Viruses

1812 KiB

Open AccessArticle

Emerging Interaction Patterns in the Emiliania huxleyi-EhV System

by Eliana Ruiz, Monique Oosterhof, Ruth-Anne Sandaa, Aud Larsen and António Pagarete

Viruses 2017, 9(3), 61; https://doi.org/10.3390/v9030061 - 22 Mar 2017

Cited by 13 | Viewed by 7739 | Correction

Viruses are thought to be fundamental in driving microbial diversity in the oceanic planktonic realm. That role and associated emerging infection patterns remain particularly elusive for eukaryotic phytoplankton and their viruses. Here we used a vast number of strains from the model system [...] Read more.

Viruses are thought to be fundamental in driving microbial diversity in the oceanic planktonic realm. That role and associated emerging infection patterns remain particularly elusive for eukaryotic phytoplankton and their viruses. Here we used a vast number of strains from the model system Emiliania huxleyi/Emiliania huxleyi Virus to quantify parameters such as growth rate (µ), resistance (R), and viral production (Vp) capacities. Algal and viral abundances were monitored by flow cytometry during 72-h incubation experiments. The results pointed out higher viral production capacity in generalist EhV strains, and the virus-host infection network showed a strong co-evolution pattern between E. huxleyi and EhV populations. The existence of a trade-off between resistance and growth capacities was not confirmed. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Resistance capacity R1 (calculated as the ratio between the number of cells that did not lyse after incubation with viruses and the number of cells in the non-inoculated controls) plotted against growth rate (μ). Error bars show standard deviation (n = 3). Full article ">Figure 2
Resistance capacity R2 (number of viral strains infecting each algal strain) plotted against growth rate (μ). Error bars show standard deviation (n = 3). Full article ">Figure 3
Viral production (Vp) plotted against resistance capacity R1. Error bars show standard deviation (n = 13). Full article ">Figure 4
Number of viral strains infecting each algal strain and maximum viral production correlation. Error bars show standard deviation (n = 3). Full article ">Figure 5
Differences between maximum viral production among EhV strains. Error bars show standard deviation (n = 49). Full article ">Figure 6
Viral-host infectivity network with a clear nested pattern (NODF value of 0.60) where specialist viruses tend to infect the most susceptible hosts, while viruses with broader host-range infect hosts with higher resistance. ■: infection; □: no infection. Sidebars represent μ, R1 and Vp parameters, respectively. Full article ">

3779 KiB

Open AccessArticle

Isolation and Characterization of a Shewanella Phage–Host System from the Gut of the Tunicate, Ciona intestinalis

by Brittany Leigh, Charlotte Karrer, John P. Cannon, Mya Breitbart and Larry J. Dishaw

Viruses 2017, 9(3), 60; https://doi.org/10.3390/v9030060 - 22 Mar 2017

Cited by 14 | Viewed by 8861

Abstract

Outnumbering all other biological entities on earth, bacteriophages (phages) play critical roles in structuring microbial communities through bacterial infection and subsequent lysis, as well as through horizontal gene transfer. While numerous studies have examined the effects of phages on free-living bacterial cells, much [...] Read more.

Outnumbering all other biological entities on earth, bacteriophages (phages) play critical roles in structuring microbial communities through bacterial infection and subsequent lysis, as well as through horizontal gene transfer. While numerous studies have examined the effects of phages on free-living bacterial cells, much less is known regarding the role of phage infection in host-associated biofilms, which help to stabilize adherent microbial communities. Here we report the cultivation and characterization of a novel strain of Shewanella fidelis from the gut of the marine tunicate Ciona intestinalis, inducible prophages from the S. fidelis genome, and a strain-specific lytic phage recovered from surrounding seawater. In vitro biofilm assays demonstrated that lytic phage infection affects biofilm formation in a process likely influenced by the accumulation and integration of the extracellular DNA released during cell lysis, similar to the mechanism that has been previously shown for prophage induction. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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(A) Viral-like particles (VLPs) enumerated via epifluorescence microscopy after mitomycin C induction of Shewanella fidelis 3313. (B) Bacterial turbidity measured at OD600. All measurements were recorded at 24 h after treatment. Full article ">Figure 2
Representative transmission electron microscopy (TEM) images of mitomycin C-induced phages recovered from the supernatant of S. fidelis 3313. Full article ">Figure 3
(A) Circular genome of lytic phage SFCi1 depicts 40 open reading frames (ORFs). (B) Mauve alignment of the SFCi1 genome against two most closely related phage genomes, VP16C and VP16T. Colored boxes indicate sequence blocks with shared sequence identity; regions lacking sequence homology are indicated in white. Full article ">Figure 4
One-step growth curve of lytic phage SFCi1 with its host, S. fidelis 3313, as determined by plaque forming units (PFUs) and bacterial turbidity measurements (OD600). Full article ">Figure 5
(A) TEM image of a pure culture of lytic phage, SFCi1, indicates that it is a myophage. (B) Magnified image of a single viral particle outlining the icosahedral head and details of the contractile tail and baseplate. Full article ">Figure 6
Development of a biofilm by S. fidelis over 48 h in stationary culture, as measured by crystal violet staining of the biofilm, in the presence (Phage) and absence (Control) of lytic phage SFCi1 and/or DNase I treatment. Full article ">Figure 7
TOTO-1 Iodide 514/533 stain and SYTO60 red counterstain reveals extracellular DNA as a major component of the stationary culture biofilm. (A) Untreated S. fidelis 3313 control culture (30.23% ± 5.1% pixel area coverage); (B) S. fidelis 3313 exposed to SFCi1 lytic phage (80.96% ± 7.9% area); (C) control culture treated with DNase I (0.754% ± 0.01% area); and (D) lytic-phage treated culture co-treated with DNase I (0.772% ± 0.017% area). Full article ">

2251 KiB

Open AccessReview

Myeloid C-Type Lectin Receptors in Viral Recognition and Antiviral Immunity

by João T. Monteiro and Bernd Lepenies

Viruses 2017, 9(3), 59; https://doi.org/10.3390/v9030059 - 22 Mar 2017

Cited by 70 | Viewed by 16005

Abstract

Recognition of viral glycans by pattern recognition receptors (PRRs) in innate immunity contributes to antiviral immune responses. C-type lectin receptors (CLRs) are PRRs capable of sensing glycans present in viral pathogens to activate antiviral immune responses such as phagocytosis, antigen processing and presentation, [...] Read more.

Recognition of viral glycans by pattern recognition receptors (PRRs) in innate immunity contributes to antiviral immune responses. C-type lectin receptors (CLRs) are PRRs capable of sensing glycans present in viral pathogens to activate antiviral immune responses such as phagocytosis, antigen processing and presentation, and subsequent T cell activation. The ability of CLRs to elicit and shape adaptive immunity plays a critical role in the inhibition of viral spread within the host. However, certain viruses exploit CLRs for viral entry into host cells to avoid immune recognition. To block CLR interactions with viral glycoproteins, antiviral strategies may involve the use of multivalent glycan carrier systems. In this review, we describe the role of CLRs in antiviral immunity and we highlight their dual function in viral clearance and exploitation by viral pathogens. Full article

(This article belongs to the Special Issue Lectins as Antiviral)

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Recognition of pathogen-associated molecular patterns (PAMPs), damage-associated molecular patterns (DAMPs) and signaling motifs of myeloid C-type lectin receptors (CLRs). CLRs expressed by antigen-presenting cells (APCs) are able to recognize PAMPs present on pathogens, including bacteria, viruses, fungi and parasites; and DAMPs in damaged host cells. Recognized ligands cover a vast type of glycan structures, such as fucose, mannose, β-glucan, galactose, GlcNAc, but also non-glycan ligands such as monosodium urate. Upon CLR engagement, a signaling cascade is initiated through binding of early adaptors and the recruitment of kinases or phosphatases. Myeloid CLRs can be subdivided in four distinct groups according to their cytoplasmic signaling motifs and early adaptors: (A) hemi-immunoreceptor tyrosine-based activating motif (hemITAM)-coupled; (B) ITAM-coupled; (C) immunoreceptor tyrosine-based inhibitory motif (ITIM)-coupled; and (D) ITAM-ITIM independent CLRs. Full article ">Figure 2
CLRs present on the surface of dendritic cells (DCs) recognize viral antigens to trigger APC activation and subsequent T cell stimulation. Upon binding, endocytosis will take place, resulting in the internalization of the antigens. Further processing in endosomes and lysosomes results in fragmented peptides that are loaded on major histocompatibility complex (MHC) class II and MHC class I molecules for efficient priming of CD4+ and CD8+ T cells, respectively. Two signals are required for T cell activation by DCs. First, the T-cell receptor (TCR) recognizes MHC/peptide complexes. Second, the costimulatory molecules CD80/CD86 interact with CD28 expressed by the T cell. In addition, DCs express and secrete cytokines. The combination of these signals determines the fate of the activated T cell. Full article ">Figure 3
Human immunodeficiency virus type 1 (HIV-1) capture and transmission by DCs. Dendritic cell-specific intercellular adhesion molecule-3-grabbing non integrin (DC-SIGN) present at the surface of DCs neutralizes HIV-1 by virion uptake and signalosome-mediated cytokine production and degradation of virions. Fragmented peptides are loaded on MHC class II molecules for antigen presentation to CD4+ T cells in order to prime T cell effector functions and induce an adaptive immune response against the virus. DC-SIGN establishment of a DC-T-cell interaction is accomplished through transient binding to intercellular adhesion molecule (ICAM)-3. However, DC-SIGN is also exploited by HIV-1 for evasion of the immune response by maintenance of intact virions in non-lysosomal endosomes. These virions undergo exocytosis and can infect CD4+ T cells in a process called trans-infection. Trans-infection can also occur without HIV-1 virion internalization. Additional subversion strategies of HIV-1 encompass augmented viral replication via the DC-SIGN signalosome and triggering of DC apoptosis via apoptosis signal-regulating kinase 1 (ASK-1). Full article ">Figure 4
CLRs in antiviral immunity: roles in viral clearance or as targets for viral exploitation. Surface glycoproteins in different enveloped viruses possess glycan moieties (highlighted area), mainly N-linked glycans, that are recognized by CLRs and lead to CLR-mediated endocytosis by APCs. Once inside the host cell, viruses may hijack the host’s cellular machinery to ensure viral spreading (left). HIV-1 interaction with DC-SIGN promotes trans-infection of T cells and tampers with signaling cascades to promote viral replication or to induce apoptosis of professional APCs to enhance viral infectivity. A different type of exploitation is mediated by viruses, such as DV, through interaction with myeloid DAP-12 associating lectin (MDL-1). In this case, viruses elicit an exacerbated pro-inflammatory response, that damages host cells and increases the susceptibility for viral infection. However, CLRs also have important functions in the clearance of viral infections (right). For instance, langerin captures viruses and directs them towards specialized degradation organelles called Birbeck granules. Langerin is also responsible for priming CD4+ and CD8+ T cells and to limit viral spread. Full article ">

223 KiB

Open AccessReview

Virus-Bacteria Interactions: An Emerging Topic in Human Infection

by Erin A. Almand, Matthew D. Moore and Lee-Ann Jaykus

Viruses 2017, 9(3), 58; https://doi.org/10.3390/v9030058 - 21 Mar 2017

Cited by 88 | Viewed by 16315

Abstract

Bacteria and viruses often occupy the same niches, however, interest in their potential collaboration in promoting wellness or disease states has only recently gained traction. While the interaction of some bacteria and viruses is well characterized (e.g., influenza virus), researchers are typically more [...] Read more.

