen tested in n = 16 MS patients during relapse and in n = 35 patients in remission. Over the course of the study, n = 16 patients were untreated, while n = 33 patients received immunomodulatory therapy. The data show that there was a moderate correlation between the frequencies of EBV- and brain-reactive B cells in MS patients in remission. In addition we could detect a correlation between the B cell response to EBV and disease activity. There was no evidence of an EBV reactivation. Interestingly, there was also a correlation between the frequencies of CMV- and brain-specific B cells in MS patients experiencing an acute relapse and an elevated B cell response to CMV was associated with higher disease activity. The trend remained when excluding seronegative subjects but was non-significant. These data underline that viral infections might impact the immunopathology of MS, but the exact link between the two entities remains subject of controversy. Full article

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14 pages, 3295 KiB

Open AccessArticle

HIV-1 Mutation and Recombination Rates Are Different in Macrophages and T-cells

by Deborah Cromer^1,2,†, Timothy E. Schlub^3,†, Redmond P. Smyth^4,5,†

, Andrew J. Grimm¹, Abha Chopra⁶, Simon Mallal⁶, Miles P. Davenport^1,2,*

and Johnson Mak^7,8,*

¹ Infection Analytics Program, Kirby Institute, UNSW Australia, Sydney NSW 2052, Australia

² Centre for Vascular Research, UNSW Australia, Sydney NSW 2052, Australia

³ Sydney School of Public Health, Sydney Medical School, University of Sydney, Sydney NSW 2006, Australia

⁴ Centre for Virology, Burnet Institute, Melbourne VIC 3004, Australia

⁵ Architecture et Réactivité de l’ARN, IBMC, CNRS, Université de Strasbourg, 67084 Strasbourg, France

⁶ Institute for Immunology and Infectious Diseases (IIID), Murdoch University, Perth WA 6150, Australia

⁷ Biosecurity Flagship, CSIRO (AAHL), Geelong VIC 3220, Australia

⁸ School of Medicine, Deakin University and CSIRO (AAHL), Geelong VIC 3216, Australia

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 118; https://doi.org/10.3390/v8040118 - 22 Apr 2016

Cited by 12 | Viewed by 7886

Abstract

High rates of mutation and recombination help human immunodeficiency virus (HIV) to evade the immune system and develop resistance to antiretroviral therapy. Macrophages and T-cells are the natural target cells of HIV-1 infection. A consensus has not been reached as to whether HIV [...] Read more.

High rates of mutation and recombination help human immunodeficiency virus (HIV) to evade the immune system and develop resistance to antiretroviral therapy. Macrophages and T-cells are the natural target cells of HIV-1 infection. A consensus has not been reached as to whether HIV replication results in differential recombination between primary T-cells and macrophages. Here, we used HIV with silent mutation markers along with next generation sequencing to compare the mutation and the recombination rates of HIV directly in T lymphocytes and macrophages. We observed a more than four-fold higher recombination rate of HIV in macrophages compared to T-cells (p < 0.001) and demonstrated that this difference is not due to different reliance on C-X-C chemokine receptor type 4 (CXCR4) and C-C chemokine receptor type 5 (CCR5) co-receptors between T-cells and macrophages. We also found that the pattern of recombination across the HIV genome (hot and cold spots) remains constant between T-cells and macrophages despite a three-fold increase in the overall recombination rate. This indicates that the difference in rates is a general feature of HIV DNA synthesis during macrophage infection. In contrast to HIV recombination, we found that T-cells have a 30% higher mutation rate than macrophages (p < 0.001) and that the mutational profile is similar between these cell types. Unexpectedly, we found no association between mutation and recombination in macrophages, in contrast to T-cells. Our data highlights some of the fundamental difference of HIV recombination and mutation amongst these two major target cells of infection. Understanding these differences will provide invaluable insights toward HIV evolution and how the virus evades immune surveillance and anti-retroviral therapeutics. Full article

(This article belongs to the Special Issue Next Generation Sequencing: New Developments and Discoveries in Virology)

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15 pages, 1917 KiB

Open AccessArticle

RNA-Sequencing Reveals the Progression of Phage-Host Interactions between φR1-37 and Yersinia enterocolitica

by Katarzyna Leskinen¹, Bob G. Blasdel², Rob Lavigne²

and Mikael Skurnik^1,3,*

¹ Department of Bacteriology and Immunology, Medicum, and Research Programs Unit, Immunobiology, University of Helsinki, P.O.Box 21 (Haartmaninkatu 3), FIN-00014 HY Helsinki, Finland

² Laboratory of Gene Technology, KU Leuven, BE-3001 Leuven, Belgium

³ Division of Clinical Microbiology, Helsinki University Hospital, HUSLAB, FIN-00270 Helsinki, Finland

Viruses 2016, 8(4), 111; https://doi.org/10.3390/v8040111 - 22 Apr 2016

Cited by 70 | Viewed by 8907

Abstract

Despite the expanding interest in bacterial viruses (bacteriophages), insights into the intracellular development of bacteriophage and its impact on bacterial physiology are still scarce. Here we investigate during lytic infection the whole-genome transcription of the giant phage vB_YecM_φR1-37 (φR1-37) and its host, the [...] Read more.

Despite the expanding interest in bacterial viruses (bacteriophages), insights into the intracellular development of bacteriophage and its impact on bacterial physiology are still scarce. Here we investigate during lytic infection the whole-genome transcription of the giant phage vB_YecM_φR1-37 (φR1-37) and its host, the gastroenteritis causing bacterium Yersinia enterocolitica. RNA sequencing reveals that the gene expression of φR1-37 does not follow a pattern typical observed in other lytic bacteriophages, as only selected genes could be classified as typically early, middle or late genes. The majority of the genes appear to be expressed constitutively throughout infection. Additionally, our study demonstrates that transcription occurs mainly from the positive strand, while the negative strand encodes only genes with low to medium expression levels. Interestingly, we also detected the presence of antisense RNA species, as well as one non-coding intragenic RNA species. Gene expression in the phage-infected cell is characterized by the broad replacement of host transcripts with phage transcripts. However, the host response in the late phase of infection was also characterized by up-regulation of several specific bacterial gene products known to be involved in stress response and membrane stability, including the Cpx pathway regulators, ATP-binding cassette (ABC) transporters, phage- and cold-shock proteins. Full article

(This article belongs to the Section Bacterial Viruses)

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12 pages, 1430 KiB

Open AccessArticle

Vaccination with the Secreted Glycoprotein G of Herpes Simplex Virus 2 Induces Protective Immunity after Genital Infection

by Karin Önnheim, Maria Ekblad, Staffan Görander, Tomas Bergström and Jan-Åke Liljeqvist^*

Section of Virology, Department of Infectious Medicine, Institute of Biomedicine, The Sahlgrenska Academy, University of Gothenburg, Guldhedsgatan 10 B, S-413 46 Gothenburg, Sweden

Viruses 2016, 8(4), 110; https://doi.org/10.3390/v8040110 - 22 Apr 2016

Cited by 8 | Viewed by 6098

Abstract

Herpes simplex virus 2 (HSV-2) infects the genital mucosa and establishes a life-long infection in sensory ganglia. After primary infection HSV-2 may reactivate causing recurrent genital ulcerations. HSV-2 infection is prevalent, and globally more than 400 million individuals are infected. As clinical trials [...] Read more.

