Replication of Marek's Disease Virus Is Dependent on Synthesis of De Novo Fatty Acid and Prostaglandin E2

Marek’s disease virus (MDV) causes deadly lymphoma and induces an imbalance of the lipid metabolism in infected chickens. Here, we discovered that MDV activates the fatty acid synthesis (FAS) pathway in primary chicken embryo fibroblasts (CEFs). In addition, MDV-infected cells contained high levels of fatty acids and showed increased numbers of lipid droplets (LDs). Chemical inhibitors of the FAS pathway (TOFA and C75) reduced MDV titers by approximately 30-fold. Addition of the downstream metabolites, including malonyl-coenzyme A and palmitic acid, completely restored the inhibitory effects of the FAS inhibitors. Furthermore, we could demonstrate that MDV infection activates the COX-2/prostaglandin E2 (PGE2) pathway, as evident by increased levels of arachidonic acid, COX-2 expression, and PGE2 synthesis. Inhibition of the COX-2/PGE2 pathway by chemical inhibitors or knockdown of COX2 using short hairpin RNA reduced MDV titers, suggesting that COX-2 promotes virus replication. Exogenous PGE2 completely restored the inhibition of the COX-2/PGE2 pathway in MDV replication. Unexpectedly, exogenous PGE2 also partially rescued the inhibitory effects of FAS inhibitors on MDV replication, suggesting that there is a link between these two pathways in MDV infection. Taken together, our data demonstrate that the FAS and COX-2/PGE2 pathways play an important role in the replication of this deadly pathogen

Viral Bacterial Artificial Chromosomes: Generation, Mutagenesis, and Removal of Mini-F Sequences

Maintenance and manipulation of large DNA and RNA virus genomes had presented an obstacle for virological research. BAC vectors provided a solution to both problems as they can harbor large DNA sequences and can efficiently be modified using well-established mutagenesis techniques in Escherichia coli. Numerous DNA virus genomes of herpesvirus and pox virus were cloned into mini-F vectors. In addition, several reverse genetic systems for RNA viruses such as members of Coronaviridae and Flaviviridae could be established based on BAC constructs. Transfection into susceptible eukaryotic cells of virus DNA cloned as a BAC allows reconstitution of recombinant viruses. In this paper, we provide an overview on the strategies that can be used for the generation of virus BAC vectors and also on systems that are currently available for various virus species. Furthermore, we address common mutagenesis techniques that allow modification of BACs from single-nucleotide substitutions to deletion of viral genes or insertion of foreign sequences. Finally, we review the reconstitution of viruses from BAC vectors and the removal of the bacterial sequences from the virus genome during this process

Generation, Mutagenesis, and Removal of Mini-F Sequences

Maintenance and manipulation of large DNA and RNA virus genomes had presented an obstacle for virological research. BAC vectors provided a solution to both problems as they can harbor large DNA sequences and can efficiently be modified using well-established mutagenesis techniques in Escherichia coli. Numerous DNA virus genomes of herpesvirus and pox virus were cloned into mini-F vectors. In addition, several reverse genetic systems for RNA viruses such as members of Coronaviridae and Flaviviridae could be established based on BAC constructs. Transfection into susceptible eukaryotic cells of virus DNA cloned as a BAC allows reconstitution of recombinant viruses. In this paper, we provide an overview on the strategies that can be used for the generation of virus BAC vectors and also on systems that are currently available for various virus species. Furthermore, we address common mutagenesis techniques that allow modification of BACs from single-nucleotide substitutions to deletion of viral genes or insertion of foreign sequences. Finally, we review the reconstitution of viruses from BAC vectors and the removal of the bacterial sequences from the virus genome during this process

IFNα and IFNγ Impede Marek’s Disease Progression

Marek’s disease virus (MDV) is an alphaherpesvirus that causes Marek’s disease, a malignant lymphoproliferative disease of domestic chickens. While MDV vaccines protect animals from clinical disease, they do not provide sterilizing immunity and allow field strains to circulate and evolve in vaccinated flocks. Therefore, there is a need for improved vaccines and for a better understanding of innate and adaptive immune responses against MDV infections. Interferons (IFNs) play important roles in the innate immune defenses against viruses and induce upregulation of a cellular antiviral state. In this report, we quantified the potent antiviral effect of IFNα and IFNγ against MDV infections in vitro. Moreover, we demonstrate that both cytokines can delay Marek’s disease onset and progression in vivo. Additionally, blocking of endogenous IFNα using a specific monoclonal antibody, in turn, accelerated disease. In summary, our data reveal the effects of IFNα and IFNγ on MDV infection and improve our understanding of innate immune responses against this oncogenic virus

A Special Issue on Marek’s Disease Virus - The Editors’ View

Marek’s disease virus (MDV), an Alphaherpesvirus belonging to the genus Mardivirus, causes T cell lymphomas in chickens and remains one of the greatest threats to poultry production worldwide. While losses caused by Marek’s disease have been reduced through live-attenuated vaccines, field strains have increased in virulence over recent decades. MDV research has led to a profound understanding of virus-induced pathogenesis and tumor development [1,2,3]. Our goal with this Microorganisms Special Issue on Marek’s disease virus was to collect manuscripts that would provide deeper insights into MDV infection, lytic replication, and latency in vitro and in vivo. Moreover, we assembled reports that provide novel data on pathogenesis, immune system interactions, as well as state-of-the-art concepts to identify approaches to control MDV infections. We were happy to edit seven research articles, three short communications, and a review article on these diverse aspects of MDV infections

