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Volume 12, November

Viruses, Volume 12, Issue 12 (December 2020) – 132 articles

Cover Story (view full-size image): Emerging RNA viruses pose a great threat to human health. Finding treatment strategies and new targets against pathogenic viruses is of high interest globally, not least in the context of the ongoing pandemic. This work presents the identification of new antiviral compounds by phenotypic screening with activity against pathogenic RNA viruses including SARS-CoV-2, Ebola virus, and Crimean-Congo hemorrhagic fever virus. A target identification strategy by thermal protein profiling revealed host factor alterations, particularly in the cellular Hsp70 pathway leading to impaired virus progeny production. Combining phenotypic screening with recently-developed target identification methods has a great potential to advance antiviral drug discovery and reveal new host cell-virus biology. View this paper
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Article
Variations in BK Polyomavirus Immunodominant Large Tumor Antigen-Specific 9mer CD8 T-Cell Epitopes Predict Altered HLA-Presentation and Immune Failure
Viruses 2020, 12(12), 1476; https://doi.org/10.3390/v12121476 - 21 Dec 2020
Cited by 1 | Viewed by 952
Abstract
Failing BK polyomavirus (BKPyV)-specific immune control is underlying onset and duration of BKPyV-replication and disease. We focused on BKPyV-specific CD8 T-cells as key effectors and characterized immunodominant 9mer epitopes in the viral large tumor-antigen (LTag). We investigated the variation of LTag-epitopes and their [...] Read more.
Failing BK polyomavirus (BKPyV)-specific immune control is underlying onset and duration of BKPyV-replication and disease. We focused on BKPyV-specific CD8 T-cells as key effectors and characterized immunodominant 9mer epitopes in the viral large tumor-antigen (LTag). We investigated the variation of LTag-epitopes and their predicted effects on HLA-class 1 binding and T-cell activation. Available BKPyV sequences in the NCBI-nucleotide (N = 3263), and the NCBI protein database (N = 4189) were extracted (1368 sequences) and analyzed for non-synonymous aa-exchanges in LTag. Variant 9mer-epitopes were assessed for predicted changes in HLA-A and HLA-B-binding compared to immunodominant 9mer reference. We identified 159 non-synonymous aa-exchanges in immunodominant LTag-9mer T-cell epitopes reflecting different BKPyV-genotypes as well as genotype-independent variants altering HLA-A/HLA-B-binding scores. Decreased binding scores for HLA-A/HLA-B were found in 27/159 (17%). This included the immunodominant LPLMRKAYL affecting HLA-B*07:02-, HLA-B*08:01- and HLA-B*51:01-presentation. In two healthy BKPyV-seropositive HLA-B*07:02 blood donors, variant LSLMRKAYL showed reduced CD8 T-cell responses compared to LPLMRKAYL. Thus, despite LTag being highly conserved, aa-exchanges occur in immunodominant CD8 T-cell epitopes of BKPyV-genotypes as well as of genotypes -independent variants, which may contribute to genotype-dependent and genotype-independent failure of cellular immune control over BKPyV-replication. The data warrant epidemiological and immunological investigations in carefully designed clinical studies. Full article
(This article belongs to the Special Issue Polyomaviruses)
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Article
The Antimalarial Compound Atovaquone Inhibits Zika and Dengue Virus Infection by Blocking E Protein-Mediated Membrane Fusion
Viruses 2020, 12(12), 1475; https://doi.org/10.3390/v12121475 - 21 Dec 2020
Cited by 4 | Viewed by 1596
Abstract
Flaviviruses bear class II fusion proteins as their envelope (E) proteins. Here, we describe the development of an in vitro quantitative mosquito-cell-based membrane-fusion assay for the E protein using dual split proteins (DSPs). The assay does not involve the use of live viruses [...] Read more.
Flaviviruses bear class II fusion proteins as their envelope (E) proteins. Here, we describe the development of an in vitro quantitative mosquito-cell-based membrane-fusion assay for the E protein using dual split proteins (DSPs). The assay does not involve the use of live viruses and allows the analysis of a membrane-fusion step independent of other events in the viral lifecycle, such as endocytosis. The progress of membrane fusion can be monitored continuously by measuring the activities of Renilla luciferase derived from the reassociation of DSPs during cell fusion. We optimized the assay to screen an FDA-approved drug library for a potential membrane fusion inhibitor using the E protein of Zika virus. Screening results identified atovaquone, which was previously described as an antimalarial agent. Atovaquone potently blocked the in vitro Zika virus infection of mammalian cells with an IC90 of 2.1 µM. Furthermore, four distinct serotypes of dengue virus were also inhibited by atovaquone with IC90 values of 1.6–2.5 µM, which is a range below the average blood concentration of atovaquone after its oral administration in humans. These findings make atovaquone a likely candidate drug to treat illnesses caused by Zika as well as dengue viruses. Additionally, the DSP assay is useful to study the mechanism of membrane fusion in Flaviviruses. Full article
(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)
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Article
Live Attenuated African Swine Fever Viruses as Ideal Tools to Dissect the Mechanisms Involved in Cross-Protection
Viruses 2020, 12(12), 1474; https://doi.org/10.3390/v12121474 - 21 Dec 2020
Cited by 11 | Viewed by 1445
Abstract
African swine fever (ASF) has become the major threat for the global swine industry. Furthermore, the epidemiological situation of African swine fever virus (ASFV) in some endemic regions of Sub-Saharan Africa is worse than ever, with multiple virus strains and genotypes currently circulating [...] Read more.