Herpes simplex virus 2 (HSV-2) infects the genital mucosa and establishes a life-long infection in sensory ganglia. After primary infection HSV-2 may reactivate causing recurrent genital ulcerations. HSV-2 infection is prevalent, and globally more than 400 million individuals are infected. As clinical trials have failed to show protection against HSV-2 infection, new vaccine candidates are warranted. The secreted glycoprotein G (sgG-2) of HSV-2 was evaluated as a prophylactic vaccine in mice using two different immunization and adjuvant protocols. The protocol with three intramuscular immunizations combining sgG-2 with cytosine-phosphate-guanine dinucleotide (CpG) motifs and alum induced almost complete protection from genital and systemic disease after intra-vaginal challenge with HSV-2. Robust immunoglobulin G (IgG) antibody titers were detected with no neutralization activity. Purified splenic CD4+ T cells proliferated and produced interferon-γ (IFN-γ) when re-stimulated with the antigen in vitro. sgG-2 + adjuvant intra-muscularly immunized mice showed a significant reduction of infectious HSV-2 and increased IFN-γ levels in vaginal washes. The HSV-2 DNA copy numbers were significantly reduced in dorsal root ganglia, spinal cord, and in serum at day six or day 21 post challenge. We show that a sgG-2 based vaccine is highly effective and can be considered as a novel candidate in the development of a prophylactic vaccine against HSV-2 infection. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

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16 pages, 2042 KiB

Open AccessArticle

Intranasal Immunization with Influenza Virus-Like Particles Containing Membrane-Anchored Cholera Toxin B or Ricin Toxin B Enhances Adaptive Immune Responses and Protection against an Antigenically Distinct Virus

by Xianliang Ji^1,2, Zhiguang Ren^2,3,4, Na Xu⁵, Lingnan Meng^2,6, Zhijun Yu^2,3, Na Feng², Xiaoyu Sang², Shengnan Li^2,6, Yuanguo Li², Tiecheng Wang², Yongkun Zhao^2,7, Hualei Wang^2,7, Xuexing Zheng^2,8, Hongli Jin², Nan Li², Songtao Yang^2,7

, Jinshan Cao¹, Wensen Liu^2,*, Yuwei Gao^2,7,* and Xianzhu Xia^2,7,*

¹ College of Veterinary Medicine, Inner Mongolia Agricultural University, Huhhot 010018, China

² Institute of Military Veterinary, Academy of Military Medical Sciences, Changchun 130122, China

³ Institute of Laboratory Animal Sciences, Chinese Academy of Medical Sciences Peking Union Medical College, Beijing 100730, China

⁴ Key Lab of Cellular and Molecular Immunology, Henan University School of Medicine, Kaifeng 475001, China

⁵ Jilin Medical University, Changchun 132013, China

⁶ College of Animal Science and Technology, Jilin Agricultural University, Changchun 130118, China

⁷ Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses, Nanjing 210009, China

⁸ School of Public Health, Shandong University, Jinan 250110, China

Viruses 2016, 8(4), 115; https://doi.org/10.3390/v8040115 - 21 Apr 2016

Cited by 8 | Viewed by 5728

Abstract

Vaccination is the most effective means to prevent influenza virus infection, although current approaches are associated with suboptimal efficacy. Here, we generated virus-like particles (VLPs) composed of the hemagglutinin (HA), neuraminidase (NA) and matrix protein (M1) of A/Changchun/01/2009 (H1N1) with or without either [...] Read more.

Vaccination is the most effective means to prevent influenza virus infection, although current approaches are associated with suboptimal efficacy. Here, we generated virus-like particles (VLPs) composed of the hemagglutinin (HA), neuraminidase (NA) and matrix protein (M1) of A/Changchun/01/2009 (H1N1) with or without either membrane-anchored cholera toxin B (CTB) or ricin toxin B (RTB) as molecular adjuvants. The intranasal immunization of mice with VLPs containing membrane-anchored CTB or RTB elicited stronger humoral and cellular immune responses when compared to mice immunized with VLPs alone. Administration of VLPs containing CTB or RTB significantly enhanced virus-specific systemic and mucosal antibody responses, hemagglutination inhibiting antibody titers, virus neutralizing antibody titers, and the frequency of virus-specific IFN-γ and IL-4 secreting splenocytes. VLPs with and without CTB or RTB conferred complete protection against lethal challenge with a mouse-adapted homologous virus. When challenged with an antigenically distinct H1N1 virus, all mice immunized with VLPs containing CTB or RTB survived whereas mice immunized with VLPs alone showed only partial protection (80% survival). Our results suggest that membrane-anchored CTB and RTB possess strong adjuvant properties when incorporated into an intranasally-delivered influenza VLP vaccine. Chimeric influenza VLPs containing CTB or RTB may represent promising vaccine candidates for improved immunological protection against homologous and antigenically distinct influenza viruses. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

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16 pages, 2897 KiB

Open AccessArticle

De Novo Transcriptome Analysis Shows That SAV-3 Infection Upregulates Pattern Recognition Receptors of the Endosomal Toll-Like and RIG-I-Like Receptor Signaling Pathways in Macrophage/Dendritic Like TO-Cells

by Cheng Xu, Øystein Evensen and Hetron Mweemba Munang’andu^*

Section of Aquatic Medicine and Nutrition, Department of Basic Sciences and Aquatic Medicine, Faculty of Veterinary Medicine and Biosciences, Norwegian University of Life Sciences, Ullevålsveien 72, P.O. Box 8146 Dep NO-0033 Oslo, Norway

Viruses 2016, 8(4), 114; https://doi.org/10.3390/v8040114 - 21 Apr 2016

Cited by 28 | Viewed by 9893

Abstract

A fundamental step in cellular defense mechanisms is the recognition of “danger signals” made of conserved pathogen associated molecular patterns (PAMPs) expressed by invading pathogens, by host cell germ line coded pattern recognition receptors (PRRs). In this study, we used RNA-seq and the [...] Read more.

A fundamental step in cellular defense mechanisms is the recognition of “danger signals” made of conserved pathogen associated molecular patterns (PAMPs) expressed by invading pathogens, by host cell germ line coded pattern recognition receptors (PRRs). In this study, we used RNA-seq and the Kyoto encyclopedia of genes and genomes (KEGG) to identify PRRs together with the network pathway of differentially expressed genes (DEGs) that recognize salmonid alphavirus subtype 3 (SAV-3) infection in macrophage/dendritic like TO-cells derived from Atlantic salmon (Salmo salar L) headkidney leukocytes. Our findings show that recognition of SAV-3 in TO-cells was restricted to endosomal Toll-like receptors (TLRs) 3 and 8 together with RIG-I-like receptors (RLRs) and not the nucleotide-binding oligomerization domain-like receptors NOD-like receptor (NLRs) genes. Among the RLRs, upregulated genes included the retinoic acid inducible gene I (RIG-I), melanoma differentiation association 5 (MDA5) and laboratory of genetics and physiology 2 (LGP2). The study points to possible involvement of the tripartite motif containing 25 (TRIM25) and mitochondrial antiviral signaling protein (MAVS) in modulating RIG-I signaling being the first report that links these genes to the RLR pathway in SAV-3 infection in TO-cells. Downstream signaling suggests that both the TLR and RLR pathways use interferon (IFN) regulatory factors (IRFs) 3 and 7 to produce IFN-a2. The validity of RNA-seq data generated in this study was confirmed by quantitative real time qRT-PCR showing that genes up- or downregulated by RNA-seq were also up- or downregulated by RT-PCR. Overall, this study shows that de novo transcriptome assembly identify key receptors of the TLR and RLR sensors engaged in host pathogen interaction at cellular level. We envisage that data presented here can open a road map for future intervention strategies in SAV infection of salmon. Full article