Bacteria and viruses often occupy the same niches, however, interest in their potential collaboration in promoting wellness or disease states has only recently gained traction. While the interaction of some bacteria and viruses is well characterized (e.g., influenza virus), researchers are typically more interested in the location of the infection than the manner of cooperation. There are two overarching types of bacterial-virus disease causing interactions: direct interactions that in some way aid the viruses, and indirect interactions aiding bacteria. The virus-promoting direct interactions occur when the virus exploits a bacterial component to facilitate penetration into the host cell. Conversely, indirect interactions result in increased bacterial pathogenesis as a consequence of viral infection. Enteric viruses mainly utilize the direct pathway, while respiratory viruses largely affect bacteria in an indirect fashion. This review focuses on some key examples of how virus-bacteria interactions impact the infection process across the two organ systems, and provides evidence supporting this as an emerging theme in infectious disease. Full article

(This article belongs to the Section Animal Viruses)

863 KiB

Open AccessReview

Cross-Regulation between Transposable Elements and Host DNA Replication

by Mikel Zaratiegui

Viruses 2017, 9(3), 57; https://doi.org/10.3390/v9030057 - 21 Mar 2017

Cited by 11 | Viewed by 6989

Abstract

Transposable elements subvert host cellular functions to ensure their survival. Their interaction with the host DNA replication machinery indicates that selective pressures lead them to develop ancestral and convergent evolutionary adaptations aimed at conserved features of this fundamental process. These interactions can shape [...] Read more.

Transposable elements subvert host cellular functions to ensure their survival. Their interaction with the host DNA replication machinery indicates that selective pressures lead them to develop ancestral and convergent evolutionary adaptations aimed at conserved features of this fundamental process. These interactions can shape the co-evolution of the transposons and their hosts. Full article

(This article belongs to the Special Issue Recent Progress in Understanding the Mechanism and Consequences of Retrotransposon Movement)

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Replication of type II DNA transposons. (A) Non-replicative transposition of Mu after infection. The Mu phage and flanking DNA are injected into the host. Cleavage and strand transfer join the Mu phage DNA to the target site, leaving single-stranded gaps. Upon arrival of a replication fork the flanking DNA is degraded, and the gaps create a double stranded end create a double stranded end. Both gaps are simultaneously filled by passive DNA replication, yielding a mature prophage; (B) Replicative transposition of Mu in the lytic phase. Strand transfer of the prophage into the target site create a Θ-shaped Shapiro intermediate, with the Mu element flanked by fork-like structures. Primosome-started replication at these structures duplicate the Mu element in a joined cointegrate; (C) Control of activator/dissociator (Ac/Ds) transposition by replication fork passage. Methylation at the inverted terminal repeats (ITRs) is depicted as filled arrows. Hemimethylated ITR depicted as half-filled arrows, with the filled portion indicating the methylated strand. Replication of the methylated 3′ ITR yields two hemimethylated daughter ITR, only one of which binds the transposase (TPase), determining which of the two daughter elements can assemble the transpososome. Full article ">Figure 2
Fork influence on target site selection. (A) Insertion patterns of the Tn7 TnsE-dependent transposition into the host chromosome. Ori = origin of replication. Ter = replication termination region. Insertions are depicted as grey arrows; (B) Insertion patterns of Ty1 and Ty3 in type III genes. Ty1 insertions in black [<a href="#B33-viruses-09-00057" class="html-bibr">33</a>], Ty3 insertions in green [<a href="#B34-viruses-09-00057" class="html-bibr">34</a>] DNA pol ε average occupancy in red [<a href="#B35-viruses-09-00057" class="html-bibr">35</a>]; (C) Insertion patterns of Tf1 in type II genes. Tf1 insertion in black [<a href="#B36-viruses-09-00057" class="html-bibr">36</a>], average DNA pol ε occupancy in red [<a href="#B37-viruses-09-00057" class="html-bibr">37</a>] and average Sap1 occupancy in green [<a href="#B38-viruses-09-00057" class="html-bibr">38</a>]. Full article ">Figure 3
Fork instability at transposable elements (TE). An LTR containing replication fork barriers (RFB) can lead to replication fork stalling and double strand break (DSB) formation (left). (A) Active transcription of the TE can cause replisome-RNA Pol II collisions and unreplicated regions (right); (B) TE with actively transcribing bidirectional promoters can cause replisome-RNA Pol II collisions and unreplicated regions. Full article ">

576 KiB

Open AccessReview

Complete and Incomplete Hepatitis B Virus Particles: Formation, Function, and Application

by Jianming Hu and Kuancheng Liu

Viruses 2017, 9(3), 56; https://doi.org/10.3390/v9030056 - 21 Mar 2017

Cited by 222 | Viewed by 15975

Abstract

Hepatitis B virus (HBV) is a para-retrovirus or retroid virus that contains a double-stranded DNA genome and replicates this DNA via reverse transcription of a RNA pregenome. Viral reverse transcription takes place within a capsid upon packaging of the RNA and the viral [...] Read more.

Hepatitis B virus (HBV) is a para-retrovirus or retroid virus that contains a double-stranded DNA genome and replicates this DNA via reverse transcription of a RNA pregenome. Viral reverse transcription takes place within a capsid upon packaging of the RNA and the viral reverse transcriptase. A major characteristic of HBV replication is the selection of capsids containing the double-stranded DNA, but not those containing the RNA or the single-stranded DNA replication intermediate, for envelopment during virion secretion. The complete HBV virion particles thus contain an outer envelope, studded with viral envelope proteins, that encloses the capsid, which, in turn, encapsidates the double-stranded DNA genome. Furthermore, HBV morphogenesis is characterized by the release of subviral particles that are several orders of magnitude more abundant than the complete virions. One class of subviral particles are the classical surface antigen particles (Australian antigen) that contain only the viral envelope proteins, whereas the more recently discovered genome-free (empty) virions contain both the envelope and capsid but no genome. In addition, recent evidence suggests that low levels of RNA-containing particles may be released, after all. We will summarize what is currently known about how the complete and incomplete HBV particles are assembled. We will discuss briefly the functions of the subviral particles, which remain largely unknown. Finally, we will explore the utility of the subviral particles, particularly, the potential of empty virions and putative RNA virions as diagnostic markers and the potential of empty virons as a vaccine candidate. Full article

(This article belongs to the Special Issue Recent Advances in Hepatitis B Virus Research)

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Schematic of hepatitis B virus (HBV) replication cycle. 1. Virus binding and entry into the host cell (large rectangle). 2. Intracellular trafficking and delivery of relaxed circular (RC) DNA to the nucleus (large circle). 3. Conversion of RC DNA to CCC DNA, or integration of the double-stranded linear (DSL) DNA into host DNA (3a). 4. and 4a. Transcription to synthesize viral RNAs (wavy lines), including the C mRNA for both the core and RT proteins; LS mRNA for the L envelope protein; S mRNA for the M and S envelope proteins; X mRNA for the X protein; and PreC mRNA for the PreCore protein. The C mRNA is also the pgRNA. 5. Translation to synthesize viral proteins. 6. Assembly of the pgRNA- (and RT-) containing NC, or alternatively, empty capsids (6a). 7. Reverse transcription of pgRNA to synthesize the (−) strand SS DNA and then RC DNA. 8. Nuclear recycling of progeny RC DNA to form more CCC DNA (intracellular CCC DNA amplification). 9. Envelopment of the RC DNA-containing NC and secretion of complete virions, or alternatively, secretion of empty virions (9b) or HBsAg spheres and filaments (9a). Processing of the PreCore protein and secretion of HBeAg are depicted in 9c. The secretion of putative RNA virions is not yet resolved (9?). The different viral particles outside the cell are depicted schematically with their approximate concentrations in the blood of infected persons indicated: the complete, empty, or RNA virions as large circles (outer envelope) with an inner diamond shell (capsid), with or without RC DNA (unclosed, double concentric circle) or RNA (wavy line) inside the capsid respectively; HBsAg spheres and filament as small circles and a cylinder. It is important to point out that the concentrations of all these particles can vary widely between different patients and over time in the same patient. Intracellular capsids are depicted as diamonds, with either viral pgRNA, SS [(−) strand] DNA (straight line), RC DNA, or empty. The letters “P” denote phosphorylated residues on the immature NCs (containing SS DNA or pgRNA) or empty capsid. The dashed lines of the diamond in the RC DNA-containing mature NCs signify the destabilization of the mature NC, which is dephosphorylated. The empty capsids, like mature NCs, are also less stable compared to immature NCs but unlike mature NCs, are phosphorylated. The soluble, dimeric HBeAg is depicted as grey double bars. The thin dashed line and arrow denote the fact that HBeAg is frequently decreased or lost late in infection. Boxed letters denote the viral proteins translated from the mRNAs. The filled circle on RC DNA denotes the RT protein attached to the 5’ end of the (−) strand (outer circle) of RC DNA and the arrow denotes the 3’ end of the (+) strand (inner circle) of RC DNA. ccc, CCC DNA; rc, RC DNA. For simplicity, synthesis of the minor DSL form of the genomic DNA in the mature NC, its secretion in virions, and infection of DSL DNA-containing virions are not depicted here, as are the functions of X. See text for details. Modified from [<a href="#B2-viruses-09-00056" class="html-bibr">2</a>]. Full article ">Figure 2
The single strand blocking hypothesis to explain selective HBV virion formation. The new hypothesis is presented in panel (B), in comparison with the classical maturation signal hypothesis depicted in panel (A). The symbol * denotes that the envelope signal for the mature NC vs. the empty capsid may or may not be the same. See text for details. Modified from [<a href="#B8-viruses-09-00056" class="html-bibr">8</a>]. Full article ">

4901 KiB

Open AccessArticle

Virological Surveillance of Influenza A Subtypes Isolated in 2014 from Clinical Outbreaks in Canadian Swine

by Helena Grgić, Jackie Gallant and Zvonimir Poljak

Viruses 2017, 9(3), 55; https://doi.org/10.3390/v9030055 - 21 Mar 2017

Cited by 3 | Viewed by 5690

Abstract

Influenza A viruses (IAVs) are respiratory pathogens associated with an acute respiratory disease that occurs year-round in swine production. It is currently one of the most important pathogens in swine populations, with the potential to infect other host species including humans. Ongoing research [...] Read more.