African swine fever (ASF) has become the major threat for the global swine industry. Furthermore, the epidemiological situation of African swine fever virus (ASFV) in some endemic regions of Sub-Saharan Africa is worse than ever, with multiple virus strains and genotypes currently circulating in a given area. Despite the recent advances on ASF vaccine development, there are no commercial vaccines yet, and most of the promising vaccine prototypes available today have been specifically designed to fight the genotype II strains currently circulating in Europe, Asia, and Oceania. Previous results from our laboratory have demonstrated the ability of BA71∆CD2, a recombinant LAV lacking CD2v, to confer protection against homologous (BA71) and heterologous genotype I (E75) and genotype II (Georgia2007/01) ASFV strains, both belonging to same clade (clade C). Here, we extend these results using BA71∆CD2 as a tool trying to understand ASFV cross-protection, using phylogenetically distant ASFV strains. We first observed that five out of six (83.3%) of the pigs immunized once with 106 PFU of BA71∆CD2 survived the tick-bite challenge using Ornithodoros sp. soft ticks naturally infected with RSA/11/2017 strain (genotype XIX, clade D). Second, only two out of six (33.3%) survived the challenge with Ken06.Bus (genotype IX, clade A), which is phylogenetically more distant to BA71∆CD2 than the RSA/11/2017 strain. On the other hand, homologous prime-boosting with BA71∆CD2 only improved the survival rate to 50% after Ken06.Bus challenge, all suffering mild ASF-compatible clinical signs, while 100% of the pigs immunized with BA71∆CD2 and boosted with the parental BA71 virulent strain survived the lethal challenge with Ken06.Bus, without almost no clinical signs of the disease. Our results confirm that cross-protection is a multifactorial phenomenon that not only depends on sequence similarity. We believe that understanding this complex phenomenon will be useful for designing future vaccines for ASF-endemic areas. Full article
(This article belongs to the Special Issue African Swine Fever Virus 2021)
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Article
The RNA Architecture of the SARS-CoV-2 3′-Untranslated Region
Viruses 2020, 12(12), 1473; https://doi.org/10.3390/v12121473 - 21 Dec 2020
Cited by 16 | Viewed by 2322
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the current COVID-19 pandemic. The 3′ untranslated region (UTR) of this β-CoV contains essential cis-acting RNA elements for the viral genome transcription and replication. These elements include an equilibrium between an extended [...] Read more.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is responsible for the current COVID-19 pandemic. The 3′ untranslated region (UTR) of this β-CoV contains essential cis-acting RNA elements for the viral genome transcription and replication. These elements include an equilibrium between an extended bulged stem-loop (BSL) and a pseudoknot. The existence of such an equilibrium is supported by reverse genetic studies and phylogenetic covariation analysis and is further proposed as a molecular switch essential for the control of the viral RNA polymerase binding. Here, we report the SARS-CoV-2 3′ UTR structures in cells that transcribe the viral UTRs harbored in a minigene plasmid and isolated infectious virions using a chemical probing technique, namely dimethyl sulfate (DMS)-mutational profiling with sequencing (MaPseq). Interestingly, the putative pseudoknotted conformation was not observed, indicating that its abundance in our systems is low in the absence of the viral nonstructural proteins (nsps). Similarly, our results also suggest that another functional cis-acting element, the three-helix junction, cannot stably form. The overall architectures of the viral 3′ UTRs in the infectious virions and the minigene-transfected cells are almost identical. Full article
(This article belongs to the Section SARS-CoV-2 and COVID-19)
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Article
Identification of an Intermediate Step in Foamy Virus Fusion
Viruses 2020, 12(12), 1472; https://doi.org/10.3390/v12121472 - 21 Dec 2020
Cited by 1 | Viewed by 1102
Abstract
Viral glycoprotein-mediated membrane fusion is an essential step for productive infection of host cells by enveloped viruses; however, due to its rarity and challenges in detection, little is known about the details of fusion events at the single particle level. Here, we have [...] Read more.
Viral glycoprotein-mediated membrane fusion is an essential step for productive infection of host cells by enveloped viruses; however, due to its rarity and challenges in detection, little is known about the details of fusion events at the single particle level. Here, we have developed dual-color foamy viruses (FVs) composed of eGFP-tagged prototype FV (PFV) Gag and mCherry-tagged Env of either PFV or macaque simian FV (SFVmac) origin that have been optimized for detection of the fusion process. Using our recently developed tracking imaging correlation (TrIC) analysis, we were able to detect the fusion process for both PFV and SFVmac Env containing virions. PFV Env-mediated fusion was observed both at the plasma membrane as well as from endosomes, whereas SFVmac Env-mediated fusion was only observed from endosomes. PFV Env-mediated fusion was observed to happen more often and more rapidly than as for SFVmac Env. Strikingly, using the TrIC method, we detected a novel intermediate state where the envelope and capsids are still tethered but separated by up to 400 nm before final separation of Env and Gag occurred. Full article
(This article belongs to the Special Issue Mechanisms of Viral Fusion and Applications in Antivirals)
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Article
Can Coronaviruses Steal Genes from the Host as Evidenced in Western European Hedgehogs by EriCoV Genetic Characterization?
Viruses 2020, 12(12), 1471; https://doi.org/10.3390/v12121471 - 20 Dec 2020
Cited by 2 | Viewed by 2852
Abstract
Due to their need for living cells, viruses have developed adaptive evolutionary strategies to survive and perpetuate in reservoir hosts that play a crucial role in the ecology of emerging pathogens. Pathogenic and potentially pandemic betacoronaviruses arose in humans in 2002 (SARS-CoV, disappeared [...] Read more.
Due to their need for living cells, viruses have developed adaptive evolutionary strategies to survive and perpetuate in reservoir hosts that play a crucial role in the ecology of emerging pathogens. Pathogenic and potentially pandemic betacoronaviruses arose in humans in 2002 (SARS-CoV, disappeared in July 2003), 2012 (MERS-CoV, still circulating in Middle East areas), and 2019 (SARS-CoV-2, causing the current global pandemic). As universally recognized, bats host ancestors of the above-mentioned zoonotic viruses. However, hedgehogs have been recently identified in Europe and Asia as possible reservoirs of MERS-CoV-like strains classified as Erinaceus coronavirus (EriCoV). To elucidate the evolution and genetics of EriCoVs, NGS (next generation sequencing) and Sanger sequencing were used to examine fecal samples collected in Northern Italy in 2018/2019 from 12 hedgehogs previously found EriCoV-positive by RT-PCR. By sequence analysis, eight complete EriCoV genomes, obtained by NGS, showed a high phylogenetic correlation with EriCoV strains previously reported in Eurasia. Interestingly, eight viral strains presented an additional ORF encoding for the CD200 ortholog located between the genes encoding for the Spike and the ORF3a proteins. The CD200 ortholog sequences were closely similar to the host CD200 protein but varying among EriCoVs. The result, confirmed by Sanger sequencing, demonstrates for the first time that CoVs can acquire host genes potentially involved in the immune-modulatory cascade and possibly enabling the virus to escape the host defence. Full article
(This article belongs to the Special Issue Drivers of Evolution of Animal RNA Viruses)
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Article
In Vitro Evaluation of a Phage Cocktail Controlling Infections with Escherichia coli
Viruses 2020, 12(12), 1470; https://doi.org/10.3390/v12121470 - 19 Dec 2020
Cited by 8 | Viewed by 1779
Abstract
Worldwide, poultry industry suffers from infections caused by avian pathogenic Escherichia coli. Therapeutic failure due to resistant bacteria is of increasing concern and poses a threat to human and animal health. This causes a high demand to find alternatives to fight bacterial [...] Read more.