(This article belongs to the Section Animal Viruses)

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20 pages, 2201 KiB

Open AccessArticle

Validation of the Filovirus Plaque Assay for Use in Preclinical Studies

by Amy C. Shurtleff^1,*, Holly A. Bloomfield², Shannon Mort³, Steven A. Orr³, Brian Audet⁴, Thomas Whitaker⁴, Michelle J. Richards⁴ and Sina Bavari¹

¹ Molecular and Translational Sciences Division, United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Frederick, MD 21702, USA

² Center for Aerobiological Studies, United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Frederick, MD 21702, USA

³ Nonclinical Development Division, United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Frederick, MD 21702, USA

⁴ Cell Culture Laboratory, Virology Division, United States Army Medical Research Institute of Infectious Diseases, 1425 Porter Street, Frederick, MD 21702, USA

Viruses 2016, 8(4), 113; https://doi.org/10.3390/v8040113 - 21 Apr 2016

Cited by 16 | Viewed by 7985

Abstract

A plaque assay for quantitating filoviruses in virus stocks, prepared viral challenge inocula and samples from research animals has recently been fully characterized and standardized for use across multiple institutions performing Biosafety Level 4 (BSL-4) studies. After standardization studies were completed, Good Laboratory [...] Read more.

A plaque assay for quantitating filoviruses in virus stocks, prepared viral challenge inocula and samples from research animals has recently been fully characterized and standardized for use across multiple institutions performing Biosafety Level 4 (BSL-4) studies. After standardization studies were completed, Good Laboratory Practices (GLP)-compliant plaque assay method validation studies to demonstrate suitability for reliable and reproducible measurement of the Marburg Virus Angola (MARV) variant and Ebola Virus Kikwit (EBOV) variant commenced at the United States Army Medical Research Institute of Infectious Diseases (USAMRIID). The validation parameters tested included accuracy, precision, linearity, robustness, stability of the virus stocks and system suitability. The MARV and EBOV assays were confirmed to be accurate to ±0.5 log₁₀ PFU/mL. Repeatability precision, intermediate precision and reproducibility precision were sufficient to return viral titers with a coefficient of variation (%CV) of ≤30%, deemed acceptable variation for a cell-based bioassay. Intraclass correlation statistical techniques for the evaluation of the assay’s precision when the same plaques were quantitated by two analysts returned values passing the acceptance criteria, indicating high agreement between analysts. The assay was shown to be accurate and specific when run on Nonhuman Primates (NHP) serum and plasma samples diluted in plaque assay medium, with negligible matrix effects. Virus stocks demonstrated stability for freeze-thaw cycles typical of normal usage during assay retests. The results demonstrated that the EBOV and MARV plaque assays are accurate, precise and robust for filovirus titration in samples associated with the performance of GLP animal model studies. Full article

(This article belongs to the Collection Advances in Ebolavirus, Marburgvirus, and Cuevavirus Research)

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30 pages, 1358 KiB

Open AccessReview

Measles Virus Fusion Protein: Structure, Function and Inhibition

by Philippe Plattet^1,*, Lisa Alves², Michael Herren^1,3 and Hector C. Aguilar^4,5

¹ Division of Experimental and Clinical Research, , Vetsuisse Faculty, University of Bern, Bremgartenstrasse 109a, Bern 3001, Switzerland

² Clinical Neurology, Department of Veterinary Medicine, University of Cambridge, CB3 0ES, Cambridge, UK

³ Graduate School for Cellular and Biomedical Sciences, University of Bern, Bern 3012, Switzerland

⁴ Paul G. Allen School for Global Animal Health, Washington State University, Pullman, WA 99164, USA

⁵ Department of Veterinary Microbiology and Pathology, Washington State University, Pullman, WA 99164, USA

Viruses 2016, 8(4), 112; https://doi.org/10.3390/v8040112 - 21 Apr 2016

Cited by 85 | Viewed by 19787

Abstract

Measles virus (MeV), a highly contagious member of the Paramyxoviridae family, causes measles in humans. The Paramyxoviridae family of negative single-stranded enveloped viruses includes several important human and animal pathogens, with MeV causing approximately 120,000 deaths annually. MeV and canine distemper virus (CDV)-mediated [...] Read more.

Measles virus (MeV), a highly contagious member of the Paramyxoviridae family, causes measles in humans. The Paramyxoviridae family of negative single-stranded enveloped viruses includes several important human and animal pathogens, with MeV causing approximately 120,000 deaths annually. MeV and canine distemper virus (CDV)-mediated diseases can be prevented by vaccination. However, sub-optimal vaccine delivery continues to foster MeV outbreaks. Post-exposure prophylaxis with antivirals has been proposed as a novel strategy to complement vaccination programs by filling herd immunity gaps. Recent research has shown that membrane fusion induced by the morbillivirus glycoproteins is the first critical step for viral entry and infection, and determines cell pathology and disease outcome. Our molecular understanding of morbillivirus-associated membrane fusion has greatly progressed towards the feasibility to control this process by treating the fusion glycoprotein with inhibitory molecules. Current approaches to develop anti-membrane fusion drugs and our knowledge on drug resistance mechanisms strongly suggest that combined therapies will be a prerequisite. Thus, discovery of additional anti-fusion and/or anti-attachment protein small-molecule compounds may eventually translate into realistic therapeutic options. Full article

(This article belongs to the Special Issue Recent Progress in Measles Virus Research)

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20 pages, 996 KiB

Open AccessReview

Constraints on the Genetic and Antigenic Variability of Measles Virus

by Shannon M. Beaty

and Benhur Lee^*

Icahn School of Medicine at Mount Sinai, New York, NY 10029, USA

Viruses 2016, 8(4), 109; https://doi.org/10.3390/v8040109 - 21 Apr 2016

Cited by 43 | Viewed by 12621

Abstract

Antigenic drift and genetic variation are significantly constrained in measles virus (MeV). Genetic stability of MeV is exceptionally high, both in the lab and in the field, and few regions of the genome allow for rapid genetic change. The regions of the genome [...] Read more.