Influenza A viruses (IAVs) are respiratory pathogens associated with an acute respiratory disease that occurs year-round in swine production. It is currently one of the most important pathogens in swine populations, with the potential to infect other host species including humans. Ongoing research indicates that the three major subtypes of IAV—H1N1, H1N2, and H3N2—continue to expand in their genetic and antigenic diversity. In this study, we conducted a comprehensive genomic analysis of 16 IAVs isolated from different clinical outbreaks in Alberta, Manitoba, Ontario, and Saskatchewan in 2014. We also examined the genetic basis for probable antigenic differences among sequenced viruses. On the basis of phylogenetic analysis, all 13 Canadian H3N2 viruses belonged to cluster IV, eight H3N2 viruses were part of the IV-C cluster, and one virus belonged to the IV-B and one to the IV-D cluster. Based on standards used in this study, three H3N2 viruses could not be clearly classified into any currently established group within cluster IV (A to F). Three H1N2 viruses were part of the H1α cluster. Full article

(This article belongs to the Section Animal Viruses)

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1993 KiB

Open AccessMeeting Report

1st Workshop of the Canadian Society for Virology

by Craig McCormick and Nathalie Grandvaux

Viruses 2017, 9(3), 54; https://doi.org/10.3390/v9030054 - 20 Mar 2017

Cited by 1 | Viewed by 5294

Abstract

The 1st Workshop of the Canadian Society for Virology (CSV2016) was a Special Workshop of the 35th Annual Meeting for the American Society for Virology, held on 18 June 2016 on the beautiful Virginia Tech campus in Blacksburg, Virginia. The workshop provided a [...] Read more.

The 1st Workshop of the Canadian Society for Virology (CSV2016) was a Special Workshop of the 35th Annual Meeting for the American Society for Virology, held on 18 June 2016 on the beautiful Virginia Tech campus in Blacksburg, Virginia. The workshop provided a forum for discussion of recent advances in the field, in an informal setting conducive to interaction with colleagues. CSV2016 featured two internationally-renowned Canadian keynote speakers who discussed translational virology research; American Society for Virology President Grant McFadden (then from University of Florida, now relocated to Arizona State University) who presented his studies of oncolytic poxviruses, while Matthew Miller (McMaster University) reviewed the prospects for a universal influenza vaccine. The workshop also featured a variety of trainee oral and poster presentations, and a panel discussion on the topic of the future of the CSV and virus research in Canada. Full article

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4029 KiB

Open AccessArticle

Porcine Epidemic Diarrhea Virus Induces Autophagy to Benefit Its Replication

by Xiaozhen Guo, Mengjia Zhang, Xiaoqian Zhang, Xin Tan, Hengke Guo, Wei Zeng, Guokai Yan, Atta Muhammad Memon, Zhonghua Li, Yinxing Zhu, Bingzhou Zhang, Xugang Ku, Meizhou Wu, Shengxian Fan and Qigai He

Viruses 2017, 9(3), 53; https://doi.org/10.3390/v9030053 - 19 Mar 2017

Cited by 71 | Viewed by 8923

Abstract

The new porcine epidemic diarrhea (PED) has caused devastating economic losses to the swine industry worldwide. Despite extensive research on the relationship between autophagy and virus infection, the concrete role of autophagy in porcine epidemic diarrhea virus (PEDV) infection has not been reported. [...] Read more.

The new porcine epidemic diarrhea (PED) has caused devastating economic losses to the swine industry worldwide. Despite extensive research on the relationship between autophagy and virus infection, the concrete role of autophagy in porcine epidemic diarrhea virus (PEDV) infection has not been reported. In this study, autophagy was demonstrated to be triggered by the effective replication of PEDV through transmission electron microscopy, confocal microscopy, and Western blot analysis. Moreover, autophagy was confirmed to benefit PEDV replication by using autophagy regulators and RNA interference. Furthermore, autophagy might be associated with the expression of inflammatory cytokines and have a positive feedback loop with the NF-κB signaling pathway during PEDV infection. This work is the first attempt to explore the complex interplay between autophagy and PEDV infection. Our findings might accelerate our understanding of the pathogenesis of PEDV infection and provide new insights into the development of effective therapeutic strategies. Full article

(This article belongs to the Special Issue Porcine Viruses)

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5240 KiB

Open AccessFeature PaperArticle

Coccolithoviruses: A Review of Cross-Kingdom Genomic Thievery and Metabolic Thuggery

by Jozef I. Nissimov, António Pagarete, Fangrui Ma, Sean Cody, David D. Dunigan, Susan A. Kimmance and Michael J. Allen

Viruses 2017, 9(3), 52; https://doi.org/10.3390/v9030052 - 18 Mar 2017

Cited by 23 | Viewed by 8070

Abstract

Coccolithoviruses (Phycodnaviridae) infect and lyse the most ubiquitous and successful coccolithophorid in modern oceans, Emiliania huxleyi. So far, the genomes of 13 of these giant lytic viruses (i.e., Emiliania huxleyi viruses—EhVs) have been sequenced, assembled, and annotated. Here, we performed [...] Read more.

Coccolithoviruses (Phycodnaviridae) infect and lyse the most ubiquitous and successful coccolithophorid in modern oceans, Emiliania huxleyi. So far, the genomes of 13 of these giant lytic viruses (i.e., Emiliania huxleyi viruses—EhVs) have been sequenced, assembled, and annotated. Here, we performed an in-depth comparison of their genomes to try and contextualize the ecological and evolutionary traits of these viruses. The genomes of these EhVs have from 444 to 548 coding sequences (CDSs). Presence/absence analysis of CDSs identified putative genes with particular ecological significance, namely sialidase, phosphate permease, and sphingolipid biosynthesis. The viruses clustered into distinct clades, based on their DNA polymerase gene as well as full genome comparisons. We discuss the use of such clustering and suggest that a gene-by-gene investigation approach may be more useful when the goal is to reveal differences related to functionally important genes. A multi domain “Best BLAST hit” analysis revealed that 84% of the EhV genes have closer similarities to the domain Eukarya. However, 16% of the EhV CDSs were very similar to bacterial genes, contributing to the idea that a significant portion of the gene flow in the planktonic world inter-crosses the domains of life. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Phylogenetic analysis of coccolithoviruses based on their DNA polymerase and serine palmitoyltransferase (SPT) genes. The evolutionary history of 13 EhV strains was inferred based on the 2604 bp long SPT (I and II) and 2921 bp long DNA polymerase (III and IV) genes, using the Neighbor-Joining (I and III) and Maximum Likelihood (II and IV) methods. Note that EhV-18 and EhV-145 are absent from the serine palmitoyltransferase tree due to the full length SPT protein being split over two separate genes in their respective genomes. Based on the DNA polymerase phylogeny, the EhVs cluster into two main clades: A and B (green). Clade A is further divided into sub-clusters A1 (red), A2 (yellow), and A3 (purple). The percentage of replicate trees in which the associated taxa clustered together in the bootstrap test (1000 replicates) are shown next to the branches. The evolutionary distances were computed using the Tamura-Nei method and are in the units of the number of base substitutions per site. Full article ">Figure 2
Whole genome alignment of sequenced coccolithovirus genomes. The genomes were aligned using MAUVE, in relation to the non-gapped backbone genome of EhV-86. Syntenous blocks are indicated in the same colours and the lines that connect them indicate the position of each block in relation to the same block of genes in the genome of EhV-86. The small red lines on each genome represent the exact positions of the gaps that separate the different contigs within each draft genome. The genomes are ordered based on their DNA polymerase phylogeny (<a href="#viruses-09-00052-f001" class="html-fig">Figure 1</a>), based on the ANI analysis of this study (<a href="#viruses-09-00052-t002" class="html-table">Table 2</a>), and based on previously published microarray data that puts them into the aforementioned groups and sub-clades [<a href="#B20-viruses-09-00052" class="html-bibr">20</a>]. Full article ">Figure 3
“Best BLAST hit” analysis of coccolithovirus CDSs in relation to the three domains of life: Eukarya, Bacteria and Archaea. Predicted genes within EhV genomes were BLASTp analyzed against possible hits in the three domains of life using a gene BitScore of >50 (A); and >100 (B). Further EhV gene hits analysis to the taxonomic level of “order” in Eukarya (C); and Bacteria (D) was performed using a BitScore of >100. Full article ">

4513 KiB

Open AccessArticle

Schrödinger’s Cheshire Cat: Are Haploid Emiliania huxleyi Cells Resistant to Viral Infection or Not?

by Gideon J. Mordecai, Frederic Verret, Andrea Highfield and Declan C. Schroeder

Viruses 2017, 9(3), 51; https://doi.org/10.3390/v9030051 - 18 Mar 2017

Cited by 7 | Viewed by 11033

Abstract

Emiliania huxleyi is the main calcite producer on Earth and is routinely infected by a virus (EhV); a double stranded DNA (dsDNA) virus belonging to the family Phycodnaviridae. E. huxleyi exhibits a haplodiploid life cycle; the calcified diploid stage is non-motile and [...] Read more.