Worldwide, poultry industry suffers from infections caused by avian pathogenic Escherichia coli. Therapeutic failure due to resistant bacteria is of increasing concern and poses a threat to human and animal health. This causes a high demand to find alternatives to fight bacterial infections in animal farming. Bacteriophages are being especially considered for the control of multi-drug resistant bacteria due to their high specificity and lack of serious side effects. Therefore, the study aimed on characterizing phages and composing a phage cocktail suitable for the prevention of infections with E. coli. Six phages were isolated or selected from our collections and characterized individually and in combination with regard to host range, stability, reproduction, and efficacy in vitro. The cocktail consisting of six phages was able to inhibit formation of biofilms by some E. coli strains but not by all. Phage-resistant variants arose when bacterial cells were challenged with a single phage but not when challenged by a combination of four or six phages. Resistant variants arising showed changes in carbon metabolism and/or motility. Genomic comparison of wild type and phage-resistant mutant E28.G28R3 revealed a deletion of several genes putatively involved in phage adsorption and infection. Full article
(This article belongs to the Section Bacterial Viruses)
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Review
HIV Vaccine Development: 35 Years of Experimenting in the Funding of Biomedical Research
Viruses 2020, 12(12), 1469; https://doi.org/10.3390/v12121469 - 19 Dec 2020
Cited by 1 | Viewed by 1556
Abstract
Funding vaccine development research is more complicated than simply putting out an announcement of funds available. The funders must decide whether product development can be accomplished by purely applied research, or whether more fundamental knowledge is needed before product development can be started. [...] Read more.
Funding vaccine development research is more complicated than simply putting out an announcement of funds available. The funders must decide whether product development can be accomplished by purely applied research, or whether more fundamental knowledge is needed before product development can be started. If additional basic knowledge is needed, identifying the specific area of the knowledge gap can be a challenge. Additionally, when there appears to be a clear path of applied research sometimes obstacles are encountered that require a return to more basic work. After deciding on the work to be done, funders must attract the scientists with the broad range of needed skills to cover all the stages of development. Collaborations must be promoted and alliances with other funders and industry must be developed. Funders use multiple tools and strategies to accomplish these tasks with varying success. Full article
(This article belongs to the Special Issue HIV and SARS-CoV-2 Pathogenesis and Vaccine Development)
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Article
Potentially Infectious Novel Hepatitis A Virus Strains Detected in Selected Treated Wastewater Discharge Sources, South Africa
Viruses 2020, 12(12), 1468; https://doi.org/10.3390/v12121468 - 19 Dec 2020
Cited by 2 | Viewed by 951
Abstract
Hepatitis A virus (HAV) is a waterborne pathogen of public health importance. In South Africa (SA), unique HAV subgenotype IB strains have been detected in surface and wastewater samples, as well as on fresh produce at the point of retail. However, due to [...] Read more.
Hepatitis A virus (HAV) is a waterborne pathogen of public health importance. In South Africa (SA), unique HAV subgenotype IB strains have been detected in surface and wastewater samples, as well as on fresh produce at the point of retail. However, due to the use of molecular-based assays, the infectivity of the detected strains was unknown. Considering the potential shift of HAV endemicity from high to intermediate, which could increase the risk of severe symptomatic disease, this study investigated the identity of HAV strains detected before and after viability treatment of selected wastewater discharge samples. For one year, 118 samples consisting of sewage, treated wastewater discharge and downstream dam water were collected from five wastewater treatment plants (WWTP 1, 2, 3, 4 and 5). Unique HAV IB strains were detected in samples from all five WWTPs, with 11 of these strains carrying amino acid mutations at the immunodominant and neutralisation epitopes. A quasispecies dynamic of HAV has also been detected in sewage samples. The subsequent application of viability PCR revealed that potentially infectious HAV strains were discharged from WWTP 1, 2, 4 and 5 into the dam. Therefore, there is a potential risk of HAV exposure to communities using water sources downstream the WWTPs. Full article
(This article belongs to the Special Issue Recent Progress in Hepatitis A Virus Research)
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Article
Enhanced Cell-Based Detection of Parvovirus B19V Infectious Units According to Cell Cycle Status
Viruses 2020, 12(12), 1467; https://doi.org/10.3390/v12121467 - 18 Dec 2020
Cited by 1 | Viewed by 1058
Abstract
Human parvovirus B19 (B19V) causes various human diseases, ranging from childhood benign infection to arthropathies, severe anemia and fetal hydrops, depending on the health state and hematological status of the patient. To counteract B19V blood-borne contamination, evaluation of B19 DNA in plasma pools [...] Read more.
Human parvovirus B19 (B19V) causes various human diseases, ranging from childhood benign infection to arthropathies, severe anemia and fetal hydrops, depending on the health state and hematological status of the patient. To counteract B19V blood-borne contamination, evaluation of B19 DNA in plasma pools and viral inactivation/removal steps are performed, but nucleic acid testing does not correctly reflect B19V infectivity. There is currently no appropriate cellular model for detection of infectious units of B19V. We describe here an improved cell-based method for detecting B19V infectious units by evaluating its host transcription. We evaluated the ability of various cell lines to support B19V infection. Of all tested, UT7/Epo cell line, UT7/Epo-STI, showed the greatest sensitivity to B19 infection combined with ease of performance. We generated stable clones by limiting dilution on the UT7/Epo-STI cell line with graduated permissiveness for B19V and demonstrated a direct correlation between infectivity and S/G2/M cell cycle stage. Two of the clones tested, B12 and E2, reached sensitivity levels higher than those of UT7/Epo-S1 and CD36+ erythroid progenitor cells. These findings highlight the importance of cell cycle status for sensitivity to B19V, and we propose a promising new straightforward cell-based method for quantifying B19V infectious units. Full article
(This article belongs to the Special Issue Advances in Parvovirus Research 2020)
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Article
An Amphipathic Alpha-Helix Domain from Poliovirus 2C Protein Tubulate Lipid Vesicles
Viruses 2020, 12(12), 1466; https://doi.org/10.3390/v12121466 - 18 Dec 2020
Cited by 2 | Viewed by 877
Abstract
Positive-strand RNA viruses universally remodel host intracellular membranes to form membrane-bound viral replication complexes, where viral offspring RNAs are synthesized. In the majority of cases, viral replication proteins are targeted to and play critical roles in the modulation of the designated organelle membranes. [...] Read more.