Antigenic drift and genetic variation are significantly constrained in measles virus (MeV). Genetic stability of MeV is exceptionally high, both in the lab and in the field, and few regions of the genome allow for rapid genetic change. The regions of the genome that are more tolerant of mutations (i.e., the untranslated regions and certain domains within the N, C, V, P, and M proteins) indicate genetic plasticity or structural flexibility in the encoded proteins. Our analysis reveals that strong constraints in the envelope proteins (F and H) allow for a single serotype despite known antigenic differences among its 24 genotypes. This review describes some of the many variables that limit the evolutionary rate of MeV. The high genomic stability of MeV appears to be a shared property of the Paramyxovirinae, suggesting a common mechanism that biologically restricts the rate of mutation. Full article

(This article belongs to the Special Issue Recent Progress in Measles Virus Research)

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21 pages, 4531 KiB

Open AccessReview

Molecular Insights into Crimean-Congo Hemorrhagic Fever Virus

by Marko Zivcec, Florine E. M. Scholte

, Christina F. Spiropoulou, Jessica R. Spengler

and Éric Bergeron^*

Viral Special Pathogens Branch, Division of High Consequence Pathogens and Pathology, Centers for Disease Control and Prevention, Atlanta, GA 30333, USA

Viruses 2016, 8(4), 106; https://doi.org/10.3390/v8040106 - 21 Apr 2016

Cited by 105 | Viewed by 20604

Abstract

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne pathogen that causes high morbidity and mortality. Efficacy of vaccines and antivirals to treat human CCHFV infections remains limited and controversial. Research into pathology and underlying molecular mechanisms of CCHFV and other nairoviruses is limited. [...] Read more.

Crimean-Congo hemorrhagic fever virus (CCHFV) is a tick-borne pathogen that causes high morbidity and mortality. Efficacy of vaccines and antivirals to treat human CCHFV infections remains limited and controversial. Research into pathology and underlying molecular mechanisms of CCHFV and other nairoviruses is limited. Significant progress has been made in our understanding of CCHFV replication and pathogenesis in the past decade. Here we review the most recent molecular advances in CCHFV-related research, and provide perspectives on future research. Full article

(This article belongs to the Special Issue Recent Progress in Bunyavirus Research)

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13 pages, 1594 KiB

Open AccessReview

Translational Control during Calicivirus Infection

by Elizabeth Royall and Nicolas Locker^*

Faculty of Health and Medical Sciences, School of Biosciences and Medicine, University of Surrey, Guildford GU2 7HX, UK

Viruses 2016, 8(4), 104; https://doi.org/10.3390/v8040104 - 20 Apr 2016

Cited by 21 | Viewed by 8023

Abstract

In this review, we provide an overview of the strategies developed by caliciviruses to subvert or regulate the host protein synthesis machinery to their advantage. As intracellular obligate parasites, viruses strictly depend on the host cell resources to produce viral proteins. Thus, many [...] Read more.

In this review, we provide an overview of the strategies developed by caliciviruses to subvert or regulate the host protein synthesis machinery to their advantage. As intracellular obligate parasites, viruses strictly depend on the host cell resources to produce viral proteins. Thus, many viruses have developed strategies that regulate the function of the host protein synthesis machinery, often leading to preferential translation of viral mRNAs. Caliciviruses lack a 5′ cap structure but instead have a virus-encoded VPg protein covalently linked to the 5′ end of their mRNAs. Furthermore, they encode 2–4 open reading frames within their genomic and subgenomic RNAs. Therefore, they use alternative mechanisms for translation whereby VPg interacts with eukaryotic initiation factors (eIFs) to act as a proteinaceous cap-substitute, and some structural proteins are produced by reinitiation of translation events. This review discusses our understanding of these key mechanisms during caliciviruses infection as well as recent insights into the global regulation of eIF4E activity. Full article

(This article belongs to the Special Issue Viral Subversion of Stress Responses and Translational Control)

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18 pages, 8118 KiB

Open AccessArticle

ESCRT Requirements for Murine Leukemia Virus Release

by Christina Bartusch and Reinhild Prange^*

Department of Medical Microbiology and Hygiene, University Medical Center of the Johannes Gutenberg University Mainz, Augustusplatz, Mainz D-55131, Germany

Viruses 2016, 8(4), 103; https://doi.org/10.3390/v8040103 - 18 Apr 2016

Cited by 21 | Viewed by 7708

Abstract

The Murine Leukemia Virus (MLV) is a gammaretrovirus that hijack host components of the endosomal sorting complex required for transport (ESCRT) for budding. To determine the minimal requirements for ESCRT factors in MLV viral and viral-like particles (VLP) release, an siRNA knockdown screen [...] Read more.

The Murine Leukemia Virus (MLV) is a gammaretrovirus that hijack host components of the endosomal sorting complex required for transport (ESCRT) for budding. To determine the minimal requirements for ESCRT factors in MLV viral and viral-like particles (VLP) release, an siRNA knockdown screen of ESCRT(-associated) proteins was performed in MLV-producing human cells. We found that MLV VLPs and virions primarily engage the ESCRT-I factor Tsg101 and marginally the ESCRT-associated adaptors Nedd4-1 and Alix to enter the ESCRT pathway. Conversely, the inactivation of ESCRT-II had no impact on VLP and virion egress. By analyzing the effects of individual ESCRT-III knockdowns, VLP and virion release was profoundly inhibited in CHMP2A- and CHMP4B-knockdown cells. In contrast, neither the CHMP2B and CHMP4A isoforms nor CHMP3, CHMP5, and CHMP6 were found to be essential. In case of CHMP1, we unexpectedly observed that the CHMP1A isoform was specifically required for virus budding, but dispensable for VLP release. Hence, MLV utilizes only a subset of ESCRT factors, and viral and viral-like particles differ in ESCRT-III factor requirements. Full article

(This article belongs to the Section Animal Viruses)

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26 pages, 989 KiB

Open AccessReview

Shutoff of Host Gene Expression in Influenza A Virus and Herpesviruses: Similar Mechanisms and Common Themes

by Hembly G. Rivas^1,2,†, Summer K. Schmaling^1,† and Marta M. Gaglia^1,3,*

¹ Department of Molecular Biology and Microbiology, Tufts University School of Medicine, Boston, MA 02111, USA

² Post-Baccalaureate Research Education Program, Tufts University School of Medicine, Boston, MA 02111, USA

³ Graduate Program in Molecular Microbiology, Tufts University School of Medicine, Boston, MA 02111, USA

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 102; https://doi.org/10.3390/v8040102 - 16 Apr 2016

Cited by 75 | Viewed by 11929

Abstract

The ability to shut off host gene expression is a shared feature of many viral infections, and it is thought to promote viral replication by freeing host cell machinery and blocking immune responses. Despite the molecular differences between viruses, an emerging theme in [...] Read more.