Emiliania huxleyi is the main calcite producer on Earth and is routinely infected by a virus (EhV); a double stranded DNA (dsDNA) virus belonging to the family Phycodnaviridae. E. huxleyi exhibits a haplodiploid life cycle; the calcified diploid stage is non-motile and forms extensive blooms. The haploid phase is a non-calcified biflagellated cell bearing organic scales. Haploid cells are thought to resist infection, through a process deemed the “Cheshire Cat” escape strategy; however, a recent study detected the presence of viral lipids in the same haploid strain. Here we report on the application of an E. huxleyi CCMP1516 EhV-86 combined tiling array (TA) that further confirms an EhV infection in the RCC1217 haploid strain, which grew without any signs of cell lysis. Reverse transcription polymerase chain reaction (RT-PCR) and PCR verified the presence of viral RNA in the haploid cells, yet indicated an absence of viral DNA, respectively. These infected cells are an alternative stage of the virus life cycle deemed the haplococcolithovirocell. In this instance, the host is both resistant to and infected by EhV, i.e., the viral transcriptome is present in haploid cells whilst there is no evidence of viral lysis. This superimposed state is reminiscent of Schrödinger’s cat; of being simultaneously both dead and alive. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Open AccessReview

A Review of Phage Therapy against Bacterial Pathogens of Aquatic and Terrestrial Organisms

by Janis Doss, Kayla Culbertson, Delilah Hahn, Joanna Camacho and Nazir Barekzi

Viruses 2017, 9(3), 50; https://doi.org/10.3390/v9030050 - 18 Mar 2017

Cited by 207 | Viewed by 38683

Abstract

Since the discovery of bacteriophage in the early 1900s, there have been numerous attempts to exploit their innate ability to kill bacteria. The purpose of this report is to review current findings and new developments in phage therapy with an emphasis on bacterial [...] Read more.

Since the discovery of bacteriophage in the early 1900s, there have been numerous attempts to exploit their innate ability to kill bacteria. The purpose of this report is to review current findings and new developments in phage therapy with an emphasis on bacterial diseases of marine organisms, humans, and plants. The body of evidence includes data from studies investigating bacteriophage in marine and land environments as modern antimicrobial agents against harmful bacteria. The goal of this paper is to present an overview of the topic of phage therapy, the use of phage-derived protein therapy, and the hosts that bacteriophage are currently being used against, with an emphasis on the uses of bacteriophage against marine, human, animal and plant pathogens. Full article

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Open AccessArticle

Viral Protein Kinetics of Piscine Orthoreovirus Infection in Atlantic Salmon Blood Cells

by Hanne Merethe Haatveit, Øystein Wessel, Turhan Markussen, Morten Lund, Bernd Thiede, Ingvild Berg Nyman, Stine Braaen, Maria Krudtaa Dahle and Espen Rimstad

Viruses 2017, 9(3), 49; https://doi.org/10.3390/v9030049 - 18 Mar 2017

Cited by 32 | Viewed by 9145

Abstract

Piscine orthoreovirus (PRV) is ubiquitous in farmed Atlantic salmon (Salmo salar) and the cause of heart and skeletal muscle inflammation. Erythrocytes are important target cells for PRV. We have investigated the kinetics of PRV infection in salmon blood cells. The findings [...] Read more.

Piscine orthoreovirus (PRV) is ubiquitous in farmed Atlantic salmon (Salmo salar) and the cause of heart and skeletal muscle inflammation. Erythrocytes are important target cells for PRV. We have investigated the kinetics of PRV infection in salmon blood cells. The findings indicate that PRV causes an acute infection of blood cells lasting 1–2 weeks, before it subsides into persistence. A high production of viral proteins occurred initially in the acute phase which significantly correlated with antiviral gene transcription. Globular viral factories organized by the non-structural protein µNS were also observed initially, but were not evident at later stages. Interactions between µNS and the PRV structural proteins λ1, µ1, σ1 and σ3 were demonstrated. Different size variants of µNS and the outer capsid protein µ1 appeared at specific time points during infection. Maximal viral protein load was observed five weeks post cohabitant challenge and was undetectable from seven weeks post challenge. In contrast, viral RNA at a high level could be detected throughout the eight-week trial. A proteolytic cleavage fragment of the µ1 protein was the only viral protein detectable after seven weeks post challenge, indicating that this µ1 fragment may be involved in the mechanisms of persistent infection. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Figure 1
Piscine orthoreovirus (PRV) RNA load in blood cells. Reverse transcription quantitative polymerase chain reaction (RT-qPCR) of PRV gene segments S1, M2 and M3 in blood cells from cohabitant fish. Individual (dots) and mean (line) Ct-values, n = 6 per time-point. wpc = weeks post challenge. Full article ">Figure 2
Expression of immune genes in blood cells. (a) Immune genes were assayed at 3–8 wpc by RT-qPCR in blood cells from cohabitant fish (n = 6 per time point). Data are normalized against EF1α and the lowest ΔCt level at 3 wpc (n = 6), and 2-ΔΔCt values are calculated. Mean relative expression is indicated. ISG = interferon-stimulated gene, PKR = double-stranded RNA (dsRNA)-activated protein kinase; (b) Correlation between Ct values for S1/M3 RNA and relative levels of antiviral gene expression for a set of immune genes. Full article ">Figure 3
Presence of PRV µNS and σ1 in blood cells. (a) Intracellular staining of µNS in blood cells analyzed by flow cytometry from three cohabitant fish sampled at 4, 5 and 6 wpc. The negative control staining is one fish sampled at 0 wpc. A total of 50,000 cells were counted per sample and 30,000 were gated for analysis; (b) Fluorescent labeling of µNS (left) and σ1 (right) displaying viral factory-like inclusions (green) in infected red blood cells sampled 0 (negative control), 4, 5 and 6 wpc. The nuclei were stained with Hoechst (blue). Full article ">Figure 4
Transmission electron microscopy (TEM) of blood cells. PRV-infected red blood cells sampled at 0 (negative control), 4, 5 and 6 wpc show small empty vesicles (cross), lamellar structures (arrowhead), reovirus-like particles (arrow) and large empty inclusions (star). Full article ">Figure 5
Detection of PRV uNS protein in blood cells compared to viral RNA load. Blood cells from 3, 4, 5 and 6 wpc (n = 6) analyzed for µNS by Western blotting. Ct-values for gene segment M3 (µNS) from the same samples are shown below each lane. M = molecular weight standard; Lane 1–6 refers to individual fish (1–6) per time point. Full article ">Figure 6
Presence of PRV proteins in blood cells. Pooled blood cell samples (n = 6) from each week were analyzed by Western blotting, targeting µNS, σ1, σ3, µ1 and λ1. M = molecular weight standard. Actin was used as control for protein load. Full article ">Figure 7
µNS interacts with multiple PRV proteins. Pooled blood cell lysate (n = 6) immunoprecipitated with µNS-antiserum, followed by Western blotting with primary antibodies detecting µNS, µ1C, σ1, σ3 and λ1 (arrows). M = molecular weight standard. Full article ">

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Open AccessReview

Porcine Rotaviruses: Epidemiology, Immune Responses and Control Strategies

by Anastasia N. Vlasova, Joshua O. Amimo and Linda J. Saif

Viruses 2017, 9(3), 48; https://doi.org/10.3390/v9030048 - 18 Mar 2017

Cited by 173 | Viewed by 18277

Abstract

Rotaviruses (RVs) are a major cause of acute viral gastroenteritis in young animals and children worldwide. Immunocompetent adults of different species become resistant to clinical disease due to post-infection immunity, immune system maturation and gut physiological changes. Of the 9 RV genogroups (A–I), [...] Read more.

Rotaviruses (RVs) are a major cause of acute viral gastroenteritis in young animals and children worldwide. Immunocompetent adults of different species become resistant to clinical disease due to post-infection immunity, immune system maturation and gut physiological changes. Of the 9 RV genogroups (A–I), RV A, B, and C (RVA, RVB, and RVC, respectively) are associated with diarrhea in piglets. Although discovered decades ago, porcine genogroup E RVs (RVE) are uncommon and their pathogenesis is not studied well. The presence of porcine RV H (RVH), a newly defined distinct genogroup, was recently confirmed in diarrheic pigs in Japan, Brazil, and the US. The complex epidemiology, pathogenicity and high genetic diversity of porcine RVAs are widely recognized and well-studied. More recent data show a significant genetic diversity based on the VP7 gene analysis of RVB and C strains in pigs. In this review, we will summarize previous and recent research to provide insights on historic and current prevalence and genetic diversity of porcine RVs in different geographic regions and production systems. We will also provide a brief overview of immune responses to porcine RVs, available control strategies and zoonotic potential of different RV genotypes. An improved understanding of the above parameters may lead to the development of more optimal strategies to manage RV diarrheal disease in swine and humans. Full article

(This article belongs to the Special Issue Porcine Viruses)