Positive-strand RNA viruses universally remodel host intracellular membranes to form membrane-bound viral replication complexes, where viral offspring RNAs are synthesized. In the majority of cases, viral replication proteins are targeted to and play critical roles in the modulation of the designated organelle membranes. Many viral replication proteins do not have transmembrane domains, but contain single or multiple amphipathic alpha-helices. It has been conventionally recognized that these helices serve as an anchor for viral replication protein to be associated with membranes. We report here that a peptide representing the amphipathic α-helix at the N-terminus of the poliovirus 2C protein not only binds to liposomes, but also remodels spherical liposomes into tubules. The membrane remodeling ability of this amphipathic alpha-helix is similar to that recognized in other amphipathic alpha-helices from cellular proteins involved in membrane remodeling, such as BAR domain proteins. Mutations affecting the hydrophobic face of the amphipathic alpha-helix severely compromised membrane remodeling of vesicles with physiologically relevant phospholipid composition. These mutations also affected the ability of poliovirus to form plaques indicative of reduced viral replication, further underscoring the importance of membrane remodeling by the amphipathic alpha-helix in possible relation to the formation of viral replication complexes. Full article
(This article belongs to the Section Animal Viruses)
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Article
SARS-CoV-2 Spike Alterations Enhance Pseudoparticle Titers and Replication-Competent VSV-SARS-CoV-2 Virus
Viruses 2020, 12(12), 1465; https://doi.org/10.3390/v12121465 - 18 Dec 2020
Cited by 14 | Viewed by 1999
Abstract
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the most recent global pandemic that has caused more than a million deaths around the world. The spike glycoprotein (S) drives the entry and fusion of this virus and is the [...] Read more.
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the causative agent of the most recent global pandemic that has caused more than a million deaths around the world. The spike glycoprotein (S) drives the entry and fusion of this virus and is the main determinant of cell tropism. To explore S requirements for entry under BSL2 conditions, S has been pseudotyped onto vesicular stomatitis virus (VSV) or retroviral particles with varied success. Several alterations to S were demonstrated to improve pseudoparticle titers, but they have not been systematically compared. In this study, we produced pseudotyped VSV particles with multiple modifications to S, including truncation, mutation, and tagging strategies. The main objective of this study was to determine which modifications of the S protein optimize cell surface expression, incorporation into pseudotyped particles, and pseudoparticle entry. Removal of the last 19 residues of the cytoplasmic tail produced a hyper-fusogenic S, while removal of 21 residues increased S surface production and VSV incorporation. Additionally, we engineered a replication-competent VSV (rVSV) virus to produce the S-D614G variant with a truncated cytoplasmic tail. While the particles can be used to assess S entry requirements, the rVSV∆G/SMet1D614G∆21 virus has a poor specific infectivity (particle to infectious titer ratio). Full article
(This article belongs to the Section SARS-CoV-2 and COVID-19)
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Review
Emerging Role of PYHIN Proteins as Antiviral Restriction Factors
Viruses 2020, 12(12), 1464; https://doi.org/10.3390/v12121464 - 18 Dec 2020
Cited by 2 | Viewed by 1528
Abstract
Innate immune sensors and restriction factors are cellular proteins that synergize to build an effective first line of defense against viral infections. Innate sensors are usually constitutively expressed and capable of detecting pathogen-associated molecular patterns (PAMPs) via specific pattern recognition receptors (PRRs) to [...] Read more.
Innate immune sensors and restriction factors are cellular proteins that synergize to build an effective first line of defense against viral infections. Innate sensors are usually constitutively expressed and capable of detecting pathogen-associated molecular patterns (PAMPs) via specific pattern recognition receptors (PRRs) to stimulate the immune response. Restriction factors are frequently upregulated by interferons (IFNs) and may inhibit viral pathogens at essentially any stage of their replication cycle. Members of the Pyrin and hematopoietic interferon-inducible nuclear (HIN) domain (PYHIN) family have initially been recognized as important sensors of foreign nucleic acids and activators of the inflammasome and the IFN response. Accumulating evidence shows, however, that at least three of the four members of the human PYHIN family restrict viral pathogens independently of viral sensing and innate immune activation. In this review, we provide an overview on the role of human PYHIN proteins in the innate antiviral immune defense and on viral countermeasures. Full article
(This article belongs to the Special Issue Intrinsic Antiviral Factors)
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Review
The VP1u of Human Parvovirus B19: A Multifunctional Capsid Protein with Biotechnological Applications
Viruses 2020, 12(12), 1463; https://doi.org/10.3390/v12121463 - 18 Dec 2020
Cited by 3 | Viewed by 1254
Abstract
The viral protein 1 unique region (VP1u) of human parvovirus B19 (B19V) is a multifunctional capsid protein with essential roles in virus tropism, uptake, and subcellular trafficking. These functions reside on hidden protein domains, which become accessible upon interaction with cell membrane receptors. [...] Read more.
The viral protein 1 unique region (VP1u) of human parvovirus B19 (B19V) is a multifunctional capsid protein with essential roles in virus tropism, uptake, and subcellular trafficking. These functions reside on hidden protein domains, which become accessible upon interaction with cell membrane receptors. A receptor-binding domain (RBD) in VP1u is responsible for the specific targeting and uptake of the virus exclusively into cells of the erythroid lineage in the bone marrow. A phospholipase A2 domain promotes the endosomal escape of the incoming virus. The VP1u is also the immunodominant region of the capsid as it is the target of neutralizing antibodies. For all these reasons, the VP1u has raised great interest in antiviral research and vaccinology. Besides the essential functions in B19V infection, the remarkable erythroid specificity of the VP1u makes it a unique erythroid cell surface biomarker. Moreover, the demonstrated capacity of the VP1u to deliver diverse cargo specifically to cells around the proerythroblast differentiation stage, including erythroleukemic cells, offers novel therapeutic opportunities for erythroid-specific drug delivery. In this review, we focus on the multifunctional role of the VP1u in B19V infection and explore its potential in diagnostics and erythroid-specific therapeutics. Full article
(This article belongs to the Special Issue Advances in Parvovirus Research 2020)
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Perspective
Homo sapiens: The Superspreader of Plant Viral Diseases
Viruses 2020, 12(12), 1462; https://doi.org/10.3390/v12121462 - 17 Dec 2020
Cited by 2 | Viewed by 1303
Abstract
Plant viruses are commonly vectored by flying or crawling animals, such as aphids and beetles, and cause serious losses in major agricultural and horticultural crops. Controlling virus spread is often achieved by minimizing a crop’s exposure to the vector, or by reducing vector [...] Read more.