The ability to shut off host gene expression is a shared feature of many viral infections, and it is thought to promote viral replication by freeing host cell machinery and blocking immune responses. Despite the molecular differences between viruses, an emerging theme in the study of host shutoff is that divergent viruses use similar mechanisms to enact host shutoff. Moreover, even viruses that encode few proteins often have multiple mechanisms to affect host gene expression, and we are only starting to understand how these mechanisms are integrated. In this review we discuss the multiplicity of host shutoff mechanisms used by the orthomyxovirus influenza A virus and members of the alpha- and gamma-herpesvirus subfamilies. We highlight the surprising similarities in their mechanisms of host shutoff and discuss how the different mechanisms they use may play a coordinated role in gene regulation. Full article

(This article belongs to the Special Issue Viral Subversion of Stress Responses and Translational Control)

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15 pages, 1979 KiB

Open AccessArticle

Nuclear Translocation of Crk Adaptor Proteins by the Influenza A Virus NS1 Protein

by Leena Ylösmäki¹, Riku Fagerlund¹, Inka Kuisma¹, Ilkka Julkunen² and Kalle Saksela^1,*

¹ Department of Virology, University of Helsinki and Helsinki University Hospital, 00014 Helsinki, Finland

² Department of Virology, University of Turku, 20520 Turku, Finland and Virology Unit, Department of Infectious Disease Surveillance and Control, National Institute for Health and Welfare (THL), 00300 Helsinki, Finland

Viruses 2016, 8(4), 101; https://doi.org/10.3390/v8040101 - 15 Apr 2016

Cited by 6 | Viewed by 6193

Abstract

The non-structural protein-1 (NS1) of many influenza A strains, especially those of avian origin, contains an SH3 ligand motif, which binds tightly to the cellular adaptor proteins Crk (Chicken tumor virus number 10 (CT10) regulator of kinase) and Crk-like adapter protein (CrkL). This [...] Read more.

The non-structural protein-1 (NS1) of many influenza A strains, especially those of avian origin, contains an SH3 ligand motif, which binds tightly to the cellular adaptor proteins Crk (Chicken tumor virus number 10 (CT10) regulator of kinase) and Crk-like adapter protein (CrkL). This interaction has been shown to potentiate NS1-induced activation of the phosphatidylinositol 3-kinase (PI3K), but additional effects on the host cell physiology may exist. Here we show that NS1 can induce an efficient translocation of Crk proteins from the cytoplasm into the nucleus, which results in an altered pattern of nuclear protein tyrosine phosphorylation. This was not observed using NS1 proteins deficient in SH3 binding or engineered to be exclusively cytoplasmic, indicating a physical role for NS1 as a carrier in the nuclear translocation of Crk. These data further emphasize the role of Crk proteins as host cell interaction partners of NS1, and highlight the potential for host cell manipulation gained by a viral protein simply via acquiring a short SH3 binding motif. Full article

(This article belongs to the Section Animal Viruses)

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11 pages, 1674 KiB

Open AccessArticle

Chlamydiaphage φCPG1 Capsid Protein Vp1 Inhibits Chlamydia trachomatis Growth via the Mitogen-Activated Protein Kinase Pathway

by Yuanli Guo, Rui Guo, Quan Zhou, Changgui Sun, Xinmei Zhang, Yuanjun Liu and Quanzhong Liu^*

Dermatology and Venereology Department, Tianjin Medical University General Hospital, No. 154 Anshan Road, Heping District, Tianjin 300052, China

Viruses 2016, 8(4), 99; https://doi.org/10.3390/v8040099 - 14 Apr 2016

Cited by 8 | Viewed by 5731

Abstract

Chlamydia trachomatis is the most common cause of curable bacterial sexually transmitted infections worldwide. Although the pathogen is well established, the pathogenic mechanisms remain unclear. Given the current challenges of antibiotic resistance and blocked processes of vaccine development, the use of a specific [...] Read more.

Chlamydia trachomatis is the most common cause of curable bacterial sexually transmitted infections worldwide. Although the pathogen is well established, the pathogenic mechanisms remain unclear. Given the current challenges of antibiotic resistance and blocked processes of vaccine development, the use of a specific chlamydiaphage may be a new treatment solution. φCPG1 is a lytic phage specific for Chlamydia caviae, and shows over 90% nucleotide sequence identity with other chlamydiaphages. Vp1 is the major capsid protein of φCPG1. Purified Vp1 was previously confirmed to inhibit Chlamydia trachomatis growth. We here report the first attempt at exploring the relationship between Vp1-treated C. trachomatis and the protein and gene levels of the mitogen-activated/extracellular regulated protein kinase (MAPK/ERK) pathway by Western blotting and real-time PCR, respectively. Moreover, we evaluated the levels of pro-inflammatory cytokines interleukin (IL)-8 and IL-1 by enzyme-linked immunosorbent assay after Vp1 treatment. After 48 h of incubation, the p-ERK level of the Vp1-treated group decreased compared with that of the Chlamydia infection group. Accordingly, ERK1 and ERK2 mRNA expression levels of the Vp1-treated group also decreased compared with the Chlamydia infection group. IL-8 and IL-1 levels were also decreased after Vp1 treatment compared with the untreated group. Our results demonstrate that the inhibition effect of the chlamydiaphage φCPG1 capsid protein Vp1 on C. trachomatis is associated with the MAPK pathway, and inhibits production of the pro-inflammatory cytokines IL-8 and IL-1. The bacteriophages may provide insight into a new signaling transduction mechanism to influence their hosts, in addition to bacteriolysis. Full article

(This article belongs to the Section Bacterial Viruses)

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17 pages, 2260 KiB

Open AccessArticle

Purification and Biochemical Characterisation of Rabbit Calicivirus RNA-Dependent RNA Polymerases and Identification of Non-Nucleoside Inhibitors

by Nadya Urakova^1,2,3, Natalie Netzler⁴, Andrew G. Kelly⁴, Michael Frese^1,2,3,5

, Peter A. White⁴

and Tanja Strive^1,2,5,*

¹ Commonwealth Scinetific and Industrial Research Organisation, Health and Biosecurity, 2601 Canberra, ACT, Australia

² Invasive Animals Cooperative Research Centre, University of Canberra, 2617 Canberra, ACT, Australia

³ Health Research Institute, University of Canberra, 2617 Canberra, ACT, Australia

⁴ School of Biotechnology and Biomolecular Sciences, Faculty of Science, University of New South Wales, 2052 Sydney, NSW, Australia

⁵ Institute for Applied Ecology, University of Canberra, 2617 Canberra, ACT, Australia

Viruses 2016, 8(4), 100; https://doi.org/10.3390/v8040100 - 14 Apr 2016

Cited by 18 | Viewed by 6849

Abstract

Rabbit haemorrhagic disease virus (RHDV) is a calicivirus that causes acute infections in both domestic and wild European rabbits (Oryctolagus cuniculus). The virus causes significant economic losses in rabbit farming and reduces wild rabbit populations. The recent emergence of RHDV variants [...] Read more.