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Potential mechanisms of rotavirus (RV) pathogenesis. RV replication inside enterocytes induces osmotic diarrhea. RV also increases the concentration of intracellular calcium (Ca2+), disrupting the cytoskeleton and the tight junctions, increasing paracellular permeability. In addition, RV produces non-structural protein 4 (NSP4), an enterotoxin that induces Ca2+ efflux from endoplasmatic reticulum via the phospholipase C dependent (PLC) mechanism further contributing to electrolyte imbalance and secretory diarrhea. RV can also stimulate the enteric nervous system (ENS, via NSP4 dependent mechanism), further contributing to secretory diarrhea and increasing intestinal motility. Agents that can inhibit the ENS could be useful in alleviating RV diarrhea in children. Following, tryptic cleavage of viral protein 8 (VP8) from VP5, the VP8 fragment alters the localization of claudin-3, ZO-1 and occludin leading to the disruption of the barrier integrity of tight junctions (TJ) [<a href="#B3-viruses-09-00048" class="html-bibr">3</a>,<a href="#B4-viruses-09-00048" class="html-bibr">4</a>,<a href="#B5-viruses-09-00048" class="html-bibr">5</a>,<a href="#B6-viruses-09-00048" class="html-bibr">6</a>]. Late in the infectious process, RV destroys mature enterocytes, further contributing to malabsorptive or osmotic diarrhoea. RV antigens, genomic RNA and infectious particles have been found in the blood of children and blood and systemic organs in animals [<a href="#B7-viruses-09-00048" class="html-bibr">7</a>,<a href="#B8-viruses-09-00048" class="html-bibr">8</a>]. The role of systemic RV translocation in disease pathogenesis is currently unknown. DLP: double-layered particles. Full article ">Figure 2
Global genotype distribution of porcine RVA strains reported in historic (1976–2011, blue figure arrows) and current (after 2000, pink figure arrows) studies. Porcine RVAs are also detected in Germany and Russia, but no genotyping data is available. Full article ">Figure 3
Global genotype distribution of porcine RVB (pink figure arrows) and RVC (blue figure arrows) strains and porcine RVE (bolded, orange circle)/RVH (bolded, purple circles) occurrence in different countries reported in historic (1976–2011) and current (after 2000) studies. Porcine RVCs are also detected in Germany and China, and porcine RVB is confirmed in Germany and Czech Republic, but no porcine RVC/RVB genotyping data is available for these countries. Full article ">Figure 4
Immune responses to RV infection in pigs. Intestinal RV VP4/VP7 secretory immunoglobulin A (sIgA) neutralizing antibodies can prevent viral binding to enterocytes and penetration (early post-infection), while viral replication can be partially inhibited by anti-VP6 sIgA during transcytosis across enterocytes. In addition, a number of immune cells contribute to RV innate and adaptive immune responses: plasmacytoid dendritic cells (pDCs) produce antiviral (IFN-α) and pro-inflammatory (IL-12) cytokines which can inhibit RV replication or induce other immune cell subsets, including natural killer (NK) cells that produce granzymes, perforins and TNF-α and can lyse RV-infected cells. After antigen presentation by conventional dendritic cells (cDCs) to T cells, cytokine-secreting (IFN-γ in particular) RV-specific Th cells can also inhibit viral replication and activate IgA production by B cells. Additionally, RV-specific CD8 cytotoxic IFN-γ producing T cells contribute to the lysis of RV infected cells. RV induces apoptosis of intestinal epithelial (enterocytes) and immune cells; however, it is unclear whether this decreases (by eliminating infected cells) or promotes (via dissemination of the infectious particles) RV replication. Although high levels of systemic RV-neutralizing antibodies may coincide with improved protection against RV challenge, they are not correlated with protection in most studies. TJ: tight junctions. DLP: double-layered particles. Full article ">

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Open AccessArticle

A Pelagic Microbiome (Viruses to Protists) from a Small Cup of Seawater

by Flavia Flaviani, Declan C. Schroeder, Cecilia Balestreri, Joanna L. Schroeder, Karen Moore, Konrad Paszkiewicz, Maya C. Pfaff and Edward P. Rybicki

Viruses 2017, 9(3), 47; https://doi.org/10.3390/v9030047 - 17 Mar 2017

Cited by 16 | Viewed by 10097

Abstract

The aquatic microbiome is composed of a multi-phylotype community of microbes, ranging from the numerically dominant viruses to the phylogenetically diverse unicellular phytoplankton. They influence key biogeochemical processes and form the base of marine food webs, becoming food for secondary consumers. Due to [...] Read more.

The aquatic microbiome is composed of a multi-phylotype community of microbes, ranging from the numerically dominant viruses to the phylogenetically diverse unicellular phytoplankton. They influence key biogeochemical processes and form the base of marine food webs, becoming food for secondary consumers. Due to recent advances in next-generation sequencing, this previously overlooked component of our hydrosphere is starting to reveal its true diversity and biological complexity. We report here that 250 mL of seawater is sufficient to provide a comprehensive description of the microbial diversity in an oceanic environment. We found that there was a dominance of the order Caudovirales (59%), with the family Myoviridae being the most prevalent. The families Phycodnaviridae and Mimiviridae made up the remainder of pelagic double-stranded DNA (dsDNA) virome. Consistent with this analysis, the Cyanobacteria dominate (52%) the prokaryotic diversity. While the dinoflagellates and their endosymbionts, the superphylum Alveolata dominates (92%) the microbial eukaryotic diversity. A total of 834 prokaryotic, 346 eukaryotic and 254 unique virus phylotypes were recorded in this relatively small sample of water. We also provide evidence, through a metagenomic-barcoding comparative analysis, that viruses are the likely source of microbial environmental DNA (meDNA). This study opens the door to a more integrated approach to oceanographic sampling and data analysis. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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(a) Map showing the location of sample collection; (b) schematics of the bioinformatics pipeline. Full article ">Figure 1 Cont.
(a) Map showing the location of sample collection; (b) schematics of the bioinformatics pipeline. Full article ">Figure 2
Analyses of the prokaryotic fraction. (a) Reduction in number of reads when filters are applied; (b) percentage and phylotype count when filter are applied; (c) presence–absence analyses at phylotype level before and after application of the filters; (d) number of phylotype analyses by replicate; (e) presence–absence analyses at phylotype level when filters are applied to each replicate. Full article ">Figure 3
Analyses of the eukaryotic fraction. (a) Reduction in number of reads when filters are applied; (b) percentage and phylotype count when filters are applied; (c) presence–absence analyses at phylotype level before and after application of the filter; (d) number of phylotypes analyses by replicate; (e) presence–absence analyses at phylotype level when filters are applied to each replicate. Full article ">Figure 4
Taxonomic assignment based on reads (a,c) and contigs (b,d) analyses. Reads (R1) were annotated using (a) the Virus database and (c) the Refseq database; contigs were annotated using (b) the virus database and (c) the Refseq nr-protein database. Full article ">Figure 5
Presence–absence analyses of the <0.45 µm fraction. Comparison of phylotypes at the level of species (a,c) and genus (b,d) using a subsample of reads (R1) versus contigs at T0 (a,b) and T10 (c,d). Full article ">Figure 6
Krona chart of contigs annotation using the Virus db. Full article ">Figure 7
Presence–absence analyses between the >0.45 µm fraction (prokaryotes and eukaryotes) and the permeate (<0.45 µm). (a) T0: Metagenomic contigs, prokaryotes, eukaryotes; (b) T0: Metagenomic contigs, T1: prokaryotes, eukaryotes; (c) T0: Metagenomic contigs, T10: prokaryotes, eukaryotes; (d) T0: Metagenomic contigs, T10-R1: prokaryotes, eukaryotes; (e) T0: Metagenomic contigs, T10-R2: prokaryotes, eukaryotes. Full article ">

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Open AccessReview

A Student’s Guide to Giant Viruses Infecting Small Eukaryotes: From Acanthamoeba to Zooxanthellae

by Steven W. Wilhelm, Jordan T. Bird, Kyle S. Bonifer, Benjamin C. Calfee, Tian Chen, Samantha R. Coy, P. Jackson Gainer, Eric R. Gann, Huston T. Heatherly, Jasper Lee, Xiaolong Liang, Jiang Liu, April C. Armes, Mohammad Moniruzzaman, J. Hunter Rice, Joshua M. A. Stough, Robert N. Tams, Evan P. Williams and Gary R. LeCleir

Viruses 2017, 9(3), 46; https://doi.org/10.3390/v9030046 - 17 Mar 2017

Cited by 34 | Viewed by 12547

Abstract

The discovery of infectious particles that challenge conventional thoughts concerning “what is a virus” has led to the evolution a new field of study in the past decade. Here, we review knowledge and information concerning “giant viruses”, with a focus not only on [...] Read more.

The discovery of infectious particles that challenge conventional thoughts concerning “what is a virus” has led to the evolution a new field of study in the past decade. Here, we review knowledge and information concerning “giant viruses”, with a focus not only on some of the best studied systems, but also provide an effort to illuminate systems yet to be better resolved. We conclude by demonstrating that there is an abundance of new host–virus systems that fall into this “giant” category, demonstrating that this field of inquiry presents great opportunities for future research. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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The scale of giant virus genomes. (A). Genome size vs. rank plot for the largest 100 complete viral genomes as of January 2016 from National Center for Biotechnology Information (NCBI). Data points noted (●) were previously used in discussion by Claverie et al. [<a href="#B3-viruses-09-00046" class="html-bibr">3</a>] to define giants viruses as having genomes > 280 kb, open circles (○) represent additional data; (B). Genome size vs. rank order of completed bacterial genomes in NCBI as of January 2016. Sizes are color-coded to match the ranges of giant virus genomes. Full article ">Figure 2
Transmission electron micrographs of giant virus particles. (A) Pithovirus, as seen in Michel et al. [<a href="#B22-viruses-09-00046" class="html-bibr">22</a>]. Originally identified as a KC5/2 parasite, the image shows the electron dense viral wall consisting of perpendicularly oriented fibers or microtubules (arrows), and a marked ostiole (os) located at the apical end of the cell. Reprinted with permission—original magnification at 85,000×; (B) Megavirus chilensis. Image courtesy of Professors Chantal Abergel and Jean-Michel Claverie. Full article ">Figure 2 Cont.
Transmission electron micrographs of giant virus particles. (A) Pithovirus, as seen in Michel et al. [<a href="#B22-viruses-09-00046" class="html-bibr">22</a>]. Originally identified as a KC5/2 parasite, the image shows the electron dense viral wall consisting of perpendicularly oriented fibers or microtubules (arrows), and a marked ostiole (os) located at the apical end of the cell. Reprinted with permission—original magnification at 85,000×; (B) Megavirus chilensis. Image courtesy of Professors Chantal Abergel and Jean-Michel Claverie. Full article ">

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Open AccessArticle

Phage Biodiversity in Artisanal Cheese Wheys Reflects the Complexity of the Fermentation Process

by Jennifer Mahony, Angelo Moscarelli, Philip Kelleher, Gabriele A. Lugli, Marco Ventura, Luca Settanni and Douwe Van Sinderen

Viruses 2017, 9(3), 45; https://doi.org/10.3390/v9030045 - 16 Mar 2017

Cited by 21 | Viewed by 7370

Abstract

Dairy fermentations constitute a perfect “breeding ground” for bacteriophages infecting starter cultures, particularly strains of Lactococcus lactis. In modern fermentations, these phages typically belong to one of three groups, i.e., the 936, P335, and c2 phage groups. Traditional production methods present fewer chemical [...] Read more.