Plant viruses are commonly vectored by flying or crawling animals, such as aphids and beetles, and cause serious losses in major agricultural and horticultural crops. Controlling virus spread is often achieved by minimizing a crop’s exposure to the vector, or by reducing vector numbers with compounds such as insecticides. A major, but less obvious, factor not controlled by these measures is Homo sapiens. Here, we discuss the inconvenient truth of how humans have become superspreaders of plant viruses on both a local and a global scale. Full article
(This article belongs to the Special Issue Plant Virus Emergence)
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Article
Influenza A H1 and H3 Transmembrane Domains Interact Differently with Each Other and with Surrounding Membrane Lipids
Viruses 2020, 12(12), 1461; https://doi.org/10.3390/v12121461 - 17 Dec 2020
Cited by 5 | Viewed by 1552
Abstract
Hemagglutinin (HA) is a class I viral membrane fusion protein, which is the most abundant transmembrane protein on the surface of influenza A virus (IAV) particles. HA plays a crucial role in the recognition of the host cell, fusion of the viral envelope [...] Read more.
Hemagglutinin (HA) is a class I viral membrane fusion protein, which is the most abundant transmembrane protein on the surface of influenza A virus (IAV) particles. HA plays a crucial role in the recognition of the host cell, fusion of the viral envelope and the host cell membrane, and is the major antigen in the immune response during the infection. Mature HA organizes in homotrimers consisting of a sequentially highly variable globular head and a relatively conserved stalk region. Every HA monomer comprises a hydrophilic ectodomain, a pre-transmembrane domain (pre-TMD), a hydrophobic transmembrane domain (TMD), and a cytoplasmic tail (CT). In recent years the effect of the pre-TMD and TMD on the structure and function of HA has drawn some attention. Using bioinformatic tools we analyzed all available full-length amino acid sequences of HA from 16 subtypes across various host species. We calculated several physico-chemical parameters of HA pre-TMDs and TMDs including accessible surface area (ASA), average hydrophobicity (Hav), and the hydrophobic moment (µH). Our data suggests that distinct differences in these parameters between the two major phylogenetic groups, represented by H1 and H3 subtypes, could have profound effects on protein–lipid interactions, trimer formation, and the overall HA ectodomain orientation and antigen exposure. Full article
(This article belongs to the Section Viral Immunology, Vaccines, and Antivirals)
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Article
Integrase-Defective Lentiviral Vectors for Delivery of Monoclonal Antibodies against Influenza
Viruses 2020, 12(12), 1460; https://doi.org/10.3390/v12121460 - 17 Dec 2020
Cited by 1 | Viewed by 1547
Abstract
Delivering rapid protection against infectious agents to non-immune populations is a formidable public health challenge. Although passive immunotherapy is a fast and effective method of protection, large-scale production and administration of monoclonal antibodies (mAbs) is expensive and unpractical. Viral vector-mediated delivery of mAbs [...] Read more.
Delivering rapid protection against infectious agents to non-immune populations is a formidable public health challenge. Although passive immunotherapy is a fast and effective method of protection, large-scale production and administration of monoclonal antibodies (mAbs) is expensive and unpractical. Viral vector-mediated delivery of mAbs offers an attractive alternative to their direct injection. Integrase-defective lentiviral vectors (IDLV) are advantageous for this purpose due to the absence of pre-existing anti-vector immunity and the safety features of non-integration and non-replication. We engineered IDLV to produce the humanized mAb VN04-2 (IDLV-VN04-2), which is broadly neutralizing against H5 influenza A virus (IAV), and tested the vectors’ ability to produce antibodies and protect from IAV in vivo. We found that IDLV-transduced cells produced functional VN04-2 mAbs in a time- and dose-dependent fashion. These mAbs specifically bind the hemagglutinin (HA), but not the nucleoprotein (NP) of IAV. VN04-2 mAbs were detected in the serum of mice at different times after intranasal (i.n.) or intramuscular (i.m.) administration of IDLV-VN04-2. Administration of IDLV-VN04-2 by the i.n. route provided rapid protection against lethal IAV challenge, although the protection did not persist at later time points. Our data suggest that administration of mAb-expressing IDLV may represent an effective strategy for rapid protection against infectious diseases. Full article
(This article belongs to the Special Issue Lentiviral Vectors)
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Article
Significantly Improved Recovery of Recombinant Sonchus Yellow Net Rhabdovirus by Expressing the Negative-Strand Genomic RNA
Viruses 2020, 12(12), 1459; https://doi.org/10.3390/v12121459 - 17 Dec 2020
Cited by 6 | Viewed by 1012
Abstract
Generation of recombinant negative-stranded RNA viruses (NSVs) from plasmids involves in vivo reconstitution of biologically active nucleocapsids and faces a unique antisense problem where the negative-sense viral genomic RNAs can hybridize to viral messenger RNAs. To overcome this problem, a positive-sense RNA approach [...] Read more.
Generation of recombinant negative-stranded RNA viruses (NSVs) from plasmids involves in vivo reconstitution of biologically active nucleocapsids and faces a unique antisense problem where the negative-sense viral genomic RNAs can hybridize to viral messenger RNAs. To overcome this problem, a positive-sense RNA approach has been devised through expression of viral antigenomic (ag)RNA and core proteins for assembly of antigenomic nucleocapsids. Although this detour strategy works for many NSVs, the process is still inefficient. Using Sonchus yellow net rhabdovirus (SYNV) as a model; here, we develop a negative-sense genomic RNA-based approach that increased rescue efficiency by two orders of magnitude compared to the conventional agRNA approach. The system relied on suppression of double-stranded RNA induced antiviral responses by co-expression of plant viruses-encoded RNA silencing suppressors or animal viruses-encoded double-stranded RNA antagonists. With the improved approach, we were able to recover a highly attenuated SYNV mutant with a deletion in the matrix protein gene which otherwise could not be rescued via the agRNA approach. Reverse genetics analyses of the generated mutant virus provided insights into SYNV virion assembly and morphogenesis. This approach may potentially be applicable to other NSVs of plants or animals. Full article
(This article belongs to the Section Viruses of Plants, Fungi and Protozoa)
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Review
Resveratrol, Rapamycin and Metformin as Modulators of Antiviral Pathways
Viruses 2020, 12(12), 1458; https://doi.org/10.3390/v12121458 - 17 Dec 2020
Cited by 6 | Viewed by 2096
Abstract
Balanced nutrition and appropriate dietary interventions are fundamental in the prevention and management of viral infections. Additionally, accurate modulation of the inflammatory response is necessary to achieve an adequate antiviral immune response. Many studies, both in vitro with mammalian cells and in vivo [...] Read more.