Rabbit haemorrhagic disease virus (RHDV) is a calicivirus that causes acute infections in both domestic and wild European rabbits (Oryctolagus cuniculus). The virus causes significant economic losses in rabbit farming and reduces wild rabbit populations. The recent emergence of RHDV variants capable of overcoming immunity to other strains emphasises the need to develop universally effective antivirals to enable quick responses during outbreaks until new vaccines become available. The RNA-dependent RNA polymerase (RdRp) is a primary target for the development of such antiviral drugs. In this study, we used cell-free in vitro assays to examine the biochemical characteristics of two rabbit calicivirus RdRps and the effects of several antivirals that were previously identified as human norovirus RdRp inhibitors. The non-nucleoside inhibitor NIC02 was identified as a potential scaffold for further drug development against rabbit caliciviruses. Our experiments revealed an unusually high temperature optimum (between 40 and 45 °C) for RdRps derived from both a pathogenic and a non-pathogenic rabbit calicivirus, possibly demonstrating an adaptation to a host with a physiological body temperature of more than 38 °C. Interestingly, the in vitro polymerase activity of the non-pathogenic calicivirus RdRp was at least two times higher than that of the RdRp of the highly virulent RHDV. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

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16 pages, 607 KiB

Open AccessReview

Human Cytomegalovirus Strategies to Maintain and Promote mRNA Translation

by Heather A. Vincent, Benjamin Ziehr and Nathaniel J. Moorman^*

Department of Microbiology & Immunology, Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill, Chapel Hill, NC 27599, USA

Viruses 2016, 8(4), 97; https://doi.org/10.3390/v8040097 - 13 Apr 2016

Cited by 18 | Viewed by 7934

Abstract

mRNA translation requires the ordered assembly of translation initiation factors and ribosomal subunits on a transcript. Host signaling pathways regulate each step in this process to match levels of protein synthesis to environmental cues. In response to infection, cells activate multiple defenses that [...] Read more.

mRNA translation requires the ordered assembly of translation initiation factors and ribosomal subunits on a transcript. Host signaling pathways regulate each step in this process to match levels of protein synthesis to environmental cues. In response to infection, cells activate multiple defenses that limit viral protein synthesis, which viruses must counteract to successfully replicate. Human cytomegalovirus (HCMV) inhibits host defenses that limit viral protein expression and manipulates host signaling pathways to promote the expression of both host and viral proteins necessary for virus replication. Here we review key regulatory steps in mRNA translation, and the strategies used by HCMV to maintain protein synthesis in infected cells. Full article

(This article belongs to the Special Issue Viral Subversion of Stress Responses and Translational Control)

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15 pages, 227 KiB

Open AccessReview

Current Approaches for Diagnosis of Influenza Virus Infections in Humans

by Sai Vikram Vemula, Jiangqin Zhao

, Jikun Liu, Xue Wang, Santanu Biswas and Indira Hewlett^*

Laboratory of Molecular Virology, Center for Biologics Evaluation and Research, Food and Drug Administration, Silver Spring, MD 20993, USA

Viruses 2016, 8(4), 96; https://doi.org/10.3390/v8040096 - 12 Apr 2016

Cited by 270 | Viewed by 22197

Abstract

Despite significant advancement in vaccine and virus research, influenza continues to be a major public health concern. Each year in the United States of America, influenza viruses are responsible for seasonal epidemics resulting in over 200,000 hospitalizations and 30,000–50,000 deaths. Accurate and early [...] Read more.

Despite significant advancement in vaccine and virus research, influenza continues to be a major public health concern. Each year in the United States of America, influenza viruses are responsible for seasonal epidemics resulting in over 200,000 hospitalizations and 30,000–50,000 deaths. Accurate and early diagnosis of influenza viral infections are critical for rapid initiation of antiviral therapy to reduce influenza related morbidity and mortality both during seasonal epidemics and pandemics. Several different approaches are currently available for diagnosis of influenza infections in humans. These include viral isolation in cell culture, immunofluorescence assays, nucleic acid amplification tests, immunochromatography-based rapid diagnostic tests, etc. Newer diagnostic approaches are being developed to overcome the limitations associated with some of the conventional detection methods. This review discusses diagnostic approaches currently available for detection of influenza viruses in humans. Full article

14 pages, 799 KiB

Open AccessReview

Viral Evasion of Natural Killer Cell Activation

by Yi Ma¹, Xiaojuan Li¹ and Ersheng Kuang^1,2,*

¹ Institute of Human Virology, Zhongshan School of Medicine, Sun Yat-Sen University, No. 74, Zhongshan 2nd Road, Guangzhou 510080, China

² Key Laboratory of Tropical Disease Control (Sun Yat-Sen University), Ministry of Education, Guangzhou 510080, China

Viruses 2016, 8(4), 95; https://doi.org/10.3390/v8040095 - 12 Apr 2016

Cited by 32 | Viewed by 8369

Abstract

Natural killer (NK) cells play a key role in antiviral innate defenses because of their abilities to kill infected cells and secrete regulatory cytokines. Additionally, NK cells exhibit adaptive memory-like antigen-specific responses, which represent a novel antiviral NK cell defense mechanism. Viruses have [...] Read more.

Natural killer (NK) cells play a key role in antiviral innate defenses because of their abilities to kill infected cells and secrete regulatory cytokines. Additionally, NK cells exhibit adaptive memory-like antigen-specific responses, which represent a novel antiviral NK cell defense mechanism. Viruses have evolved various strategies to evade the recognition and destruction by NK cells through the downregulation of the NK cell activating receptors. Here, we review the recent findings on viral evasion of NK cells via the impairment of NK cell-activating receptors and ligands, which provide new insights on the relationship between NK cells and viral actions during persistent viral infections. Full article

(This article belongs to the Section Animal Viruses)

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15 pages, 2334 KiB

Open AccessArticle

Virus-Like Particle Vaccination Protects Nonhuman Primates from Lethal Aerosol Exposure with Marburgvirus (VLP Vaccination Protects Macaques against Aerosol Challenges)

by John M. Dye^1,*,†, Kelly L. Warfield^2,*,†,‡, Jay B. Wells¹, Robert C. Unfer^2,‡, Sergey Shulenin², Hong Vu², Donald K. Nichols¹, M. Javad Aman² and Sina Bavari¹

¹ United States Army Medical Research Institute of Infectious Diseases, Fort Detrick, MD 21702, USA

² Integrated Biotherapeutics, Inc., Gaithersburg, MD 20878, USA

^† These authors contributed equally to this work.

^‡ Current affiliation: Emergent BioSolutions, Gaithersburg, MD 20879, USA (K.L.W.); NIH/NIAID, Rockville, MD 20852, USA (R.C.U.)

Viruses 2016, 8(4), 94; https://doi.org/10.3390/v8040094 - 8 Apr 2016

Cited by 20 | Viewed by 6913

Abstract

Marburg virus (MARV) was the first filovirus to be identified following an outbreak of viral hemorrhagic fever disease in Marburg, Germany in 1967. Due to several factors inherent to filoviruses, they are considered a potential bioweapon that could be disseminated via an aerosol [...] Read more.