Dairy fermentations constitute a perfect “breeding ground” for bacteriophages infecting starter cultures, particularly strains of Lactococcus lactis. In modern fermentations, these phages typically belong to one of three groups, i.e., the 936, P335, and c2 phage groups. Traditional production methods present fewer chemical and physical barriers to phage proliferation compared to modern production systems, while the starter cultures used are typically complex, variable, and undefined. In the current study, a variety of cheese whey, animal-derived rennet, and vat swab samples from artisanal cheeses produced in Sicily were analysed for the presence of lactococcal phages to assess phage diversity in such environments. The complete genomes of 18 representative phage isolates were sequenced, allowing the identification of 10 lactococcal 949 group phages, six P087 group phages, and two members of the 936 group phages. The genetic diversity of these isolates was examined using phylogenetic analysis as well as a focused analysis of the receptor binding proteins, which dictate specific interactions with the host-encoded receptor. Thermal treatments at 63 °C and 83 °C indicate that the 949 phages are particularly sensitive to thermal treatments, followed by the P087 and 936 isolates, which were shown to be much less sensitive to such treatments. This difference may explain the relatively low frequency of isolation of the so-called “rare” 949 and P087 group phages in modern fermentations. Full article

(This article belongs to the Special Issue Viruses of Microbes)

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Unrooted phylogenetic tree of the complete nucleotide sequences of the 949 (green branches) and P087 (blue branches) isolates from this study and previously sequenced members 949, L47, WRP3, and P087. Whole phage nucleotide alignment was performed via ClustalW V2.1. Subsequently, the phylogenetic tree was generated using the neighbor-joining method and bootstrapped (×1000) replicates. Visualisation of the phylogenetic tree was performed using the Itol software (<a href="http://itol.embl.de/" target="_blank">http://itol.embl.de/</a>). Clusters of phage genomes by source sample are indicated in coloured text and corresponding coloured text boxes with the sample from which the phages were derived. The 949 phages isolated from W1.16 (green) are genetically distinct from those isolated from samples W2.16 (hot pink), W4.16 (purple) and W8.14 (pale pink). Similarly, the P087 phage isolates are genetically distinct based on the sample source with the exception of LW33 (isolated from W3.14, orange), which groups more closely to the 2016 isolates from W10.16 (blue) and P087, while phage LW4 occupies a distinct position (grey) highlighting its more distant relationship to the other P087 isolates. Full article ">Figure 2
Heatmap indicating the presence (red) or absence (black) of individual protein families encoded by members of the 949 (A) and P087 (B) groups. Distinct phylogenetic clusters of the 949 and P087 phages can be identified using this approach that are consistent with the phylogeny based on overall nucleotide content presented in <a href="#viruses-09-00045-f001" class="html-fig">Figure 1</a>. Full article ">Figure 3
Rooted phylogenetic tree (rooted by the first sequenced member, 949) of the receptor binding proteins (RBPs) encoded by the 949 phage isolates from this and previous studies. The tree was constructed based on a multiple alignment using ClustalW software with the neighbour-joining method with a bootstrap value of 1000. Visualisation of the phylogenetic tree was performed using the ITOL software (<a href="http://itol.embl.de/" target="_blank">http://itol.embl.de/</a>). The names of the phage isolates derived from this study are colour-coded to indicate the cell wall polysaccharide (CWPS)-type presented by host(s) of these phages as indicated in the text, while in purple text are previously described isolates whose host CWPS types were not assessed in this study. Full article ">Figure 4
Unrooted phylogenetic tree of the RBP sequences of R31 and R3.4 and previously sequenced representative members of the five 936 RBP groups. R31 and R3.4 RBPs are identical to each other and occupy a distinct node within the RBP group I clade. The CWPS types of the host strains infected by these phages are presented along with the RBP grouping information. Full article ">Figure 5
Thermal inactivation assays of selected 949, P087, and 936 isolates at room temperature (control, black bars), 63 °C (grey) and 83 °C (patterned). The 949 isolates display almost complete inactivation at 83 °C and reduced titres at 63 °C. The P087 isolates exhibit approximately 5 log titre at 83 °C and only minor titre reductions (one to two logs) at 63 °C. Full article ">

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Open AccessReview

Reverse Transcription in the Saccharomyces cerevisiae Long-Terminal Repeat Retrotransposon Ty3

by Jason W. Rausch, Jennifer T. Miller and Stuart F. J. Le Grice

Viruses 2017, 9(3), 44; https://doi.org/10.3390/v9030044 - 15 Mar 2017

Cited by 4 | Viewed by 6187

Abstract

Converting the single-stranded retroviral RNA into integration-competent double-stranded DNA is achieved through a multi-step process mediated by the virus-coded reverse transcriptase (RT). With the exception that it is restricted to an intracellular life cycle, replication of the Saccharomyces cerevisiae long terminal repeat (LTR)-retrotransposon [...] Read more.

Converting the single-stranded retroviral RNA into integration-competent double-stranded DNA is achieved through a multi-step process mediated by the virus-coded reverse transcriptase (RT). With the exception that it is restricted to an intracellular life cycle, replication of the Saccharomyces cerevisiae long terminal repeat (LTR)-retrotransposon Ty3 genome is guided by equivalent events that, while generally similar, show many unique and subtle differences relative to the retroviral counterparts. Until only recently, our knowledge of RT structure and function was guided by a vast body of literature on the human immunodeficiency virus (HIV) enzyme. Although the recently-solved structure of Ty3 RT in the presence of an RNA/DNA hybrid adds little in terms of novelty to the mechanistic basis underlying DNA polymerase and ribonuclease H activity, it highlights quite remarkable topological differences between retroviral and LTR-retrotransposon RTs. The theme of overall similarity but distinct differences extends to the priming mechanisms used by Ty3 RT to initiate (−) and (+) strand DNA synthesis. The unique structural organization of the retrotransposon enzyme and interaction with its nucleic acid substrates, with emphasis on polypurine tract (PPT)-primed initiation of (+) strand synthesis, is the subject of this review. Full article

(This article belongs to the Special Issue Recent Progress in Understanding the Mechanism and Consequences of Retrotransposon Movement)

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4697 KiB

Open AccessReview

Marine Prasinoviruses and Their Tiny Plankton Hosts: A Review

by Karen D. Weynberg, Michael J. Allen and William H. Wilson

Viruses 2017, 9(3), 43; https://doi.org/10.3390/v9030043 - 15 Mar 2017

Cited by 34 | Viewed by 10138

Abstract

Viruses play a crucial role in the marine environment, promoting nutrient recycling and biogeochemical cycling and driving evolutionary processes. Tiny marine phytoplankton called prasinophytes are ubiquitous and significant contributors to global primary production and biomass. A number of viruses (known as prasinoviruses) that [...] Read more.

Viruses play a crucial role in the marine environment, promoting nutrient recycling and biogeochemical cycling and driving evolutionary processes. Tiny marine phytoplankton called prasinophytes are ubiquitous and significant contributors to global primary production and biomass. A number of viruses (known as prasinoviruses) that infect these important primary producers have been isolated and characterised over the past decade. Here we review the current body of knowledge about prasinoviruses and their interactions with their algal hosts. Several genes, including those encoding for glycosyltransferases, methyltransferases and amino acid synthesis enzymes, which have never been identified in viruses of eukaryotes previously, have been detected in prasinovirus genomes. The host organisms are also intriguing; most recently, an immunity chromosome used by a prasinophyte in response to viral infection was discovered. In light of such recent, novel discoveries, we discuss why the cellular simplicity of prasinophytes makes for appealing model host organism–virus systems to facilitate focused and detailed investigations into the dynamics of marine viruses and their intimate associations with host species. We encourage the adoption of the prasinophyte Ostreococcus and its associated viruses as a model host–virus system for examination of cellular and molecular processes in the marine environment. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Open AccessReview

Non-Canonical Roles of Dengue Virus Non-Structural Proteins

by Julianna D. Zeidler, Lorena O. Fernandes-Siqueira, Glauce M. Barbosa and Andrea T. Da Poian

Viruses 2017, 9(3), 42; https://doi.org/10.3390/v9030042 - 13 Mar 2017

Cited by 29 | Viewed by 10160

Abstract

The Flaviviridae family comprises a number of human pathogens, which, although sharing structural and functional features, cause diseases with very different outcomes. This can be explained by the plurality of functions exerted by the few proteins coded by viral genomes, with some of [...] Read more.

The Flaviviridae family comprises a number of human pathogens, which, although sharing structural and functional features, cause diseases with very different outcomes. This can be explained by the plurality of functions exerted by the few proteins coded by viral genomes, with some of these functions shared among members of a same family, but others being unique for each virus species. These non-canonical functions probably have evolved independently and may serve as the base to the development of specific therapies for each of those diseases. Here it is discussed what is currently known about the non-canonical roles of dengue virus (DENV) non-structural proteins (NSPs), which may account for some of the effects specifically observed in DENV infection, but not in other members of the Flaviviridae family. This review explores how DENV NSPs contributes to the physiopathology of dengue, evasion from host immunity, metabolic changes, and redistribution of cellular components during infection. Full article

(This article belongs to the Special Issue Advances in Flavivirus Research)