Balanced nutrition and appropriate dietary interventions are fundamental in the prevention and management of viral infections. Additionally, accurate modulation of the inflammatory response is necessary to achieve an adequate antiviral immune response. Many studies, both in vitro with mammalian cells and in vivo with small animal models, have highlighted the antiviral properties of resveratrol, rapamycin and metformin. The current review outlines the mechanisms of action of these three important compounds on the cellular pathways involved with viral replication and the mechanisms of virus-related diseases, as well as the current status of their clinical use. Full article
(This article belongs to the Special Issue Unconventional Antiviral Agents)
Article
Monitoring Viral Entry in Real-Time Using a Luciferase Recombinant Vesicular Stomatitis Virus Producing SARS-CoV-2, EBOV, LASV, CHIKV, and VSV Glycoproteins
Viruses 2020, 12(12), 1457; https://doi.org/10.3390/v12121457 - 17 Dec 2020
Cited by 9 | Viewed by 2208
Abstract
Viral entry is the first stage in the virus replication cycle and, for enveloped viruses, is mediated by virally encoded glycoproteins. Viral glycoproteins have different receptor affinities and triggering mechanisms. We employed vesicular stomatitis virus (VSV), a BSL-2 enveloped virus that can incorporate [...] Read more.
Viral entry is the first stage in the virus replication cycle and, for enveloped viruses, is mediated by virally encoded glycoproteins. Viral glycoproteins have different receptor affinities and triggering mechanisms. We employed vesicular stomatitis virus (VSV), a BSL-2 enveloped virus that can incorporate non-native glycoproteins, to examine the entry efficiencies of diverse viral glycoproteins. To compare the glycoprotein-mediated entry efficiencies of VSV glycoprotein (G), Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) spike (S), Ebola (EBOV) glycoprotein (GP), Lassa (LASV) GP, and Chikungunya (CHIKV) envelope (E) protein, we produced recombinant VSV (rVSV) viruses that produce the five glycoproteins. The rVSV virions encoded a nano luciferase (NLucP) reporter gene fused to a destabilization domain (PEST), which we used in combination with the live-cell substrate EndurazineTM to monitor viral entry kinetics in real time. Our data indicate that rVSV particles with glycoproteins that require more post-internalization priming typically demonstrate delayed entry in comparison to VSV G. In addition to determining the time required for each virus to complete entry, we also used our system to evaluate viral cell surface receptor preferences, monitor fusion, and elucidate endocytosis mechanisms. This system can be rapidly employed to examine diverse viral glycoproteins and their entry requirements. Full article
(This article belongs to the Section SARS-CoV-2 and COVID-19)
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Review
Application of a Sanger-Based External Quality Assurance Strategy for the Transition of HIV-1 Drug Resistance Assays to Next Generation Sequencing
Viruses 2020, 12(12), 1456; https://doi.org/10.3390/v12121456 - 17 Dec 2020
Cited by 1 | Viewed by 908
Abstract
The National Institute of Allergy and Infectious Diseases (NIAID) Virology Quality Assurance (VQA) established a robust proficiency testing program for Sanger sequencing (SS)-based HIV-1 drug resistance (HIVDR) testing in 2001. While many of the lessons learned during the development of such programs may [...] Read more.
The National Institute of Allergy and Infectious Diseases (NIAID) Virology Quality Assurance (VQA) established a robust proficiency testing program for Sanger sequencing (SS)-based HIV-1 drug resistance (HIVDR) testing in 2001. While many of the lessons learned during the development of such programs may also apply to next generation sequencing (NGS)-based HIVDR assays, challenges remain for the ongoing evaluation of NGS-based testing. These challenges include a proper assessment of assay accuracy and the reproducibility of low abundance variant detection, intra- and inter-assay performance comparisons among laboratories using lab-defined tests, and different data analysis pipelines designed for NGS. In collaboration with the World Health Organization (WHO) Global HIVDR Laboratory Network and the Public Health Agency of Canada, the Rush VQA program distributed archived proficiency testing panels to ten laboratories to evaluate internally developed NGS assays. Consensus FASTA files were submitted using 5%, 10%, and 20% variant detection thresholds, and scored based on the same criteria used for SS. This small study showed that the SS External Quality Assurance (EQA) approach can be used as a transitional strategy for using NGS to generate SS-like data and for ongoing performance while using NGS data from the same quality control materials to further evaluate NGS assay performance. Full article
(This article belongs to the Special Issue Next Generation Sequencing for HIV Drug Resistance Testing)
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Review
The Unknown Unknowns: Recovering Gamma-Delta T Cells for Control of Human Immunodeficiency Virus (HIV)
Viruses 2020, 12(12), 1455; https://doi.org/10.3390/v12121455 - 17 Dec 2020
Cited by 2 | Viewed by 1240
Abstract
Recent advances in γδ T cell biology have focused on the unique attributes of these cells and their role in regulating innate and adaptive immunity, promoting tissue homeostasis, and providing resistance to various disorders. Numerous bacterial and viral pathogens, including human immunodeficiency virus-1 [...] Read more.
Recent advances in γδ T cell biology have focused on the unique attributes of these cells and their role in regulating innate and adaptive immunity, promoting tissue homeostasis, and providing resistance to various disorders. Numerous bacterial and viral pathogens, including human immunodeficiency virus-1 (HIV), greatly alter the composition of γδ T cells in vivo. Despite the effectiveness of antiretroviral therapy (ART) in controlling HIV and restoring health in those affected, γδ T cells are dramatically impacted during HIV infection and fail to reconstitute to normal levels in HIV-infected individuals during ART for reasons that are not clearly understood. Importantly, their role in controlling HIV infection, and the implications of their failure to rebound during ART are also largely unknown and understudied. Here, we review important aspects of human γδ T cell biology, the effector and immunomodulatory properties of these cells, their prevalence and function in HIV, and their immunotherapeutic potential. Full article
(This article belongs to the Special Issue HIV Infection and Spread between T Cells)
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Review
Exposure Risk of Chronic Wasting Disease in Humans
Viruses 2020, 12(12), 1454; https://doi.org/10.3390/v12121454 - 17 Dec 2020
Cited by 4 | Viewed by 1932
Abstract
The majority of human prion diseases are sporadic, but acquired disease can occur, as seen with variant Creutzfeldt–Jakob disease (vCJD) following consumption of bovine spongiform encephalopathy (BSE). With increasing rates of cervid chronic wasting disease (CWD), there is concern that a new form [...] Read more.