Marburg virus (MARV) was the first filovirus to be identified following an outbreak of viral hemorrhagic fever disease in Marburg, Germany in 1967. Due to several factors inherent to filoviruses, they are considered a potential bioweapon that could be disseminated via an aerosol route. Previous studies demonstrated that MARV virus-like particles (VLPs) containing the glycoprotein (GP), matrix protein VP40 and nucleoprotein (NP) generated using a baculovirus/insect cell expression system could protect macaques from subcutaneous (SQ) challenge with multiple species of marburgviruses. In the current study, the protective efficacy of the MARV VLPs in conjunction with two different adjuvants: QS-21, a saponin derivative, and poly I:C against homologous aerosol challenge was assessed in cynomolgus macaques. Antibody responses against the GP antigen were equivalent in all groups receiving MARV VLPs irrespective of the adjuvant; adjuvant only-vaccinated macaques did not demonstrate appreciable antibody responses. All macaques were subsequently challenged with lethal doses of MARV via aerosol or SQ as a positive control. All MARV VLP-vaccinated macaques survived either aerosol or SQ challenge while animals administered adjuvant only exhibited clinical signs and lesions consistent with MARV disease and were euthanized after meeting the predetermined criteria. Therefore, MARV VLPs induce IgG antibodies recognizing MARV GP and VP40 and protect cynomolgus macaques from an otherwise lethal aerosol exposure with MARV. Full article

(This article belongs to the Collection Advances in Ebolavirus, Marburgvirus, and Cuevavirus Research)

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27 pages, 726 KiB

Open AccessReview

Next-Generation Sequencing in the Understanding of Kaposi’s Sarcoma-Associated Herpesvirus (KSHV) Biology

by Roxanne Strahan^†, Timsy Uppal^†

and Subhash C. Verma^*

¹ Department of Microbiology and Immunology, School of Medicine, University of Nevada, 1664 N, Virginia Street, MS 320, Reno, NV 89557, USA

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 92; https://doi.org/10.3390/v8040092 - 31 Mar 2016

Cited by 9 | Viewed by 10706

Abstract

Non-Sanger-based novel nucleic acid sequencing techniques, referred to as Next-Generation Sequencing (NGS), provide a rapid, reliable, high-throughput, and massively parallel sequencing methodology that has improved our understanding of human cancers and cancer-related viruses. NGS has become a quintessential research tool for more effective [...] Read more.

Non-Sanger-based novel nucleic acid sequencing techniques, referred to as Next-Generation Sequencing (NGS), provide a rapid, reliable, high-throughput, and massively parallel sequencing methodology that has improved our understanding of human cancers and cancer-related viruses. NGS has become a quintessential research tool for more effective characterization of complex viral and host genomes through its ever-expanding repertoire, which consists of whole-genome sequencing, whole-transcriptome sequencing, and whole-epigenome sequencing. These new NGS platforms provide a comprehensive and systematic genome-wide analysis of genomic sequences and a full transcriptional profile at a single nucleotide resolution. When combined, these techniques help unlock the function of novel genes and the related pathways that contribute to the overall viral pathogenesis. Ongoing research in the field of virology endeavors to identify the role of various underlying mechanisms that control the regulation of the herpesvirus biphasic lifecycle in order to discover potential therapeutic targets and treatment strategies. In this review, we have complied the most recent findings about the application of NGS in Kaposi’s sarcoma-associated herpesvirus (KSHV) biology, including identification of novel genomic features and whole-genome KSHV diversities, global gene regulatory network profiling for intricate transcriptome analyses, and surveying of epigenetic marks (DNA methylation, modified histones, and chromatin remodelers) during de novo, latent, and productive KSHV infections. Full article

(This article belongs to the Special Issue Next Generation Sequencing: New Developments and Discoveries in Virology)

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16 pages, 964 KiB

Open AccessReview

Enterovirus Control of Translation and RNA Granule Stress Responses

by Richard E. Lloyd

Department of Molecular Virology and Microbiology, Baylor College of Medicine, Houston, TX 77030, USA

Viruses 2016, 8(4), 93; https://doi.org/10.3390/v8040093 - 30 Mar 2016

Cited by 31 | Viewed by 10431

Abstract

Enteroviruses such as poliovirus (PV) and coxsackievirus B3 (CVB3) have evolved several parallel strategies to regulate cellular gene expression and stress responses to ensure efficient expression of the viral genome. Enteroviruses utilize their encoded proteinases to take over the cellular translation apparatus and [...] Read more.

Enteroviruses such as poliovirus (PV) and coxsackievirus B3 (CVB3) have evolved several parallel strategies to regulate cellular gene expression and stress responses to ensure efficient expression of the viral genome. Enteroviruses utilize their encoded proteinases to take over the cellular translation apparatus and direct ribosomes to viral mRNAs. In addition, viral proteinases are used to control and repress the two main types of cytoplasmic RNA granules, stress granules (SGs) and processing bodies (P-bodies, PBs), which are stress-responsive dynamic structures involved in repression of gene expression. This review discusses these processes and the current understanding of the underlying mechanisms with respect to enterovirus infections. In addition, the review discusses accumulating data suggesting linkage exists between RNA granule formation and innate immune sensing and activation. Full article

(This article belongs to the Special Issue Viral Subversion of Stress Responses and Translational Control)

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16 pages, 3537 KiB

Open AccessArticle

Natural History of Aerosol Exposure with Marburg Virus in Rhesus Macaques

by Evan C. Ewers^1,†, William D. Pratt^2,*,†, Nancy A. Twenhafel^1,2, Joshua Shamblin², Ginger Donnelly², Heather Esham², Carly Wlazlowski², Joshua C. Johnson³

, Miriam Botto², Lisa E. Hensley³ and Arthur J. Goff^1,2,†

¹ Department of Medicine, Tripler Army Medical Center, Honolulu, HI 96859, USA

² US Army Medical Research Institute of Infectious Diseases, Fort Detrick, Frederick, MD 21702, USA

³ Integrated Research Facility, National Institute of Allergy and Infectious Diseases, National Institutes of Health, Frederick, MD 21702, USA

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 87; https://doi.org/10.3390/v8040087 - 30 Mar 2016

Cited by 19 | Viewed by 7287

Abstract

Marburg virus causes severe and often lethal viral disease in humans, and there are currently no Food and Drug Administration (FDA) approved medical countermeasures. The sporadic occurrence of Marburg outbreaks does not allow for evaluation of countermeasures in humans, so therapeutic and vaccine [...] Read more.

Marburg virus causes severe and often lethal viral disease in humans, and there are currently no Food and Drug Administration (FDA) approved medical countermeasures. The sporadic occurrence of Marburg outbreaks does not allow for evaluation of countermeasures in humans, so therapeutic and vaccine candidates can only be approved through the FDA animal rule—a mechanism requiring well-characterized animal models in which efficacy would be evaluated. Here, we describe a natural history study where rhesus macaques were surgically implanted with telemetry devices and central venous catheters prior to aerosol exposure with Marburg-Angola virus, enabling continuous physiologic monitoring and blood sampling without anesthesia. After a three to four day incubation period, all animals developed fever, viremia, and lymphopenia before developing tachycardia, tachypnea, elevated liver enzymes, decreased liver function, azotemia, elevated D-dimer levels and elevated pro-inflammatory cytokines suggesting a systemic inflammatory response with organ failure. The final, terminal period began with the onset of sustained hypotension, dehydration progressed with signs of major organ hypoperfusion (hyperlactatemia, acute kidney injury, hypothermia), and ended with euthanasia or death. The most significant pathologic findings were marked infection of the respiratory lymphoid tissue with destruction of the tracheobronchial and mediastinal lymph nodes, and severe diffuse infection in the liver, and splenitis. Full article

(This article belongs to the Collection Advances in Ebolavirus, Marburgvirus, and Cuevavirus Research)

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11 pages, 1237 KiB

Open AccessArticle

Recombinant Pseudorabies Virus (PRV) Expressing Firefly Luciferase Effectively Screened for CRISPR/Cas9 Single Guide RNAs and Antiviral Compounds

by Yan-Dong Tang^1,†, Ji-Ting Liu^1,2,†, Qiong-Qiong Fang^1,†, Tong-Yun Wang^1,†, Ming-Xia Sun¹, Tong-Qing An¹, Zhi-Jun Tian¹ and Xue-Hui Cai^1,*

¹ The Key Laboratory of Veterinary Public Health, Ministry of Agriculture, State Key Laboratory of Veterinary Biotechnology, Harbin Veterinary Research Institute of the Chinese Academy of Agricultural Sciences, Harbin 150001, China

² College of Animal Science and Technology, Jilin Agriculture University, Changchun 130018, China

^† These authors contributed equally to this work.