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Figure 1
Schematic representation of the flaviviruses’ polyprotein. Viral RNA encodes a polyprotein that is co- and post-translationally processed by host proteases (black scissors) or by the viral protease NS2B/NS3 (red scissors) to generate the structural (C, PrM, and E) and non-structural proteins (NS1, NS2A, NS2B, NS3, NS4A, NS4B, and NS5), represented in different colors. Full article ">Figure 2
Involvement of DENV NSPs in the evasion of host innate immune response. (A) Pathways of immune response affected by DENV NSPs. (1) Activation of the complement system is triggered by different pathways that converge to the cleavage of factor C3 by the protease C3 convertase. This enzyme formed by the association of two other cleavage products: C4b, a fragment of C4, and C2a, a fragment of C2. The cleavage of C4 and C2 may be catalyzed by two different pathways: the classical pathway, triggered by C1 binding to antigen-antibody complexes, or by the lectin pathway, in which a carbohydrate recognition receptor, such as mannose binding lectin (MBL), associates to a serine protease after binding to carbohydrates. One of the products of C3 cleavage, C3b, binds to C3 convertase changing its substrate specificity, so that the enzyme becomes a C5 convertase. The fragment C5b, generated from the cleavage of C5, binds to the infected cell membrane, initiating the assembly of a complex formed by C6, C7, C8, and C9, which promotes cell lysis. (2) Viral dsRNAs produced during the replication of RNA viruses are recognized by PRRs. Binding of dsRNA leads the cytosolic PPRs to associate with the mitochondrial antiviral signaling protein (MAVS) through its caspase-recruitment domain (CARD), recruiting the TANK-protein kinase 1 (TBK1) and IκB kinase-ε (IKKε). These kinases phosphorylate IRF-3, which forms homodimers or heterodimers with IRF-7, which, in turn, translocate to the nucleus, inducing the expression of type I IFN and pro-inflammatory cytokines. This pathway also includes the participation of the adaptor protein STING, which acts in mitochondrial-associated membrane (MAM) to mediate RIG-I downstream signaling. (3) Type I IFN-mediated antiviral responses occurs via the expression of several IFN-stimulated genes (ISGs). IFN binding to its heterodimeric IFN-α receptor (IFNAR1/2) activates Janus kinase 1 (JAK1) or tyrosine kinase 2 (TYK2), leading to the phosphorylation of the signal transducer and activator of transcription (STAT) proteins, which dimerize and translocate to the nucleus. STAT1–STAT2 heterodimer binds to IFN regulatory factor 9 (IRF9) and migrates to the nucleus, inducing the expression of ISGs through its binding to the IFN-stimulated response elements (ISRE). Type I and type II IFNs can also induce dimerization of STAT3, which translocate to the nucleus, where it binds to gamma-activated sequences (GAS), stimulating the production of both pro- and anti-inflammatory cytokines; (B) Participation of DENV NSPs in the evasion of the host immune response. (1) DENV NS1 inhibits complement activation by interacting with different components of the complement system, including C1 proenzyme, C1s, C4, C4b, and MBL. The formation of the complex C4-NS1-C1s/C1 results in degradation of C4, impairing the formation of C3 convertase. NS1 binding to MBL protects DENV against MBL-mediated virus neutralization by the lectin pathway of complement activation. (2) DENV NSPs impair the innate immune response mediated by viral dsRNA recognition. NS4B interacts with the CARD domain of MAVS, impairing its binding to the cytoplasmic PRRs. Moreover, this protein, by inducing the formation of convoluted membranes (CM) and promoting mitochondrial elongation, inhibits the translocation of PRRs to MAMs. NS2B/NS3 interacts with IKKε and cleaves STING and NS2A together with NS4B, inhibiting the phosphorylation of TBK1 and its substrate IRF3. These steps impair the activation of transcription factors IRF-3 and IRF-7. (3) DENV NSPs inhibit INF-stimulated signaling in different points. NS4B interacts with STAT1, blocking its phosphorylation, and NS5 mediates STAT2 degradation, so both proteins inhibit the expression of ISGs by interfering in ISRE activation. Additionally, NS1 interacts with STAT3, inhibiting the formation of its homodimers, thus preventing GAS-induced gene expression. Red arrows represent the events induced by NSPs, while dashed red arrows represent those ones that are blocked by NSPs. ER, endoplasmic reticulum; MT, mitochondria. Full article ">Figure 2 Cont.
Involvement of DENV NSPs in the evasion of host innate immune response. (A) Pathways of immune response affected by DENV NSPs. (1) Activation of the complement system is triggered by different pathways that converge to the cleavage of factor C3 by the protease C3 convertase. This enzyme formed by the association of two other cleavage products: C4b, a fragment of C4, and C2a, a fragment of C2. The cleavage of C4 and C2 may be catalyzed by two different pathways: the classical pathway, triggered by C1 binding to antigen-antibody complexes, or by the lectin pathway, in which a carbohydrate recognition receptor, such as mannose binding lectin (MBL), associates to a serine protease after binding to carbohydrates. One of the products of C3 cleavage, C3b, binds to C3 convertase changing its substrate specificity, so that the enzyme becomes a C5 convertase. The fragment C5b, generated from the cleavage of C5, binds to the infected cell membrane, initiating the assembly of a complex formed by C6, C7, C8, and C9, which promotes cell lysis. (2) Viral dsRNAs produced during the replication of RNA viruses are recognized by PRRs. Binding of dsRNA leads the cytosolic PPRs to associate with the mitochondrial antiviral signaling protein (MAVS) through its caspase-recruitment domain (CARD), recruiting the TANK-protein kinase 1 (TBK1) and IκB kinase-ε (IKKε). These kinases phosphorylate IRF-3, which forms homodimers or heterodimers with IRF-7, which, in turn, translocate to the nucleus, inducing the expression of type I IFN and pro-inflammatory cytokines. This pathway also includes the participation of the adaptor protein STING, which acts in mitochondrial-associated membrane (MAM) to mediate RIG-I downstream signaling. (3) Type I IFN-mediated antiviral responses occurs via the expression of several IFN-stimulated genes (ISGs). IFN binding to its heterodimeric IFN-α receptor (IFNAR1/2) activates Janus kinase 1 (JAK1) or tyrosine kinase 2 (TYK2), leading to the phosphorylation of the signal transducer and activator of transcription (STAT) proteins, which dimerize and translocate to the nucleus. STAT1–STAT2 heterodimer binds to IFN regulatory factor 9 (IRF9) and migrates to the nucleus, inducing the expression of ISGs through its binding to the IFN-stimulated response elements (ISRE). Type I and type II IFNs can also induce dimerization of STAT3, which translocate to the nucleus, where it binds to gamma-activated sequences (GAS), stimulating the production of both pro- and anti-inflammatory cytokines; (B) Participation of DENV NSPs in the evasion of the host immune response. (1) DENV NS1 inhibits complement activation by interacting with different components of the complement system, including C1 proenzyme, C1s, C4, C4b, and MBL. The formation of the complex C4-NS1-C1s/C1 results in degradation of C4, impairing the formation of C3 convertase. NS1 binding to MBL protects DENV against MBL-mediated virus neutralization by the lectin pathway of complement activation. (2) DENV NSPs impair the innate immune response mediated by viral dsRNA recognition. NS4B interacts with the CARD domain of MAVS, impairing its binding to the cytoplasmic PRRs. Moreover, this protein, by inducing the formation of convoluted membranes (CM) and promoting mitochondrial elongation, inhibits the translocation of PRRs to MAMs. NS2B/NS3 interacts with IKKε and cleaves STING and NS2A together with NS4B, inhibiting the phosphorylation of TBK1 and its substrate IRF3. These steps impair the activation of transcription factors IRF-3 and IRF-7. (3) DENV NSPs inhibit INF-stimulated signaling in different points. NS4B interacts with STAT1, blocking its phosphorylation, and NS5 mediates STAT2 degradation, so both proteins inhibit the expression of ISGs by interfering in ISRE activation. Additionally, NS1 interacts with STAT3, inhibiting the formation of its homodimers, thus preventing GAS-induced gene expression. Red arrows represent the events induced by NSPs, while dashed red arrows represent those ones that are blocked by NSPs. ER, endoplasmic reticulum; MT, mitochondria. Full article ">Figure 3
DENV NSPs-induced host metabolic alterations. During infection, DENV NS4B induces the formation of the convoluted membrane (CM), the membranous structure where viral polyprotein is processed and the resulting proteins accumulate. NS3 recruits the enzyme fatty acid synthase (FASN) to the virus replication sites, besides stimulating its enzymatic activity, increasing de novo FA biosynthesis, which provides lipids for inducing the formation of CM, as well as for the assembly of the viral envelope. Fatty acids (FA) are mobilized from lipid droplets (LD) to undergo β-oxidation mitochondria, providing energy to the high-energy demanding virus replication process. NS4B is able to inhibit phosphorylation of cytoplasmic protein dynamin-related protein 1 (Drp1), preventing the mitochondrial fission process. Fused mitochondria accumulate in infected cells, increasing the efficiency of the oxidative metabolism. Red arrows represent the events induced by NSPs, while dashed red arrows represent those ones that are blocked by NSPs. MFN1, mitofusin 1; MFN2, mitofusin 2; Opa1, optic atrophy protein 1; ER, endoplasmic reticulum; MT, mitochondria. Full article ">

1704 KiB

Open AccessArticle

Isolation and Characterization of a Double Stranded DNA Megavirus Infecting the Toxin-Producing Haptophyte Prymnesium parvum

by Ben A. Wagstaff, Iulia C. Vladu, J. Elaine Barclay, Declan C. Schroeder, Gill Malin and Robert A. Field

Viruses 2017, 9(3), 40; https://doi.org/10.3390/v9030040 - 9 Mar 2017

Cited by 20 | Viewed by 9882

Abstract

Prymnesium parvum is a toxin-producing haptophyte that causes harmful algal blooms globally, leading to large-scale fish kills that have severe ecological and economic implications. For the model haptophyte, Emiliania huxleyi, it has been shown that large dsDNA viruses play an important role [...] Read more.