The majority of human prion diseases are sporadic, but acquired disease can occur, as seen with variant Creutzfeldt–Jakob disease (vCJD) following consumption of bovine spongiform encephalopathy (BSE). With increasing rates of cervid chronic wasting disease (CWD), there is concern that a new form of human prion disease may arise. Currently, there is no evidence of transmission of CWD to humans, suggesting the presence of a strong species barrier; however, in vitro and in vivo studies on the zoonotic potential of CWD have yielded mixed results. The emergence of different CWD strains is also concerning, as different strains can have different abilities to cross species barriers. Given that venison consumption is common in areas where CWD rates are on the rise, increased rates of human exposure are inevitable. If CWD was to infect humans, it is unclear how it would present clinically; in vCJD, it was strain-typing of vCJD prions that proved the causal link to BSE. Therefore, the best way to screen for CWD in humans is to have thorough strain-typing of harvested cervids and human CJD cases so that we will be in a position to detect atypical strains or strain shifts within the human CJD population. Full article
(This article belongs to the Special Issue The Future of the Chronic Wasting Disease Epizootic)
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Brief Report
Genetic Diversity of Rift Valley Fever Strains Circulating in Namibia in 2010 and 2011
Viruses 2020, 12(12), 1453; https://doi.org/10.3390/v12121453 - 16 Dec 2020
Cited by 2 | Viewed by 869
Abstract
Outbreaks of Rift Valley fever (RVF) occurred in Namibia in 2010 and 2011. Complete genome characterization was obtained from virus isolates collected during disease outbreaks in southern Namibia in 2010 and from wildlife in Etosha National Park in 2011, close to the area [...] Read more.
Outbreaks of Rift Valley fever (RVF) occurred in Namibia in 2010 and 2011. Complete genome characterization was obtained from virus isolates collected during disease outbreaks in southern Namibia in 2010 and from wildlife in Etosha National Park in 2011, close to the area where RVF outbreaks occurred in domestic livestock. The virus strains were sequenced using Sanger sequencing (Namibia_2010) or next generation sequencing (Namibia_2011). A sequence-independent, single-primer amplification (SISPA) protocol was used in combination with the Illumina Next 500 sequencer. Phylogenetic analysis of the sequences of the small (S), medium (M), and large (L) genome segments of RVF virus (RVFV) provided evidence that two distinct RVFV strains circulated in the country. The strain collected in Namibia in 2010 is genetically similar to RVFV strains circulating in South Africa in 2009 and 2010, confirming that the outbreaks reported in the southern part of Namibia in 2010 were caused by possible dissemination of the infection from South Africa. Isolates collected in 2011 were close to RVFV isolates from 2010 collected in humans in Sudan and which belong to the large lineage containing RVFV strains that caused an outbreak in 2006–2008 in eastern Africa. This investigation showed that the RVFV strains circulating in Namibia in 2010 and 2011 were from two different introductions and that RVFV has the ability to move across regions. This supports the need for risk-based surveillance and monitoring. Full article
(This article belongs to the Special Issue Drivers of Evolution of Animal RNA Viruses)
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Article
Discovery and Characterization of a Novel Ampelovirus on Firespike
Viruses 2020, 12(12), 1452; https://doi.org/10.3390/v12121452 - 16 Dec 2020
Cited by 2 | Viewed by 766
Abstract
A novel RNA virus was identified in firespike (Odontonema tubaeforme) plants exhibiting leaf curling and chlorosis. The molecular features of the viral genomic RNA and proteins resemble those of ampeloviruses. Based on sequence comparisons and phylogenetic analysis, we propose a new [...] Read more.
A novel RNA virus was identified in firespike (Odontonema tubaeforme) plants exhibiting leaf curling and chlorosis. The molecular features of the viral genomic RNA and proteins resemble those of ampeloviruses. Based on sequence comparisons and phylogenetic analysis, we propose a new species in the genus Ampelovirus, which we have tentatively named Firespike leafroll-associated virus (FLRaV). Bioassays showed that the virus is mechanically transmissible to Nicotiana benthamiana. In addition, a full-length cDNA clone of FLRaV could successfully infect N. benthamiana via agroinfiltration. Full article
(This article belongs to the Special Issue Closteroviridae)
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Article
Mining the Unmapped Reads in Bovine RNA-Seq Data Reveals the Prevalence of Bovine Herpes Virus-6 in European Dairy Cows and the Associated Changes in Their Phenotype and Leucocyte Transcriptome
Viruses 2020, 12(12), 1451; https://doi.org/10.3390/v12121451 - 16 Dec 2020
Cited by 2 | Viewed by 886
Abstract
Microbial RNA is detectable in host samples by aligning unmapped reads from RNA sequencing against taxon reference sequences, generating a score proportional to the microbial load. An RNA-Seq data analysis showed that 83.5% of leukocyte samples from six dairy herds in different EU [...] Read more.
Microbial RNA is detectable in host samples by aligning unmapped reads from RNA sequencing against taxon reference sequences, generating a score proportional to the microbial load. An RNA-Seq data analysis showed that 83.5% of leukocyte samples from six dairy herds in different EU countries contained bovine herpes virus-6 (BoHV-6). Phenotypic data on milk production, metabolic function, and disease collected during their first 50 days in milk (DIM) were compared between cows with low (1–200 and n = 114) or high (201–1175 and n = 24) BoHV-6 scores. There were no differences in milk production parameters, but high score cows had numerically fewer incidences of clinical mastitis (4.2% vs. 12.2%) and uterine disease (54.5% vs. 62.7%). Their metabolic status was worse, based on measurements of IGF-1 and various metabolites in blood and milk. A comparison of the global leukocyte transcriptome between high and low BoHV-6 score cows at around 14 DIM yielded 485 differentially expressed genes (DEGs). The top pathway from Gene Ontology (GO) enrichment analysis was the immune system process. Down-regulated genes in the high BoHV-6 cows included those encoding proteins involved in viral detection (DDX6 and DDX58), interferon response, and E3 ubiquitin ligase activity. This suggested that BoHV-6 may largely evade viral detection and that it does not cause clinical disease in dairy cows. Full article
(This article belongs to the Section Animal Viruses)
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Communication
Sea Lice (Lepeophtheirus salmonis) Infestation Reduces the Ability of Peripheral Blood Monocytic Cells (PBMCs) to Respond to and Control Replication of Salmonid Alphavirus in Atlantic Salmon (Salmo salar L.)
Viruses 2020, 12(12), 1450; https://doi.org/10.3390/v12121450 - 16 Dec 2020
Cited by 1 | Viewed by 618
Abstract
Here we have studied the impact of lice (Lepeophtheirus salmonis) infestation of donor fish on the ability of isolated peripheral blood monocytes (PBMCs) to control the replication of salmonid alphavirus (SAV) ex vivo. PBMCs were collected by Percoll gradients at eight [...] Read more.