Viruses 2016, 8(4), 90; https://doi.org/10.3390/v8040090 - 29 Mar 2016

Cited by 31 | Viewed by 8712

Abstract

A Pseudorabies virus (PRV) variant has emerged in China since 2011 that is not protected by commercial vaccines, and has not been well studied. The PRV genome is large and difficult to manipulate, but it is feasible to use clustered, regularly interspaced short [...] Read more.

A Pseudorabies virus (PRV) variant has emerged in China since 2011 that is not protected by commercial vaccines, and has not been well studied. The PRV genome is large and difficult to manipulate, but it is feasible to use clustered, regularly interspaced short palindromic repeats (CRISPR)/Cas9 technology. However, identification of single guide RNA (sgRNA) through screening is critical to the CRISPR/Cas9 system, and is traditionally time and labor intensive, and not suitable for rapid and high throughput screening of effective PRV sgRNAs. In this study, we developed a recombinant PRV strain expressing firefly luciferase and enhanced green fluorescent protein (EGFP) as a reporter virus for PRV-specific sgRNA screens and rapid evaluation of antiviral compounds. Luciferase activity was apparent as soon as 4 h after infection and was stably expressed through 10 passages. In a proof of the principle screen, we were able to identify several PRV specific sgRNAs and confirmed that they inhibited PRV replication using traditional methods. Using the reporter virus, we also identified PRV variants lacking US3, US2, and US9 gene function, and showed anti-PRV activity for chloroquine. Our results suggest that the reporter PRV strain will be a useful tool for basic virology studies, and for developing PRV control and prevention measures. Full article

(This article belongs to the Special Issue Replication-Competent Reporter-Expressing Viruses)

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8 pages, 679 KiB

Open AccessArticle

Frequency of Natural Resistance within NS5a Replication Complex Domain in Hepatitis C Genotypes 1a, 1b: Possible Implication of Subtype-Specific Resistance Selection in Multiple Direct Acting Antivirals Drugs Combination Treatment

by Sabrina Bagaglio^1,2,*, Andrea Andolina¹, Marco Merli¹, Caterina Uberti-Foppa¹ and Giulia Morsica¹

¹ Infectious Diseases Department, Scientific Institute Ospedale San Raffaele, 20127 Milan, Italy

² Vita-Salute University, 20132 Milan, Italy

Viruses 2016, 8(4), 91; https://doi.org/10.3390/v8040091 - 25 Mar 2016

Cited by 18 | Viewed by 5441

Abstract

Different HCV subtypes may naturally harbor different resistance selection to anti-NS5a inhibitors. 2761 sequences retrieved from the Los Alamos HCV database were analyzed in the NS5a domain 1, the target of NS5a inhibitors. The NS5a resistance-associated polymorphisms (RAPs) were more frequently detected in [...] Read more.

Different HCV subtypes may naturally harbor different resistance selection to anti-NS5a inhibitors. 2761 sequences retrieved from the Los Alamos HCV database were analyzed in the NS5a domain 1, the target of NS5a inhibitors. The NS5a resistance-associated polymorphisms (RAPs) were more frequently detected in HCV G1b compared to G1a. The prevalence of polymorphisms associated with cross-resistance to compounds in clinical use (daclatasvir, DCV, ledipasvir, LDV, ombitasvir, and OMV) or scheduled to come into clinical use in the near future (IDX719, elbasvir, and ELV) was higher in G1b compared to G1a (37/1552 (2.4%) in 1b sequences and 15/1209 (1.2%) in 1a isolates, p = 0.040). Interestingly, on the basis of the genotype-specific resistance pattern, 95 (6.1%) G1b sequences had L31M RAP to DCV/IDX719, while 6 sequences of G1a (0.5%) harbored L31M RAP, conferring resistance to DCV/LDV/IDX719/ELV (p < 0.0001). Finally, 28 (2.3%) G1a and none of G1b isolates harbored M28V RAP to OMV (p < 0.0001). In conclusion, the pattern of subtype-specific resistance selection in the naturally occurring strains may guide the treatment option in association with direct acting antivirals (DAAs) targeting different regions, particularly in patients that are difficult to cure, such as those with advanced liver disease or individuals who have failed previous DAAs. Full article

(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)

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1 pages, 139 KiB

Open AccessCorrection

Correction: Shuang, L.; et al. Autophagy Activated by Bluetongue Virus Infection Plays a Positive Role in Its Replication, Viruses 2015, 7, 4657–4675

by Viruses Editorial Office

MDPI AG, Klybeckstrasse 64, CH-4057 Basel, Switzerland

Viruses 2016, 8(4), 89; https://doi.org/10.3390/v8040089 - 24 Mar 2016

Viewed by 3906

Abstract

The Viruses Editorial Office wishes to notify its readers of a correction in [1].[...] Full article

7 pages, 528 KiB

Open AccessReview

Determinants of the Bovine Leukemia Virus Envelope Glycoproteins Involved in Infectivity, Replication and Pathogenesis

by Alix De Brogniez^1,*, Jan Mast² and Luc Willems^1,*

¹ Molecular and Cellular Epigenetics (GIGA) and Molecular Biology (Gembloux Agro-Bio Tech), University of Liège (ULg), 4000 Liège, Belgium

² Veterinary and Agrochemical Research Center CODA-CERVA, 1180 Brussels, Belgium

Viruses 2016, 8(4), 88; https://doi.org/10.3390/v8040088 - 24 Mar 2016

Cited by 18 | Viewed by 6042

Abstract

Interaction of viral envelope proteins with host cell membranes has been extensively investigated in a number of systems. However, the biological relevance of these interactions in vivo has been hampered by the absence of adequate animal models. Reverse genetics using the bovine leukemia [...] Read more.

Interaction of viral envelope proteins with host cell membranes has been extensively investigated in a number of systems. However, the biological relevance of these interactions in vivo has been hampered by the absence of adequate animal models. Reverse genetics using the bovine leukemia virus (BLV) genome highlighted important functional domains of the envelope protein involved in the viral life cycle. For example, immunoreceptor tyrosine-based activation motifs (ITAM) of the envelope transmembrane protein (TM) are essential determinants of infection. Although cell fusion directed by the aminoterminal end of TM is postulated to be essential, some proviruses expressing fusion-deficient envelope proteins unexpectedly replicate at wild-type levels. Surprisingly also, a conserved N-linked glycosylation site of the extracellular envelope protein (SU) inhibits cell-to-cell transmission suggesting that infectious potential has been limited during evolution. In this review, we summarize the knowledge pertaining to the BLV envelope protein in the context of viral infection, replication and pathogenesis. Full article

(This article belongs to the Special Issue Host Membranes and the Viral Infection Cycle)

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