Prymnesium parvum is a toxin-producing haptophyte that causes harmful algal blooms globally, leading to large-scale fish kills that have severe ecological and economic implications. For the model haptophyte, Emiliania huxleyi, it has been shown that large dsDNA viruses play an important role in regulating blooms and therefore biogeochemical cycling, but much less work has been done looking at viruses that infect P. parvum, or the role that these viruses may play in regulating harmful algal blooms. In this study, we report the isolation and characterization of a lytic nucleo-cytoplasmic large DNA virus (NCLDV) collected from the site of a harmful P. parvum bloom. In subsequent experiments, this virus was shown to infect cultures of Prymnesium sp. and showed phylogenetic similarity to the extended Megaviridae family of algal viruses. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Figure 1
(Top) control culture; (Bottom) ‘Cleared’ culture 96 h post viral infection. Full article ">Figure 2
(A) Thin-sections of healthy P. parvum 946/6 cells; (B) Thin-sections of P. parvum 946/6 48 h post infection. (C) Free Prymnesium parvum DNA virus (PpDNAV) particles in culture supernatant 72 h post infection. C: chloroplast; V: contractile vacuole; N: nucleus; S: scales; M: mitochondria, P: pyrenoid. Full article ">Figure 3
PpDNAV infection cycle propagated on P. parvum 946/6. Graph shows the average number of algal cells in control cultures (squares) and PpDNAV infected cultures (circles). Error bars represent the standard error for triplicate cultures. Full article ">Figure 4
Phylogenetic clustering of PpDNAV with other large algal Megaviridae. Alignment was performed using the default settings of multiple sequence alignment software version 7 (MAFFT) [<a href="#B33-viruses-09-00040" class="html-bibr">33</a>], and the neighbour-joining method (midpoint-rooted) [<a href="#B34-viruses-09-00040" class="html-bibr">34</a>] was used to construct a tree from 19 viral DNA Polymerase Beta (polB) sequences using Molecular Evolutionary Genetics Analysis version 7.0 (MEGA7) [<a href="#B35-viruses-09-00040" class="html-bibr">35</a>]. The final tree was based on 630 ungapped positions, 500 resampling permutations, and was collapsed for bootstrap values <50. The tree shows that PpDNAV clusters with the well-defined clade of Megaviridae and the algal-infecting Megaviridae (red), and not with the Phycodnaviridae (blue). Full article ">

5590 KiB

Open AccessArticle

Change in Emiliania huxleyi Virus Assemblage Diversity but Not in Host Genetic Composition during an Ocean Acidification Mesocosm Experiment

by Andrea Highfield, Ian Joint, Jack A. Gilbert, Katharine J. Crawfurd and Declan C. Schroeder

Viruses 2017, 9(3), 41; https://doi.org/10.3390/v9030041 - 8 Mar 2017

Cited by 11 | Viewed by 9588

Abstract

Effects of elevated pCO₂ on Emiliania huxleyi genetic diversity and the viruses that infect E. huxleyi (EhVs) have been investigated in large volume enclosures in a Norwegian fjord. Triplicate enclosures were bubbled with air enriched with CO₂ to 760 ppmv [...] Read more.

Effects of elevated pCO₂ on Emiliania huxleyi genetic diversity and the viruses that infect E. huxleyi (EhVs) have been investigated in large volume enclosures in a Norwegian fjord. Triplicate enclosures were bubbled with air enriched with CO₂ to 760 ppmv whilst the other three enclosures were bubbled with air at ambient pCO₂; phytoplankton growth was initiated by the addition of nitrate and phosphate. E. huxleyi was the dominant coccolithophore in all enclosures, but no difference in genetic diversity, based on DGGE analysis using primers specific to the calcium binding protein gene (gpa) were detected in any of the treatments. Chlorophyll concentrations and primary production were lower in the three elevated pCO₂ treatments than in the ambient treatments. However, although coccolithophores numbers were reduced in two of the high-pCO₂ treatments; in the third, there was no suppression of coccolithophores numbers, which were very similar to the three ambient treatments. In contrast, there was considerable variation in genetic diversity in the EhVs, as determined by analysis of the major capsid protein (mcp) gene. EhV diversity was much lower in the high-pCO₂ treatment enclosure that did not show inhibition of E. huxleyi growth. Since virus infection is generally implicated as a major factor in terminating phytoplankton blooms, it is suggested that no study of the effect of ocean acidification in phytoplankton can be complete if it does not include an assessment of viruses. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Figure 1
Temporal changes over the period of the experiment in (a) pH, which was calculated from measurements of (b) pCO2 in µatmospheres; (c) nitrate concentration, µmol·N·L−1; (d) phosphate concentration µmol·P·L−1; (e) chlorophyll concentration µg·L−1; and (f) depth-integrated primary production as mg·C·m−2·d−1. Enclosures M1 (▲), M2 (■), M3 (●), M4 (■), M5 (●), M6 (▲). Full article ">Figure 2
Total coccolithophore numbers assessed by flow cytometry. Enclosures M1 (▲), M2 (■), M3 (●), M4 (■), M5 (●), M6 (▲). Full article ">Figure 3
TEM images of identical Emiliania huxleyi morphologies (typical type A) present in both pCO2 treatments throughout the experiment. Full article ">Figure 4
DGGE gels of EhV mcp-PCR products during the experiment from (a) high pCO2-treatment mesocosms, 1, 2, and 3 and (b) ambient pCO2-treatment mesocosms 4, 5, and 6. Bands that migrated at the same position when run on the same gel are indicated with the same symbol. Full article ">Figure 5
Bray–Curtis multidimensional plots based on the DGGE profiles (<a href="#viruses-09-00041-f004" class="html-fig">Figure 4</a>) for EhV from (a) the high pCO2-treatment mesocosms 1, 2, and 3 and (b) ambient pCO2-treatment mesocosms 4, 5, and 6. “Early stage” corresponds to 7–9 May when coccolithophore numbers were <1000 cells mL−1 in ambient enclosures and “mid/late stage” corresponds to 12–14 May when coccolithophore numbers exceeded 1500 cells mL−1 in ambient enclosures. Contours indicate the percentage similarity, as indicated. Full article ">

1989 KiB

Open AccessArticle

Virus Resistance Is Not Costly in a Marine Alga Evolving under Multiple Environmental Stressors

by Sarah E. Heath, Kirsten Knox, Pedro F. Vale and Sinead Collins

Viruses 2017, 9(3), 39; https://doi.org/10.3390/v9030039 - 8 Mar 2017

Cited by 10 | Viewed by 6443

Abstract

Viruses are important evolutionary drivers of host ecology and evolution. The marine picoplankton Ostreococcus tauri has three known resistance types that arise in response to infection with the Phycodnavirus OtV5: susceptible cells (S) that lyse following viral entry and replication; resistant cells (R) [...] Read more.

Viruses are important evolutionary drivers of host ecology and evolution. The marine picoplankton Ostreococcus tauri has three known resistance types that arise in response to infection with the Phycodnavirus OtV5: susceptible cells (S) that lyse following viral entry and replication; resistant cells (R) that are refractory to viral entry; and resistant producers (RP) that do not all lyse but maintain some viruses within the population. To test for evolutionary costs of maintaining antiviral resistance, we examined whether O. tauri populations composed of each resistance type differed in their evolutionary responses to several environmental drivers (lower light, lower salt, lower phosphate and a changing environment) in the absence of viruses for approximately 200 generations. We did not detect a cost of resistance as measured by life-history traits (population growth rate, cell size and cell chlorophyll content) and competitive ability. Specifically, all R and RP populations remained resistant to OtV5 lysis for the entire 200-generation experiment, whereas lysis occurred in all S populations, suggesting that resistance is not costly to maintain even when direct selection for resistance was removed, or that there could be a genetic constraint preventing return to a susceptible resistance type. Following evolution, all S population densities dropped when inoculated with OtV5, but not to zero, indicating that lysis was incomplete, and that some cells may have gained a resistance mutation over the evolution experiment. These findings suggest that maintaining resistance in the absence of viruses was not costly. Full article

(This article belongs to the Special Issue Marine Viruses 2016)

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Figure 1
Mean (± SE) cell density mL−1 of resistant (R), resistant producer (RP) and susceptible (S) O. tauri lines three days after OtV5 inoculation in five environments. Points represent the average of the three assay replicates for each evolved population. Inoculated = populations inoculated with OtV5, Not inoculated = negative control populations that were grown for the same period without OtV5 inoculation. There were three evolved populations of each line. The dashed line represents the starting cell density at 100,000 cell mL−1. Full article ">Figure 2
Change in cell density of the susceptible lines NG’2, NG’3 and NG’4 after OtV5 inoculation one week into the selection experiment (Start) and after 32 transfer cycles of evolution (End). The dashed line represents no change. Full article ">Figure 3
Growth rates as measured by mean cell divisions per day for each evolving population over four time points (1, 14, 20 and 32 transfer cycles). The dashed line represents one cell division per day. T1 is the growth rate following acclimation at the beginning of the experiment. There are no growth measurements for the randomized environment at T1 because lines had only been growing for one transfer cycle. Full article ">Figure 4
Mean cell divisions per day (±SEM). R = resistant, RP = resistant producer, S = susceptible. Each panel represents a growth assay, with cells evolved in the selection environment (top label) and growth rates measured in the assay environment (bottom label). The dashed line indicates, for reference, one cell division per day. Full article ">Figure 5
Competitive ability, as measured by fold difference in growth relative to a roGFP-modified O. tauri line, of evolved populations and control populations assayed in the selection environments. R = resistant, RP= resistant producer, S = susceptible. Each panel represents one assay, with populations evolved in the selection environment (top label) and competitiveness measured in the assay environment (bottom label). The dashed line represents no change (i.e., equal proportions of roGFP and competitor populations). Full article ">

1761 KiB

Open AccessArticle

A New Strategy to Reduce Influenza Escape: Detecting Therapeutic Targets Constituted of Invariance Groups

by Julie Lao and Anne Vanet

Viruses 2017, 9(3), 38; https://doi.org/10.3390/v9030038 - 2 Mar 2017

Cited by 7 | Viewed by 5880

Abstract

The pathogenicity of the different flu species is a real public health problem worldwide. To combat this scourge, we established a method to detect drug targets, reducing the possibility of escape. Besides being able to attach a drug candidate, these targets should have [...] Read more.

The pathogenicity of the different flu species is a real public health problem worldwide. To combat this scourge, we established a method to detect drug targets, reducing the possibility of escape. Besides being able to attach a drug candidate, these targets should have the main characteristic of being part of an essential viral function. The invariance groups that are sets of residues bearing an essential function can be detected genetically. They consist of invariant and synthetic lethal residues (interdependent residues not varying or slightly varying when together). We analyzed an alignment of more than 10,000 hemagglutinin sequences of influenza to detect six invariance groups, close in space, and on the protein surface. In parallel we identified five potential pockets on the surface of hemagglutinin. By combining these results, three potential binding sites were determined that are composed of invariance groups located respectively in the vestigial esterase domain, in the bottom of the stem and in the fusion area. The latter target is constituted of residues involved in the spring-loaded mechanism, an essential step in the fusion process. We propose a model describing how this potential target could block the reorganization of the hemagglutinin HA2 secondary structure and prevent viral entry into the host cell. Full article

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

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