Here we have studied the impact of lice (Lepeophtheirus salmonis) infestation of donor fish on the ability of isolated peripheral blood monocytes (PBMCs) to control the replication of salmonid alphavirus (SAV) ex vivo. PBMCs were collected by Percoll gradients at eight and nine weeks post copepodid infestation of Atlantic salmon post smolt. Uninfested fish were controls. PBMCs were then infected ex vivo with SAV (subtype 3), and samples were collected for analysis at two, four, and six days post virus infection. Virus titer in the supernatant was assayed in CHH-1 cells, and in addition, the relative expression of the virus structural protein E2 and selected host antiviral genes, IRF9, ISG15, Mx, and IFIT5, were assayed using real-time PCR. Significantly higher virus replication was detected in cells collected from lice-infested fish compared to controls. Higher virus titer coincided with an inability to upregulate the expression of different immune genes, IFIT5, IRF9, and Mx. These findings point towards compromised ability of PBMCs from lice-infested fish to control virus replication, and, to our knowledge, is the first report showing the direct effect of lice infestation on the interplay between viruses and immune cells. There is a possible impact on the dynamic spread of viral diseases in the aquatic environment. Full article
(This article belongs to the Section Animal Viruses)
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Article
Autochthonous Transmission of West Nile Virus by a New Vector in Iran, Vector-Host Interaction Modeling and Virulence Gene Determinants
Viruses 2020, 12(12), 1449; https://doi.org/10.3390/v12121449 - 16 Dec 2020
Cited by 2 | Viewed by 856
Abstract
Using molecular techniques and bioinformatics tools, we studied the vector-host interactions and the molecular epidemiology of West Nile virus (WNV) in western Iran. Mosquitoes were collected during 2017 and 2018. DNA typing assays were used to study vector-host interactions. Mosquitoes were screened by [...] Read more.
Using molecular techniques and bioinformatics tools, we studied the vector-host interactions and the molecular epidemiology of West Nile virus (WNV) in western Iran. Mosquitoes were collected during 2017 and 2018. DNA typing assays were used to study vector-host interactions. Mosquitoes were screened by RT-PCR for the genomes of five virus families. WNV-positive samples were fully sequenced and evolutionary tree and molecular architecture were constructed by Geneious software and SWISS-MODEL workspace, respectively. A total of 5028 mosquito specimens were collected and identified. The most prevalent species was Culex (Cx.) pipiens complex (57.3%). Analysis of the blood-feeding preferences of blood-fed mosquitoes revealed six mammalian and one bird species as hosts. One mosquito pool containing non-blood-fed Cx. theileri and one blood-fed Culex pipiens pipiens (Cpp.) biotype pipiens were positive for WNV. A phylogram indicated that the obtained WNV sequences belonged to lineage 2, subclade 2 g. Several amino acid substitutions suspected as virulence markers were observed in the Iranian WNV strains. The three-dimensional structural homology model of the E-protein identified hot spot domains known to facilitate virus invasion and neurotropism. The recent detection of WNV lineage 2 in mosquitoes from several regions of Iran in consecutive years suggests that the virus is established in the country. Full article
(This article belongs to the Special Issue West Nile Virus 2019)
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Article
Relocation of the attTn7 Transgene Insertion Site in Bacmid DNA Enhances Baculovirus Genome Stability and Recombinant Protein Expression in Insect Cells
Viruses 2020, 12(12), 1448; https://doi.org/10.3390/v12121448 - 16 Dec 2020
Cited by 3 | Viewed by 1684
Abstract
Baculovirus expression vectors are successfully used for the commercial production of complex (glyco)proteins in eukaryotic cells. The genome engineering of single-copy baculovirus infectious clones (bacmids) in E. coli has been valuable in the study of baculovirus biology, but bacmids are not yet widely [...] Read more.
Baculovirus expression vectors are successfully used for the commercial production of complex (glyco)proteins in eukaryotic cells. The genome engineering of single-copy baculovirus infectious clones (bacmids) in E. coli has been valuable in the study of baculovirus biology, but bacmids are not yet widely applied as expression vectors. An important limitation of first-generation bacmids for large-scale protein production is the rapid loss of gene of interest (GOI) expression. The instability is caused by the mini-F replicon in the bacmid backbone, which is non-essential for baculovirus replication in insect cells, and carries the adjacent GOI in between attTn7 transposition sites. In this paper, we test the hypothesis that relocation of the attTn7 transgene insertion site away from the mini-F replicon prevents deletion of the GOI, thereby resulting in higher and prolonged recombinant protein expression levels. We applied lambda red genome engineering combined with SacB counterselection to generate a series of bacmids with relocated attTn7 sites and tested their performance by comparing the relative expression levels of different GOIs. We conclude that GOI expression from the odv-e56 (pif-5) locus results in higher overall expression levels and is more stable over serial passages compared to the original bacmid. Finally, we evaluated this improved next-generation bacmid during a bioreactor scale-up of Sf9 insect cells in suspension to produce enveloped chikungunya virus-like particles as a model vaccine. Full article
(This article belongs to the Special Issue The Application of Viruses to Biotechnology)
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Article
Spatial Distribution Patterns of Parthenolecanium corni (Hemiptera, Coccidae) and of the Ampelovirus GLRaV-1 and the Vitivirus GVA in a Commercial Vineyard
Viruses 2020, 12(12), 1447; https://doi.org/10.3390/v12121447 - 16 Dec 2020
Cited by 1 | Viewed by 938
Abstract
Distribution patterns of the European fruit lecanium Parthenolecanium corni (Bouché) and of grapevine leafroll-associated virus-1 (GLRaV-1) and grapevine virus A (GVA) were monitored from 2003 to 2015 in a Riesling vine plot in the northeast of France. Virus spread was compared between two [...] Read more.
Distribution patterns of the European fruit lecanium Parthenolecanium corni (Bouché) and of grapevine leafroll-associated virus-1 (GLRaV-1) and grapevine virus A (GVA) were monitored from 2003 to 2015 in a Riesling vine plot in the northeast of France. Virus spread was compared between two periods: 2003–2008 and 2009–2014. The percentage of infected vines increased from 54 to 78% for GLRaV-1 and from 14 to 26% for GVA. The spatial distribution of viruses and of P. corni was analysed using permutation tests and revealed an aggregative pattern. Virus distribution was not associated with the density of P. corni population on grapevines. However, GLRaV-1 and GVA spread mainly from initially infected vines. New GLRaV-1 and GVA infections were more frequent on vines near primarily infected vines, first anisotropically along the row, then between neighbouring rows. Virus spread was similar to those described in literature with grapevine mealybug species. This slow vine-to-vine progression suggests that P. corni was responsible for the virus spread, in accordance with the low mobility and low transmission capacities of its local population. Full article
(This article belongs to the Special Issue Closteroviridae)
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