The Stellar Parameters and Evolutionary State of the Primary in the d'-Symbiotic System StH\alpha190

We report on a high-resolution, spectroscopic stellar parameter and abundance analysis of a d' symbiotic star: the yellow component of StH\alpha190. This star has recently been discovered, and confirmed here, to be a rapidly rotating (vsini=100 km/s) subgiant, or giant, that exhibits radial-velocity variations of probably at least 40 km/s, indicating the presence of a companion (a white dwarf star). It is found that the cool stellar component has Teff=5300K and log g=3.0. The iron and calcium abundances are close to solar, however, barium is overabundant, relative to Fe and Ca, by about +0.5 dex. The barium enhancement reflects mass-transfer of s-process enriched material when the current white dwarf was an asymptotic giant branch (AGB) star. The past and future evolution of this binary system depends critically on its current orbital period, which is not yet known. Concerted and frequent radial-velocity measurements are needed to provide crucial physical constraints to this d' symbiotic system.Comment: 9 pages, 1 table, 3 figures. In press to Astrophysical Journal Letter

The Transcriptional Landscape of Marek’s Disease Virus in Primary Chicken B Cells Reveals Novel Splice Variants and Genes

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

First Stellar Abundances in the Dwarf Irregular Galaxy Sextans A

We present the abundance analyses of three isolated A-type supergiant stars in the dwarf irregular galaxy Sextans A from high-resolution spectra the UVES spectrograph at the VLT. Detailed model atmosphere analyses have been used to determine the stellar atmospheric parameters and the elemental abundances of the stars. The mean iron group abundance was determined from these three stars to be [(FeII,CrII)/H]=-0.99+/-0.04+/-0.06. This is the first determination of the present-day iron group abundances in Sextans A. These three stars now represent the most metal-poor massive stars for which detailed abundance analyses have been carried out. The mean stellar alpha element abundance was determined from the alpha element magnesium as [alpha(MgI)/H]=-1.09+/-0.02+/-0.19. This is in excellent agreement with the nebular alpha element abundances as determined from oxygen in the H II regions. These results are consistent from star-to-star with no significant spatial variations over a length of 0.8 kpc in Sextans A. This supports the nebular abundance studies of dwarf irregular galaxies, where homogeneous oxygen abundances are found throughout, and argues against in situ enrichment. The alpha/Fe abundance ratio is [alpha(MgI)/FeII,CrII]=-0.11+/-0.02+/-0.10, which is consistent with the solar ratio. This is consistent with the results from A-supergiant analyses in other Local Group dwarf irregular galaxies but in stark contrast with the high [alpha/Fe] results from metal-poor stars in the Galaxy, and is most clearly seen from these three stars in Sextans A because of their lower metallicities. The low [alpha/Fe] ratios are consistent with the slow chemical evolution expected for dwarf galaxies from analyses of their stellar populations.Comment: 40 pages, 8 figures, accepted for publication in A

Three-Dimensional Normal Human Neutral Progenitor Tissue-Like Assemblies: A Model for Persistent Varicella-Zoster Virus Infection and Platform to Study Oxidate Stress and Damage in Multiple Hit Scenarios

The environment of space results in a multitude of challenges to the human physiology that present barriers to extended habitation and exploration. Over 40 years of investigation to define countermeasures to address space flight adaptation has left gaps in our knowledge regarding mitigation strategies partly due to the lack of investigative tools, monitoring strategies, and real time diagnostics to understand the central causative agent(s) responsible for physiologic adaptation and maintaining homeostasis. Spaceflight-adaptation syndrome is the combination of space environmental conditions and the synergistic reaction of the human physiology. Our work addresses the role of oxidative stress and damage (OSaD) as a negative and contributing Risk Factor (RF) in the following areas of combined spaceflight related dysregulation: i) radiation induced cellular damage [1], [2] ii) immune impacts and the inflammatory response [3], [4] and iii) varicella zoster virus (VZV) reactivation [5]. Varicella-zoster (VZV)/Chicken Pox virus is a neurotropic human alphaherpes virus resulting in varicella upon primary infection, suppressed by the immune system becomes latent in ganglionic neurons, and reactivates under stress events to re-express in zoster and possibly shingles. Our laboratory has developed a complex three-dimensional (3D) normal human neural tissue model that emulates several characteristics of the human trigeminal ganglia (TG) and allows the study of combinatorial experimentation which addresses, simultaneously, OSaD associated with Spaceflight adaptation and habitation [6]. By combining the RFs of microgravity, radiation, and viral infection we will demonstrate that living in the space environment leads to significant physiological consequences for the peripheral and subsequently the central nervous system (PNS, CNS) associated with OSaD generation and consequentially endangers long-duration and exploration-class missions